Classification system for inherited epidermolysis bullosa. Based on Fine et al. [4].
\r\n\tAntiphospholipid syndrome is likely to be involved in a small percentage of infertility. It is mainly associated with repeated miscarriages, secretion of cytokines, and growth factors that affect both ovulation, fertilization, and implantation, as well as a previous ectopic pregnancy in the fallopian tubes. Ectopic pregnancy is linked to taking drugs to induce ovulation, but the exact pathogenetic mechanism associated with hormone intake and fallopian tube damage is not yet known.
\r\n\tFertilization of the egg with the sperm under normal conditions occurs in the fibrous part of the fallopian tube. After 3-4 days the fertilized egg reaches the uterus, where the blastocyst within 2 to 4 days may be in a state of immersion in the endometrial tissue. Thus, the implantation takes place on the 20th-21st day of the 4-week menstrual cycle. The pathology of the function of the fallopian tubes is most often associated with inflammatory processes of any etiology. Non-specific infection plays a predominant role, the spread of which contributes to abortion, intrauterine contraception, diagnostic intervention and the occurrence of obstetric and perinatal complications.
\r\n\tIn recent years, the increasing incidence of chlamydial infection has been linked to the occurrence of ectopic pregnancy. Along with the inflammatory nature of the structure and function of the fallopian tubes, endometriosis also seems to play an important e localization may be ovarian, cervical or intra-abdominal, but in most cases it is tubal. The incidence of ectopic pregnancies in the USΑ has at least quadrupled in recent years with the probability nowadays standing at 20 per 1000 pregnancies. Ectopic pregnancy in the USA is reported to be the cause of death in 10% of obstetric deaths, but it is important to note that most of these deaths are bleeding-related and preventable.
\r\n\tA clear trend of increasing the frequency of ectopic pregnancies has been observed in the last ten years, which is triggered by two main reasons. On the one hand, the localization of inflammatory processes in the internal genitals is constantly increasing, while at the same time, the number of surgeries on the fallopian tubes is increasing in order to improve a woman's reproductive capacity. Τhe number of women using intrauterine and hormonal methods of contraception is also increasing and ovulation inducers are increasingly being introduced into the practice of infertility treatment.
\r\n\tOn the other hand, diagnostic capabilities have been improved in recent years by allowing detection of intolerance and even remission of ectopic pregnancy. Currently, an ectopic pregnancy occurs from 0.8-2.4% of cases that are born. In 4-10% of cases, it is repeated. Ectopic pregnancy often occurs as a result of tubal pathology.
\r\n\tRisk factors include smoking, inflammation of the pelvic organs as a result of chlamydia or gonorrhea, and endometriosis, conditions that lead to the formation of scar tissue in the fallopian tubes.role.
\r\n\tThe role of invasive procedures for the treatment of the causative agents of structural abnormalities of the fallopian tubes in the occurrence of ectopic pregnancy is becoming increasingly crucial and even to such an extent that the introduction of microsurgery does not exclude the risks.
",isbn:null,printIsbn:"979-953-307-X-X",pdfIsbn:null,doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!1,isSalesforceBook:!1,isNomenclature:!1,hash:"46740c30c279d7ad0334167a63819809",bookSignature:"Prof. Panagiotis Tsikouras, Prof. Nikolaos Nikolettos, Prof. Werner Rath and Prof. Georg-Friedrich Von Tempelhoff",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/11282.jpg",keywords:"Differential Diagnosis, Early Diagnosis, Abortion, Intrauterine Pregnancy, Surgery, Methotrexate, Salpingostomy, New Aspects, Markers, Endometriosis, Diagnosis, Management",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:0,numberOfDimensionsCitations:0,numberOfTotalCitations:0,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"September 9th 2021",dateEndSecondStepPublish:"October 7th 2021",dateEndThirdStepPublish:"December 6th 2021",dateEndFourthStepPublish:"February 24th 2022",dateEndFifthStepPublish:"April 25th 2022",dateConfirmationOfParticipation:null,remainingDaysToSecondStep:"9 months",secondStepPassed:!0,areRegistrationsClosed:!0,currentStepOfPublishingProcess:5,editedByType:null,kuFlag:!1,biosketch:"Dr. Panagiotis Tsikouras has authored and co-authored multiple peer-reviewed scientific papers and presented works at many national and international conferences. 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From 2007 to 2011, Dr.\nNikos served as director of the In Vitro Fertilization Unit, University Regional General Hospital of Alexandroupolis, Greece. From\n2012 to 2018, he was director of the Laboratory of Reproductive\nPhysiology - Artificial Fertilization. In 2018 he became Professor of Assisted Reproduction at the Democritus University of Thrace, Greece. In 2019 he took over the\nmanagement of the University Obstetrics-Gynecology Clinic, Medical Department,\nDemocritus University of Thrace. 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The inheritance of the affected genes can occur in a dominant or recessive way depending on the subform of the disease. In general, epidermolysis bullosa is caused by mutations in genes encoding structural proteins within the basal membrane zone of the skin. Absence or functional loss of one of these proteins results in a lack of stability of the microarchitecture of the connection between dermis and epidermis leading to a loss of coherence [1]. The basement membrane between the dermis and the epidermis is a complex membrane produced by basal keratinocytes and dermal fibroblasts that acts as mechanical support for the connection of both skin layers. The basal membrane also regulates the metabolic exchange between the two skin compartments [2]. Up to date, there are at least 15 genes associated with EB causing different forms of the disease. Numerous mutations in these genes that encode for structural proteins within keratinocytes or within mucocutaneous basement membranes have been identified up to now [1].
Mutations in the genes, encoding for the keratins 5 and 14 and plectin, lead to epidermolysis bullosa simplex (EBS) characterized by the cytolysis within basal keratinocytes. Junctional epidermolysis bullosa (JEB) is caused by the absence or loss of function of laminin-332, type XVII collagen or integrin-β4. JEB is a severe EB form and is characterized by the separation of the skin within the lamina lucida. Mutations in type VII collagen (encoded by
\n\t\t\t\t | \n\t\t||
Major EB type | \n\t\t\tMajor EB subtypes | \n\t\t\tAffected proteins | \n\t\t
EB simplex (EBS) | \n\t\t\tSuprabasal EBS | \n\t\t\tplakophilin-1, desmoplakin; | \n\t\t
\n\t\t\t | \n\t\t\t | others? | \n\t\t
\n\t\t\t | Basal EBS | \n\t\t\tkeratins 5 & 14; plectin, | \n\t\t
\n\t\t\t | \n\t\t\t | α6β4 integrin, BPAG1 | \n\t\t
Junctional EB (JEB) | \n\t\t\tJEB, Herlitz (JEB-H) | \n\t\t\tlaminin-332, (laminin-5) | \n\t\t
\n\t\t\t | JEB, other | \n\t\t\tlaminin-332, type XVII collagen | \n\t\t
\n\t\t\t | \n\t\t\t | α6β4 integrin, α3 integrin | \n\t\t
Dystrophic EB (DEB) | \n\t\t\tDominant DEB (DDEB) | \n\t\t\ttype VII collagen | \n\t\t
\n\t\t\t | Recessive DEB (RDEB) | \n\t\t\ttype VII collagen | \n\t\t
Kindler syndrome | \n\t\t\t\n\t\t\t | kindlin-1 | \n\t\t
\n\t\t\t\t | \n\t\t||
Major types | \n\t\t\tEBS subtypes | \n\t\t\tAffected proteins | \n\t\t
EBS suprabasal | \n\t\t\tdesmoplakin | \n\t\t|
\n\t\t\t | plakophilin-1 | \n\t\t|
\n\t\t\t | ? | \n\t\t|
EBS basal | \n\t\t\tEBS, localized (EBS-loc)a | \n\t\t\tK5, K14 | \n\t\t
\n\t\t\t | EBS, Dowling Meara (EBS-DM) | \n\t\t\tK5, K14 | \n\t\t
\n\t\t\t | EBS, other generalized (EBS,gen-nonDM)b | \n\t\t\tK5, K14, BPAG1 | \n\t\t
\n\t\t\t | \n\t\t\t | \n\t\t |
\n\t\t\t | K5 | \n\t\t|
\n\t\t\t | EBS with muscular dystrophy (EBS-MD) | \n\t\t\tplectin | \n\t\t
\n\t\t\t | \n\t\t\t | \n\t\t |
\n\t\t\t | plectin, α6β4 integrin | \n\t\t|
\n\t\t\t | K14 | \n\t\t|
\n\t\t\t | plectin | \n\t\t|
\n\t\t\t | K5 | \n\t\t|
\n\t\t\t\t | \n\t\t||
\n\t\t\t\ta Previously called EBS, Weber-Cockayne \n\t\t\t\tb Includes patients previously classified as EBS-Koebner | \n\t\t||
\n\t\t\t\t | \n\t\t||
Major JEB subtype | \n\t\t\tSubtypes | \n\t\t\tAffected proteins | \n\t\t
JEB, Herlitz (JEB-H) | \n\t\t\t\n\t\t\t | laminin-332 | \n\t\t
JEB, other (JEB-O) | \n\t\t\tJEB, non-Herlitz, generalized (JEB-nH gen)a | \n\t\t\tlaminin-332, type XVII collagen | \n\t\t
\n\t\t\t | JEB, non-Herlitz localized (JEB-nH loc) | \n\t\t\ttypeXVII collagen | \n\t\t
\n\t\t\t | \n\t\t\t | \n\t\t |
\n\t\t\t | JEB with pyloric atresia (JEB-PA) | \n\t\t\tα6β4 integrin | \n\t\t
\n\t\t\t | laminin-332 | \n\t\t|
\n\t\t\t | \n\t\t | |
\n\t\t\t | laminin-332 α3 chain | \n\t\t|
\n\t\t\t | ? | \n\t\t\tα3 integrin | \n\t\t
\n\t\t\t\t | \n\t\t||
\n\t\t\t\ta Formerly known as generalized atrophic benign EB (GABEB) | \n\t\t||
\n\t\t\t\tb Formerly known as EB progressive | \n\t\t||
\n\t\t\t\t | \n\t\t||
Major DEB subtype | \n\t\t\tSubtypes | \n\t\t\tAffected protein | \n\t\t
DDEB | \n\t\t\tDDEB, generalized (DDEB-gen) | \n\t\t\ttype VII collagen | \n\t\t
\n\t\t\t | \n\t\t | |
\n\t\t\t | \n\t\t | |
\n\t\t\t | \n\t\t | |
\n\t\t\t | \n\t\t | |
\n\t\t\t | \n\t\t | |
RDEB | \n\t\t\tRDEB, severe generalized (RDEB-sev gen)a | \n\t\t\ttype VII collagen | \n\t\t
\n\t\t\t | RDEB, generalized other (RDEB-O) | \n\t\t\t\n\t\t |
\n\t\t\t | \n\t\t | |
\n\t\t\t | \n\t\t | |
\n\t\t\t | \n\t\t | |
\n\t\t\t | \n\t\t | |
\n\t\t\t | \n\t\t | |
\n\t\t\t\t | \n\t\t||
\n\t\t\t\ta Previously called RDEB, Hallopeau-Siemens | \n\t\t
Classification system for inherited epidermolysis bullosa. Based on Fine et al. [4].
Mutations in the gene
The DDEB phenotype is mostly generalized but mild and clinically characterized by recurrent blistering, milia, atrophic scarring, nail dystrophy and eventual loss of nails [3]. See Figure 1A,B. The fact that the defective and wildtype alleles are expressed equally explains the relative mild phenotype in comparison to RDEB [3]. Missense mutations or in frame deletions in
Clinical phenotype of DEB.
The blisters characteristic for EB arise within the dermal-epidermal junction. Having a look at this compartment of the skin helps to understand the cause of blistering in EB. The dermal-epidermal junction is a complex basement membrane synthesized by dermal fibroblasts and basal keratinocytes. Adhesion of the epidermis to the underlying dermis is mechanically supported by the so called basement membrane zone (BMZ). Moreover it regulates the metabolic exchange between these two compartments. Up to now more than 20 macromolecules situated in the dermal-epidermal-junction have been detected and characterized at biochemical and genomic level [2].
Three protein-junction complexes stabilize the adherence of the basal keratinocytes to the dermis. See Figure 2. The hemidesmosomes built up by plectin, the bullous pemphigoid antigen 1 (BPAG1), α6β4 integrin and type XVII collagen (bullous pemphigoid antigen 2 - BPAG2) link the basal keratinocytes with the basement membrane, spanning the lamina lucida and anchored in the lamina densa [2]. Different laminin isoforms are located in the lamina lucida (laminin-332, laminin 6, laminin 10) and contribute along with BPAG2 to the formation of the anchoring filaments. The lamina densa is mainly built up by type VII collagen anchoring the lamina densa to the underlying dermis by the formation of anchoring fibrils [2]. Some other antigens as uncein (19-DEJ-1 antigen), NU-T2 antigen, KF1 antigen, LDA1 antigen, nidogen, heparin-sulfate, proteoglycan, antigens AF1 and AF2, thrombospondin, type V collagen and osteonectin/BM-40 have been detected in the lamina densa but have not yet been adequately characterized [2].
Schematic setup of the cutaneous dermal-epidermal junction zone and localization of structural proteins affected in inherited EB (Diagram by R. Hametner). laminin 5 = laminin 332; EBS = epidermolysis bullosa simplex; JEB = junctional epidermolysis bullosa; DEB = dystrophic epidermolysis bullosa
Type VII collagen is classified in the superfamily of collagens [7]. A protein domain in triple-helical conformation, which provides stability and integrity between connective tissues, is a common structural feature of all collagens [7]. Type VII collagen is a minor collagen in human skin and demonstrates spatially restricted location but it plays a critical role in providing integral stability to the skin because it is the major component of the anchoring fibrils [6,7].
Type VII collagen molecules are characterized by the two non-collagenous NC-1 and NC-2 domains flanking a central collagenous, triple-helical segment [7]. In contrast to other interstitial collagens the repeating Gly-X-Y collagenous sequence is interrupted by 19 imperfections due to insertions or deletions of amino acids. There is a 39 amino acid non-collagenous hinge region susceptible to proteolytic digestion with pepsin in the middle of the triple-helical domain [15]. The amino terminal NC-1 domain (approximately 145kD in size), is built up of sub-modules with homology to known adhesive proteins, including segments with homology to cartilage matrix protein (CMP), nine consecutive fibronectin type III-like (FN-III) domains, a segment with homology to the A domain of von Willebrand factor, and a short cysteine and proline-rich region [15]. The C-terminal non-collagenous NC-2 domain is with 30kD in size relatively small, and contains a segment with homology to Kuniz protease inhibitor molecule [16,17].
The 32kb gene encoding a 9,2kb mRNA has been mapped to the short-arm of chromosome 3p21.1 [18]. The encoding primary sequence and the gene structure of type VII collagen are well conserved. The mouse gene shows 90.4% identity at the protein level and 84.7% homology at the nucleotide level, indicating the importance of type VII collagen as a structural protein [19].
The expression pattern of
Type VII collagen is synthesized by two cell types in the skin: keratinocytes and fibroblasts [22]. After synthesis of complete pro-α1 (VII) polypeptides, three polypeptides are associated through their carboxy-terminal ends to a trimer molecule, which is then folded in its collagenous segment into the triple-helical formation. Past to secretion into the extracellular milieu two type VII collagen molecules are aligned into an anti-parallel dimer with the amino-terminal domains present at both ends of the molecule [6]. During dimer-assembly stabilization by inter-molecular disulfide bond formation and a proteolytic removal of a part of the carboxy-terminal ends (NC-2 domain) of both type VII collagen molecules take place [23]. Large numbers of these anti-parallel dimers aggregate laterally to form anchoring fibrils, which then can be identified by their characteristic, centro-symmetric banding patterns in transmission electron microscopy [7].
The affinity of the NC-1 domain to bind the principal components of the cutaneous basement membrane, laminin-332, laminin-311 and type IV collagen provides stability to the dermo-epidermal adhesion on the dermal site at the lamina lucida/papillary dermis interface [6,24,25]. Arg-Gly-Asp sequences in the NC-1 domain serve as integrin mediated attachment sites for cells to adhere to extracellular matrix components such as fibronectin [26].
Mutations in
RDEB, generalized other, the milder phenotype, is mostly caused by PTCs, small deletions, substitutions of glycine residues in the collagenous domain, splice-site mutations within NC-2 [32-35], delayed termination codons [36], in frame exon skipping [29,36], or missense substitution mutations involving amino acids other than glycine [29,37,38], the majority involving arginine residues resulting either in the loss of an ionic charge or in the introduction of a bulky chain at an external position of the triple helix [27]. Thereby these mutations usually concern a critical amino acid and change the conformation of the protein, which then might still be able to assemble into a small number of anchoring fibrils but is likely to be unstable when they laterally aggregate. Anyhow some full length type VII collagen polypeptides can still be built up [39].
DDEB is caused by glycine substitutions within the triple helical domain of
So far there are only two viable mouse models with defects in the
Due to the size of the
\n\t\t\t\t | \n\t\t\t\n\t\t\t\t | \n\t\t\t\n\t\t\t\t | \n\t\t
Woodley et al. | \n\t\t\tType VII collagen minigene | \n\t\t\t2000 | \n\t\t
Sat et al. | \n\t\t\tCosmid clone containing the entire | \n\t\t\t2000 | \n\t\t
Mecklenbeck et al. | \n\t\t\tMicroinjection of a | \n\t\t\t2002 | \n\t\t
Urda et al. | \n\t\t\tФC31 bacteriophage integrase | \n\t\t\t2002 | \n\t\t
Chen et al. | \n\t\t\tMinimal lentiviral vectors | \n\t\t\t2002 | \n\t\t
Baldeschi et al | \n\t\t\tCanine type VII collagen | \n\t\t\t2003 | \n\t\t
Woodley et al | \n\t\t\tTargeting fibroblasts instead of keratinocytes (lentivirally) | \n\t\t\t2003 | \n\t\t
Gache et al. | \n\t\t\tFull-length cDNA (retrovirally) | \n\t\t\t2004 | \n\t\t
Woodley et al. | \n\t\t\tIntradermal injection of recombinant type VII collagen | \n\t\t\t2004 | \n\t\t
Woodley et al. | \n\t\t\tIntradermal injection of lentiviral vectors in vivo | \n\t\t\t2006 | \n\t\t
Goto et al. | \n\t\t\tTargeting fibroblasts instead of keratinocytes (retrovirally) | \n\t\t\t2006 | \n\t\t
Goto et al. | \n\t\t\tTargeted exon skipping using antisense | \n\t\t\t2006 | \n\t\t
Wong et al. | \n\t\t\tIntradermal injection of allogenic wildtype fibroblasts into a patient | \n\t\t\t2007 | \n\t\t
Fritsch et al. | \n\t\t\tIntradermal injection of murine wildtype fibroblasts in a DEB mouse model | \n\t\t\t2008 | \n\t\t
Remington et al. | \n\t\t\tIntradermal injection of human type VII collagen in mice | \n\t\t\t2009 | \n\t\t
Titeux et al. | \n\t\t\tMinimal self-inactivating retroviral vectors harbouring the full length human | \n\t\t\t2010 | \n\t\t
Wagner et al. | \n\t\t\tAllogeneic bone marrow transplantation | \n\t\t\t2010 | \n\t\t
Siprashvili et al. | \n\t\t\tFull-length cDNA (retrovirally) | \n\t\t\t2010 | \n\t\t
Murauer et al. | \n\t\t\t3´ Trans-splicing of | \n\t\t\t2011 | \n\t\t
Therapy approaches to restore type VII collagen expression
Woodley et al. used a type VII collagen minigene, which contains the intact noncollagenous domains NC1 and NC2 and part of the central collagenous domain. This approach resulted after transduction into DEB keratinocytes in persistent synthesis and secretion of a 230kDa recombinant minicollagen VII [47]. However deletions in
Until now, no
RNA
In SMaRT constructs that are engineered to bind the introns of specific pre-mRNAs – RNA
Schematic overview on different applications of SMaRT.
Arrowheads indicate mutations.
The efficiency of
\n\t\t\t\t | \n\t\t\t\n\t\t\t\t | \n\t\t\t\n\t\t\t\t | \n\t\t
\n\t\t\t\t | \n\t\t\t\n\t\t\t | \n\t\t |
Puttaraju et al. | \n\t\t\t3´ repair of lacZ in a tractable system | \n\t\t\t2001 | \n\t\t
Chao et al. | \n\t\t\t3´ repair of haemophilia A mice in vivo | \n\t\t\t2003 | \n\t\t
Dallinger et al. | \n\t\t\t3´ repair in a lacZ model system in a keratinocyte specific background | \n\t\t\t2003 | \n\t\t
Liu et al. | \n\t\t\t3´ repair of CFTR mRNA (adenovirally) | \n\t\t\t2005 | \n\t\t
Rodriguez-Martin et al. | \n\t\t\t3´ reprogramming of tau alternative splicing in a model system | \n\t\t\t2005 | \n\t\t
Zayed et al. | \n\t\t\t3´ repair of DNA-PKcs in SCID (delivery via sleeping beauty) | \n\t\t\t2007 | \n\t\t
Chen et al. | \n\t\t\t3´ repair dystrophia myotonica type 1 pre-mRNA | \n\t\t\t2008 | \n\t\t
Coady et al. | \n\t\t\t3´ SMN2 trans-splicing in combination with blocking an cis-splice sit in mice in vivo | \n\t\t\t2010 | \n\t\t
Murauer et al. | \n\t\t\tFunctional 3´ repair of the COL7A1 gene | \n\t\t\t2010 | \n\t\t
Wang et al. | \n\t\t\t3´ introduction of therapeutic proteins in highly abundant albumin transcripts in mice in vivo | \n\t\t\t2009 | \n\t\t
Gruber et al. | \n\t\t\t3´ reprogramming of tumor marker genes to introduce suicide genes | \n\t\t\t2011 | \n\t\t
\n\t\t\t\t | \n\t\t||
Mansfield et al. | \n\t\t\t5´ repair of CFTR mRNA | \n\t\t\t2000 | \n\t\t
Kierlin-Duncan et al. | \n\t\t\t5´ repair of β-globin mRNA | \n\t\t\t2007 | \n\t\t
Wally et al. | \n\t\t\t5´ repair of the PLEC1 gene | \n\t\t\t2007 | \n\t\t
Wally et al. | \n\t\t\t5´ K14 mRNA reprogramming | \n\t\t\t2010 | \n\t\t
Rindt et al. | \n\t\t\t5´ trans-splicing repair of huntingtin at mRNA level | \n\t\t\t2012 | \n\t\t
\n\t\t\t\t | \n\t\t||
Lorain et al. | \n\t\t\tExon exchange approach to repair Duchenne dystrophin transcripts in a minigene | \n\t\t\t2010 | \n\t\t
Koller et al. | \n\t\t\tA screening system for IER molecules | \n\t\t\t2011 | \n\t\t
Overview on functional
RNA
So a tractable lacZ model repair system, in which user defined target introns can be
5’
Lorain et al. primarily published the methodology of internal exon replacement (IER) to correct a dystrophin minigene on mRNA level [87]. Recently, Koller et al. developed a new RTM screening system to improve double RNA
So far, there are no general rules for the design of highly efficient
We started to establish 5´
Intron 15 was chosen as target intron because its size of about 1,5kb allows to create a large number of different binding domains. To generate a large amount of different RTMs, containing binding domains with different binding affinities to the target intron, the target exon/intron was digested out of the artificial target used in the screen with HindIII and BamHI and digested with CviJI*. The resulting fragments with a length of 50-750bp were cloned into the RTM backbone. Binding domains were identified by colony PCR using a forward primer situated in the 5´ half of the split GFP and a vector specific reverse primer. Possible binding domains with different lengths were detected on a 2% agarose gel after gel electrophoresis. To check orientation and location of the binding domain, clones with inserts were sequenced. To evaluate the created RTM library the artificial target containing the target intron (intron15) and the 3´ half of the split AcGFP instead of the 3´ part of murine
Fluorescence microscopy of with RTM library and target double transfected HEK293 cells.
Flow cytometric analysis: 5´ screen for murine
The RTM with the highest AcGFP/DsRED ratio (RTM+3A)
Endogenous
RNA
We have established all three modes of
We want to thank Prof. Leena Bruckner-Tuderman for providing the murine keratinocytes. Moreover we thank the Austrian Science Fund (FWF) for financing the project “Development of a 5´
Water molecules, which reside on the surfaces of proteins or lipid bilayers or in tissues and cells, exhibit properties that are significantly different from those found in pure or bulk water as water molecules in such biological systems face additional interactions [1, 2, 3, 4, 5, 6, 7]. Unique properties of such water molecules, which are confined in microenvironments of biological surfaces or interfaces are popularly termed as ‘biological water’. This water drives many biological processes, in which it plays wide varieties of roles at different levels of complexity making its participation increasingly evident as an active agent and not simply as the spectator solvent [8, 9, 10, 11, 12]. Biological water differs from bulk water in a number of ways. First, clustering of the water molecules at the surface of a protein increases the local density by as much as 25% compared to that in bulk water [13, 14]. MD simulation by Smith et al. has revealed that about half of this density increase arises from the shortening of the average water – water distances and the other half from an increase in the coordination number [15]. Second, replacement of the water–water hydrogen bonds by the water–protein hydrogen bonds in the protein hydration layer lowers the freezing point of the hydration water (i.e. prevents formation of ice). The hydration layer of many proteins does not freeze even at sub-zero temperatures and thus life may sustain even at low temperatures [16, 17, 18]. Third, in bulk water, mutual polarization of the hydrogen bonded water molecules increases the dipole moment and dielectric constant. Such polarization is absent for water molecules in hydration layers of biological molecules, i.e. biological water is less polar than bulk water [19]. Further, measurements of water dynamics suggest that around 10–25% of water molecules in cells have slower reorientation dynamics, by around an order of magnitude, than those in the bulk [2, 20]. Centrality of liquid water to life can be easily understood from the fact that condition for the search for the possibility of life elsewhere is the presence of trace of water [21]. In spite of this status, role of water in sustaining life is still not understood perfectly [12]. It is now, however, clear that water plays an active role in the life of the cell over many scales of time and distance and exhibits diverse structural and dynamical roles in molecular cell biology [3, 22].
In liquid (or bulk) water, the molecules form a tetrahedrally coordinated motif, which is the building block of ephemeral six-membered (ice-like) or five-membered (clathrate-like) ring structure, consisting of fluctuating network of hydrogen bonds, but each bond has an average lifetime of about a picosecond [23]. Thermodynamics of hydration in water are generally governed by a balance between the enthalpic and entropic consequences, namely the enthalpies of water–water and water–solute interactions (hydrogen bonding, electrostatic, and van der Waals) and the entropies of disrupting the relatively ordered hydrogen-bonded networks of bulk water and forming new hydrogen bonds to suit the geometric factors of the biological interfaces [15, 24, 25].
Similarities in water dynamics in hydration shells of various proteins [26] suggest that the dynamics are determined by rather general features of surface chemistry and topology, which induce excluded volume effects and hinder the approach of new hydrogen bond acceptors within the hydration network. There is now universal recognition that the dynamical behavior of biological macromolecules cannot be decoupled from that of water and the dynamical behavior of the hydration shell largely controls the chemical function of the biological molecules [27, 28].
Since the development of this understanding that biomolecules are surrounded by a sheath of hydration water, which takes active part in all of their normal activities, extensive efforts have been made to perceive the detailed structure and dynamics of the hydration layer using a variety of spectroscopic methods, such as X-ray and neutron scattering [14, 29, 30], NMR [31, 32, 33], second harmonic generation [34, 35] and ultrafast fluorescence and IR spectroscopies [1, 2, 22, 26, 36, 37], assisted by ab initio and molecular dynamics simulations [24, 25, 38, 39]. However, these measurements could provide information about the dynamics of water and the biomolecules at a low-hydration level corresponding to the first layer of hydration water only. Since fluctuations of protein and solvent dynamics take place over a wide range of length scales (in the range of a few nm) and timescales (from milliseconds to picoseconds) influencing several aspects of protein function, no single technique can span so many spatial and temporal orders of magnitude. Therefore, there has obviously been a debate about how to reconcile the results of different experimental methods that explore dynamics [1, 2, 22, 40, 41].
Recent realization that low-frequency and large-amplitude modes of water molecules in the hydration shell are particularly important in controlling the conformational changes that dominate protein function, has led to using the terahertz (THz) spectroscopy to observe water dynamics around biological molecules [42, 43]. THz radiation (1 THz = 1012 Hz = 33.33 cm−1) excites the low frequency vibrations of the solvated protein and directly probes the collective hydrogen bond dynamics of the coupled protein-water system in (sub-) ps time domain. Librational, translational and intermolecular, collective motions of hydration water, as well as large amplitude motions of biomolecules, also occur in similar time scale (see Figure 1). In frequency domain, this corresponds to low-frequency modes in the 1–10 THz frequency region. In spite of the fact that water molecules strongly attenuate the THz radiation in this frequency region, laser or accelerator-based THz sources (vide Section 2) are now powerful enough to penetrate water layers. Modern THz spectroscopic techniques can provide valuable information about water dynamics in the frequency region of the electromagnetic spectrum (0.3–20 THz or 10 to 600 cm−1, the so-called “terahertz gap” between the dielectric and the infrared regimes). This has offered a unique view of the hydration water in fully solvated biomolecules.
The hierarchy of time scales for motions of proteins and their hydration environment. (Adopted with permission from ref. [
Two kinds of THz spectroscopy techniques, namely THz transmission and THz time-domain spectroscopy (THz-TDS), have extensively been used to record the THz absorption spectra of biological materials. THz transmission spectroscopy is a simple single or dual beam steady state spectrometer using THz radiation beam to estimate the THz absorption coefficient. THz transmission or absorption spectroscopy of fully solvated biomolecules in water yields direct information on the global dynamical correlations among solvent (water) molecules. However, application of THz spectroscopy to study solvated biomolecules was not possible because of huge absorption of water in the THz frequency range. This problem was solved by the development of the p-Ge laser, which was a strong the THz emitter [44, 45, 46, 47]. Prior to this development, weak radiation power of standard sources like a globar or an arc lamp in the far IR and THz region was the cause for poor signal to noise ratio and the measurements were limited to powders or hydrated films of biomolecules, but not in their native aqueous environment. Free-electron lasers [48, 49] and synchrotrons [50], which are also sources of high power far-IR and THz pulses, have also been reported to be used for spectroscopy studies of biomolecules but are not easily accessible. THz transmission spectroscopy of water in biological systems received a momentum after the discovery of the p-Ge laser. This laser is a powerful table-top THz source (up to about 400 W) and has the tunability in the 80–100 cm−1 (2.4-3.3 THz) frequency region. Development of a table top dual beam (reference and sample arms) THz transmission spectrometer ensured accurate measurements of absorption of water of thickness as large as 100 μm [51].
Accelerators delivering ultrashort pulses of relativistic electrons have been widely used as a source of high intensity THz radiation. Free electron lasers and synchrotrons are actually seeded by short duration electron pulses. An easier method of generating high intensity and tuneable THz radiation using an ultrafast electron accelerator is the coherent transition radiation (CTR). CTR occurs when a charged particle passes through an interface between media with different dielectric constants [52, 53, 54, 55]. Sudden change in the dielectric constants along the electron’s path causes a discontinuity in the electric field at the interface and this discontinuity readjusts itself as radiation spreading out from the point where the electron passes through the discontinuity. The angular spectral energy density of the CTR depends on the dielectric constants of the two media [56, 57]. Since the dielectric constants of aluminum and gold are much more than that of vacuum in far-IR region, these two metals are frequently used as perfect conductors. Direction of propagation and the intensity distribution of the CTR with respect to the angle of incidence of the electron pulse on the target has been discussed in detail in Ref. [53].
At wavelengths shorter than the bunch length, the emitted radiation field is incoherent and total intensity is proportional to the number of electrons (N). But at wavelengths longer than the electron bunch length, the radiation emitted from the bunch is coherent. With a typical number of electrons per bunch on the order of 108–109, the coherent radiation intensity greatly exceeds that of incoherent radiation. Especially, at long wavelengths compared to the bunch length, the radiation intensity is proportional to the square of the number of electrons in the bunch. Therefore, it is possible to generate coherent radiation in the far-IR or THz spectral range from short electron bunches with bunch lengths of hundred micron or less. Moreover, the shorter the bunch length, the broader is the radiation spectrum that can be generated. With larger number of electrons in the bunch, the total intensity (both coherent and incoherent components) of the CTR at frequency ω is given by,
In this equation, contribution of the coherent component, Icoherent(ω), is given by,
Here,
and Ie(ω) is the radiation intensity generated from one electron in the bunch and Eqs. (1) and (2) have been derived assuming that all electrons in the bunch have the same energy.
In THz transmission measurements at AIST, Tsukuba, Japan, a femtosecond linear accelerator system, which delivered electron pulses of about 300 fs duration, and electron energy of 42 MeV and ∼ 100 pC charge (∼108 electrons per one macro-pulse) [58], was used. Electron beam was directed to hit a thin gold foil (thickness is about 500 μm) at an incidence angle of 45°. CTR, which contained the component of THz radiation, was collected in the direction perpendicular to the electron beam using a gold coated parabolic mirror and the intensity of the THz radiation was measured using a Schottky diode detector [54]. Intensity of the THz pulse generated by this method was of high intensity (about 100 nJ/micropulse and covered a wide frequency range (0.3–2.5 THz), which was practically determined by the frequency response of the Schottky diode detector. Band pass filters (Tydex, Russia) were used to select a band of THz frequencies for estimation of absorption of protein solutions. Absorbance of the solutions of different path lengths were measured. Samples were taken in Bruker liquid sample cells, path lengths of which were varied using Teflon spacers of required thicknesses. Fused silica windows having thickness of 1.5 mm were used in the Bruker sample cell. Thicknesses of the Teflon spacers used were in the range of 20 to 150 μm. Absorption coefficient value (in cm−1) of the solution was estimated from the slope of the linear plot of absorbance vs. path length of the cell (Eq. (4)) [59]. Experimental arrangement used here corresponded to that of a single beam absorption spectrometer.
Here, I0 and Id represent the beam intensities in the absence (i.e. using the blank cell) and presence of the sample solution, respectively. αsol represents the absorption coefficient of the sample solution, d is the path length or thickness of the sample.
The absorption coefficient of the sample, α, is defined by,
The real part
Presently, the THz-TDS in the 0.1–10 THz (3.33 cm−1– 333 cm−1) region is the most popular technique, which has been extensively applied in the fields of research in chemistry, materials science, physics, engineering, medicine as well as in industry [60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74]. THz -TDS has also been proved to be a valuable technique for investigation of low frequency dielectric relaxation and vibrational spectroscopy of hydrogen bonded liquids, such as water, alcohols, and others [75, 76, 77]. While various configurations of the THz time-domain spectrometer have been used depending on the kind of applications, we will describe here the general principle of the technique and the spectrometer configuration used for investigation of biomolecules.
The core principle of the THz-TDS technique, which uses a short duration (say a few tens of femtosecond) pulses of THz radiation, is measurement of the transient electric field associated with the THz pulse rather than variation of intensity of the frequency components of the THz pulse. However, since the response or the rise times of the electronic components and detectors are slower than a few ps, it is the normal practice to use the pump-and-probe kind of optical configurations in ultrafast spectrometers in order to overcome the slow response of the detectors. To achieve sub-picosecond time resolution in THz-TDS, the ultrashort NIR optical pulse (typically shorter than 100 fs) is beam-split along two paths to generate pump and probe pulses and to detect the time dependent THz field using a time-delay stage. Schematic diagram presented in Figure 2 represents the principle of a THz- time-domain spectrometer.
(a): Schematic diagram of a typical THz - time-domain spectrometer. (b): Temporal variation of electric field of the THz pulse and its Fourier transform in air and in presence of water.
The pump pulse, which contains the major amount of energy (more than 70% of the total energy of the laser beam), is used for generating broadband pulses of THz radiation using a photoconductive antenna consisting of a low temperature grown GaAs or InGaAs film covered with metallic contacts for application of bias voltage, or by optical rectification in ZnTe(110) crystal. The pulsed beam of THz radiation is collimated and focused onto the sample using gold coated parabolic mirrors. After transmission through the sample, the THz beam is recollimated and refocused on another photoconductive antenna or ZnTe(110) crystal, which works as the THz detector. Recently, ZnTe (110) crystal are being used extensively both as THz emitter as well as in detection because of possibility of its application in much wider THz frequency region as compared to that has been possible using photoconductive antenna.
The probe beam is used to gate the detector and measure the instantaneous THz electric field using the method of electro-optic sampling. A high precision linear motion stage is used to delay the probe pulses to arrive at detector crystal with respect to the pump pulses. The THz pulse and the gate (NIR) pulse are propagated collinearly through the ZnTe crystal. The THz pulse induces a birefringence in ZnTe crystal, which is read out by a linearly polarized gate pulse. When both the gate pulse and the THz pulse are present in the crystal at the same time, the polarization of the gate pulse will be rotated by the THz pulse depending on the strength of the electric field of the THz pulse. Using a λ/4 waveplate and a beam splitting polarizer together with a set of balanced photodiodes, the THz pulse amplitude is mapped by monitoring the gate pulse polarization rotation after the ZnTe crystal at various delay times with respect to the THz pulse (Figure 2(a)). A Fourier transform is then used to convert this time domain electrical signal to a frequency domain spectrum (Figure 2(b)). The optical path from THz generation to THz detection is purged with dry nitrogen gas to avoid the effect of the humidity.
Dielectric relaxation measurement is a sound method to investigate intermolecular interactions and is capable of monitoring cooperative processes at the molecular level [78, 79, 80]. This method is appropriate to monitor molecular motions in widely varied time scales and has been used extensively to study the structure, dynamics, and macroscopic behavior of complex systems [81]. A brief discussion (more details may be found elsewhere [81]) on the basic principle of dielectric polarization is presented below.
(a) Polarization of dielectric. (b) Polarization vector.
If a static electric field of strength,
where ε0 is the dielectric permittivity of the vacuum. Applying the macroscopic Maxwell theory, the electric displacement (electric induction) vector,
Now, in the linear regime, the relative dielectric permittivity (or
There are different types of polarization mechanisms by an applied electric field in different frequency regions: i)
Therefore,
For
The orientation polarization is given by the dipole density due to the dipoles
where
where
where
where
Eq. (17) gives equivalent information on dielectric relaxation properties of the sample both in frequency domain and time domain measurements. Therefore, the dielectric response might be measured experimentally as a function of either frequency or time, providing data in the form of a dielectric spectrum ε*(ω) or the macroscopic relaxation function
Where, τm indicates the relaxation time. Eqs. (17) and (19) give rise to the following relation,
i.e,
This is known as the Debye model for frequency dependent dielectric permittivity. According to the Debye model, the complex frequency dependent dielectric response [
where,
The dynamics of water around small as well as complex molecules changes owing to their specific interaction with the solute surface; the specific nature of the interaction could mostly be electrostatic or hydrogen bonding. Such interaction expectedly ruptures/modifies the tetrahedral water network structure as well as its dynamics, a clear imprint of that gets reflected in the THz frequency window. Measurement of optical parameters α(ν) and n(ν) enables one to determine the change in water dynamics at the vicinity of (bio)surface and consequently one can infer on the changes of the corresponding solute also.
Metal ions are perhaps the simplest solutes that can induce perturbation in water dynamics, the interaction being mostly electrostatic in nature. A systematic investigation using alkali monovalent cations [85] and alkaline earth bivalent cations [86] in aqueous solutions have been put forward by the group of Martina Havenith using far-IR and THz FTIR measurements coupled with classical MD simulation studies. Their study concluded that ion rattling motions can account for the observed changes in the THz absorption. They have shown that the spectrum of the salt solutions can be approximated by a linear superposition of concentration weighted neat water and ion contributions. Ab initio MD simulation study by Marx et al. [87] has successfully reproduced the spectral responses of the solvation shell around the ions in infrared and THz frequency range. Their study has shown that the solute-solvent dipolar couplings and the dipole–dipole correlations are the important factors that govern the absorption features in this frequency regions. The group of Martina Havenith has subsequently put forward a series of studies on the hydration dynamics of heavy metal ions, e.g. lithium [88], manganese [89], iron [90] and ytterbium [91]. An elaborate description of the process involved has recently been put forward by this group [92]. The overall THz effect of the ions could be apprehended in terms of the contributions from various modes as depicted in Figure 4.
Schematic representation of ion hydration. (a): Bulk-like water (blue), water around ions (yellow and green), and hydration water (lighter shades of yellow and green). The total absorption of the solution (αsol) has contribution from all. (b): Concentration-weighted bulk-water values are subtracted from αsol. (c): This yields the effective absorption of the solvated ion or ion pair (αeffion).
In a systematic study using THz-TDS in the frequency region (0.3–2.1 THz; 10–70 cm−1), experimental evidences for the ultrafast collective hydrogen bond dynamics of water in the extended hydration layers of alkali metal chlorides have been obtained [93]. The real and imaginary part of the permittivity (ε), as obtained from the THz-TDS measurements was fitted using a triple Debye relaxation model. The time scales obtained for bulk water are of the order of ∼8–9 ps (τ1), 200 fs (τ2) and 80 fs (τ3). The ∼9 ps and ∼ 200 fs timescales are due to the well-known cooperative rearrangement of the H-bonded network structure and the small angular rotational modes of individual polar water molecules, respectively. It has been reported that (τ1) decreases with increasing salt concentration, which identifies an acceleration of the cooperative hydrogen bond dynamics affirming a positive support towards the most debated notion of these ions to act as water structure breakers (see Figure 5(a)). The extent of this effect has been found to be mostly ion specific, K+ being the most effective ion and a simple consideration of ionic charge density is insufficient to account for the observed changes. Very recently, Havenith and Marx [94] have put forward a combined experimental and simulation investigation to provide a detailed mechanistic analyses based on cross-correlation analysis (CCA) technique to understand the minute details of water-solute interactions.
(a) Cooperative relaxation dynamics (τ1) and relaxation strength (S1) of aqueous solutions of different alkaline metal cations. (b) Relaxation dynamics (as defined by the timescales τ1 and τ2 of aqueous urea solution as a function of urea concentration. A sharp change in the τ1 is observed at ∼4 M urea. (c) Cooperative hydrogen bond relaxation time constant (τ2) of aqueous solutions of amino acids as a function of their concentration. The dotted line is the time constant for the buffer solution. The inset shows the change in τ2 (measured at maximum amino acid concentration) as a function of the SASA of amino acid. (d) Relaxation time scale τ3 corresponding to the jump orientation of water (obtained by Debye fitting of THz data) of different amino acids solutions. The dotted line is the time constant for that of the buffer. The inset shows the change in the timescale (Δτ3 = τsolution-τbuffer) at the maximum concentration of the amino acids.
While the interaction of metal ions with water is more straight-forward, complexity arises when an ion is associated with a hydrophobic moiety. Such molecules are often of biological importance and therefore their interaction with water is essential to establish. There have been several reports on the THz studies of sucrose and saccharides [95, 96, 97, 98, 99]. In a pioneering work Heyden et al. [100] have described the nature of hydration around small carbohydrate molecules and a detailed simulation study has revealed that the extent of the hydration shell and their absorption co-efficient change with the solute specificity. One such molecule is urea, which is well known to be a protein denaturant, however, the exact molecular mechanism of the processes involved still remains a strongly debatable issue, specially the argument of whether the interaction is water mediated (the water structure breakers notion of urea) or a direct interaction of urea with protein surface.
An elaborate attempt has been made to understand the effect of urea (and its derivatives) on the ultrafast solvation dynamics of water using a Debye type dielectric relaxation model in the 0.3–2.0 THz frequency region using THz-TDS technique [101]. The relaxation dynamics shows considerable acceleration beyond a threshold concentration. Such acceleration is possibly associated with a disruption of the tetrahedral water network structure. It seems also intriguing that the observed collapse occurs at a certain urea concentration (see Figure 5(b)), which strikingly coincides with the denaturation concentration of urea for many proteins. Another molecule of such biological interest is guanidinium chloride (GdmCl), which offers remarkable protein denaturation ability beyond a certain threshold concentration (∼2–3 M) [102, 103]. Like in the case of urea, protein denaturation mechanism of GdmCl has also been debatable concerning a direct or indirect interaction. We have investigated the collective hydrogen bond dynamics around GdmCl in aqueous solutions as well as in the presence of a model globular protein human serum albumin (HSA) using the THz-TDS technique. It is found that the relaxation dynamics gets faster which renders support to the previously speculated notion that GdmCl acts as a water structure breaker. A similar but more prominent trend is observed in case of NaCl, which, however, does not interact with proteins.
The change in hydration dynamics in presence of HSA has been found to be in complete contrast in these two salts unambiguously pointing out towards a possibility of the collective hydration dynamics to share a pivotal contribution in the protein unfolding phenomenon. These studies seem to conjecture that ions with complex hydrophobic moiety acts as water structure breaker, and this very nature of such molecules makes them protein denaturant. To investigate the validity of such conjecture we investigated the ultrafast (sub-ps to ps) collective hydrogen bond dynamics of water in the extended hydration layers in a series of alkylammonium chloride salts using THz -TDS technique [104]. We found these salts to transform from being a water ‘structure breaker’ to ‘structure maker’ with increasing carbon content. For example, the THz-TDS measurements reveal that ammonium chloride (AC) acts as a water network structure breaker while tri-ethyl ammonium chloride (EAC) and higher carbon containing salts are distinct water structure makers. The change in protein hydration is also found to trace its secondary structure rupture and the exposure of hydrophobic moieties accordingly changes the protein hydration. Our study strongly concludes that it is the hydrophobic effect, at least in the case of this type of salts, that plays the decisive role in determining their interaction with biomolecules.
Amino acids, which are the building blocks of proteins, often act as intermediates in metabolism as well as osmolytes that can stabilize proteins. Depending upon the ‘side group’, the amino acids are classified as hydrophilic or hydrophobic. Hydrophobicity of amino acids is believed to be a key parameter that regulates phenomena like protein folding - unfolding, aggregation, activity, protein-ligand binding and protein hydration in aqueous environments. While to analyze the solvation of hydrophobic and hydrophilic parts of a protein separately, one needs to take into consideration that the environment of amino acid residues is heterogeneous in nature as they are often composed of hydrophobic alkyl chains and hydrophilic groups. It is therefore essential to study the hydration of amino acids of various side chains in the exposure to solvents. Niehues et al. [105] have studied the hydration of a series of amino acids in ∼2.4 THz window and observed that the THz absorption coefficient, α(νTHz), of hydrated amino acids can be correlated with the hydrophobicity and the fraction of polar volume of amino acids. They have shown that glycine has the largest positive THz slope followed by serine, whereas for the other amino acids, the slope becomes gradually negative with increasing hydrophobicity. In another study [106], the same group, by analyzing the corresponding THz spectrum in terms of the correlated dynamics of solute and solvent molecules, demonstrates the line shape of the low-frequency vibrational response of glycine in water. Recently, they [107] have shown that hydrophilic solvation of the zwitterionic groups in valine and glycine leads to similar THz responses, which are fully decoupled from the side chain. This result concludes that the hydrophilic groups and their solvation shells dominate the THz absorption difference, while on the same intensity scale, the influence of hydrophobic water can be neglected. Shiraga et al. [108] concludes a correlation between the extent of H-bonding and the hydrophobicity of the solute. Glycine (Gly) and the L- isomers of five different amino acids: serine (Ser), aspartic acid (Asp), lysine (Lys), arginine (Arg) and tryptophan (Trp) of varying hydrophobicity and solvent accessible surface area (SASA) have been used by Samanta et al. [109] to study the dielectric relaxation up to their maximum water solubility in the frequency window of GHz-THz at neutral pH. The various rotational dynamics of the solutes and water are obtained by fitting the dielectric data in a multiple Debye relaxation model. From GHz study, we observed that the molecular rotation of amino acids correlates their respective molecular volume. Also, it was concluded that the amino acids do not usually aggregate even at high concentrations. From THz study, the authors found that Gly is a water structure breaker while the other amino acids are structure makers. Consequently, Gly accelerates cooperative hydration, while the others retard (see Figure 5(c) and (d)). It has been established that hydration in amino acids depends both on its hydrophobic as well as its hydrophilic nature of side chains. Born et al. [110] have studied the hydration dynamics of some small model peptides using THz spectroscopy. They observed that the cooperative hydration dynamics changes as the extent of hydration of such molecules change.
Addition of otherwise indifferent organic solvents can influence marked changes in aqueous environments as well as they can induce misfunction in biologically important molecules e.g. protein, DNA, etc. Water - dimethyl sulfoxide (DMSO) binary mixture has been found to be of potential interest as it plays a significant role in the field of chemistry, physics, biology and pharmacology. Das Mahanta et al. have investigated the change in the collective hydration dynamics in presence of DMSO at different concentrations using THz-TDS. We found that α(ν) of the mixed solvents shows a non-linear change with XDMSO [111]. This change has been correlated with the water-DMSO structural heterogeneity in the mixed solvents. Luong et al. have also studied the evolution of water hydrogen bonded collective network dynamics in water −1,4-dioxane (Dx) mixtures as the mole fraction of water (Xw) increases from 0.005 to 0.54 [112]. The inter- and intra-molecular vibrations of water be observed using THz-TDS in the frequency range 0.4–1.4 THz (13–47 cm−1) and Fourier transform infrared (FTIR) spectroscopy in the far-infrared (30–650 cm−1) regions. From the absorption coefficient measurements, they infer that the mixtures are not ideal in nature, which suggests a significant change in the network by the addition of the solute. The authors found an increase in the collective hydrogen bond network as evidenced from dielectric relaxation studies in which τ1 has been found to be small at low Xw (where Xw is the mole fraction of water in the mixture), but at Xw > 0.1, it increases rapidly to reach a value identical to that in bulk water. It has been concluded that hetero-molecular (water - Dx) hydrogen bond dominates in the water diluted region in water-Dx mixtures, and with progressive addition of water, bulk-like intermolecular three-dimensional hydrogen bonded water network dynamics evolves beyond Xw = 0.1. In a separate study [113], a combined experimental (mid- and far-infrared FTIR spectroscopy, THz-TDS (0.3–1.6 THz)) and molecular dynamics (MD) simulation technique has been carried out to understand the evolution of the structure and dynamics of water in its binary mixture with 1,2-dimethoxy ethane (DME) over the entire concentration range. Debye relaxation data reveals a non-monotonous behavior in which the collective dynamics is much faster in the low Xw region, whereas in the Xw ∼ 0.8 region, the dynamics gets slower than that of pure water. The concentration dependence of the reorientation times of water, estimated from the MD simulations, also captures this non-monotonous character. Bohm et al. [114] have demonstrated that the THz response of alcohols can be decomposed into the spectrum of bulk water, tetrahedral hydration water, and more disordered (or interstitial) hydration water. They also concluded that it is not the tetrahedrally ordered component, rather it is the interstitial hydration water which is responsible for the temperature-dependent change in ΔCp and ΔG in such mixtures.
The surface of proteins is extremely heterogeneous owing to the presence of amino acids of varying types of charges. The pioneering studies by the group of Havenith et al. have shown that protein molecules are hydrated and the cooperative dynamics of water changes accordingly. An earlier study using a small protein ubiquitin [115] shows how the measurement of α(νTHz) reveals a change in the protein hydration dynamics as the authors termed it as the “THz dance”. In a subsequent ever important simulation paper, the same group has established different THz absorption of the hydrophobic and hydrophilic residues of a protein as they interact differently with water [116]. In the later years, this group has put forward substantial contribution on the hydration dynamics of proteins using THz measurements [117, 118] as well as simulation studies [42, 119]. Their studies have unambiguously suggested that fast protein motion and solvent dynamics are correlated with enzymatic reactions [120]. In a recent study this group has established the pivotal role of collective motion of water during an electron transfer reaction between flavoenzyme ferredoxin - NADP+ − reductase and ferredoxin-1 [121]. In a seminal paper, Marklez et al. [122] have used THz-TDS measurements to show that the protein (hen egg white lysozyme) dynamical transition (the rapid increase in protein dynamics occurring at ∼200 K) needs neither tertiary nor secondary structure. Their results revealed that the temperature dependency essentially arises from the protein side-chain interaction with the solvent. He et al. [123] have investigated the presence of structural collective motions on a picosecond timescale for the heme protein, cytochrome C, as a function of oxidation and hydration, using THz-TDS and molecular dynamics simulations. Marklez group has developed a novel measurement technique (anisotropy THz microscopy) [124] wherein they were able to detect the long range protein vibration modes in chicken egg white lysozyme single crystals. They found the underdamped modes to exist for frequencies >10 cm−1. Such underdamped vibrational modes have also been identified using optical Kerr effect measurements in the THz frequency window [125]. In a recent study, Niessen et al. [126] have used anisotropy THz microscopy, which is found to be sensitive to inhibitor binding and unique vibrational spectra for several proteins and an RNA G-quadruplex. There have been other reports from several experimental groups: Sun et al. [127] have reported the application of a new machine learning methods for quantitative characterization of bovine serum albumin (BSA) deposited thin-films detected by THz-TDS. The group of Emma Pickwell-MacPherson [128] have used THz spectroscopy to study the hydration shell formation around H9 subtype influenza A virus’s HA protein (H9 HA). They have also detected antigen binding of H9 HA with the broadly neutralizing monoclonal antibody. They observed a remarkable concentration dependent nonlinear response of the H9 HA, which reveals the formation process of the hydration shell around H9 HA molecules. The same group has also reported the dielectric properties of two different antibodies in water-glycerol mixtures using THz-TDS measurements [129]. Sun et al. [130] used THZ-TDS to investigate the molecular processes involved above and below the transition temperature (TD) for GP2 peptide.
While much research has been focused on proteins, relatively less attention has been paid on the other biologically important molecule, DNA. There had been a few preliminary studies to understand the vibronic bands in the THz frequency region using single and double stranded DNAs and RNAs [131, 132, 133]. Arora et al. [134] have presented a label free quantitative detection method for DNA samples amplified by polymerase chain reaction (PCR) in aqueous medium using THz-TDS in the frequency range from 0.3 to 1.2 THz. Tang et al. [135] have recently investigated the feasibility of THz spectroscopy combined with microstructures for marker-free detection of DNA and oligonucleotides. Polley et al. have investigated the collective dynamics of two DNA molecules extracted from salmon sperm and calf thymus and observed that the dynamics did not differ much at the concentration range of the experiments [136].
There have been only limited studies on the THz studies on lipid membranes and/or vesicles. One of the preliminary results was due to Tielrooij et al. [137] who had studied the dielectric relaxation in mixed system of hydrated DOPC lipid bilayers in the THz frequency domain. They could identify three distinct water types: fast, bulk and irrotational. The relative content of those change with the extent of hydration. Later, Yamamoto et al. [138] have studied the temperature and hydration dependent low frequency spectra of lipid bilayers of 1,2-dimyristoyl-sn-glycero-3-phosphoryl-3′- rac-glycerol (DMPG) and 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) using THz-TDS. They found that the THz absorption patterns reflect the lipid packing pattern in the bilayers. They subsequently extended their investigations towards purple membrane (PM, a complex of lipids and a membrane protein, bacteriorhodopsin) [139] and lipid bilayer of DMPC [140]. Pal et al. have recently studied the microstructure and collective dynamics of the membrane interfacial hydration shell in zwitterionic and negatively charged phospholipid membrane bilayers using THz-TDS [141]. They observed a dependence of the critical lipid concentration corresponding to the inflection point on the charge of the lipid head-group, thereby implicating membrane electrostatics as a major factor in the microstructure and dynamics of water at the membrane interface.
Hydration dynamics of biomolecules is significantly influenced upon changing its physical conditions as well as in the presence of additional chemical agents, like alcohols, glycols, etc. Protein molecules undergo various physical and chemical changes, which could induce conformational modifications in their secondary as well as tertiary structures. It can be noted here that as protein structures get disrupted (during unfolding or denaturation) the hydrophobic moieties (amino acid residues), which are otherwise buried inside in the native structure, get exposed, and this produces a definite alteration in THz absorption coefficient, α(νTHz). It therefore suggests that estimation of α(νTHz) provides a direct evidence of the structural perturbation in proteins.
Several experimental techniques are available to determine the structural evolution of proteins during unfolding, while THz provides with the estimation of the associated hydration changes. In a pioneering experimental work, in which a stopped-flow techniques was synchronized with THz-TDS, which the authors termed as Kinetic terahertz absorption (KITA) spectroscopy, Kim et al. [142] have shown that the pH induced unfolding-refolding kinetics (in real time) of Ub* could easily be traced by the associated α(νTHz) measurements. The THz results very well reconcile with the results obtained from circular dichroism (CD) and fluorescence measurements.
Another important physical environment that induces protein unfolding is temperature. In a study using HSA as a model protein, Mitra and Havenith [143] have shown that water dynamics associated with the protein during its reversible unfolding pathway up to 55°C as well as its irreversible denaturation pathway up to 70°C traces the protein’s structural rupture pathway. The THz measurements do support the conventional CD and fluorescence measurements. Sudden increase in the environment (like temperature or pressure) for a very short period of time (often termed as T-jump experiments) leads protein molecules to be structurally ruptured but upon removal of the intense pulse, the protein refolds. T-jump experiments have previously been characterized using conventional CD and fluorescence measurements. However, THz measurements was demanded to obtain explicit information of water dynamics. The first report of such experiment was from the group of Havenith [144], wherein the authors put forward a coupled KITA setup with a T-jump attachment. The authors monitored changes in the THz absorption λ∗6 − 85protein with a time resolution of >50 μs. They reported that the spectral changes are correlated with the hydrophobic collapse of the protein. In a subsequent study from the same group, Wirtz et al. [145] used an even better time resolution of about 500 ns to reveal the coupled ubiquitin−solvent dynamics in the initial phase of hydrophobic collapse (temperature induced unfolding). They propose that, in the case of ubiquitin, a rapid (∼500 ns) initial phase of the hydrophobic collapse from the elongated protein to a molten globule structure precedes secondary structure formation. Recently there have been a few reports of using THz spectroscopy technique to underline thermal denaturation of BSA [146], temperature- and pH-dependent protein conformational changes in pepsin A [147]. In a very recent study Cao et al. [148] have successfully employed THz-TDS to track the hydrolysis of BSA protein by pepsin. The results indicate that protein hydrolysis can be easily monitored over time by focusing on the variation of the absorption coefficient from a macroscopic perspective. The authors explored the use of the Debye model to analyze the dielectric properties of the solution during protein hydrolysis. The results of the Debye analysis prove that it is possible to investigate in detail the microscopic dynamics of bio-macromolecule solutions at the molecular level by THz-TDS.
Samanta et al. have been involved in determining the changes in protein hydration in various distressed environments using the THz-TDS measurements. They have investigated the hydration dynamics around HSA in presence of short chain polyethylene glycols of different chain lengths (PEG 200, PEG 400, and PEG 10000) at different concentrations [149]. FIR-FTIR studies conclude that the protein hydration is affected in a distinct way below and above the critical PEG concentration of 30% (v/v). THz-TDS study unambiguously confirmed a retardation of the solvation dynamics by PEGs. This study clearly shows an independent behavior of protein hydration at low PEG concentrations and a noticeable interaction between protein and PEG hydration beyond a critical PEG concentration. In another study, Das et al. have made an attempt to understand whether the DMSO induced conformation changes in lysozyme conformation perturbs its hydration dynamics [150]. CD study establishes a marked change in the protein tertiary structure in presence of DMSO. The relative change in the THz absorption coefficient (Δα/α0) shows a negative minimum at XDMSO = 0.05 and a positive value at XDMSO = 0.15. The observed minimum is found to be due to the increased size of the protein while the positive value is attributed to the increased SASA and consequent increased hydration of the protein surface. In a recent report, Das et al. put forward an experimental observation of nonmonotonic changes in the collective hydration of BSA in the presence of alcohols of varying carbon-chain lengths, that is, ethanol, 2-propanol, and tert-butyl alcohol (TBA), by using THz – TDS [151]. They observe an anomalous hydration behavior of the protein hydration with the alcohol concentration, which correlates the alcohol-induced α-helix to random coil transition of the protein secondary structure, as revealed by CD spectroscopy measurements. Recently, Das Mahanta have investigated the effect of alkyl-ammonium chloride salts on BSA and found a systematic trend towards disrupting the protein secondary structure [104]. The associated changes in the protein hydration in the presence of these salts have also been investigated using THz-TDS. The change in protein hydration is also found to trace its secondary structure rupture, and the exposure of hydrophobic moieties accordingly change the protein hydration. The THz-TDS measurements strongly conclude that it is the hydrophobic effect, at least in the case of this type of salts, that plays the decisive role in determining their interaction with biomolecules.
Aggregated protein is toxic to functioning of living systems and many of human diseases are associated with misfolded protein disorders [152, 153, 154, 155, 156]. This is why understanding of mechanisms of interactions between protein molecules in solutions have been the subject of extensive investigations during the last two decades [157, 158, 159, 160, 161]. Additionally, understanding the protein aggregation propensity may offer novel design principles for producing aggregation-resistant proteins for biotherapeutics.
Globular proteins, e.g. HSA and BSA, in their native states are present in living cells at concentrations as high as about 200 mg mL−1 and bimolecular interactions are significant. We explained earlier that stability of the three - dimensional structure of a monomeric protein molecule in physiological environments is the result of an intricate interplay between electrostatic, hydrophobic, hydrogen bonding and other interactions and any variation of temperature, pH of the medium or any other physiochemical conditions including concentration of protein in the cell may result in imbalance of the stabilization forces leading to misfolding [162, 163], thus triggering aggregation [164, 165, 166, 167, 168].
A native and structurally stable protein molecule is strongly hydrated with a well-defined hydration layer of thickness of about a few tens of Å (20–40 Å) around it [50, 59, 169]. The role of water molecules in the hydration shell could be crucial for the intermolecular interactions and the overall protein hydrophobicity, which may be defined by its hydration free energy, which may play an important role in protein aggregation in aqueous solution [162, 163, 166]. However, the role of hydration water in protein aggregation has largely been unexplored owing to the perception that protein – protein interaction is the major factor and the surrounding water is just a spectator playing no role in aggregation of protein, ignoring water as an active constituent of biological systems. In fact, whether a protein remains soluble or forms aggregation should intrinsically rely on its state of hydration in the monomeric state.
We have described in the earlier sections of this article that how the properties of water molecules in the hydration layer on an average are different from those in the bulk, which determines the stability of a monomer protein in aqueous environment. Protein aggregation proceeds through a multistep process initiated by conformational transitions, called protein misfolding, of monomer species towards aggregation-prone structures. Chong and Ham applied the fluctuating thermodynamic analysis method to understand the variations of the thermodynamic functions, which occur during the course of misfolding and dimerization of the Amylase-β protein. They suggest that the time variation of the solvent-averaged effective energy, F = Eu + Gsolv, describes the protein dynamics on the free energy landscape [170]. Here, the protein potential energy (Eu), comprises both intra and intermonomer contributions and the solvation free energy (Gsolv), represents the interaction of the protein with surrounding water, which plays a critical role in protein aggregation. The free energy, F, decreases as the dimerization proceeds, but the decrease in F has different origins in the approach and structural adjustment regimes. The thermodynamic force driving the approach of two monomers is the decrease in Gsolv and hence, the misfolded monomers acquire a large hydrophobicity, which leads to conformational changes in monomers. This drives two monomers to approach each other to a contact distance. In absence of this thermodynamic driving forces, two negatively charged protein monomers (the total charge of Amylase-β and BSA monomer proteins at neutral pH are −3, and − 16, respectively [170, 171]) would never approach each other by overcoming the electrostatic repulsion. On the other hand, decrease of the protein potential energy Eu, due to direct protein–protein interactions, such as intermonomer van der Waals contacts and hydrogen bonds, drives the structural rearrangement required for formation of compact dimer structure leading to energetic stabilization. Interestingly, structural rearrangements are also associated with an increase in the solvation free energy, which originates from the dehydration of the protein surface and of the interfacial region. On contrast to other spectroscopic techniques, THz spectroscopy probes directly the collective intermolecular vibrations of the hydrogen bond network, and is thus able to detect sensitively solute induced changes in the solvation dynamics. Extensive works on THz absorption of protein solutions have demonstrated that the absorption coefficient of the protein solutions (αsol) are dependent on the concentration of protein [50, 59, 169, 170, 171]. For example, at the low concentration regime (e.g. in the case of HSA, <0.5 x 10−3 mol dm−3) αsol value increases linearly and this has been explained by increasing concentration of hydrated monomer protein molecules since water in the hydration shell has larger αsol value as compared to that of bulk water. However, on further increase of the protein concentration, αsol value starts decreasing.
To delineate this issue in more detail, Manna et al. made a detailed investigation on the concentration dependence of THz absorption of the aqueous buffered solutions of HSA protein (up to 2.6 mM of protein concentration) at three THz frequencies, namely, 0.1, 1.5 and 2 THz [59]. Similar results were obtained from these three measurements. αsol value was expected to change linearly to follow Eqs. (23) and (24), which could be derived assuming that the protein solution is a two-component system.
Here, αpr is the absorption coefficient of the protein, αbw is the absorption coefficient of bulk water,
Therefore, the plots of
(a): Plots of
To understand the reasons for significant decrease of THz absorption coefficient of the protein solutions with increasing concentration beyond 6 x 10−4 mol dm−3, the possibility of aggregation of proteins at higher concentration regime was explored. To delineate this aspect, dynamic light scattering (DLS) measurements (Figure 6) were carried out using concentrations of proteins covering the entire range of THz absorption measurements. The DLS data recorded for the solution containing protein concentration of 0.4 x 10−3 mol dm−3 revealed the existence of only monomeric protein molecules with the most probable diameter of about 6–8 nm in the solution. This is quite in good agreement with the diameter of the hydrated monomer protein molecules. However, at higher concentrations (say, >1 x10−3 mol dm−3 of protein), the DLS data revealed two important features. Firstly, the size distribution of the monomer band became wider indicating the presence of particles of diameter in the range 12–15 nm, possibly suggesting formation of dimers or trimers of HSA, in addition to the monomeric species. Secondly, at higher concentrations of the protein, DLS data also revealed the presence of large size aggregates of the most probable diameter of about 700 nm. However, a quantitative estimation of the relative percentages of monomer, dimer and aggregates was not possible from the DLS data because the intensity distribution of the scattered radiation was not directly proportional to the number of the particles. However, this experiment confirmed the presence of protein aggregates in solutions with higher concentrations of protein and the formation of aggregates may possibly be held responsible for nonlinear dependence of THz.
On the other hand, CD measurements confirmed that the tertiary structure of HSA remained unchanged through the entire range of HSA concentrations used for THz measurements. This suggested that the native structure of HSA molecules remained unaltered through the entire range of concentrations of HSA. Therefore, hydration states of proteins, even in the aggregated state, remain unchanged and hence possibly justifies the assumption made in the earlier works regarding overlapping of hydration shells at higher concentrations of proteins.
Patro and Przybycien have simulated the structures of reversible protein aggregates as a function of protein surface characteristics, protein–protein interaction energies and assessed the aggregate properties [174]. Results of their simulation reveals that aggregate particles have the kind of organization of the hydrophobic and hydrophilic domains as they are present in HSA protein monomer molecules and aggregation of protein molecules causes the loss in the total solvent accessible surface area (SAS) is about 67% and the mean solvent content for these aggregates vary in the range of 0.37–0.55 volume fraction depending on the conformation of the monomer protein [175]. Therefore, at higher concentrations of protein, volume fraction of hydration water decreases due to formation of aggregates. In addition, proteins are THz transparent and as we increase the protein concentration, protein aggregates replace the water molecules and leads to lowering of total THz absorbance of the solution.
A method of analysis was adopted to analytically fit the data in the regime of higher concentrations of the protein to predict the relative concentrations of the monomer and aggregated particles. In this analysis, the value of αhl, which was estimated from the linear regime of the plot of
Here, x is the concentration of monomer in the unit of 1 × 10−3 mol dm−3 and
Manna et al. estimated the percentage of the number of HSA molecules, which exist as monomer (or dimer or trimer) in solution without being associated with aggregate formation, for each of the HSA concentrations used for THz absorption measurements [59]. We find that for the protein solution with its concentration of 8 x 10−4 mol dm−3, about 40% of the protein molecules exists as monomer (or dimer or trimer) in solution and 60% protein molecules are part of aggregates. The monomer (or dimer or trimer) concentration becomes <1% at the protein concentration of 2.6 x 10−3 mol dm−3, i.e. nearly all the protein molecules are associated with aggregate formation. These calculations suggest that formation of aggregates at higher concentrations of protein may be the responsible factor for the turnover of the THZ absorbance of the protein solution at ∼6 x 10−4 mol dm−3 concentration.
The absorption coefficient of a complex system is the weighted sum of the absorption of its components. However, determination of the complex dielectric function of proteins in solution using THZ-TDS spectroscopy allowed detailed analysis beyond what was possible from simple absorption measurements. Following this approach, Novelli et al. determined both the phase and amplitude of the induced dipole in the volume containing the protein and hydration water [170]. This result revealed that not only the amplitude of the induced dipole varied with the concentration of protein, but also its phase changed above a concentration threshold (Figure 7). They proposed a phenomenological model, which explained that the phase of the induced dipole in the protein-solvent interaction region began to vary when there was significant overlap between hydration layers of the neighboring protein. This result suggested that indirect electromagnetic protein–protein interactions could take place if mediated by the extended hydration layers surrounding each protein.
(a) Cartoon of a human lysozyme protein (red sphere) in water. Water molecules tightly-bound to the protein surface (white), extended hydration layers (blue) and unperturbed bulk water (light blue). (b) Sketch of the evolution of the modulus and the phase of the induced dipole in a unit volume of solution versus protein concentration. The effect of phase gain at larger concentrations is represented by darker colors on the bottom right panel (adopted with permission from Ref. [
THz spectroscopy, from its very inception, has mostly been used by scientists studying cosmology, condensed matter physics and materials. The huge absorption of water in this frequency range used to be treated as a drawback of this technique; however, for chemists and biologists this point serves as an advantage since all the biological function is somehow or other related to water dynamics. Due to its inherent sensitivity to water hydrogen bonding dynamics, THz spectroscopy has become an indispensable tool for direct observation of fast and coupled biomolecule-water network. The initial studies by the groups of E. W. Heilweil, P.U. Jepsen, A. Marklez, C. Schmuttenmaer and M. Havenith in the late 1990s and early 2000s have established THz spectroscopy at a concrete platform to be recognized as a potential tool to label free detection of water dynamics in the vicinity of biomolecules, the effect being extended to several layers and remains practically undetected using conventional spectroscopic methods. The last decade has witnessed a huge leap towards exploiting this frequency window in biophysical studies, a few of such results have been depicted in this article. Now that the phenomenon has been established beyond any doubt, new sort of experimental studies, where the role of hydration in ultrafast processes (like electron transfer or proton transfer) could explicitly be determined, is the new challenge to the researchers.
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\\n\\nAs a firm believer in the wider dissemination of knowledge, IntechOpen supports the Open Access Initiative Protocol for Metadata Harvesting (OAI-PMH Version 2.0). Read more
\\n\\nLicense
\\n\\nBook chapters published in edited volumes are distributed under the Creative Commons Attribution 3.0 Unported License (CC BY 3.0). IntechOpen upholds a very flexible Copyright Policy. There is no copyright transfer to the publisher and Authors retain exclusive copyright to their work. All Monographs/Compacts are distributed under the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0). Read more
\\n\\nPeer Review Policies
\\n\\nAll scientific works are Peer Reviewed prior to publishing. Read more
\\n\\nOA Publishing Fees
\\n\\nThe Open Access publishing model employed by IntechOpen eliminates subscription charges and pay-per-view fees, enabling readers to access research at no cost. In order to sustain operations and keep our publications freely accessible we levy an Open Access Publishing Fee for manuscripts, which helps us cover the costs of editorial work and the production of books. Read more
\\n\\nDigital Archiving Policy
\\n\\nIntechOpen is committed to ensuring the long-term preservation and the availability of all scholarly research we publish. We employ a variety of means to enable us to deliver on our commitments to the scientific community. Apart from preservation by the Croatian National Library (for publications prior to April 18, 2018) and the British Library (for publications after April 18, 2018), our entire catalogue is preserved in the CLOCKSS archive.
\\n\\nOpen Science is transparent and accessible knowledge that is shared and developed through collaborative networks.
\\n\\nOpen Science is about increased rigour, accountability, and reproducibility for research. It is based on the principles of inclusion, fairness, equity, and sharing, and ultimately seeks to change the way research is done, who is involved and how it is valued. It aims to make research more open to participation, review/refutation, improvement and (re)use for the world to benefit.
\\n\\nOpen Science refers to doing traditional science with more transparency involved at various stages, for example by openly sharing code and data. It implies a growing set of practices - within different disciplines - aiming at:
\\n\\nWe aim at improving the quality and availability of scholarly communication by promoting and practicing:
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The Open Access publishing movement started in the early 2000s when academic leaders from around the world participated in the formation of the Budapest Initiative. They developed recommendations for an Open Access publishing process, “which has worked for the past decade to provide the public with unrestricted, free access to scholarly research—much of which is publicly funded. Making the research publicly available to everyone—free of charge and without most copyright and licensing restrictions—will accelerate scientific research efforts and allow authors to reach a larger number of readers” (reference: http://www.budapestopenaccessinitiative.org)
\n\nIntechOpen’s co-founders, both scientists themselves, created the company while undertaking research in robotics at Vienna University. Their goal was to spread research freely “for scientists, by scientists’ to the rest of the world via the Open Access publishing model. The company soon became a signatory of the Budapest Initiative, which currently has more than 1000 supporting organizations worldwide, ranging from universities to funders.
\n\nAt IntechOpen today, we are still as committed to working with organizations and people who care about scientific discovery, to putting the academic needs of the scientific community first, and to providing an Open Access environment where scientists can maximize their contribution to scientific advancement. By opening up access to the world’s scientific research articles and book chapters, we aim to facilitate greater opportunity for collaboration, scientific discovery and progress. We subscribe wholeheartedly to the Open Access definition:
\n\n“By “open access” to [peer-reviewed research literature], we mean its free availability on the public internet, permitting any users to read, download, copy, distribute, print, search, or link to the full texts of these articles, crawl them for indexing, pass them as data to software, or use them for any other lawful purpose, without financial, legal, or technical barriers other than those inseparable from gaining access to the internet itself. The only constraint on reproduction and distribution, and the only role for copyright in this domain, should be to give authors control over the integrity of their work and the right to be properly acknowledged and cited” (reference: http://www.budapestopenaccessinitiative.org)
\n\nOAI-PMH
\n\nAs a firm believer in the wider dissemination of knowledge, IntechOpen supports the Open Access Initiative Protocol for Metadata Harvesting (OAI-PMH Version 2.0). Read more
\n\nLicense
\n\nBook chapters published in edited volumes are distributed under the Creative Commons Attribution 3.0 Unported License (CC BY 3.0). IntechOpen upholds a very flexible Copyright Policy. There is no copyright transfer to the publisher and Authors retain exclusive copyright to their work. All Monographs/Compacts are distributed under the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0). Read more
\n\nPeer Review Policies
\n\nAll scientific works are Peer Reviewed prior to publishing. Read more
\n\nOA Publishing Fees
\n\nThe Open Access publishing model employed by IntechOpen eliminates subscription charges and pay-per-view fees, enabling readers to access research at no cost. In order to sustain operations and keep our publications freely accessible we levy an Open Access Publishing Fee for manuscripts, which helps us cover the costs of editorial work and the production of books. Read more
\n\nDigital Archiving Policy
\n\nIntechOpen is committed to ensuring the long-term preservation and the availability of all scholarly research we publish. We employ a variety of means to enable us to deliver on our commitments to the scientific community. Apart from preservation by the Croatian National Library (for publications prior to April 18, 2018) and the British Library (for publications after April 18, 2018), our entire catalogue is preserved in the CLOCKSS archive.
\n\nOpen Science is transparent and accessible knowledge that is shared and developed through collaborative networks.
\n\nOpen Science is about increased rigour, accountability, and reproducibility for research. It is based on the principles of inclusion, fairness, equity, and sharing, and ultimately seeks to change the way research is done, who is involved and how it is valued. It aims to make research more open to participation, review/refutation, improvement and (re)use for the world to benefit.
\n\nOpen Science refers to doing traditional science with more transparency involved at various stages, for example by openly sharing code and data. It implies a growing set of practices - within different disciplines - aiming at:
\n\nWe aim at improving the quality and availability of scholarly communication by promoting and practicing:
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On September, 29th 2006 he has won a post PhD fellowship from the university of Bologna (from October 2006 to October 2008), at the competitive examination he was ranked first in the industrial engineering area. He extensively served as referee for several international journals. He is author/coauthor of more than 100 research papers. He has been involved in some projects supported by MURST and European Community. 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From 1985 to 1986, he was a Research Fellow in the Research Institute for Electronic Equipment, ZZU AD, Plovdiv, Bulgaria. In 1986, he joined the Department of Control Systems, Technical University of Sofia at the Plovdiv campus, where he is presently a Full Professor. He has held long-term visiting Professor/Scholar positions at various institutions in South Korea, Turkey, Mexico, Greece, Belgium, UK, and Germany. And he has coauthored one book and authored or coauthored more than 80 research papers in conference proceedings and journals. 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After finishing his P. hD degree in 1992, he served in the Industry as a Scientific Officer and continued his academic career as a visiting scholar for a number of educational institutions. In 1996 he joined National University of Science & Technology Pakistan (NUST) as an Associate Professor; NUST is one of the top few universities in Pakistan. In 1999 he joined an International Company Lineo Inc, Canada as Manager Compiler Group, where he headed the group for developing Compiler Tool Chain and Porting of Operating Systems for the BLACKfin processor. The processor development was a joint venture by Intel and Analog Devices. In 2002 Lineo Inc., was taken over by another company, so he joined Aalborg University Denmark as an Assistant Professor.\nProfessor Akbar has truly a multi-disciplined career and he continued his legacy and making progress in many areas of his interests both in teaching and research. 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It is important to understand how these facets interact, given that those diagnosed with AN often fluctuate and relapse–as opposed to maintaining a stable diagnosis—between Diagnostic and Statistical Manual version 5 (DSM-5) categories, over the life course. The National Institute of Health’s Research Domain Criteria (NIH RDoC) subscribes to the transdiagnostic view of mental disorders and provides progressive guidelines for neuroscience research. As such, using the RDoC guidelines may help to pinpoint how impulsivity and compulsivity contribute to the cognitive mechanisms underlying variations in appetite restraint in eating disorders and common psychiatric comorbidities such as anxiety and obsessive-compulsive disorder. Exploring impulsivity and compulsivity in AN from the perspective of the RDoC cognitive systems domain is aided by measures of genetic, molecular, cellular, neural, physiological, behavioural and cognitive task paradigms. Thus, from the standpoint of the RDoC measures, this chapter will describe some of the ways in which impulsivity and compulsivity contribute to the cognitive systems associated with appetite restraint in AN, with the aim of further clarifying a model of appetite restraint to improve treatment interventions.",book:{id:"7885",slug:"anorexia-and-bulimia-nervosa",title:"Anorexia and Bulimia Nervosa",fullTitle:"Anorexia and Bulimia Nervosa"},signatures:"Samantha Jane Brooks and Helgi Schiöth",authors:[{id:"273344",title:"Dr.",name:"Samantha",middleName:null,surname:"Brooks",slug:"samantha-brooks",fullName:"Samantha Brooks"},{id:"293440",title:"Prof.",name:"Helgi",middleName:null,surname:"Schioth",slug:"helgi-schioth",fullName:"Helgi Schioth"}]},{id:"52677",doi:"10.5772/65695",title:"EMDR in Anorexia Nervosa: From a Theoretical Framework to the Treatment Guidelines",slug:"emdr-in-anorexia-nervosa-from-a-theoretical-framework-to-the-treatment-guidelines",totalDownloads:2262,totalCrossrefCites:4,totalDimensionsCites:4,abstract:"Studies on the risks and on the positive factors implied in the onset of anorexia nervosa (AN) have reported the role of an insecure or disorganized state of mind (SoM) with respect to attachment. We compare the effects of eyes movement desensitization and reprocessing (EMDR) approach with cognitive behavioral therapy (CBT) in the treatment of AN in terms of SoMs, reflective function (RF), and narrative coherence (Coh). Our results are part of a broader observational clinical comparative study of the two approaches, and it is based on the Adult Attachment Interview (AAI) outcomes. Differences in terms of belongingness to a secure group and an unsecure group before and after the treatments in EMDR and CBT group have been reported through McNemar's test. The generalized linear model (GLM) repeated‐measures multivariate ANOVA (RM‐MANOVA) has been selected. Our results suggest that EMDR allows an active reprocessing of traumatic memories related to family dynamics and to eating behaviors, which could enable a positive resolution of eating disorder (ED) symptoms. The emotional reprocessing of unresolved attachment issues can allow a better modulation of the control‐related rigidity that is a commonality between AN patients.",book:{id:"5372",slug:"eating-disorders-a-paradigm-of-the-biopsychosocial-model-of-illness",title:"Eating Disorders",fullTitle:"Eating Disorders - A Paradigm of the Biopsychosocial Model of Illness"},signatures:"Maria Zaccagnino, Cristina Civilotti, Martina Cussino, Chiara\nCallerame and Isabel Fernandez",authors:[{id:"186530",title:"Ph.D.",name:"Maria",middleName:null,surname:"Zaccagnino",slug:"maria-zaccagnino",fullName:"Maria Zaccagnino"},{id:"194184",title:"Dr.",name:"Cristina",middleName:null,surname:"Civilotti",slug:"cristina-civilotti",fullName:"Cristina Civilotti"},{id:"194185",title:"Dr.",name:"Martina",middleName:null,surname:"Cussino",slug:"martina-cussino",fullName:"Martina Cussino"},{id:"194186",title:"Dr.",name:"Chiara",middleName:null,surname:"Callerame",slug:"chiara-callerame",fullName:"Chiara Callerame"},{id:"194187",title:"Dr.",name:"Isabel",middleName:null,surname:"Fernandez",slug:"isabel-fernandez",fullName:"Isabel Fernandez"}]},{id:"53353",doi:"10.5772/65305",title:"Communication Challenges Within Eating Disorders: What People Say and What Individuals Hear",slug:"communication-challenges-within-eating-disorders-what-people-say-and-what-individuals-hear",totalDownloads:2140,totalCrossrefCites:4,totalDimensionsCites:4,abstract:"Communication challenges are apparent in many different ways when working with individuals who struggle with eating disorders. These issues can include the influence of parenting styles to society’s weight messages to comments by professionals as they interact with those struggling with eating disorders. Other challenges come from the skewed interpretations that individuals with eating disorders can place on messages that they receive. This chapter examines the literature on many of these issues, highlights challenges with clinical examples, and proposes potential tools to ameliorate some of the impact of these issues on communication.",book:{id:"5372",slug:"eating-disorders-a-paradigm-of-the-biopsychosocial-model-of-illness",title:"Eating Disorders",fullTitle:"Eating Disorders - A Paradigm of the Biopsychosocial Model of Illness"},signatures:"Martha Peaslee Levine",authors:[{id:"186919",title:"Dr.",name:"Martha",middleName:null,surname:"Peaslee Levine",slug:"martha-peaslee-levine",fullName:"Martha Peaslee Levine"}]},{id:"52740",doi:"10.5772/65844",title:"Eating Disorders with Comorbidity Anxiety Disorders",slug:"eating-disorders-with-comorbidity-anxiety-disorders",totalDownloads:1788,totalCrossrefCites:1,totalDimensionsCites:3,abstract:"Although eating disorders and anxiety disorders (AD) are under different diagnosis categories, it is striking that they have high comorbidity and similar clinical features. The most frequently informed anxiety disorders are obsessive-compulsive disorder (OCD), social anxiety disorder (SAD) and generalized anxiety disorder (GAD). Moreover, in cases with a tendency of perfectionism, concern and harm avoidance before the diagnosis of eating disorder, the anxiety disorder is able to be failed to notice. The existence of anxiety disorder or eating disorder makes these syndromes worse. Until today, the relation in between eating disorder and AD has tried to be clarified by phenomenological, neurobiological and family studies. But even if a significant relation has been specified in phenomenological aspect in between OCD and eating disorders, the relation in between eating disorders and other AD is not clear. The existence of AD may be a risk factor in the arise of eating disorders. Therefore, diagnosis and treatment of childhood-adolescence occurring AD may prevent the development of eating disorders. The comorbidity of eating disorders and AD is negatively affecting the treatment and prognosis of the disorder. Moreover, there is limited evidence regarding the effectiveness of treatment options (medication, cognitive behavioral therapy (CBT), family therapy, dialectic behavioral therapy, interpersonal therapy) used in the treatment of cases with a diagnosis of concurrent eating disorder and anxiety disorder. In this chapter, a review of the literature on the comorbidity between eating disorders and the anxiety disorders of OCD, posttraumatic stress disorder (PTSD), SAD, GAD, simple phobia, agoraphobia and panic disorder.",book:{id:"5372",slug:"eating-disorders-a-paradigm-of-the-biopsychosocial-model-of-illness",title:"Eating Disorders",fullTitle:"Eating Disorders - A Paradigm of the Biopsychosocial Model of Illness"},signatures:"Cicek Hocaoglu",authors:[{id:"28322",title:"Prof.",name:"Cicek",middleName:null,surname:"Hocaoglu",slug:"cicek-hocaoglu",fullName:"Cicek Hocaoglu"}]},{id:"66979",doi:"10.5772/intechopen.86083",title:"Patients’ and Carers’ Perspectives of Psychopharmacological Interventions Targeting Anorexia Nervosa Symptoms",slug:"patients-and-carers-perspectives-of-psychopharmacological-interventions-targeting-anorexia-nervosa-s",totalDownloads:937,totalCrossrefCites:0,totalDimensionsCites:2,abstract:"In clinical practice, patients with anorexia nervosa (AN), their carers and clinicians often disagree about psychopharmacological treatment. We developed two corresponding questionnaires to survey the perspectives of patients with AN and their carers on psychopharmacological treatment. These questionnaires were distributed to 36 patients and 37 carers as a quality improvement project on a specialist unit for eating disorders at the South London and Maudsley NHS Foundation Trust. Although most patients did not believe that medication could help with AN, the majority thought that medication for AN should help with anxiety (61.1%), concentration (52.8%), sleep problems (52.8%) and anorexic thoughts (55.6%). Most of the carers shared the view that drug treatment for AN should help with anxiety (54%) and anorexic thoughts (64.8%). Most patients had concerns about potential weight gain, increased appetite, changes in body shape and metabolism during psychopharmacological treatment. By contrast, the majority of carers were not concerned about these specific side effects. Some of the concerns expressed by the patients seem to be AN-related. However, their desire for help with anxiety and anorexic thoughts, which is shared by their carers, should be taken seriously by clinicians when choosing a medication or planning psychopharmacological studies.",book:{id:"7885",slug:"anorexia-and-bulimia-nervosa",title:"Anorexia and Bulimia Nervosa",fullTitle:"Anorexia and Bulimia Nervosa"},signatures:"Amabel Dessain, Jessica Bentley, Janet Treasure, Ulrike Schmidt and Hubertus Himmerich",authors:[{id:"231568",title:"Dr.",name:"Hubertus",middleName:null,surname:"Himmerich",slug:"hubertus-himmerich",fullName:"Hubertus Himmerich"},{id:"240670",title:"Ms.",name:"Jessica",middleName:null,surname:"Bentley",slug:"jessica-bentley",fullName:"Jessica Bentley"},{id:"284593",title:"Dr.",name:"Amabel",middleName:null,surname:"Dessain",slug:"amabel-dessain",fullName:"Amabel Dessain"},{id:"295428",title:"Prof.",name:"Janet",middleName:null,surname:"Treasure",slug:"janet-treasure",fullName:"Janet Treasure"},{id:"295429",title:"Prof.",name:"Ulrike",middleName:null,surname:"Schmidt",slug:"ulrike-schmidt",fullName:"Ulrike Schmidt"}]}],mostDownloadedChaptersLast30Days:[{id:"53036",title:"Neurobiology and the Changing Face of Eating Disorder Treatment: Healing the Eating Disordered Brain",slug:"neurobiology-and-the-changing-face-of-eating-disorder-treatment-healing-the-eating-disordered-brain",totalDownloads:2037,totalCrossrefCites:0,totalDimensionsCites:0,abstract:"By recognizing eating disorders (EDs) as disruptions in brain circuitry, neuroscience has begun to shed light on how people make changes in psychotherapy. The clinician who treats the eating disordered patient also treats the eating disordered brain. It is time for practitioners to become better acquainted with the organ they treat, and to apply neuroplasticity research findings to clinical practice. Eating disorders and body image disturbances signify the loss of integrity of the core self. Twenty-first century research and technology has validated the age‐old notion that healthy neuronal connectivity within, and between, mind(s), brain(s), and body(s) reintegrates and defines the healthy self. The concept of the “self” as embodied (grounded in somatic reality) expands the scope of effective healing practices. Neurophysiological (somatosensory education and mindful psychotherapeutic attachments) interventions that support the emergence of embodied mindfulness and sensory awareness facilitate the reintegration of the eating disordered brain, and of the fragmented core self. Both lie at the heart of eating disorder recovery. Nowhere in the field of mental health are the concepts of the embedded self and embodied healing as significant as in the treatment of eating disorders and body image disturbances. This article discusses the healing impact of neurophysiological connections, intrapersonal and interpersonal, that foster recovery of the self.",book:{id:"5372",slug:"eating-disorders-a-paradigm-of-the-biopsychosocial-model-of-illness",title:"Eating Disorders",fullTitle:"Eating Disorders - A Paradigm of the Biopsychosocial Model of Illness"},signatures:"Abigail H. Natenshon",authors:[{id:"186482",title:"M.A.",name:"Abigail H.",middleName:null,surname:"Natenshon",slug:"abigail-h.-natenshon",fullName:"Abigail H. Natenshon"}]},{id:"67092",title:"Bulimia Nervosa and Body Dissatisfaction in Terms of Self-Perception of Body Image",slug:"bulimia-nervosa-and-body-dissatisfaction-in-terms-of-self-perception-of-body-image",totalDownloads:1038,totalCrossrefCites:0,totalDimensionsCites:0,abstract:"Bulimia nervosa is characterized by disturbed body image, repetitive binge eating, and compensatory behaviours such as self-induced vomiting, laxative abuse, or fasting. Body image dissatisfaction and eating disordered behaviours (e.g. food restriction, purging, and binge eating) can affect men and women of varied ages, races, and cultural backgrounds. Body dissatisfaction is defined as a negative subjective evaluation of the weight and shape of one’s own body. Body dissatisfaction predicts the onset, severity, and treatment outcomes of eating disorders. A core component of body dissatisfaction is appearance-based social comparisons. In this context a study on self-perception of body image of women in Riyadh in 2018 revealed that a sudden spurt in obesity after marriage is leading to shift of higher percentage of women from positive to negative perception. Overall, an underestimation of body weight in terms of BMI was found among the participants. Such misconceptions should be addressed in view of the high obesity prevalence. It was also evident that positive and negative body image perception will lead to eating disorders in adolescents.",book:{id:"7885",slug:"anorexia-and-bulimia-nervosa",title:"Anorexia and Bulimia Nervosa",fullTitle:"Anorexia and Bulimia Nervosa"},signatures:"Layam Anitha, Asma Abdulaziz Alhussaini, Hessah Ibrahim Alsuwedan, Hessa Faleh Alnefaie, Rehab Abduallah Almubrek and Shima Abdulaziz Aldaweesh",authors:[{id:"276645",title:"Dr.",name:"Anitha",middleName:null,surname:"Layam",slug:"anitha-layam",fullName:"Anitha Layam"},{id:"408346",title:"Dr.",name:"Asma",middleName:null,surname:"Abdulaziz Alhussaini",slug:"asma-abdulaziz-alhussaini",fullName:"Asma Abdulaziz Alhussaini"},{id:"408347",title:"Dr.",name:"Hessah",middleName:null,surname:"Ibrahim Alsuwedan",slug:"hessah-ibrahim-alsuwedan",fullName:"Hessah Ibrahim Alsuwedan"},{id:"408348",title:"Dr.",name:"Hessa",middleName:null,surname:"Faleh Alnefaie",slug:"hessa-faleh-alnefaie",fullName:"Hessa Faleh Alnefaie"},{id:"408349",title:"Dr.",name:"Rehab",middleName:null,surname:"Abduallah Almubrek",slug:"rehab-abduallah-almubrek",fullName:"Rehab Abduallah Almubrek"},{id:"408350",title:"Dr.",name:"Shima",middleName:null,surname:"Abdulaziz Aldaweesh",slug:"shima-abdulaziz-aldaweesh",fullName:"Shima Abdulaziz Aldaweesh"}]},{id:"64858",title:"The Neurobiology of Anorexia Nervosa",slug:"the-neurobiology-of-anorexia-nervosa",totalDownloads:1557,totalCrossrefCites:1,totalDimensionsCites:1,abstract:"Anorexia nervosa is considered the most deadly psychological illness. Individuals with and recovered from anorexia nervosa experience numerous physical and mental health difficulties, and treatment outcomes remain unpromising. Anorexia nervosa is rare in the general population, but common among individuals with a first-degree relative with the disorder. In addition, the onset of anorexia nervosa is developmentally specific, which suggests a partly biological etiology. A better understanding of the biological and neurobiological etiology of anorexia nervosa is direly needed to inform new therapies and to identify individuals at risk for the disorder. This paper summarizes the research related to neurotransmitter abnormalities, aberrant brain activity, and genetic and epigenetic mechanisms that may contribute to the etiology of this deadly disorder.",book:{id:"7885",slug:"anorexia-and-bulimia-nervosa",title:"Anorexia and Bulimia Nervosa",fullTitle:"Anorexia and Bulimia Nervosa"},signatures:"Ashley Higgins",authors:[{id:"274781",title:"Dr.",name:"Ashley",middleName:null,surname:"Higgins",slug:"ashley-higgins",fullName:"Ashley Higgins"}]},{id:"53353",title:"Communication Challenges Within Eating Disorders: What People Say and What Individuals Hear",slug:"communication-challenges-within-eating-disorders-what-people-say-and-what-individuals-hear",totalDownloads:2140,totalCrossrefCites:4,totalDimensionsCites:4,abstract:"Communication challenges are apparent in many different ways when working with individuals who struggle with eating disorders. These issues can include the influence of parenting styles to society’s weight messages to comments by professionals as they interact with those struggling with eating disorders. Other challenges come from the skewed interpretations that individuals with eating disorders can place on messages that they receive. This chapter examines the literature on many of these issues, highlights challenges with clinical examples, and proposes potential tools to ameliorate some of the impact of these issues on communication.",book:{id:"5372",slug:"eating-disorders-a-paradigm-of-the-biopsychosocial-model-of-illness",title:"Eating Disorders",fullTitle:"Eating Disorders - A Paradigm of the Biopsychosocial Model of Illness"},signatures:"Martha Peaslee Levine",authors:[{id:"186919",title:"Dr.",name:"Martha",middleName:null,surname:"Peaslee Levine",slug:"martha-peaslee-levine",fullName:"Martha Peaslee Levine"}]},{id:"53044",title:"Oral Implications of Eating Disorders",slug:"oral-implications-of-eating-disorders",totalDownloads:1321,totalCrossrefCites:0,totalDimensionsCites:0,abstract:"Eating disorders (EDs) are defined as persistent behavioural problems related to food and weight control, which significantly damage the physical and mental health with dramatic effects on the oral cavity. 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Professor Derbel has published many articles in journals and collaborates intensively with IntechOpen Access Publisher as an editor.",institutionString:"Clinique les Oliviers",institution:null},{id:"300144",title:"Dr.",name:"Meriem",middleName:null,surname:"Braiki",slug:"meriem-braiki",fullName:"Meriem Braiki",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/300144/images/system/300144.jpg",biography:"Dr. Meriem Braiki is a specialist in pediatric surgeon from Tunisia. She was born in 1985. She received her medical degree from the University of Medicine at Sousse, Tunisia. She achieved her surgical residency training periods in Pediatric Surgery departments at University Hospitals in Monastir, Tunis and France.\r\nShe is currently working at the Pediatric surgery department, Sidi Bouzid Hospital, Tunisia. Her hospital activities are mostly concerned with laparoscopic, parietal, urological and digestive surgery. She has published several articles in diffrent journals.",institutionString:"Sidi Bouzid Regional Hospital",institution:null},{id:"229481",title:"Dr.",name:"Erika M.",middleName:"Martins",surname:"de Carvalho",slug:"erika-m.-de-carvalho",fullName:"Erika M. de Carvalho",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/229481/images/6397_n.jpg",biography:null,institutionString:null,institution:{name:"Oswaldo Cruz Foundation",country:{name:"Brazil"}}},{id:"186537",title:"Prof.",name:"Tonay",middleName:null,surname:"Inceboz",slug:"tonay-inceboz",fullName:"Tonay Inceboz",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/186537/images/system/186537.jfif",biography:"I was graduated from Ege University of Medical Faculty (Turkey) in 1988 and completed his Med. PhD degree in Medical Parasitology at the same university. I became an Associate Professor in 2008 and Professor in 2014. I am currently working as a Professor at the Department of Medical Parasitology at Dokuz Eylul University, Izmir, Turkey.\n\nI have given many lectures, presentations in different academic meetings. I have more than 60 articles in peer-reviewed journals, 18 book chapters, 1 book editorship.\n\nMy research interests are Echinococcus granulosus, Echinococcus multilocularis (diagnosis, life cycle, in vitro and in vivo cultivation), and Trichomonas vaginalis (diagnosis, PCR, and in vitro cultivation).",institutionString:"Dokuz Eylül University",institution:{name:"Dokuz Eylül University",country:{name:"Turkey"}}},{id:"71812",title:"Prof.",name:"Hanem Fathy",middleName:"Fathy",surname:"Khater",slug:"hanem-fathy-khater",fullName:"Hanem Fathy Khater",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/71812/images/1167_n.jpg",biography:"Prof. Khater is a Professor of Parasitology at Benha University, Egypt. She studied for her doctoral degree, at the Department of Entomology, College of Agriculture, Food and Natural Resources, University of Missouri, Columbia, USA. She has completed her Ph.D. degrees in Parasitology in Egypt, from where she got the award for “the best scientific Ph.D. dissertation”. She worked at the School of Biological Sciences, Bristol, England, the UK in controlling insects of medical and veterinary importance as a grant from Newton Mosharafa, the British Council. Her research is focused on searching of pesticides against mosquitoes, house flies, lice, green bottle fly, camel nasal botfly, soft and hard ticks, mites, and the diamondback moth as well as control of several parasites using safe and natural materials to avoid drug resistances and environmental contamination.",institutionString:null,institution:{name:"Banha University",country:{name:"Egypt"}}},{id:"99780",title:"Prof.",name:"Omolade",middleName:"Olayinka",surname:"Okwa",slug:"omolade-okwa",fullName:"Omolade Okwa",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/99780/images/system/99780.jpg",biography:"Omolade Olayinka Okwa is presently a Professor of Parasitology at Lagos State University, Nigeria. She has a PhD in Parasitology (1997), an MSc in Cellular Parasitology (1992), and a BSc (Hons) Zoology (1990) all from the University of Ibadan, Nigeria. She teaches parasitology at the undergraduate and postgraduate levels. She was a recipient of a Commonwealth fellowship supported by British Council tenable at the Centre for Entomology and Parasitology (CAEP), Keele University, United Kingdom between 2004 and 2005. She was awarded an Honorary Visiting Research Fellow at the same university from 2005 to 2007. \nShe has been an external examiner to the Department of Veterinary Microbiology and Parasitology, University of Ibadan, MSc programme between 2010 and 2012. She is a member of the Nigerian Society of Experimental Biology (NISEB), Parasitology and Public Health Society of Nigeria (PPSN), Science Association of Nigeria (SAN), Zoological Society of Nigeria (ZSN), and is Vice Chairperson of the Organisation of Women in Science (OWSG), LASU chapter. She served as Head of Department of Zoology and Environmental Biology, Lagos State University from 2007 to 2010 and 2014 to 2016. She is a reviewer for several local and international journals such as Unilag Journal of Science, Libyan Journal of Medicine, Journal of Medicine and Medical Sciences, and Annual Research and Review in Science. \nShe has authored 45 scientific research publications in local and international journals, 8 scientific reviews, 4 books, and 3 book chapters, which includes the books “Malaria Parasites” and “Malaria” which are IntechOpen access publications.",institutionString:"Lagos State University",institution:{name:"Lagos State University",country:{name:"Nigeria"}}},{id:"273100",title:"Dr.",name:"Vijay",middleName:null,surname:"Gayam",slug:"vijay-gayam",fullName:"Vijay Gayam",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/273100/images/system/273100.jpeg",biography:"Dr. Vijay Bhaskar Reddy Gayam is currently practicing as an internist at Interfaith Medical Center in Brooklyn, New York, USA. He is also a Clinical Assistant Professor at the SUNY Downstate University Hospital and Adjunct Professor of Medicine at the American University of Antigua. He is a holder of an M.B.B.S. degree bestowed to him by Osmania Medical College and received his M.D. at Interfaith Medical Center. His career goals thus far have heavily focused on direct patient care, medical education, and clinical research. He currently serves in two leadership capacities; Assistant Program Director of Medicine at Interfaith Medical Center and as a Councilor for the American\r\nFederation for Medical Research. As a true academician and researcher, he has more than 50 papers indexed in international peer-reviewed journals. He has also presented numerous papers in multiple national and international scientific conferences. His areas of research interest include general internal medicine, gastroenterology and hepatology. He serves as an editor, editorial board member and reviewer for multiple international journals. His research on Hepatitis C has been very successful and has led to multiple research awards, including the 'Equity in Prevention and Treatment Award” from the New York Department of Health Viral Hepatitis Symposium (2018) and the 'Presidential Poster Award” awarded to him by the American College of Gastroenterology (2018). He was also awarded 'Outstanding Clinician in General Medicine” by Venus International Foundation for his extensive research expertise and services, perform over and above the standard expected in the advancement of healthcare, patient safety and quality of care.",institutionString:"Interfaith Medical Center",institution:{name:"Interfaith Medical Center",country:{name:"United States of America"}}},{id:"93517",title:"Dr.",name:"Clement",middleName:"Adebajo",surname:"Meseko",slug:"clement-meseko",fullName:"Clement Meseko",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/93517/images/system/93517.jpg",biography:"Dr. Clement Meseko obtained DVM and PhD degree in Veterinary Medicine and Virology respectively. He has worked for over 20 years in both private and public sectors including the academia, contributing to knowledge and control of infectious disease. Through the application of epidemiological skill, classical and molecular virological skills, he investigates viruses of economic and public health importance for the mitigation of the negative impact on people, animal and the environment in the context of Onehealth. \r\nDr. Meseko’s field experience on animal and zoonotic diseases and pathogen dynamics at the human-animal interface over the years shaped his carrier in research and scientific inquiries. He has been part of the investigation of Highly Pathogenic Avian Influenza incursions in sub Saharan Africa and monitors swine Influenza (Pandemic influenza Virus) agro-ecology and potential for interspecies transmission. He has authored and reviewed a number of journal articles and book chapters.",institutionString:"National Veterinary Research Institute",institution:{name:"National Veterinary Research Institute",country:{name:"Nigeria"}}},{id:"158026",title:"Prof.",name:"Shailendra K.",middleName:null,surname:"Saxena",slug:"shailendra-k.-saxena",fullName:"Shailendra K. Saxena",position:null,profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bRET3QAO/Profile_Picture_2022-05-10T10:10:26.jpeg",biography:"Professor Dr. Shailendra K. Saxena is a vice dean and professor at King George's Medical University, Lucknow, India. His research interests involve understanding the molecular mechanisms of host defense during human viral infections and developing new predictive, preventive, and therapeutic strategies for them using Japanese encephalitis virus (JEV), HIV, and emerging viruses as a model via stem cell and cell culture technologies. His research work has been published in various high-impact factor journals (Science, PNAS, Nature Medicine) with a high number of citations. He has received many awards and honors in India and abroad including various Young Scientist Awards, BBSRC India Partnering Award, and Dr. JC Bose National Award of Department of Biotechnology, Min. of Science and Technology, Govt. of India. Dr. Saxena is a fellow of various international societies/academies including the Royal College of Pathologists, United Kingdom; Royal Society of Medicine, London; Royal Society of Biology, United Kingdom; Royal Society of Chemistry, London; and Academy of Translational Medicine Professionals, Austria. He was named a Global Leader in Science by The Scientist. He is also an international opinion leader/expert in vaccination for Japanese encephalitis by IPIC (UK).",institutionString:"King George's Medical University",institution:{name:"King George's Medical University",country:{name:"India"}}},{id:"94928",title:"Dr.",name:"Takuo",middleName:null,surname:"Mizukami",slug:"takuo-mizukami",fullName:"Takuo Mizukami",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/94928/images/6402_n.jpg",biography:null,institutionString:null,institution:{name:"National Institute of Infectious Diseases",country:{name:"Japan"}}},{id:"233433",title:"Dr.",name:"Yulia",middleName:null,surname:"Desheva",slug:"yulia-desheva",fullName:"Yulia Desheva",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/233433/images/system/233433.png",biography:"Dr. Yulia Desheva is a leading researcher at the Institute of Experimental Medicine, St. Petersburg, Russia. She is a professor in the Stomatology Faculty, St. Petersburg State University. She has expertise in the development and evaluation of a wide range of live mucosal vaccines against influenza and bacterial complications. Her research interests include immunity against influenza and COVID-19 and the development of immunization schemes for high-risk individuals.",institutionString:'Federal State Budgetary Scientific Institution "Institute of Experimental Medicine"',institution:null},{id:"238958",title:"Mr.",name:"Atamjit",middleName:null,surname:"Singh",slug:"atamjit-singh",fullName:"Atamjit Singh",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/238958/images/6575_n.jpg",biography:null,institutionString:null,institution:null},{id:"333753",title:"Dr.",name:"Rais",middleName:null,surname:"Ahmed",slug:"rais-ahmed",fullName:"Rais Ahmed",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/333753/images/20168_n.jpg",biography:null,institutionString:null,institution:null},{id:"252058",title:"M.Sc.",name:"Juan",middleName:null,surname:"Sulca",slug:"juan-sulca",fullName:"Juan Sulca",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/252058/images/12834_n.jpg",biography:null,institutionString:null,institution:null},{id:"191392",title:"Dr.",name:"Marimuthu",middleName:null,surname:"Govindarajan",slug:"marimuthu-govindarajan",fullName:"Marimuthu Govindarajan",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/191392/images/5828_n.jpg",biography:"Dr. M. Govindarajan completed his BSc degree in Zoology at Government Arts College (Autonomous), Kumbakonam, and MSc, MPhil, and PhD degrees at Annamalai University, Annamalai Nagar, Tamil Nadu, India. He is serving as an assistant professor at the Department of Zoology, Annamalai University. His research interests include isolation, identification, and characterization of biologically active molecules from plants and microbes. He has identified more than 20 pure compounds with high mosquitocidal activity and also conducted high-quality research on photochemistry and nanosynthesis. He has published more than 150 studies in journals with impact factor and 2 books in Lambert Academic Publishing, Germany. He serves as an editorial board member in various national and international scientific journals.",institutionString:null,institution:null},{id:"274660",title:"Dr.",name:"Damodar",middleName:null,surname:"Paudel",slug:"damodar-paudel",fullName:"Damodar Paudel",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/274660/images/8176_n.jpg",biography:"I am DrDamodar Paudel,currently working as consultant Physician in Nepal police Hospital.",institutionString:null,institution:null},{id:"241562",title:"Dr.",name:"Melvin",middleName:null,surname:"Sanicas",slug:"melvin-sanicas",fullName:"Melvin Sanicas",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/241562/images/6699_n.jpg",biography:null,institutionString:null,institution:null},{id:"337446",title:"Dr.",name:"Maria",middleName:null,surname:"Zavala-Colon",slug:"maria-zavala-colon",fullName:"Maria Zavala-Colon",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Puerto Rico, Medical Sciences Campus",country:{name:"United States of America"}}},{id:"338856",title:"Mrs.",name:"Nur Alvira",middleName:null,surname:"Pascawati",slug:"nur-alvira-pascawati",fullName:"Nur Alvira Pascawati",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Universitas Respati Yogyakarta",country:{name:"Indonesia"}}},{id:"441116",title:"Dr.",name:"Jovanka M.",middleName:null,surname:"Voyich",slug:"jovanka-m.-voyich",fullName:"Jovanka M. Voyich",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Montana State University",country:{name:"United States of America"}}},{id:"330412",title:"Dr.",name:"Muhammad",middleName:null,surname:"Farhab",slug:"muhammad-farhab",fullName:"Muhammad Farhab",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Agriculture Faisalabad",country:{name:"Pakistan"}}},{id:"349495",title:"Dr.",name:"Muhammad",middleName:null,surname:"Ijaz",slug:"muhammad-ijaz",fullName:"Muhammad Ijaz",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Veterinary and Animal Sciences",country:{name:"Pakistan"}}}]}},subseries:{item:{id:"26",type:"subseries",title:"Machine Learning and Data Mining",keywords:"Intelligent Systems, Machine Learning, Data Science, Data Mining, Artificial Intelligence",scope:"The scope of machine learning and data mining is immense and is growing every day. 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