Main characteristics of HHVs.
\r\n\t
",isbn:"978-1-83962-547-3",printIsbn:"978-1-83962-546-6",pdfIsbn:"978-1-83962-548-0",doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!0,hash:"e5ba02fedd7c87f0ab66414f3b07de0c",bookSignature:" John P. Tiefenbacher",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/10765.jpg",keywords:"Managing Urbanization, Managing Development, Managing Resource Use, Drought Management, Flood Management, Water Quality Monitoring, Air Quality Monitoring, Ecological Monitoring, Modeling Extreme Natural Events, Ecological Restoration, Restoring Environmental Flows, Environmental Management Perspectives",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:null,numberOfDimensionsCitations:null,numberOfTotalCitations:null,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"January 12th 2021",dateEndSecondStepPublish:"February 9th 2021",dateEndThirdStepPublish:"April 10th 2021",dateEndFourthStepPublish:"June 29th 2021",dateEndFifthStepPublish:"August 28th 2021",remainingDaysToSecondStep:"20 days",secondStepPassed:!1,currentStepOfPublishingProcess:2,editedByType:null,kuFlag:!1,biosketch:"A geospatial scholar working at the interface of natural and human systems, collaborating internationally on innovative studies about hazards and environmental challenges. Dr. Tiefenbacher has published more than 200 papers on a diverse array of topics that examine perception and behaviors with regards to the application of pesticides, releases of toxic chemicals, environments of the U.S.-Mexico borderlands, wildlife hazards, and the geography of wine.",coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"73876",title:"Dr.",name:"John P.",middleName:null,surname:"Tiefenbacher",slug:"john-p.-tiefenbacher",fullName:"John P. Tiefenbacher",profilePictureURL:"https://mts.intechopen.com/storage/users/73876/images/system/73876.jfif",biography:"Dr. John P. Tiefenbacher (Ph.D., Rutgers, 1992) is a professor of Geography at Texas State University. His research has focused on various aspects of hazards and environmental management. Dr. Tiefenbacher has published on a diverse array of topics that examine perception and behaviors with regards to the application of pesticides, releases of toxic chemicals, environments of the U.S.-Mexico borderlands, wildlife hazards, and the geography of wine. More recently his work pertains to spatial adaptation to climate change, spatial responses in wine growing regions to climate change, the geographies of viticulture and wine, artificial intelligence and machine learning to predict patterns of natural processes and hazards, historical ethnic enclaves in American cities and regions, and environmental adaptations of 19th century European immigrants to North America's landscapes.",institutionString:"Texas State University",position:null,outsideEditionCount:0,totalCites:0,totalAuthoredChapters:"1",totalChapterViews:"0",totalEditedBooks:"6",institution:{name:"Texas State University",institutionURL:null,country:{name:"United States of America"}}}],coeditorOne:null,coeditorTwo:null,coeditorThree:null,coeditorFour:null,coeditorFive:null,topics:[{id:"12",title:"Environmental Sciences",slug:"environmental-sciences"}],chapters:null,productType:{id:"1",title:"Edited Volume",chapterContentType:"chapter",authoredCaption:"Edited by"},personalPublishingAssistant:{id:"194667",firstName:"Marijana",lastName:"Francetic",middleName:null,title:"Ms.",imageUrl:"https://mts.intechopen.com/storage/users/194667/images/4752_n.jpg",email:"marijana@intechopen.com",biography:"As an Author Service Manager my responsibilities include monitoring and facilitating all publishing activities for authors and editors. From chapter submission and review, to approval and revision, copyediting and design, until final publication, I work closely with authors and editors to ensure a simple and easy publishing process. I maintain constant and effective communication with authors, editors and reviewers, which allows for a level of personal support that enables contributors to fully commit and concentrate on the chapters they are writing, editing, or reviewing. I assist authors in the preparation of their full chapter submissions and track important deadlines and ensure they are met. I help to coordinate internal processes such as linguistic review, and monitor the technical aspects of the process. As an ASM I am also involved in the acquisition of editors. 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Venkateswarlu",coverURL:"https://cdn.intechopen.com/books/images_new/371.jpg",editedByType:"Edited by",editors:[{id:"58592",title:"Dr.",name:"Arun",surname:"Shanker",slug:"arun-shanker",fullName:"Arun Shanker"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"878",title:"Phytochemicals",subtitle:"A Global Perspective of Their Role in Nutrition and Health",isOpenForSubmission:!1,hash:"ec77671f63975ef2d16192897deb6835",slug:"phytochemicals-a-global-perspective-of-their-role-in-nutrition-and-health",bookSignature:"Venketeshwer Rao",coverURL:"https://cdn.intechopen.com/books/images_new/878.jpg",editedByType:"Edited by",editors:[{id:"82663",title:"Dr.",name:"Venketeshwer",surname:"Rao",slug:"venketeshwer-rao",fullName:"Venketeshwer Rao"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"4816",title:"Face Recognition",subtitle:null,isOpenForSubmission:!1,hash:"146063b5359146b7718ea86bad47c8eb",slug:"face_recognition",bookSignature:"Kresimir Delac and Mislav Grgic",coverURL:"https://cdn.intechopen.com/books/images_new/4816.jpg",editedByType:"Edited by",editors:[{id:"528",title:"Dr.",name:"Kresimir",surname:"Delac",slug:"kresimir-delac",fullName:"Kresimir Delac"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"3621",title:"Silver Nanoparticles",subtitle:null,isOpenForSubmission:!1,hash:null,slug:"silver-nanoparticles",bookSignature:"David Pozo Perez",coverURL:"https://cdn.intechopen.com/books/images_new/3621.jpg",editedByType:"Edited by",editors:[{id:"6667",title:"Dr.",name:"David",surname:"Pozo",slug:"david-pozo",fullName:"David Pozo"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}]},chapter:{item:{type:"chapter",id:"51491",title:"Ectopic Expression of Human Herpesvirus 1 Thymidine Kinase Induces Male Infertility",doi:"10.5772/64390",slug:"ectopic-expression-of-human-herpesvirus-1-thymidine-kinase-induces-male-infertility",body:'\nHerpesviruses belong to a family of double‐stranded DNA (dsDNA) viruses commonly causing herpes in animals. They present a unique four‐layered structure. The outermost layer corresponds to the envelope, a lipid bilayer membrane interspersed with glycoproteins. This encases the tegument, a protein coat that surrounds the icosahedral nucleocapsid containing the viral linear DNA genome (Figure 1) [1]. At present, over 130 herpesvirus species have been identified, eight of which are known as human herpesviruses (HHVs): HHV‐1 and HHV‐2 (commonly called herpes simplex virus‐1 and ‐2; HSV‐1 and HSV‐2), HHV‐3 (varicella‐zoster virus; VZV), HHV‐4 (Epstein‐Barr virus; EBV), HHV‐5 (cytomegalovirus; CMV), HHV‐6 (human herpesvirus 6, including HHV‐6A and HHV‐6B), HHV‐7 (human herpesvirus 7), and HHV‐8 (Kaposi’s sarcoma‐associated herpesvirus; KSHV). HHVs are divided into three subfamilies (α‐, β‐ and γ‐herpesviruses) based on their unique properties (Table 1) [2]. HHVs are widespread among humans to the extent that more than 90% of adults are thought to be infected with at least one variety [2]. HHVs generally infect ectoderm‐derived tissues, such as skin, mucoepithelial tissue and nervous tissue; however, they usually show preference for specific target cells/tissues and ensuing clinical outcomes (Table 1) [2]. In addition, many people carrying the virus may be asymptomatic due to the latency of associated transcripts, which help HHVs evade the host\'s immune response [3]. Moreover, HHVs hide in specific cells during latent infection periods (Table 1).
HHVs structure. HHVs have a unique four‐layered structure: a core of double‐stranded DNA is surrounded by the nucleocapsid, which is enclosed by the envelope. The envelope is attached to the capsid through the tegument. Glycoproteins cover the surface of the envelope.
During the last decade, numerous studies have demonstrated that genital infections are often associated with human infertility, especially in males [4]. Most recently, genital infections caused by herpesviruses have attracted considerable attention [5–7]. Although herpes genitalis can be mainly caused by HHV‐1 and HHV‐2 [8], other HHVs have also been frequently detected in the genital organs of infertile male patients [9–11]. Although not fully demonstrated, there is a possible correlation between human male infertility and HHVs.
\nOur laboratory recently developed a line of transgenic (Tg) rats expressing a chimeric gene with the promoter of porcine follicle‐stimulating hormone β subunit (FSHβ) and the open reading frame of HHV‐1 thymidine kinase (TK). Unexpectedly, we observed that Tg rats showed infertility upon ectopic expression of HHV‐1‐TK in the testis, independent of the FSHβ promoter [12]. Indeed, we found that the accumulation of HHV‐1‐TK protein in postmeiotic spermatids was the cause of male infertility [13]. In this study, we used the Tg rat model to elucidate the HHV‐mediated mechanism responsible for male infertility.
\nVirus | \nCommon name | \nMajor target | \nDiseases | \nClinical syndromes | \nSite of latency | \n
---|---|---|---|---|---|
HHV‐1α | \nHSV‐1 Herpes simplex virus‐1 | \nMucoepithelial | \nOral herpes, genital herpes | \nOral and genital herpes, gingivostomatitis, keratoconjunctivitis, encephalitis, pneumonitis, esophagitis, hepatitis | \nSensory and cranial nerve ganglia | \n
HHV‐2α | \nHSV‐2 Herpes simplex virus‐2 | \nSame as above | \nsame as above | \nOral and genital herpes herpes, gingivostomatitis, keratoconjunctivitis, meningitis, encephalitis, pneumonitis, esophagitis, hepatitis | \nSame as above | \n
HHV‐3α | \nVZV Varicella zoster virus | \nSame as above | \nChickenpox, shingles | \nShingles (extradermatomal), pneumonitis, disseminated infection, hepatitis, retinitis, meningitis, hemolysis, leukopenia, thrombocytopenia | \nSame as above | \n
HHV‐4γ | \nEBV Epstein‐Bar virus | \nB cells and epithelial cells | \nInfectious mononucleosis, Burkitt’s lymphoma, CNS lymphoma, posttransplant lymphoproliferative syndrome (PTLD), nasopharyngeal carcinoma, HIV‐associated hairy leukoplakia | \nMononucleosis, posttransplant lymphoproliferative disorders, pneumonitis, hepatitis, encephalitis, hemolysis, leukopenia, thrombocytopenia | \nMemory B cells | \n
HHV‐5β | \nCMV Cytomegalovirus | \nMonocyte, lymphocyte and epithelial cells | \nInfectious mononucleosis‐like syndrome, retinitis | \nLymphadenopathy, hepatitis, pneumonitis, cns vasculitis/ encephalitis, retinitis, esophagitis, hemolysis, leukopenia, thrombocytopenia | \nMonocytes, macrophages, lymphocytes, others | \n
HHV‐6A/Bβ | \n\n | T cells, NK cell, epithelial cells and others | \nHHV‐6A: multiple sclerosis, encephalitis, glioma HHV‐6B: sixth disease (roseola infantum or exanthem subitum), multiple sclerosis, encephalitism, hepatitis, pneumonitis, glioma | \nHHV‐6A: meningoencephalitis, encephalitis, perceptual‐motor dysfunction, hemiplegia, lymphoproliferative disorder HHV‐6B: rash, fever, meningoencephalitis, encephalitis, perceptual‐motor dysfunction, hemiplegia, lymphoproliferative disorder, pneumonitis, hepatitis, thrombocytopenia, leukopenia, | \nT, B, NK cells, monocytes, macrophages, liver, salivary endothelial, neuronal cells | \n
HHV‐7γ | \n\n | T cells | \nSixth disease (roseola infantum or exanthem subitum), pityriasis rosea | \nRash, fever, encephalitis?, hepatitis? | \nCD4+ T cells, salivary epithelial, lung, skin cells | \n
HHV‐8γ | \nKSHV Kaposi’s sarcoma‐associated herpesvirus | \nLymphocyte and others | \nKaposi’s sarcoma, primary effusion lymphoma, some types of multicentric Castleman’s disease | \nFever, mononucleosis, skin lesions, encephalitis? | \nB cells | \n
Main characteristics of HHVs.
1. α‐herpesvirus: rapid reproduction and cell lysis in vitro, rapid cell lysis and spread in vivo, primary target mucoepithelial cells, latency in sensory ganglia.
2. β‐herpesvirus: long replication cycle in vivo and in vitro, limited host range, large infected cells, latency in mononuclear cells, secretory cells, some epithelial cells, and others.
3. γ‐herpesvirus: replication in lymphoblastoid cells, lytic cycle in some fibroblasts and epithelial cells.
Human infertility is a widespread problem that has been increasing in recent decades. It affects 20–30% of couples in the world [14], and 40–50% is attributed to male infertility [15, 16]. Many risk factors can disrupt male reproductive capacity and cause infertility. Among these are infections with pathogens [17], such as viruses [7], mycoplasma [18], chlamydia [19], and bacteria [20].
\nRecently, a number of studies have detected HHVs in the semen and/or spermatozoa of ∼90% of infertile male patients, although regional variations must be taken into account (Table 2) [6, 9–11, 21–31]. Many investigators have tried to confirm the relationship between male infertility and HHVs infection by combining viral infection rates with statistical analysis of semen and/or sperm samples derived from PCR‐based viral DNA analysis, antigen‐antibody reaction, and immunocytochemistry of spermatozoa. However, only in a few cases, a significant difference in sperm counts, motility, and/or abnormality has been found between HHVs carriers and non‐carriers [6, 9, 11, 27].
\nAlthough many studies have revealed a high prevalence of HHVs in male infertility patients, it is still difficult to conclude that HHVs infection is the causative agent. To address this issue, the capacity of HHVs to infect the testis and interfere with spermatogenesis should be proven regardless of the presence or absence of HHVs in the semen and spermatozoa. This would require direct evidence of a molecular mechanism for HHV‐induced infertility. In this respect, studies in Tg animal models offer the opportunity to better understand human male infertility.
\nCountry or area (year) | \nCarrier% (carrier/subject) | \nHHV‐1 | \nHHV‐2 | \nHHV‐3 | \nHHV‐4 | \nHHV‐5 | \nHHV‐6A/B | \nHHV‐7 | \nHHV‐8 | \n
---|---|---|---|---|---|---|---|---|---|
\n | \n | (HSV‐1) | \n(HSV‐2) | \n(VZV) | \n(EBV) | \n(CMV) | \n\n | \n | \n |
Italian (1999) [22] | \n91 | \n– | \n– | \n– | \n– | \n– | \n– | \n– | \n91 | \n
\n | (30/33) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Germany (2001) [6] | \n17.1 | \n3.2 | \n0 | \n7.1 | \n3.6 | \n4 | \n0.4** | \n0 | \n|
\n | (43/252) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Athens (2003) [9] | \n56.6 | \n49.5* | \n– | \n16.8 | \n7.1 | \n– | \n– | \n– | \n|
\n | (64/113) | \n\n | \n | \n | \n | \n | \n | \n | \n |
UK (2006) [25] | \n3 | \n\n | \n | \n | 3 | \n\n | \n | \n | \n |
\n | (1/33) | \n\n | \n | \n | \n | \n | \n | \n | \n |
USA (2007) [10] | \n18.7 | \n3.7 | \n– | \n0.4 | \n8.7 | \n3.7 | \n– | \n– | \n|
\n | (45/241) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Coast (2007) [31] | \n14.3 | \n\n | \n | \n | \n | 14.3 | \n\n | \n | \n |
\n | (9/63) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Greece (2009) [11] | \n– | \n2.1 | \n– | \n3.2* | \n39.1 | \n56.5 | \n66.3 | \n0 | \n– | \n
Germany (2009) [23] | \n6.5 | \n\n | \n | \n | \n | 6.5 | \n\n | \n | \n |
\n | (11/170) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Russia (2011) [26] | \n11 | \n– | \n– | \n– | \n– | \n11 | \n– | \n– | \n– | \n
\n | (10/91) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Spain (2012) [24] | \n54 | \n29 | \n– | \n45 | \n43 | \n8.2 | \n3.6 | \n– | \n|
\n | (59/109) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Denmark (2012) [28] | \n27.8 | \n0.4 | \n0.1 | \n0 | \n6.3 | \n2.7 | \n13.5 | \n4.2 | \n0 | \n
\n | (55/198) | \n\n | \n | \n | \n | \n | \n | \n | \n |
China (2013) [21] | \n38.5 | \n25.4 | \n– | \n– | \n3.9 | \n21.6 | \n1.9 | \n– | \n– | \n
\n | (59/153) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Iran (2013) [27] | \n22.9 | \n22.9* | \n14.3 | \n– | \n– | \n– | \n– | \n– | \n– | \n
\n | (16/70) | \n\n | \n | \n | \n | \n | \n | \n | \n |
France (2015) [29] | \n1.7 | \n– | \n– | \n– | \n– | \n– | \n1.7 | \n– | \n– | \n
\n | (3/184) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Russia (2014) [30] | \n17.7 | \n– | \n– | \n– | \n3.4 | \n5.2* | \n6.5 | \n– | \n– | \n
\n | (41/232) | \n\n | \n | \n | \n | \n | \n | \n | \n |
Summary of HHVs infection profiles from selected studies.
HHV‐6A/B does not discriminate between HHV‐6A and HHV‐6B in all of the references.
* Significant difference of sperm count or sperm motility in infertile and fertile group.
TK (EC 2.7.1.21) is a key enzyme in the pyrimidine salvage pathway that catalyzes the transfer of the ATP γ‐phosphate to thymidine to produce dTMP. HHV‐1‐TK is a phosphotransferase specifically required for viral DNA synthesis. HHV‐1‐TK shows broad substrate specificity, including pyrimidines (thymidine, deoxycytidine) and their analogs (azidothymidine), as well as purines (guanosine) and their analogs (acyclovir, ganciclovir [GCV], buciclovir and penciclovir). The ability to transform synthetic precursors, such as GCV, into toxic nucleotide analogs has resulted in effective cancer therapy agents [32, 33]. GCV consists of a guanine linked to an acyclic sugar moiety at position 3’. HHV‐1‐TK converts GCV into GCV‐monophosphate, which is further phosphorylated into GCV‐triphosphate by the host\'s kinases. GCV‐triphosphate has high affinity for DNA polymerase and functions as a competitive inhibitor of guanosine triphosphate (dGTP), becoming incorporated into the nascent DNA strand (Figure 2). GCV causes a distortion in the DNA sugar phosphate backbone, which blocks DNA replication [34–36] and induces cell apoptosis, thus selectively eliminating HHV‐1‐TK‐positive cells [37, 38]. Moreover, HHV‐1‐TK‐expressing cells trigger apoptosis of neighboring cells due to the transfer of GCV‐triphosphate via gap junctions in a phenomenon called “bystander killing” [39]. Therefore, HHV‐1‐TK can be used as a marker/reporter gene for removing specific target cells.
Incorporation of GCV disrupts extension of replicating DNA strands. In HHV‐1‐TK‐positive cells, GCV is phosphorylated into GCV‐monophosphate and then converted into GCV‐triphosphate. GCV‐triphosphate has high affinity for DNA polymerase and is utilized as a dGTP analog. However, the acyclic sugar moiety distorts the sugar phosphate backbone, resulting in irregular extension of the replicating DNA strand.
The use of HHV‐1‐TK offers a valuable tool for the ablation of specific cell types as well as in gene therapy. In the early 1990s, Tg rats were engineered to express HHV‐1‐TK under the control of the FSHβ promoter (Figure 3A) in the anterior lobe of the pituitary gland [12]. However, the experiment was interrupted due to observed infertility of male rats. The same happened with Tg mice [40]. Moreover, Ellison and Bishop showed that ectopic expression of HHV‐1‐TK fused to the human immunodeficiency virus long terminal repeat (HIV‐1‐LTR) gene caused male infertility in Tg mice [41]. In the present work, we observed ectopic expression of HHV‐1‐TK gene in the testis of Tg rats (Figure 3B).
Ectopic expression of HHV‐1‐TK in transgenic (Tg) rats. (A) HHV‐1‐TK (blue box) was fused to the 5’‐upstream region (-852/+10 bp) of porcine FSHβ (light blue). Arrows indicate the region amplified using specific primer sets (FSH‐tk‐I and FSH‐tk‐II); product sizes are shown in parentheses. (B) Total RNAs of pituitary, gonads (testis and ovary), cerebellum, liver, kidney, adrenal glands, prostate, and uterus of Tg rats (male and female) were analyzed by real‐time polymerase chain reaction (RT‐PCR) using specific primers for rat FSHβ subunit, HHV‐1‐TK (I), HHV‐1‐TK (II) and cyclophilin A (control). Reproduced and modified from [12] with permission from the Society for Reproduction and Development.
Whereas the testes of 3‐month‐old normal rats exhibited morphologically normal germ cells through all stages of maturation, histological analysis of Tg rats revealed developmentally arrested spermatozoa and multinuclear cells, together with altered elongated spermatozoa (Figure 4) [12]. Loss of sperm motility and viability was also observed (Figure 4F). Compared with normal rats, testis and epididymis weights were also decreased by 35 and 57%, respectively. In contrast, prostate and seminal vesicles weights were similar [12].
\nHistological analysis of rat testis and motility of epididymal spermatozoa. (A–D) Sections stained with hematoxylin and eosin showing testicular spermatogenesis in normal (A and B) and Tg (C and D) 3‐month‐old rats. Arrowhead, elongated arrested round spermatid; arrow, multinucleated cell. (E and F) Spermatozoa obtained from the epididymis of normal (E) and Tg (F) rats stained with SYBR‐14 and propidium iodide from a LIVE/DEAD sperm viability kit. Green and red indicate live and dead spermatozoa, respectively. Reproduced and modified from [12] with permission from the Society for Reproduction and Development.
Histological analysis of testis and epididymis in normal and Tg rats. A–D. Sections of normal (A) and 3‐, 6‐, and 12‐month‐old (B–D, respectively) Tg rat testis stained with hematoxylin and eosin. (E–H) Sections of normal (E) and 3‐, 6‐ and 12‐month‐old (F–H, respectively) Tg rat epididymis stained with hematoxylin and eosin. Abbreviations: spermatogonium (Sg), pachytene spermatocyte (pSc), immature spermatozoon (iSz), spermatid (Sp), round spermatid (rSp), elongated spermatid (eSp), degenerated spermatid (dSp), multinucleated round cell (mRc), and degenerated round cell (dRc). Reproduced and modified from [13] with permission from the European Teratology Society.
Histological analysis revealed that in Tg rats abnormal spermatogenesis could be observed as early as 3 months in development. Although spermatocytogenesis and meiosis seemed to progress normally (Figure 5B), the number of spermatozoa in the epididymis decreased dramatically (Figure 5F). Furthermore, large numbers of degenerated germ cells and immature/malformed spermatozoa were present (Figure 5F), indicating that maturation was likely to be disrupted. Degenerated germ cells with large vacuoles and cells lost throughout the tubules were observed in the testis of 6‐month‐old Tg rats (Figure 5C). In 12‐month‐old Tg rats there was a complete loss of germ cells and only Sertoli cells remained in the tubules (Figure 5D). In the epididymis of 6‐ and 12‐month‐old Tg rats, spermatozoa could hardly be observed (Figure 5G–H). Thus, Tg rats showed a spermatid‐stage‐specific defect in maturation and an age‐dependent loss of germ cells.
\nWhereas spermatocytogenesis and meiosis were confirmed in Tg rats, degeneration of germ cells by necrosis and apoptosis was frequently observed in the seminiferous tubules. In addition, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay showed a stage‐independent increase in TUNEL‐positive tubules and cells (Figure 6).
\nTerminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay of spermatozoa. (A–D) TUNEL assay of normal (A and C) and Tg (B and D) 3‐month‐old rats. Representative TUNEL‐positive cells are indicated by arrowheads. (E) Percentage of TUNEL‐positive tubules. (F) Mean number of TUNEL‐positive cells per TUNEL‐positive tubule (**p < 0.01). (G) TUNEL‐positive tubules of normal (N) and Tg (T) rats at stages XI‐I and II‐X (*p < 0.05). Reproduced and modified from [13] with permission from the European Teratology Society.
Tg rat testes (3‐ and 6‐month‐old) were simultaneously stained with anti‐HHV‐1‐TK and nuclear DAPI stain (Figure 7). We observed that HHV‐1‐TK protein accumulated in the cytoplasm of postmeiotic spermatids while it was absent from spermatogonia, spermatocytes, Leydig cells and Sertoli cells. Furthermore, germ cells were obviously reduced in 6‐month‐old Tg rat testes (Figure 7B-C).
Immunohistochemistry of Tg rat testis. Immunostaining of 3‐month‐old Tg rat testis with anti‐HHV‐1‐TK antibody and nuclear DAPI stain. HHV‐1‐TK localizes to the cytoplasm of round spermatids, but not to other testis cells. (B and C). Immunostaining of 6‐month‐old Tg rat testis. Germ cells decrease markedly. HHV‐1‐TK is observed in round spermatids (B) but not in Sertoli cells (C). Abbreviations: spermatogonium (Sg), spermatocyte (Sc), elongated spermatid (eSp), round spermatid (rSp), Leydig cell (LC), and Sertoli cell (SerC). Reproduced and modified from [13] with permission from the European Teratology Society.
Ectopic expression of HHV‐1‐TK protein was confirmed in postmeiotic spermatids by immunohistochemistry. This suggested the existence of alternative promoters directing specific and ectopic expression in postmeiotic spermatids. Western blot analysis of HHV‐1‐TK in Tg rat testis revealed for the first time two bands at 37 and 39 kDa, corresponding to truncated products of the full‐size 43 kDa HHV‐1‐TK protein (Figure 8A) [13].
\nAnalysis of the transcription start site by RNA ligase‐mediated rapid amplification of cDNA ends (5’‐RLM RACE) (Figure 8B) showed that the start site corresponded to the first intron of the HHV‐1‐TK gene (Figure 8C). This suggested that the second in frame ATG might have produced the 37 kDa band. Ellison and Bishop reported that HIV‐1‐LTR‐driven HHV‐1‐TK ectopic expression was abolished by deleting the region between the multiple cloning site and the second ATG [41]. They suggested that removing this portion conferred DNA methylation‐independent expression in the testis. Indeed, the corresponding region (144 bases) is GC‐rich (67%) and contains 17 CpG sites. Currently, a HHV‐1‐TK vector free of the GC‐rich region is available.
Identification of HHV‐1‐TK isoforms and transcription initiation sites in Tg rat testis. (A) Detection of HHV‐1‐TK in Tg rat testis by western blot analysis reveals two truncated isoforms (37 and 39 kDa) that do not correspond to the canonical form (43 kDa). (B) 5’‐RLM‐RACE analysis. Lane 1, PCR using a 5’‐RACE outer primer set; lane 2, PCR with a 5’‐RACE inner primer set. Products (approximately 300 bp) are indicated with an arrowhead. (C) Diagram showing the HHV‐1‐TK transcription initiation site (arrow), translation start sites (arrowheads and ATG with residue number), stop codon (grey box) and poly A signal (solid box). The closed box indicates a putative testis‐specific promoter region within HHV‐1‐TK. Reproduced and modified from [13] with permission from the European Teratology Society.
Ectopic, testis‐specific, expression of HHV‐1‐TK in Tg rats and mice raises the possibility that the same mechanism could also affect humans carrying HHV‐1. Therefore, the transcriptional regulation exhibited by ectopic expression of HHV‐1‐TK in Tg rats and the effects of HHV‐1‐TK accumulation on spermatogenesis may be revelatory of the same mechanisms in humans.
\nScanning electron microscopy revealed abnormal ultrastructure of epididymal spermatozoa in 10‐week‐old Tg rats [13]. We observed that the head and tail regions of spermatozoa from 10‐week‐old normal rats exhibited smooth surface and a regular morphological conformation (Figure 9A). Indeed, the morphology of these spermatozoa was different from those of Tg rats (Figure 9B–F). Spermatozoa with malformed heads, the consequence of a defective acrosome or reduced genome, were observed at high frequency among Tg rats (Figure 9B–F). Transmission electron microscopy (TEM) revealed that the cell membrane was missing from segments of the head (Figure 9C), midpiece (Figure 9D), and flagellum (Figure 9F). There were spermatozoa with looped tails (Figure 9D).
Scanning electron microscopy of Tg rat spermatozoa. Spermatozoa from caudal epididymis of 10‐week‐old rats visualized by scanning electron microscopy. (A) Normal rat sperm shows smooth head and tail. B–F. Tg rats present a deformed head (B), a microhead and absence of cell membrane (C), looped flagella and absence of cell membrane (D), defective acrosome and reduced genome (E), and absence of cell membrane in the flagellum (F). Reproduced and modified from [13] with permission from the European Teratology Society.
TEM of 3‐month‐old rat testis showed that in contrast with normal rats (Figure 10A–C), Tg rats displayed massive vacuoles within the seminiferous epithelium (Figure 10D–E, asterisks). These frequently presented degenerated spermatocytes in the seminiferous tubules (Figure 10E–F, arrowheads), in spite of confirmed spermatocytogenesis and meiosis. Moreover, TEM also showed elongated spermatids with disorganized heads partially devoid of cell membrane (Figure 10G, arrowhead), multiple flagella (Figure 10H, arrow), and absence of the inner arms of flagellar axonemes (Figure 10I, arrowheads) in the lumen of seminiferous tubules. In addition, vacuolization was also observed in the cytoplasm of spermatocytes (Figure 10F).
TEM of rat germ cells. Sections of 3‐month‐old normal (A–C) and Tg (D–I) rats analyzed by TEM. (A–C) Normal condensed nuclear, midpiece, and principle piece are indicated by arrows. (D–F) Numerous phagocytic vacuole compartments and massive vacuoles are present within the cytoplasm of Sertoli cells (D–E, asterisks), together with degeneration of spermatocytes (E–F, arrowheads). (G–I) Spermatozoa within the lumen of seminiferous tubules show various ultrastructural abnormalities, such as disorganized heads with a disrupted cell membrane (G, arrowhead), multiple flagella (H–I, arrow), and absence of inner arms of flagellar axonemes (I, arrowheads). Abbreviations: normal condensed nuclear (Nu), midpiece (Mp), principle piece (Pp), spermatogonium (Sg), spermatocyte (Sc), Sertoli cell (SerC), round spermatid (rSp), elongated spermatid (eSp). Reproduced and modified from [42] with permission from the Society for Reproduction and Development.
TEM also revealed the low number of spermatozoa in the epididymis of Tg rats (Figure 11B) as compared to normal rats (Figure 11A). Spermatozoa from normal rats displayed intact cell membranes and normal‐shaped heads with complete chromatin condensation (Figure 11A1). Instead, those of Tg rats presented a number of defects: several immature spermatids detaching from Sertoli cells and sloughing into the epididymis (Figure 11B), degenerated round spermatids (Figure 11B1) and many types of abnormal elongated spermatids (Figure 11B2–B4). In addition, whereas normal rats presented the typically assembled flagellar axonemes composed of nine outer doublet microtubules and a pair of central microtubules (Figure 11A2), most spermatozoa from Tg rats were dead and showed various ultrastructural defects, including breakage of the surface membrane (Figure 11B5) and a decline in the number of outer dense fibers (Figure 11B6).
TEM of epididymal spermatozoa. TEM sections of epididymal spermatozoa from 3‐month‐old rats. (A) Spermatozoa from normal rats show intact acrosomes with condensed nuclear material (A1) as well as proper assembled flagellar axonemes displaying nine outer doublet microtubules and a pair of central microtubules (A2, arrowhead). (B) Spermatozoa in the epididymis of Tg rats appear dead, and display various ultrastructural defects, such as absence of acrosomes, disorganization of surface membranes, and degeneration of midpiece and tail (B1–B4). In addition, residual bodies of degenerated germ cells (B5) and missing outer doublet microtubules are frequently observed (B6, arrowhead). Reproduced and modified from [42] with permission from the Society for Reproduction and Development.
Hematoxylin and eosin staining revealed various abnormal morphologies in Tg rat testis. Cytoplasmic vacuolation of Sertoli cells (Figure 12A, open arrowhead), disconnection between Sertoli and germ cells (Figure 12B, arrows), and disordered arrangement of elongated spermatids (Figure 12C, arrowhead) were present in many seminiferous tubules. In addition, immature spermatozoa falling off from the seminiferous epithelium were observed during all stages of spermatogenesis (Figure 12D–H, arrowheads). Furthermore, sperm development was disrupted and germ cells with different levels of maturity were observed simultaneously (Figure 12D–H). These results indicate that cell junctions between germ and Sertoli cells may have been affected. This hypothesis may be confirmed by analyzing gene expression profiles of Tg rat testis.
Histological analysis of Tg rat seminiferous tubules. Sections of 3‐month‐old Tg rat testis stained with hematoxylin and eosin. The following defects are observed: cytoplasmic vacuolation of Sertoli cells (A, open arrowhead), disconnection between Sertoli and germ cells (B, arrows), and disordered arrangement of elongated spermatids (C, solid arrowhead). (D–H) Immature spermatozoa falling off from the seminiferous epithelium (arrowheads) are observed at different maturation stages (I–XIV) and germ cell development is disrupted.
Changes in gene expression evoked by HHV‐1‐TK accumulation in Tg rat testis were examined by cDNA microarray analysis. We found that 200 genes, 0.67% of all transcripts on DNA chips, were differently expressed between Tg and normal rats. We sorted the genes by their functional categories, such as apoptosis, cell cycle, development, oxidative stress, proteolysis, signal transduction, transcription, translation, transport, metabolism, immune response, and cell adhesion. The highest number of affected genes was linked to metabolism, with 8 genes up‐regulated and 16 down‐regulated by at least 1.5‐fold in Tg rat testis (Table 3) [42].
\nRT‐PCR was performed for 10 genes involved in cell adhesion, signal transduction, and transport: Cnot7, Fgf7, Egfl6, Testin, Ostf1, and Atp2a2 (all up‐regulated); and Sh3bp4, Lamc2, Versican, and Mamdc1 (all down‐regulated). As listed in Table 4, Cnot7, Fgf7, Testin, Ostf1, Versican, and Mamdc1 were significantly up‐ or down‐regulated, confirming microarray data. Testin is a signaling molecule responsible for monitoring Sertoli‐germ cell adherens junctions. Increased secretion of testin by Sertoli cells causes disruption of these junctions [43, 44]. Versican is expressed in adult mouse brain, heart, lung, spleen, skeletal muscle, skin, tail, kidney, and testis. It is suggested to play a role in cellular attachment, migration, and proliferation by interacting with cell surfaces and extracellular matrix molecules [45]. Mamdc1, a novel member of adhesion molecules of the immunoglobulin superfamily, is expressed in human Leydig cells of the testis. Mamdc1 mRNA was shown to be up‐regulated by pro‐inflammatory cytokines, such as tumor necrosis factor‐α and interferon‐γ, involved in cell adhesion, migration, and recruitment to inflammatory sites [46]. However, the function of versican and mamdc1 in spermatogenesis has not yet been established. Expression of Fgf7, Ostf1, and Cnot7 [47] also increased significantly, indicating a stress response in Tg rat testis. The roles of these genes are listed in Table 5.
\nBiological process | \nGene number of expression changed | \n||
---|---|---|---|
Decreased | \nIncreased | \nTotal | \n|
Apoptosis | \n1 | \n4 | \n5 | \n
Cell cycle | \n2 | \n4 | \n6 | \n
Development | \n3 | \n16 | \n19 | \n
Oxidative stress | \n0 | \n1 | \n1 | \n
Proteolysis | \n3 | \n7 | \n10 | \n
Signal transduction | \n2 | \n11 | \n13 | \n
Transcription | \n6 | \n6 | \n12 | \n
Translation | \n1 | \n1 | \n2 | \n
Transport | \n12 | \n11 | \n23 | \n
Immune response | \n1 | \n2 | \n3 | \n
Metabolism | \n8 | \n16 | \n24 | \n
Cell adhesion | \n3 | \n4 | \n7 | \n
Others | \n54 | \n21 | \n75 | \n
Total | \n96 | \n104 | \n200 | \n
Number of affected genes in testes of HHV‐1‐TK Tg rats revealed by cDNA microarray.
Note: The affected genes were classified into 12 functional categories according to “Gene ontology”.
Gene ontology classification of genes whose expression changes by at least .5‐fold in Tg with respect to normal rats, as revealed by cDNA microarray analysis. Reproduced and modified from [42] with permission from the Society for Reproduction and Development.
Notably, the contraceptive adjudin has been reported to induce morphological alterations in the seminiferous tubules similar to the ones we observed here, and target directly testin and actin‐myosin [48]. However, the exact mechanism by which HHV‐1‐TK disrupts spermatogenesis and adherens junctions will require further work.
\nGene product | \n\n | Microarray analysis | \nReal‐time PCR results | \n||
---|---|---|---|---|---|
Gene title | \nGene symbol | \nCategory | \nRatio of TG value/N value | \nExpression levels TG/N | \np value | \n
Up‐regulated genes in TG rat testes | \n\n | \n | \n | \n | \n |
CCR4‐NOT transcription complex, subunit 7 | \nCnot7 | \nSignal transduction | \n18.40 | \n1.30 | \n<0.05 | \n
Fibroblast growth factor 7 | \nFgf7 | \nDevelopment | \n6.10 | \n2.00 | \n<0.05 | \n
EGF‐like‐domain, multiple 6 | \nEgfl6 | \nCell adhesion | \n3.70 | \n0.91 | \nNS | \n
Testin gene | \nTestin | \nCell adhesion | \n2.30 | \n2.00 | \n<0.05 | \n
Osteoclast stimulating factor 1 | \nOstf1 | \nTranscription | \n1.68 | \n1.30 | \n<0.05 | \n
ATPase, Ca2+ transporting, cardiac muscle, slow twitch 2 | \nAtp2a2 | \nTransport | \n1.67 | \n0.96 | \nNS | \n
Down‐regulated genes in TG rat testes | \n\n | \n | \n | \n | \n |
SH3‐domain binding protein 4 | \nSh3bp4 | \nTransport | \n0.06 | \n0.91 | \nNS | \n
Lamimin, gamma 2 | \nLamc2 | \nCell adhesion | \n0.09 | \n0.90 | \nNS | \n
Chondroitin sulfate proteoglycan 2 | \nVersican | \nCell adhesion | \n0.10 | \n0.55 | \n<0.05 | \n
MAM domain containing glycosylphosphatidylinositol anchor 2 | \nMamdc1 | \nCell adhesion | \n0.62 | \n0.67 | \n<0.05 | \n
Expression level of genes assayed by microarray analysis and real‐time PCR.
Reproduced and modified from [42] with permission from the Society for Reproduction and Development.
Gene name | \nGene symbol | \nCategory | \nMolecular function | \nBiological process | \n
---|---|---|---|---|
Up‐regulated genes in TG rat testes | \n||||
CCR4‐NOT transcription complex, subunit 7 | \nCnot7 | \nSignal transduction | \nHas 3‐5 poly(A) exoribonuclease activity, nucleic acid binding, transcription factor activity, catalytic component of the CCR4‐NOT complex, cell proliferation factor, mRNA degradation, miRNA‐mediated repression | \nspermatogenesis, tumor cell metastasis suppressor, bone metabolism, embryonic development, tumor suppressor | \n
Fibroblast growth factor 7 | \nFgf7 | \nDevelopment | \nchemoattractant activity, growth factor activity, mitogenic and cell survival activities activation of MAPKK activity actin cytoskeleton reorganization neurotrophin TRK receptor signaling pathway | \nembryonic development, cell growth, morphogenesis, tissue repair, tumor growth and invasion | \n
Testin gene | \nTestin | \nCell adhesion | \nregulation of cell motility, poly(A) RNA binding, zinc ion binding | \ncell adhesion, cell spreading, reorganization of the actin cytoskeleton, negative regulation of cell proliferation, tumor suppressor | \n
Osteoclast stimulating factor 1 | \nOstf1 | \nTransport | \nSH3 domain binding | \ninduces bone resorption, enhancing osteoclast formation | \n
Down‐regulated genes in TG rat testes | \n\n | |||
Chondroitin sulfate proteoglycan 2 | \nVersican | \nCell adhesion | \ninteraction of integrins, anti‐cell adhesion, regulation of cell motility, growth and differentiation, a key factor in inflammation hyaluronic acid binding, | \nembryonic development, embryonic cell migration important in the formation of the heart, neural crest cell migration, inhibit nervous system regeneration, axonal growth, tumor growth | \n
MAM domain containing glycosyphosphatidylinositol anchor 2 | \nMamdc1 | \nCell adhesion | \nGPI‐anchored protein, cell‐cell interaction | \nneuron development and differentiation, neuronal migration, axon outgrowth, axon‐target recognition | \n
The role of genes changed expression level significantly in Tg rats.
Nested PCR for 4 types of HHVs was performed on 153 DNA samples prepared from human semen (Figure 13). Bands of 99, 150, 165, and 135 bp were observed for HHV‐1, HHV‐4, HHV‐5, and HHV‐6A/B (HHV‐6A and 6B were detected together by common domains), respectively (Figure 13A). Samples showing positive bands by more than one PCR target indicated concomitant infection with multiple types of HHVs (Figure 13A). We identified nucleotide sequences corresponding to HHV‐1, HHV‐4, HHV‐5, and HHV‐6A/B (Figure 13B) in 39, 6, 33, and 3 patients, respectively, as summarized in Table 6. We observed double infection with HHV‐1/HHV‐5 (15 carriers), HHV‐1/HHV‐4 (1), HHV‐1/HHV‐6A/B (2) and HHV‐4/HHV‐5 (4). HHV‐4 carriers presented a double infection 83% of the time, while for other HHVs it ranged from 46 to 66%.
Detection of HHVs genomic DNA in human semen by PCR. (A) Agarose gel electrophoresis of PCR products using HHV‐specific primers and human semen as template DNA. PC (positive control) and NC (negative control) indicate reactions with control viral fragments and without DNA template, respectively. M, size marker. (B) Sequence profiles of representative PCR products. Viral sequences are shown below the nucleotide sequence. Reproduced and modified from [21] with permission from the Society for Reproduction and Development.
Number of virus infection in patients (153) | \n||||
---|---|---|---|---|
Type | \nHHV‐1 | \nHHV‐4 | \nHHV‐5 | \nHHV‐6A/B | \n
Total | \n39 (25%) | \n6 (4%) | \n33 (22%) | \n3 (2%) | \n
Number of double virus infection | \n||||
Type | \nHHV‐1 | \nHHV‐4 | \nHHV‐5 | \nHHV‐6A/B | \n
HHV‐1 | \n– | \n1 | \n15 | \n2 | \n
HHV‐4 | \n1 | \n– | \n4 | \n0 | \n
HHV‐5 | \n15 | \n4 | \n– | \n0 | \n
HHV‐6A/B | \n2 | \n0 | \n0 | \n– | \n
Total | \n18/39 (46%) | \n5/6 (83%) | \n19/33 (58%) | \n2/3 (66%) | \n
Number of cases with single and double viral infection.
HHV‐6A and 6B were detected together by common domains.
Reproduced and modified from [21] with permission from the Society for Reproduction and Development.
The viral DNA‐positive group showed a higher incidence of oligozoospermia compared with the viral DNA‐negative group (38.5 vs. 16.7%, P < 0.05; Figure 14). Therefore, to further confirm the relationship between HHVs infection and changes to semen parameters, large‐scale analysis according to WHO Guidelines (5th edition) is strongly suggested.
Comparison of oligozoospermia between viral DNA‐negative and ‐positive groups. Analysis of oligozoospermia rates between viral DNA‐negative (36 patients) and ‐positive (39 patients) according to WHO Guidelines (5th edition). Reproduced and modified from [21] with permission from the Society for Reproduction and Development.
HHV‐1 infection might strongly associate with human male infertility, possibly by testis‐specific expression of the viral TK gene. However, conclusive evidence that HHV‐1‐TK causes male infertility is still missing.
\nAs described above, HHV‐1‐TK is known as a suicide gene that kills target cells specifically in the presence of GCV. However, male infertility normally occurs in the presence of HHV‐1 but absence of GCV. Degeneration of spermatogenesis was observed as early as 3 months in Tg rats ectopically expressing HHV‐1‐TK. With the exception of Sertoli cells, it later resulted in loss of germ cells, possibly by “bystander killing.” In contrast, no abnormality in the pituitary gland of Tg rats was observed. This difference may indicate tissue‐specific action of HHV‐1‐TK in the testis, which does not occur in the pituitary gland. At present, it is unclear why HHV‐1‐TK targets testis over other tissues and how it induces degeneration of spermatogenesis. Further work is required to elucidate this and other features of HHV‐related infertility.
\nExpression of HHV‐1‐TK may be the main cause of infertility in HHV‐1‐infected human males. To firmly establish a causative link, several players require identification: (1) target molecule(s) of HHV‐1‐TK; (2) the HHV‐1‐TK‐dependent mechanism responsible for failed spermatogenesis; and (3) the mechanism dictating postmeiotic spermatid‐specific expression of HHV‐1‐TK. The identification of target molecule(s) is particularly important, since there is no evidence that enzymatically active HHV‐1‐TK is required, raising speculation it may only serve as a binding protein. Even if HHV‐1‐TK functioned as an enzyme, the substrate(s) might be testis‐specific. Currently, the only known activity of HHV‐1‐TK is to catalyze the transfer of the γ‐phosphate from ATP to substrate nucleotides and analogs. Recently, a stable‐isotope substrate ([γ‐18O4]ATP) for kinases has been developed [49]. Mass spectrometry could reveal the identity of [γ‐18O4]ATP‐labeled products obtained by incubating recombinant HHV‐1‐TK with testis homogenates. The identified HHV‐1‐TK target(s) may help elucidate the mechanisms determining failed spermatogenesis and postmeiotic spermatid‐specific expression of HHV‐1‐TK. Moreover, knowing the target(s) molecular properties may help design appropriate analogs for pharmacological use. The significance of our results is particularly clear in view of expected novel male contraceptives and drugs for HHV‐related male infertility.
\nTextiles are susceptible to micro-organisms and their products (e.g. toxins) known to cause infection and intoxication. Myriad textile products such as herbal textiles and clothing are available in the market to support healthy lifestyle and hygiene. Most textile products are meant to come in contact with the skin; hence, herbally treated fabrics are better choice as they do not harbor harmful chemicals and thus are ecofriendly. People of yore used to live in harmony with nature and passionately protected their environment; however, nowadays, excessive exploitation of natural resources has led to a phenomenal increase in environmental pollution. This is taking a serious toll on the lives of all living beings on the planet earth. Right kind of fabric is of paramount importance to a healthy lifestyle, and synthetic fabrics’ teeming with chemicals and dyes poses severe health threat. One would not like to eat a bowl of pesticide-drenched food. Then, why wear clothing doused in chemicals? Textile dyes obtained from natural sources are usually friendly to human skin and are biodegradable and ecofriendly. Some natural dyes have medicinal properties and impart healing qualities to the fabric. They are ecofriendly, save energy, provide rural employment, and preserve traditional craftsmanship [1].
Clothing is our second skin and plays an important role in human life. Textiles finished or dyed from extracts of various medicinal herbs are called herbal textiles. Textiles and clothing infused with medicinal herbs are becoming popular, especially in urban India. The use of such products helps in providing protection against myriad biological toxins and pathogens. Customers demand for new ranges of technology, owing to upward trend in enhancing beauty, that provide better esthetics along with improved health and a sense of relief and satisfaction during use. This trend has produced Cosmetotextiles.
Among technical textiles, medical textiles are a promising product, which plays an important role in maintaining the human health. Medical textiles consist of textiles used in operative and postoperative tasks on and around the patient and medical practitioners. They are broadly classified as nonimplantable materials, implantable materials, extracorporeal devices, hygiene products, protective, and healthcare textiles. Healthcare, disposable, and nondisposable hygiene products mainly used in hospitals to reduce the chances of contamination by biological toxins and infectious organisms.
India has good potential to produce natural dyes commercially as it is considered as 1 of the 17 mega diversity countries globally [2]. Traditional knowledge facilitates the use of plants and their products for healing the wounds and burn injuries and treating the skin infections. Herbal extracts are being used successfully for imparting antimicrobial finish to textiles [3].
Herbal products derived from plants are gaining popularity worldwide for use in textiles on account of their easy availability, green approach, low toxicity, biocompatibility, and ecofriendly nature [2, 3]. Owing to market demand for textile products having added comfort and functional properties, herbal products may be explored for their potential benefits to produce such products. This mandates scientists to devise methods and techniques to use plants containing bioactive agent in producing value added and more attractive functional textile substrates. Insect repellent, deodorizing/aroma, UV protection, antimicrobial, and flame retardant are some of the new properties, which have been imparted recently to textiles. In recent years, the growing demand for herbal products has led to the idea of developing healthcare textile products using herbal extracts.
Currently, herbal colorants are the most attractive option because of high biodegradability, low toxicity, green chemistry, and potential application in the textile dyeing and finishing industry. Natural colorants from plant sources are considered as novel agents in imparting multifunctional properties such as antimicrobial, insect repellent, deodorizing, and UV protection to textiles. Many natural colorants possess some inherent functions in addition to their coloring attribute. These inherent properties are transferred to the textile substrate dyed with an herbal colorant. Alternatively, dyeing textiles with these herbal colorants can combine dyeing with functionality finishes, an ecofriendly process using less water and energy than currently used separate wet treatments. Recently, increased interest in the use of natural dyes in textile dyeing has been observed due to enforcement of the stringent environmental regulations by many countries because of toxic effects of synthetic dyes. This review gives an overview of the herbal methods available at present for dyeing textile materials with aim to provide other useful finishes [4].
Textiles dyed exclusively with herbal extracts without using any chemicals are called herbal textiles. The herbs utilized in dyeing are different from vegetable dyes as they also have medicinal values. These herbs are applied to the fabric using natural ingredients in order to preserve medicinal properties. Further, bleaching of cloth is done by exposing it to sunlight without the use of any chemical bleach. The concept of herbal textiles has been derived from Ayurveda, the ancient Indian method of vedic healthcare. Ayurvastra is a branch of Ayurveda. In Sanskrit, ayur means health, and veda means wisdom, while vastra means clothing when it combines the meaning life cloth [5].
It is made by embedding essential oils and herbs into textiles. A wide range of herbs such as turmeric, tulsi, neem, rose, Indigo, lemon grass, castor, and ber can be used for this process providing medicinal value and aroma to the fabric. Ayurvastra (marketed as cosmetotextiles) has been used for curing several diseases such as diabetes, arthritis, skin infections, hypertension, and hay fever [5].
Herbal clothing is an ancient technique of dyeing textiles in medicinal herbs. This alternative method of treatment originated more than 5000 years ago in India. Its core principles are rooted in the Vedas, which are an ancient book of Hindus. The Vedas date back to around 3000 BC. The science of life or Ayurveda dates back to around 1000 BC. The origins of Ayurveda are also found in the Atharvaveda. It contains incantations and hymns that cure various diseases through mantra. Mythology says that Ayurveda was given to Dhanvantri by Lord Brahma. Charaka Samhita and Sushruta Samhita are the oldest known Ayurvedic texts [6]. About 100 years ago, people from different parts of India were using various forms of natural dyeing in which they repeatedly dipped clothes in herb-based preparations following each wash [7]. These herbs are used traditionally in Ayurvedic treatment and many medical applications. When this fabric is worn, the medicinal property of the fabric is then transferred to the skin. The process of herbal dyeing was developed through extensive research during the age-old dyeing methods practiced since the days of the Indus civilization. Herbs were used to create different colors and to provide various health benefits. The natural/herbal fabrics are 100% organic, completely free of synthetic chemicals, and toxic irritants and are biodegradable [8].
Herbal clothing is prepared from organic cotton fabric impregnated with special herbs and oils for health benefits. Since ancient times, herbal clothing (Ayurvastra) is believed to cure diabetes, skin infections, psoriasis, hypertension, asthma, arthritis, rheumatism, high blood pressure, eczema, and cancer. Antimicrobial property or natural dyes such as indigo, pomegranate, myrobalan, and Indian madder of against bacteria such as Escherichia coli and Klebsiella pneumoniae have been reported [9]. Herbs such as harad, rakta chandana, shweta chandana, and kumkuma are used in Ayurveda for protecting the skin from harmful rays of sun. The herbal preparations act on melanocytes present in the skin, modulate their function, and block the effect of UV rays on the skin. References in different texts of Ayurveda are replete with examples of wearing pure cotton clothes impregnated with herbs such as chandana, haridra usheera, manjista, and nimbi aragwadha to safeguard human body from inclement weather and to keep it healthy. They used herbal products such as soap nut and lime for washing cloths before soaps and detergents were discovered. Herbal textiles and dyes have great vista of applications in diverse walks of human life. These herbal dyes have been used for centuries to produce colors for fabrics, yarns, leather, food, and so on. Natural dyes exhibit better biodegradation and have a better compatibility with the environment. The application of natural dyes has the potential to earn carbon credit by consumption of fossil fuel-based synthetic dye [10].
Skin is the largest sense organ in the human body (Figure 1). It can act as a barrier as well as a medium for entry of certain substances in the body. Several toxins and chemicals present in the environment get accumulated in the conventional clothing and may enter into the body through the skin. Hence, enhancing skin’s ability to resist entry of harmful chemicals and toxins into the body will improve the health. The radiated heat activates herbal molecules of the cloth and is reflected back to the body along with the herbal molecules. Through the blood, the herbal molecules enter deep into the body, show its effects at different levels of the body, and help in curing various diseases [5]. The skin can act not only as a protective barrier but also as a medium for outside substances to enter into the body. In the same way, the skin has the ability to absorb herbs found in our natural dyes. These herbs release their medicinal qualities in the body and strengthen the skin’s ability to block and resist the harmful substances. The close contact of herbal clothing next to the skin enhances the body metabolism, which leads to effective elimination of body toxins. The herb-infused and herb-dyed organic fabrics act as healing agents after the herbs being absorbed through the skin [6].
Structure of the human skin [6].
Skin temperature helps the herbal molecules to be absorbed into the body through sweat pores to manifest the desired medicinal effect.
The cloth should be in direct contact with the skin to manifest its curative effects.
Herbs are garden plants that are grown and harvested for culinary, aromatic, medicinal, and fibrous uses. Plant herbs are placed in the garden for their unmistakable fragrances, attractive textures, appealing colors, and variety of home uses. Due to current ecoconsciousness, there has been a revival of inherent in ecofriendly natural dyes throughout the world. The application of herbal products has given a new direction toward the treatments of various diseases through textile products. The herbs also do not pollute the environment through contamination of water resources in areas close to processing units. All kinds of shades of red, yellow, brown, orange, and green can be obtained with the help of the herbs.
Functional finishes using natural dyes have been in the vogue that promotes an ecofriendly lifestyle. Natural dyes are environmental friendly, nontoxic, noncarcinogenic, nonallergic, and renewable resources. Biosphere is being gifted with more than 500 plant species that yield the natural dyes [11]. The roots, flowers, leaves, seeds, and barks of more than 200 medicinal herbs, plants, flowers, roots, and barks are used to make the dyes. In herbal textiles, the color is gained from the medicinal preparation only, and no other colorants are used. As a result, its properties will last as long as the color is there. Since the herbs make beautiful shades, herbal wears are also becoming very popular for its primordial elegance.
Only herbal preparations devoid of any chemical are used in dyeing herbal textiles, whereas chemicals such as copper sulfate and ferrous sulfate are used as catalysts when dyeing is done with vegetable dyes. Thus, herbal dyes are different from vegetable dyes as they also impart some medicinal value. Herbal clothing is believed to help restore the balance within the body’s systems and strengthen the immune system. Some of the medi-herbs used in herbal textiles give the magical healing quality to the dyed fabric or yarn (Table 1) [12, 13, 14, 15]. Table 1 shows the diseases cured by different herbs.
S. no. | Name of the herb | Medicinal properties | Diseases claimed to be cured |
---|---|---|---|
1. | Neem (Azadirachta indica) | Antibacterial and antifungal | Common skin diseases, skin allergy, controls blood sugar levels |
2. | Curry leaves (Murraya koenigii) | Medicinal value | Beneficial in curing arthritis suppresses pain and reduces inflammation |
3. | Shoe flower (Hibiscus rosa-sinensis) | Controls blood sugars | Diabetes |
4. | Fenugreek/Methi (Trigonella foenum-graecum) | Medicinal value | Controls cholesterol, hypertension |
5. | Amla (Emblica officinalis) | Antifungal, antibacterial, antiviral | Boosts immunity |
6. | Acacia (Catechu) | Antifungal, antibacterial | Used for treatment of parasitic infestation and itching |
7. | Manjistha (Rubia cordifolia) | Medicinal value | Blood purifier and diuretic, leprosy |
8. | Cuscus/Khus khus grass (Chrysopogon zizanioides) | Medicinal value | It helps in fighting asthma |
9. | Arjuna (Terminalia arjuna) | Medicinal value | Cardiovascular support, hypertension, asthma |
10. | Pomegranate (Punica granatum) | Antibacterial, antiviral, astringent | Antimicrobial treatment, hypertension |
11. | Harad/myrobalan (Terminalia chebula) | Antiseptic, anti-inflammatory, controls cholesterol | Hypertension, anemia, asthma cures, wounds, ulcers |
12. | Aloe vera (Aloe barbadensis) | Antibacterial, antifungal | Burns, wounds, common skin diseases, psoriasis, anemia |
13. | Gurhal (Hibiscus rosa-sinensis) | Controls blood sugar levels | Diabetes |
14. | Jamun (Syzygium cumini) | Controls blood sugar levels | Diabetes controls blood sugar levels |
15. | Mahuva (Madhuca longifolia) | Controls blood sugar levels | Control diabetes |
16. | Champa flower (Magnolia champaca) | Controls blood sugar levels | Diabetes, anemia |
17. | Touch-me-not (Mimosa pudica), | Medicinal value | Controls blood sugar levels, diabetes |
18. | Eucalyptus (Eucalyptus globules) | Antiseptic | Relieve congestion, prevent infection, ease muscle soreness |
19. | Tulsi/basil (Ocimum tenuiflorum) | Antibacterial, antiseptic, antispasmodic, diaphoretic, febrifuge, nervine, controls cholesterol | Respiratory disorders, viral, bacterial infections. Controls blood sugar levels, boosts immunity, hypertension, HIV/AIDS |
20. | Chitosan | Antimicrobial | Naturally occurring biopolymer having antimicrobial properties |
21. | Balloon vine (Cardiospermum halicacabum) | Antiphlogistic, analgesic, blood refrigerant, anti-infectious | Cold, fever, renal edema, urinary tract infections,·furuncle, carbuncle, eczema, sprains, external wounds |
22. | Sandalwood (Santalum album) | Alterative, antibacterial, anti-inflammatory, antiseptic, astringent, carminative, disinfectant, diuretic, expectorant, hemostatic, refrigerant, sedative, stimulant | Headache, acute dermatitis, bronchitis, cystitis, eye diseases, gonorrhea, herpes zoster, infection, palpitations, sunstroke, urethritis cools the skin, curing skin itching, burns, its mild fragrance has a soothing effect that helps in fighting stress |
23. | Onion (Allium cepa) | Antioxidant, antimicrobial | Skin diseases, control diabetes |
24. | Indian Madder (Rubia cordifolia) | Antibacterial, anti-inflammatory, antiseptic, astringent, sedative, stimulant | It removes blood impurities and cures various skin diseases, HIV/AIDS |
25. | Indian Mulberry (Morinda pubescens) | Antibacterial, anti-inflammatory | It is blood purifier and used for wound healing, control diabetes |
26. | Saffron (Crocus sativus) | Medicinal value | Controls heart disease |
27. | Indigo (Indigofera tinctoria) | Antibacterial, antiallergy, antiseptic and aromatic, anti-inflammatory properties | Controls cholesterol, helps reducing the insulin resistance, thus controls diabetes, boosts immunity, anemia, skin diseases |
28. | Turmeric (Curcuma longa) | Antihepatotoxic, antioxidant, antimicrobial antiseptic, antiallergic | Fighting skin diseases and helps fight skin disease, anemia, psoriasis, liver disorders |
29. | Henna (Lawsonia inermis) | Blood purifier, anti-irritant, deodorant, antiseptic | Skin irritations such as heat rashes and skin allergies and to cool the body. Because of this cooling property, henna used as a prophylactic against skin diseases like burns, bruises, and skin inflammations, including sores from leprosy |
30. | Brazil wood (Caesalpinia echinata) | Blood purifier, anti-irritant | Skin diseases such as burns, bruises, and skin inflammations |
31. | Castor (Ricinus communis) | Blood purifier | Skin diseases |
32. | Baheda (Terminalia bellirica) | Controls cholesterol | Hypertension, anemia |
33. | Guar (Cyamopsis tetragonoloba) | Controls cholesterol | Hypertension |
34. | Safflower (Carthamus tinctorius) | Controls cholesterol | Hypertension, HIV/AIDS |
35. | Reetha (Sapindus trifoliatus) | Medicinal value | Psoriasis, skin disease |
36. | Amaltas (Cassia fistula) | Therapeutic effect | Psoriasis |
37. | Cluster fig (Ficus racemosa) | Therapeutic effect | Psoriasis |
38. | Winter cherry (Withania somnifera) | Therapeutic effect | Psoriasis, rheumatism |
Some of the medi-herbs used for herbal textiles.
Different parts of the plants namely root, bark, leaf, fruit, wood, seed, and flower are used to extract dyes. It is estimated that over 2000 pigments are found in various parts of plants; however, only about 150 have been commercially utilized [16]. Herbal cloth production by a particular method is accomplished by utilizing extracts of selected herbs for dyeing cotton/silk/linen, yarn, and fabric. Synthetic/chemical dyes are not used at any stage, and for washing, specific products are used [17].
The process of herbal dyeing was developed through extensive research during the age-old dyeing methods practiced since the days of Indus civilization. Manufacturing herbal textile begins with 100% hand-loomed organic cotton or silk wool, linen, jute, hemp, and so on and their natural blends that are dyed exclusively in herbal infusions for both color and health purposes. No chemical process is involved while dyeing and processing the fabric. The fabrics have to pass through various stages of treatment before they are turned into finished products. Herbal textiles are extra smooth and good for transpiration that helps in recovering from various diseases. Each fabric is infused with specific herbs, which are known to cure diseases. It may help to treat a broad range of diseases and skin infections such as eczema and psoriasis; the fabric also helps with conditions such as heart ailments, rheumatism, arthritis, blood pressure, diabetes, and respiratory conditions, such as asthma and sleeping disorders. It also helps in boosting immunity. The antibacterial and anti-inflammatory properties of herbal medicated clothing extend its use as dressings and bandages. It can also relieve the general body aches, stimulate the weight loss, and strengthen the immune system. It can also be used as energy booster; mood enhancer, for overall wellbeing, calming, and blood purification; and cooling [18]. The most effective time to wear herbal cured clothing is when the body is at rest, such as during sleep, relaxation, or meditation because this is when the body is naturally healing and re-establishing balance. Hence, most of herbal clothing products are sleepwear, bed sheets, towels, meditation clothes, and cotton mats [19].
The herbal textile production begins with pure organic yarn/fabric, and each step is carefully and meticulously controlled without the use of any chemical process in dyeing and processing. The process starts with the gray cloth going through various stages of treatment before it is dyed and becomes ready to wear. Only certified organic cotton, natural cotton, silk wool, linen, jute, hemp, and their natural blends should be used for herbal dyeing. The herbal dyeing process has been developed following the extensive research and is in practice since the Indus civilization. Machines are not used to prepare herbal clothing, and all the processes namely bleaching, dyeing, and other processes are accomplished in big vessels fabricated according to need. Further, no chemicals are added during spinning, weaving, or enhancing the appearance [20].
Most common fabric used is organic cotton, the cotton that is cultivated without chemical pesticides and fertilizers. Silk, wool, coir, linen, hemp banana, nettle, and bamboo are also used to prepare herbal clothing. For handloom, the process starts with the cotton yarn and goes through a process named desizing (to remove loose particles, debris, etc.). The common processes used in manufacturing of herbal clothing are detailed in the following paragraphs.
It involves washing of the processed gray cloth with mineral-rich water and sea salt to remove sizing, gums, and oils used in the weaving.
This process is actualized with biodegradable, organic cleaning agents, and surfactants and sun dried. The bleaching of cloth is done naturally by exposing it to sunlight on a natural grass base and by using animal manure. The fabric or yarn is first bleached using cow urine, milk, honey, along with biodegradable, naturally derived, organic cleaning agents, and surfactants such as Saptala (Acacia sinuata) and Phenila (Sapindus mukorossi) [21]. Many Ayurvedic doctors consider cow’s urine highly beneficial in balancing an individual’s “doshas” or basic constituents of an individual’s physiology and psychology, strengthening the immune systems and as an elixir in giving life [22]. The process ends with exposing the fabric or yarn directly to sunlight.
To make the colors look bright and retain color fastness, natural mordants such as myroballans, rubhabs leaves, oils, alum, fruit extracts of haritaki, bark of lodhra, minerals, and iron are used. Use of heavy metallic mordants such as copper, chrome, zinc, and tin is avoided as they are not ecofriendly. Mordanting is done before dyeing the fabric or yarn to make the colors look bright and retain fastness.
The word medication is used instead of dyeing because the medicinal herbs are used to impart required color to fabric or yarn [20, 23]. The yarn or fabric is then medicated in a carefully controlled mixture of herbal medicinal preparations depending on the disease or ailment being treated. Required herbs are applied directly to the fabric with the help of natural ingredients so as to keep the medicinal value of herbs intact [7, 11]. Steps involved are as follows:
The organic cotton yarn/fabric is dyed carefully with concoction of herbal dyes indicated for the treatment of desired disease(s).
The herbal dyes are prepared by careful blending of medicinal herbs derived from plants, flowers, roots, and barks.
The temperature, duration, number of the soaks, blend of herbs, and equipment are carefully controlled.
Desired herbs with the help of natural ingredients are applied directly on fabric in order to preserve the medicinal value of herbs.
Shades of red, yellow, brown, orange, and green can be obtained with careful use of herbs.
Finally, the medicated cloth is cooled and washed several times to remove the nonabsorbed particles and always dried in shade.
The finishing process used after herbal dyeing is also organic and is given by sprinkling pure water on the cloth and then stretching under pressure, using hand rolls, Aloe vera, castor oil, and so on [7, 11].
After dyeing, residue can be recycled. Solid and liquid wastes are separated by the filtration and used for farming purposes—as biomanure and for watering the fields and also to generate the biogas [11].
Herbal textile is made by embedding essential oils and herbal extract finishes into textiles. Such textiles have their applications in providing medicinal value and aroma to the garment. Microencapsulation, sensory perception technology, liposomes, dyes, and coated textile technology are used for incorporating the herbal drugs and oils in textiles. The temperatures of the finish, the duration and concentration of plant extract, the blend of herbs, and even the equipment used are carefully controlled [24].
Spray drying, air suspension coating, solvent evaporation, sol-gel microencapsulation, and in-situ polymerization are some commonly used techniques used for textile finishing by microencapsualtion. For encapsulation of flavor, fragrance, and oils, spray drying is used in which core particles dispersed in a polymer solution and sprayed into a hot chamber leading to solidification of shell particles on core material and polynuclear or matrix-type microcapsules are obtained. Air suspension coating is done by suspending solid particles of core material in a hot or cold upward moving steam of air. Coating material may be derived from cellulose derivatives, dextrins, emulsifiers, lipids, protein derivatives, and starch derivatives, which is atomized through nozzles into the chamber and deposits as a thin layer on the surface of suspended particles. Solvent evaporation is a process in which the drug is dissolved, dispersed, or emulsified in an external aqueous or oil phase; it has been used to impart antifungal finish to textiles. Sol-gel microencapsulation is used to produce flame retardant, water/oil repellent, UV protecting, antiwrinkle, self-cleaning, flavor and odor containing, and antibacterial textiles. Sol-gel synthesis involves the hydrolysis of silicone monomer and the condensation of silica into a porous structure with a three-dimensional networked structure, which can be tailored for a wide range of useful properties.
Microencapsulation, widely used in textile finishing, is a process in which small capsules having desired properties are prepared using small particles/droplets surrounded by a coating material. The material inside the microcapsule is known as core material, whereas outer coating is called a shell. Microcapsules may have diameter ranging a few micrometers to a few millimeters. Many special and functional properties can be imparted to the fabrics by microencapsulating the core material. This core material can be any substance having a special function to perform for the fabric. Encapsulation has allowed moisturizers, therapeutic oils, and insecticides to be incorporated into fabrics. Microencapsulation of antimicrobial agents is also gaining popularity in sportswear and medical textiles [25].
For preparation of herbal textile by microencapsulation, the “core material” along with the wall material should give a durable finish. This wall material can be certain substances like gum acacia, and the core material is the desired herbal extract. The microencapsulated herbal extracts have shown antimicrobial activity and wash durability up to 20 washes [26]. Microencapsulation is a very popular technique for incorporating fragrance into fabrics. These capsules applied are unbreakable under the normal conditions, and the application of pressure on fabric releases fragrance, which causes healing in aromatherapy [27]. Microencapsulated herbal extracts such as Aloe barbadensis Mill, Bitter Gourd, Ginger, and Cuminum cyminum Linn have successful application in which the herbal extracts are used as core material and gum acacia as wall material [28]. According to the study, microencapsulated extracts of Andrographis paniculata, when applied on the fabric to impart mosquito repellency finish, gave good mosquito repellent activity up to 30 washes [29].
New textile technologies have enabled the application of cosmetic ingredients on fabric to provide its functional benefit to the end-use product, and therefore, cosmetic textiles are moving from laboratory to showrooms. Fragrance finish is one such finish that falls under this category. A fragrance is made from a pleasant smelling aroma compound. Aromachology is a science that studies the effects of fragrances on the human body and mind. It researches how scents can be used to induce relaxation and make life more pleasant. Fragrances can be used in various forms such as essential oils, perfumes, colognes, household products, and potpourri. Some areas in textiles where fragrance finish can find application include Cosmetotextiles.
The major use of microencapsulation in cosmetic textiles is in the application of vitamins, essential oils, skin moisturizing agents, skin cooling, and antiaging agents [30]. Yamato et al. studied microcapsules having active substances that can improve the physiological conditions of human skin. The microcapsule gradually released its content when the textile structure was subjected to light pressure created due to the movement of human body [31].
Aroma finish is a process by which textile material is treated with the pleasant odor producing essential oils and aromatic compounds so that the wearer gets beneficial effects. Various essential oils such as lavender, rosemary, and jasmine are used.
The fragrance compounds and the essential oils are volatile substances, and prolonging life span of their odor is most difficult task in preparation of the textiles for aromatherapy. Microencapsulation is an effective technique to achieve this task. Microcapsules are tiny spherical enclosures containing a liquid or gas and assume the shape of the enclosed particle if containing a solid [32]. With microencapsulation method, fragrance lasts for more number of washes since microencapsulation makes the finished particles more securely attached to the internal structure of the fiber and thus releasing them slowly [33].
Bacteria often cause decay of fabrics leading to loss of various useful properties of fabrics. This problem can be prevented by the use of antimicrobial finishes that can be applied with the help of microencapsulation. This finish is especially beneficial for textiles for medical and technical uses [34].
Herbal extracts prepared from Chamomile, sage, and green tea were applied to cotton fabric using pad-dry-cure method. Treated fabrics exhibited antimicrobial activity against Gram-negative, Gram-positive bacteria and yeasts. This antimicrobial activity was retained even after 10 cyclic washes [35].
Textile materials having antimicrobial activity are used in hygienic, health, personal care, and surgical products. Some products include baby and adult diapers and sanitary pads. Most of the diapers are made by synthetic materials, which are nonbiodegradable and poor antimicrobial properties, and may cause rashes and dermatitis. Natural antimicrobial coatings on diapers obtained by using curcumin, Aloe vera, tulsi, and neem have same antimicrobial effect when compared with synthetic antimicrobial coatings [36]. Curcumin or diferuloylmethane (1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione) is the main phytochemical present in turmeric. This polyphenolic compound has several biological activities including antimicrobial activity. Curcumin is suspected to affect the bacterial cytoskeleton, thus producing the antimicrobial effect [37]. The Aloe vera leaves secrete a colorless gel-like substance, which contains about 98–99% water and 1–2% active compounds such as aloesin, aloin, aloe emodin, aloe mannan, flavonoids, saponin, sterols, amino acids, and vitamins. Anthraquinones and their derivatives such as barbaloin-IO-aloe emodin-9 anthrone, isobarbaloin, and chromones, which inhibit the bacterial protein synthesis, are potent antimicrobial agents [38]. Antimicrobial property of tulsi (holy basil) may be attributed to essential oils found in leaves. These essential oils contain germacrene-A, clemene, caryophyllene, eugenol, and caryophylline oxide. Additionally, tulsi leaves also have other biologically active phytochemicals such as rosmarinic acid, oleanolic acid, and ursolic acid. These essential oils and phytochemicals have antibacterial properties owing to their damaging effect of bacterial cell membrane and thus causing leakage of cellular potassium [39]. Neem leaf, bark, and seeds have antimicrobial properties, and it is high, moderate, and low, respectively. Neem contains several bioactive constituents like azadirachtin and other compounds such as nimbolinin, nimbin, nimbidin, nimbidol, sodium nimbinate, gedunin, salannin, quercetin, nimbanene, 6-desacetylnimbinene, nimbandiol, nimbolide, ascorbic acid, n-hexacosanol and amino acid, 7-desacetyl-7-benzoylazadiradione, 7-desacetyl-7-benzoylgedunin, 17-hydroxyazadiradione gedunin, and nimbiol. Quercetin and β-sitosterol, polyphenolic flavonoids purified from leaves of neem have antifungal and antibacterial activities. Neem (Azadirachta indica) in plant parts shows antimicrobial property through bacterial cell wall breakdown [40].
Wet processing of textile substrates from its preparatory stage to dyeing and then finishing is important for enhancing its esthetic value, removal of impurities, color shade, color pattern, and requisite functionality. Some of the traditional processes used in herbal finishing involve an excessive use of water, energy, and chemicals. Recently, due to global concerns on environmental pollution, sustainability mandates the development of sustainable dyeing and finishing processes using plant waste and nonfood plant extracts [41]. Based on environmental friendly, plant-based products having biocompatibility, biodegradability, and nontoxicity along with insect repellent, deodorizing, flame retardant, UV protection, and antimicrobial activity are in great demand all around the world for producing more appealing and highly functional value-added textiles [42, 43, 44, 45]. A wide variety of finishing chemicals from plants are now available in the market that matches the expectations of consumers [46]. Various plants reported as source of natural dyes are teak, mahogany, ketapang, tamarind, mangosteen, mango, suji, pandan, indigofera, guava, banana, and onion [47]. Plant parts including roots, leaves, twigs, stems, heartwood, bark, wood shavings, flowers, fruits, rinds, hulls, husks, and the like used to produce herbal dye. Additionally, most of the herbal dyes have inherent medicinal properties [48].
Natural fibers such as cotton and wool are susceptible to microbial growth and even dust mites because they retain oxygen, water, and nutrients. Hence, antimicrobial finishes should be applied to textiles to destroy or suppress the growth of microorganisms and also to protect the textiles from strength and color loss, unpleasant odor, and quality deterioration [49]. Micro-organisms may deteriorate the clothes in a closet, curtains, carpets, bed, bath and kitchen linens, pillows, and mattresses. Several microorganisms also thrive on the skin, while dust mites live on skin cells sheded on sheets, towels, and clothing. A hospital houses an immense amount of textiles with high volumes of traffic. Because of the constant flow of people, especially those with infectious diseases, specific finish hospital uses are required. Patients, caretakers, and hospital staff are at risk of acquiring infection as inherent properties of the textile fibers susceptible to the growth of micro-organisms.
In past, natural dyes were applied to textiles for simultaneous coloration and antimicrobial finishing successfully. Finish imparted by catechu on wool was found to be effective against Escherichia coli, Staphylococcus aureus, Candida albicans, and Candida tropicalis. Observed antimicrobial characteristics and negligible cytotoxicity of catechu indicated the dye as a promising antimicrobial agent for developing bioactive textile materials and herbal clothing [50, 51]. Several natural, nonmetallic, and antimicrobial finishes exist, and one of such antimicrobial finish is chitosan, a deacetylated form of chitin, which is a main component in crustacean shells and reported to be effective against both Gram-positive and Gram-negative bacteria [52]. Many antimicrobial textiles are produced with the combinations of bioactive substances to enhance the antimicrobial efficacy of the finishes and counteract the negative aspects of the treatments. By combining finishes, the occurrence of drug-resistant strains arising from the finish is reduced.
Pure cotton woven/knitted fabrics treated with the selected medicinal 16 medicinal herb extracts such as neem, turmeric, holy basil, and sandal wood have been evaluated for curing seven different diseases such as allergic dermatitis, psoriasis, asthma, liver disorders, headache, joint pain, and sinus trouble/cold. Seven different types of curative garments have been developed. The antibacterial assessments of the medicinal herb extracts treated fabrics and clinical trials have confirmed the correlation between the curative performance and its antibacterial activity. The curative property of the garments in all the cases was found to be significant and lasted for 10–15 washes [53].
Although known since long for dyeing and medicinal value, the protective properties of herbal dyes have been noticed only recently. Several plants used for dye production are classified as medicinal, and some have recently been shown to possess significant antimicrobial activity. Several natural dyes have demonstrated to possess antimicrobial activity like curcumin from turmeric; naphthoquinones such as lawsone from Lawsonia inermis, juglone from walnut, lapachol from taigu, and catechin from Acacia catechu; and several anthraquinones from Rubia tinctorum, Rubia cordifolia, Rheum emodi, Punica granatum, and Quercus infectoria [54, 55]. These natural antimicrobial agents may significantly reduce the risk of infections especially when they are used in close contact. Natural bioactive compounds present in natural dyes/pigments have also been reported as significant antimicrobial agents for finishing in green dyeing procedures.
Ultraviolet (UV) rays, electromagnetic spectrum between visible light and X-rays, influence the physiology of living organisms. Exposure may cause tanning to skin cancers. Sunscreen lotions and clothing provide protection from dangers of UV rays. Change in construction parameters of fabrics with appropriate UV absorbers and adequate finishing methods may be used to prepare the UV protective fabrics.
Textiles, as a protective shield against solar radiation, have been since ancient times. Textile structures provide the desired characteristics such as pliability, good mechanical strength, softness, esthetics, and other engineered properties, which are required for preparing good sunscreening apparels. Textiles themselves are not able to provide effective protection against UV rays; hence, UV blocking agents should be applied over the fabric to make them UV protective. Quantum of protection required for different skin types is determined by UV radiation intensity and distribution subject to geographical location, time of day, and season. Several UV protection agents have been developed to enhance the UV protection offered by different textiles. Both organic and inorganic UV blocking agents are available. The organic blockers are also termed as UV absorbers because they absorb UV rays, whereas inorganic blockers scatter them [47].
Three yellow dyes obtained from Rheum emodi, Gardenia yellow, and curcumin have been successfully used for simultaneous dyeing and functionalization of silk to render its UV protective [56]. Wool fabric dyed with dye obtained from the eucalyptus leaves using pad-batch and pad-dry techniques showed ultraviolet protection factor (UPF) values ranging between very good and excellent [57]. UV protection properties of chlorogenic acid, the main ingredient of water extract from honeysuckle, on wool have been studied. The wool treated with honeysuckle extract showed good UV absorbance; thus, extract of honeysuckle may be developed as a natural UV-absorbing agent [58]. Natural plant colorants such as madder (Rubia tinctorum) and indigo (Indigofera tinctoria) and of insect origin cochineal (Dactylopius coccus) were applied on cotton fabrics, and indigo was found to have higher UPF values [59]. UV rays induce DNA damage, oxidative stress, and inflammatory processes in skin. Rheum emodi contains anthraquinones (rhein, emodin, aloe-emodin, chrysophanol, physcion, and rhein), stilbenoids (resveratrol and piceatannol), and flavonoids. UV protective effect of Rheum emodi is supposed to be due to flavonoids, which absorb the UV radiation [60]. Main coloring ingredient of the dye extracted from Gardenia yellow is crocin, which is a water soluble carotene, which absorbs the UV rays [61]. Curcumin alleviates the effect of UV rays by reducing the transmittance of UV light across fabric [62]. Madder roots contain anthraquinone derivatives mainly purpurin and munjistin and also small amounts of rubiadin, nordamncanthal, pseudopurpurin, and xanthopurpurin. Anthraquinones are considered to be strong UV absorbers [22].
Bacterial growth and/or waste released from human body are the main causes for odor in garments. Recent advances in textile manufacturing have improved the performance of textiles with respect to odor with antimicrobial and UV protection properties. To meet the consumer’s growing demand for hygienic clothing, extensive research has been done to deodorizing property to textiles with the application of natural colorants. The deodorizing performance of fabrics dyed with herbal colorants has been compared, and pomegranate was found to be best among gardenia, Cassia tora L., coffee sludge, and pomegranate rind [23]. Fabric dyed with gallnut recorded a better deodorizing function against ammonia, trimethyl amine, and acetaldehyde than control fabrics. Dyed fabrics also found to possess an excellent antimicrobial activity against Staphylococcus aureus and Klebsiella pneumoniae [20]. Young-Hee Lee and co-workers dyed cotton, silk, and wool fabrics with pomegranate (Punica granatum) and showed excellent deodorizing property in the dyed fabrics [21]. Cotton fabrics dyed with C.I. Direct Blue 200, a copper complex direct dye, and pre- and post-mordanted with Cu(II) sulfate for deodorization of ethyl mercaptan showed that deodorization effects increased quadratically with the copper ion uptake [63]. These studies indicate that natural and synthetic dyes can be utilized for deodorizing functionalization of textiles.
Carpets, blankets, and shawls are prepared from wool, and other similar fibers have properties such as warmth, softness, and flame retardancy protein content of the fiber, which are susceptible to attack by moth as its larvae thrive on the protein present in the wool. Cloth moth (Tineola bisselliella) and carpet beetle (Anthrenus verbasci) are common moths, which attack the wool items. Dichlorodiphenyltrichloroethane (DDT), permethrin, permethrin/hexahydro pyrimidine derivative, and cyhalothrin are some of the insecticides used to provide antimoth finish. Nano titanium oxide particles have also been utilized as an antifeeding compound on wool fabric to kill larvae of the carpet beetle, Anthrenus verbasci, which feeds on wool fibers [64]. Chemicals used for antimoth finishing are detrimental to the environment; hence, natural colorants may be good alternatives. Shakyawar et al. used saffron flower waste, onion skin, henna, myrobolan, silver oak leaf, madder, walnut, dholkanali, and yellow root natural dyes for antimoth finishing and recorded the best results for silver oak leaves, walnut husk, and pomegranate rind [65]. Natural dyes from cochineal, madder, walnut (quinines), chestnut, fustic, indigo, and logwood (flavonoids) were also applied on wool, and their antimoth properties against black carpet beetles were tested, all except indigo, which increased the insect resistance of the wool fabric. Metallic mordants were found to have no significant effect on insect resistance with all natural dyes used. The anthraquinone dyes including cochineal, madder, and walnut were found to be quite effective in protecting wool fabric against black carpet beetles [66].
Synthetic/chemical dyes are increasingly being banned for use in the textile industry, which have led to severe blow to the handloom industry. At the same time, textiles dyed with natural herbal dyes having medicinal attributes are commanding a huge market due to their inherent advantages. Nowadays, the use of herbal dyes in the textile industry is preferred owing to the advantage of dyes extracted from the medicinal plants, origin from the renewable resources, limited chemical reactions involved in their preparation, biodegradable properties, disease-curing properties, and ecofriendly in nature [18, 67].
Herbal textiles are of two kinds: one category is for curing diseases and the other is for wellbeing. The first category of cloths, which are used for curing diseases, includes sleep wear, bed sheets, towels, meditation clothes, and coir mats. Herbal textiles are mostly used in making such garments that stay close to human skin, so that all the medicinal components could be absorbed through the skin. Herbal textiles can fetch more competitive than conventional fabrics owing to low production cost. For example, a common chemical dye costs around $75 per kg, and a herbal dye costs approximately $5 per kg. Most of the herbs used in dyeing are cultivated in South Asian countries, namely India, Pakistan, and Bangladesh, India being the largest producer. The Handloom Weavers Development Society (HLWDS) of Kerala, India has exported herbal and organic textile worth Rs. 50 Lakh to the United States, Europe, and Japan. The herbal textile is gaining such popularity that the Japanese Government has accorded HLWDS a $40,218 grant. Large consignments of herbal textiles have been shipped to international markets of the United States, Canada, France, Denmark, Italy, Poland, Maldives, Mauritius, Japan, and Sri Lanka [17, 68, 69, 70].
Most effective time to wear the herbal-infused clothing is while resting, sleeping, or meditating, when the body is naturally healing and re-establishing balance, so many of the products are created with these factors in mind. The technique for producing the herbal textiles is also used for floor coverings and coir mats. For coir mats, the fibers are first soaked in herbal dyes and then woven into coir mats. Apart from medicated handloom clothes such as sarees, T-shirts, shirts, trousers, kurtas, dhods, chudidhars, nightwear, fashion wear, sportswear, and inner wears, caps for healing headaches, goosery caps for head balance, cooling caps, hair shining caps, hair strengthening caps, bandages, and mask are also prepared [68, 71].
Plant dyes are increasingly being incorporated by designers into their designs contributing to revival of Ayurvastra or herbal clothing. Ayurganic is a line of loungewear inspired from the concept of Ayurvastra. Designer duo Lecoanet and Hemant have revived the tradition of Ayurvastra in Ayurganic, their line of medicinal clothing. The multi-award winning designers who began their journey as couturiers in Paris are now refining the concept of Ayurvastra. Anjelika dreams organic produces fair trade, organic handmade clothing that follows this tradition. Gibie towels specialize in Ayurvastra towels, Ayurvastram in Ayurvastra textiles and yoga mats, and niraamaya in Ayurvastra yoga mats. The handloom weavers’ development society in India is producing a wide range of herbal fabrics including sarees, bed sheets, and dress materials using medicinal dyes and exporting them to the United States, Germany, the United Kingdom, Singapore, and Taiwan [6, 11].
“Herbalfab” ecofriendly fabric brand has developed a unique technique to dye with flowers, fruits, roots, and so on. Turmeric, myrabolams, onion, kesu and dhavadi flowers, and natural indigo are used for preparing the natural dyes. This prevents the water pollution by replacing the petrochemical dyes with herbal dyes and imparts the medicinal value to cloth. The colors obtained are unique, which can kick start a new fashion trend. Herbalfab also encompasses a range of other ecofriendly fabrics like khadi, which is handspun and handwoven, nonviolent silk, organic denim, and so on [72].
The Handloom Weavers’ Development Society, India produces a variety of home textiles using herbal fabrics in India. They also export herbal clothing such as sarees, bed sheets, and dress materials to other countries. With increasing demand for herbal clothing, companies such as Ayurvastra online have started offering the online products. Many textile industries are getting inclined toward Cosmetotextiles. The brands making herbal clothes are Aura, Cognis, Pantaloons, Quiospheres, and so on. These brands have a range of clothing namely bandee, vest, camisole, men and women’s tee, coirs, and so on [73].
It acts as a revitalizing tonic and helps in keeping the body fresh and healthy. The uses of proper herbs in the textiles have proven to cure diseases such as arthritis and fever. We breathe through our body more than we do through nose. It can improve the skin’s natural ability to block and resist the harmful chemicals and toxins from entering the body, which will be beneficial to health.
Herbal cloths have the ability to protect us from various skin diseases and provide relief from infectious diseases and mental ailments.
No synthetic/chemical dye is used at any stage of herbal cloth production. For washing and cleaning purpose, the nut or nut powder of Sapindus laurifolia is recommended [74].
Medicinal herbs treated fabrics also have a lot of therapeutic value; thus, the fabric has been found very helpful for people suffering from ailments such as skin allergies, breathing problems, sleeping disorders, and blood pressure.
The health benefits of herbal clothing and its usage depend on the theory of touch. The body loses toxins when it comes in contact with herbal clothing, and this improves the metabolism. Herbal clothing is also known to help fight against many common diseases such as diabetes, hypertension, skin allergies, asthma, and heart ailments.
The human body naturally heals itself during sleep or meditation. Thus, when the body is at rest, herbal clothing will work most effectively. The skin is known as the largest organ of the body. Not only it acts as a fence and protects the body, but it can also be the channel for outside germs and toxins to enter into the body. Herbal clothing guards against the harmful toxins trying to enter the body through the skin [75, 76].
Herbal textiles are mainly used in making sleepwear, undergarments, bed coverings, towels, meditation clothing, and so on, which remain close to the skin absorbing all the benefits it gives out. Herbal textiles are also used in home textile products such as mattresses, coir mats, door mats, bath towels, bed spreads, and carpets.
Some of herbal constituents are antiallergens and hence are safe for skin contact and are mostly nonhazardous to human beings.
Natural dyes are usually moth proof and can replace the synthetic dyes in kids’ garments.
We breathe through our skin more than we do through nose, and chemically dyed textiles having carcinogenic amines and chemicals may be allergic and dangerous to human skin. Organic clothing can help reduce the exposure to allergens and other irritants and give a comfortable feeling [77, 78, 79].
Preparation of herbal cloths is a labor-intensive industry and hence will provide the job opportunities for manpower engaged in cultivation, extraction, and application on textile, food, leather, and so on.
Use of natural dyes may earn the carbon credit as it reduces the consumption of petroleum-based synthetic dyes.
It promotes the agriculture and balances the ecological cycle.
Waste can be used as manure.
No heavy metals such as chrome and copper are used in natural/herbal Ayurvedic dyeing.
Manual farming and organic practices have a lower carbon footprint as the entire process consumes less fuel and energy and emits fewer greenhouse gases than chemical textiles.
Herbal dyes are extracted from the plants cultivated without the use of synthetic pesticides and fertilizers, and also no chemical defoliants are used.
Processing is in green environment; hence, workers’ health is not compromised, and also water and electric use and toxic runoff are reduced.
Stringent testing ensures that product is free from contaminants such as nickel, lead, formaldehyde, amines, and pesticides.
It improves the soil fertility and soil structure by increasing the soil activity, thus reducing the risk of erosion.
It is the massive saving of precious water.
It promotes the development of earthworms and other arthropods leading to improvement in the growth conditions of the crop. Furthermore, organic crops thrive on root symbioses and are better equipped to exploit the soil; hence, fields can accommodate a more number of plants, animals, and microorganisms [80, 81, 82].
Natural fibers and natural dyes used in the preparation of herbal clothing allow its natural breakdown without damaging the environment compared to the synthetic material, which takes hundreds of years to breakdown and leaves the pollutant material in soil [66, 67, 83].
Herbal textiles are ecofriendly, and also residues they produce can be further used for making the other green products.
Solid and liquid wastes from herbal dyeing are amenable to recycling producing the organic manure.
Some of the ecofriendly industries producing the herbal textiles produce organically recycled paper bags by utilizing the residual organic and herbal dyed fabrics that can reduce the pollution due to plastic bags [84].
It also promotes the cultivation of herbs required for the production of herbal textile products.
Herbal dyes add value to the cotton fabrics in ecofriendly manner.
Herbal fabrics are lightweight and so used to construct the perfect breezy clothes. The biggest advantage of cotton herbal cloth is that it is cool in summer and warm in winter [85, 86].
The color shades produced by herbal dyes are unique, hence commanding high demand in market.
Herbal fabrics and dyes are not good substitutes for synthetic dyes offering broader range and variations.
Retaining color and its herbal benefits demands utmost care in washing; herbal cloths should be washed separately, preferably hand wash or gentle machine wash with bleach-free detergents and should be shed dried. This causes inconvenience to the user.
The dyeing process is cumbersome and time taking, and each stage should be carefully controlled.
Medicinal herbs impart colors; hence, color choice for consumer is very limited as only few herbs can be used for dyeing a fabric that meant to be used in a particular disease condition.
Herbal cloths show the curative effects, but progress is slow.
Though there are no complaints on allergic reaction by the consumers, some people think that the herbal clothing shows side effects on children [87, 88, 89].
Textile industry has realized its negative contribution to the environmental pollution through the harmful chemicals used in manufacturing processes and is continuously trying to find solution for this problem. Herbal dyeing is one important step toward achieving the organic lifestyle, thus reducing the environmental pollution. Herbal textile products are devoid of pesticides, fertilizers, antibiotics, growth hormones, genetically modified organisms, additives, irradiation, or sewage sludge, hence avoiding many ailments that are common with conventional clothing.
Modern consumer’s demand for novel ecomaterials is expected to increase in future. Recently, efforts have been made toward the development of commercially viable natural colorants by making advances in aspects such as identification of new sources, formulations, extraction, purification, and stability techniques. In spite of enthusiastic studies discussing the socioeconomic viability of natural dye production and applications at commercial scale for sustainable utilization of bioresources, there is a need for investigations related to hygiene and ecosafety, which have great future for the discovery of relatively better and more stable natural pigments that may have wider industrial applications.
Herbal textiles have tremendous scope in world textile market and may become a major textile product in future. The blend of herbs and textiles to achieve the health in an ecofriendly manner is the great way of adopting the healthy life. Herbal clothing is receiving the increased awareness all over the world. Herbally treated textiles are one of the great solutions to revive and increase the share of Indian handloom industry in world textile market. Furthermore, herbal clothing is nontoxic, noncarcinogenic, biodegradable, and ecofriendly; these characteristics make them an obvious choice for modern informed costumer looking for a healthy product. Additionally, the production of biocolorants to meet the rising demand shall force entrepreneurs to take up this venture for greater profits leading to more employment generation.
Supporting women in scientific research and encouraging more women to pursue careers in STEM fields has been an issue on the global agenda for many years. But there is still much to be done. And IntechOpen wants to help.
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