Thyroid fine needle aspiration diagnostic category.
\\n\\n
Released this past November, the list is based on data collected from the Web of Science and highlights some of the world’s most influential scientific minds by naming the researchers whose publications over the previous decade have included a high number of Highly Cited Papers placing them among the top 1% most-cited.
\\n\\nWe wish to congratulate all of the researchers named and especially our authors on this amazing accomplishment! We are happy and proud to share in their success!
Note: Edited in March 2021
\\n"}]',published:!0,mainMedia:{caption:"Highly Cited",originalUrl:"/media/original/117"}},components:[{type:"htmlEditorComponent",content:'IntechOpen is proud to announce that 191 of our authors have made the Clarivate™ Highly Cited Researchers List for 2020, ranking them among the top 1% most-cited.
\n\nThroughout the years, the list has named a total of 261 IntechOpen authors as Highly Cited. Of those researchers, 69 have been featured on the list multiple times.
\n\n\n\nReleased this past November, the list is based on data collected from the Web of Science and highlights some of the world’s most influential scientific minds by naming the researchers whose publications over the previous decade have included a high number of Highly Cited Papers placing them among the top 1% most-cited.
\n\nWe wish to congratulate all of the researchers named and especially our authors on this amazing accomplishment! We are happy and proud to share in their success!
Note: Edited in March 2021
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Issues can arise at this step for various reasons, including a low concentration of analytes, incompatibility of the sample with the analytical instrument, and matrix interferences. This volume discusses the basics of sample preparation and examines modern techniques that can be used by both novice and expert analytical chemists. 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To address a significant variability in the reporting of cytological findings in thyroid FNA samples, a consensus recommendation known as the Bethesda System for Reporting Thyroid Cytology (TBSRTC) was provided in 2007. It includes six diagnostic categories, which are associated with varying risks of malignancy: I = nondiagnostic (ND), II = benign, III = atypia/follicular lesion of undetermined significance (AUS/FLUS), IV = follicular neoplasm/suspicious for a follicular neoplasm (FN/SFN), V = suspicious for malignancy, and VI = malignant [3]. This reporting system is compatible with that recommended by the British Thyroid Association [4] (Table 1). The BSRTC has proven to be an effective and robust thyroid FNA classification scheme to guide the clinical treatment of patients with thyroid nodules [5–9]; it is endorsed as a standard practice in reporting thyroid aspiration cytology by the 2015 American Thyroid Association (ATA) guidelines [2]. The Bethesda reporting system for thyroid FNA was used in 90% of practices in a recent survey [10].
\nBethesda diagnostic category [3] | \nBritish Thyroid Association [4] | \nCancer Risk [3] | \nManagement [3] | \n||
---|---|---|---|---|---|
Nondiagnostic or unsatisfactory | \nNondiagnostic | \n1–4% | \nRepeat FNA with U/S | \n||
Benign | \nNonneoplastic | \n1–3% | \nFollow-up clinically | \n||
Atypia of undetermined significance (AUS) or follicular lesion of undermined significance (FLUS) | \nAtypical features present | \n~5–15%* | \nRepeat FNA | \n||
Follicular neoplasm (FN) or suspicious for a follicular neoplasm (SFN) | \nFollicular neoplasm suspected | \n20–30%* | \nLobectomy | \n||
Suspicious for malignancy (SM) | \nSuspicious of Malignancy | \n60–75%* | \nLobectomy or total thyroidectomy | \n||
Malignant | \nDiagnostic of malignancy | \n97–99% | \nTotal thyroidectomy | \n
Thyroid fine needle aspiration diagnostic category.
* Reclassifying non-invasive follicular variant papillary thyroid carcinoma (NI-FVPTC) as neoplasm, and renaming it as noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP) would have most pronounced effect on the indeterminate categories: a decrease of cancer risk 5.2-17.6% in AUS/FLUS, 8-15.1% in SF/FN and 17.6-41.5% in suspicious for malignancy [22, 23] (see 3.3).
However, with implementation of BSRTC there is a tendency of increasing diagnosis of AUS/FLUS [6–12], and the risk of malignancy (ROM) associated with AUS/FLUS seems to be higher than estimated before [6–17]. These trends are commensurate with the incorporation of new molecular tests for classifying indeterminate thyroid nodules [16–20], and increased diagnosis of follicular variant papillary thyroid carcinoma (FVPTC) by surgical pathology [21]. Of note, a significant portion of AUS/FLUS has a histologic diagnosis of FVPTC [11, 12, 22]. If non-invasive FVPTC (NI-FVPTC) is considered as a neoplasm rather than a carcinoma, e.g. noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP) as recently suggested, the risk of malignancy for AUS/FLUS would further change [16, 22, 23].
\nAlthough “indeterminate” thyroid nodules include AUS/FLUS (III), FN/SF (IV), and suspicious for malignancy (V) categories, AUS/FLUS remains the most “
Hürthle cells are oncocytic cells, present in normal thyroid (increase with age), goiter, Hashimoto thyroiditis, Graves disease, radiation, chemotherapy, adenoma and carcinoma [29]. A Hürthle cell carcinoma is diagnosed if morphologic features indicative of malignancy are obvious. Otherwise, it is categorized as SN/FSN if the nodule is composed exclusively of Hürthle cells. It is downgraded to AUS/FLUS in a background of Hashimoto thyroiditis [3, 28, 29]. Hürthle cell lesions are considered “non-predictable” or “with higher malignant risk” [30], despite the fact that the risk of malignancy diagnosed by FNA in Hürthle cell lesions is similar to non-Hürthle cell lesions by other studies [28, 31].
\nWe would discuss the scenarios triggering AUS/FLUS diagnosis, characters helpful for subclassifying AUS/FLUS, cytological features of FVPTC, and Hürthle cell lesion. An algorithm with integrated approaches of clinical, radiologic, pathologic, and molecular findings will be proposed for indeterminate thyroid nodules.
\nThe AUS/FLUS thyroid nodules represent those not clearly benign or malignant [3]. The cytologic findings are not convincingly benign, yet the degree of cellular or architectural atypia is not sufficient for an interpretation of SF/FN, suspicious for malignancy, or malignancy [28]. It is a diagnosis of exclusion (Figure 1).
\nCase 1 (Figure 2): The aspiration of a 2.6 cm × 2.5 cm × 1.7-cm isthmus solid nodule is from a 46-year-old female. The smears are cellular with scant colloid, unlike a benign thyroid nodule. Nuclear enlargement and overlapping resembling SF/FN are present, but are focal. Rare microfollicles are identified in cell-block section. There are no papillary carcinoma features such as nuclear groove, pseudoinclusion, or papillae. This case was categorized cytologically as AUS (case courtesy of Dr. Jaklyn McClendon, Anaheim Health Medical Center, CA).
\nThe specimen was forwarded for ThyGenX Thyroid Oncogene Panel and ThyraMIR Thyroid miRNA Classifier. ThyGenXTM Oncogene Panel detected one NRAS point mutation (Q61R). The ThyraMIRTM microRNA Classifier was negative. Thyroid nodules with AUS diagnosis and NRAS mutation are at increased risk for malignancy (42–65% compared to 7–37% in NRAS-negative AUS,
AUS/FLUS in the Bethesda Thyroid Diagnostic Category. The cytological interpretation is on the left, the corresponding histological diagnosis is on the right. Category I nondiagnostic or unsatisfactory is not illustrated. Category II is a group of benign lesions including nodular goiter with abundant colloid and honeycomb sheets of follicular cells (IIA), and Hashimoto thyroiditis characterized by follicular cells, Hürthle cells, lymphocytes and plasmacytes (IIB). Category IV indicates follicular lesions (SF/FN) featured by increased cellularity and microfollicles. Category V applies to aspirations with some but not all features of malignancy. Category VI includes lesions meeting diagnostic criteria of malignancy. \nAUS/FLUS: Atypia of undetermined significance/follicular lesion of undetermined significance SF/FN: suspicious for follicular neoplasm/follicular neoplasm.
Atypia of undetermined significance. Focal crowded follicular cell clusters with scant colloid. (Case courtesy of Dr. Jaklyn McClendon, Anaheim Health Medical Center, CA.)
This case has also brought up a question of how ancillary studies would help subclassify AUS/FLUS. The various performances in molecular tests could be due to the nature of AUS/FLUS, which is a group of heterogeneous borderline lesions in transit from hyperplastic or adenomatoid nodule to adenoma, adenoma with atypia, and carcinoma. There is no single magic marker for subclassifying AUS/FLUS. Most tests use a panel of markers. New tests are emerging and extensive validation is required. Therefore, the 2015 American Thyroid Association (ATA) guidelines recommend ancillary studies but do not specify particular test(s) for AUS/FLUS follow-up [2].
\nRecognizing that some of the equivocal FNA cases are the result of inadequate number of cells or poorly visualized cells, the Bethesda System for Reporting Thyroid Cytopathology (TBSRTC) has recommended adequacy evaluation with category I being nondiagnostic [3]. Blood diluted and obscured specimen may push the follicular cells into microfollicle-like structure, making the colloid imperceivable and masking the benign cytological details. A sparse preparation or interpretation hindered by sampling preparation artifact may not be justified for a category of AUS/FLUS [3, 35]. It is important not to place those cases in AUS/FLUS group for the reasons: although re-biopsy is the same immediate outcome for ND or AUS/FLUS, the risk of malignancy for two AUS/FLUS diagnoses is higher than that for one AUS/FLUS diagnosis [10, 36], and most cases with two AUS/FLUS diagnoses will go to surgery [36].
\nClinical findings such as spiculated margin, microcalcification, hypoechogenicity of ultrasonography, larger size of mass, male gender, and increased thyroid-stimulating hormone (TSH) level may be associated with increased risk of malignancy in AUS/FLUS diagnosis. On the other hand, cystic or complex nodules have a lower risk of malignancy [37–42]. Considering those features, a more definitive diagnosis could be reached [35].
\nExpert consultation and group consensus reviews have been reported to minimize the diagnosis of FLUS [43]. A second opinion has an overall diagnostic resolution rate of 42.5% for “indeterminate” thyroid nodules and 71.5% reclassification accuracy in AUS/FLUS category [44].
\nThe use of the individual diagnostic categories within BSRTC varied up to 12.7-fold. The ratio of "AUS/FLUS” (A) to "malignant" (M) diagnoses varied at a range of 0.5–4.9 with a median ratio of 2.0. Based on this, an A:M ratio of 1.0–3.0 is proposed for the proper use of AUS/FLUS diagnosis. AUS:M ratios of >3.0 are likely because of overdiagnosis of AUS or underdiagnosis of M. AUS:M ratios of <1.0 are mostly due to low AUS rates, at the likely expense of sensitivity [45]. Some study supports this quality measure, but disputes exist [46].
\nReactive atypia in thyroid are associated with cystic degeneration, thyroiditis (inflammation), physical or chemical trauma, radiation, and other nonspecific causes, in a very similar way to cervical cytology [28, 29]. They present as metaplastic Hürthle cells, squamous cells, degenerative/regenerative follicular cells, or proliferative fibroblasts. Cells can be alarmingly bizarre with nuclear grooves or nucleoli but never have high N:C ratio, crowded nuclei, papillae, or microfollicle [47].
\nCase 2 (Figure 3): The smear is prepared from a 4-cm left-neck complex lesion on a 65-year-old female. The significant finding is a cluster of hemosiderin-associated cells with large elongated or polygonal nuclei in contrast to a background of honeycomb-round follicular cells. Despite the presence of nuclear groove, no intranuclear pseudoinclusion or architectural disorder such as crowded nuclei or papillae is identified to further suggest malignancy. Concurrent Afirma gene expression classifier result is benign, and medullary thyroid cancer classifier is negative (Veracyte, Inc., San Francisco, CA).
\nAtypical cells suggestive of benign cystic lining. The cells (upper) are larger compared to the rest of the follicular cells (lower), showing partially streaming appearance, irregular nuclear membrane and nuclear groove, pale chromatin, and small nucleoli. No nuclear overlapping or inclusion is present.
Atypical cystic lining cells need to be distinguished from cystic papillary carcinoma, which constitutes 10% of papillary carcinoma. The former does not have nuclear overlapping and inclusion [47].
\nEpithelioid cells in Hashimoto thyroiditis (Figure 4) or reparative fibroblasts can have atypical features such as large nuclei, taking an appearance of ugly ducklings among benign follicular cells. The low N:C ratio and non-clumping chromatin distinguish them from malignancy.
\nGranulomatous inflammation. (A) Epithelioid cells (left) in contrast to sheets of bland-appearing follicular cells mixed with several plasmacytes (arrow), Diff Quick stain. (B) Epithelioid cells in spindle-shaped and multinucleated form, Pap stain. (C) Resection shows Hashimoto thyroiditis with diffuse inflammation.
Perifollicular fibrosis refers to basement membrane-like material outlining follicles. Perifollicular fibrosis has been described in sporadic colloid nodule, adenomatoid hyperplasia, and in pediatric thyroid cancer following the Chernobyl disaster [24, 28, 48–50]. It is associated with benign or low-grade thyroid lesions. The incidence of perifollicular fibrosis seems to be higher in elderly people (>60) than in other age population (5–10 vs 2%) ([51] and unpublished data). In the elderly, it is often linked to a paucicellular aspiration with fibrosis.
\nCases 3 and 4 (Figure 5): Perifollicular fibrosis is illustrated in two cases with Bethesda category of atypia and benign, respectively. The first case (A) is a 38-year-old male with a left thyroid nodule status post chemoradiation for Hodgkin disease. Surgical resection shows an atypical adenomatoid hyperplasia. The second case (B) is a 61-year-old female with a benign 1.3-cm complex thyroid nodule.
\nPerifollicular fibrosis (A) in a cellular specimen with atypical cells. Follicular cells are expelled off a balloon-like fibrous rind (perifollicular fibrosis, left, Diff Quick stain). The follicular cells are atypical with slightly larger, crowded nuclei and rare nuclear groove and inclusion (right, Pap stain). (B) In a benign thyroid nodule with sparser cells. A glassy hyaline rind is partially peeled off a follicle with hemosiderin macrophages (upper, Pap stain) and scattered benign follicular cells (lower, Diff-Quik stain) are seen in the background.
Perifollicular fibrosis was first described in post-Chernobyl nonneoplastic thyroid tissue [50]. The basement membrane-like structure surrounding follicles may inhibit tumor genesis and progression, although the mechanism underlying perifollicular fibrosis in radiation exposure might be different from that in natural aging.
\nParathyroid gland is occasionally aspirated during thyroid nodule workup. In one molecular study for indeterminate thyroid nodules, three patients out of 441 who had a diagnosis of AUS/FLUS were found to have parathyroid rather than thyroidal origin after biochemical study and surgery [19].
\n\nParathyroid gland mimicking thyroid nodule. (A) An elongated nodule in the left posterior “thyroid” close to vessels. (B) Aggregates of structureless uniform cells with no colloid. (C) Immunohistochemistry performed on cell block confirmed parathyroid origin. PTH: parathyroid hormone; TG: thyroglobulin; CD45: white blood cell common antigen.
Case 5 (Figure 6): A 65-year-old female complained of “a lump in the throat.” Ultrasonography identified a right isthmus movable 1.9-cm nodule and a left posterior 2-cm nodule. Image and cytological features of the left posterior nodule are shown in A and B. The monomorphic small cells are bland appearing with scant cytoplasm and no identifiable colloid. The immunohistochemistry (IHC) confirms parathyroid origin.
\nIf IHC information is not available, this case may be mistaken for FLUS in the absence of colloid or benign thyroid nodule if colloid from adjacent thyroid is mixed up.
\nUltimobranchial body or solid cell nest is a developmental remnant derived from fourth to fifth pharyngeal pouch, considered as a normal component of thyroid. It is found in about 60% of serially sectioned thyroid with a male predominance [52–54]. Incidental aspiration of them may lead to AUS diagnosis [55].
\n\nUltimobranchial body. The aspiration shows cohesive polygonal cells. Histologic section shows a nest composed predominantly of similar polygonal or oval to spindle cells (main cells) mixed with a minor population of small cells with compact nuclei and clear cytoplasm.
Case 6 (Figure 7): The illustrated cell cluster is from a thyroid aspiration of a 46-year-old male. Cytological features distinguishing from malignancy include the absence of hyperchromatic nuclei or prominent nucleoli in tightly cohesive squamoid cells. Those cells are positive for Galectin-3, but negative for HMBE-1 [55].
\nIt seems that nuclear atypia (a subgroup of AUS) is more predictive of malignancy than microfollicle (a subgroup of FLUS) [56]. Nuclear atypia suggestive of papillary carcinoma carry a higher risk of malignancy than AUS/FLUS interpretations made for other reasons, such as microfollicles, Hürthle cells, and suboptimal specimens [57]. This might be due to higher incidence of papillary carcinoma or follicular variant papillary carcinoma than follicular carcinoma, and incapability of cytology to distinguish adenoma from follicular carcinoma.
\n\nAUS upgraded to papillary carcinoma follicular pattern. (A) A combination of nuclear overlapping, groove, and pale chromatin indicates AUS, suspicious for malignancy. (B) A follow-up core biopsy confirmed papillary carcinoma follicular pattern with invasion.
Nuclear pseudoinclusion and papillae are diagnostic of papillary carcinoma. Other nuclear features such as enlarged nuclei, pale chromatin, nuclear groove, and crowded nuclei are more commonly seen in AUS/FLUS category. More prominence of these findings or a great of them should increase the suspicion of malignancy [57].
\nCase 7 (Figure 8): This is a case of AUS upgraded to malignancy in a 32-year-old female. The initial aspiration of a thyroid nodule demonstrates groups of follicular cells with various features of nuclear atypia including pale chromatin, nuclear groove, and nuclear overlapping. A core-needle biopsy (CNB) 1 month later confirmed papillary carcinoma of follicular pattern with invasion.
\nFVPTC is defined by histology as a tumor with PTC-type nuclei and follicular rather than papillary pattern (papillary architecture of <1%) [21]. Based on the presence or absence of invasion of capsule or parenchyma, it is subgrouped as invasive FVPTC (I-FVPTC) and noninvasive FVPTC. The NI-FVPTC is clinically indolent similar to adenoma [22, 58]. It is recently renamed as noninvasive follicular thyroid neoplasm with papillary-like nuclear features to avoid overtreatment [22, 23, 58]. The impact of reclassifying NI-FVPTC on the risk of malignancy would be most pronounced in the indeterminate categories: a decrease of ROM, 5.2–17.6%, in AUS/FLUS, 8–15.1% in SF/FN, and 17.6–41.5% in suspicious for malignancy [16, 22].
\n\nNIFNP/NI-FVPTC. NI-FVPTC characterized by pale nuclei with marginated chromatin (yellow arrow), nuclear groove (black arrow), and microfollicle (orange circle) in an encapsulated nodule (left-upper corner) without capsule or parenchyma invasion.
NIFTP or NI-FVPTC has a molecular file of RAS mutation and PAX8/PPARr translocation but lacking BRAF V600E mutation in keeping with follicular adenoma. I-FVPTC has an opposite molecular profile closer to classical papillary carcinoma than to follicular adenoma or NIFTP/NI-FVPTC (BRAFV600E > RAS mutations) [58].
\nMost NIFTP or NI-FVPTC are categorized as AUS/FLUS, SF/FN, or suspicious for malignancy in Bethesda system. The NIFTP/NI-FVPTC in AUS/FLUS category shows atypical PTC-type nuclear feature and microfollicles (Figure 9). However, I-FVPTC versus NIFTP/NI-FVPTC is a histologic diagnosis, not a cytology stratification. Whether the degree of nuclear atypia in FVPTC is predictive of invasion is unclear.
\nHürthle cell tumor of thyroid is a group of neoplasm with distinct biology, morphology, and natural history. However, it is not an independent entity in World Health Organization (WHO) category and therefore has been diagnosed as either benign adenoma, or papillary carcinoma or follicular carcinoma or poorly differentiated based on cytomorphology, architecture of papillae, follicle and solid/trabeculae, and evaluation of mitosis and necrosis [59]. Capsule invasion or vascular invasion is the criterion for malignancy, while pleomorphism by itself is not a feature of malignancy [28, 59]. Cytomorphological criteria have been shown to be helpful in distinguishing Hürthle cell neoplasm from Hürthle cell metaplasia (in nodular goiter or Hashimoto thyroiditis), but not Hürthle cell carcinoma from adenoma [60].
\nHürthle cell lesion. (A) Predominant oncocytic cells on thyroid aspiration (left) and well-circumscribed nodule in the background of lymphocytic thyroiditis (Hashimoto disease) (right). (B) Loosely cohesive eosinophilic cells with binucleation on thyroid smears (left) and metastatic disease to the lung composed of the same cells (right) (case courtesy of Dr. Terry Welsh, Anaheim Health Medical Center, CA).
Lymphocytic thyroiditis is diagnosed based on a mixed cell population of lymphocytes, plasma cells, and follicular cells with Hürthle cell metaplasia. However, the differentiation of benign hyperplastic Hürthle cell nodule (or Hürthle cell adenoma) from Hürthle cell carcinoma can be very difficult based on cytology alone.
\nCase 8 (A and B) (Figure 10): A is a 56-year-old female presented with bilateral thyroid nodules. Surgical resection revealed hyperplastic oncocytic nodules in the background of lymphocytic thyroiditis. B is an 85-year-old female with one left thyroid nodule. Left lung wedge resection demonstrates metastatic lesion from thyroid (case courtesy of Dr. Terry Welsh, Anaheim Health Medical Center, CA). The smear preparations from A and B show similar oncocytic cells with the same cytological diagnosis of AUS/FLUS, Hürthle cell lesion.
\nAfter the lung lesion is diagnosed as thyroid metastasis (case B), a retrospective comparison of these two cytology cases was done. Hürthle cells in case B seem to be more loosely cohesive and have more binucleation. These architectural differences and cytological atypia are very subtle, requiring high grade of alert. Moreover, Hürthle cell metaplasia shows more pleomorphism than Hürthle cell carcinoma [27].
\nRetrospectively, the Hürthle cells in case A is positive for p27, but negative for HBME-1 and Galectin-3, in support of benign [54]. The triple immunostaining is not available for case B. The application of p27/HBME-1/Galectin-3 immunostaining in distinguishing benign Hürthle cell from Hürthle cell carcinoma needs to be studied.
\nA suspicious result from Afirma gene expression classifier does not increase the probability of malignancy in the Hürthle cell nodules. Patient should be counseled for the high possibility of unnecessary surgery based on suspicious interpretation of Afirma test for a Hürthle cell nodule [61, 62]. Molecular tests with both high sensitivity and specificity are needed for Hürthle cell nodules.
\nRepeat FNA is the recommendation for AUS/FLUS in TBSRTC [3]. For an initial aspirate diagnosed as AUS/FLUS, repeat FNA is specifically endorsed by the American Thyroid Association as a suitable follow-up option, perhaps proving especially useful when limited cellularity contributes to the initial AUS/FLUS interpretation [2, 63].
\nFor initial AUS/FLUS diagnosis, reclassification rate with repeat FNA is 56 and 69% [11, 64]. Most of the repeat FNA diagnoses are benign (69%, 47/74) [64]. The malignancy rate after surgery with or without repeat FNA for initial AUS/FLUS is 38.6 versus 15.6% [42, 65]. Repeat FNA helps the selection of patients with AUS/FLUS to triage to surgery, significantly reducing unnecessary surgery. Therefore, repeat FNA for nodules with AUS/FLUS on initial FNA is suggested.
\nTwo consecutive AUS/FLUS diagnoses have higher malignancy risk of at least 31.0% than one AUS/FLUS diagnosis and a higher proportion of FVPTC [11, 36, 65]. Solid structure, increased nodular size (>2 cm), and irregular/microlobulated margins are found to be risk factors in two studies [42, 66] but not in the other [36]. Most cases with repeated AUS/FLUS will require surgery [11, 36, 42, 65].
\nFor benign aspirates following initial AUS/FLUS, the malignancy risk is low (2 and 2.8%) so that clinical follow-up instead of surgical excision or continuous repeat FNA may be enough [64, 67], although one study has suggested a still higher risk of malignancy with one AUS/FLUS diagnosis compared to none [11]. The ultrasound features might be insignificant in predicting malignancy in this scenario [67].
\nA meta-analysis study has suggested that a core-needle biopsy has a higher conclusive rate than repeat FNA when the initial FNA produced inconclusive results [68]. This might be due to reduction in nondiagnostic category. Further prospective studies are necessary before follow-up CNB can be applied in daily practice.
\nCurrently available commercial molecular tests for indeterminate (AUS/FLUS) thyroid nodules on fine needle aspiration have different strengths (Table 2). The Afirma gene expression classifier developed by Veracyte, Inc., is a “rule-out malignancy” test for the preoperative identification of benign thyroid nodules with indeterminate cytology. The Afirma test assesses gene expression from mRNA isolated from thyroid FNA samples by comparing the mRNA expression detected in a thyroid FNA against a panel of 167 molecular genes [17]. The other three later-developed assays test for 17 known thyroid cancer-related mutations and translocations [18], combined with miRNA, mRNA, and DNA mutation [19] or 14 thyroid cancer-related genes and 42 types of gene fusion related to thyroid cancer [20], aiming to increase diagnostic yields on positive-predictive value.
\n\n | Sensitivity (%) | \nSpecificity (%) | \nPositive-predictive value (%) | \nNegative-predictive value (%) | \n
---|---|---|---|---|
Afirma gene expression classifier (Veracyte, Inc.) [17] | \n90 | \n53 | \n37.7 | \n95 | \n
miRInform™ (Asuragen Inc.) [18] | \n63 | \n99 | \n94 | \n88 | \n
Multi-Gene ThyroSeq Next-Generation Sequencing Assay (ThyroSeq®) [19] | \n90.9 | \n92.1 | \n76.9 | \n97.2 | \n
ThyGenX and ThyraMIRTM [20] | \n94 | \n80 | \n68 | \n97 | \n
Commercial molecular tests for indeterminate (AUS/FLUS) thyroid nodules on fine needle aspiration.
The test performance may change when applied to individual clinics. One independent study has reported lower than previously reported negative- and positive-predictive value (75 compared to 95%, and 16 compared to 38%) for cytology diagnosis of AUS/FLUS and SN/FN combined [62]. One reason might be due to the lower malignancy rate for indeterminate thyroid nodules in the specialized academic center compared to the validation settings from Afirma (17 vs 24%). Adjusting the malignancy rate on Bethesda categories III and IV, another study from the same institution demonstrated still lower actual performance of Afirma™, miRInform™, and ThyroSeq™ v2 tests compared to published sensitivity and specificity [69]. Assessing the institutional performance of each test is necessary along with the prevalence of malignancy. This has called for attention that customization is needed for the application of the molecular tests.
\nDiagnostic excision has been performed for high-risk thyroid nodules, and surgery is an indication for large goiter with symptoms. The synthesis of cytological interpretation, clinical factors including age, gender, nodular size, results of molecular testing, ultrasound findings, personal and family history, and the presence of additional nodules will impact the determination of the appropriate extent of initial surgical management [2, 42, 63, 70].
\nQuantitative methods are available to assemble clinical, ultrasonographic, and cytological findings into a scoring system to evaluate the malignant risk of thyroid nodules, especially in cases with indeterminate or repeated nondiagnostic FNA [70]. A similar scoring system integrating molecular test results would be desirable.
\nCases are from the University of Chicago, Mercy Hospitals at Bakersfield, the University of California at Irvine, Anaheim Health Medical Center, and the University of California at Los Angeles. We thank Dr. McClendon and Dr. Welsh for the courtesy of cases and helpful discussion.
\nPseudocereal grains are considered as good sources of protein with a balanced amino acid profile. Proteins from pseudocereal grains have recently gained increasing popularity due to their nutritional, functional, and biological properties. Proteins from quinoa, amaranth, and chia are among the most extensively studied pseudocereal proteins in terms of characterization of physicochemical, functional, and biological properties. The functionality of proteins from other less known pseudocereals, such as kiwicha and cañihua, still remains to be explored. Although proteins from pseudocereal grains are indicated to show good functionality, some processes may be required to modify the structure and improve the functionality of pseudocereal proteins. Structural and functional properties of various pseudocereal proteins are recently reviewed [1, 2, 3]. This chapter presents an overview of the structural and functional properties of pseudocereal proteins, the effects of methods used for protein extraction and fractionation on protein functionality, and several methods applied for modification of structure and optimizing the functionality of pseudocereal proteins.
Quinoa (
Various physical, chemical, and biological modification methods are applied to pseudocereal proteins to improve functionality. Enzymatic hydrolysis is a commonly applied strategy to improve not only the functional but also the bioactive properties of plant-based proteins. Guo et al. [7] recently reviewed the biological activities of quinoa protein hydrolysate and peptides. In a recent study, Daliri et al. [8] applied enzymatic hydrolysis to quinoa protein concentrate with pancreatin and investigated the changes in emulsifying, foaming, and antioxidant properties. Quinoa protein concentrate was obtained from defatted quinoa flour with alkaline extraction followed by the isoelectric precipitation method. Hydrolysis with pancreatin at 40°C for 180 min was reported to result in the highest degree of hydrolysis (∼19%). Fourier-transform infrared spectroscopy analysis revealed that different functional groups, such as free regions of hydroxylic amino acids, aromatic amino acids, and free amino groups, originated in the hydrolysate due to the hydrolyzing action of pancreatin. The obtained hydrolysate was reported to show better antioxidant properties in terms of 2,2-diphenyl-1-picrylhydrazyl free radical scavenging activity. Solubility, emulsifying and foaming activities of the hydrolysate were found to be higher than that of the native protein. On the other hand, the native protein showed better emulsion and foam stabilizing properties [8].
Maillard reaction is used as a tool to modify structural properties and improve the functionality and biological activity of proteins. In a recent study, Teng et al. [9] investigated the effect of glycosylation with xylose on the structural and functional properties of quinoa protein. Quinoa protein isolate (96% protein) was obtained from defatted quinoa flour with alkaline extraction followed by an isoelectric precipitation method. Glycosylation via Maillard reaction was performed by mixing quinoa protein isolate with mannose or xylose with varying proportions in phosphate buffer and heating at 60°C for 4 h. The optimum ratio of quinoa protein to monosaccharide was determined to be 2:1 based on the degree of grafting and browning index analyses. The electrophoretic profile of samples revealed that glycosylation had significant effects on the depolymerization and remodeling of molecular aggregates of quinoa protein. The specific surface area and absorption capacity of quinoa protein were indicated to increase after glycosylation. Solubility, water and fat absorption capacities, emulsifying activity, and stability of glycosylated quinoa protein were reported to be significantly higher than that of the native protein. Moreover, anti-inflammatory and anti-proliferative activities of quinoa protein were indicated to increase after the glycosylation reaction [9].
Amaranth (
Figueroa-González et al. [12] investigated the effects of pH-shifting and ultrasonication treatments on the structure, physicochemical, and foaming properties of amaranth protein. Amaranth protein isolate (83% protein) was obtained from defatted amaranth flour with alkaline extraction-isoelectric precipitation method. Amaranth protein dispersions were prepared in distilled water (30 mg/mL, pH 7.0) and protein was modified by five different treatments—pH-shifting at pH 2.0 and 12.0, sonication (750 W) for 10 min at an amplitude of 50%, and pH-shifting (at pH 2.0 and 12.0) followed by sonication. After the modification treatments, amaranth protein dispersions were dried at 35°C for 45 h in the oven to avoid protein denaturation. Alkaline pH-shifting followed by sonication was reported to result in a significant decrease in the hydrodynamic diameter of amaranth protein. On the other hand, hydrodynamic diameter of protein was observed to increase after the acidic pH-shifting treatment. The isoelectric point of amaranth protein increased from 4.0 to 4.2 after the alkaline pH-shifting treatment and to 4.5 after the combined alkaline pH-shifting and ultrasound treatments. However, ultrasound treatment alone was reported to decrease the isoelectric point of amaranth protein to 3.5. Alkaline pH-shifting and ultrasound treatments were reported to induce changes in the secondary structure fractions of amaranth protein. Moreover, both pH-shifting treatments and combination of pH-shifting and ultrasound treatments resulted in changes in the sulfhydryl groups and disulfide bonds of amaranth protein. Both pH-shifting treatments were reported to improve the solubility of amaranth protein, where the highest protein solubility was observed in the sample treated with a combination of alkaline pH-shifting and ultrasound. The foaming capacity and stability of amaranth protein were reported to increase significantly after all treatments except for the acidic pH-shifting treatment. Moreover, treatments applied were indicated to improve the
Das et al. [13] investigated the effects of pH treatment and the extraction pH on the physicochemical and functional properties of amaranth protein isolate. Amaranth protein isolate was obtained from defatted amaranth flour with alkaline extraction at different pH values (9.0, 10.0, 11.0, and 12.0) followed by isoelectric precipitation (pH 4.5). Amaranth protein isolates were subjected to pH treatments at pH 3.0–9.0 and tested for functionality. The protein content of amaranth proteins extracted at different pH values changed between 56 and 85%, where the isolate obtained at pH 9.0 showed the highest purity, solubility, and particle size but the lowest yield. The authors reported that maintaining the extraction and treatment pH values at 9.0 resulted in significant improvements in functional properties including solubility, water and oil binding capacities, emulsifying and foaming properties. Extraction and treatment at pH 9.0 were also indicated to result in better thermal properties and improved gelation characteristics. Moreover, emulsifying, foaming, and gelation properties of amaranth protein isolates were reported to be affected by the particle size, wettability, and solubility [13].
Enzymatic modification is recently applied as a useful tool to improve the functionality and biological activity of amaranth protein. Kamal et al. [14] prepared amaranth protein hydrolysates using bromelain, chymotrypsin, and pronase E enzymes at three different hydrolysis durations (2, 4, and 6 h). Bioactive peptides were identified by LC-MS-QToF analysis. Amaranth protein hydrolysates were reported to contain bioactive peptides with inhibitory properties against enzymatic markers linked with hypertension and diabetes [14].
In another recent study, Karimi et al. [15] investigated the effects of using selective hydrolyzed protein of amaranth on sourdough fermentation, bread quality, and shelf life. Amaranth protein isolate was obtained from defatted amaranth flour using the alkaline extraction-isoelectric precipitation method. Protein hydrolysates were prepared with Alcalase® treatment for 3 h. The authors reported that amaranth protein hydrolysates increased the growth of
Kiwicha (
The protein content of chia (
Julio et al. [22] prepared different protein fractions of albumins, globulins, glutelins, and prolamins from chia protein-rich fraction of chia seeds, a by-product of chia oil extraction process. The solubility profile of chia protein-rich fraction, globulins, and prolamins was observed to be similar and made a peak at pH 9.0. On the other hand, maximum solubility was observed at pH 5.0 for glutelin and albumin fractions. Detailed emulsion characterization tests including destabilization kinetics and particle size distributions revealed that globulin fraction resulted in the most stable emulsion systems. The authors reported that higher pH values resulted in improved stability in emulsions stabilized with globulins, glutelins, and chia protein-rich fraction [22].
Urbizo-Reyes et al. [23] prepared chia protein hydrolysates with ultrasound treatment followed by microwave-assisted hydrolysis. For this purpose, chia seed mucilage and chia seed oil were extracted from the seeds prior to protein hydrolysis. Chia protein hydrolysates were prepared using Alcalase® or sequential hydrolysis with Alcalase® and Flavourzyme®. Enzymatic hydrolysis reaction was conducted using a conventional or microwave-assisted system. Chia protein hydrolysates obtained using sequential hydrolysis with microwave treatment were reported to show significantly higher
Teff (
Teff flour is widely used in formulations of gluten-free bread and bakery products. Adebowale et al. [25] compared the characteristics of protein fractions in three different teff types with sorghum with the main focus on bread-making quality. The major protein fraction in teff was reported to be prolamin. Aqueous alcohol-soluble protein fraction was indicated to be rich in glutamine and leucine. The authors suggested that differences in the electrophoretic profile of proteins indicated that teff prolamin is less polymerized compared to sorghum prolamin. Functional properties of teff prolamins useful in bread making were attributed to the differences in thermal profile, lower polymerization, and hydrophobicity [25].
Common buckwheat (
Xue et al. [29] investigated the effects of high-intensity ultrasound treatment and Maillard reaction on structural, interfacial, and emulsifying properties of buckwheat protein. Buckwheat protein isolate was prepared from defatted buckwheat flour with alkaline extraction method followed by isoelectric precipitation. Buckwheat protein isolate-dextran conjugates were prepared via Maillard reaction combined with ultrasound treatment. The secondary and tertiary structures and surface hydrophobicity of buckwheat protein isolate-dextran conjugates obtained with ultrasonication were observed to be different than those of conjugates obtained with classical heating. As a result of the modifications in protein structure, emulsifying properties and surface activity of conjugates obtained with ultrasonication were reported to be improved compared to classical heating [29].
In another recent study, Wu et al. [30] investigated the effect of extraction pH on structure, functional properties, and digestibility of tartary buckwheat protein. Protein isolates were prepared from defatted tartary buckwheat flour using alkaline extraction at different pH values (pH 7.0–13.0) followed by isoelectric precipitation. Tartary buckwheat flour and protein isolates were separated into albumin, globulin, prolamin, and glutenin fractions based on Osborne-type protein fractionation. Protein extraction at alkaline conditions was reported to increase protein extraction yield. Increased extraction pH was indicated to decrease the albumin content of tartary buckwheat protein isolate while glutenin content increased. The solubility of isolates extracted at pH > 12.0 was observed to decrease. On the other hand, emulsion stability increased at the same conditions that were attributed to increased surface hydrophobicity. The differences observed in
In addition to functional properties, buckwheat protein and derived bioactive peptides are reported to show various biological properties, including cholesterol-lowering activity, blood pressure controlling enzyme inhibitory activity, antimicrobial and antioxidant activities that suggest the potential use of buckwheat protein and peptides as functional food ingredients [3].
Cañihua (
Enzymatic hydrolysis was applied to cañihua protein for obtaining peptides with biological activities. Chirinos et al. [34] derived hydrolysates and peptides from cañihua protein concentrate. Protein concentrate (79% protein) was obtained from defatted cañihua meal with alkaline extraction-isoelectric precipitation method. Cañihua protein concentrate was subjected to enzymatic hydrolysis with Alcalase®, Neutrase®, and Flavourzyme® at 50°C up to 240 min. The hydrolysates obtained were purified via ultrafiltration and size exclusion chromatography to obtain three peptide fractions. The authors reported that cañihua protein can be considered as a good source of bioactive peptides with antioxidant and angiotensin-I converting enzyme (ACE) inhibitory activities. Specifically, cañihua protein hydrolysate obtained with Neutrase®-Alcalase® sequential hydrolysis for 180 min was indicated to show good
In another recent study, Moscoso-Mujica et al. [35] also applied enzymatic hydrolysis to cañihua protein. Cañihua flour was obtained from the seeds of two different varieties (Ramis and Cupi-Sayhua) and defatted prior to protein extraction. Protein fractions of albumins, 7S globulins, 11S globulins, and glutelins were obtained based on solubility differences and subjected to sequential hydrolysis with Alcalase® and pepsin-pancreatin. Hydrolysates with varying degrees of hydrolysis were obtained and tested for antimicrobial activity against
Pseudocereals are indicated as good protein sources with a balanced amino acid profile. Nutritional composition and protein characteristics of pseudocereal grains change depending on the seed variety and growing conditions. Moreover, the methods used for protein extraction and fractionation affect protein structure, composition, and hence, functionality. Enzymatic hydrolysis has been shown to be a useful tool for obtaining peptides from pseudocereal proteins with biological activities, including antioxidant, antimicrobial, and antihypertensive properties. Proteins and peptides from pseudocereal grains can be potentially utilized as ingredients in innovative product formulations due to their nutritional quality, functional properties, and biological activities. More research is needed to investigate the effects of pseudocereal proteins on end-product quality to elucidate the potential and increase the utilization of pseudocereal proteins as food ingredients.
The author declares no conflicts of interest.
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His studies in robotics lead him not only to a PhD degree but also inspired him to co-found and build the International Journal of Advanced Robotic Systems - world's first Open Access journal in the field of robotics.",institutionString:null,institution:{name:"TU Wien",country:{name:"Austria"}}},{id:"441",title:"Ph.D.",name:"Jaekyu",middleName:null,surname:"Park",slug:"jaekyu-park",fullName:"Jaekyu Park",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/441/images/1881_n.jpg",biography:null,institutionString:null,institution:{name:"LG Corporation (South Korea)",country:{name:"Korea, South"}}},{id:"465",title:"Dr",name:"Christian",middleName:null,surname:"Martens",slug:"christian-martens",fullName:"Christian Martens",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:null},{id:"479",title:"Dr.",name:"Valentina",middleName:null,surname:"Colla",slug:"valentina-colla",fullName:"Valentina Colla",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/479/images/358_n.jpg",biography:null,institutionString:null,institution:{name:"Sant'Anna School of Advanced Studies",country:{name:"Italy"}}},{id:"494",title:"PhD",name:"Loris",middleName:null,surname:"Nanni",slug:"loris-nanni",fullName:"Loris Nanni",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/494/images/system/494.jpg",biography:"Loris Nanni received his Master Degree cum laude on June-2002 from the University of Bologna, and the April 26th 2006 he received his Ph.D. in Computer Engineering at DEIS, University of Bologna. On September, 29th 2006 he has won a post PhD fellowship from the university of Bologna (from October 2006 to October 2008), at the competitive examination he was ranked first in the industrial engineering area. He extensively served as referee for several international journals. He is author/coauthor of more than 100 research papers. He has been involved in some projects supported by MURST and European Community. His research interests include pattern recognition, bioinformatics, and biometric systems (fingerprint classification and recognition, signature verification, face recognition).",institutionString:null,institution:null},{id:"496",title:"Dr.",name:"Carlos",middleName:null,surname:"Leon",slug:"carlos-leon",fullName:"Carlos Leon",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Seville",country:{name:"Spain"}}},{id:"512",title:"Dr.",name:"Dayang",middleName:null,surname:"Jawawi",slug:"dayang-jawawi",fullName:"Dayang Jawawi",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Technology Malaysia",country:{name:"Malaysia"}}},{id:"528",title:"Dr.",name:"Kresimir",middleName:null,surname:"Delac",slug:"kresimir-delac",fullName:"Kresimir Delac",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/528/images/system/528.jpg",biography:"K. Delac received his B.Sc.E.E. degree in 2003 and is currentlypursuing a Ph.D. degree at the University of Zagreb, Faculty of Electrical Engineering andComputing. His current research interests are digital image analysis, pattern recognition andbiometrics.",institutionString:null,institution:{name:"University of Zagreb",country:{name:"Croatia"}}},{id:"557",title:"Dr.",name:"Andon",middleName:"Venelinov",surname:"Topalov",slug:"andon-topalov",fullName:"Andon Topalov",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/557/images/1927_n.jpg",biography:"Dr. Andon V. Topalov received the MSc degree in Control Engineering from the Faculty of Information Systems, Technologies, and Automation at Moscow State University of Civil Engineering (MGGU) in 1979. He then received his PhD degree in Control Engineering from the Department of Automation and Remote Control at Moscow State Mining University (MGSU), Moscow, in 1984. From 1985 to 1986, he was a Research Fellow in the Research Institute for Electronic Equipment, ZZU AD, Plovdiv, Bulgaria. In 1986, he joined the Department of Control Systems, Technical University of Sofia at the Plovdiv campus, where he is presently a Full Professor. He has held long-term visiting Professor/Scholar positions at various institutions in South Korea, Turkey, Mexico, Greece, Belgium, UK, and Germany. And he has coauthored one book and authored or coauthored more than 80 research papers in conference proceedings and journals. His current research interests are in the fields of intelligent control and robotics.",institutionString:null,institution:{name:"Technical University of Sofia",country:{name:"Bulgaria"}}},{id:"585",title:"Prof.",name:"Munir",middleName:null,surname:"Merdan",slug:"munir-merdan",fullName:"Munir Merdan",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/585/images/system/585.jpg",biography:"Munir Merdan received the M.Sc. degree in mechanical engineering from the Technical University of Sarajevo, Bosnia and Herzegovina, in 2001, and the Ph.D. degree in electrical engineering from the Vienna University of Technology, Vienna, Austria, in 2009.Since 2005, he has been at the Automation and Control Institute, Vienna University of Technology, where he is currently a Senior Researcher. His research interests include the application of agent technology for achieving agile control in the manufacturing environment.",institutionString:null,institution:null},{id:"605",title:"Prof",name:"Dil",middleName:null,surname:"Hussain",slug:"dil-hussain",fullName:"Dil Hussain",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/605/images/system/605.jpg",biography:"Dr. Dil Muhammad Akbar Hussain is a professor of Electronics Engineering & Computer Science at the Department of Energy Technology, Aalborg University Denmark. Professor Akbar has a Master degree in Digital Electronics from Govt. College University, Lahore Pakistan and a P-hD degree in Control Engineering from the School of Engineering and Applied Sciences, University of Sussex United Kingdom. Aalborg University has Two Satellite Campuses, one in Copenhagen (Aalborg University Copenhagen) and the other in Esbjerg (Aalborg University Esbjerg).\n· He is a member of prestigious IEEE (Institute of Electrical and Electronics Engineers), and IAENG (International Association of Engineers) organizations. \n· He is the chief Editor of the Journal of Software Engineering.\n· He is the member of the Editorial Board of International Journal of Computer Science and Software Technology (IJCSST) and International Journal of Computer Engineering and Information Technology. \n· He is also the Editor of Communication in Computer and Information Science CCIS-20 by Springer.\n· Reviewer For Many Conferences\nHe is the lead person in making collaboration agreements between Aalborg University and many universities of Pakistan, for which the MOU’s (Memorandum of Understanding) have been signed.\nProfessor Akbar is working in Academia since 1990, he started his career as a Lab demonstrator/TA at the University of Sussex. After finishing his P. hD degree in 1992, he served in the Industry as a Scientific Officer and continued his academic career as a visiting scholar for a number of educational institutions. In 1996 he joined National University of Science & Technology Pakistan (NUST) as an Associate Professor; NUST is one of the top few universities in Pakistan. In 1999 he joined an International Company Lineo Inc, Canada as Manager Compiler Group, where he headed the group for developing Compiler Tool Chain and Porting of Operating Systems for the BLACKfin processor. The processor development was a joint venture by Intel and Analog Devices. In 2002 Lineo Inc., was taken over by another company, so he joined Aalborg University Denmark as an Assistant Professor.\nProfessor Akbar has truly a multi-disciplined career and he continued his legacy and making progress in many areas of his interests both in teaching and research. 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This study was carried out under field conditions, over a period of four years, where the effect of indaziflam and glyphosate herbicides on roots of Coffee and Citrus plants was evaluated. The results demonstrate that the methodology used to assess the effect of herbicides on the roots was important to validate and qualify safe herbicide selectivity towards crops. 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