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Year : 2019  |  Volume : 16  |  Issue : 3  |  Page : 140-143

Diagnostic issue and caveats of immunoreactivity for Ki67 in hyalinizing trabecular neoplasm with a review of literature

1 Department of Pathology, LLRM Medical College, Meerut, Uttar Pradesh, India
2 Department of Pathology, Subharti Medical College, Meerut, Uttar Pradesh, India

Date of Submission16-Jun-2018
Date of Acceptance04-Oct-2019
Date of Web Publication18-Nov-2019

Correspondence Address:
Dr. Veer Karuna
Department of Pathology, LLRM Medical College, Meerut, Uttar Pradesh
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/trp.trp_27_18

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Few benign and malignant tumors of thyroid lesion share many histological features, including papillary and trabecular pattern, hyalinized stroma, calcification, nuclear grooving, and nuclear inclusion. The lesions those have these common features are hyalinizing trabecular tumor, papillary thyroid carcinoma, medullary carcinoma, paraganglioma, trabecular follicular adenoma (TFA), and poorly differentiated tumor (PDT). These tumors are derived from thyroid follicular cell except PDT and paraganglioma. Hyalinized trabecular tumor and TFA are rare tumors. Due to these common features, they create a diagnostic pitfall for a false-positive or false-negative diagnosis. Therefore, histopathological and immunohistochemical studies are needed to make a definite diagnosis. The present case describes a relatively rare thyroid tumor – hyalinizing trabecular tumor – with its varied morphological features creating diagnostic difficulties and role of immunohistochemistry in formulating definitive diagnosis.

Keywords: Hyalinizing trabecular tumor, Ki67 (clone MIB-1 and SP6), papillary thyroid carcinoma

How to cite this article:
Karuna V, Gupta P, Gupta M, Grover K. Diagnostic issue and caveats of immunoreactivity for Ki67 in hyalinizing trabecular neoplasm with a review of literature. Thyroid Res Pract 2019;16:140-3

How to cite this URL:
Karuna V, Gupta P, Gupta M, Grover K. Diagnostic issue and caveats of immunoreactivity for Ki67 in hyalinizing trabecular neoplasm with a review of literature. Thyroid Res Pract [serial online] 2019 [cited 2022 Jun 30];16:140-3. Available from: https://www.thetrp.net/text.asp?2019/16/3/140/271152

  Introduction Top

Hyalinizing trabecular tumor (HTT) was first described by Carney et al. in 1987. The term hyalinized trabecular adenoma was replaced by tumor due to its features consisting of large trabeculae with intratrabecular and intertrabecular hyaline material.[1] The majority of these cases have a benign course, but according to 2017 WHO Classification of thyroid tumors described this tumor as borderline/precursor tumors.[2] In the largest series of cases, the tumor was noted more common in the right lobe of the thyroid. Prevalent in females between the fourth and fifth decades of age was originally defined as infrequent but potentially a confusing neoplasm.[3]

  Case Report Top

A 35-year-old Indian male patient complained of midline neck swelling of 2 months duration. Physical examination revealed a mass in the right lobe of thyroid measuring 3 cm × 3 cm in size. There was no significant family or previous history of surgery or radiation exposure. Serum thyroid profile assay was within the normal limits. Magnetic resonance imaging showed an enlarged right lobe of the thyroid gland with well-defined oval lobulated lesion measuring 3.8 cm × 4.8 cm × 5.2 cm in size, suggesting the possibility of thyroid adenoma [Figure 1]a. Fine-needle aspiration cytology was performed; however, findings were nonspecific with only few clusters of benign follicular epithelial cells. The patient underwent right hemithyroidectomy. The extirpated tumor was 5 cm × 4 cm × 3 cm in size showed clear margins. It was circumscribed but not encapsulated [Figure 1]b. The parenchymal tumor cells were homogeneous and arranged in trabecular pattern and nests separated by hyalinized stroma. The tumor cells were elongated to ovoid with round nucleus, focal nuclear grooving, and moderate amphophilic cytoplasm [Figure 2]. Occasional trabeculae had round to irregularly shaped follicles that were either empty or contained colloid. No mitotic figures were detected. The margins were sharp, revealing no invasion of the surrounding tissues with no capsular and vascular invasion. Granular calcific material was present. A diagnosis of HTT was considered. However, due to overlapping features with other thyroid neoplasms such as papillary thyroid carcinoma (PTC), medullary thyroid carcinoma, trabecular follicular adenoma (TFA), poorly differentiated tumor (PDT), and paraganglioma. Histochemistry with Congo red stain was done revealing the absence of apple-green birefringence on polarized microscopy, thereby excluding the possibility of medullary carcinoma [Figure 3]a. Immunohistochemistry (IHC) for TTF, thyroglobulin, and Ki67 Clone MIB1 (Dako) was performed using polymer labeling technique on automated immunostainer, whereas Ki67 Clone SP6 (cell marque) was done in semiautomated antigen retrieval system. IHC for Ki67 using both MIB-1 and SP6 clone showed nuclear positivity in <1% of follicular cells, thereby differentiating from other thyroid malignancies [Figure 3]b. TTF showed nuclear positivity with moderate intensity and thyroglobulin were strongly positive, the latter two differentiating from paraganglioma [Figure 4].
Figure 1: (a) An enlarged right lobe of thyroid gland with well-defined oval lobulated lesion measuring 3.8 cm × 4.8 cm × 5.2 cm in size, (b) gross photograph showing well-circumscribed globular mass of 5 cm × 4 cm × 3 cm in size. The cut surface was homogeneously pale and firm with focal hemorrhagic areas

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Figure 2: (a and b) Abrupt onset of tumor with prominent trabeculae of follicular epithelium in abundant hyalinized stroma (H and E, ×40), (c) remarkable inter and intratrabecular hyalinization (H and E, ×100), (d) elongated tumor cell with round nuclei and moderate amphophilic cytoplasm. Focal granular calcification within the tumor noted (H and E, ×400)

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Figure 3: (a) The hyaline material of the tumor is Congo red negative (×100), (b) immunohistochemistry Ki-67 stain - focal nuclear positivity (×400)

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Figure 4: Immunohistochemistry - (a and b) thyroid transcription factor shows nuclear positivity ([a] ×40; [b] ×400) with moderate intensity of staining, (c and d) thyroglobulin exhibiting diffuse strong positivity

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  Discussion Top

We are presenting a case report that shares few but common architectural features, including trabecular growth pattern, nuclear grooving, hyalinized material, and focal granular calcification. The differential diagnoses considered with these features were HTT, PTC, medullary carcinoma (MC), paraganglioma, TFA, and PDT.

HTT and TFA are conceivably rare type of follicular cell neoplasm and frequently misinterpreted as PTC, MC, and paraganglioma due to some common histological features. The diagnosis of HTT is challenging due to many controversial aspects in regard to its diagnosis, classification, differential diagnosis, and malignant potential.[3],[4] HTT contain hyaline material, mimic amyloid that confuses it for medullary carcinoma thyroid. HTT was also designated as “paraganglioma-like adenoma of the thyroid” because of its histologic architectural resemblance to a neuroendocrine tumor. However, the HTT is derived from the follicular epithelium.[3],[5] Resemblances of morphological features among these tumors make the diagnosis difficult.

The PDT of the thyroid is characterized by tumor cells having enlarged nuclei with prominent nucleoli, presence of invasion, adhesion, and metastases in the surrounding tissue and absence of follicle or colloid by light microscopy in the trabeculae.[6] In the present case, there was no evidence of adhesion or invasion. The nuclei were uniform and small, with no visible nucleoli. Occasional trabeculae showed follicles that were either empty or contained colloid.

The presence of RET/PTC translocations in HTT strengthens the fact that HTN is a variant of PTC. Some molecular features that distinguishing HTT and PTC include, first is mRNA, which failed to reveal the characteristic upregulation in HTT and is expressed in PTC, and second mutations of the BRAF and N-RAS genes are found in PTC.[3]

The histological features in our case were strongly in favor of HTT, but Ki 67 (MIB-1) showed focal nuclear positivity rather than characteristic cytoplasmic/membranous expression, as described by Hirokawa and Carney.[5],[7]

A nonhistone protein Ki-67 antigen is expressed in G1, S, G2, and M phases, except G0. Ki 67 is available in clones such as MIB-1, SP2, and SP6 by different manufacturers. The MIB-1 monoclonal antibody has been frequently used to stain Ki-67 antigen.[5] Ki-67/MIB-1 has been identified as a useful marker in delineating between HTT and PTC. The positivity of cell membrane or cytoplasm is in favor of HTT, but its absence or nonreactivity does not exclude the diagnosis of HTT. This feature of MIB-1 is absent in PTC. Hirokawa and Carney stated that unique cytoplasmic MIB-1 (Ki-67) expression, in HTT is useful in making the distinction from PTC that is negative. However, negative MIB-1 stain has no diagnostic value.[7] Immunoreactivity for Ki67 varies according to the IHC methods and with different clones of Ki67. In our case, we applied clone MIB-1 (Ki67) and clone SP6 immunohistochemical stain, both revealing focal nuclear positivity in <1% of cells.

Various pitfalls and troubles can cause false-negative results with Ki 67; it may be due to type of method, under-fixation or overfixation, degradation of antibody, dewaxing, antigen retrieval temperature, few epitopes, tumor heterogeneity, technical error, and misinterpretation.[8]

Leonardo et al., in 2007, applied MIB-1 and different anti-Ki67 antibodies in HTT and other human tumors. They observed that the cell membrane reactivity in HTT by MIB-1 is obtained only if the reaction is performed at room temperature; because automated immunostainers which operate at 37° do not produce any MIB-1 membrane localization.[9] Therefore, MIB-1 focal nuclear positivity in our case could be due to variation in temperature during processing.

Takada et al.[1] used fully and semi-automated strainers in 10 cases of HTT. They observed that MIB-1 using fully-automated strainers may be negative, but in semi-automated strainers when antigen retrieval is carried out manually by using autoclave, pressure cooker, or microwave, membrane positivity will be raised. A similar phenomenon was reported by Park et al.,[10] in which fully -automated (BOND-MAX), semi-automated (BOND-X) immune strainer (IHC) systems and a manual method of MIB-1 were done. BOND-X showed MIB-1 Monoclonal antibody revealing membranous positivity, but BOND–MAX did not. The differences were in the IHC steps before the incubation with the primary antibody. These findings indicate that antigen retrieval by fully-automated stainers is insufficient for cell membranous staining of MIB-1 for HTTs.

This lack of true specificity suggests that alternative staining conditions be applied to detect the membranous immunoreactivity of Ki67 when the case suspected as HTT appears negative for Ki67 immunostaining.

Positivity of thyroglobulin, TTF, and negativity for congo red exclude the possibility of paraganglioma and medullary thyroid carcinoma, respectively.[11]

  Conclusion Top

In our case, the common histological features are trabecular pattern, presence of nuclear grooving, and hyalinized stroma. These features are also seen in other thyroid tumors. Exclusion of PDT, MC, and paraganglioma can be achieved using histochemistry and IHC in addition to morphology. IHC for Ki67 could produce unexpected results during the diagnosis of HTT. When staining for Ki67 is negative despite typical histological features of HTT, staining conditions should be altered to detect the characteristic membranous expression of Ki67.

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Conflicts of interest

There are no conflicts of interest.

  References Top

Takada N, Hirokawa M, Ohbayashi C, Nishikawa T, Itoh T, Imagawa N, et al. Re-evaluation of MIB-1 immunostaining for diagnosing hyalinizing trabecular tumour of the thyroid: Semi-automated techniques with manual antigen retrieval are more accurate than fully automated techniques. Endocr J 2018;65:239-44.  Back to cited text no. 1
Lloyd RV. Borderline Thyroid Tumors in WHO Classification of Endocrine Organs. 4th ed. Springer; 2017.  Back to cited text no. 2
Jadhav DS, Dukare SR, Kale P. Hyalinizing trabecular tumor of the thyroid: A rare case report. IJSR 2015;4;817-20.  Back to cited text no. 3
Andreas K, Efthimios P, Georgios G, Dimitrios L. Thyroid hyalinizing trabecular tumor: A case series. IJMPCR 2017;9:1-4.  Back to cited text no. 4
Yufei L, Xin H, Yuchang H, Fei W, Tingting D, Weiwen H, et al. Hyalinizing trabecular tumor of the thyroid: A clinicopathological analysis of four cases and review of the literature. Int J Clin Exp Pathol 2017;10:7616-26.  Back to cited text no. 5
Yuji O, Masashi K, Gang HL, Mitsuyoshi H. A purely trabecular follicular adenoma of the thyroid gland, harboring extraordinarily long trabeculae. Biomed Res 2015;26:380-3.  Back to cited text no. 6
Hirokawa M, Carney JA. Cell membrane and cytoplasmic staining for MIB-1 in hyalinizing trabecular adenoma of the thyroid gland. Am J Surg Pathol 2000;24:575-8.  Back to cited text no. 7
Jones DJ, Kieliszak CR, Patel SS, Selinsky CR. Hyalinizing trabecular tumor of the thyroid gland and its significant diagnostic issue. Thyroid Res 2017;10:7.  Back to cited text no. 8
Leonardo E, Volante M, Barbareschi M, Cavazza A, Dei Tos AP, Bussolati G, et al. Cell membrane reactivity of MIB-1 antibody to ki67 in human tumors: Fact or artifact? Appl Immunohistochem Mol Morphol 2007;15:220-3.  Back to cited text no. 9
Park HS, Kim KM, Bae JS, Chung MJ, Lee H, Moon WS, et al. Diagnostic caveats of immunoreactivity for ki67 and chromogranin a in hyalinizing trabecular tumour of the thyroid. J Clin Pathol 2014;67:835-9.  Back to cited text no. 10
Liu H, Lin F. Application of immunohistochemistry in thyroid pathology. Arch Pathol Lab Med 2015;139:67-82.  Back to cited text no. 11


  [Figure 1], [Figure 2], [Figure 3], [Figure 4]


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