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ORIGINAL ARTICLE
Year : 2020  |  Volume : 17  |  Issue : 1  |  Page : 14-18

A comparison of liquid-based cytology and conventional smears in fine needle aspiration cytology of thyroid lesions: Diagnostic efficacy and pitfalls


Department of Pathology, VMMC and Safdarjung Hospital, New Delhi, India

Date of Submission23-Nov-2019
Date of Acceptance07-Jan-2020
Date of Web Publication24-Apr-2020

Correspondence Address:
Dr. Mukul Singh
Room No. 408, Department of Pathology, VMMC and Safdarjung Hospital, New Delhi
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/trp.trp_42_19

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  Abstract 


Introduction: Fine needle aspiration cytology is an unrivalled method of preoperative triage of thyroid lesions. The use of liquid-based cytology (LBC) is now extended to nongynecological samples due to the advantages of lesser screening time, clearer background, and potential use of leftover material for ancillary investigations.
Aims and Objectives: The aim of the study is cytomorphological comparison of LBC and conventional method and cytohistopathological correlation in surgically resected cases.
Materials and Methods: Fine needle aspiration was done in 100 patients. Conventional smears were prepared followed by LBC from the remaining material. The smears from both the methods were scored semiquantatively and were compared cytomorphologically. Pearson's Chi-square test and fisher's exact test were used for P value calculation using SPSS software. A P < 0.05 was considered as significant. A cytohistopathological correlation was done in operated cases. The remaining material from LBC was used for immunocytochemistry wherever required.
Results: LBC had more unsatisfactory as well as suspicious for malignancy cases. The cells in LBC were in three-dimensional clusters and singly scattered with loss of cellular architecture. The nuclei were shrunkun and hyperchromatic with spilling of cytoplasm. The background colloid, blood as well as lymphocytes were mostly washed off. Immunocytochemisrty results were not affected by the LBC procedural steps.
Conclusion: It was very difficult to come to a conclusive diagnosis with the help of LBC alone. It can be useful for ancillary investigations like immunocytochemistry as in this study. Thus, it can be used as a supplement to conventional method rather than replacing it.

Keywords: Conventional method, fine needle aspiration cytology, immunocytochemistry, liquid-based cytology, thyroid lesions


How to cite this article:
Kumari M, Singh M. A comparison of liquid-based cytology and conventional smears in fine needle aspiration cytology of thyroid lesions: Diagnostic efficacy and pitfalls. Thyroid Res Pract 2020;17:14-8

How to cite this URL:
Kumari M, Singh M. A comparison of liquid-based cytology and conventional smears in fine needle aspiration cytology of thyroid lesions: Diagnostic efficacy and pitfalls. Thyroid Res Pract [serial online] 2020 [cited 2020 Aug 14];17:14-8. Available from: http://www.thetrp.net/text.asp?2020/17/1/14/283220




  Introduction Top


Fine needle aspiration cytology (FNAC) is an extremely useful, simple, safe, cost-effective, accurate and widely used diagnostic tool for preoperative evaluation of palpable thyroid nodules.[1] The worldwide prevalence of palpable thyroid nodules is 12.2% which is further increased to 10%–14% due to availability of imaging techniques.[2],[3] However, 95% of these are benign in nature which do not require any surgical management. The preoperative FNAC categorization of benign or malignant thyroid nodules has increased malignant thyroid resections from 14% to 50%.[4],[5]

Liquid-based cytology (LBC) was initially introduced for gynecological cervical smears. Recently, its utility has been studied in both nongynecological and FNAC material.[6] The two most common methods of LBC are: SurePath™ and Thinprep2000™. Both the methods have two basic steps: (1) Collection of material in an alcohol based medium (2) Automated preparation of a monolayer of cells from the material. SurePath™ the cells are collected in an ethanol-based solution (CytoRich™ red), centrifuged twice then slowly sedimentated onto a poly-l-lysinated slide and eventually stained with a specific hematoxylin–eosin stain, whereas in thinprep2000™, the cells are aspirated in a methanol-based solution (Cytolyt™) then filtered and transferred onto a positively charged slide with a gentle positive pressure. Both the processes have a circular smear in the center of the slide as an result with a sensitivity of 77% and specificity of 81%.[7]

The process of sample collection for LBC of thyroid can be done by two methods: Split sample or direct to vial. In split sample a single pass is done within the nodule and the sample is divided in two parts, first part is used for smear preparation and second part is used for LBC. In direct to vial method two different passes are done within the nodule to prepare smears and for LBC.

In most of the centers conventional smears (CSs) with Giemsa and Papanicolaou (PAP) staining is the standard method used for thyroid FNAC. The aim of the present study is to compare the cytomorphological features of CSs and LBC as well as to elaborate the utility of immunocytochemistry in LBC smears.


  Materials and Methods Top


A total of 100 patients with palpable thyroid nodules were included in the study. The study was conducted from March 2019 to September 2019. FNAC was performed under sterile conditions using 24G needle, 2–3 passes were given and 2–6 direct smears were prepared (both PAP and Giemsa stained). The remaining aspirate in the needle was transferred to 6 ml of Becton Dickinson (BD) Cytorich Red preservative liquid (minimum fixation time: 30 min; preservation time period: 30 days). It was processed using BD SurePath™ method (Tripath imaging-Headquarters in Burlington, US. Patent organization-BD). Fixation is followed by centrifugation for 10 min at 600 g and then the supernatant was discarded. The resultant pellet was washed with 6 ml of distill water and then was thawed and again centrifuged at 600 g for 5 min. The supernatant was again discarded and the tube was loaded in the BD PrepStain™ slide processor, which is a fully automated software. One PAP stained LBC and 1–2 unstained smears were obtained. The unstained smears were used for immunocytochemistry wherever required.

Conventional  Pap smear More Detailss and LBC smears were compared cytomorphologically, a semiquantitative scoring was done [Table 1] and P values were calculated using IBM SPSS software version 24, Chicago, USA. Also sensitivity, specificity and positive predictive values were calculated for both LBC and conventional method. The final diagnosis was made with the help of CS and LBC smear according to Bethesda system for reporting thyroid cytopathology (2007), which includes six diagnostic categories: (a) Category 1: Nondiagnostic, (b) Category 2: Benign, (c) Category 3: Follicular lesion of undetermined significance, (d) Category 4: Suspicious for a follicular neoplasm, (e) Category 5: Suspicious for malignancy, and (f) Category 6: Malignant.[4]
Table 1: Semiquantitative scoring criteria of various parameters

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Adequacy criteria

At least six groups of well-visualized follicular epithelial cells with at least ten cells in each group in a single slide. Abundant colloid in absence of follicular epithelial cells was diagnostic of colloid goiter.

Cytohistological correlation was done in operated cases.


  Results Top


The cellularity on smears was more on CSs than on LBC. This can be attributed to the method of sample collection (split sample). However, three-dimensional collection of cells as well as singly scattered cells was more often in LBC than in CS. It was difficult to appreciate the nuclear and cytological features in these clusters in LBC smears, whereas these were crisper and were very well seen in CS. The individual cells had smaller shrunken falsely hyperchromatic nucleus and the cytoplasm was spilled out of the cells. This is the result of processing steps used to prepare the LBC smears. The background, hemorrhage, inflammatory cells, and colloid in LBC were washed off in most of the cases with some having dense deposits or folded sheet of colloid. The lymphocytic impinging was difficult to appreciate in cases of thyroiditis in LBC. All these differences in between LBC and CS were statistically significant with P < 0.05 [Figure 1] and [Figure 2]; [Table 2].
Figure 1: (a) Colloid in napkin fold pattern on liquid-based cytology (Papanicolaou stain, ×400). (b) Hyperplastic nodule on liquid-based cytology (Papanicolaou stain, ×400). (c) Medullary carcinoma of thyroid showing scattered cells on liquid-based cytology (Papanicolaou stain, ×400). (d) Follicular carcinoma of thyroid showing vague microfollicular pattern (Papanicolaou stain, ×400). (e) Papillary carcinoma of thyroid showing papillary architecture along with singly scattered cells on liquid-based cytology (Papanicolaou stain, ×400). (f) Cytoplasmic positivity of calcitonin in tumor cells in medullary carcinoma on liquid-based cytology (Immunocytochemistry, ×400). (g) Nuclear positivity of thyroid transcription factor one in tumor cells in follicular carcinoma on liquid-based cytology. (Immunocytochemistry, ×400)

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Figure 2: (a) Smear showing sheets of follicular cells in hyperplastic nodule on conventional smear (Giemsa, ×400). (b) Conventional Smear showing lymphocytic impingement in thyroid follicular clusters in lymphocytic thyroiditis (Giemsa, ×200). (c) Conventional smear showing back to back microfollicular arrangement of cells in follicular carcinoma (Giemsa, ×200). (d) Conventional smear showing nuclear inclusions and grooves (arrow) in papillary carcinoma (Pap stain, oil immersion). (e) Conventional smear showing sheets of plasmacytoid cells in medullary carcinoma (Giemsa, ×400)

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Table 2: Semiquantative scoring of various parameters and statistical comparision of these parameters in conventional smears and liquid based cytology

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The number of unsatisfactory cases was higher in LBC in comparison to the conventional method [Table 3]. Furthermore, the LBC smears had higher frequency of suspicious for malignancy cases. This may be due to high N:C ratio and noncrisp nuclear and cytoplasmic features. This distribution of cases among different categories in LBC and CS was statistically significant (P = 0.006). It was very difficult to categorize the thyroid lesions on LBC alone.
Table 3: Category wise comparision of conventional method and Liquid based cytology

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Out of 100 cases 21 underwent surgery and cytohistopathological correlation was done in these [Table 4]. Ten cases were of multinodular goiter, two of follicular carcinoma, eight of papillary carcinoma and one of medullary carcinoma. All the malignant cases were reported as malignant on conventional method whereas overdiagnosis of malignancy was done on LBC smears as most of the cases were reported as suspicious for malignancy on LBC. The sensitivity of CSs and LBC 80.95% and 36.36% for histologically confirmed cases in the present study whereas the positive predictive value was 100% on CSs. The diagnostic correlation between LBC and CS was 80%. Diagnostic accuracy of LBC and CS were 40.7% and 100% respectively for surgically resected cases.
Table 4: Cytohistological correlation of surgically resected cases

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Immunocytochemistry applied on unstained LBC smears showed positivity for thyroid transcription factor one in follicular epithelial cells in hyperplastic nodule, follicular carcinoma and papillary carcinoma whereas calcitonin showed cytoplasmic positivity in follicular epithelial cells in case of medullary carcinoma.


  Discussion Top


LBC is now well established and has achieved great success for gynecological specimens. It has now also gaining popularity for nongynecological specimens due to its possible utility in ancillary investigations and various other reasons.[8] In the present study, we have compared the cytomorphological features as well as diagnostic utility of conventional method and LBC.

LBC has lesser screening time with single slide for screening. The background had almost no red blood cells whereas the CSs had hemorrhagic background although it was not hampering the final impression in all the cases.[9],[10] The colloid was significantly reduced in LBC smears which can be even missed by inexperienced person. As colloid nodule is a common thyroid lesion in general population as well as in this study, the loss of colloid in these cases can falsely label it as nondiagnostic. Lymphocytic impinging was not well appreciated in the cases of thyroidtis in LBC which can lead to diagnostic pitfall in these cases.

The number of unsatisfactory cases were higher in LBC which were all due to hypocellularity. This can be attributed to split sample method of smear preparation in the present study.[10] The number of suspicious for malignancy cases were also higher in LBC due to false hyperchromasia, overlapping dense three dimensional clusters, alteration in nuclear and cytoplasmic characteristics. In contrast to present study, Biscotti et al. found that cell types and cellular arrangements were well preserved in LBC slides and the diagnostic accuracy of both LBC and CS was same.[11] Also, Mesonero and Sickel found no nuclear and cytoplasmic differences in LBC and CS though their diagnostic correlation of both the methods was 90% which was 80% in the present study.[12] Whereas Frost and Cochand-Priollet et al. found higher number of unsatisfactory cases in LBC and they also found variation in nuclear and cytoplasmic features among LBC and CS.[13],[14] The malignant cases are more conveniently diagnosed on CS as their cellular architecture are well maintained and nuclear features like nuclear grooves, inclusions are very well appreciated in CS than in LBC.

CS is an established method with well-known diagnostic criteria for different categories of Bethesda system reporting. It requires a training period so that both the methods can be done in parallel so that the reporting cytopathologists gain self-confidence and can become familiar about the differences of various cytomorphological features of benign and malignant lesions in both LBC and CS.[15]

LBC has an advantage of preparation of additional smears which can be used for immunocytochemistry and special stains. Furthermore, it provides material for possible ancillary investigations which need to be further explored. In the present study, immunocytochemistry was applied on various follicular lesions including hyperplastic nodule, follicular carcinoma, papillary carcinoma, and medullary carcinoma which were positive.


  Conclusion Top


In the present scenario, CS is a cost-effective and more specific as well as sensitive diagnostic method in comparison to LBC. Conventional method does not require any special equipment or personnel training as in case of LBC. LBC can be used for ancillary investigation in conjunction to CS to avoid diagnostic pitfalls but cannot replace the conventional method completely.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Gharib H, Goellner JR, Johnson DA. Fine-needle aspiration cytology of the thyroid. A 12-year experience with 11,000 biopsies. Clin Lab Med 1993;13:699-709.  Back to cited text no. 1
    
2.
Usha Menon V, Sundaram KR, Unnikrishnan AG, Jayakumar RV, Nair V, Kumar H. High prevalence of undetected thyroid disorders in an iodine sufficient adult South Indian population. J Indian Med Assoc 2009;107:72-7.  Back to cited text no. 2
    
3.
Frates MC, Benson CB, Charboneau JW, Cibas ES, Clark OH, Coleman BG, et al. Management of thyroid nodules detected at US: Society of radiologists in ultrasound consensus conference statement. Radiology 2005;237:794-800.  Back to cited text no. 3
    
4.
Cibas ES, Ali SZ; NCI Thyroid FNA State of the Science Conference. The Bethesda system for reporting thyroid cytopathology. Am J Clin Pathol 2009;132:658-65.  Back to cited text no. 4
    
5.
Hamberger B, Gharib H, Melton LJ 3rd, Goellner JR, Zinsmeister AR. Fine-needle aspiration biopsy of thyroid nodules. Impact on thyroid practice and cost of care. Am J Med 1982;73:381-4.  Back to cited text no. 5
    
6.
Yassa L, Cibas ES, Benson CB, Frates MC, Doubilet PM, Gawande AA, et al. Long-term assessment of a multidisciplinary approach to thyroid nodule diagnostic evaluation. Cancer 2007;111:508-16.  Back to cited text no. 6
    
7.
Rossi ED, Fadda G. Thin-layer liquid-based preparation of exfoliative non-gynaecologic and fine-needle aspiration biopsy cytology. Diagn Histopathol 2008;14:563-70.  Back to cited text no. 7
    
8.
Geers C, Bourgain C. Liquid-based FNAC of the thyroid: A 4-year survey with SurePath. Cancer Cytopathol 2011;119:58-67.  Back to cited text no. 8
    
9.
Saleh H, Bassily N, Hammoud MJ. Utility of a liquid-based, monolayer preparation in the evaluation of thyroid lesions by fine needle aspiration biopsy: Comparison with the conventional smear method. Acta Cytol 2009;53:130-6.  Back to cited text no. 9
    
10.
Jung CK, Lee A, Jung ES, Choi YJ, Jung SL, Lee KY. Split sample comparison of a liquid-based method and conventional smears in thyroid fine needle aspiration. Acta Cytol 2008;52:313-9.  Back to cited text no. 10
    
11.
Biscotti CV, Hollow JA, Toddy SM, Easley KA. ThinPrep versus conventional smear cytologic preparations in the analysis of thyroid fine-needle aspiration specimens. Am J Clin Pathol 1995;104:150-3.  Back to cited text no. 11
    
12.
Mesonero CE, Sickel J. Thyroid fine needle aspiration: A comparison of thin-layer slide preparation with conventional smears (abstr). Acta Cytol 1993;37:795.  Back to cited text no. 12
    
13.
Frost AR, Sidawy MK, Ferfelli M, Tabbara SO, Bronner NA, Brosky KR, et al. Utility of thin-layer preparations in thyroid fine-needle aspiration. Cancer Cytopathol 1998;84:17-25.  Back to cited text no. 13
    
14.
Cochand-Priollet B, Prat JJ, Polivka M, Thienpont L, Dahan H, Wassef M, et al. Thyroid fine needle aspiration: The morphological features on ThinPrep slide preparations. Eighty cases with histological control. Cytopathology 2003;14:343-9.  Back to cited text no. 14
    
15.
Rossi ED, Raffaelli M, Minimo C, Mulè A, Lombardi CP, Vecchio FM, et al. Immunocytochemical evaluation of thyroid neoplasms on thin-layer smears from fine-needle aspiration biopsies. Cancer Cytopathol 2005;105:87-95.  Back to cited text no. 15
    


    Figures

  [Figure 1], [Figure 2]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]



 

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