INTRODUCTION
Fine needle aspiration (FNA) is a widely and successfully used method all over the world in the diagnosis of thyroid lesions. FNA is an initial diagnostic test because of its simplicity, safety, and cost-effectiveness. Although its main task is to distinguish between benign and malignant thyroid lesions, it also benefits the patient management process of benign lesions such as thyroiditis.[ 1 , 2 ]
The thyroid FNAs were examined with conventional smears for many years and still are studied with this method in some laboratories. After liquid-based cytology (LBC) technique developed for cervical cytology, it has also been used for nongynecological samples and has been very successful. It provides more convenient slides to evaluate that have less artifacts and clean background. Furthermore, it offers to have extra slides of the samples for further examination and ancillary tests. In addition, cell block can be performed from the residue material.[ 1 , 3 , 4 ]
The purpose of all these techniques was to obtain sufficient material to evaluate and reduce the nondiagnostic (ND) cases. The Bethesda System for Reporting Thyroid Cytopathology (TBSRTC) recommends repeat FNA with ultrasound guidance for ND cases.[ 5 ] Although thyroid FNA is a minimally invasive procedure, it can cause complications.[ 6 ] Decreasing of ND cases will protect patients from complications and morbidity that may arise from repeated FNA.
The study aimed to evaluate the importance of preparing a second LBC slide or cell block from the residue material of thyroid FNA samples in reaching a definitive diagnosis in cases diagnosed as ND after the first slide.
MATERIAL AND METHODS
Seventy five cases diagnosed as ND after the first slide were included in study. For 50 cases, the second LBC slides were prepared (LBC group); for 25 cases, cell block was made from residue material (CB group). None of the cases in both groups had previously formed a cell block.
Cytological preparation
BD SurePath pap test kit (BD Diagnostics-tripath, Burlington, NC, USA) was used for additional LBC slides. The sample in ethanol-based fixative was centrifuged two times. Then cellular sample was mixed with a vortex homogeneously. The sample spread as a thin layer in a circular area of the microscope slide and stained with PAP. For cell block, the residue material in the vial was centrifuged one time.
The tissues at the bottom of the tube were fixed in formalin and stained with hematoxylin and eosin for further histological examination.
Cytological examination
The PAP-stained second LBC slides and hematoxylin and eosin slides of cell blocks were evaluated under the light microscope. The cases were categorized as per TBSRTC into the following six categories such as Nondiagnostic, Benign, Atypia of undetermined significance/follicular lesion of undetermined significance (AUS/FLUS), Follicular neoplasm/Suspicious for follicular neoplasm, Suspicious for malignancy (SM), and Malignancy.[ 5 ]
Histological follow-up was available in eight cases (six in LBC group, two in CB group). Results of resection were noted.
Statistical analysis
SPSS.25 program was used for statistical evaluation. A descriptive analysis was carried out in which nominal variables were shown as number of cases and the percentage (%). The independent sample test was used to compare proportions within the two groups. A P value <.05 was considered significant.
RESULTS
A total of 75 ND cases were included in the study. At the end of secondary procedures, a definitive diagnosis was reached in a total of 24 cases (32%) [Table 1 ]. Twenty one cases were categorized as benign and three cases were categorized as AUS/FLUS as per TBSRTC.
Table 1: Comparison of groups in terms of reaching a definitive diagnosis
The second LBC slides were prepared for 50 cases (LBC group). Cell blocks were made from the remaining material for 25 cases (CB group). Twenty of 50 cases (40%) in LBC group reached a definitive diagnosis, while four cases (16%) in the CB group reached a definitive diagnosis. Achieving a definitive diagnosis was found higher in the LBC group in which the second slide was formed compared to the group in which the cell block was formed. There was statistically significant difference in the comparing of groups in terms of reaching a definitive diagnosis (P =0.036) [Table 1 ].
Among the cases that reached a definitive diagnosis, in LBC group, three cases were categorized as AUS/FLUS and 17 cases categorized as benign and all the four cases in CB group were categorized as benign as per TBSRTC. There were no cases in other TBSRTC category such as Follicular neoplasm/Suspicious for follicular neoplasm, Suspicious for malignancy, and Malignancy [Table 2 ].
Table 2: The Bethesda categories of cases after secondary procedures
Among the cases that reached a definitive diagnosis, three cases diagnosed as AUS/FLUS with focal cytological atypia. In benign category, lymphocytic thyroiditis was seen in one case and four cases diagnosed as follicular nodule with cystic degeneration. Although they were insufficient for diagnosis, in 18 of ND cases follicular epithelium was seen, three of ND cases had only cystic content. Diagnostic cells could not be obtained in 30 cases. Only fibrinous material or inflammatory cells were present [Table 3 ]. Histological follow-up was available in eight cases (six in LBC group, two in CB group). Four out of eight cases were malignant (50%). Malignancy rates were 33.3% (2/6) and 100% (2/2) in LBC and CB groups, respectively.
Table 3: Diagnostic distribution of cases after secondary procedures
DISCUSSION
In TBSRTC, ND category applies to samples that are nondiagnostic or unsatisfactory due to obscuring blood, overly thick smears, air drying of alcohol-fixed smears, or an inadequate number of follicular cells. In LBC method, these situations that cause inadequacy are less common and the number of ND cases is low when compared to the conventional method.[ 7 , 8 ]
There are so many factors that influence nondiagnostic rates of FNA results, including the skill of the operator, vascularity of the nodule, and the cystic component of the nodule.[ 7 ] In our study, we only investigated whether it would be possible to reduce the ND rates in the LBC method with different applications (additional slides or cell blocks). Also, we were able to reach a definitive diagnosis in 32% of the total cases.
In a recent study, it was stated that a definitive diagnosis was reached in 23 of 39 cases in which the second slide was created with the liquid-based method in thyroid FNA specimens with ND on the first slide.[ 1 ] In our study, a definitive diagnosis was reached in 40% of LBC group that second slide created. When we compared the results with the CB group, statistically higher definitive diagnoses were obtained in LBC group. More follicle epithelium was obtained in the LBC group [Figure 1 ]. In the LBC technique, if there is technique-related degeneration on the first slide, the decrease rate on the second slide is high. In addition, if there is a technical reason such as a pipette clogging on the first slide, the probability of seeing sufficient follicle epithelium on the second slide is quite high. However, there may be an increase in fibrinous material in second slide secondary to the waiting of residual material and bleeding during FNA.
Figure 1: In the case where no follicle epithelium was observed on the first LBC slide (1A- PAPx100), the second slide showed sufficient follicle epithelium for evaluation (1B- PAPx200, 1C-PAPx200). In another case, there is only a few follicles epithelium in the first LBC slide (1D- PAPx400), but in second slide there are follicle epithelium, whose cellular details can be easily seen and colloidal material are observed (1E- PAPx400, 1F- PAPx400)
In cervical cytology, the using of LBC techniques has been evaluated broadly and stated that a single LBC slide can represent the specimen as a whole and cell block not to proceed with extra information which is not represented on the original slide.[ 9 , 10 ] However, in nongynecological samples, additional LBC slides or cell blocks reduce ND rates and increase definitive diagnosis.[ 1 , 10 ]
Preparing cell blocks may provide diagnostic benefits in diagnosis malignant lesions such as papillary thyroid carcinoma.[ 11 ] In our study, a definitive diagnosis was reached in 16% of CB group. Cell blocks from residue LBC material are no more advantageous than a second LBC slide. When both groups were compared, epithelium was observed in 10 of 30 cases in LBC group and eight of 21 in CB group in cases that remained ND category after secondary procedures [Table 3 ].
Nondiagnostic or unsatisfactory results cause anxiety to the clinicians and/or the patient because of the high risk of malignancy rates of ND cases.[ 5 , 7 ] The risk of malignancy for ND category is difficult to evaluate because only a small group of cases undergo surgery. In the literature, the malignancy rates of ND cases show obvious variation, ranging from 2% to 51%, and it is 5%-10% in TBSRTC.[ 5 , 12–15 ] In present study, histopathological follow-up was available only in eight cases (six cases in LBC group, two cases in CB group). Four of 8 cases (50%) were diagnosed as malignant after surgery.
This study has a few limitations. The number of cases were limited, and histopathological follow-up was available only in eight cases.
CONCLUSION
The present study shows that preparing a second slide with LBC method is more purposive than preparing a cell block from residue material of thyroid FNA samples.
Although there were a few numbers of cases with histopathological confirmation, the malignancy risk was very high in nondiagnostic cases.
Reducing the percentage of ND cases will protect patients from complications and morbidity that may arise from repeated FNA.
Data availability
The datasets generated and/or analyzed during the present study are not publicly available but are available from the corresponding author on reasonable request.
Ethical approval
This study was found appropriate ethically as per the ethical committee of the Basaksehir Cam and Sakura City Hospital with its decision number 2021-228.
Author contributions
S.E.D. made substantial contributions to conception, data curation, methodology, analysis and data interpretation, wrote the final manuscript, and managed the overall progress of the study. Z.B.E. conceptualized the study design, analyzed and interpreted the data, and checked the final version of the article. O.Y. helped in providing and analyzing data, literature research, and edited the final version of the article.
Financial support and sponsorship
Nil.
Conflicts of interest
There are no conflicts of interest.
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