Prostate-specific antigen: An unfamiliar protein in the human salivary glands.

OBJECTIVES: The presence of prostate-specific antigen (PSA) in saliva and salivary glands has been reported. Nevertheless, its release pathway in these glands remains to be elucidated. Here, we showed PSA subcellular distribution focusing on its plausible route in human salivary parenchyma. MATERIALS AND METHODS: Sections of parotid and submandibular glands were subjected to the immunohistochemical demonstration of PSA by the streptavidin-biotin method revealed by alkaline phosphatase. Moreover, ultrathin sections were collected on nickel grids and processed for immunocytochemical analysis, to visualize the intracellular distribution pattern of PSA through the observation by transmission electron microscopy. RESULTS: By immunohistochemistry, in both parotid and submandibular glands PSA expression was detected in serous secretory acini and striated ducts. By immunocytochemistry, immunoreactivity was retrieved in the cytoplasmic compartment of acinar and ductal cells, often associated with small cytoplasmic vesicles. PSA labeling appeared also on rough endoplasmic reticulum and in the acini's lumen. A negligible PSA labeling appeared in most of the secretory granules of both glands. CONCLUSIONS: Our findings clearly support that human parotid and submandibular glands are involved in PSA secretion. Moreover, based on the immunoreactivity pattern, its release in oral cavity would probably occur by minor regulated secretory or constitutive-like secretory pathways.

FK962 protects retinal ganglion cell under hypoxia/reoxygenation: Possible involvement of glial cell line-derived neurotrophic factor signaling pathway.

Loss of retinal ganglion cells (RGCs) is the cause of visual impairment and blindness in glaucoma. Previously, our studies showed that FK962 (N-[1-acetylpiperidin-4-yl]-4-fluorobenzamide) promoted neurite elongation in rat RGCs and trigeminal ganglion (TG) cells. In TG cells, glial cell line-derived neurotrophic factor (GDNF) is known to be involved in the mechanism. The purpose of the present study is to investigate whether, 1) FK962 shows an RGC-protective effect under hypoxia/reoxygenation (H/R) and 2) GDNF is involved in the neuroprotective mechanism of FK962. Rat primary retinal cells were cultured under 24-h hypoxia/24-h reoxygenation conditions, with or without FK962, recombinant GDNF, GDNF antibody and RET receptor tyrosine kinase inhibitor, GSK3179106. Cells were co-immunostained with RBPMS and Neurofilament 200 as a RGC marker, and the number of survived RGCs was counted. Results showed H/R treatment decreased the number of survived RGCs. FK962 promoted RGC survival under H/R by a bell-shaped dose response, with the highest RGC-protective effect of 10-8 M. The protective effect was the same level with 10-12 M exogenous GDNF. Addition of GDNF antibody or GSK3179106 counteracted the neuroprotective effect of FK962. From these results, it is suggested that FK962 ameliorates RGC death under H/R, possibly via a GDNF signaling pathway.

Comparative examination of a rapid immunocytochemical test for the detection of highly pathogenic avian influenza virus in domestic birds in field outbreaks.

The quantitative real-time reverse polymerase chain reaction (RRT-PCR) is the preferred test method for the diagnosis of avian influenza (AI), but can be performed only in specialized laboratories. Different antigen detection methods for the diagnosis of AI were previously reported to be specific and sensitive in field outbreaks. These tests can be performed in basic countryside labs. Brain smears of domestic birds (n = 105) collected during AI field outbreaks were examined with immunocytochemistry (IC). The results were statistically analysed by comparing IC to brain histology (BH), and immunohistochemistry (IHC), to gross pathological examination (GP) (n = 105), and RRT-PCR (n = 91). AI was diagnosed with RRT-PCR in 66 cases. IC and IHC were positive in 59/66 (90%) and 60/66 (91%) cases, respectively. Lesions suspicious for AI were detected with GP and HP in 66/66 (100%) and 61/66 (92%) cases, respectively. An almost perfect agreement was found between RRT-PCR, IC, IHC, and HP. Substantial agreement was found between IC and GP, between IHC and GP, between HP and GP, and between RRT-PCR and GP. The chromogen-based IC test presented in this study produces durable staining, which can be evaluated using a simple brightfield microscope. The test is rapid (can be completed in 2 h), sensitive (90%), specific (100%), and cost-effective, which makes the method suitable for routine diagnostic tests in AI epidemics.RESEARCH HIGHLIGHTSAvian influenza virus (AIV) antigen detection was examined in field outbreaks.Bird brain smears were tested using immunocytochemistry (IC).IC results strongly correlated with real-time RT-PCR results.The IC method was rapid, specific, sensitive, and cost-effective in AIV field outbreaks.

Expression of Tshb and Tshr in the ricefield eel Monopterus albus: Potential paracrine/autocrine roles in gonads.

Thyroid stimulating hormone (TSH), a glycoprotein synthesized and secreted from thyrotrophs of the pituitary gland, is composed of a glycoprotein hormone common alpha subunit (CGA) and a specific beta subunit (TSHB). The major biological function of TSH is to stimulate thyroidal follicles to synthesize and secrete thyroid hormones through activating its cognate receptor, the thyroid stimulating hormone receptor (TSHR). In the present study, polyclonal antisera against ricefield eel Tshb and Tshr were generated respectively, and the expression of Tshb and Tshr was examined at mRNA and protein levels. RT-PCR analysis showed that tshb mRNA was expressed mainly in the pituitary as well as in some extrapituitary tissues including the ovary and testis. Tshr mRNA was also expressed in a tissue-specific manner, with transcripts detected in tissues including the kidney, ovary, and testis. The immunoreactive Tshb signals in the pituitary were shown to be localized to the inner areas of adenohypophysis which are close to the neurohypophysis of adult ricefield eels. Tshb-immunoreatvie cells in the pituitary of ricefield eel larvae were firstly observed at hatching. The expression of immunoreactive Tshb and Cga was also detected in ricefield eel ovary and testis together with Tshr. In the ovary, immunoreactive Tshb, Cga, and Tshr were observed in oocytes and granulosa cells. In the testis, immunoreactive Tshb was mainly observed in Sertoli cells while immunoreactive Cga and Tshr were detected in germ cells as well as somatic cells. Results of the present study suggest that Tsh may be synthesized both in the ovary and testis locally, which may play paracrine and/or autocrine roles in gonadal development in ricefield eels.

Establishment and characterization of a novel cell line and xenotransplant mouse model derived from feline colorectal adenocarcinoma.

Colorectal adenocarcinoma is an aggressive malignant tumor in cats that frequently metastasizes to the lymph nodes and/or distant organs. However, research on feline colorectal adenocarcinoma is limited, and experimental models have not been established. A novel cell line, FeLeco-G7, was established from the lymph node of a 12-year-old spayed female Maine Coon cat with metastatic colorectal adenocarcinoma. FeLeco-G7 cells were polygonal with abundant cytoplasm and adherent growth. The population-doubling time was approximately 28.3 hours, and the mean number of chromosomes was 37.6±0.1 per cell (ranging between 32 and 41). Consistent with the original tumor, FeLeco-G7 cells were immunopositive for cytokeratin (CK) 20 and CDX2, and immunonegative for CD10 and CK7. Nuclear accumulation of ß-catenin was rarely observed. Mutation analysis suggested TP53 gene alterations. A subcutaneous injection of FeLeco-G7 cells into immunodeficient mice resulted in the formation of a mass at the injection site without the development of metastatic lesions. An orthotopic (intrarectal) transplantation of FeLeco-G7 cells caused cachexia and diffuse involvement of the rectal mucosa in one of the 3 mice and the formation of masses around the rectum in the other 2 mice. Metastases to the regional lymph nodes and lungs were detected in three of the 3 and one of the 3 mice, respectively. The histological findings and immunohistochemical features of these masses were similar to those of the original tumor. These results suggest that FeLeco-G7 cells and the orthotopically transplanted mouse model are valuable tools for further molecular and therapeutic research on feline colorectal adenocarcinoma.

Practical issues related to immunocytochemistry on cytological smears: Tips and recommendations.

OBJECTIVE: Immunocytochemistry (ICC) is essential for enhancing diagnostic accuracy and identifying markers for diagnosis, prognosis and targeted therapies. While cell blocks (CBs) are preferred for standardization and optimized staining, cytological smears are an alternative when CBs are unavailable. However, the literature on ICC protocols for smears is sparse. This review addresses preparation, fixation and protocols for nuclear and cytoplasmic antibodies on smears, drawing from our laboratory's experience. METHODS: We reviewed procedures for ICC on cytological smears using existing literature and practical insights from our laboratory. RESULTS: Commercially available antibodies were found to be reliable for ICC on smears if specimens are properly prepared and fixed. Protocols developed in our laboratory maintained antigenicity and provided clear staining results. CONCLUSIONS: Although ICC on CBs is the gold standard for standardization, cytological smears are a viable alternative when CBs are unavailable. Success in ICC on smears depends on proper preparation and fixation. This review offers practical protocols and insights to help laboratories optimize ICC on cytological smears. Further research and standardization are necessary to enhance reproducibility and reliability of ICC on smears. The practical information provided is based on personal experience in our laboratory.

Immunocytochemical markers, molecular testing and digital cytopathology for aspiration cytology of metastatic breast carcinoma.

Fine-needle aspiration cytology (FNAC) is a versatile diagnostic procedure uniquely suited for tissue biopsy of breast carcinomas and axillary metastases and/or recurrences. With the expanding treatment options and accompanying theragnostic tests, it is crucial to recognize the developments on ancillary testing and digital cytopathology techniques related to aspiration cytology of metastatic breast carcinoma. In this review, we aim to summarize and update the evidence of immunocytochemistry, for the detection of carcinoma cells (epithelial markers), confirmation of breast primary (breast-specific markers), assessment of surrogate immunostains (hormone receptors, ki-67 proliferative index and HER2) and theragnostic biomarkers, with discussion on potential diagnostic pitfalls, followed by the application of molecular tests, and digital cytopathologic techniques for assessing metastatic breast carcinoma in cytology.

Myelomatous meningitis diagnosed by cerebrospinal fluid cytology examination.

Myelomatous meningitis diagnosed by CSF cytology. The combined use of cytology with immunocytochemistry can identify the presence of multiple myeloma cells in cerebrospinal fluid specimens.

A sub-hepatic nodule in a young female: Chase the case.

Sertoli-Leydig cell tumours (SLCTs) are rare, mixed sex-cord stromal tumours composed of varying proportions of both Sertoli and Leydig cells, which account for <0.5% of all ovarian tumours. The cytomorphologic features of SLCTs are not well described in literature. Herein, we describe the cytomorphologic features of an SLCT at an uncommon metastatic site in a young female. Sertoli-Leydig cell tumours (SLCTs) are rare, mixed sex-cord stromal tumours composed of varying proportions of both Sertoli and Leydig cells, which account for <0.5% of all ovarian tumours. The cytomorphologic features of SLCTs are not well described in literature. Herein, we describe the cytomorphologic features of an SLCT at an uncommon metastatic site in a young female.

Some uncommon cystic lesions in the anterior head and neck region: Pitfalls to be avoided on cytology.

Cystic lesions of the anterior head and neck region are a challenging and frequent finding on cytological smears. The scant amount of cellular material in cystic slides poses the greatest difficulty to interpretation, so that frequently they are diagnosed as inadequate or with minimal cellular component. Despite the majority of cystic lesions being benign, a minor portion consist of malignant cystic entities. In these latter cases, the evidence of very scant malignant cells can be misdiagnosed and/or underestimated, leading to a false negative diagnosis. Many papers have already described and detailed the range of possible benign and malignant cystic lesions in head and neck. In the current review we have focused on the less common entities that often lead to serious misinterpretation.

Dual Sec62/Ki67 immunocytochemistry of liquid-based cytological preparations represents a highly valid biomarker for non-invasive detection of head and neck squamous cell carcinomas.

BACKGROUND: Head and neck squamous cell carcinomas (HNSCC) are frequently diagnosed in advanced stages, which limits therapeutic options and results in persistently poor patient outcomes. The aim of this study was to use liquid-based swab cytology (LBC) in combination with dual immunocytochemical detection of migration and proliferation markers Sec62 and Ki67 in order to allow non-invasive early detection of HNSCC as well as to analyse the diagnostic validity of this method for predicting the malignancy of suspicious oral lesions. METHODS: 104 HNSCC patients and 28 control patients, including healthy patients (n = 17), papilloma (n = 1) and leukoplakia patients (n = 10), were included in this study. For all patients, an LBC swab followed by simultaneous immunocytochemical detection of Sec62 and Ki67 was performed. Immunocytochemical as well as cytopathological results were correlated with histological diagnoses and clinical findings. RESULTS: All HNSCC patients (100%) showed dual Sec62/Ki67 positivity, and all control patients except for the papilloma patient were negative for Sec62/Ki67 (96.4%), resulting in a 100% sensitivity and 96.4% specificity of Sec62/Ki67 dual stain for non-invasive detection of HNSCC. The positive predictive value was 99% and the negative predictive value was 100%. Sec62 expression levels showed a positive correlation with tumour de-differentiation (p = 0.0489). CONCLUSION: Simultaneous immunocytochemical detection of Sec62/Ki67 using LBC represents a promising non-invasive and easy-to-apply tool for the early detection of HNSCC in routine clinical practice. This novel technique can help to avoid incisional biopsies and reduce the frequency with which general anaesthesia is used in diagnostic procedures in patients with suspicious oral lesions.

Novel Molecular Mechanisms Underlying the Ameliorative Effect of Platelet-Rich Plasma against Electron Radiation-Induced Premature Ovarian Failure.

Radiotherapy is one of the risk factors for radiation-induced premature ovarian failure and infertility in cancer patients. The development of methods for ovarian radioprotection remains relevant. Moreover, electrons are a little-studied and promising method of radiation with the least toxic effect on normal tissues. The assessment of intracellular mechanisms regulating the protective effects of leukocyte-poor platelet-rich plasma in a model of radiation-induced premature ovarian failure caused by electron irradiation. Wistar rats were divided into four groups, namely a control group, irradiation group (electron exposure), irradiation + leukocyte-poor platelet-rich plasma group, and only leukocyte-poor platelet-rich plasma group. Fragments of ovaries were removed and hormonal, oxidant, histological, and morphometric studies were carried out. The cell cycle of ovarian follicles and the inflammatory and vascular response were assessed using immunohistochemistry. The activity of MAPK, ERK, and PI3K pathways was also assessed using the RT-qPCR. We found that electron irradiation causes a decrease in the functional activity of the ovaries and the death of follicular cells through apoptosis. The administration of LP-PRP led to a partial restoration of the cytokine balance. In addition, minor ovarian damage and mild inflammation were observed in this group. Leukocyte-poor platelet-rich plasma components have anti-inflammatory, angiogenetic, and radioprotective effects, reducing the activation of the NOX4, caspase and cytokine cascades, and inflammatory response severity through the MAPK/p38/JNK signaling pathway. This leads to the induction of endogenous antioxidant protection, the repair of post-radiation follicular damage, and slowing down the development of radiation-induced premature ovarian failure after electron irradiation.

The polychromatism of postmortem cerebrospinal fluid.

Based on the assumption that postmortem cerebrospinal fluid (CSF) is contaminated depending on the chosen sampling technique in the forensic setting resulting in bloody or at least hemolytic CSF samples, we systematically documented a total of 183 postmortem CSF samples. These samples were all assessed for their quality and color, regardless of the cause of death or the postmortem interval. The investigations were carried out through subjective assessment of color and turbidity, as well as objective measurements of the optical density (OD) of the CSF supernatants after centrifugation of each sample, with standardized photographic documentation. The observations revealed that in 28 cases the CSF was absolutely (crystal-) clear and transparent. Most of our samples showed color changes ranging from xanthrochromic to rose. Intensive staining of the supernatants was only found in a small proportion of the examined collective. We found that postmortem CSF has no uniform appearance but rather a diverse range of color spectra, and the color, as well as the OD of the CSF, correlates significantly with the postmortem interval (p < 0.001) when sampled using the proposed standard procedure.

SARS-CoV-2 virus associated angiotensin converting enzyme 2 expression modulation in colorectal cancer: Insights from mRNA and protein analysis COVID-19 associated (ACE2) expression in colorectal cancer.

The SARS-CoV-2 virus gains entry into human cells by exploiting the angiotensin-converting enzyme 2 (ACE2), a key component known as the spike protein (S), as a point of entry. Initially, SARS-CoV-2 suppresses the natural function of ACE2, leading to a gradual decline in cell health. Additionally, individuals with cancer are considered more susceptible to COVID-19. This study investigates the expression patterns of ACE2 in colorectal cancer (CRC) patients with and without a history of COVID-19 infection. RT-PCR was used to analyze samples from both cancerous and adjacent non-affected colorectal tissues of 47 CRC patients, comprising two groups: 24 CRC patients with no history of COVID-19 and 23 CRC patients with a recent history of COVID-19 infection. Epithelial CR cells were isolated from both types of tissues and cultured to evaluate cell adhesion. Immunohistochemistry analyses were conducted to examine ACE2 protein expression using various ACE2 antibodies for both cell types. The study revealed ACE2 mRNA expression in all CRC tissues of patients with and without a history of COVID-19. ACE2 expression was significantly higher in CRC patients without a history of COVID-19. Notably, the non-affected colorectal cancer (NACRC) tissues of patients without a history of COVID-19 also showed ACE2 expression, whereas no ACE2 expression was detected in the biopsies of CRC patients with a positive COVID-19 history. ACE2 antibodies were employed to validate ACE2 protein expression at the mRNA level. COVID-19 appears to downregulate ACE2 expression in both CRC and NACRC tissues of CRC patients with a positive history of COVID-19 infection.

Identification and characterisation of PRXamide peptides in the western flower thrips, Frankliniella occidentalis.

Insect CAPA-PVK (periviscerokinin) and pyrokinin (PK) neuropeptides belong to the PRX family peptides and are produced from capa and pyrokinin genes. We identified and characterised the two genes from the western flower thrips, Frankliniella occidentalis. The capa gene transcribes three splice variants, capa-a, -b, and -c, encoding two CAPA-PVKs (EVQGLFPFPRVamide; QGLIPFPRVamide) and two PKs (ASWMPSSSPRLamide; DSASFTPRLamide). The pyrokinin mRNA encodes three PKs: DLVTQVLQPGQTGMWFGPRLamide, SEGNLVNFTPRLamide, and ESGEQPEDLEGSMGGAATSRQLRTDSEPTWGFSPRLamide, the most extended pheromone biosynthesis activating neuropeptide (PBAN) ortholog in insects. Multiple potential endoproteolytic cleavage sites were presented in the prepropeptides from the pyrokinin gene, creating ambiguity to predict mature peptides. To solve this difficulty, we used three G protein-coupled receptors (GPCRs) for CAPA-PVK, tryptophan PK (trpPK), and PK peptides, and evaluated the binding affinities of the peptides. The binding activities revealed each subfamily of peptides exclusively bind to their corresponding receptors, and were significant for determining the CAPA-PVK and PK peptides. Our biological method using specific GPCRs would be a valuable tool for determining mature peptides, particularly with multiple and ambiguous cleavage sites in those prepropeptides. Both capa and pyrokinin mRNAs were strongly expressed in the head/thorax, but minimally expressed in the abdomen. The two genes also were clearly expressed during most of the life stages. Whole-mounting immunocytochemistry revealed that neurons contained PRXamide peptides throughout the whole-body: four to six neurosecretory cells in the head, and three and seven pairs of immunostained cells in the thorax and abdomen, respectively. Notably, the unusual PRXamide profiles of Thysanoptera are different from the other insect groups.

The earliest events in BRAF-mutant colorectal cancer: exome sequencing of sessile serrated lesions with a tiny focus dysplasia or cancer reveals recurring mutations in two distinct progression pathways.

Around 15-30% of colorectal cancers (CRC) develop from sessile serrated lesions (SSLs). After many years of indolent growth, SSLs can develop dysplasia and rapidly progress to CRC through events that are only partially understood. We studied molecular events at the very early stages of progression of SSLs via the MLH1-proficient and deficient pathways to CRC. We collected a cohort of rare SSLs with a small focus (<10 mm) of dysplasia or cancer from the pathology archives of three hospitals. Whole-exome sequencing was performed on DNA from nonprogressed and progressed components of each SSL. Putative somatic driver mutations were identified in known cancer genes that were differentially mutated in the progressed component. All analyses were stratified by MLH1 proficiency. Forty-five lesions with a focus dysplasia or cancer were included, of which 22 (49%) were MLH1-deficient. Lesions had a median diameter of 10 mm (interquartile range [IQR] 8-15), while the progressed component had a median diameter of 3.5 mm (IQR 1.75-4.75). Tumor mutational burden (TMB) was high in MLH1-deficient lesions (23.9 mutations per MB) as compared to MLH1-proficient lesions (6.3 mutations per MB). We identified 34 recurrently mutated genes in MLH1-deficient lesions. Most prominently, ACVR2A and RNF43 were affected in 18/22 lesions, with mutations clustered in three hotspots. Most lesions with RNF43 mutations had concurrent mutations in ZNRF3. In MLH1-proficient lesions APC (10/23 lesions) and TP53 (6/23 lesions) were recurrently mutated. Our results show that the mutational burden is exceptionally high even in the earliest MLH1-deficient lesions. We demonstrate that hotspot mutations in ACVR2A and in the RNF43/ZNRF3 complex are extremely common in the early progression of SSLs along the MLH1-deficient serrated pathway, while APC and TP53 mutations are early events in the the MLH1-proficient pathway. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Pericardial malignant mesothelioma diagnosed in a dog by immunocytochemistry of the pericardial fluid: a case report.

BACKGROUND: Pericardial effusions are one of the most common cardiac diseases in dogs. Common causes of haemorrhagic pericardial effusions include neoplasia, such as hemangiosarcoma, mesothelioma, chemodectoma, and ectopic thyroid tumours, and benign idiopathic pericardial effusion. Distinguishing among reactive mesothelial cells, malignant mesothelioma, and adenocarcinoma in body effusions is a diagnostic challenge. Therefore, the author aimed to discover whether the observed cells were reactive mesothelial, mesothelioma, or adenocarcinoma cells through immunocytochemistry using five markers (cytokeratin, vimentin, desmin, E-cadherin, and calretinin) in a canine patient. CASE PRESENTATION: A 2.1 kg, spayed female, 10-year-old Yorkshire Terrier dog presented to a local hospital with dyspnoea and was evaluated for pericardial effusion. The presence of pericardial fluid was confirmed, and she was referred to our hospital for further evaluation. In cytological evaluation, cells shed individually or in clusters were observed, along with numerous non-degenerative neutrophils and macrophages. The cells showed binucleation, anisocytosis, anisokaryosis, abnormal nucleoli, abundant basophilic cytoplasm, high nuclear-cytoplasmic ratio, and coarse chromatin. Large atypical multinucleate cells were also observed. Erythrophagia was observed, indicating chronic haemorrhage. Immunocytochemistry using pericardial fluid was positive for cytokeratin, vimentin, desmin, E-cadherin, and calretinin. Therefore, malignant mesothelioma was diagnosed. CONCLUSIONS: Immunocytochemistry is a very useful diagnostic technique because it can determine whether several fluorescent markers are simultaneously expressed in the same cell. Further, E-cadherin and calretinin can be used for the differential diagnosis of reactive mesothelial cells, malignant mesothelioma, and adenocarcinoma in dogs.

Clinicopathological significance of TUBB3 in upper tract urothelial carcinoma and possible application in urine cytology.

ßIII-Tubulin, encoded by the TUBB3 gene, is a microtubule protein. We previously reported that TUBB3 is overexpressed in renal cell carcinoma. We investigated the clinicopathological significance of TUBB3 in upper tract urothelial carcinoma (UTUC) by immunohistochemistry. In normal tissue, TUBB3 expression was weak or absent. In contrast, TUBB3 overexpression was observed in urothelial carcinoma (UC) tissues in 51 (49%) of 103 UTUC cases. TUBB3 overexpression was associated with nodular/flat morphology, high-grade disease, high T stage, and a poor prognosis. Similar results were obtained in The Cancer Genome Atlas bladder cancer cohort. TUBB3 expression was also associated with high Ki-67 labeling index, CD44v9, HER2, EGFR, and p53 expression in UTUC. Among representative cancer-related molecules, TUBB3 was an independent predictor of progression-free survival and high-grade UC. Finally, using urine cytology samples, we analyzed TUBB3 expression by immunocytochemistry. TUBB3 expression was more frequently found in UC cells than in nonneoplastic cells. The diagnostic accuracy of urine cytology was improved when combined with TUBB3 immunostaining. The findings suggest the importance of TUBB3 in tumor progression and its potential application as a biomarker for high-grade disease and the prognosis of UC. Moreover, combination with TUBB3 immunostaining might improve the diagnostic accuracy of urine cytology.

Quality of cell blocks prepared from residual pleural effusion and bronchial washing samples for immunocytochemistry.

INTRODUCTION: Cell blocks (CBs) enable the long-term preservation of cytological samples. The aim of this study was to analyse the quality of CBs prepared from leftover fluid from lung adenocarcinoma pleural effusion samples and residual bronchial washing sediment for immunocytochemistry. METHODS: The residual part of 455 lung adenocarcinoma pleural effusion samples and sediment from 384 bronchial washing samples were used to prepare CBs following the agarose method. The quality of CBs was evaluated based on the quantity of malignant cells in haematoxylin and eosin-stained slides and interpreted as optimal or insufficient for immunocytochemistry. Immunocytochemistry on CBs was performed using the Dako EnVision™ FLEX detection visualisation system. The CB results for TTF-1, ALK, and PD-L1 immunocytochemistry were compared with the corresponding cytological smears. RESULTS: Among all CBs, 202 (44.4%) from leftover pleural effusion fluid and 85 (22.1%) from residual bronchial washing sediment had an optimal number of lung adenocarcinoma cells. Eight pleural effusion CBs were stained for TTF-1. Four pleural effusion and two bronchial washing CBs were stained for ALK and PD-L1. All tested pleural effusion CBs were confirmed positive for TTF-1 and negative for ALK. The PD-L1 tumour proportion score (TPS) was ≥ 50% in two pleural effusions. ALK was confirmed negative in bronchial washing CBs. One bronchial washing CB was interpreted as PD-L1-negative while the corresponding smear was positive (TPS ≥1%; 2%). CONCLUSION: The CB results of TTF-1, ALK, and PD-L1 corresponded to the findings for the smears. The inclusion of CBs prepared from leftover fluid from pleural effusion samples and residual bronchial washing sediment in routine cytological practice could provide a source of high-quality material for immunocytochemistry in addition to smears and cytospins.

Utility of parathyroid hormone immunocytochemistry in fine needle aspiration diagnosis of parathyroid tissue.

OBJECTIVE: The cytomorphological features of parathyroid tissue (PTT) may overlap with those of thyroid lesions, thus posing a diagnostic challenge. In this retrospective study, we reviewed our institutional experience in using parathyroid hormone (PTH) immunocytochemistry (ICC) to substantiate the diagnosis of PTT on fine needle aspiration (FNA). METHODS: Our pathology database was searched for FNA cases in which PTH ICC was performed between 1 January 2015 and 31 March 2022. PTH ICC was performed on a ThinPrep slide in cases with a clinical suspicion of PTT or with cytomorphological features raising the possibility of PTT. Patients' clinicopathological characteristics, PTH ICC results, cytological diagnoses, and surgical follow-ups, if available, were reviewed and analysed. RESULTS: The study cohort included 103 cases clinically designated as thyroid (n = 85, 82.5%), parathyroid (n = 11, 10.7%) and neck soft tissue (n = 7, 6.8%). PTH immunostaining was negative, positive, and indeterminate in 53 (51.5%), 27 (26.2%), and 23 (22.3%) cases, respectively. Surgical follow-up was available in 27 (26.2%) cases, including 17 thyroid lesions and 10 PTT cases. All positive PTH cases were confirmed to be PTT, while all but one of the negative PTH cases were non-PTT on follow-up. The calculated sensitivity, specificity, positive and negative predictive values were 85.7%, 100%, 100% and 93.3%, respectively. CONCLUSION: Our study demonstrates that PTH ICC performed on additional ThinPrep slides is a valuable adjunct test in FNA samples with a differential diagnosis of PTT vs non-PTT. Low cellularity may be a limiting factor in the accurate assessment of PTH by ICC.