Methods in molecular biology (Clifton, N.J.)Natália G Salomão, Kíssila Rabelo
Histopathological analysis of tissues infected with Chikungunya virus (CHIKV), including biopsy and autopsy specimens, can provide valuable insights into the pathogenesis of atypical and fatal cases. By examining tissue architecture and cellular alterations under the microscope, it is possible to identify patterns of injury, inflammation, and cellular degeneration. These morphological findings provide evidence of how the pathogen interacts with host cells and tissues. Moreover, immunohistochemistry may be performed in tissue sections for the detection of viral antigens. Importantly, this technique allows the characterization of diverse biologically relevant targets in tissue samples, such as distinct immune cell subsets, cytokines, and additional molecular markers. This is accomplished by the use of antibodies selected for their specificity toward the antigen of interest. In this chapter, we present a concise overview of how histological analysis and immunohistochemical approaches can enhance the understanding of CHIKV-associated pathological mechanisms.
Methods in molecular biology (Clifton, N.J.)Samna Sagadevan, Paloma Ordóñez-Morán
Intestinal stem cell (ISC)-derived organoids provide a physiologically relevant 3D culture system to model intestinal biology, regeneration, and disease mechanisms. Their complex architecture, recapitulating crypt structures, poses challenges for downstream histological and immunohistochemical analyses, necessitating optimized processing, and staining protocols. Here, we describe robust workflows for the fixation, embedding, sectioning, and staining of ISC-derived organoids. Each method is detailed stepwise, with emphasis on preserving morphology and antigenicity. We discuss reagent choices, critical technical considerations, and troubleshooting tips to maximize data reproducibility and quality. We have integrated recent advances and best practices, serving as a comprehensive guide for researchers working with intestinal organoids.
Methods in molecular biology (Clifton, N.J.)Shivam Patel, Paloma Ordóñez-Morán
Human intestinal organoids (HIOs) recapitulate the architecture and cell diversity of intestinal epithelium, hence providing a system for modeling processes like regeneration and tumorigenesis. Here, we describe a detailed whole-mount immunostaining protocol for HIOs to visualize proliferative cell population. Briefly, this protocol includes preparation of HIOs, fixation and permeabilization of tissue, blocking of non-specific binding, antibody crosslinking, and visualization using a confocal microscope. Additionally, the protocol is broadly adaptable for investigating different antibodies, enabling the exploration of other cell signaling processes in HIO models. By eliminating embedding and sectioning steps, this method is both time-efficient and preserves spatial tissue architecture.
Veterinary medicine and scienceHossein Lak, Amir Amniattalab
BACKGROUND: Mammary tumours are the most common neoplasm in female dogs. Canine mammary tumours (CMTs) are appropriate models for human studies. OBJECTIVES: The present study aimed to diagnose CMTs pathologically and compare different tumour types by immunohistochemistry (IHC). METHODS: Histological features of various tumours were recorded. Moreover, haematological, biochemical and immunohistochemical changes were evaluated in affected animals. RESULTS: Among 130 mastectomized dogs in veterinary hospitals in Tehran, Iran, 74 dogs were affected by various types of mammary tumours. The highest frequency of malignant tumours was related to carcinoma types (38/74; 51.4%) and the highest frequency of benign tumours was related to fibroadenoma (6/74; 8.1%). In malignant tumours, anaemia and monocytosis were observed compared to benign tumours. Serum ALT and ALP levels were also significantly increased in malignant tumours (p < 0.05) compared to benign tumours. The IHC showed the profile of adenoma (ductal and intraductal papillary) ER+/PR-/HER2-, benign mixed tumour ER-/PR-/HER2-, carcinoma (mucinous, anaplastic) ER-/PR-/HER2- and metastatic mast cell tumour ER-/PR+/HER2-. The highest expression of αSMA was recorded in benign tumours and the highest expression of Ki67 was recorded in metastatic mast cell tumour, benign mixed tumour and adenoma (p < 0.05). CONCLUSION: Together, although the dog is a good model for studying mammary tumours, the variability of results in CMTs requires further studies in this field. An interesting finding of this study was the triple-negative profile in the benign mixed tumour, which had previously been reported only for malignant mammary tumours. In addition, the significant positivity of Ki67 expression (34%-66%) in benign CMTs is noted for the first time.
Epidermal growth factor receptor (EGFR), cellular mesenchymal-epithelial transition factor (c-MET), and B7 homolog 3 (B7H3) are important targets for antibody-based drug development in colorectal cancer (CRC). To overcome the limitations of single-target antibody therapies - such as limited efficacy, widespread resistance, narrow patient populations, and treatment-related toxicities - various bispecific antibodies or antibody-drug conjugate (ADCs) targeting two targets are being extensively validated in clinical settings. A deeper understanding of their expression profiles and co-expression patterns of actionable therapeutic targets may guide more effective treatment strategies. We evaluated the immunohistochemical expression of B7H3, c-MET, and EGFR in a cohort of 193 CRC patients. Among which, B7H3 exhibited the highest positivity rate (80.83%) in CRC tissues, followed by c-MET (77.20%) and EGFR (47.15%). The positive rates of B7H3 showed no significant differences across patient gender, age, TNM stage, differentiation grade, or tumor site. In contrast, the c-MET positivity rate was elevated in patients with stage I-II disease, and the expression of EGFR was significantly higher in female patients than in male patients. Concurrent assessment of the three targets in metastatic lesions revealed that B7H3 expression levels were comparable between primary and metastatic tumors, whereas c-MET expression was significantly higher in primary lesions. Co-expression analysis indicated that the group with the highest patient coverage was the B7H3/c-MET dual-positive group, followed by the B7H3/EGFR, and c-MET/EGFR combinations, and the proportion of patients with at least one positive target exceeded 80%. Finally, mRNA profile from TIMER database reveals that CD276 and MET expression in CRC tumor tissues were higher than those in the majority of normal tissues. Therefore, utilizing B7H3/c-MET combination therapies that integrate intracellular signaling inhibition with immune checkpoint modulation could potentially extend therapeutic benefits to a wider range of CRC patients.
BACKGROUND: Claudin 18.2 (CLDN18.2) is a promising therapeutic target in gastric/esophagogastric junction adenocarcinoma (GC/EGJC), but its dynamic expression during systemic therapy is unclear. METHODS: CLDN18.2 expression by immunohistochemistry in paired tumor samples from 155 patients with advanced GC/EGJC was retrospectively evaluated before and after systemic therapy. CLDN18.2-positive was defined as ≥75% of viable tumor cells exhibiting moderate-to-strong (2+ or 3+) membranous staining. RESULTS: Among all patients, 54 (34.84%) were CLDN18.2-positive. Following treatment, the proportion of CLDN18.2-positive patients increased significantly to 61 (39.35%) (p < .001). Among patients who were CLDN18.2-negative at baseline, 57 (56.44%) showed increased expression after treatment, including 22 (21.78%) who converted to positive status. In contrast, among those CLDN18.2-positive at baseline, expression decreased in 35 patients (64.81%), with 15 (27.78%) converting to negative status (p < .001). Among patients receiving chemotherapy alone (n = 25), 1 (4.00%) converted to negative and 5 (20.00%) converted to positive, whereas among those receiving chemotherapy combined with immunotherapy (n = 130), 14 (10.77%) converted to negative and 17 (13.08%) converted to positive (p = .45). A total of 131 patients (84.52%) demonstrated changes in CLDN18.2 expression following therapy, encompassing both conversions across the positivity threshold and fluctuations within the same classification. CONCLUSION: CLDN18.2 expression in GC/EGJC is dynamic and frequently altered after systemic therapy. These findings support reassessment of CLDN18.2 status before initiating CLDN18.2-targeted therapies.
INTRODUCTION: The response to corticosteroid therapy in idiopathic nephrotic syndrome (INS) is variable. Glucocorticoid receptors (GR), which are essential for steroid action, may partly explain this variability. OBJECTIVE: To investigate the relationship between podocyte glucocorticoid receptor expression assessed by immunohistochemistry and the response to corticosteroid therapy in INS. PATIENTS AND METHODS: This was a retrospective study conducted over a 22-year period, including 29 patients with minimal change disease or primary focal segmental glomerulosclerosis. Podocyte GR expression was evaluated by immunohistochemistry and correlated with treatment response. RESULTS: In total, 18 patients were steroid-sensitive, 6 were steroid-dependent, and 5 were steroid-resistant. No conventional clinical, laboratory, or histological factor was predictive. However, GR expression was higher in steroid-sensitive than in steroid-resistant patients (p = 0.046). CONCLUSION: Podocyte glucocorticoid receptor expression may predict the response to corticosteroid therapy in idiopathic nephrotic syndrome.
World journal of urologyJiayu Yang, Wentao Yu, Shengwang Zhang, Qian Chen
OBJECTIVES: This study aimed to classify upper tract urothelial carcinoma (UTUC) using GATA3, KRT5, and P16 immunohistochemistry and to evaluate associations of these molecular subtypes with clinicopathological features and outcomes. PATIENTS/MATERIALS AND METHODS: Tissue samples from 102 UTUC patients who underwent radical nephroureterectomy were analyzed. Tumors were classified into genomically unstable (GU), urothelial-like (URO), and basal subtypes based on GATA3, KRT5, and P16 expression. Associations with pathological features, recurrence, and survival were analyzed. Kaplan-Meier and Cox regression analyses were used. RESULTS: The GU, URO, and basal subtypes accounted for 33%, 45%, and 22% of cases, respectively. Subtype was significantly associated with infiltrative pattern (p = 0.009), perineural invasion (p = 0.016), histological variant (p < 0.001), T stage (p = 0.004), N stage (p = 0.017), and vital status (p = 0.005). The basal subtype showed the poorest prognosis, with 3-year overall survival of 41% vs. 85% for the URO subtype (p < 0.001). Multivariable analysis identified diffuse infiltrative pattern (HR 6.84, P = 0.004), lymphovascular invasion (HR 2.12, p = 0.046), and T4 stage (HR 9.80, p = 0.023) as independent risk factors for overall survival, and diffuse infiltrative pattern (HR 6.37, p = 0.006) and T4 stage (HR 12.6, p = 0.012) for recurrence-free survival. CONCLUSIONS: IHC-based subtyping using GATA3, KRT5, and P16 is significantly associated with aggressive clinicopathological features in UTUC, with the basal subtype showing poor prognosis in univariate analysis. However, it was not an independent prognostic factor in multivariable analysis. While this exploratory strategy may aid in risk stratification, its clinical utility requires further validation in larger, prospective cohorts.
Journal of visualized experiments : JoVELavanya Mendu, Chunda Feng
Reniform nematodes (Rotylenchulus reniformis) are a major constraint on cotton production, particularly in the southern United States. The reniform nematode is the second most important pest among more than 400 nematode species. These nematodes pose a threat to global food security. Increasing environmental and human health concerns and the cost of nematicides have intensified interest in resistant cotton germplasm. The plant cell wall, especially lignin, is a critical biopolymer involved in resistance to biotic stress. Because nematodes infect root tissues, cell wall components such as lignin may contribute to defense; however, lignin-mediated barriers in cotton roots are not well characterized. This study employed a standardized phloroglucinol-HCl histochemical staining protocol to visualize lignin deposition differences in reniform nematode-resistant (A2-514, A2-995) and susceptible cotton (Gossypium arboreum) genotypes at the true leaf stage under non-infected conditions. This stain binds specifically to hydroxy cinnamylaldehyde such as coniferyl aldehyde and sinapyl aldehyde of lignin in the presence of acid and generates a pinkish purple color to visualize lignin. Distinct lignin staining patterns were observed between resistant and susceptible genotypes, supporting a potential role for lignin-associated structural defenses.
PURPOSE: The purpose of this review paper is to provide an integrative algorithmic approach to the diagnosis of salivary gland tumors, including pertinent histologic features, immunohistochemical patterns and molecular driver mutations and/or fusions. METHODS: Review of the literature was performed to provide an up-to-date synopsis of salivary gland tumors, in a concise and summarized format. RESULTS: Diagnostic workflow algorithms are created and included to complement the histomorphologic subgroups, to provide a user-friendly tool when approaching salivary gland tumors in routine surgical pathology. CONCLUSION: Histomorphologic features and immunophenotype of majority of salivary gland tumors, including select molecular surrogate immunohistochemical stains, are the most important starting point in diagnostic interpretation. Molecular data can provide an additional data point for further supporting evidence and/or to aid in challenging cases.
Veterinary research communicationsMaíra Meira Nunes, Eric Santos Oliveira, Júlia Gabriela Wronski, Rayane de Souza Ruas Lopes, Marcella Letícia Melo Souza da Rocha, Karen Yumi Ribeiro Nakagaki,…
Secretory carcinoma of the breast is a rare histological subtype in women and is uncommon in female dogs and cats, with only two cases described in female dogs and one case described in a cat. This report describes the histopathological and immunohistochemical features of secretory carcinoma of the mammary gland in a domestic rabbit (Oryctolagus cuniculus). Microscopic evaluation revealed a non-encapsulated, well-demarcated neoplastic proliferation composed of epithelial cells arranged in solid and tubular patterns, supported by a delicate fibrous connective tissue stroma. The neoplastic cells had slightly eosinophilic cytoplasm, frequently containing large, well-demarcated vacuoles that displaced the nucleus to the periphery, giving the cells a signet-ring-like appearance. Periodic acid-Schiff (PAS) histochemical staining, performed with and without diastase digestion, showed positivity in the intracellular and extracellular secretory material in both approaches. Immunohistochemical evaluation was performed for the markers α-lactalbumin, Ki-67, estrogen receptor, progesterone receptor, and COX-2. Positive cytoplasmic immunolabeling for α-lactalbumin was observed in neoplastic cells and in intracellular and extracellular secretory material, confirming their secretory nature. The diagnosis of secretory carcinoma was established based on the association of histopathological, histochemical, and immunohistochemical features. Considering the secretory features of the mammary gland in rabbits, secretory carcinoma should be included in the differential diagnosis of mammary carcinomas with prominent secretory activity in this species.
MedicineDongjun Lee, Young Jun Choi, Joon Seon Song, Yoon Se Lee, Sae Rom Chung, Jung Hwan Baek, Jeong Hyun Lee
With the increasing prevalence of human papillomavirus (HPV)-driven oropharyngeal squamous cell carcinoma (OPSCC), there is a need for efficient, minimally invasive methods to evaluate p16 status. This study aimed to assess the concordance of p16 status between ultrasound (US)-guided core needle biopsy (CNB) of cervical lymph nodes and surgical specimens, and to describe the numbers of CNB and surgical sessions required to establish the histopathologic diagnosis. This retrospective study included 39 patients diagnosed with OPSCC or head and neck squamous cell carcinoma of unknown primary (SCCUP) at a single tertiary hospital between March 2019 and March 2024. All included patients underwent US-guided CNB of cervical lymph nodes and a surgical procedure (biopsy or resection), and had evaluable p16 immunohistochemistry (IHC) results on both specimen types. The concordance rate for p16 status between US-guided CNB and surgical specimens was 100.0% (39/39; 95% CI, 91.0-100.0%; 35 p16-positive and 4 p16-negative cases). Sensitivity was 100.0% (35/35; 95% CI, 90.0-100.0%) and specificity was 100.0% (4/4; 95% CI, 39.8-100.0%). In all 39 included patients, a diagnosis of metastatic squamous cell carcinoma and an evaluable p16 result were obtained at the first CNB session. Among the 35 patients with OPSCC, 10 (28.6%) required more than 1 surgical session at the primary tumor site. No immediate complications were observed following US-guided CNB. In this cohort, US-guided CNB of cervical lymph nodes showed complete observed concordance with surgical specimens for p16 status, and a diagnosis with an evaluable p16 result was obtained at the first CNB session in all patients. Prospective, multicenter validation including more p16-negative cases is required before wider adoption.
Histochemistry and cell biologyRachelle Honeywell, Maggie Donner, Sheila Criswell
Tissue under- and over-processing in the histology laboratory, while decreasing in frequency, is still a valid concern and typically results from user or instrument error. This study evaluated the effects of extended exposure of histological tissues to processing reagents on specimen quality, including embedding, microtomy, histochemical staining, and immunohistochemistry (IHC). All specimens were fixed in neutral buffered formalin for 3-5 weeks prior to processing except for a subset of organs which underwent prolonged fixation for 1 year. Although overprocessing is traditionally associated with tissue hardening, brittleness, and shrinkage, these effects are often noted when fixation timing is not clearly defined. To simulate processing delays, specimens were subjected to an additional 24 h in absolute ethanol, xylene, molten paraffin, or a heated embedding station. Extended exposure to these reagents did not result in observable morphologic changes. Embedding quality, microtomy performance, and histochemical staining were consistent across all experimental conditions. Additionally, IHC antigenicity was largely preserved under most conditions. However, reduced or absent immunoreactivity was observed in select markers in organs fixed for 1 year, consistent with prior reports of fixation-related antigen masking and deterioration of antigenic sites. Unexpectedly, extended exposure to ethanol, xylene, or paraffin did not independently compromise IHC performance in adequately fixed specimens.
NMR in biomedicineIrene Garcia-Bocanegra, Jesus David Urbano-Gamez, Antonio Perez-Cardona, Laura Barrios, Maria Isabel Somoza-Ramírez, Luis Vicioso, Elena Pardo-Susacasa, Gema D…
Advances in molecular characterization have significantly redefined our understanding of breast tumor heterogeneity, enabling improved diagnostics and the development of tailored therapeutic strategies. In particular, integrative omics approaches offer a broader scope for exploring the molecular and biochemical complexity of breast cancer. Notably, metabolomics through high-resolution magic angle spinning (HR-MAS) NMR spectroscopy enables nondestructive metabolic profiling of intact tissue samples, preserving the native biochemical context and offering unique potential for refining breast cancer subtyping and gaining deeper insight into disease biology. However, the reproducibility and comparability of HR-MAS-based metabolomics across studies are still limited by methodological variations, underscoring the need for standardized and reliable protocols to manage the complexity of breast cancer tissue spectra. In this study, LCModel analysis in combination with the Electronic REference To access In vivo Concentrations (ERETIC2) method as a quantitative reference was implemented for absolute metabolite quantification of breast cancer samples, providing a robust metabolic profiling approach. Generalized linear models were applied to evaluate the associations between quantified metabolites and tumor immunohistochemical classification, and transcriptomic correlation analysis was used to explore gene expression profiles underlying the observed metabolic alterations. A total of 27 metabolites were quantified from breast tumor samples spanning the Luminal A, Luminal B (human epidermal growth factor receptor 2-negative, HER2-), Luminal B (HER2+), and triple-negative breast cancer (TNBC) subtypes. Strong positive associations were observed between phosphocholine (PCho), betaine (Bet), total choline (tCho), and total creatine (tCr) and immunohistochemical subtype, whereas alanine, glucose, and myo-inositol showed negative associations. Notably, Luminal B (HER2-) tumors exhibited the highest PCho levels, linked to upregulated expression of choline kinase alpha (CHKA) and other enzymes of the Kennedy pathway. In contrast, Luminal B (HER2+) tumors exhibited elevated levels of alanine (Ala), glucose (Glc), and myo-inositol (Ins), indicating distinct metabolic reprogramming within the luminal subgroups. Additionally, this study highlights the potential role of betaine (Bet) in interconnecting metabolic and epigenetic pathways in breast cancer, as its concentrations were found to vary across subtypes and correlate with genes involved in fatty acid metabolism, DNA methylation, and estrogen signaling. Robust HR-MAS NMR profiling captured subtype-dependent metabolic signatures, and integrative analysis with transcriptomics identified gene expression differences associated with these signatures, thereby advancing the understanding of breast cancer heterogeneity.
Cancer medicinePartha Basu, Anuradha Talukdar, Poulome Mukherjee, Richard Muwonge, Eric Lucas, Monoj Kumar Deka, Amanjit Bal, Upasana Gautam, Manish Rohilla, Baneet Bansal, V…
BACKGROUND: Accurate detection of estrogen and progesterone receptors (ER/PR), HER2, and Ki-67 is essential for molecular classification of breast cancer and in deciding upon tailored treatment. In many low- and middle-income countries (LMICs), access to histopathology and immunohistochemistry (IHC) is limited. STRAT4, an automated RT-qPCR assay performed on the widely available GeneXpert platform, offers a potential alternative. We evaluated the performance of STRAT4 on core biopsy, unstained smear of fine-needle aspiration biopsy (FNAB-US), and cellblock samples compared with conventional IHC in 178 histopathology-confirmed breast cancer patients. METHODS: Matched core biopsy, cellblock, and FNAB-US samples were analyzed using STRAT4. Concordance with IHC for ER, PR, HER2, and Ki-67 was assessed using percentage agreement and kappa statistics. The effect of alternative dCt cut-offs used in an earlier study was also examined. Molecular subtype classification derived from STRAT4 was compared with IHC-based classification. RESULTS: Concordance between STRAT4 and IHC was high for ER across all sample types (κ: 0.78-0.86). PR detection showed substantial agreement for core biopsies (κ = 0.78) and cellblocks (κ = 0.63), but only fair for FNAB-US samples (κ = 0.53). HER2 detection showed moderate agreement (κ = 0.41-0.58), improving with modified cut-offs (κ = 0.57-0.66), especially for FNAB-US. Ki-67 concordance was low (agreement 58.9%-71.0%). Kappa values indicating agreement between molecular subtype classifications using STRAT4 on various samples and routine IHC were as follows: core biopsies = 0.60; cellblocks = 0.61; FNAB-US samples = 0.45. CONCLUSION: Detection of ER from FNAB-US, cellblocks, and core biopsies using STRAT4 showed high concordance with IHC. However, the concordance was low to moderate for other markers.
The Journal of comparative neurologyNatalia Kasica, Jerzy Kaleczyc, Waldemar Sienkiewicz, Ewa Lepiarczyk
The objective of the current investigation was to determine the effect of neonatal castration on the expression of androgen receptor (AR), estrogen receptor alpha (ERα), and estrogen receptor beta (ERβ) in the neurons of the anterior pelvic ganglion (APG) in male pigs. Receptor expression was quantitatively assessed using real-time polymerase chain reaction (qPCR) and immunohistochemistry. Ganglia were sampled from intact pigs at 7 days of age (the day of castration) and from both intact and castrated animals at 3 and 6 months postsurgery. qPCR analysis of the intact boars revealed a significantly elevated gene expression level (GEL) of AR at 6 months of age compared to 3 months and 7 days of age. Conversely, the GEL of ERα was comparable across all age groups in the intact animals. Gene expression analysis of ERβ in the pelvic ganglia of both intact and castrated pigs consistently indicated very low levels. In the 3-month-old castrated cohort, the GELs of AR and ERα were significantly elevated compared to those in age-matched intact pigs. Conversely, the GEL of ERβ did not exhibit a significant difference between the castrated and intact groups. Immunofluorescence investigations complemented the qPCR data, revealing that numerous APG neurons across all control and experimental groups demonstrated immunoreactivity for AR, ERα, and ERβ, confirming the translation and presence of all three receptor types within the neuronal somata. These data collectively suggest that, in the male pig, pelvic neurons constitute a component of autonomic circuits that are highly dependent on sex steroids and are implicated in the regulation of urogenital function. Furthermore, the sensitization of these neurons to sex hormones, likely involving not only androgens but also estrogens (primarily mediated via ERα), appears to be a dynamic, maturational process that occurs during the prepubertal period.
Investigative ophthalmology & visual scienceLeticia Ussem, Karin U Loeffler, Verena Tischler, Marieta Toma, Natalie Pelusi, Frank G Holz, Anne Gulbins, Daniela Süsskind, Roberta Noberini, Tiziana Bonaldi…
PURPOSE: Uveal melanoma (UM) is the most common primary intraocular malignancy in adults, with a unique genetic and epigenetic profile and a high metastatic risk. The lack of experimental models to replicate tumor biology, including its epigenetic landscape, limits progress in translational research. Recently, patient-derived organoid (PDO) models have emerged as a promising tool for the modeling of various tumors, including UM. This study aims to establish an air-liquid interface (ALI) PDO culture system for UM, evaluate its similarity to the primary tumors, and assess its utility for drug screening. METHODS: Fresh and cryopreserved tissue samples from primary UM of 5 patients were cultured using a protocol previously validated for other malignancies. The subsequent characterization of the ALI PDOs included histologic and immunohistochemical evaluation, DNA and RNA sequencing, and histone posttranslational modification (HPTM) analysis. The model's drug-screening potential was evaluated by treating ALI PDOs with the histone deacetylase inhibitor quisinostat and comparing their responses with those of controls. RESULTS: UM ALI PDOs were successfully established and retained their original tumors' immunophenotype and genetic variants. Gene expression profiling revealed similarities between PDOs and primary tumors, with main differences consisting of microenvironment-related signals. Epigenetic analysis confirmed that the overall HPTM landscape remained preserved. Treatment of the PDOs with quisinostat led to transcriptional reprogramming and the expected epigenetic response, with increased acetylation marks. CONCLUSIONS: The UM ALI PDO model recapitulates the genetic and epigenetic features of primary UM and holds promise as a platform for future molecular studies and the testing of personalized therapy strategies.
Rhode Island medical journal (2013)Tanin Zadeh, Komeil Mirzaei Baboli, Sara Salehiazar, Truc Tran
Malignant transformation of a mature cystic teratoma (MCT) is rare, with mucinous adenocarcinoma being an even more uncommon subtype. Accurate diagnosis requires distinction from metastatic gastrointestinal tumors. A woman in her 30s presented with a 19 cm right ovarian mass. She underwent exploratory laparotomy with right salpingo-oophorectomy, left ovarian cystectomy, omentectomy, peritoneal biopsies, and bilateral pelvic and para-aortic lymphadenectomy with intraoperative rupture of the mass. Frozen section suggested immature teratoma, but final pathology revealed mucinous adenocarcinoma arising within a mature teratoma. The tumor showed CK20, CDX2, SATB2 positivity, patchy CK7, and loss of PAX8. All lymph nodes and staging biopsies were negative for carcinoma. Final stage: FIGO IC1. This case highlights the diagnostic and staging challenges in malignant transformation (MT) of a MCT. Thorough sampling, immunohistochemistry (IHC), and clinical correlation are critical to differentiate primary ovarian neoplasms from metastatic disease. Accurate classification is essential to guide appropriate management.
Cancer reports (Hoboken, N.J.)Phat Thi Hong Ho, Duy Duc Nguyen, Vuong Dinh Thy Hao, Giang Huong Tran, Lam Viet Trung, Thinh Van Hoang, Hoang Bac Nguyen
BACKGROUND: Aberrant p53 expression may have prognostic significance in colorectal cancer; however, its role in patients with colorectal liver metastases (CRLM) who receive neoadjuvant chemotherapy but do not achieve pathological complete response (pCR) remains unclear. AIMS: We evaluated whether aberrant p53 expression assessed by immunohistochemistry (IHC) predicts recurrence-free survival (RFS) in patients with CRLM receiving neoadjuvant FOLFOXIRI followed by resection without achieving pCR. METHODS AND RESULTS: A prospective cohort study was conducted on 39 patients with CRLM who met the inclusion criteria. All patients received neoadjuvant chemotherapy with the FOLFOXIRI regimen. Following surgical resection of both primary colorectal tumors and liver metastases, none of the patients achieved pCR. Immunohistochemistry was performed to assess p53 expression in resected liver metastasis specimens using a pattern-based interpretation model. RFS was analyzed using univariate and multivariate Cox proportional hazards models. Aberrant p53 expression was observed in 20 patients (51.3%). In univariate analysis, aberrant p53 expression (HR = 2.20; 95% CI: 1.02-4.71; p = 0.043), total tumor size > 6 cm (HR = 5.62; 95% CI: 2.31-13.68; p < 0.001), number of liver metastases > 4 (HR = 4.41; 95% CI: 1.59-12.26; p = 0.004), and positive surgical margin (HR = 2.62; 95% CI: 1.16-5.91; p = 0.021) were significantly associated with recurrence. In multivariate analysis, aberrant p53 expression (HR = 3.78; 95% CI: 1.56-9.15; p = 0.003) and total tumor size > 6 cm (HR = 5.23; 95% CI: 1.56-17.55; p = 0.007) remained independent prognostic factors for shorter RFS.cnr270701_RedLinePDF. CONCLUSION: Aberrant p53 expression, as assessed by immunohistochemistry on post-neoadjuvant chemotherapy specimens, is an independent prognostic factor for early recurrence in patients with CRLM who did not achieve pCR. Together with total tumor size, aberrant p53 expression may serve as a useful marker for risk stratification and personalized postoperative treatment strategies in this high-risk patient population.
The journal of obstetrics and gynaecology researchFajrin Mammadova Bahitli, Levent Akman, Gurdeniz Serin, Mert Acar, Nuri Yildirim, Osman Zekioglu, Mustafa Cosan Terek, Erdem Goker, Ahmet Aydin Ozsaran
OBJECTIVE: To evaluate and compare programmed cell death-ligand 1 (PD-L1) expression across benign and malignant uterine mesenchymal tumors using immunohistochemical analysis. METHODS: This retrospective study included 100 patients who underwent hysterectomy for uterine mesenchymal tumors at a single tertiary center between 2000 and 2022. Cases included leiomyoma (LM; n = 20), leiomyosarcoma (LMS; n = 20), low-grade endometrial stromal sarcoma (LG-ESS; n = 20), high-grade endometrial stromal sarcoma (HG-ESS; n = 20), and endometrial stromal nodule (ESN; n = 20). Immunohistochemical staining was performed using the PD-L1 (22C3) antibody, and PD-L1 expression was evaluated using the combined positive score. Clinicopathological parameters and PD-L1 expression were compared among tumor subtypes. RESULTS: Patients with HG-ESS demonstrated a significantly higher mean age at diagnosis compared with the other histopathological subtypes. PD-L1 expression was detected in 9 (45%) LMS, 10 (50%) HG-ESS, 3 (15%) LG-ESS, and 1 (5%) ESN cases, whereas no LM cases demonstrated positivity. PD-L1 expression rates were significantly higher in LMS and HG-ESS than in LM, LG-ESS, and ESN cases (p < 0.0001). Combined group analyses also demonstrated significantly higher PD-L1 expression rates in the combined LMS/HG-ESS group compared with the LG-ESS group and the combined LM/ESN group (p < 0.0001). CONCLUSION: These findings demonstrate increased PD-L1 expression in aggressive uterine mesenchymal tumors; however, the clinical and therapeutic significance of these immunohistochemical findings remains unclear. Further studies integrating PD-L1 expression with clinical findings and treatment outcomes are needed to determine the potential therapeutic relevance of PD-L1 in these neoplasms.
Neuropathology and applied neurobiologyAlessia Andrea Ricci, Filippo Nozzoli, Alessandro Biale, Cristian Tampieri, Ludovica Verdun di Cantogno, Diego Garbossa, Mario Levis, Roberta Rudà, Camilla Bon…
AIMS: Homozygous deletion (HD) of CDKN2A/B represents an adverse prognostic biomarker in meningiomas and a diagnostic criterion for CNS WHO grade 3 assignment. However, fluorescence in situ hybridisation (FISH) thresholds and the role of immunohistochemistry (IHC) surrogates remain uncertain. This study evaluated multiple CDKN2A/B FISH cutoffs and assessed the diagnostic and prognostic performance of MTAP and p16 IHC. METHODS AND RESULTS: Ninety-two meningiomas (CNS WHO grades 1-3) were analysed by FISH evaluating ≥ 10%, ≥ 20% and ≥ 30% CDKN2A/B HD thresholds. CDKN2A/B HD was identified in 18.5%, 6.5% and 4.3% of cases using the respective cutoffs and was significantly associated with shorter disease-specific survival across all thresholds (HD ≥ 10% p = 0.001; HD ≥ 20% p = 0.0001; HD ≥ 30% p = 0.043). Patients with HD ≥ 20% and ≥ 30% experienced universal disease-specific mortality within a median survival time of approximately 2 years. MTAP and p16 IHC were performed in matched areas. MTAP IHC showed high specificity (90%-92%) but limited sensitivity (30%-50%), while p16 IHC displayed variable sensitivity (41%-75%) and lower specificity (63%). Combined MTAP/p16 negativity improved sensitivity (up to 100%) but reduced specificity (57%). Neither MTAP nor p16 loss correlated significantly with survival. Spatially heterogeneous IHC patterns corresponded to regional variability in CDKN2A/B deletion by FISH. CONCLUSIONS: CDKN2A/B HD detected by FISH, particularly using thresholds equal or greater than 20%, is strongly associated with poor outcome in meningiomas. MTAP and p16 IHC, alone or combined, lack sufficient accuracy as independent surrogates but may guide tissue selection for molecular testing in heterogeneous tumours.
BACKGROUND/AIM: Microvessel density (MVD) is an inconsistent prognostic factor in colorectal cancer (CRC). We evaluated the vascular proliferation index (VPI), a qualitative marker of proliferating endothelial cells, as an alternative biomarker. This study aimed to comprehensively compare the prognostic performance of MVD and VPI in Stage II/III CRC patients. PATIENTS AND METHODS: MVD and VPI were examined using dual immunohistochemistry (IHC) for CD31/Ki-67 in 85 patients with Stage II/III CRC. Single-cell flow cytometry (FCM) was performed in 72 patients whose samples were suitable for single-cell evaluation among the 85 patients. RESULTS: High VPI (>16.5%) via IHC correlated with a significantly shorter disease-free survival (DFS) (p=0.020), whereas MVD showed no prognostic correlation. Consistently, a high Ki-67/CD31 ratio via single-cell FCM predicted a poorer DFS (p=0.002). In the multivariate analysis, high VPI was an independent poor prognostic factor for the DFS (p=0.011). Neither marker was correlated with overall survival. CONCLUSION: VPI via single-cell analysis is a superior, independent prognostic biomarker to MVD for the DFS in Stage II/III CRC, offering an objective platform for postoperative recurrence risk stratification.
PLoS neglected tropical diseasesFadwa M Arafa, Mohamed Hagar, Ahmed Zakaria, Eman Sheta, Nehal N Hezema
Given the grave risk posed by intestinal cryptosporidiosis to vulnerable populations, the current research focused on a new therapeutic avenue. Olibanum (OL) extract and probiotic Bacillus clausii (B. clausii) and their combination were evaluated against Cryptosporidium in vivo in immunosuppressed mice via parasitological, ultrastructural, biochemical, and histopathological analysis. Liquid chromatography-mass spectrometry (LC-MS) analysis of OL revealed the predominance of incensole isomers in its chemical composition. Regarding oocyst count in stool, the highest percentage of oocyst reduction was achieved after combined therapy (98.4%) on the 19th day post-infection, followed by B. clausii-treated group (87.6%), then the OL-treated group (87%). Similar ultrastructural alterations were noticed by scanning electron microscopy (SEM) following treatment by both OL and B. clausii, ranging from shrunken oocysts to severely deformed ones with extensive pits or protrusions. Nonetheless, their combination resulted in complete distortion of the oocysts, leading to their rupture. Biochemically, both OL and B. clausii treatment, either alone or in combination, provoked a decrease in the serum MDA levels, together with a corresponding rise in the mean levels of GSH. Concerning the histopathological findings, the best results were achieved following the combined OL and B. clausii treatment, which has restored villus architecture and remarkably ameliorated detrimental intestinal inflammation and goblet cell count. Finally, immunohistochemical analysis of the lymphocytic population of the intestinal villi revealed a statistically significant rise in the CD4+ /CD8+ ratio after treatment with either OL, B. clausii or their combination, which signals their proficient use for this therapeutic endeavour.
Breast cancer research and treatmentPayu Raval, Qingqing Ding, Puneet Singh, Alaaeddin Alrohaibani, Hongxia Sun, Di Ai, Kerollos Nashat Wanis, Ming Zhao, Hui Chen, Vicente Valero, Mariana Chavez-…
PURPOSE: Assessment of HER2 status change in HER2-positive breast carcinoma (BC) prior to and after HER2-targeted neoadjuvant therapy (NAT) may potentially impact adjuvant HER2-targeted therapy. Reported rates of HER2 status change vary widely (< 10-56%), underscoring the need for further study. This study evaluated the frequency of HER2 status change and its clinicopathologic correlations in HER2-positive BC cases. DESIGN: Residual breast carcinomas were retrospectively assessed at a single institution after HER2-targeted NAT. HER2 status on residual tumor was evaluated by immunohistochemistry (IHC) and, for IHC 2 + cases, confirmed by fluorescence in situ hybridization (FISH). Tumors were categorized as HER2-retained (IHC 3 + or 2+/FISH amplified) or HER2-change (IHC 0/1 + or 2+/FISH non-amplified). Clinicopathologic features were correlated with post-treatment HER2 status. RESULTS: Among 161 HER2-positive BC patients treated with HER2-targeted NAT between 2016 and 2025, 65 had adequate residual invasive tumor for HER2 assessment. Invasive ductal carcinoma was the predominant histologic type. Retained HER2 overexpression was observed in 28/65 (43%) cases, while 37/65 (57%) showed HER2 status change. Pre-NAT hormone receptor positivity was present in 23 (82.1%) HER2-retained and 33 (89.2%) HER2- status changed cases. Clinicopathologic parameters were comparable between the groups. CONCLUSION: HER2 status change from HER2 overexpression to loss of overexpression occurred in more than half of residual breast carcinomas following HER2-targeted NAT. The current national and international guidelines recommend adjuvant HER2-targeted therapy regardless of post-treatment HER2. Prospective randomized phase II/III trials are needed to determine the benefit of adjuvant HER2-targeted therapy in this subgroup of patients with HER2-positive BC.
Diagnostic pathologyAlaa Saad Abd Elhamid Mohamed, Heba Ashraf Hosni, Maram Salaheldeen Mahmoud, Alzahraa Abdelmoktader Mohammed
BACKGROUND: Endometrial carcinoma (EC) is a leading gynecological malignancy predominantly driven by hormonal imbalances. While the roles of estrogen and progesterone are well established, the influence of androgenic signaling, specifically through the androgen receptor, remains an underexplored diagnostic dimension. This study aims to evaluate the immunohistochemical (IHC) expression of the androgen receptor (AR) in EC tissue samples and correlate its presence with key clinicopathological parameters. METHODS: A retrospective cohort study was conducted utilizing 40 formalin-fixed, paraffin-embedded (FFPE) tissue specimens from patients diagnosed with EC who had undergone total hysterectomies. Cases were histologically categorized according to World Health Organization (WHO) criteria into endometrioid endometrial carcinoma (EEC), n = 32 and serous endometrial carcinoma, n = 8, and classified by the International Federation of Gynecology and Obstetrics (FIGO) for staging and grading. IHC staining was performed to evaluate nuclear AR expression, defined as positive when > 10% of neoplastic cells exhibited distinct nuclear staining. Data were statistically analyzed to identify associations between AR status and patient age, histological subtype, tumor grade, and stage. RESULTS: Distinct nuclear AR positivity was observed in 50% (20 of 40) of the total cohort. Expression was exclusively restricted to EEC (62.5% positive), whereas all serous carcinomas lacked expression entirely (P = 0.002). Notably, positive receptor expression demonstrated a strong inverse relationship with advanced FIGO stage and high tumor grade. Immunoreactivity was significantly enriched in early-stage disease (62.1% in Stages I and II versus 18.2% in Stage III; P = 0.013) and low-grade tumors (76.0% in Grades 1 and 2 versus 6.7% in Grade 3; P < 0.001). Patient age at diagnosis yielded no statistically significant correlation with expression status (P = 0.206). CONCLUSION: The AR is frequently expressed in low-grade, early-stage EEC, suggesting that AR may be associated with well-differentiated tumors and favorable clinicopathological parameters. The integration of AR immunohistochemistry into routine pathology panels could serve as a hypothesis-generating foundation for future clinical trials investigating targeted antiandrogen therapies.
The Fc receptor-like 4 (FcRL4)/immunoglobulin superfamily receptor translocation-associated 1 (IRTA1) immunohistochemical marker has emerged in recent literature as a highly specific-albeit moderately sensitive-marker for marginal zone lymphoma with near consistent negativity reported in mantle cell lymphoma (MCL) cohorts. We report a MCL patient with classic clinical, morphologic, immunophenotypic and genetic findings, but having aberrant FcRL4/IRTA1 co-expression. The aberrant expression and its possible favourable prognostic impact in MCL are discussed.
PURPOSE: Claudin 18 (CLDN18) is a protein that functions to maintain the integrity of tight junctions in epithelial cells. Recent development of anti-Claudin 18.2 targeted therapies have yielded positive survival outcomes for patients with Claudin 18 expression in gastric carcinomas. This study aims to investigate whether expression of Claudin 18 is present in a variety of benign and malignant salivary gland neoplasms. METHODS: Forty-eight cases of salivary gland neoplasms of various subtypes were included for this pilot study. Ancillary data including tumor location, grade, and stage of tumor as applicable was collected. Claudin 18 immunohistochemistry was performed on a representative tumor block of each case. RESULTS: No aberrant Claudin 18 expression was seen in the benign as well as malignant salivary gland neoplasms in this pilot study. CONCLUSIONS: CLDN18 expression was uniformly absent across the tumor types, locations, grades, and stages represented in this cohort. The absence of expression in all cases indicates that Claudin 18 has limited potential as a therapeutic marker in patients with salivary gland neoplasms. This study contributes to the extremely limited literature regarding Claudin 18 expression within salivary gland neoplasms.
ImmunoHorizonsLea Maristela, Alyssa Longworth, David F Ackart, Faye Lanni, Joseph Gibb, Ashley Freedman, Olivia R Miller, Ling Cai, Hong Qi, Susan L Baldwin, Sasha E Larsen,…
The guinea pig model of Mycobacterium tuberculosis (Mtb) infection offers distinct advantages compared to conventional mouse models. Namely, the guinea pig model develops circumscribed granulomas with central necrosis which most closely resembles that of human granulomas. However, a limitation of this model is the lack of available reagents to characterize the Mtb immune response. To better leverage this representative model, a set of recombinant rabbit monoclonal antibodies targeting guinea pig cytokines were generated and validated. Using established stimulation conditions for primary guinea pig cells capable of producing specific cytokine profiles, the reactivity and specificity of these antibodies to their biological target were evaluated. These antibodies were then evaluated individually for direct ELISA and immunohistochemistry or part of a pair for sandwich ELISA and ELISpot techniques. Utilized in a low-dose aerosol model of Mtb in the guinea pig, the utility of these antibodies to detect specific immune cytokines generated in response to infection is demonstrated. Collectively, these antibodies will provide a more comprehensive understanding of immune kinetics, vaccine response, and comparative immunology across the guinea pig model of tuberculosis and other infectious diseases.
Head and neck pathologySara C Degerholm, Tuula A Salo, Susanna Juteau, Matti O Mauramo
BACKGROUND: Mucosal melanoma is a rare disease with a poor prognosis, and its clinical signs and symptoms are nonspecific. This study aims to identify histopathological features and immunohistochemical markers of head and neck mucosal melanoma that characterize the tumor immunophenotype and are associated with patient survival. METHODS: The study analysed head and neck mucosal melanoma diagnosed at Helsinki University hospital between January 1, 2011, and June 8, 2023. Samples were obtained from the Helsinki Biobank in Finland. New slides were prepared and stained for hematoxylin and eosin (H&E), CD3, CD4, CD8, Granzyme B (GrzB), CD68, CD117, p16INK4a, p53, SOX10 and PRAME. RESULTS: The study included 23 patients, 10 men and 13 women, aged 60-92 years (mean 75 years). The tumors were predominantly sinonasal. Ulceration was present in 15 cases, also 15 cases were amelanotic. Immunohistochemical staining SOX10 was positive in all tumors. PRAME was strongly positive in majority (20 cases, 87%). Strong p16 positivity was observed in 13 cases (57%), with a longer median survival compared to cases with weakened p16 expression (32.8 vs 13.8 months, p = 0.176). The median survival was also longer in the cases with high score of tumor infiltrating CD3-, CD8- and CD68-positive immune cells, though not statistically significant. CONCLUSIONS: Immunohistochemical markers routinely applied in cutaneous melanoma (SOX-10 and PRAME) are also expressed in head and neck mucosal melanoma. Tumour-infiltrating immune cells showed trend toward better survival, as has been previously found out in cutaneous melanoma. Strong p16 positivity may be an independent predictor of favorable prognosis in head and neck mucosal melanoma, irrespective of disease stage. Larger multicenter studies are warranted to validate these findings in this rare disease.
PURPOSE: To investigate the protective effects of green tea extract on ovarian ischemia-reperfusion injury in a rat model using histopathological and immunohistochemical analyses. METHODS: Twenty-one female Wistar-Albino rats were randomly divided into three groups (n = 7 each): sham, saline-treated ischemia-reperfusion, and green tea extract-treated ischemia-reperfusion. Ovarian torsion was induced by 720° clockwise rotation of the adnexa for three hours, followed by three hours of reperfusion. Green tea extract (200 mg/kg) or saline was administered intraperitoneally immediately before reperfusion. Ovarian tissues were evaluated histopathologically for congestion, hemorrhage, edema, polymorphonuclear leukocyte infiltration, and follicular degeneration. Bax and caspase-3 expression levels were assessed immunohistochemically. RESULTS: The saline group demonstrated significantly increased congestion, edema, follicular degeneration, and total injury scores compared with the sham group. Green tea extract administration significantly reduced congestion, edema, and follicular degeneration compared with the saline group, indicating partial histopathological protection against ovarian ischemia-reperfusion injury. However, Bax and caspase immunoreactivity remained elevated in the treatment group. CONCLUSION: Green tea extract attenuated histopathological ovarian damage following ischemia-reperfusion injury, suggesting a protective effect against ovarian tissue injury. Although treatment was associated with increased expression of apoptotic markers, the overall findings support the potential of green tea-derived antioxidants as a therapeutic strategy for ovarian ischemia-reperfusion injury.
International journal of molecular sciencesEbru Karci, Sabin Goktas Aydin, Osman Erinc, Taskin Erkin Uresin, Sevinc Dagistanli, Nilay Bakoglu Malinowski, Ahmet Aydın, Onur Tanrikulu
SOX9 is a transcription factor linked to cellular plasticity and aggressive behavior in breast cancer. We investigated whether nuclear SOX9 expression correlates with short-term (5-year) survival outcomes in luminal breast cancer. In a retrospective cohort of 60 patients with estrogen receptor-positive invasive breast carcinoma, nuclear SOX9 expression was evaluated via immunohistochemical H-scores (0-300). Survival was analyzed continuously, by median-split dichotomy, and using a 1% nuclear-staining threshold via Kaplan-Meier and Cox regression models. Most tumors were luminal B (73.3%), and 45% were node-positive. Nuclear SOX9 was detected in 51.7% of cases (median positive H-score: 30). Over follow-up, 13 disease-free survival (DFS) and 12 overall survival (OS) events occurred. Continuous H-score did not correlate with clinicopathological variables, DFS (p = 0.576), or OS (p = 0.613). A median-split showed higher 5-year DFS for high-expression tumors (89.7% vs. 74.2%, p = 0.048), but this borderline protective trend was unconfirmed by Cox regression (p = 0.063) or the 1% threshold; ROC analysis showed no discrimination (AUC: 0.34-0.36). Standalone nuclear SOX9 expression has no significant impact on short-term (5-year) prognosis in early-stage luminal breast cancer; extended follow-up is warranted to assess potential late recurrence.
Breast cancer is the most common cancer and the second leading cause of cancer-related death among women worldwide. For accurate diagnosis, pathologists use immunohistochemical (IHC) staining for biomarkers such as human epidermal growth factor receptor 2 (HER2) as an auxiliary test, in addition to hematoxylin and eosin (H&E) staining for evaluating tissue morphology. Traditional IHC staining is limited by high cost, time, and labor, with a shortage of pathologists to meet demand. In this work, we propose a digital HER2 IHC staining algorithm based on tumor masks. The proposed model builds on the Dual Contrastive Learning Generative Adversarial Network (DCLGAN), adding tumor mask loss, structural similarity index measure (SSIM) loss, and a modified identity loss that differentiates between active and inactive regions. Performance was quantitatively assessed by comparing HER2 scores using the Fréchet Inception Distance (FID) and a grade classification model. Compared to DCLGAN, the proposed model achieved a 27.2% decrease in FID and a 2.57% increase in soft accuracy. Visual evaluation further showed that it prevents yellow discoloration and suppresses nonspecific region staining seen in unsupervised learning. These results indicate that effective learning is achievable even with limited and unaligned data, thereby accelerating digital pathology workflows.
Parkinson's disease (PD) is associated with systemic inflammatory and metabolic alterations that may extend beyond the central nervous system. Although 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) reproduces several pathological features of Parkinsonian neurodegeneration, its effects on pancreatic tissue remain poorly characterized. This study investigated MPTP-associated pancreatic histopathological and immunohistochemical alterations and evaluated the potential attenuating effects of Hexarelin. Fifty male BALB/c mice were randomly assigned to five groups (n = 10/group): Sham, MPTP, Hexarelin, PreHexarelin, and PostHexarelin. Pancreatic tissues were examined using hematoxylin and eosin staining and immunohistochemistry for amylin, β-amyloid, caspase-3, glucagon, insulin, CD11b, CD68, interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α). MPTP administration was associated with marked pancreatic injury characterized by acinar degeneration, cytoplasmic vacuolization, vascular congestion, interstitial edema, inflammatory cell infiltration, and degeneration of the islets of Langerhans. Immunohistochemical analysis demonstrated increased inflammatory (CD11b, CD68, IL-1β, and TNF-α) and apoptosis-associated (caspase-3) marker immunoreactivity, together with increased β-amyloid immunoreactivity and decreased insulin and amylin immunoreactivities. Hexarelin treatment attenuated these histopathological and immunohistochemical alterations in both treatment groups, although the magnitude of improvement varied among individual markers. These findings suggest that MPTP exposure is associated with pancreatic inflammation, apoptosis-associated changes, and alterations in endocrine-marker immunoreactivity. Hexarelin treatment was associated with attenuation of these pathological changes and partial preservation of islet morphology and endocrine-marker immunoreactivity; however, the PostHexarelin findings should be interpreted cautiously because the absence of a sequence- and time-matched post-MPTP vehicle control prevents complete separation of Hexarelin-associated effects from temporal changes after MPTP administration.
Journal of molecular histologySeyda Belli, İsmail Safa Poyrazoğlu, Eren Altun, Nida Sünnetçi Arkan
Ionizing radiation is widely used in the treatment of head and neck malignancies. However, its subacute effects on adjacent endocrine tissues have not yet been fully elucidated. This study aimed to investigate the structural changes that develop in the thyroid and parathyroid glands following irradiation and to evaluate HSP-90 immunoexpression in these tissues. The researchers initially randomized eighteen male Wistar-Hannover rats into control and irradiation groups (n = 9 per group). Rats in the irradiation group underwent 6-MV photon irradiation of the head and neck target region using a nominal prescribed single-fraction dose of 18 Gy with a Varian Clinac iX linear accelerator. One of the irradiated animals died on day 8; consequently, the day-21 analyses included nine control and eight irradiated animals. On day-21, the researchers performed histopathological examination and immunohistochemical evaluation of HSP-90 immunoexpression in thyroid and parathyroid tissues. Irradiation significantly increased vascular congestion (p = 0.031), inflammatory cell infiltration (p = 0.040), and epithelial desquamation (p = 0.006) in the thyroid tissue compared to the control group. It also significantly reduced HSP-90 immunoexpression in the thyroid follicular cells (p = 0.011). In contrast, irradiation did not cause marked disruption in the tissue architecture of the parathyroid glands. HSP-90 immunoexpression also showed no significant difference between the control and irradiation groups (p = 0.914). A single dose of high-energy photon irradiation induced subacute microstructural damage in the thyroid gland and reduced HSP-90 immunoexpression. The parathyroid gland, however, largely preserved its overall morphology and HSP-90 immunoexpression at day-21. These results indicate that thyroid and parathyroid tissues exhibit organ-specific subacute responses to irradiation.
BMJ case reportsYuan-Yin Zheng, Ying-Yu Mao, Mao-Hua Lin, Xiao-Bin Liu
NTRK (neurotrophic receptor tyrosine kinase)-rearranged spindle cell tumours are rare sarcomas predominantly occurring in the extremities and trunk, with only rare gastrointestinal tract cases reported to date. A man in his late 30s presented with abdominal pain. Given the CT evidence of a large transmural tumour requiring radical resection irrespective of histological subtype, the multidisciplinary team proceeded directly to surgery without preoperative colonoscopy. Diagnosis was established through histopathology, immunohistochemistry (IHC) and next-generation sequencing (NGS). Right hemicolectomy revealed an 8 cm transmural spindle cell neoplasm with necrosis, vascular invasion and perineural infiltration. IHC showed diffuse strong CD31 positivity but negativity for CD117, DOG-1, S100 and smooth muscle markers. NGS identified an ETS Translocation Variant 6-Neurotrophic Tyrosine Receptor Kinase 3 gene (ETV6-NTRK3) fusion. Applying the American Joint Committee on Cancer (AJCC) 8th edition criteria for soft tissue sarcoma by analogy, the tumour was staged pT2N0M0 (stage IIIA, high grade). Adjuvant chemotherapy was discussed after multidisciplinary review of the complete (R0) resection but was declined by the patient for financial reasons; close surveillance was adopted. The patient remained recurrence-free at the 8-month follow-up. This case adds to the limited number of reported gastrointestinal NTRK-rearranged sarcomas and expands the anatomic and molecular spectrum of these tumours. CD31 expression was an unexpected observational finding in this case and requires independent validation before it can be considered a characteristic immunohistochemical feature. Molecular confirmation is essential for accurate diagnosis and for identifying patients who may be eligible for tropomyosin receptor kinase (TRK) inhibitor therapy.
Veterinary research communicationsJoanna Klećkowska-Nawrot, Aleksandra Kroczak-Zdańkowska, Adam Urantówka, Grzegorz Zaniewicz, Shaw Badenhorst, Aleksander Chrószcz, Krzysztof Stegmann, Karolina…
The Harderian and lacrimal glands are the main orbital glands of the avian lacrimal apparatus and contribute to the tear film, nictitating membrane function, and local protection of the ocular surface. Although the ocular adnexa have been well described in domestic birds, the morphology and function of these organs remain poorly characterized in free-living species. This study aimed to characterize the morphology and histochemistry of these glands in the Great Spotted Woodpecker. The material consisted of 18 individuals found dead in the natural environment in Poland. Gross anatomical, morphometric, histological, and histochemical analyses were performed. Both glands occupied typical orbital positions. The secretory units of the Harderian gland consisted of one predominant epithelial cell type, contrasting with the more heterogeneous epithelial organization reported in some other avian species. Both glands were compound tubuloacinar structures. The Harderian gland opened into the lower conjunctival sac, whereas the smaller lacrimal gland drained into the dorsal conjunctival sac. The Harderian gland had a more homogeneous secretory parenchyma and a well-developed excretory duct system. The overall morphology of both glands was comparable to that described in other birds. However, the histochemical secretory profile differed from that reported in several species: the Harderian gland produced mainly acidic mucosubstances, whereas the lacrimal gland showed a mixed secretory profile. The immune component of both glands had a diffuse and periductal distribution, which contrasted with the pattern described in some birds. These findings show that the Harderian and lacrimal glands are structurally and histochemically distinct and suggest complementary roles in ocular surface maintenance in this species. At the same time, the results indicate selected morphological and functional differences compared with both domestic and other free-living birds.
Oral and maxillofacial surgeryCornelius Busse, Marius Hörner, Babak Saravi, Lukas Haug, Sebastian Gubik, Alexander Kübler, Urs Müller-Richter, Andreas Rosenwald, Stefan Hartmann
BACKGROUND: PD-L1 expression is an established predictive biomarker for immune checkpoint inhibitor therapy in recurrent or metastatic head and neck squamous cell carcinoma (HNSCC), including oral squamous cell carcinoma (OSCC). However, the spatial heterogeneity of PD-L1 and additional biomarkers such as c-MET between primary tumors and lymph node metastases remains insufficiently characterized. Given the potential for compartment-specific biomarker differences between primary tumors and metastatic lesions, we investigated PD-L1 and c-MET expression patterns in matched primary tumors and lymph node metastases of OSCC patients. METHODS: In this retrospective cohort study, PD-L1 expression (Tumor Proportion Score [TPS], Combined Positive Score [CPS]) and c-MET expression (H-score, percentage-based scoring) were analyzed by immunohistochemistry in primary tumors from 59 OSCC patients. Matched primary tumor-lymph node metastasis pairs were available for 24 patients and served as the basis for the paired spatial heterogeneity analyses. Biomarker concordance between primary tumors and metastases as well as associations with clinicopathologic parameters were evaluated. RESULTS: Both PD-L1 and c-MET were frequently expressed in primary tumors but showed significantly lower expression levels in matched lymph node metastases. Higher PD-L1 expression in primary tumors was associated with nodal metastasis in exploratory analyses, whereas c-MET expression showed no significant association with nodal status. No significant correlation between PD-L1 and c-MET expression was observed. Furthermore, discordance of PD-L1 CPS status between primary tumors and lymph node metastases resulted in different biomarker classifications in a substantial subset of matched cases. In exploratory analyses, biomarker expression and primary tumor-lymph node discordance were not significantly associated with recurrence, although the limited number of events precludes definitive conclusions. CONCLUSIONS: Our findings demonstrate pronounced spatial heterogeneity of PD-L1 and c-MET expression in OSCC and reveal compartment-specific biomarker expression patterns between primary tumors and lymph node metastases. The observed discordance between tumor compartments warrants further investigation in larger prospective studies to determine its biological and potential clinical relevance for biomarker assessment in OSCC.
PURPOSE: In this study, we aimed to investigate the immunohistochemical expression of apelin in tissue samples obtained from patients with Cushing's syndrome (CS) due to cortisol-producing adrenal adenoma (CPA) and to compare these findings with normal adrenal cortex tissue. Additionally, we sought to evaluate the association of apelin expression with clinical, hormonal, and histopathological parameters. The expression profile of vascular endothelial growth factor (VEGF) was also assessed to explore potential angiogenic interactions. METHODS: This retrospective, single-center study included 11 patients diagnosed with CPA and 15 control subjects with histologically normal adrenal cortex tissue. Formalin-fixed, paraffin-embedded tissue sections were subjected to immunohistochemical staining for apelin and VEGF. Immunohistochemical evaluation was performed by assessing staining intensity and the percentage of positively stained cells. An immunoreactivity score (IRS) was calculated by combining these parameters to provide a semi-quantitative measure of protein expression. RESULTS: Apelin staining intensity was significantly higher in CPA tissues compared with controls (p = 0.026), whereas the percentage of apelin-positive cells and IRS values did not differ significantly between groups. In an exploratory ROC analysis, apelin staining intensity showed moderate discrimination between CPA and control adrenal cortex tissues (AUC = 0.776, 95% CI: 0.581-0.971, p = 0.018). In bias-reduced univariable logistic regression, apelin staining intensity was associated with CPA (OR = 2.96, 95% CI: 1.02-8.60, p = 0.046), although the wide confidence interval indicated considerable uncertainty. VEGF staining parameters did not differ significantly between CPA and control tissues. No statistically significant correlations could be demonstrated between apelin or VEGF IRS and the evaluated clinicopathological or hormonal parameters. CONCLUSION: Apelin staining intensity was higher in CPA than in normal adrenal cortex, whereas the proportion of apelin-positive cells and IRS did not differ significantly between groups. VEGF staining parameters also did not differ significantly between CPA and control tissues. These findings suggest altered apelin immunoreactivity in CPA; however, given the exploratory nature and small sample size of the study, larger studies are required to confirm these observations and clarify their biological significance. CLINICAL TRIAL NUMBER: Not applicable.
International journal of molecular sciencesClaudia Giampietri, Francesca Somma, Elisa Pizzichini, Daniela D'Arcangelo, Siavash Rahimi, Cinzia Fabrizi, Antonio Facchiano
Considerable interest has recently grown toward the role Schwann cells (SCs) play in controlling the growth of several tumors. TCGA human melanoma database was interrogated focusing on 30 genes preferentially expressed in SCs and not present in melanoma (here referred to as SC-signature). The KM-plotter platform was exploited to address the relation of SC-signature to survival in melanoma patients and in other tumors as specificity controls. In addition, PCA in the GEPIA2 portal was used to address whether the SC-signature discriminates melanomas vs. healthy controls. Finally, the expression of one of the components of the SC-signature (Growth-Associated Protein 43, GAP43) was analyzed by immunohistochemistry. High levels of SC-signature expression were found to correlate inversely with survival in melanoma (423 patients) with strong statistical significance (HR = 2.12, p = 0.000003). A permutation test and validation on an independent melanoma database supported the relevance of these findings. PCA of the SC-signature revealed its capacity to discriminate melanomas from healthy controls. Consistently, immunohistochemical analysis revealed high expression of GAP43 in melanomas compared with control nevi. We have identified a group of 30 genes (SC-signature) whose expression negatively correlates with survival in melanoma patients, representing a promising prognostic biomarker candidate.
Ampullary somatostatin-producing D-cell neuroendocrine tumors are rare neoplasms that may be associated with type 1 neurofibromatosis. The molecular features of sporadic ampullary somatostatin-producing D-cell neuroendocrine tumors (SAMSOM-NETs) remain poorly characterized. We performed an integrated morphological, immunohistochemical, and genomic analysis of a multicenter series of SAMSOM-NETs. Eleven cases were included (73% male; median age: 63 years). All six patients who underwent lymphadenectomy were staged as pN1, and liver metastases were found in three cases; however, no tumor-related deaths occurred (median follow-up: 104 months). Common histologic features that can pose diagnostic challenges in the differential diagnosis with adenocarcinoma included a tubulo-glandular architecture (100%), periodic Acid-Schiff (PAS)-positive intraluminal mucin (73%), MUC1 expression (100%), and carcinoembryonic antigen (45%) expression. All tumors exhibited dot-like cytoplasmic reactivity for cytokeratins (CK) CAM5.2 or CK AE1/AE3, and 82% were CK7-positive. ISL1 and PDX1 were diffusely expressed in all cases, while CDX2 was positive in 54% and ARX showed only focal expression in four tumors. Genomic profiling revealed microsatellite stability and low tumor mutational burden. Alterations in the RAS pathway, including HRAS mutations (4 cases, 36%), a KRAS mutation (1 case), and NF1 alterations (one case), were identified in 54% of cases and in all tumors with liver metastases. Additional molecular findings included a CDK12 splice-site alteration and an NTRK3::PRDM4 fusion. Potentially actionable alterations affecting kinase-related pathways were detected in 64% of tumors. Our findings support SAMSOM-NET as a peculiar neuroendocrine tumor subtype showing distinctive histologic and molecular characteristics with potential diagnostic and therapeutic implications.