Cytokeratin Immunostaining Research Articles

Reproducibility of Assessment of Lepidic (Noninvasive) Patterns in Lung Adenocarcinoma With Cytokeratin Immunostain Compared With Hematoxylin and Eosin and the Proposed New International Association for the Study of Lung Cancer (IASLC) Algorithm

IntroductionLepidic growth is considered noninvasive in lung nonmucinous adenocarcinoma, whereas other patterns are invasive. Considerable interobserver variability in assessing “invasion” has been reported. We assessed the utility of cytokeratin 7 (CK7) stain and recently proposed International Association for the Study of Lung Cancer criteria to improve assessment of noninvasion in lung adenocarcinoma. MethodsFour pathologists (two staff, two trainees) assessed 158 hematoxylin and eosin (HE)- and CK7-stained slides of 108 pT1N0-2 nonmucinous lung adenocarcinoma cases. Scoring took place in four rounds. First, sections were independently scored for percentage of noninvasive or probable noninvasive and invasive or probable invasive patterns. Second, after a consensus scoring algorithm for CK7 was formulated, the slides were rescored. Subsequent third-round scoring was conducted only on HE slides using the 2023 International Association for the Study of Lung Cancer proposed criteria, and fourth-round scoring on both HE and CK7 slides simultaneously. Intraclass correlation coefficient (ICC) was calculated for each round. Recurrence-free survival was assessed using Cox proportional hazards regression methods. ResultsIn the first two rounds, interobserver concordance was consistently higher with CK7 (ICC range = 0.44–0.6) than HE (range = 0.24–0.49) scores. The IASLC proposed algorithm improved ICC of HE scores to 0.60 (95% confidence interval: 0.52–0.67), and round 4 HE and CK7 combined improved ICC to 0.75 (95% confidence interval: 0.70–0.80). Continuous measures of averaged noninvasive and probable noninvasive scores on HE were associated with improved recurrence-free survival (hazard ratio: 0.83–0.86). ConclusionsCK7 staining consistently increased interobserver concordance in assessment of invasive versus noninvasive patterns than HE. Combining CK7 with the 2023 IASLC criteria for morphologic features of invasion may further improve the interobservers’ concordance for the recognition of lepidic growth in nonmucinous lung adenocarcinoma.

Abstract 4248: Patient-derived MTAP-deleted bladder cancer organoid model: A unique platform for drug development

Abstract Introduction: Approximately 30% of advanced bladder cancer (BCa) tumors exhibit MTAP gene deletion, which has been associated with luminal subtype BCa, poor response to chemotherapy, and shorter progression to metastatic disease (De Souza 2023). MTAP deletion has been proposed as a potential biomarker for response to pemetrexed and an emerging class of PRMT5 and MAT2A inhibitors in development for MTAP-deleted tumors. Here, we describe the first patient-derived 3D model of BCa with MTAP deletion and its translational potential as a drug sensitivity platform. Methods: A patient (pt) with metastatic BCa harboring MTAP deletion underwent a pelvic lymph node biopsy. A fresh tumor sample was processed under collagenase/dispase solution for 30 minutes and filtered through a 70-um cell strainer. Cells were cultures in low attachment plates with MammoCult™ Basal Medium (Stem Cell Technologies). After 5 passages and 3D spheroid formation, immunostaining for cytokeratin (CK) 5 and 6, GATA3, CK20, and Ki67 staining were performed to confirm the urothelial origin. The MTAP-del was confirmed by a western blot of the organoid specimen and compared to other BLCa cell lines (UMUC, RT4, J82, and 5637). The viability assays were performed using the CellTiter-Glo® 3D Cell kit. Next-generation sequencing (NGS) of primary BCa tumor revealed deletion of MTAP, CDKN2A, CDKN2B, and mutations in p53, TERT, and KDM6A. Results: The patient organoid sample was positive for the luminal CK5/6 markers and negative for GATA3. Ki67 and CK20 were primarily expressed in the nucleus of the same sample. The organoid retained morphologic and immunohistochemical features of the patient’s primary bladder tumor, showing the same squamous differentiation and similar Ki-67 proliferative index. The organoid displayed no evidence of the MTAP protein by western blot (similar findings with the MTAP-del cell lines UMUC and RT4). The organoid showed sensitivity to pemetrexed [pemetrexed IC50=0.12 uM (other MTAP-del BLCa IC50: 0.1-0.23 uM)] and resistance to gemcitabine and cisplatin [Gem IC50=55 nM (other BLCa IC50: 1-29 nM), Cis IC50=90.5 uM (other BLCa IC50: 2-17 uM). Conclusions: We describe the first patient-derived MTAP-deleted BCa organoid recapitulating the features of the primary tumor. The organoid displayed sensitivity to pemetrexed as described with MTAP-deleted BCa. The organoid represents a unique platform for testing novel therapeutic strategies for MTAP-deleted BCa. Citation Format: Maximilian Schwermann, Benedito Carneiro, Shaolei Lu, Andre De Souza, Matthew Hadfield, Anthony Mega, Galina Lagos, Sari Khaleel, Dragan Golijanin, Sheldon Holder, Praveen Srinivasan, William MacDonald, Andrew George, Lanlan Zhou, Leiqing Zhang, Wafik El-Deiry. Patient-derived MTAP-deleted bladder cancer organoid model: A unique platform for drug development [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4248.

Deep learning approach for accurate prostate cancer identification and stratification using combined immunostaining of cytokeratin, p63, and racemase.

Prostate cancer (PCa) is the most frequently diagnosed cancer in men worldwide, affecting around 1.4 million individuals. Current PCa diagnosis relies on histological analysis of prostate biopsy samples, an activity that is both time-consuming and prone to observer bias. Previous studies have demonstrated that immunostaining of cytokeratin, p63, and racemase can significantly improve the sensitivity and the specificity of PCa detection compared to traditional H&E staining. This study introduces a novel approach that combines diagnosis-specific immunohistochemical (IHC) staining and deep learning techniques to provide reliable stratification of prostate glands. Our approach leverages a customized segmentation network, called K-PPM, that incorporates adaptive kernels and multiscale feature integration to enhance the functional information of IHC. To address the high class-imbalance problem in the dataset, we propose a weighted adaptive patch-extraction and specific-class kernel update. Our system achieved noteworthy results, with a mean Dice Score Coefficient of 90.36% and a mean absolute error of 1.64 % in specific-class gland quantification on whole slides. These findings demonstrate the potential of our system as a valuable support tool for pathologists, reducing workload and decreasing diagnostic inter-observer variability. Our study presents innovative approaches that have broad applicability to other digital pathology areas beyond PCa diagnosis. As a fully automated system, this model can serve as a framework for improving the histological and IHC diagnosis of other types of cancer.

Modeling an ascending infection by Ureaplasma parvum and its cell signaling and inflammatory response at the feto-maternal interface.

Ascending bacterial infection is associated with ∼ 40% of spontaneous preterm birth (PTB), and Ureaplasma spp. is one of the most common bacteria isolated from the amniotic fluid. Developing novel in vitro models that mimic in vivo uterine physiology is essential to study microbial pathogenesis. We utilized the feto-maternal interface organ-on-chip (FMi-OOC) device and determined the propagation of Ureaplasma parvum, and its impact on cell signaling and inflammation. FMi-OOC is a microphysiologic device mimicking fetal membrane/decidua interconnected through microchannels. The impact of resident decidual CD45+ leukocytes was also determined by incorporating them into the decidual chamber in different combinations with U. parvum. We tested the propagation of live U. parvum from the decidual to the amniochorion membranes (immunocytochemistry and quantitative PCR), determined its impact on cytotoxicity (LDH assay), cell signaling (JESSTM Western Blot), cellular transition (immunostaining for vimentin and cytokeratin), and inflammation (cytokine bead array). U. parvum transversed the chorion and reached the amnion epithelium after 72hours but did not induce cell signaling kinases (p38MAPK and JNK) activation, or cellular transition (epithelial-mesenchymal), regardless of the presence of immune cells. The inflammatory response was limited to the choriodecidual interface and did not promote inflammation in the amnion layer. Our data suggest that U. parvum is poorly immunogenic and does not produce massive inflammatory changes at the feto-maternal interface. We speculate that the presence of U. parvum may still compromise the feto-maternal interface making it susceptible to other pathogenic infection.

P-20 Pseudoglandular pattern in hepatocellular neoplasms and its diagnostic value with cytokeratin immunostain

P-20 Pseudoglandular pattern in hepatocellular neoplasms and its diagnostic value with cytokeratin immunostain

Comparative Study of Cytokeratin Immunostaining of Parotid Gland Parenchyma in Normal, Diabetic, and Excretory Duct Ligation of Mongrel Dogs.

The present study aimed to give a glimpse of the normal distribution of intermediate filaments within the parotid gland parenchyma of mongrel dogs and to reveal the pathological changes that may occur as a result of the effects of diabetes mellitus or atrophy of the gland caused by the ligation of the excretory duct to discover whether there is a similarity in these pathological behaviors. Twelve healthy mongrel dogs were used in the experiment and were divided into three groups: group I (the control group), group II (dogs with alloxan-induced diabetes), and group III (dogs with the right-side duct-ligated parotid gland). The dogs were sacrificed 45 days after the parotid excretory duct were tied. The right parotid gland of all groups was dissected and prepared for histological and immunohistochemical expression of cytokeratin 17 assay. Histological findings confirmed that the parotid gland parenchyma of the diabetic group had glandular atrophy characterized by the loss of gland structure, degenerated acini, and dilatation of the duct system. Moreover, there is a predominance of the fibrous component with the presence of fat cells within the gland compartments. On the contrary, the excretory duct-ligated group undergoes severe glandular atrophy of the previous character with the presence of duct-like structure as well as extravasation and vasodilatation. Immunohistochemical expression of cytokeratin 17 in control parotid using an immunoperoxidase technique showed that cytokeratin expression varies from negative to mild in all ducts and some serous acinar cells. The gland parenchyma of the diabetic group showed mild to strong cytokeratin expression of duct cells more concentrated in the apical part with moderate to strong expression of diffuse type in some serous acini. The intensity of cytokeratin 17 in gland compartments of the excretory duct-ligated group revealed a variation in expression that ranged from negative to strong diffuse staining throughout the gland. The severity and prevalence of cytokeratin 17 in our results are predictive of the pathological influence of both diabetes mellitus and duct ligation on the cytokeratin intracellular filaments of the salivary gland parenchyma in a different way that interferes with saliva production and/or secretion leading to xerostomia.

Apparent diffusion coefficient and intravoxel incoherent motion-diffusion kurtosis model parameters in invasive breast cancer: Correlation with the histological parameters of whole-slide imaging

ObjectivesTo evaluate the correlation of ADC and IVIM-diffusion kurtosis (DK) model parameters with quantitative histological parameters of whole-slide imaging (WSI). MethodsThis retrospective study (September 2015–July 2016) included 37 consecutive patients (all females; median age: 64 years, range, 41–82 years), each with a single invasive breast ductal carcinoma lesion with a mass appearance on preoperative MRI. DWI with b-values of 0, 50, 100, 300, 550, 850, and 1000 s/mm2 was performed. ADC maps were generated with b-values of 50 and 850 s/mm2. The IVIM-DK model was analysed using the following formula:Sb=S0fe−bD∗+1−fe−bD+1/6kbD2where S is the signal intensity, b is the b-value, f is the perfusion fraction, D* is the pseudo-diffusion coefficient of the vascular component, D is the diffusion coefficient of the non-vascular component, and k is the diffusion kurtosis. Whole tumour segmentation was performed to obtain the mean ADC, f, D*, D, and k. Quantitative histological parameters were obtained using cytokeratin immunostaining and WSI. The correlation of ADC and IVIM-DK model parameters with quantitative histological parameters was examined by Pearson's correlation. ResultsThe ADC was significantly correlated with the area ratio of interstitium (r = 0.53, p = 0.00082) and entropy (r = −0.58, p = 0.00019). k was significantly correlated with the area ratio of cancer cell nuclei (r = 0.53, p = 0.00079). ConclusionsSince the ADC reflected the area ratio of interstitium and entropy, and diffusion kurtosis reflected the area ratio of cancer cell nuclei, these parameters may be effective in distinguishing between benign and malignant breast tumours and in grading breast cancer.

Cytokeratin 19 expression in intrathoracic neoplasms: First study utilizing cellblocks, evaluating the role of a rarely used cytokeratin for lung cancers.

Cytokeratin 19 (CK19) is low-molecular weight CK with widespread expression in various malignancies including those of lung, colon, and breast. Occasional studies have also indicated prognostic utility of CK19 in nonsmall cell lung carcinomas, with better prognosis among squamous cell carcinoma cases with strong CK19 immunostaining. The aim of this study was to evaluate the expression and potential roles of CK19 immunostaining in cases of carcinoma lung as well as other metastatic intrathoracic neoplasms. This was a prospective single-center study of 2 years duration, during which 233 cases of endobronchial ultrasound guided aspiration of the intrathoracic lesions were examined. After excluding the cases without cellblock, with inadequate samples, with benign diagnosis, and excluding the lymphoma cases, finally 31 neoplastic cases were evaluated for expression of cytokeratin 19. Diffuse cytoplasmic staining for CK19 was noted in nonsmall cell lung carcinomas (18/19 cases), while the expression in small cell carcinomas was low (2/5 cases). CK19 was also positive in mediastinal lymph node metastasis from a case each of papillary thyroid carcinoma, prostatic adenocarcinoma and infiltrating ductal carcinoma breast (1/2 cases). It showed focal faint staining in a case of mediastinal mesenchymal neoplasm and was negative in both the cases of primitive neuroectodermal tumor. CK19 is a highly sensitive but nonspecific marker for nonsmall cell lung cancers. However, owing to diffuse staining pattern in most of these cases, role of its staining intensity as a predictor of prognosis could not be evaluated.

Florid Mesothelial Hyperplasia Associated with Abdominal Wall Endometriosis Mimicking Invasive Carcinoma

Florid mesothelial hyperplasia typically occurs in the pelvis, abdomen, or chest associated with an underlying neoplastic or inflammatory process. These lesions are of clinical significance because they can mimic a neoplasm. Early reports were published in the 1970s, but only a few case series of such lesions have been published in the gynecologic pathology literature. Here, we report a case of florid mesothelial hyperplasia with an infiltrative growth pattern, mimicking an invasive carcinoma. The lesion was associated with endometriosis forming a mass lesion in the abdominal wall. Histologically, tubular arrangements and nests of mesothelial cells, some with artifactual slit-like spaces, formed a stellate lesion adjacent to endometrial glands and stroma. Cytologic atypia was mild and reactive, and positive immunostaining for calretinin, WT-1, and cytokeratin 5 identified the lesion as mesothelial and benign. We describe in detail the histologic findings in this case and review the pertinent literature. We discuss the clinically importance of this diagnostic pitfall and the path to arriving at the correct diagnosis.

Characterization of a macrophagic-like cell line derived from rabbit fish (Siganus fuscescens): An illustration of anti-inflammatory responses of the herbal extract of Scutellaria baicalensis

A new cell line was isolated and characterized from the head kidney of Siganus fuscescens (rabbit fish). The new macrophagic-like cell line was named as rabbit fish macrophage (RFM), and which could be sub-cultured for over 50 cycles since the development. RFM cell line was tested for growth in different temperatures and serum concentrations: the best growing condition was optimized at 20% serum under 28 °C. In cultured RFM cells, sequencing of 18S rRNA, as well as immunostaining of cytokeratin and CD 68, confirmed the identity as macrophagic cell of S. fuscescens. Cultured RFM cells were exposed to challenge of inflammation, as triggered by LPS, showing highly sensitive responses to inflammation, including release of nitric oxide, expression of cytokine, and activation of phagocytosis. The water extract of aerial part of Scutellaria baicalensis, named as SBA, has been shown anti-inflammatory property in S. fuscescens fish. In order to extend the application of SBA in aquaculture, the extract and its effective flavonoids, i.e. baicalin and scutellarin, were applied in LPS-treated RFM cells. Application of SBA extract, baicalin or scutellarin, inhibited the expressions of LPS-induced inflammatory cytokines, i.e. IL-1β, TNF-α, as well as the signaling of transcription factor NF-κB. The results support the established RFM cell line could be an ideal in vitro model in drug screening relating to inflammation. Additionally, the notion of SBA herbal extract in fish aquaculture is supported by its efficacy against inflammation.

P–369 Three-dimensional co-culture of human endometrial cells with aneuploid and euploid embryos

Abstract Study question Is implantation different in euploid and aneuploid embryos? Summary answer By simulating the human endometrium using a three-dimensional scaffold, aneuploid embryos were unable to attach to the endometrial cells, while euploid embryos attached. What is known already Although embryo selection for transfer is usually based on morphology, 70% of embryos with high morphological quality have chromosomal abnormalities. The results of implantation and pregnancy rate assessments following Preimplantation Genetic Screening (PGS) are controversial. There is still no in vitro study to compare the implantation of human euploid and aneuploid embryos. Study design, size, duration After informed consent, 10 endometrial biopsies were taken from fertile women. For scaffolding, the stromal cells were resided within the matrix, after 24 hours, the epithelial cells were seeded on the scaffold. Cell culture continued for 5 days to reach the appropriate confluence. The embryos were also examined by performing PGS following CGH Array. 10 euploid and 10 aneuploid blastocysts were selected and co-cultured for 72 hours with the 3D structure of human endometrial cells. Participants/materials, setting, methods Endometrial cells were isolated and expanded in 2D cultures to achieve enough cells. The fibrin-agarose scaffold was made and stromal and epithelial cells were cultured into and on the scaffold, respectively. Then, cell proliferation was assessed by MTT assay. The simulated endometrial construct was confirmed by H&E and IHC. Partial hatching of blastocysts was performed using a laser system. The blastocyst’s attachment to the endometrial-like structure was examined under a phase-contrast microscope and SEM. Main results and the role of chance The MTT OD of scaffolds increased during 5 days of cell culture (P < 0.05). The histological evaluation of the co-culture systems was done under light microscopy by H&E staining. On the top of the 3D culture system, epithelial cells shaped a constricted cell monolayer. Stromal cells combined with the fibrin-agarose scaffold got lengthened and expanded, displaying that the 3D culture systems supplied a suitable environment for the growth of endometrial cells. In the 3D culture, the origins and locations of epithelial and stromal cells were defined by cytokeratin and vimentin immunostaining, respectively. IHC for cytokeratin was only positive for epithelial cells in the surface epithelium. IHC for the vimentin was positive for the stromal cells located in the 3D matrix. These results showed that fibrin-agarose scaffold could simulate the human endometrial structure. Using SEM and phase-contrast microscopy, it was found that only euploid embryos were able to attach to the endometrial construct while aneuploid embryos weren’t. Limitations, reasons for caution Since the co-culture does not contain a unique cell type, and the MTT OD standard curve against cell number is specified for cell type, the number of growing cells in the co-culture cannot be calculated; therefore it is reported as OD. Wider implications of the findings: Our findings determined that PGS allows us to transfer top-quality embryos with higher implantation potential. It improves implantation and pregnancy rate during ART cycles, especially in patients with recurrent implantation failure. Trial registration number Not applicable

Sarcomatoid Squamous Cell Carcinoma of the Conjunctiva: A Rare Entity and A Brief Review of Literature

Sacrcomatoid variant of squamous cell carcinoma has been known to involve numerous tissues like an aerodigestive tract, skin. A reddish mass of the left bulbar conjunctiva, with intraocular extension on ultra-bio-microscopy (UBM) was present. Histopathologic examination showed the characteristic spindle-shaped cells in continuity with the overlying epithelium with positive immunostains for cytokeratin, epithelial membrane antigen and vimentin. The presence of such an entity in the conjunctiva is rare and here we report such a case, this being the 37th case to be documented in literature. We also give a brief review of literature.

Sentinel Nodal Metastasis Detection in Endometrial Carcinoma With Microcystic, Elongated, and Fragmented (MELF) Pattern by Cytokeratin Immunostaining.

The microcystic, elongated, and fragmented (MELF) pattern of myoinvasion in endometrial carcinoma (EC) is associated with an increased risk of lymph node metastasis. Our aim is to assess the role of cytokeratin immunohistochemical (IHC) stains in detecting sentinel nodal metastasis in MELF pattern tumors. We recovered 19 MELF pattern EC hysterectomies with lymphadenectomy from our files. Negative nodes were subjected to cytokeratin AE1/AE3 IHC. Ten additional cases with sentinel lymph node (SLN) biopsies primarily assessed by IHC were also analyzed. Of the 19 cases of EC, 6 had positive lymph nodes based on H&E-stained sections at the time of their initial diagnosis. With the addition of IHC stains, 8 previously negative cases were found to have node metastases, and 3 of these were SLNs. Among the 10 cases primarily assessed by IHC, 5 had malignant cells in their SLNs. Cytokeratin IHC staining detected malignant cells in 9 of 16 cases with SLNs in our sample of women with MELF pattern of myoinvasion. Immunohistochemical stains should be routinely performed on SLNs from all MELF-positive cases to detect occult lymph node metastases and isolated tumor cells.

Preoperative prediction of granulation pattern subtypes in GH-secreting pituitary adenomas.

The aim of this study was to establish a preoperative prediction method for sparsely granulated (SG) growth hormone (GH)-secreting pituitary adenoma, an aggressive tumour subtype with high recurrence risk, in acromegaly patients. Eighty-three patients with GH-secreting pituitary adenomas were included in this study. GH measurements, cytokeratin immunostaining and electron microscopy were performed to detect granulation patterns. Preoperative factors, including general, radiological and endocrinological features and acute octreotide suppression test outcomes, were compared between SG and densely granulated (DG) groups. The predictive capabilities of these features were analysed using a receiver operating characteristic (ROC) curve, and the most predictive features were combined to establish a grading scale. Thirty-nine of the 83 patients had SG GH-secreting pituitary adenomas; 44 had DG tumours. SG tumours tended to occur in younger patients and have larger diameters and volumes, higher Knosp grades, lower GH indexes and normalized insulin-like growth factor-1 (IGF-1) level, and a lower ∆GH% after octreotide treatment. The tumour size, Knosp grade, GH index and ∆GH% after octreotide treatment had good predictive performance, with area under the curve (AUC) values ranging from 0.70 to 0.80. Combining four parameters, including diameter, Knosp grade, GH index and ∆GH% after octreotide treatment, we established a grading scale for predicting SG GH-secreting pituitary adenomas with an AUC of 0.84 and relatively high sensitivity and specificity. We propose a predictive method for distinguishing SG and DG GH-secreting pituitary adenomas preoperatively. This method will help physicians identify candidates for presurgical medical treatment and neurosurgeons determine radical surgical strategies for high-risk tumours.

Silent Pituitary Adenoma in Nasal Cavity

Introduction: Silent corticotroph adenomas (SCA) represent 3-6% of all pituitary adenomas. SCA are confirmed on post-surgical histological tumour staining and can either have no prior clinical or biochemical evidence of excess ACTH or alternatively only biochemical (not clinical) evidence of Cushing’s disease. SCA are potentially large and aggressive tumours with a greater tendency to post-surgical recurrence. We present a case of a giant SCA which pre-operatively didn’t exhibit any clinical signs of cortisol excess but showed signs of early recurrence on follow up imaging. Case: A 40-year-old man presented with frontal and maxillary sinus pain and nasal stuffiness. Nasal endoscopy revealed a mass in the nasal cavity and sphenoid sinus. Subsequent CT scan of his sinuses and MRI Pituitary reported a 4 x 3.5 x 2.5 cm sellar mass extending through the floor of sella into the sphenoid sinus and nasal cavity. Despite no clinical features of any pituitary hormone excess (including Cushing’s) or deficiency, his biochemical parameters were consistent with hypercortisolaemia. Post-low dose dexamethasone (2 mg over 48 hours) suppression cortisol was inadequate: 106 nmol/L (<50nmol/L). His corresponding serum dexamethasone level was 8.6 nmol/L (>3nmol/L), ACTH 46 ng/L (0-46 ng/L) and 24-hour urinary free cortisol levels were 502 and 260 nmol/24 hours (<165 nmol/24 hours)) on 2 separate occasions. The remainder of his pituitary profile was normal. He therefore underwent expanded endonasal endoscopic resection of tumour without significant complication. His post-operative cortisol was <50nmol/L (other pituitary hormone measurements normal) so hydrocortisone replacement therapy was commenced. A 3-month post-operative MRI pituitary scan revealed near total resection of the tumour with peripheral enhancement within the tumour resection site representing post-surgical changes. Histopathology showed sparsely granulated, diffuse cytokeratin immunostaining and immunopositivity for ACTH and T-pit consistent with a corticotroph PitNet of the sparsely granulated variety with low Ki 67 index. His six month post-operative scan showed recurrent sellar mass of 2.2 x 0.9 x 1.4 cm. Again there was an absence of clinical features of Cushing’s syndrome. After multidisciplinary discussion clinical, biochemical and radiological surveillance was continued with a view to further transsphenoidal surgery and/or radiotherapy depending on the progress of the tumour. Conclusion: Our patient solely had biochemical (not clinical) evidence of Cushing’s disease despite a large pituitary tumour in keeping with an SCA, which are aggressive, have high recurrence rate and shorter time to recurrence after surgery. Close and frequent post-operative clinical, biochemical and radiological surveillance is critical to detect and manage early recurrence that may require multimodal therapy.

Comparative ultrastructural-functional characterizations of the skin in three reptile species; Chalcides ocellatus, Uromastyx aegyptia aegyptia, and Psammophis schokari aegyptia (FORSKAL, 1775): Adaptive strategies to their habitat.

The current investigation was planned utilizing SEM, histological, and furthermore cytokeratin immunohistochemical to give a full depiction of skin of three reptiles species; Chalcides ocellatus (Scincidae), Uromastyx aegyptia aegyptia (Agamidae), and Psammophis schokari aegyptia (Colubridae) captured from Egypt with various ecological environment. Our SEM results showed different scales covered epidermis of three reptile's species with diverse surface microstructure. Overlapped rhomboid scales with numerous lenticular sense organs with numerous pores and oberhäutchen layer with microridges in C. ocellatus. In U. aegyptia, scales were overlapped elliptical-shaped possess lens-like sense structure with several scattered pits and oberhäutchen layer with polygonal outlined cells. While in P. schokari aegyptia, smooth scales flattened with two large dome-shaped scale receptors/sensilla and lens-like sense structure, moreover polygonal-shaped micro-ornamentation in scale hinge joints were observed. Histologically, skin of three species had outer epidermis with stratum germinativum, stratum corneum (α-keratin, β-keratin layer) capped by surface Oberhäutchen and inner dermis. Osteoderms were observed with dermis of C. ocellatus. There are marked variation within pigment cells types among examined species. Melanophores observed in dermal layer of C. ocellatus, while in U. aegyptia, three pigment cells in tegument dermis; melanophores, xanthophores, and iridophores whereas, P. schokari aegyptia had two forms of chromatophore cells (melanophores and iridophores) in dermis and few melanophores scattered between stratum germinativum cells. The highest cytokeratin immunostaining observed in epidermal cell layer of U. aegyptia aegyptia than two other species. Conclusion, dry scaly skin of reptiles reflects a great range of functional aspects and success to adapt with terrestrial life.

RARE-58. CONGENITAL METASTATIC CHORDOMA OF THE CLIVUS

Chordomas are rare midline axial skeletal neoplasms that typically present in adults. They are infrequent in childhood with typical localization in the spheno-occipital skull base. They are derived from remnants of the embryonic notochord. We present the case of 4 months old girl, who was born with „blueberry muffin” syndrome and was first negatively diagnosed for neuroblastoma and leukemia (two negative skin biopsies were performed) was admitted with axial laxity. In imaging testes there was a tumor of the scull base, metastases in the lungs and kidneys (that were not seen at previous assessments) and a small lesion in the heart. The third biopsy of skin lesion was performed and pathological examination revealed a neoplasm composed of cords, clusters, and chains of multivacuolated cells embedded within a myxoid matrix and separated by fibrous septa. No atypical and dedifferentiated features were present. Mitotic activity was not observed. Neoplastic cells showed the typical cytoplasmic immunostaining for EMA, S100 and cytokeratin AE1/AE3, strong nuclear brachyury expression, and retention of nuclear INI-1 expression. The diagnosis of chordoma was established. Neoplastic tissue and blood samples were obtained for molecular analysis using next generation sequencing, including germline mutations assessment (are ongoing). Chemotherapy as for soft tissue sarcomas was undertaken. Currently a patient is on treatment with improvement of neurological status.

Synovial Metaplasia of the Endometrium

Abstract Introduction/Objective Synovial-like metaplasia is well known to accompany breast implants, but it can also rarely be found in the endometrium, associated with intrauterine devices. We present two cases of endometrial synovial-like metaplasia. Methods The hysterectomy specimens were diagnosed at Northwell Health and reviewed at the departmental Consensus Conference. Immunostains for cytokeratin AE1/AE3 were performed on the endometrium. The histopathologic findings were correlated with the clinical information. Results Both patients had had intrauterine devices placed for birth control. One, a 47-year-old female with a family history of Lynch syndrome and newly diagnosed with a corresponding mutation, underwent prophylactic hysterectomy. The second patient, a 50-year-old female with a history of uterine prolapse, was experiencing stress urinary incontinence, for which she underwent supracervical hysterectomy. Both patients had intrauterine devices in place at the time of hysterectomy. Both uteri showed endometrial synovial-like metaplasia. It was associated with a complete absence of the endometrial surface epithelium. The synovial-like cells were eosinophilic, plump and elongated, were palisaded perpendicular to the endometrial surface and were negative for cytokeratin AE1/AE3, which immunostain also confirmed the absence of the endometrial surface epithelium and the presence of some residual deeper endometrial glands. Conclusion Endometrial synovial-like metaplasia is a rare entity described in patients with intrauterine devices. The mesodermal-derived endometrium is apparently able, when exposed to a semi-mobile foreign body, to develop a way to minimize friction by mimicking synovium, similar to the connective tissue facing breast implants. Such surface must be better at withstanding chronic friction than the regular endometrial epithelial surface, leading to this unusual and fascinating change. Since the resulting surface mimics synovium perfectly, both morphologically and immunohistochemically, and serves a very similar mechanical purpose, we propose to call this phenomenon, wherever it is found, “synovial metaplasia” instead of the current term “synovial-like metaplasia.”

MiR-18a Contributes to Preeclampsia by Downregulating Smad2 (Full Length) and Reducing TGF-β Signaling

The study investigated the regulation of Smad2 by miR-18a and its role in preeclampsia (PE). Bioinformatics analysis showed that both Smad2 and Smad3 were the predicted targets for miR-18a. Mass spectrum analysis showed that two mature Smad2 isoforms existed in human placenta: full length, Smad2(FL), and that lacking exon3, Smad2(Δexon3). The protein level of Smad2(FL), but not Smad2(Δexon3) or Smad3, was significantly increased in severe PE (sPE) placenta, which was inversely correlated with the level of miR-18a. Elevated Smad2(FL) phosphorylation level appeared in sPE placenta, and Smad2 was colocalized with miR-18a in various subtypes of trophoblasts in human placenta. Smad2(FL) was validated as the direct target of miR-18a in HTR8/SVneo cells. miR-18a enhanced trophoblast cell invasion, which was blocked by the overexpression of Smad2(FL). Furthermore, overexpression of miR-18a repressed Smad2 activation and the inhibition of trophoblast cell invasion by transforming growth factor-β (TGF-β). In conclusion, our results suggest that miR-18a inhibits the expression of Smad2(FL), but not Smad2(Δexon3) or Smad3, which can reduce TGF-β signaling, leading to the enhancement of trophoblast cell invasion. A lack of miR-18a, which results in the upregulation of Smad2(FL), contributes to the development of PE.

The herbal extract deriving from aerial parts of Scutellaria baicalensis shows anti-inflammation and anti-hypoxia responses in cultured fin cells from rabbit fish

A new cell line derived from dorsal fin of rabbit fish Siganus fuscescens was developed and characterized. The cell line was isolated from the dorsal fin, named as rabbit fish fin (RFF) cell line, and which was sub-cultured for 50 cycles since the development. This cell line was tested for growth in different temperatures and serum concentrations, and the best growing condition was at 20% serum at 28 °C. In cultured RFF cells, amplification of 18S rRNA from genomic DNA and immunostaining of cellular cytokeratin confirmed the proper identity of S. fuscescens fish. After 30th passage of cultures, the cells were exposed to challenge of inflammation, triggered by LPS, and hypoxia, mimicked by CoCl2. Cultured RFF cells showed robust sensitive responses to inflammation and hypoxia in directing the expressions of cytokines and hypoxia inducible factor-1α (HIF-1α). The water extract of aerial part of Scutellaria baicalensis (SBA) has been shown in rabbit fish to prevent inflammation. Here, we extended this notion of testing the efficacy of SBA extract in the developed cultured RFF cells. Application of SBA extract inhibited the expression of LPS-induced inflammatory cytokines, i.e. IL-1β, IL-6, as well as the signaling of NF-κB. The application of CoCl2 in cultured RFF cells triggered the hypoxia-induced cell death and up regulation of HIF-1α. As expected, applied SBA extract in the cultures prevented the hypoxia-induced signaling. Our results show the established RFF cell line may be served as an ideal in vitro model in drug screening relating to inflammation and hypoxia. Additionally, we are supporting the usage of SBA herbal extract in fish aquaculture, which possesses efficacy against inflammation and hypoxia.