Stratified Epithelium Research Articles

For Better or Worse: Modulation of the Host DNA Damage Response by Human Papillomavirus.

High-risk human papillomaviruses (HPVs) are associated with several human cancers. HPVs are small, DNA viruses that rely on host cell machinery for viral replication. The HPV life cycle takes place in the stratified epithelium, which is composed of different cell states, including terminally differentiating cells that are no longer active in the cell cycle. HPVs have evolved mechanisms to persist and replicate in the stratified epithelium by hijacking and modulating cellular pathways, including the DNA damage response (DDR). HPVs activate and exploit DDR pathways to promote viral replication, which in turn increases the susceptibility of the host cell to genomic instability and carcinogenesis. Here, we review recent advances in our understanding of the regulation of the host cell DDR by high-risk HPVs during the viral life cycle and discuss the potential cellular consequences of modulating DDR pathways.

Keloid Co-Existing with epidermoid Cyst in a Black Elderly Patient: A Case Report

Keloid is an abnormal scar that can be due to tissue trauma or occur spontaneously. It commonly occurs in dark pigmented individuals with positive family history. Epidermoid cyst (EC) on the other hand is an adnexal cysts that contain keratin and stratified epithelium. The co-existence of both keloid and EC is rare and very few have been reported in the literature especially among the Negroids. The objective is to report a keloid mass co-existing with an EC in a Negroid elderly patient. A 76-year-old man presented with right sided upper back swelling of a year duration with no prior trauma or family history of keloid. Physical examination showed a 2 × 7 cm raised keloid scar with an intraoperative incidental finding of a 1 × 1 cm hemispheric epidermoidcyst. The significance of this case is in the rarity of co-existing keloid and EC especially in the elderly and Negroids.

Whiteboard animation simulation for dental education.

Whiteboard animation is a specific type of animation where line drawing and narration are used to explain concepts. The combination of visual thinking and storytelling makes whiteboard animations a unique and effective educational tool. Storytelling is a multimodal teaching approach that simultaneously engages listeners’ thinking, emotions, and imagination.1 However, this powerful tool is underutilized in higher education, particularly science education.1 Whiteboard animations, powered by storytelling, have successfully explained complex health-related topics to pharmacology students2 and positively impacted the retention, engagement, and enjoyment of university-level students.3 We planned to create whiteboard animations for dental students to explain complex science topics using graphics and storytelling. However, we have encountered two challenges while attempting to make whiteboard animations as an educator: (1) Making a good quality whiteboard animation requires paid software, which can be expensive. (2) Most whiteboard animation software is designed for business advertising and does not provide the graphics gallery needed to explain health science-related topics. The graphics gallery of the software do not have images representing the microanatomy of the tooth, oral cavity, and cellular processes. As a solution, we have used traditional PowerPoint and hand-drawn digital images to mimic whiteboard animation. Procreate, a stylus pen, and an iPad were used for drawings that were exported into PowerPoint. The built-in animations of PowerPoint were used to animate the images and supporting text (Figure 1). Screen-capture software was used to record the animation screen as a video with a voice-over narration. Our whiteboard animation explains the types of epithelial cells in a storytelling manner. First-year dentistry students learn about the epithelium as part of histology. The animation starts with the story of a boy who wants to eat cereal and milk with a fork. The narration then draws a connection between the structure-function relationships of a utensil to the structure-function relations of cells in the epithelium (Figure 2). We have incorporated interactive self-assessment questions allowing viewers to assess their learning. The learning objective of the animation and corresponding assessment questions are shown in Table 1. Learning objective whiteboard animation: Epithelium of the gastrointestinal (GI) tract Identify the types of epithelial cells based on their shape and function. Differentiate between simple and stratified epithelium. Define the type of epithelium found in different parts of the gastrointestinal tract. Describe the correlation between the shape of an epithelial cell and its function in the gastrointestinal tract. Q1. Which epithelium comprises cells whose height is two–three times greater than their width and are arranged in an ordered layer? Simple cuboidal epithelium Simple columnar epithelium *** Simple squamous epithelium Stratified epithelium Q2. Which epithelium comprises several layers, such that cells high up in the epithelium are not in contact with the underlying extracellular matrix? Simple cuboidal epithelium Simple columnar epithelium Simple squamous epithelium Stratified epithelium *** Characterize the epithelium of the oral cavity. Explain how the epithelium of the oral cavity is different from the rest of the gastrointestinal tract. Q3. What type of epithelium is found in our oral cavity? Stratified squamous epithelium *** Simple columnar epithelium Stratified columnar epithelium Simple squamous epithelium Traditionally whiteboard animations use hand motion to draw and write on a whiteboard. However, PowerPoint does not have that option. As an alternative, we used the animated “human hand” to bring and stick the picture on a white screen or color the important regions instead of appearing it to draw. Adding background music and sound effect can make the video engaging and entertaining. According to Cognitive Load Theory, the learning load of working memory can be reduced by alternating how a topic is presented.4 Whiteboard animations offer an alternative learning scaffold, therefore expected to reduce the extrinsic load of understanding complex concepts. Stories with real-world examples will also help students understand and retain the information. The authors declare no conflicts of interest.

AGS3 antagonizes LGN to balance oriented cell divisions and cell fate choices in mammalian epidermis.

Oriented cell divisions balance self-renewal and differentiation in stratified epithelia such as the skin epidermis. During peak epidermal stratification, the distribution of division angles among basal keratinocyte progenitors is bimodal, with planar and perpendicular divisions driving symmetric and asymmetric daughter cell fates, respectively. An apically restricted, evolutionarily conserved spindle orientation complex that includes the scaffolding protein LGN/Pins/Gpsm2 plays a central role in promoting perpendicular divisions and stratification, but why only a subset of cell polarize LGN is not known. Here, we demonstrate that the LGN paralog, AGS3/Gpsm1, is a novel negative regulator of LGN and inhibits perpendicular divisions. Static and ex vivo live imaging reveal that AGS3 overexpression displaces LGN from the apical cortex and increases planar orientations, while AGS3 loss prolongs cortical LGN localization and leads to a perpendicular orientation bias. Genetic epistasis experiments in double mutants confirm that AGS3 operates through LGN. Finally, clonal lineage tracing shows that LGN and AGS3 promote asymmetric and symmetric fates, respectively, while also influencing differentiation through delamination. Collectively, these studies shed new light on how spindle orientation influences epidermal stratification.

Abstract 594: Effects of interstitial pressure on lung cancer cells

Abstract Chronic inflammation is a risk factor for various types of cancer. The pressure in the interstitial tissues has been reported to be elevated in chronic inflammation and cancer. However, little is known about the biological effect of pressure from the basal side on epithelia, and a significance of the elevated interstitial fluid pressure in cancer biology is still poorly understood. In this study, we investigated the effect of hydrostatic pressure on cultured lung cancer cells grown on Transwell permeable filters by changing the height of the surfaces of culture medium in the apical and basal sides. A gradient of the hydrostatic pressure from the basal to the apical side induced stratification of epithelia. The elimination of the hydrostatic pressure gradient restored the stratification of epithelia. Interestingly, cavities were found within the stratified epithelia, and the characteristics of apical cell polarity including microvilli and tight junctions were formed in the surfaces the cavities. The cavities with similar aberrant cell polarity were also observed in human lung cancer tissues. The hydrostatic pressure gradient from the basal side also promoted cell proliferation and suppressed apoptosis. On the other hand, the expression levels of phosphorylated ERK and AKT were not significantly affected by the hydrostatic pressure. These results indicate that the pressure from the basal side to the epithelia promotes cell proliferation, suppresses apoptosis and induces aberrant cell polarity, which results in epithelial stratification. The findings in this study suggest that the elevated interstitial fluid pressure in chronic inflammation and cancer tissues facilitates cancer growth and plays a significant role in cancer biology. Citation Format: Shinsaku Tokuda, Keisuke Onoi, Tadaaki Yamada, Koichi Takayama. Effects of interstitial pressure on lung cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 594.

Morphological analysis of the digestive tract of Hypophthalmus marginatus (Valenciennes, 1840).

The mapará, Hypophthalmus marginatus, is a rheophilic and planktophagous catfish and it is one of the main fishing resources in the Amazon region. To understand its nutritional characteristics, this work described the morphology and histochemistry of the digestive tract of H. marginatus. The oropharyngeal cavity has abundant, long and thin gill rakers, responsible for retaining plankton, while the short and muscular oesophagus promotes the transport of these items to the stomach and prevents water ingestion. The stratified oesophageal epithelium has goblet cells with mucins predominantly neutral that could facilitate the food passage the food passage. The U-shaped siphonal stomach shows a columnar epithelium with neutral mucins to protect against autodigestion. The cardiac and fundic regions have gastric glands, while the pyloric region has a thick muscular layer with a sphincter. The coiled intestine with intestinal quotient of 2.14 ± 0.5 shows long longitudinal folds in the anterior region, decreasing in height in the aboral direction, and highlighting its function in digestion and nutrient absorption. Goblet cells are abundant in posterior intestine and rectum, and rectum shows epithelial cells with mucins in the apical cytoplasm, for protection and defecation. Intraepithelial lymphocytes also are abundant in posterior intestine and rectum, and it acts in immunological defence.

Warty Dyskeratoma with involvement of multiple hair follicles in a dog

The warty dyskeratoma is a neoplasm of follicular origin, with uncommon incidence in veterinary medicine, and benign biological behavior. An excisional biopsy sample was received from a dog, female, Shih Tzu, 12 years old, from the left axillary region. Grossly, after section, green and friable multifocal areas were identified, streaked by solid white areas. Microscopically, the analysis revealed multifocal cystic structures, lined by stratified epithelium, with basal area including villous epithelial projections, toward the dermis, and the luminal region featured acantholysis and dyskeratosis, as well as parakeratosis. Those findings were compatible with a warty dyskeratoma affecting multiple hair follicles, thus being the first case report in veterinary medicine with such this unique pattern. Up to now, the literature encompassed only human medicine. This report, besides collaborating to the description of a rare presentation of an uncommon neoplasm, also contributes with new information found for epidemiological studies of this tumor.

Morphological and ultrastructural study of the palpebral conjunctiva of healthy domestic cats.

To describe the morphology of the meibomian glands and goblet cells in the palpebral conjunctiva of healthy cats. Five healthy domestic cats without ocular changes that had died from causes unrelated to the study were evaluated. Forty samples were collected from upper and lower palpebral conjunctiva and 20 from palpebral fornix region in the nasal corner. The samples were processed for scanning electron microscopy (SEM), transmission electron microscopy (TEM), and histopathology. In the SEM analysis of the palpebral fornix, numerous points of mucous extrusion between the cell junctions were visualized, along with the presence of microvilli in the apical portions with small secretory vesicles. A homogeneous surface was highlighted, formed by the arrangement of cell contours in the form of hexagons. The grouping of goblet cells and their cytoplasmic vesicles filled with homogeneous content was visualized using TEM. Histopathology showed goblet cells interspersed with stratified epithelium accompanied by well-vascularized connective tissue. In the samples stained with hematoxylin and eosin, the meibomian glands, formed by acinar cells and with the presence of individual openings of the ducts in the eyelid margin, were easily visualized in the eyelid margins. This study describes the ultrastructural form of goblet cells and the morphology of the palpebral conjunctiva of healthy cats by the histopathology of the meibomian glands. This description can serve as a parameter of normality and aid in the detection of morphological alterations in these structures, as well as a parameter for comparison with other animal species.

Single-cell transcriptomics uncovers the differentiation of a subset of murine esophageal progenitors into taste buds in vivo

Mouse esophagus is lined with a stratified epithelium, which is maintained by the constant renewal of unipotent progenitors. In this study, we profiled mouse esophagus by single-cell RNA sequencing and found taste buds specifically in the cervical segment of the esophagus. These taste buds have the same cellular composition as the ones from the tongue but express fewer taste receptor types. State-of-the-art transcriptional regulatory network analysis allowed the identification of specific transcription factors associated to the differentiation of immature progenitors into the three different taste bud cell types. Lineage tracing experiments revealed that esophageal taste buds arise from squamous bipotent progenitor, thus demonstrating that all esophageal progenitors are not unipotent. Our cell resolution characterization of cervical esophagus epithelium will enable a better understanding of esophageal progenitor potency and insights into the mechanisms involved in the development of taste buds.

Human papillomavirus in the setting of immunodeficiency: Pathogenesis and the emergence of next-generation therapies to reduce the high associated cancer risk.

Human papillomavirus (HPV), a common sexually transmitted virus infecting mucosal or cutaneous stratified epithelia, is implicated in the rising of associated cancers worldwide. While HPV infection can be cleared by an adequate immune response, immunocompromised individuals can develop persistent, treatment-refractory, and progressive disease. Primary immunodeficiencies (PIDs) associated with HPV-related disease include inborn errors of GATA, EVER1/2, and CXCR4 mutations, resulting in defective cellular function. People living with secondary immunodeficiency (e.g. solid-organ transplants recipients of immunosuppression) and acquired immunodeficiency (e.g. concurrent human immunodeficiency virus (HIV) infection) are also at significant risk of HPV-related disease. Immunocompromised people are highly susceptible to the development of cutaneous and mucosal warts, and cervical, anogenital and oropharyngeal carcinomas. The specific mechanisms underlying high-risk HPV-driven cancer development in immunocompromised hosts are not well understood. Current treatments for HPV-related cancers include surgery with adjuvant chemotherapy and/or radiotherapy, with clinical trials underway to investigate the use of anti-PD-1 therapy. In the setting of HIV co-infection, persistent high-grade anal intraepithelial neoplasia can occur despite suppressive antiretroviral therapy, resulting in an ongoing risk for transformation to overt malignancy. Although therapeutic vaccines against HPV are under development, the efficacy of these in the setting of PID, secondary- or acquired- immunodeficiencies remains unclear. RNA-based therapeutic targeting of the HPV genome or mRNA transcript has become a promising next-generation therapeutic avenue. In this review, we summarise the current understanding of HPV pathogenesis, immune evasion, and malignant transformation, with a focus on key PIDs, secondary immunodeficiencies, and HIV infection. Current management and vaccine regimes are outlined in relation to HPV-driven cancer, and specifically, the need for more effective therapeutic strategies for immunocompromised hosts. The recent advances in RNA-based gene targeting including CRISPR and short interfering RNA (siRNA), and the potential application to HPV infection are of great interest. An increased understanding of both the dysregulated immune responses in immunocompromised hosts and of viral persistence is essential for the design of next-generation therapies to eliminate HPV persistence and cancer development in the most at-risk populations.

Adverse pulmonary effects after oral exposure to copper, manganese and mercury, alone and in mixtures, in a Spraque-Dawley rat model

ABSTRACT The rise in respiratory disease has been attributed to an increase in environmental pollution. Heavy metals contribute to environmental contamination via air, water, soil and food. The effects of atmospheric exposure to heavy metals on pulmonary structure and function have been researched, but the effects through drinking water have been neglected. The aim of this study was to investigate the potential in vivo alterations in the pulmonary tissue of male Sprague-Dawley rats after a 28-day oral exposure to copper (Cu), manganese (Mn) and mercury (Hg), alone and in mixtures, at 100 times the World Health Organization’s (WHO) safety limit for each heavy metal in drinking water. Forty-eight male Sprague-Dawley rats were randomly divided into eight groups (n = 6): control, Cu, Mn, Hg, Cu + Mn, Cu + Hg, Mn + Hg and Cu, Mn + Hg. The morphology of lung tissue and the bronchioles were evaluated using light- and transmission electron microscopy. For all exposed groups, morphological changes included thickened inter- and intra-alveolar spaces, stratified epithelium, disrupted smooth muscle and early fibrosis and desquamation of the epithelia of the bronchioles to varying degrees. In all exposed groups, ultrastructurally, an increase in disarranged collagen and elastin fibers, nuclear membrane detachment, chromatin condensation, indistinct nucleoli and an increase in collagen fiber disarrangement was observed. This study has identified that oral exposure to Cu, Mn and Hg and as part of mixtures caused pathogenesis due to inflammation, cellular damage and fibrosis with Mn + Hg being the most potent heavy metal group.

Innovative Therapeutic Approaches for the Treatment of the Ocular Morbidities in Patients with EEC Syndrome.

Ectrodactyly-Ectodermal dysplasia-Clefting (EEC) syndrome is caused by heterozygous missense point mutations in the p63 gene, an important transcription factor during embryogenesis and for stem cell differentiation in stratified epithelia. Most of the cases are sporadic, related to de novo mutations arising during early-stage development. Familial cases show an autosomic dominant inheritance. The major cause of visual morbidity is limbal stem cell failure, which develops in the second to third decade of life. Patients often show ocular surface alterations, such as recurrent blepharitis and conjunctivitis, superficial microlesions of the cornea, and spontaneous corneal perforation and ulceration, leading to progressive corneal clouding and eventually visual loss. No definitive cures are currently available, and treatments to alleviate symptoms are only palliative. In this review, we will discuss the proposed therapeutic strategies that have been tested or are under development for the management of the ocular defects in patients affected by EEC syndrome: (i) gene therapy-based approaches by means of Allele-Specific (AS) siRNAs to correct the p63 mutations; (ii) cell therapy-based approaches to replenish the pool of limbal stem cells; and (iii) drug therapy to correct/bypass the genetic defect. However, as the number of patients with EEC syndrome is too limited, further studies are still necessary to prove the effectiveness (and safety) of these innovative therapeutic approaches to counteract the premature differentiation of limbal stem cells.

Spread of premalignant mutant clones and cancer initiation in multilayered tissue

Over 80% of human cancers originate from the epithelium, which covers the outer and inner surfaces of organs and blood vessels. In stratified epithelium, the bottom layers are occupied by stem and stem-like cells that continually divide and replenish the upper layers. In this work, we study the spread of premalignant mutant clones and cancer initiation in stratified epithelium using the biased voter model on stacked two-dimensional lattices. Our main result is an estimate of the propagation speed of a premalignant mutant clone, which is asymptotically precise in the cancer-relevant weak-selection limit. We use our main result to study cancer initiation under a two-step mutational model of cancer, which includes computing the distributions of the time of cancer initiation and the size of the premalignant clone giving rise to cancer. Our work quantifies the effect of epithelial tissue thickness on the process of carcinogenesis, thereby contributing to an emerging understanding of the spatial evolutionary dynamics of cancer.

The Epstein-Barr Virus Glycoprotein BDLF2 Is Essential for Efficient Viral Spread in Stratified Epithelium.

Epstein-Barr virus (EBV) is a ubiquitous human pathogen that infects the majority of the adult population regardless of socioeconomic status or geographical location. EBV primarily infects B and epithelial cells and is associated with different cancers of these cell types, such as Burkitt lymphoma and nasopharyngeal carcinoma. While the life cycle of EBV in B cells is well understood, EBV infection within epithelium is not, largely due to the inability to model productive replication in epithelium in vitro. Organotypic cultures generated from primary human keratinocytes can model many aspects of EBV infection, including productive replication in the suprabasal layers. The EBV glycoprotein BDLF2 is a positional homologue of the murine gammaherpesvirus-68 protein gp48, which plays a role in intercellular spread of viral infection, though sequence homology is limited. To determine the role that BDLF2 plays in EBV infection, we generated a recombinant EBV in which the BDLF2 gene has been replaced with a puromycin resistance gene. The ΔBDLF2 recombinant virus infected both B cell and HEK293 cell lines and was able to immortalize primary B cells. However, the loss of BDLF2 resulted in substantially fewer infected cells in organotypic cultures compared to wild-type virus. While numerous clusters of infected cells representing a focus of infection are observed in wild-type-infected organotypic cultures, the majority of cells observed in the absence of BDLF2 were isolated cells, suggesting that the EBV glycoprotein BDLF2 plays a major role in intercellular viral spread in stratified epithelium. IMPORTANCE The ubiquitous herpesvirus Epstein-Barr virus (EBV) is associated with cancers of B lymphocytes and epithelial cells and is primarily transmitted in saliva. While several models exist for analyzing the life cycle of EBV in B lymphocytes, models of EBV infection in the epithelium have more recently been established. Using an organotypic culture model of epithelium that we previously determined accurately reflects EBV infection in situ, we have ascertained that the loss of the viral envelope protein BDLF2 had little effect on the EBV life cycle in B cells but severely restricted the number of infected cells in organotypic cultures. Loss of BDLF2 has a substantial impact on the size of infected areas, suggesting that BDLF2 plays a specific role in the spread of infection in stratified epithelium.

Transglutaminase Activity Is Conserved in Stratified Epithelia and Skin Appendages of Mammals and Birds.

The cross-linking of structural proteins is critical for establishing the mechanical stability of the epithelial compartments of the skin and skin appendages. The introduction of isopeptide bonds between glutamine and lysine residues depends on catalysis by transglutaminases and represents the main protein cross-linking mechanism besides the formation of disulfide bonds. Here, we used a fluorescent labeling protocol to localize the activity of transglutaminases on thin sections of the integument and its appendages in mammals and birds. In human tissues, transglutaminase activity was detected in the granular layer of the epidermis, suprabasal layers of the gingival epithelium, the duct of sweat glands, hair follicles and the nail matrix. In the skin appendages of chickens, transglutaminase activity was present in the claw matrix, the feather follicle sheath, the feather sheath and in differentiating keratinocytes of feather barb ridges. During chicken embryogenesis, active transglutaminase was found in the cornifying epidermis, the periderm and the subperiderm. Transglutaminase activity was also detected in the filiform papillae on the tongue of mice and in conical papillae on the tongue of chickens. In summary, our study reveals that transglutaminase activities are widely distributed in integumentary structures and suggests that transglutamination contributes to the cornification of hard skin appendages such as nails and feathers.

New insight into Epstein-Barr virus infection using models of stratified epithelium.

Epstein-Barr virus (EBV) is a ubiquitous human pathogen that is transmitted in saliva. EBV transits through the oral epithelium to infect B cells, where it establishes a life-long latent infection. Reinfection of the epithelium is believed to be mediated by virus shed from B cells, but whether a latent reservoir can exist in the epithelia is unknown. We previously developed an in vitro organotypic model of stratified epithelium where EBV can readily replicate within the suprabasal layers of the epithelium following apical infection mediated by virus-producing B cells. Given that infected epithelial cells and cell-free virus are observed in saliva, we examined the ability of both of these to mediate infection in organotypic cultures. Epithelial-derived cell-free virus was able to infect organotypic cultures from the apical surface, but showed enhanced infection of B cells. Conversely, B cell-derived virus exhibited enhanced infection of epithelial cells. While EBV has been detected in basal cells in oral hairy leukoplakia, it is unknown whether EBV can be seen in undifferentiated primary keratinocytes in the basal layer. Undifferentiated epithelial cells expressed proposed EBV receptors in monolayer and were susceptible to viral binding and entry. Integrins, and occasionally ephrin A2, were expressed in the basal layer of gingiva and tonsil derived organotypic cultures, but the known B-cell receptors HLAII and CD21 were not detected. Following infection with cell-free virus or virus-producing B cells at either the apical or basolateral surface of preformed organotypic cultures, abundant infection was detected in differentiated suprabasal cells while more limited but readily detectable infection was observed in the undifferentiated basal cells. Together, our data has provided new insight into EBV infection in stratified epithelium.

Somatic BAP1 Loss in Ovarian Serous Borderline Tumor and Recurrent Low-grade Serous Carcinoma From a Germline BAP1 Mutation Carrier.

To the Editor: In a recent issue, Dr. Oliva and colleagues proposed the use of the immunohistochemical markers, Claudin-4 and BAP1, to distinguish between low-grade serous neoplasms and peritoneal mesothelioma 1. This article is of particular interest to us, as we have recently published on a cohort of 119 clinically annotated low-grade serous carcinomas, confirmed by central pathology review, with molecular profiling by targeted next-generation sequencing 2. In this cohort, we encountered 1 patient who represents an unusual exception to relying on loss of BAP1 expression to exclude the diagnosis of a low-grade serous neoplasm. This was a 59-yr-old female who underwent comprehensive surgical staging for bilateral ovarian masses. Final pathology revealed a stage IIIC serous borderline tumor involving bilateral ovaries (Fig. 1A), associated with noninvasive peritoneal implants and involvement of pelvic lymph nodes. After 7 yrs, she developed recurrent disease involving the sigmoid colon, in the form of metastatic low-grade serous carcinoma (Fig. 1B). Both primary and recurrent tumors showed a similar immunophenotype, characterized by strong diffuse expression of epithelial/Mullerian markers, including Claudin-4 (Fig. 1C), BerEP4, PAX8, ER, and PR, while negative for markers of mesothelial differentiation, namely, calretinin, and D2-40. Somatic mutational profiling by targeted next-generation sequencing revealed the low-grade serous carcinoma to harbor a BAP1 frameshift mutation (Y241Lfs*2). Germline testing further revealed a heterozygous BAP1 germline mutation (c.1110dupC, p.Met371Hisfs*27). Loss of BAP1 expression by immunohistochemistry in the low-grade serous carcinoma, as well as the prior serous borderline tumor (Fig. 1D), was consistent with bi-allelic inactivation of BAP1 as an early event in the molecular pathogenesis of this case.FIG. 1: Low-grade serous neoplasms associated with germline BAP1 mutation. (A) Ovarian serous borderline tumor showing characteristic hierarchical branching papillae lined by stratified columnar epithelium (inset: ciliated cells observed at high-magnification, consistent with tubal differentiation). (B) Recurrent invasive low-grade serous carcinoma involving muscularis propria of the sigmoid colon, showing infiltrating nests and micropapillae composed of tumor cells with small monotonous nuclei, and scattered psammoma bodies. Immunohistochemical analysis of the serous borderline tumor showing (C) diffuse expression of Claudin-4 and (D) loss of BAP1 expression in tumor cells with retained nuclear staining in non-neoplastic stromal cells.To our knowledge, this represents the first reported case of a BAP1-deficient low-grade serous neoplasm. It illustrates how genetic alterations of oncogenes and tumor suppressor genes are not tissue or disease-specific and emphasizes the importance of applying several cell lineage markers, along with morphologic correlation, for appropriate diagnostic classification of tumor type.

P63 as an Ideal Diagnostic Marker for Pleomorphic Adenoma: An Immunohistochemical Study

Introduction: Pleomorphic Adenoma (PA) histopathologically represents a heterogeneous lesion with a varying proportion of epithelial and mesenchymal components. Due to its morphological diversity, a diagnostic dilemma often arises when identifying its various patterns. This diverse morphology is considered to be a function of the neoplastic myoepithelium. Tumor protein 63 (p63) is expressed in stratified epithelia and in the basal cells of salivary glands. Aim: To investigate the immunoreactivity of p63 as a reliable myoepithelial marker in PA. Materials and Methods: A cross-sectional study was performed in the Department of Oral and Maxillofacial Pathology at PGIDS, Rohtak, Haryana, India. The duration of the study was 14 months, from March 2015 to April 2016. A total of 15 tissue blocks of histopathologically diagnosed cases of PA were included from departmental archives and subjected to Immunohistochemistry (IHC) using a monoclonal p63 antibody. The myoepithelium was classified as myoepithelial-like (abluminal and spindled), modified myoepithelium (myxoid and chondroid), and transformed myoepithelium (solid epithelioid, squamous, and basaloid cribriform). IHC for p63 was assessed in each myoepithelial component, as well as in non neoplastic Myoepithelial Cells (MEC) and inner tubular epithelial cells. Only nuclear staining for p63 was considered positive. The obtained data were subjected to statistical analysis, and the Chi-square test was applied. Results: The PA samples subjected to IHC showed 100% p63 reactivity. Transformed MEC, abluminal, and modified MEC revealed variable immunostaining with a significant difference (p=0.049). There was no immunostaining in luminal/inner layer cells in all cases of PA. p63 was also expressed in the nuclei of MEC of acini and intercalated ducts of normal salivary gland tissue. Conclusion: p63 is a sensitive and specific myoepithelial marker to identify MEC in PA. Additionally, the expression of MECassociated markers in acini and intercalated ducts of normal salivary glands has confirmed their role in the histogenesis of this tumour.

Structural and histochemical aspects of the Gymnotus carapo tegument (Teleostei: Gymnotiformes).

The present study aims to structurally and histochemically characterize the Gymnotus carapo tegument. 30 specimens were captured and slaughtered by spinal section with anesthesia. The observation was carried out with a stereoscopic microscope and the body surface was photographed. Fragments of the dorsal, ventral and lateral region were fixed in Bouin's solution for 12 hours and subsequently preserved in 70% alcohol. They were subsequently observed in the scanning electron microscope (SEM). The preparation for SEM was performed following the standardized protocol. Histological preparations were made, and the cuts were colored with H-E, PAS and Coomassie Blue. The images were obtained in an Olympus BX41-ENUTV-4 microscope. From the observations in SEM a plain tegument with pores of different sizes could be evidenced. The scales of the different regions of the body have different ornaments. Microscopically it was composed of a stratified non-keratinized epithelium consisting of two types of morphologically distinct cells: epidermal cells and mucous cells (PAS-Commassie Blue positive). Under the epithelium there is a layer of dense irregular connective tissue with associated chromatophores and more deeply scales. These analyzes are the basis for future studies that will focus on elucidating the events related to integumentary healing in this species.

Placental development in the early stages of red-rumped agouti pregnancy (Dasyprocta leporina Linnaeus, 1758).

Hystricomorpha rodents display a similar placentation model to humans. The present study was carried out considering the scarcity of information concerning the placental development in agouti. Describe the microscopy of the placenta, subplacenta and yolk sac of agoutis in early pregnancy and report on the inversion of the yolk sac. Fifteen females between the 14th-32nd day of gestation were used following euthanasia. Gestational buttons were collected, fixed, processed, stained to optical microscopy or immunohistochemistry. Chorioallantoic placenta (CP) ranged from conical to a half-sphere, as follows: from the 14th to 17th day, the CP displays an inverted "V" shape, predominantly formed by cytotrophoblasts; from 20 to 22 days, formed almost entirely by cytotrophoblasts; at 28 days, a half sphere, with distinct lobes and interlobular area, numerous maternal gaps delimited by syncytiotrophoblasts and trophoblast giant cells; at 32 days, globose and undergoing the maturation process. Subplacenta, located between decidua and CP, initially presents septa consisting of simple columnar epithelium and after 17 days, comprising stratified epithelium. Visceral yolk sac (VYS) is attached to two CP projections between 14 and 17 days, formed by a simple cubic epithelium and inverted. Between 20 and 22 days, the epithelium displays apical villous projections with cytoplasmic vacuoles and a vascularized mesoderm. After the 24th day, the VYS near the placenta is pleated, very vascularized and villous, with decreased villi sizes further away from the placenta. The agouti CP displays similar characteristics to other hystricomorpha, including placenta lobulation, a subplacenta and an inverted vitelline placenta.