56,753 research outputs found

    Cellular and molecular characterization of the corneal epithelium in Xenopus frogs

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    The vertebrate cornea is a transparent, avascular tissue that forms the front window of the eye. Specifically, the corneal epithelium is exposed to detrimental conditions including infections, solar (Ultraviolet, UV) irradiation, mechanical injuries, and undergoes constant self-renewal. Corneal epithelial stem cells (CESCs) and their progeny, the transit amplifying cells (TACs), play a prominent role in the maintenance of corneal homeostasis, transparency, and wound repair processes. Towards this end, my thesis focuses on understanding the molecular signature of the corneal epithelium using the frog, Xenopus laevis, with the goal to explore novel markers that will reliably identify populations of CESCs and TACs. In Chapter 2, I start by examining the expression of known corneal biomarkers (previously reported in literature from corneal studies of various vertebrates) in the vertebrate species, the African clawed frog. Here, I used antibody-based immunohistochemical staining to molecularly characterize the expression of nine proteins in the corneas of both Xenopus larvae and post-metamorphic adults. I found that localization of some markers changes between tadpole and juvenile adult stages. Markers such as p63, Keratin19, and beta1-integrin are restricted to basal corneal epithelial cells of the larvae. After metamorphosis their expression is found in basal and intermediate layer cells of the adult frog cornea epithelium. Another protein, Pax6 was expressed in the larval corneas, but surprisingly it was not detectable in the adult corneal epithelium. For the first time we report that Tcf7l2 can be used as a marker to differentiate cornea vs. skin in frogs. Tcf7l2 is present only in the frog skin, which differs from reports indicating that the protein is expressed in the human cornea. Furthermore, I identified the transition between the inner, and the outer surface of the adult frog eyelid as a key boundary in terms of marker expression. Although these markers are useful to identify different regions and cellular layers of the frog corneal epithelium, none was unique to CESCs or TACs. The results of this study substantiate findings from other studies in the field indicating that there may not be a single conserved, specific CESC marker in vertebrates. To overcome the limitation of candidate-based biomarker characterization, however, I undertook single-cell genomics approaches to understand the temporal cell atlas of the frog cornea (Chapter 3). Using ~22,000 corneal cells isolated from two distinct developmental time-points, this work provides key insights about the amphibian corneal transcriptome. The data also reveals several novel genes expressed in corneal cells and spatiotemporal changes in gene expression during corneal differentiation. In addition, the data helps in understanding the developmental trajectory of corneal cells during development and differentiation, and identifies key gene regulatory networks that are involved in corneal maturation. In conclusion, this work provides a detailed molecular characterization of the Xenopus corneal epithelium and establishes distinct and newly identified biomarkers of corneal cellular layers. Although a single biomarker for CESCs was not identified in this work (and may not even exist), it helps identify a range of proteins that could be tested for their functional role in regulating the population of corneal stem cells in vertebrates, including humans. Furthermore, this work will be valuable for future studies to understand the critical factors that regulate cornea epithelial cells and lens regeneration, the latter phenomenon being unique to the larval stages of Xenopus frogs.LimitedAuthor requested closed access (OA after 2yrs) in Vireo ETD syste

    Comparative histomorphometric and histochemical analysis of gastrointestinal tract of five rodent species

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    Prehrana ┼żivotinja odra┼żava se u gra─Ĺi probavnog sustava, a razlike se mogu o─Źitovati u debljini pojedinih slojeva te broju i tipu ┼żljezdanih vr─Źastih stanica koje izlu─Źuju mukozni sekret za probavu i za┼ítitu. U ovom radu izabrala sam pet vrsta glodavaca koji ┼żive u simpatriji na kr┼íkom podru─Źju centralne Dalmacije, s ciljem utvr─Ĺivanja postoje li histomorfometrijske i histokemijske razlike u gra─Ĺi probavnog sustava koje su rezultat razlika u prehrani ovih vrsta. Uklju─Źene vrste su Apodemus epimelas, Apodemus sylvaticus, Dinaromys bogdanovi, Eliomys quercinus i Glis glis. Histomorfometrijski rezultati su pokazali ve─çu debljinu oro┼żenog epitela jednjaka i manju debljinu vanjskog mi┼íi─çnog sloja jednjaka kod karnivornih vrsta, dok vrste koje se hrane visoko kalori─Źnom i lako probavljivom hranom imaju manju debljinu sluznice ┼żlijezdanog dijela ┼żeluca i deblji epitel debelog crijeva. Vrste kod kojih prehrana sadr┼żi otrovne tvari imaju specifi─Źne prilagodbe debljine pojedinih slojeva. Histokemijska analiza pokazala je da karnivorne vrste imaju vi┼íe kiselih vr─Źastih stanica u tankom crijevu, a granivorne vi┼íe mje┼íovitih. Ovi rezultati predstavljaju nove podatke za uklju─Źene vrste i daju novi uvid u postojanje razlika izme─Ĺu srodnih vrsta istra┼żivanih glodavaca s razli─Źitom prehranom, ali i izme─Ĺu razli─Źitih porodica, mi┼íeva iz roda Apodemus i puhova, sa sli─Źnom prehranom.Animal diet is reflected in the structure of the digestive tract. Some of the differences can be manifested in the thickness of individual layers or the number and type of glandular goblet cells that secrete mucose for digestion and protection. For this thesis, I selected five species of rodents that live in a sympatry in the karstic area of central Dalmatia, with the aim of determining if there are any histomorphometric and histochemical differences in the structure of the gastrointestinal tract, as a result of distinct diets of the species. Species that were included are Apodemus epimelas, Apodemus sylvaticus, Dinaromys bogdanovi, Eliomys quercinus i Glis glis. The histomorphometric results showed thicker esophageal keratenized epithelium and thinner esophageal outer muscular layer in carnovorous species, while species with highcalorie and easily digestible diets have thinner mucose layer of the glandular part of the stomach and thicker epithelium of the large intestine. Species whose diet contains toxic subtances showed specific adaptations in the thicknesses of certain layers. Histochemical analysis showed that carnivorous species have more acidic goblet cells in the small intestine, while granivorous species have more mixed ones. These results represent new data for included species and provide new insight into the existence of both differences between related species of rodents with distinct diets, and between species of different families but with similar diet, as between Apodemus genus and dormice species

    A Comparison of the Human Umbilical Cord's Histomorphometric and Histological Structure in Pregnant Diabetic and Non-Diabetic Women

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    This study aimed to explore the histomorphometrical and histopathological alterations of umbilical cord (UC) vessels caused by gestational diabetes mellitus (GDM) and pre-gestational diabetes mellitus (PGDM). A total of thirty UC samples were obtained from full term pregnant women without any complications. Ten out of thirty UCs were obtained from non-diabetic pregnant women (normal group), 10 from GDM and 10 from PGDM pregnant women. Segments from the placental attachment, center and fetal side of UCs were taken for each group. These segments were processed for paraffin blocks, sectioned, and stained with H&E, Masson trichrome (MT) and Periodic Acid Shift (PAS). The results of the histomorphometric study showed no significant differences in the UC mean weight among these three groups. In three different segments, GDM resulted in a significant decrease in artery and vein wall thickness compared to the control group. GDM and PGDM resulted in a non-significant difference in the diameter of artery and vein in fetal segment, the vein in placental segment, and artery in the central segment compared with normal. All the UCs in the three groups contained two arteries and one vein but only one cord recorded in the GDM group contained one artery and one vein. Histological study of diabetic UC segments showed extravasation of blood, artery discordance, degeneration of WartonÔÇÖs jelly (WJ) fibers with formation of honeycomb like empty spaces, formation of multiple spaces between smooth muscle cells of tunica media and detachment of the umbilical arteries from surrounding WJ. In both diabetic groups, there was a marked decrease in collagen fibers in tunica intima and media with their irregular arrangement in both arteries and vein especially in placental segment. The results also showed there was a rich carbohydrate content in the intima and media in all three groups. In conclusion, the current results proved that GDM and PGDM have an adverse effect on the structure of UC and its vessels. Keywords: Umbilical cord; Diabetes mellitus; Histological study; Special stains; Blood vessels DOI: 10.7176/JBAH/13-6-04 Publication date: April 30th 2023

    Analysis of Wild Type and Variant B Cystatin C Interactome in Retinal Pigment Epithelium Cells Reveals Variant B Interacting Mitochondrial Proteins

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    Cystatin C, a secreted cysteine protease inhibitor, is abundantly expressed in retinal pigment epithelium (RPE) cells. A mutation in the protein's leader sequence, corresponding to formation of an alternate variant B protein, has been linked with an increased risk for both age-related macular degeneration (AMD) and Alzheimer's disease (AD). Variant B cystatin C displays intracellular mistrafficking with partial mitochondrial association. We hypothesized that variant B cystatin C interacts with mitochondrial proteins and impacts mitochondrial function. We sought to determine how the interactome of the disease-related variant B cystatin C differs from that of the wild-type (WT) form. For this purpose, we expressed cystatin C Halo-tag fusion constructs in RPE cells to pull down proteins interacting with either the WT or variant B form, followed by identification and quantification by mass spectrometry. We identified a total of 28 interacting proteins, of which 8 were exclusively pulled down by variant B cystatin C. These included 18 kDa translocator protein (TSPO) and cytochrome B5 type B, both of which are localized to the mitochondrial outer membrane. Variant B cystatin C expression also affected RPE mitochondrial function with increased membrane potential and susceptibility to damage-induced ROS production. The findings help us to understand how variant B cystatin C differs functionally from the WT form and provide leads to RPE processes adversely affected by the variant B genotype

    Mushroom ╬▓-glucan and polyphenol formulations as natural immunity boosters and balancers: nature of the application

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    Mushrooms are experiencing a kind of renaissance as a part of the contemporary human diet. These valuable organisms are more than food, they fi t in perfectly as a novel market group known as nutra-mycoceuticals. Immune-balancing mushroom dietary fibers and secondary metabolites such as polyphenols are the main focus of the healthcare industry. Wellness and cosmetic companies are increasingly using mushroom extracts rich in these ingredients. This review considers the basic molecular immunomodulatory mechanisms of action of the most commonly used mushroom dietary fibers, ╬▓-glucans. The literature data on their bioavailability, metabolic transformations, preclinical and human clinical research, and safety are discussed. Immunomodulatory mechanisms of polyphenol ingredients are also considered. These molecules present great potential in the design of the new immunity balancer formulations according to their widespread structural diversity. Finally, we draw attention to the perspectives of modern trends in mushroom nutraceutical and cosmeceutical formulations to strengthen and balance immunity

    Cutaneous anaplastic large T-cell lymphoma in a cat - case report

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    ABSTRACT Lymphoma is the most common lymphoproliferative disorder in cats. However, cutaneous lymphomas are uncommon in this species and can be classified as epitheliotropic or non-epitheliotropic. Epitheliotropic lymphomas are typically of T-cells and have tropism for epidermis and/or adnexal epithelium. Non-epitheliotropic lymphomas can be of either T-cell or B-cell and involve predominantly the dermis. The present study describes a case of multifocal cutaneous anaplastic large T-cell lymphoma. The skin nodules were multiple and variable in size, alopecic and erythematous in appearance and randomly distributed. Immunohistochemistry revealed positivity for the CD3 antigen, consistent with T-lymphocytes. This neoplasm should be remembered whenever multiple cutaneous nodules are observed in cats

    Pathogenesis and treatment of chronic rhinosinusitis from the perspective of sinonasal epithelial dysfunction

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    BackgroundChronic rhinosinusitis (CRS) is a clinical syndrome primarily characterized by long-term mucosal inflammation of the nasal cavity and sinuses. The pathogenesis of CRS is still unclear due to its high heterogeneity. A number of studies have recently focused on the sinonasal epithelium. Thus, there has been a quantum leap in awareness of the role of the sinonasal epithelium, which is now understood as an active functional organ rather than simply an inert mechanical barrier. Undoubtedly, epithelial dysfunction plays a vital role in the onset and development of CRS.ObjectiveIn this article, we discuss the potential contribution of sinonasal epithelium dysfunction to CRS pathogenesis and explore a few current and developing therapeutic options targeting the sinonasal epithelium.ResultsImpaired mucociliary clearance (MCC) and an abnormal sinonasal epithelial barrier are usually considered to be the main causative factors in CRS. Epithelial-derived bioactive substances, such as cytokines, exosomes, and complements, play a vital role in the regulation of innate and adaptive immunity and contribute to the pathophysiological alterations of CRS. The phenomena of epithelialÔÇômesenchymal transition (EMT), mucosal remodeling, and autophagy observed in CRS offer some novel insights into the pathogenesis of this disease. In addition, existing treatment options targeting disorder of sinonasal epithelium can help to relieve the main symptoms associated with CRS to some extent.ConclusionThe presence of a normal epithelium is fundamental for maintaining homeostasis in the nasal and paranasal sinuses. Here, we describe various aspects of the sinonasal epithelium and highlight the contributions of epithelial dysfunction to CRS pathogenesis. Our review provides sound evidence of the need for in-depth study of the pathophysiological alterations of this disease and for the development of novel epithelium-targeting alternative treatments

    Galectin-3 promotes secretion of proteases that decrease epithelium integrity in human colon cancer cells.

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    Galectin-3 is a galactoside-binding protein that is commonly overexpressed in many epithelial cancers. It is increasingly recognized as a multi-functional, multi-mode promoter in cancer development, progression, and metastasis. This study reports that galectin-3 secretion by human colon cancer cells induces cancer cell secretion, in an autocrine/paracrine manner, of a number of proteases including cathepsin-B, MMP-1 and MMP-13. The secretion of these proteases causes disruption of epithelial monolayer integrity, increases its permeability and promotes tumour cell invasion. This effect of galectin-3 is shown to be mediated through induction of cellular PYK2-GSK3╬▒/╬▓ signalling and can be prevented by the presence of galectin-3 binding inhibitors. This study thus reveals an important mechanism in galectin-3-mediated promotion of cancer progression and metastasis. It provides further evidence to the increased realization of galectin-3 as a potential therapeutic target for the treatment of cancer
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