Effect of cellular senescence on the response of human peritoneal mesothelial cells to TGF-ß.

Transforming growth factor ß (TGF-ß) is implicated in both mesothelial-to-mesenchymal transition (MMT) and cellular senescence of human peritoneal mesothelial cells (HPMCs). We previously showed that senescent HPMCs could spontaneously acquire some phenotypic features of MMT, which in young HPMCs were induced by TGF-ß. Here, we used electron microscopy, as well as global gene and protein profiling to assess in detail how exposure to TGF-ß impacts on young and senescent HPMCs in vitro. We found that TGF-ß induced structural changes consistent with MMT in young, but not in senescent HPMCs. Of all genes and proteins identified reliably in HPMCs across all treatments and states, 4,656 targets represented overlapping genes and proteins. Following exposure to TGF-ß, 137 proteins and 46 transcripts were significantly changed in young cells, compared to 225 proteins and only 2 transcripts in senescent cells. Identified differences between young and senescent HPMCs were related predominantly to wound healing, integrin-mediated signalling, production of proteases and extracellular matrix components, and cytoskeleton structure. Thus, the response of senescent HPMCs to TGF-ß differs or is less pronounced compared to young cells. As a result, the character and magnitude of the postulated contribution of HPMCs to TGF-ß-induced peritoneal remodelling may change with cell senescence.

Curcumin and gallic acid have a synergistic protective effect against ovarian surface epithelium and follicle reserve damage caused by autologous intraperitoneal ovary transplantation in rats.

The objective of this study to examine the effects of curcumin and gallic acid use against oxidative stress damage in the autologous intraperitoneal ovarian transplantation model created in rats on ovarian follicle reserve, ovarian surface epithelium, and oxidant-antioxidant systems. 42 adult female Sprague Dawley rats (n=7) were allocated into 6 groups. Group 1 served as the control. In Group 2, rats underwent ovarian transplantation (TR) to their peritoneal walls. Group 3 received corn oil (CO) (0.5 ml/day) one day before and 14 days after transplantation. Group 4 was administered curcumin (CUR) (100 mg/kg/day), Group 5 received gallic acid (GA) (20 mg/kg/day), and Group 6 was treated with a combination of curcumin and gallic acid via oral gavage after transplantation. Rats were sacrificed on the 14th postoperative day, and blood along with ovaries were collected for analysis. The removed ovaries were analyzed at light microscopic, fluorescence microscopic, and biochemical levels. In Group 2 and Group 3, while serum and tissue Total Oxidant Levels (TOS) and Oxidative Stress Index (OSI) increased, serum Total Antioxidant Levels (TAS) decreased statistically significantly (p˂0.05) compared to the other groups (Groups 1, 4, 5, and 6). The ovarian follicle reserve was preserved and the changes in the ovarian surface epithelium and histopathological findings were reduced in the antioxidant-treated groups (Groups 4, 5, and 6). In addition, immunofluorescence examination revealed that the expression of Cytochrome C and Caspase 3 was stronger and Ki-67 was weaker in Groups 2 and 3, in comparison to the groups that were given antioxidants. It can be said that curcumin and gallic acid have a histological and biochemical protective effect against ischemia-reperfusion injury due to ovarian transplantation, and this effect is stronger when these two antioxidants are applied together compared to individual use.

Epithelium-derived 6-nitrodopamine modulates noradrenaline-induced contractions in human seminal vesicles.

AIMS: To evaluate the basal release of 6-nitrodopamine (6-ND) from human isolated seminal vesicles (HISV) and to characterize its action and origin. MAIN METHODS: Left HISV obtained from patients undergoing prostatectomy surgery was suspended in a 3-mL organ bath containing warmed (37 °C) and gassed (95%O2:5%CO2) Krebs-Henseleit's solution (KHS) with ascorbic acid. An aliquot of 2 mL of the supernatant was used to quantify catecholamines by LC-MS/MS. For functional studies, concentration-responses curves to catecholamines were obtained, and pEC50 and Emax values were calculated. Detection of tyrosine hydroxylase and S100 protein were also carried out by both immunohistochemistry and fluorescence in-situ hybridization assays (FISH). KEY FINDINGS: Basal release of 6-ND was higher than the other catecholamines (14.76 ± 14.54, 4.99 ± 6.92, 3.72 ± 4.35 and 5.13 ± 5.76 nM for 6-ND, noradrenaline, adrenaline, and dopamine, respectively). In contrast to the other catecholamines, the basal release of 6-ND was not affected by the sodium current (Nav) channel inhibitor tetrodotoxin (1 µM; 10.4 ± 8.9 and 10.4 ± 7.9 nM, before and after tetrodotoxin, respectively). All the catecholamines produced concentration-dependent HISV contractions (pEC50 4.1 ± 0.2, 4.9 ± 0.3, 5.0 ± 0.3, and 3.9 ± 0.8 for 6-ND, noradrenaline, adrenaline, and dopamine, respectively), but 6-ND was 10-times less potent than noradrenaline and adrenaline. However, preincubation with very low concentration of 6-ND (10-8 M, 30 min) produced significant leftward shifts of the concentration-response curves to noradrenaline. Immunohistochemical and FISH assays identified tyrosine hydroxylase in tissue epithelium of HISV strips. SIGNIFICANCE: Epithelium-derived 6-ND is the major catecholamine released from human isolated seminal vesicles and that modulates smooth muscle contractility by potentiating noradrenaline-induced contractions.

Exploring Salivary Epithelial Dysfunction in Sjögren's Disease.

Sjögren's Disease (SjD) is an autoimmune disease of the exocrine tissues. Etiological events result in the loss of epithelial homeostasis alongside extracellular matrix (ECM) destruction within the salivary and lacrimal glands, followed by immune cell infiltration. In this review, we have assessed the current understanding of epithelial-mesenchymal transition (EMT)-associated changes within the salivary epithelium potentially involved in salivary dysfunction and SjD pathogenesis. We performed a PubMed literature review pertaining to the determination of pathogenic events that lead to EMT-related epithelial dysfunction and signaling in SjD. Molecular patterns of epithelial dysfunction in SjD salivary glands share commonalities with EMT mediating wound healing. Pathological changes altering salivary gland integrity and function may precede direct immune involvement while perpetuating MMP9-mediated ECM destruction, inflammatory mediator expression, and eventual immune cell infiltration. Dysregulation of EMT-associated factors is present in the salivary epithelium of SjD and may be significant in initiating and perpetuating the disease. In this review, we further highlight the gap regarding mechanisms that drive epithelial dysfunction in salivary glands in the early or subclinical pre-lymphocytic infiltration stages of SjD.

LRBA, a BEACH protein mutated in human immune deficiency, is widely expressed in epithelia, exocrine and endocrine glands, and neurons.

Mutations in LRBA, a BEACH domain protein, cause severe immune deficiency in humans. LRBA is expressed in many tissues and organs according to biochemical analysis, but little is known about its cellular and subcellular localization, and its deficiency phenotype outside the immune system. By LacZ histochemistry of Lrba gene-trap mice, we performed a comprehensive survey of LRBA expression in numerous tissues, detecting it in many if not all epithelia, in exocrine and endocrine cells, and in subpopulations of neurons. Immunofluorescence microscopy of the exocrine and endocrine pancreas, salivary glands, and intestinal segments, confirmed these patterns of cellular expression and provided information on the subcellular localizations of the LRBA protein. Immuno-electron microscopy demonstrated that in neurons and endocrine cells, which co-express LRBA and its closest relative, neurobeachin, both proteins display partial association with endomembranes in complementary, rather than overlapping, subcellular distributions. Prominent manifestations of human LRBA deficiency, such as inflammatory bowel disease or endocrinopathies, are believed to be primarily due to immune dysregulation. However, as essentially all affected tissues also express LRBA, it is possible that LRBA deficiency enhances their vulnerability and contributes to the pathogenesis.

Analysis of long-range chromatin contacts, compartments and looping between mouse embryonic stem cells, lens epithelium and lens fibers.

BACKGROUND: Nuclear organization of interphase chromosomes involves individual chromosome territories, "open" and "closed" chromatin compartments, topologically associated domains (TADs) and chromatin loops. The DNA- and RNA-binding transcription factor CTCF together with the cohesin complex serve as major organizers of chromatin architecture. Cellular differentiation is driven by temporally and spatially coordinated gene expression that requires chromatin changes of individual loci of various complexities. Lens differentiation represents an advantageous system to probe transcriptional mechanisms underlying tissue-specific gene expression including high transcriptional outputs of individual crystallin genes until the mature lens fiber cells degrade their nuclei. RESULTS: Chromatin organization between mouse embryonic stem (ES) cells, newborn (P0.5) lens epithelium and fiber cells were analyzed using Hi-C. Localization of CTCF in both lens chromatins was determined by ChIP-seq and compared with ES cells. Quantitative analyses show major differences between number and size of TADs and chromatin loop size between these three cell types. In depth analyses show similarities between lens samples exemplified by overlaps between compartments A and B. Lens epithelium-specific CTCF peaks are found in mostly methylated genomic regions while lens fiber-specific and shared peaks occur mostly within unmethylated DNA regions. Major differences in TADs and loops are illustrated at the ~ 500 kb Pax6 locus, encoding the critical lens regulatory transcription factor and within a larger ~ 15 Mb WAGR locus, containing Pax6 and other loci linked to human congenital diseases. Lens and ES cell Hi-C data (TADs and loops) together with ATAC-seq, CTCF, H3K27ac, H3K27me3 and ENCODE cis-regulatory sites are shown in detail for the Pax6, Sox1 and Hif1a loci, multiple crystallin genes and other important loci required for lens morphogenesis. The majority of crystallin loci are marked by unexpectedly high CTCF-binding across their transcribed regions. CONCLUSIONS: Our study has generated the first data on 3-dimensional (3D) nuclear organization in lens epithelium and lens fibers and directly compared these data with ES cells. These findings generate novel insights into lens-specific transcriptional gene control, open new research avenues to study transcriptional condensates in lens fiber cells, and enable studies of non-coding genetic variants linked to cataract and other lens and ocular abnormalities.

Basal actomyosin pulses expand epithelium coordinating cell flattening and tissue elongation.

Actomyosin networks constrict cell area and junctions to alter cell and tissue shape. However, during cell expansion under mechanical stress, actomyosin networks are strengthened and polarized to relax stress. Thus, cells face a conflicting situation between the enhanced actomyosin contractile properties and the expansion behaviour of the cell or tissue. To address this paradoxical situation, we study late Drosophila oogenesis and reveal an unusual epithelial expansion wave behaviour. Mechanistically, Rac1 and Rho1 integrate basal pulsatile actomyosin networks with ruffles and focal adhesions to increase and then stabilize basal area of epithelial cells allowing their flattening and elongation. This epithelial expansion behaviour bridges cell changes to oocyte growth and extension, while oocyte growth in turn deforms the epithelium to drive cell spreading. Basal pulsatile actomyosin networks exhibit non-contractile mechanics, non-linear structures and F-actin/Myosin-II spatiotemporal signal separation, implicating unreported expanding properties. Biophysical modelling incorporating these expanding properties well simulates epithelial cell expansion waves. Our work thus highlights actomyosin expanding properties as a key mechanism driving tissue morphogenesis.

Developmental relationship between junctional epithelium and epithelial rests of Malassez.

Keratin 17 (K17) is thought to be a candidate target gene for regulation by Lymphoid Enhancer Factor-1 (Lef-1). K17 is a marker that distinguishes junctional epithelium (JE) from epithelial rests of Malassez (ERM). However, the relationship of Lef-1 to K17 is not clear in this context. Moreover, the expression of other keratins such as K5, K6, K7 and K16 is not reported. Therefore, the aim of our study was to assay the expression of K5, K6, K7, K14, K16, K17 and Lef-1 in postnatal developing teeth, and clarify the corresponding immunophenotypes of the JE and ERM. Upper jaws of Wistar rats aged from postnatal (PN) day 3.5 to PN21 were used and processed for immunohistochemistry. K5 and K14 were intensely expressed in inner enamel epithelium (IEE), reduced enamel epithelium (REE), ERM and JE. There was no staining for K16 in the tissue, except for strong staining in the oral epithelium. Specifically, at PN3.5 and PN7, K17 was initially strongly expressed and then negative in the IEE. At PN16 and PN21, both REE and ERM were strongly stained for K17, whereas K17 was negative in the JE. In addition, K6, K7 and Lef-1 were not detected in any tissue investigated. REE and ERM have an identical keratin expression pattern before eruption, while JE differs from ERM in the expression of K17 after eruption. The expression of K17 does not coincide with that of Lef-1. These data indicate that JE has a unique phenotype different from ERM, which is of odontogenic origin.

Disruption of epithelium integrity by inflammation-associated fibroblasts through prostaglandin signaling.

Inflammation-associated fibroblasts (IAFs) are associated with progression and drug resistance of chronic inflammatory diseases such as inflammatory bowel disease (IBD), but their direct impact on epithelial cells is unknown. Here, we developed an in vitro model whereby human colon fibroblasts are induced by specific cytokines and recapitulate key features of IAFs in vivo. When cocultured with patient-derived colon organoids (colonoids), IAFs induced rapid colonoid expansion and barrier disruption due to swelling and rupture of individual epithelial cells. Colonoids cocultured with IAFs also show increased DNA damage, mitotic errors, and proliferation arrest. These IAF-induced epithelial defects are mediated by a paracrine pathway involving prostaglandin E2 and its receptor EP4, leading to protein kinase A -dependent activation of the cystic fibrosis transmembrane conductance regulator. EP4-specific chemical inhibitors effectively prevented IAF-induced colonoid swelling and restored normal proliferation and genome stability. These findings reveal a mechanism by which IAFs could promote and perpetuate IBD and suggest a therapeutic avenue to mitigate inflammation-associated epithelial injury.

Blue light induced ferroptosis via STAT3/GPX4/SLC7A11/FTH1 in conjunctiva epithelium in vivo and in vitro.

The prevalence of Light-emitting diodes (LEDs) has exposed us to an excessive amount of blue light (BL) which causes various ophthalmic diseases. Previous studies have shown that conjunctiva is vulnerable to BL. In this study, we aimed to investigate the underlying mechanism of BL-induced injury in conjunctiva. We placed C57BL/6 mice and human conjunctival epithelial cell lines (HCECs) under BL (440 nm ± 15 nm, 0.2 mW/cm2) to establish a BL injury model in vivo and in vitro. Immunohistochemistry and MDA assay were used to identify lipid peroxidation (LPO) in vivo. HE staining was applied to detect morphological damage of conjunctival epithelium. DCFH-DA, C11-BODIPY 581/591, Calcein-AM, and FeRhoNox™-1 probes were performed to identify ferroptosis levels in vitro. Real-time qPCR and Western blotting techniques were employed to uncover signaling pathways of blue light-induced ferroptosis. Our findings demonstrated that BL affected tear film instability and induced conjunctival epithelium injury in vivo. Ferrostatin-1 significantly alleviated blue light-induced ferroptosis in vivo and in vitro. BL downregulates the levels of solute carrier family 7 member 11 (SLC7A11), Ferritin heavy chain (FTH1), and glutathione peroxidase (GPX4) by inhibiting the activation and translocation of the Signal transducer and activator of transcription 3 (STAT3) from inducing Fe2+ burst, ROS and LPO accumulation, ultimately resulting in ferroptosis. This study will offer new insight into BL-induced conjunctival injury and LED-induced dry eye.

Characteristics of spatial protein expression in the mouse cochlear sensory epithelia: Implications for age-related hearing loss.

Hair cells in the cochlear sensory epithelia serve as mechanosensory receptors, converting sound into neuronal signals. The basal sensory epithelia are responsible for transducing high-frequency sounds, while the apex handles low-frequency sounds. Age-related hearing loss predominantly affects hearing at high frequencies and is indicative of damage to the basal sensory epithelia. However, the precise mechanism underlying this site-selective injury remains unclear. In this study, we employed a microscale proteomics approach to examine and compare protein expression in different regions of the cochlear sensory epithelia (upper half and lower half) in 1.5-month-old (normal hearing) and 6-month-old (severe high-frequency hearing loss without hair cell loss) C57BL/6J mice. A total of 2,386 proteins were detected, and no significant differences in protein expression were detected in the upper half of the cochlear sensory epithelia between the two age groups. The expression of 20 proteins in the lower half of the cochlear sensory epithelia significantly differed between the two age groups (e.g., MATN1, MATN4, and AQP1). Moreover, there were 311 and 226 differentially expressed proteins between the upper and lower halves of the cochlear sensory epithelia in 1.5-month-old and 6-month-old mice, respectively. The expression levels of selected proteins were validated by Western blotting. These findings suggest that the spatial differences in protein expression within the cochlear sensory epithelia may play a role in determining the susceptibility of cells at different sites of the cochlea to age-related damage.

Droplet-based proteomics reveals CD36 as a marker for progenitors in mammary basal epithelium.

Deep proteomic profiling of rare cell populations has been constrained by sample input requirements. Here, we present DROPPS (droplet-based one-pot preparation for proteomic samples), an accessible low-input platform that generates high-fidelity proteomic profiles of 100-2,500 cells. By applying DROPPS within the mammary epithelium, we elucidated the connection between mitochondrial activity and clonogenicity, identifying CD36 as a marker of progenitor capacity in the basal cell compartment. We anticipate that DROPPS will accelerate biology-driven proteomic research for a multitude of rare cell populations.

Defining the contribution of Troy-positive progenitor cells to the mouse esophageal epithelium.

Progenitor cells adapt their behavior in response to tissue demands. However, the molecular mechanisms controlling esophageal progenitor decisions remain largely unknown. Here, we demonstrate the presence of a Troy (Tnfrsf19)-expressing progenitor subpopulation localized to defined regions along the mouse esophageal axis. Lineage tracing and mathematical modeling demonstrate that Troy-positive progenitor cells are prone to undergoing symmetrical fate choices and contribute to esophageal tissue homeostasis long term. Functionally, TROY inhibits progenitor proliferation and enables commitment to differentiation without affecting fate symmetry. Whereas Troy expression is stable during esophageal homeostasis, progenitor cells downregulate Troy in response to tissue stress, enabling proliferative expansion of basal cells refractory to differentiation and reestablishment of tissue homeostasis. Our results demonstrate functional, spatially restricted progenitor heterogeneity in the esophageal epithelium and identify how dynamic regulation of Troy coordinates tissue generation.

The transcription factor BMI1 increases hypoxic signaling in oral cavity epithelia.

The tongue epithelium is maintained by a proliferative basal layer. This layer contains long-lived stem cells (SCs), which produce progeny cells that move up to the surface as they differentiate. B-lymphoma Mo-MLV insertion region 1 (BMI1), a protein in mammalian Polycomb Repressive Complex 1 (PRC1) and a biomarker of oral squamous cell carcinoma, is expressed in almost all basal epithelial SCs of the tongue, and single, Bmi1-labelled SCs give rise to cells in all epithelial layers. We previously developed a transgenic mouse model (KrTB) containing a doxycycline- (dox) controlled, Tet-responsive element system to selectively overexpress Bmi1 in the tongue basal epithelial SCs. Here, we used this model to assess BMI1 actions in tongue epithelia. Genome-wide transcriptomics revealed increased levels of transcripts involved in the cellular response to hypoxia in Bmi1-overexpressing (KrTB+DOX) oral epithelia even though these mice were not subjected to hypoxia conditions. Ectopic Bmi1 expression in tongue epithelia increased the levels of hypoxia inducible factor-1 alpha (HIF1α) and HIF1α targets linked to metabolic reprogramming during hypoxia. We used chromatin immunoprecipitation (ChIP) to demonstrate that Bmi1 associates with the promoters of HIF1A and HIF1A-activator RELA (p65) in tongue epithelia. We also detected increased SC proliferation and oxidative stress in Bmi1-overexpressing tongue epithelia. Finally, using a human oral keratinocyte line (OKF6-TERT1R), we showed that ectopic BMI1 overexpression decreases the oxygen consumption rate while increasing the extracellular acidification rate, indicative of elevated glycolysis. Thus, our data demonstrate that high BMI1 expression drives hypoxic signaling, including metabolic reprogramming, in normal oral cavity epithelia.

Exposure to PM2.5 modulate the pro-inflammatory and interferon responses against influenza virus infection in a human 3D bronchial epithelium model.

Epidemiological studies showed a positive association between exposure to PM2.5 and the severity of influenza virus infection. However, the mechanisms by which PM2.5 can disrupt antiviral defence are still unclear. From this perspective, the objective of this study was to evaluate the effects of PM2.5 on antiviral signalling in the respiratory epithelium using the bronchial Calu-3 cell line grown at the air-liquid interface. Pre-exposure to PM2.5 before infection with the influenza virus was investigated, as well as a co-exposure. Although a physical interaction between the virus and the particles seems possible, no effect of PM2.5 on viral replication was observed during co-exposure, although a downregulation of IFN-ß release was associated to PM2.5 exposure. However, pre-exposure slightly increased the viral nucleoprotein production and the pro-inflammatory response. Conversely, the level of the myxovirus resistance protein A (MxA), an interferon-stimulated gene (ISG) induced by IFN-ß, was reduced. Therefore, these results suggest that pre-exposure to PM2.5 could alter the antiviral response of bronchial epithelial cells, increasing their susceptibility to viral infection.

Interferon lambda in respiratory viral infection: immunomodulatory functions and antiviral effects in epithelium.

Type III interferon (IFN-λ), a new member of the IFN family, was initially considered to possess antiviral functions similar to those of type I interferon, both of which are induced via the JAK/STAT pathway. Nevertheless, recent findings demonstrated that IFN-λ exerts a nonredundant antiviral function at the mucosal surface, preferentially produced in epithelial cells in contrast to type I interferon, and its function cannot be replaced by type I interferon. This review summarizes recent studies showing that IFN-λ inhibits the spread of viruses from the cell surface to the body. Further studies have found that the role of IFN-λ is not only limited to the abovementioned functions, but it can also can exert direct and/or indirect effects on immune cells in virus-induced inflammation. This review focuses on the antiviral activity of IFN-λ in the mucosal epithelial cells and its action on immune cells and summarizes the pathways by which IFN-λ exerts its action and differentiates it from other interferons in terms of mechanism. Finally, we conclude that IFN-λ is a potent epidermal antiviral factor that enhances the respiratory mucosal immune response and has excellent therapeutic potential in combating respiratory viral infections.

MAP3K1 regulates female reproductive tract development.

Mitogen-activated protein 3 kinase 1 (MAP3K1) has a plethora of cell type-specific functions not yet fully understood. Herein, we describe a role for MAP3K1 in female reproductive tract (FRT) development. MAP3K1 kinase domain-deficient female mice exhibited an imperforate vagina, labor failure and infertility. These defects corresponded with shunted Müllerian ducts (MDs), the embryonic precursors of FRT, that manifested as a contorted caudal vagina and abrogated vaginal-urogenital sinus fusion in neonates. The MAP3K1 kinase domain is required for optimal activation of the Jun-N-terminal kinase (JNK) and cell polarity in the MD epithelium, and for upregulation of WNT signaling in the mesenchyme surrounding the caudal MD. The MAP3K1-deficient epithelial cells and MD epithelium had reduced expression of WNT7B ligands. Correspondingly, conditioned media derived from MAP3K1-competent, but not -deficient, epithelial cells activated a TCF/Lef-luciferase reporter in fibroblasts. These observations indicate that MAP3K1 regulates MD caudal elongation and FRT development, in part through the induction of paracrine factors in the epithelium that trans-activate WNT signaling in the mesenchyme.

Computational Models of Claudin Assembly in Tight Junctions and Strand Properties.

Claudins are one of the major components of tight junctions (TJs) that polymerize within the cell membrane and form interactions between cells. Some claudins seal the paracellular space, limiting paracellular flux, while others form selectively permeable ion channels that control the paracellular permeability of small ions. Claudin strands are known to be dynamic and reshape within TJs to accommodate large-scale movements and rearrangements of epithelial tissues. Here, we summarize the recent computational and modeling studies on claudin assembly into tetrameric ion channels and their polymerization into µm long strands within the membrane. Computational studies ranging from all-atom molecular dynamics, coarse-grained simulations, and hybrid-resolution simulations elucidate the molecular nature of claudin assembly and function and provide a framework that describes the lateral flexibility of claudin strands.

Mechanosensitive TRPV4 channel guides maturation and organization of the bilayered mammary epithelium.

Biophysical cues from the cell microenvironment are detected by mechanosensitive components at the cell surface. Such machineries convert physical information into biochemical signaling cascades within cells, subsequently leading to various cellular responses in a stimulus-dependent manner. At the surface of extracellular environment and cell cytoplasm exist several ion channel families that are activated by mechanical signals to direct intracellular events. One of such channel is formed by transient receptor potential cation channel subfamily V member, TRPV4 that is known to act as a mechanosensor in wide variaty of tissues and control ion-influx in a spatio-temporal way. Here we report that TRPV4 is prominently expressed in the stem/progenitor cell populations of the mammary epithelium and seems important for the lineage-specific differentiation, consequently affecting mechanical features of the mature mammary epithelium. This was evident by the lack of several markers for mature myoepithelial and luminal epithelial cells in TRPV4-depleted cell lines. Interestingly, TRPV4 expression is controlled in a tension-dependent manner and it also impacts differentation process dependently on the stiffness of the microenvironment. Furthermore, such cells in a 3D compartment were disabled to maintain normal mammosphere structures and displayed abnormal lumen formation, size of the structures and disrupted cellular junctions. Mechanosensitive TRPV4 channel therefore act as critical player in the homeostasis of normal mammary epithelium through sensing the physical environment and guiding accordingly differentiation and structural organization of the bilayered mammary epithelium.

Bile acid metabolites enhance expression of cathelicidin antimicrobial peptide in airway epithelium through activation of the TGR5-ERK1/2 pathway.

Signals for the maintenance of epithelial homeostasis are provided in part by commensal bacteria metabolites, that promote tissue homeostasis in the gut and remote organs as microbiota metabolites enter the bloodstream. In our study, we investigated the effects of bile acid metabolites, 3-oxolithocholic acid (3-oxoLCA), alloisolithocholic acid (AILCA) and isolithocholic acid (ILCA) produced from lithocholic acid (LCA) by microbiota, on the regulation of innate immune responses connected to the expression of host defense peptide cathelicidin in lung epithelial cells. The bile acid metabolites enhanced expression of cathelicidin at low concentrations in human bronchial epithelial cell line BCi-NS1.1 and primary bronchial/tracheal cells (HBEpC), indicating physiological relevance for modulation of innate immunity in airway epithelium by bile acid metabolites. Our study concentrated on deciphering signaling pathways regulating expression of human cathelicidin, revealing that LCA and 3-oxoLCA activate the surface G protein-coupled bile acid receptor 1 (TGR5, Takeda-G-protein-receptor-5)-extracellular signal-regulated kinase (ERK1/2) cascade, rather than the nuclear receptors, aryl hydrocarbon receptor, farnesoid X receptor and vitamin D3 receptor in bronchial epithelium. Overall, our study provides new insights into the modulation of innate immune responses by microbiota bile acid metabolites in the gut-lung axis, highlighting the differences in epithelial responses between different tissues.