Activation of TRPM8 promotes K+ secretion in rat epididymal epithelium.

Intraepithelial lymphocytosis is a hallmark of microscopic colitis commonly presenting with debilitating chronic watery diarrhea. The exact pathophysiology remains unclear but is believed to involve an abnormal immune response to the luminal microenvironment in genetically predisposed individuals. Medications like nonsteroidal anti-inflammatory drugs, proton pump inhibitors, and antidepressants, as well as alcohol consumption and smoking have been regarded as risk factors for microscopic colitis. Lymphocytic colitis often responds well to anti-inflammatory treatment, with budesonide being a first line therapy. Relapse is common after discontinuing budesonide, often necessitating low-dose maintenance therapy. Advanced therapies including anti-TNF-alpha (e.g. infliximab) or anti-integrin (e.g. vedolizumab) agents as well as JAK inhibitors (e.g. upadacitinib) may be considered in refractory cases. However, confirmation of the diagnosis and ruling out other etiologies are necessary before intensifying treatment. Here, we present the case of a 78-year-old female patient with a twenty month history of chronic refractory diarrhea, repeatedly diagnosed as microscopic lymphocytic colitis. Following further deterioration and the development of hepatosplenomegaly with atypical liver lesions, a review of colonic biopsies and additional work-up led to the correct diagnosis.

There is an intricate relationship between pig nutrition and gut microbiome. For technical, cost and societal reasons, in vitro gut models can be advantageously used as an alternative to in vivo experiments to perform mechanistic studies on the interactions between gut microbes and nutrients. In this context, the aim of the present study was to develop a new in vitro model of the healthy adult pig colon integrating both luminal and mucosal microenvironments. The model was further used to evaluate feed ingredients (lactose and lactose + sucrose). Gut Microbiota composition and metabolic activities were followed by 16S Metabarcoding and short chain fatty acid (SCFA) / gas measurement, respectively. Then, the effect of the both feedstuffs on skatole production, responsible for boar taint in male pigs, was also investigated. Based on in vivo data, the new MPigut-IVM was set-up to reproduce the main physicochemical (pH, transit time, self-maintained anaerobiosis), nutritional (composition of ileal effluents, bile salts) and microbial (lumen and mucus-associated microbiota) parameters of the large intestine in healthy adult pigs. The model was validated through in vitro-in vivo comparisons regarding SCFA concentrations and bacterial profiles at the phylum and family levels. Lactose and lactose + sucrose had no significant impact on SCFAs but increased gas production (P < 0.01 with lactose). Both sugars, particularly lactose + sucrose, tended to reduce skatole concentrations while increasing indole levels (P > 0.05). This was associated to a slight reduction of the numbers of skatole-producing bacteria Olsenella scatoligenes (P > 0.05). Both feed ingredients induced a decrease in bacterial α-diversity (P < 0.05). Despite obvious limitations such as lack of host interactions, the adult MPigut-IVM represents a powerful platform for Microbiome studies in the pig colonic environment. In Line with the 3R regulations, this in vitro model can be useful to perform preliminary screening of innovative feed strategies to improve pig health and help to elucidate their mechanisms of action in relation with gut microbiota, taking into account inter-individual variabilities.

In 1998, Arlette Darfeuille-Michaud, Christel Neut and Jean-Frederic Colombel discovered a novel pathovar of Escherichia coli, adherent and invasive Escherichia coli (AIEC), in the ileum of patients with Crohn's disease (CD), that was genetically distinct from diarrheagenic E. coli, could adhere to and invade intestinal epithelial cells and survive in macrophages. The consistent association between AIEC and CD (approximately 30% across the world), their ability to exploit CD-associated genetic traits, and virulence in preclinical colitis models but not healthy hosts spurred global research to elucidate their pathogenicity. Research focused on integrating AIEC with the microbiome, metabolome, metagenome, host response and the impact of diet and antimicrobials has linked the luminal microenvironment and AIEC metabolism to health and disease. This deeper understanding has led to therapeutic trials and precision medicine targeting AIEC-colonised patients. In November 2023, prominent members of the AIEC research community met to present and discuss the many facets of basic, translational and clinical AIEC fields at 'AIEC: past, present and future' in NYC. This review is a summary of this international meeting highlighting the history of AIEC, knowledge accumulated over the past 25 years about its pathogenic properties and proposes a standardised approach for screening patients for AIEC.

The female reproductive organs present with the earliest aging characteristics, such as a decline in fertility and estrous cyclicity. While age-related changes in the ovary are well documented, it is unclear if any age-associated changes occur in the other female reproductive organs, such as the oviduct/Fallopian tube. At the distal end of aged oviducts in mice, we found vacuolated multi-ciliated cells (MCCs) with a severely apically displaced and deformed nucleus. This phenotype was unique to the distal oviduct epithelium-the infundibulum (INF) and ampulla (AMP). Ovariectomy did not affect the timeline of MCC vacuolation, suggesting little involvement of ovulation and hormonal regulation. MCC vacuolation was induced in hypoxia or hydroxyurea treatments in invitro organotypic culture of all oviduct regions, not limited to the INF/AMP epithelium. This suggests a high oxygen demand in MCCs, compared to other cell types, and a uniquely stressed INF/AMP epithelial microenvironment invivo. We found that the blood circulation of INF/AMP depended on the ovarian artery, different from the rest of the oviduct epithelium, and its circulation declined along with ovarian activities. We conclude that a decline in local blood circulation and distinct cellular identity of the INF/AMP epithelium caused age-associated MCC vacuolation, reflecting its mild, chronically stressed microenvironment.

Diabetes mellitus has increased significantly over the past decades. This disease affects the reproductive competence of diabetic men by disrupting spermatogenesis, fertility potential, penile erection, and ejaculation. However, hyperglycemic conditions' effects on the epididymis remain elusive despite its importance for sperm maturation. We aimed to determine the effects of diabetes on the epididymis, using qualitative (systematic review) and quantitative (meta-analysis) approaches, to address the question: Can diabetes disrupt epididymal structure and function? We performed an extensive literature search identifying 66 eligible studies through PubMed, Web of Science, and Embase databases. Outcomes extracted from the studies included alterations in epididymal cell metabolism and morphology under hyperglycemic conditions. Pre-clinical studies published in murine were evaluated under a meta-analytical approach, whereas clinical investigations in humans were analyzed qualitatively (PROSPERO number is CRD42020208658). Type 1 diabetic patients presented post-ejaculatory epididymal hypotonia/atonia, whereas type 1 and 2 diabetic patients exhibited perturbation in the epididymal advanced glycation end-product axis. In murine, high glucose levels disturb the metabolism of epididymal cells, the androgenic profile, and the expression of hormone receptors within the organ. The low activity of antioxidant enzymes promoted an elevation of oxidative metabolite levels, creating a pro-oxidant microenvironment toxic to spermatozoa. All these deleterious mechanisms of diabetes trigger molecular and biochemical responses contributing to the deterioration of epididymis structure and function. Our data indicated that diabetes may affect epididymis morphology and function through hormonal imbalance, glucose metabolism disturbance, and oxidative stress generation. These mechanisms may alter the luminal microenvironment and epithelial function, impairing organ functionality with consequences for sperm maturation. This review also highlighted several points that need investigation by further studies associating diabetes and epididymis to fill the knowledge gaps better.

Life relies on redox reactions to sustain; however, imbalanced redox exerts stress on living beings. This is particularly pivotal as mammalian gamete sperm cells undergo functional maturation in the epididymis, preparing themselves for the long and challenging journey to begin a new life by successfully fertilizing an oocyte. Defects in epididymal sperm maturation are closely related to male infertility and reproductive health. In the epididymis, the halted translational machinery in spermatozoa while maturing in a quiescent state prompts them to immensely depend on epithelial cells for nutritional supports and information exchange. Extensive intercellular communication is therefore required between sperm and epithelial cells, and extracellular vesicles (EVs) play a crucial role as intercellular carriers. Epididymal luminal microenvironment is spatially specialized to be an acidic, decalcitonic, and pro-reductive protective milieu to prevent premature sperm dysfunction. The uniqueness of epididymal microenvironment also requires the EVs to have adaptive characteristics. Although the biogenesis and compositions of EVs in the epididymis have been reviewed elsewhere, this chapter discusses the potential role of EVs from the perspective of cellular nutrient balance of vitamins and minerals, redox metabolism, and intercellular interplay during sperm maturation to better understand the biology of epididymis in male reproduction.

Despite the huge pool of ideas on how diet can be manipulated to ameliorate or prevent illnesses, our understanding of how specific changes in diet influence the gastrointestinal tract is limited. This review aims to describe two innovative investigative techniques that are helping lift the veil of mystery about the workings of the gut. First, the gas-sensing capsule is a telemetric swallowable device that provides unique information on gastric physiology, small intestinal microbial activity, and fermentative patterns in the colon. Its ability to accurately measure regional and whole-gut transit times in ambulant humans has been confirmed. Luminal concentrations of hydrogen and carbon dioxide are measured by sampling through the gastrointestinal tract, and such application has enabled mapping of the relative amounts of fermentation of carbohydrates in proximal-versus-distal colon after manipulation of the types and amounts of dietary fiber. Second, changes in the smell of feces, via analysis of volatile organic compounds, occur in response to the diet, and by the presence and therapy of irritable bowel syndrome and inflammatory bowel disease. Such information is likely to aid our understanding of what dietary change can do to the colonic luminal microenvironment, and may value-add to diagnosis and therapeutic design. In conclusion, such methodologies enable a more complete physiological profile of the gastrointestinal tract to be created. Systematic description in various cohorts and effects of dietary interventions, particularly when co-ordinated with the analysis of microbiome, are needed.

As a component of the nucleosome remodeling and deacetylating (NuRD) complex, metastasis-associated protein 1 (MTA1) has been reported to be abundant in male reproductive system and might participate in spermatogenesis and sperm maturation, whereas the precise functional role of MTA1 in these processes is still undetermined. To investigate the effect and potential function of MTA1 in male fertility. Mta1 knockout mice (Mta1-/-) were employed to detect their reproductive phenotype. The pH value of Mta1-/- epididymal luminal fluid was measured, and the potential mechanism of MTA1 involved in regulating luminal acidification was detected in vivo and in vitro. A vasectomy model with abnormal pH of epididymal lumen was established to further detect the effect of MTA1 on epididymal luminal microenvironment. Mta1-/- mice were fertile without any detectable defects in spermatogenesis or sperm motility while the deficiency of MTA1 could acidify the initial segment of epididymis to a certain extent. MTA1 could interact with estrogen receptor alpha (ERα) and inhibit the transcription of ERα target gene, hydrogen exchanger 3 (NHE3), and ultimately affect the epididymal luminal milieu. After vasectomy, the Mta1-/- mice presented a more acidic epididymal lumen which was closer to the normal state compared to the wild-type model. MTA1 is dispensable for male fertility in mice, but plays a potentially important function in regulating luminal acidification of the epididymis.

Functional delineation of the luminal epithelial microenvironment in breast using cell-based screening in combinatorial microenvironments.

Mitochondrial-rich cells (MRCs) are one of the most significant canceled type of epithelial cells. Morphologically these cells are totally different from other epithelial cells. These cells primarily implicated in sea-water and fresh-water adaptation, and acid-base regulation. However, in this review paper, we explored some of the most intriguing biological and immune-related functional developmental networks of MRCs. The main pinpoint, MRCs perform a dynamic osmoregulatory and immunological functional role in the gut and male reproductive system. The Na+/K+_ATPase (NKA) and Na+/K+/2Cl cotransporter (NKCC) are key acidifying proteins of MRCs for the ion-transporting function for intestinal homeostasis and maintenance of acidifying the luminal microenvironment in the male reproductive system. Further more importantly, MRCs play a novel immunological role through the exocrine secretion of nano-scale exosomes and multivesicular bodies (MVBs) pathway, which is very essential for sperm maturation, motility, acrosome reaction, and male sex hormones, and these an essential events to produce male gametes with optimal fertilizing ability. This effort is expected to promote the novel immunological role of MRCs, which might be essential for nano-scale exosome secretion.

Residing between the testes and the vas deferens, the epididymis is a highly convoluted tubule whose unique luminal microenvironment is crucial for the functional maturation of spermatozoa. This microenvironment is created by the combined secretory and resorptive activity of the lining epididymal epithelium, including the release of extracellular vesicles (epididymosomes), which encapsulate fertility modulating proteins and a myriad of small non-coding RNAs (sncRNAs) that are destined for delivery to recipient sperm cells. To enable investigation of this intercellular communication nexus, we have previously developed an immortalized mouse caput epididymal epithelial cell line (mECap18). Here, we describe the application of label-free mass spectrometry to characterize the mECap18 cell proteome and compare this to the proteome of native mouse caput epididymal epithelial cells. We report the identification of 5,313 mECap18 proteins, as many as 75.8% of which were also identified in caput epithelial cells wherein they mapped to broadly similar protein classification groupings. Furthermore, key pathways associated with protein synthesis (e.g., EIF2 signaling) and cellular protection in the male reproductive tract (e.g., sirtuin signaling) were enriched in both proteomes. This comparison supports the utility of the mECap18 cell line as a tractable in-vitro model for studying caput epididymal epithelial cell function.

Conserved in female reproduction across all mammalian species is the estrous cycle and its regulation by the hypothalamic-pituitary-gonadal (HPG) axis, a collective of intersected hormonal events that are crucial for ensuring uterine fertility. Nonetheless, knowledge of the direct mediators that synchronously shape the uterine microenvironment for successive yet distinct events, such as the transit of sperm and support for progressive stages of preimplantation embryo development, remain principally deficient. Toward understanding the timed endometrial outputs that permit luminal events as directed by the estrous cycle, we used Bovidae as a model system to uniquely surface sample and study temporal shifts to in vivo endometrial transcripts that encode for proteins destined to be secreted. The results revealed the full quantitative profile of endometrial components that shape the uterine luminal microenvironment at distinct phases of the estrous cycle (estrus, metestrus, diestrus, and proestrus). In interpreting this comprehensive log of stage-specific endometrial secretions, we define the "uterine secretory cycle" and extract a predictive understanding of recurring physiological actions regulated within the uterine lumen in anticipation of sperm and preimplantation embryonic stages. This repetitive microenvironmental preparedness to sequentially provide operative support was a stable intrinsic framework, with only limited responses to sperm or embryos if encountered in the lumen within the cyclic time period. In uncovering the secretory cycle and unraveling realistic biological processes, we present novel foundational knowledge of terminal effectors controlled by the HPG axis to direct a recurring sequence of vital functions within the uterine lumen.NEW & NOTEWORTHY This study unravels the recurring sequence of changes within the uterus that supports vital functions (sperm transit and development of preimplantation embryonic stages) during the reproductive cycle in female Ruminantia. These data present new systems knowledge in uterine reproductive physiology crucial for setting up in vitro biomimicry and artificial environments for assisted reproduction technologies for a range of mammalian species.

Previous in vitro studies have shown that the intestinal luminal content, including metabolites, possibly regulates epithelial layer responses to harmful stimuli and promotes disease. Therefore, we aimed to test the hypothesis that fecal supernatants from patients with colon cancer (CC), ulcerative colitis (UC) and irritable bowel syndrome (IBS) contain distinct metabolite profiles and establish their effects on Caco-2 cells and human-derived colon organoids (colonoids). The metabolite profiles of fecal supernatants were analyzed by liquid chromatography–mass spectrometry and distinguished patients with CC (n = 6), UC (n = 6), IBS (n = 6) and healthy subjects (n = 6). Caco-2 monolayers and human apical-out colonoids underwent stimulation with fecal supernatants from different patient groups and healthy subjects. Their addition did not impair monolayer integrity, as measured by transepithelial electrical resistance; however, fecal supernatants from different patient groups and healthy subjects altered the gene expression of Caco-2 monolayers, as well as colonoid cultures. In conclusion, the stimulation of Caco-2 cells and colonoids with fecal supernatants derived from CC, UC and IBS patients altered gene expression profiles, potentially reflecting the luminal microenvironment of the fecal sample donor. This experimental approach allows for investigating the crosstalk at the gut barrier and the effects of the gut microenvironment in the pathogenesis of intestinal diseases.

The stomach is notorious for having the steepest proton gradient in the body, which pathogenic Helicobacter pylori must traverse to infect the epithelium. This gradient across the 200‐300 μm‐thick gastric mucus layer enables the stomach to maintain a near‐neutral pH at the epithelial surface and a strongly acidic environment in the stomach lumen. In this study, we use human gastric organoids to study the maintenance and regulation of this pH gradient. Human gastric organoids are 3D cellular constructs derived from patient biopsies that mimic the microanatomy and physiology of the stomach, as they contain both mucus and gastric acid. Organoids have been used to investigate H. pylori‐induced gastritis, peptic ulcers, and stomach cancer, which claims 700,000 lives each year. Gastric mucus is critical for protection against H. pylori infection, which is the leading cause of gastric cancer. It is presently unclear how the gastric mucosal pH gradient is maintained, and how closely organoid models can mimic this microenvironment. First, we detected mucus within human gastric organoids using histological staining and bead microrheology. We also used a FITC‐conjugated lectin to visualize the mucosal matrix inside of the organoids. These data were supported by our observation that a clear viscous material was produced on the apical side of two‐dimensional, organoid‐derived cell monolayers. Additionally, we developed a novel method for the measurement of intraluminal pH of the gastric organoids. Using a micromanipulator and stereomicroscope to maneuver a glass microelectrode into the organoid lumen, we measured and manipulated luminal pH using histamine stimulation. We determined that pH remains consistent across organoid lines, and that the intraluminal space is more acidic than the surrounding environment. In ongoing studies, we will develop the spatial resolution of our organoid microprofiling technique to fully characterize the proton gradient within the mucus‐filled lumens gastric organoids. Our studies are the first to utilize pH microsensors in these models, and will lead to improved molecular and biophysical understanding of gastric physiology and potential new treatment approaches for disease conditions (such as peptic ulcers and gastric cancer) that involve dysregulated gastric acid secretion.

In the male reproductive tract, the epididymis is an essential organ for sperm maturation, in which sperm cells acquire mobility and the ability to fertilize oocytes while being stored in a protective microenvironment. Epididymal function involves a specialized luminal microenvironment established by the epithelial cells of epididymal mucosa. Low-calcium concentration is a unique feature of this epididymal luminal microenvironment, its relevance and regulation are, however, incompletely understood. In the rat epididymis, the vitamin D-related calcium-dependent TRPV6-TMEM16A channel-coupler has been shown to be involved in fluid transport, and, in a spatially complementary manner, vitamin K2-related γ-glutamyl carboxylase (GGCX)-dependent carboxylation of matrix Gla protein (MGP) plays an essential role in promoting calcium-dependent protein aggregation. An SNP in the human GGCX gene has been associated with asthenozoospermia. In addition, bioinformatic analysis also suggests the involvement of a vitamin B6-axis in calcium-dependent MGP-mediated protein aggregation. These findings suggest that vitamins interact with calcium homeostasis in the epididymis to ensure proper sperm maturation and male fertility. This review article discusses the regulation mechanisms of calcium homeostasis in the epididymis, and the potential role of vitamin interactions on epididymal calcium homeostasis, especially the role of matrix calcium in the epididymal lumen as a cofactor for the carboxylated MGP-mediated scavenging function.

The spermatozoa become mature in the epididymis which is divided into initial segment and caput, corpus, and cauda epididymis. The water movement across the epididymal epithelium is important for creating luminal microenvironment for sperm maturation. Aquaporins (Aqps) are water channel proteins, and expression of Aqps is regulated by androgens. The current research was focused to examine expressional regulation of Aqp1 and Aqp9 by an androgenic-anabolic steroid, nandrolone decanoate (ND). The ND at the low dose (2 mg/ kg body weight/week) or high dose (10 mg) was subcutaneously administrated into male rats for 2 or 12 weeks. Transcript levels of Aqp1 and Aqp9 were determined by quantitative real-time polymerase chain reaction (PCR) analyses. In the initial segment, level of Aqp1 was decreased with 12 week-treatment, while Aqp9 level was decreased by the high dose treatment for 12 weeks. In the caput epididymis, Aqp9 expression was decreased by the low dose treatment. The 2 week-treatment resulted in an increase of Aqp1 level but a decrease of Aqp9 expression in the corpus epididymis. In the corpus epididymis, the 12 week-treatment at the low dose caused the reduction of Aqp1 and Aqp9 levels, but the high dose treatment resulted in an increase of Aqp1 expression and a decrease of Aqp9 level. In the cauda epididymis, Aqp1 expression was decreased by 2 and 12 week-treatments, while increases of Aqp9 levels was detected with the high dose treatment for 2 weeks and with 12 week-treatment. These findings indicate differential regulation of Aqp1 and Aqp9 expression among epididymal segments by ND.

Spermatozoa released from testes undergo a maturation process and acquire the capacity to fertilize ova through epididymal transit. The epididymis is divided into four regions, each with unique morphology, gene profile, luminal microenvironment and distinct function. To study the functions of relevant genes in the epididymal initial segment (IS), a novel IS-specific mouse model, Lcn9-Cre knock-in (KI) mouse line was generated via CRISPR/Cas9 technology. The TAG stop codon was replaced by a 2A-NLS-Cre cassette, resulting in the co-expression of Lcn9 and Cre recombinase. IS-specific Cre expression was first observed from postnatal day 17. Using the Rosa26tdTomato reporter mice, the Cre-mediated DNA recombination was detected exclusively in principal cells. The epididymal IS-specific Cre activity in vivo was further confirmed using Lcn9-Cre mice crossed with a mouse strain carrying Tsc1 floxed alleles (Tsc1flox/+). Cre expression did not affect either normal development or male fecundity. Different from any epididymis-specific Cre mice reported previously, the novel Lcn9-Cre mouse line can be used to introduce entire IS-specific conditional gene editing for gene functional study.

The gut epithelium is a mechanical barrier that protects the host from the luminal microenvironment and interacts with the gut microflora, which influences the development and progression of ulcerative colitis (UC). Licochalcone A (LA) exerts anti-inflammatory effects against UC; however, whether it also regulates both the gut barrier and microbiota during colitis is unknown. The current study was conducted to reveal the regulatory effects of LA on the intestinal epithelium and gut microflora in C57BL/6 mice subjected to dextran sodium sulfate (DSS). Sulfasalazine (SASP) was used as the positive control. Results of clinical symptoms evaluation, hematoxylin, and eosin (H&E) staining, and enzyme-linked immunosorbent (ELISA) assays showed that LA significantly inhibited DSS-induced weight loss, disease activity index (DAI) increase, histological damage, and gut inflammation. Additionally, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and immunohistochemical (IHC) analysis showed that LA maintained the integrity of the intestinal barrier by suppressing cell apoptosis and preserving the expression of tight junction (TJ) proteins. Notably, the optimal dose of LA for gut barrier preservation was low, while that for anti-inflammatory effects was high, indicating that LA might preserve gut barrier integrity via direct effects on the epithelial cells (ECs) and TJ proteins. Furthermore, 16S rRNA analysis suggested that the regulatory effect of LA on the gut microbiota differed distinctly according to dose. Correlation analysis indicated that a low dose of LA significantly modulated the intestinal barrier-associated bacteria as compared with a moderate or high dose of LA. Western blot (WB) analysis indicated that LA exhibited anti-UC activity partly by blocking the mitogen-activated protein kinase (MAPK) pathway. Our results further elucidate the pharmacological activity of LA against UC and will provide valuable information for future studies regarding on the regulatory effects of LA on enteric diseases.

Molecular perspective concerning fluoride and arsenic mediated disorders on epididymal maturation of spermatozoa: A concise review.