Luminal Fluid Research Articles

Hypertonic water reabsorption with a parallel-current system via the glandular and saccular renal tubules of Ruditapes philippinarum.

We investigated the renal function of the brackish water clam, Ruditapes philippinarum, employing magnetic resonance imaging (MRI). The R. philippinarum kidney consists of two renal tubules, a glandular (GT) and a saccular (ST) tubule. After exposure to seawater containing manganese ion (Mn2+) at 20°C, the intensity of the T1-weighted MRI and longitudinal relaxation rates (1/T1=R1) of the kidney were increased. In the ST, haemolymph containing Mn2+ entered directly from the auricle, and the Mn2+ concentration ([Mn2+]) increased in the initial part of the ST. Thereafter, [Mn2+] was almost constant until the posterior end of the kidney. The GT received haemolymph from the pedal sinus via the visceral sinus. The GT runs parallel inside the ST, and [Mn2+] increased progressively until it merged with the ST. In a range of seawater with [Mn2+] from 1 to 30 µmoll-1, the [Mn2+] increased 12-fold in the posterior part of the ST, compared with the ambient [Mn2+]. Based on these results, the epithelium of the initial part of the ST reabsorbs water from luminal fluid, building up a higher osmotic pressure. Using this osmotic gradient, hypertonic water is reabsorbed via the epithelium of the GT to the ST, and then transferred to the haemolymph via the epithelium of the ST. Excess water is excreted as urine. This model was supported by the increases in [Mn2+] in the ST when the clams were exposed to seawater containing Mn2+ at salinity from 26.0 to 36.0‰, showing that the parallel-current system works in hypotonic seawater.

Effects of intrauterine extracellular vesicle microRNAs on embryonic gene expression in low-fertility cows.

Embryo survival and pre-implantation development depend on uterine luminal fluid, which is believed to play a role in early embryonic death and infertility in cows. Extracellular vesicles (EVs) in the uterine luminal fluid contain microRNAs (miRNAs), crucial mediators of intercellular communication. miRNAs regulate conceptus-maternal interactions and participate in embryonic development by suppressing gene expression. Therefore, we hypothesized that miRNAs in the intrauterine EVs of low-fertility cows would hinder embryonic survival and development. EVs were collected from the bovine uterine luminal fluid of both normal- and low-fertility cows 7 days post-estrus. Small RNA-sequencing analysis of miRNAs isolated from these EVs identified eight miRNAs that were highly expressed in normal-fertility cows (normal-fertility miRNAs) and eight with elevated expression in low-fertility cows (low-fertility miRNAs). These two sets of miRNAs were transfected into hatched blastocysts via lipofection. RNA-seq following lipofection with low-fertility miRNAs identified 424 differentially expressed genes (DEGs) relative to the control; in contrast, following lipofection with normal-fertility miRNAs, seven DEGs were identified. Pathway analysis of the DEGs identified following lipofection with low-fertility miRNAs revealed substantial enrichment of mitogen-activated protein kinase (MAPK) signaling. Expression of activator protein 1 (AP1) and interferon-tau (IFNT) mRNA was significantly lower in the low-fertility miRNA transfection group than in the control. IFNT is essential for maternal pregnancy recognition. Therefore, miRNAs in intrauterine EVs from low-fertility cows at 7 days post-estrus may inhibit embryo development and suppress IFNT expression by altering MAPK signaling.

Fructose metabolism is unregulated in cancers and placentae.

Fructose and lactate are present in high concentrations in uterine luminal fluid, fetal fluids and fetal blood of ungulates and cetaceans, but their roles have been ignored and they have been considered waste products of pregnancy. This review provides evidence for key roles of both fructose and lactate in support of key metabolic pathways required for growth and development of fetal-placental tissues, implantation and placentation. The uterus and placenta of ungulates convert glucose to fructose via the polyol pathway. Fructose is sequestered within the uterus and cannot be transported back into the maternal circulation. Fructose is phosphorylated by ketohexokinase to fructose-1-PO4 (F1P) by that is metabolized via the fructolysis pathway to yield dihydoxyacetone phosphate and glyceraldehyde-3-PO4 that are downstream of phosphofructokinase. Thus, there is no inhibition of the fructolysis pathway by low pH, citrate or ATP which allows F1P to continuously generate substrates for the pentose cycle, hexosamine biosynthesis pathway, one-carbon metabolism and tricarboxylic acid cycle, as well as lactate. Lactate sustains the activity of hypoxia-inducible factor alpha and its downstream targets such as vascular endothelial growth factor to increase utero-placental blood flow critical to growth and development of the fetal-placental tissues and a successful outcome of pregnancy. Pregnancy has been referred to as a controlled cancer and this review addresses similarities regarding metabolic aspects of tumors and the placenta.

Taurine is essential for mouse uterine luminal fluid resorption during implantation window via the SCNN1A and AQP8 signaling.

Uterine fluid homeostasis during peri-implantation is crucial for successful embryo implantation. Taurine (Tau) plays a crucial role in regulating osmotic pressure and ion transport. However, the precise mechanisms underlying Tau-mediated regulation of uterine fluid homeostasis during peri-implantation in mice remain unclear. In this study, we generated a Tau-deficient mouse model by administering Tau-free diet to Csad knockout (Csad-/-) mice to block endogenous Tau synthesis and exogenous Tau absorption (Csad-/--Tau free). Our findings demonstrated that Csad-/-Tau free mice with diminished level of Tau exhibited decreased rates of embryo implantation and impaired fertility. Further analysis revealed that the expression of Scnn1a was down-regulated during the implantation window, while Aqp8 was upregulated in Csad-/-Tau free mice, leading to uterine luminal fluid retention and defects in luminal closure, resulting in failed embryo implantation. Additionally, it was also found that E2 inhibited uterine Csad expression and Tau synthesis, while P4 promoted them. Therefore, our findings suggest that ovarian steroid hormones regulate Csad expression and Tau synthesis, thereby affecting release and resorption of uterine luminal fluid, ultimately impacting embryo implantation success.

Epididymis-specific RNase A family genes regulate fertility and small RNA processing

Sperm small RNAs are implicated in intergenerational transmission of paternal environmental effects. Small RNAs generated by cleavage of tRNAs, known as tRNA fragments (tRFs) or tRNA-derived RNAs (tDRs or tsRNAs), are an abundant class of RNAs in mature sperm and can be modulated by environmental conditions. The biogenesis of tRFs in the male reproductive tract remains poorly understood. Angiogenin, a member of the Ribonuclease A superfamily (RNase A), cleaves tRNAs to generate tRFs in response to cellular stress. Four paralogs of Angiogenin, namely Rnase9, Rnase10, Rnase11, and Rnase12, are specifically expressed in the epididymis -a long, convoluted tubule where sperm mature and acquire fertility and motility. Here, by generating mice deleted for all four genes (Rnase9-12-/-, termed "KO" for Knock Out), we report that these genes regulate fertility and small RNA levels. KO male mice are sterile; KO sperm fertilized oocytes in vitro but failed to efficiently fertilize oocytes in vivo due to an inability of sperm to pass through the utero-tubular junction. Intriguingly, there were decreased levels of fragments of tRNAs (tRFs) and rRNAs (rRNA-derived small RNAs or rsRNAs) in the KO epididymis and epididymal luminal fluid, although RNases 9-12 did not show ribonucleolytic activity in-vitro. Importantly, KO sperm showed a dramatic decrease in the levels of tRFs, demonstrating a role of epididymis-specific Rnase9-12 genes in regulating sperm small RNA composition. Together, our results reveal an unexpected role of four epididymis-specific non-canonical RNase A family genes in regulating fertility and small RNA processing.

Dissolution, phase behavior and mass transport of amorphous solid dispersions in aspirated human intestinal fluids

Amorphous solid dispersions (ASDs) typically show improved dissolution and generate supersaturated solutions, enhancing the oral bioavailability of poorly soluble drugs. To gain insights into intraluminal ASD behavior, we utilized two poorly soluble drugs with different crystallization tendencies, atazanavir and posaconazole, prepared as ASDs at a 10% drug loading with hydroxypropyl methylcellulose acetyl succinate (HPMCAS). We evaluated their release in aspirated fasted-state human intestinal fluid (FaHIF), and multi-component fasted-state simulated intestinal fluid (composite-FaSSIF), characterizing the supersaturation profiles and drug-rich nanodroplets that formed. Complete release was observed for atazanavir ASDs over a 90 min period. Flux for dissolved atazanavir ASDs remained high over the experimental time period of 3 h. In contrast, posaconazole solution concentrations were initially high and then decreased. Likewise, flux was initially high and then decreased where these changes are attributed to crystallization of the drug. Generation of spherical nano-sized amorphous droplets of ∼100-150 nm was found to occur in ex vivo FaHIF media for both ASDs, maximizing the diffusive flux during the supersaturation window. Moreover, buffer capacity differences were postulated to influence release rates of ASDs in simulated vs aspirated fluids. Importantly, the solution phase phenomena observed during ASD release in simulated fluids, namely amorphous nanodroplet formation and drug crystallization, were also found to occur in aspirated luminal fluids.

Epididymis-specific RNase A family genes regulate fertility and small RNA processing.

Sperm small RNAs are implicated in intergenerational transmission of paternal environmental effects. Small RNAs generated by cleavage of tRNAs, known as tRNA fragments (tRFs), are an abundant class of RNAs in mature sperm, and can be modulated by environmental conditions. The ribonuclease(s) responsible for the biogenesis of tRFs in the male reproductive tract remains unknown. Angiogenin, a member of the Ribonuclease A superfamily (RNase A), cleaves tRNAs to generate tRFs in response to cellular stress. Four paralogs of Angiogenin, namely Rnase9, Rnase10, Rnase11, and Rnase12, are specifically expressed in the epididymis-a long, convoluted tubule where sperm mature and acquire fertility and motility. The biological functions of these genes remain largely unknown. Here, by generating mice deleted for all four genes (Rnase9-12-/-, termed "KO" for Knock Out), we report that these genes regulate fertility and RNA processing. KO mice showed complete male sterility. KO sperm fertilized oocytes in vitro but failed to efficiently fertilize oocytes in vivo, likely due to an inability of sperm to pass through the utero-tubular junction. Intriguingly, there were decreased levels of fragments of tRNAs (tRFs) and rRNAs (rRNA-derived small RNAs or rsRNAs) in the KO epididymis and epididymal luminal fluid, implying that Rnase9-12 regulate the biogenesis and/or stability of tRFs and rsRNAs. Importantly, KO sperm showed a dramatic decrease in the levels of tRFs, demonstrating a role of Rnase9-12 in regulating sperm RNA composition. Together, our results reveal an unexpected role of four epididymis-specific non-canonical RNase A family genes in fertility and RNA processing.

Spatially and temporally precise microbiome profiling in the small intestine using the SIMBA capsule with X-ray tracking

IntroductionFew minimally invasive options for sampling the small intestinal (SI) luminal fluid exist to study the SI microbiota in health and disease. To address the lack of tools and methods to study GI regions that are difficult to access, Nimble Science developed a fully autonomous and passive sampling method, the Small Intestine MicroBiome Aspiration (SIMBA™) capsule, for convenient, high-quality, and reliable sampling to study the diet-microbiota interactions in the SI. MethodsThe sealing efficacy and microbial DNA preservation capacity of the SIMBA capsules was first validated through in vitro simulation assays. Then, a clinical study was conducted with 20 healthy participants to validate the in vivo use of SIMBA capsules to reliably capture samples for SI microbiome analysis before and after an intervention (NCT04489329). Briefly, participants ingested the capsules at baseline and 7 days later, with a probiotic capsule containing a blend of L. rhamnosus R0011 and B. longum R0175. Following baseline SIMBA capsule ingestion, multiple low-dosage x-ray scans were performed to track the sampling location. Fecal samples corresponding with the baseline and intervention capsule were analyzed for comparison. ResultsThe SIMBA capsules’ performance in vitro demonstrated the potential for contamination-free sampling with preservation of the microbial communities. Within the clinical study, the capsules performed safely and reliably for collection of SI content. X-ray tracking confirmed that 97.2% of the capsules completed sample collection in the SI regions before reaching the colon. Importantly, our data showed that the capsules sampled in the right area of the intestines and that baseline SIMBA microbiome profile is significantly different from fecal microbiome profile. SIMBA successfully detected a concurrent probiotic intervention in the small intestine, which was not detectable using stool samples. DiscussionsThe high accuracy of sampling location and sealing efficacy of the SIMBA capsules makes them potentially useful research tools in clinical trials for studying diet-microbiota interactions in health and disease, and perhaps eventually for the clinical diagnosis of GI tract conditions affecting the SI such as SIBO.

P-117 Extracellular vesicles from fallopian tubal fluid benefit human embryo development in vitro

Abstract Study question Do extracellular vesicles (EVs) from Fallopian tubes benefit human embryo development, and how do they affect preimplantation embryo development? Summary answer The Fallopian tubal EVs can be internalized by human embryos, and they contain miR-21-5p that downregulated P53 in blastocysts. What is known already EVs in the Fallopian tubes are identified as key mediators in embryo-oviduct interactions that contribute to a successful pregnancy in vivo. We previously showed Fallopian tubal EVs can benefit mouse embryo development in vitro. However, if a similar phenomenon can be observed in human embryos and what is the key component in Fallopian tubal EVs remains elusive. Study design, size, duration The EVs were isolated from the luminal fluid of human Fallopian tubes (n = 23). Immature human oocytes were collected and in vitro matured and fertilized, and 2-pronuclei embryos were included in this study and randomly divided (n = 87). We cocultured these zygotes with Fallopian tubal EVs until the blastocyst stage. Further, two-cell mouse embryos were used to explore the expression levels of miRNAs in Fallopian tubal EVs. The effects of miR-21-5p on mouse embryos were analyzed. Participants/materials, setting, methods EVs were isolated using ultracentrifugation. The uptake of labeled EVs by human embryos was analyzed using confocal microscopy. The fragmentation of the day-3 embryos and the blastocyst rate were compared between the control and the EV-treatment group. Then the stage-specific expression of miRNAs was compared in mouse embryos that were either developed in vivo or cultured in vitro. miR-21-5p were supplemented to the culture medium and the p53-related apoptosis in embryos was evaluated. Main results and the role of chance EVs were measured less than 200 nm in diameter with a typical bi-layer membrane structure and expressed protein markers Heat shock protein 70 (HSP70) and CD9. Most of the fluorescence-labeled EVs were attached to the zona pellucida within 2 h of co-culture. However, human embryos internalized EV within 4 h, with fluorescence signals observed in embryonic cells. The fragmentation rate was decreased and the blastocyst rate was increased by EVs. miR-21-5p, which was highly enriched in EVs, was found to be expressed at a significantly lower level in in-vitro-cultured mouse embryos compared to the in-vivo-developed embryos, through the two-cell stage to the eight-cell stage. miR-21-5p agomir supplementation to the culture medium was found to increase the mouse blastocyst formation rate, decrease apoptosis, and downregulate the P53 expression level in mouse blastocysts. Limitations, reasons for caution Though we observed that Fallopian tubal EVs could be internalized by human embryos and improve embryo development morphologically, the subsequent mechanism exploration was conducted using mouse embryos. Whether the stage-specific expression level of miR-21-5p and the effects of miR-21-5p are the same in human embryos remains to be determined. Wider implications of the findings This study showed miRNAs in Fallopian tubal EVs can act as embryonic survival signals, which opens the insight into further explorations to demonstrate the molecular mechanisms during early embryo development, as well as holds the potential to optimize the culture medium used in ART in the future. Trial registration number 2022YFC2702503

Characterization of luminal contents from the fasted human proximal colon

To treat colonic diseases more effectively, improved therapies are urgently needed. In this respect, delivering drugs locally to the colon is a key strategy to achieve higher local drug concentrations while minimizing systemic side effects. Understanding the luminal environment is crucial to efficiently develop such targeted therapies and to predict drug disposition in the colon. In this clinical study, we collected colonic contents from an undisturbed fasted proximal colon via colonoscopy and characterized their composition with regard to drug disposition. Colonic pH, osmolality, protein content, bile salts, lipids, phospholipids and short-chain fatty acids were investigated in 10 healthy volunteers (8 male and 2 female, age 19–25). The unique environment of the proximal colon was reflected in the composition of the sampled luminal fluids and the effect of the microbiota could be observed on the pH (median 6.55), the composition of bile salts (majority deconjugated and secondary), and the abundance of short-chain fatty acids. At the same time, an increase in phospholipid concentration, osmolality and total protein content compared to reported ileal values was seen, likely resulting from desiccation. Lipids could only be found in low quantities and mainly in the form of cholesterol and free fatty acids, showing almost complete digestion and absorption by the time luminal contents reach the colon. All characteristics also displayed the considerable intersubject variability found in different regions of the gastrointestinal tract. This study contributes to an improved understanding of the luminal conditions in the proximal colon and facilitates the development of new predictive tools to study colonic drug absorption.

TAKOTSUBO SYNDROME AND SMALL BOWEL OBSTRUCTION: WHEN THE HEART–GUT AXIS GOES HAYWIRE

Abstract Case Report A 66 years old woman was admitted to our emergency department for the onset of intense epigastric pain, nausea and asthenia after breakfast. She had a history of hypertension, dyslipidemia, COPD and deep venous thrombosis with MTHFR gene mutation. Moreover, she had a positive family history for coronary artery disease (CAD). On arrival, she was apyretic, hemodynamically stable, complaining chest and persisting epigastric pain. A mild and diffuse abdominal tenderness was noted without peritoneal signs. The ECG showed sinus tachycardia, ST elevation (STE) in leads D1, aVL, V2 and V3 and ST depression in leads D3, aVR and V1. Blood test revealed a marked increase of PCR with leukocytosis and elevation of Hs–cTnI (840 ng/L) while serum electrolytes were normal. Bed–side echocardiography revealed apical and mid–ventricular akinesia with a severe reduction of left ventricular ejection fraction (LVEF). InterTAK score was 62 (62,6% probability of Takotsubo syndrome – TTS). An urgent coronary angiography (CA) revealed non– obstructive CAD, while ventriculography showed a typical apical ballooning and hyperkinetic base of the LV suggestive of TTS. Subsequently, the CT abdomen showed extra–luminal free fluid and a dilated proximal small bowel (SB) with air–fluid levels and a bubbly pattern of gas in the wall. These radiological findings were consistent with a mechanical SB obstruction, complicated by mesenteric venous thrombosis and intestinal pneumatosis. Exploratory laparotomy revealed SB necrosis as consequence of bridle–mediated obstruction and therefore an ileum resection was performed. The patient clinically improved post–operatively with a complete recovery of the LVEF. Discussion We described a rare presentation of SB obstruction which induced a TTS in a setting of severe psychophysical stress. The management of this emergency situation in which cardiac injury and acute abdomen occurred simultaneously can be challenging. The consulting cardiologist needs to quickly decide whether to prioritize the heart with urgent CA or the gastroenteric condition with surgery. Despite the likelihood of TTS can be high (even calculating InterTak score), new STE and positive troponins can’t be disregarded especially in patients with significant risk factors as seen in our case. Therefore, the strong evidence of a possible acute coronary syndrome may trump a life–saving treatment for SB obstruction.

Effects of alimentary-derived bacterial metabolites on energy metabolism in colonic epithelial cells and inflammatory bowel diseases

In this review, we present the main luminal fuels that are responsible for energy production in colonocytes, namely the bacterial metabolites short-chain fatty acids and lactate, which are produced from undigestible polysaccharides and proteins, and hydrogen sulfide that is mainly produced from undigested proteins. In addition to these luminal fuels, colonocytes can use glutamine, and to a lower extent glucose, as energy substrates provided by arterial capillaries. The effects of excessive concentrations of bacterial metabolites within the colonic luminal fluid (including butyrate, hydrogen sulfide, p-cresol, indole derivatives, ammonia, 4-hydroxyphenylacetic acid, and acetaldehyde) on the mitochondrial energy metabolism in colonic epithelial cells and the consequences of altered ATP production on the colonic epithelium renewal and barrier function are detailed, as well as consequences for water and electrolyte absorption. The relationships between modifications of these latter processes and development of colitis are then discussed. Finally, several mechanisms that are considered as adaptive against deleterious effects of bacterial metabolites on colonic epithelial cell energy metabolism are presented.

Endometrial responsiveness to interferon-tau and its association with subsequent reproductive performance in dairy heifers

Our objectives were to evaluate the endometrial responsiveness of dairy heifers to an intrauterine infusion of recombinant bovine interferon-tau (rbIFN-τ) and to associate endometrial responses to rbIFN-τ with subsequent reproductive performance. In experiments 1 and 2, cyclic heifers were enrolled in a program for synchronization of the estrous cycle, and blood sampling and ultrasonography examinations were performed on d 0, 4, 7, 11, and 14 of the estrous cycle. In experiment 1, heifers were randomly assigned to receive an intrauterine infusion containing 2 µg of rbIFN-τ (rbIFN-τ = 19) or saline control (CTRL = 19) into the uterine horn ipsilateral to the corpus luteum (CL) on d 14 of the estrous cycle. Then, 6 hours after the infusion, the infused uterine horn was flushed for sampling of the uterine luminal fluid (ULF) for composition analysis, and the endometrium was biopsied for transcriptomics. In experiment 2, 100 heifers received an intrauterine infusion of rbIFN-τ, and the same procedures for uterine sample collection were performed as described in experiment 1. After the intrauterine test, heifers were enrolled in a breeding program and classified as highly fertile (HF; pregnant at first AI) or subfertile (SF; not pregnant at first AI). Statistical analyses were performed using regression models, which included the effects of treatment (experiment 1: CTRL vs. rbIFN-τ) or fertility group (experiment 2: HF vs. SF) and block of samples. Intrauterine infusion of rbIFN-τ increased the expression of classical interferon-stimulated genes in the endometrium (e.g., ISG15, MX1, OAS2, IRF9, and USP18), and an antiviral response was predicted to be the main downstream effect of the transcriptome changes. In addition, rbIFN-τ increased the abundance of cholesterol, glycerol, and the overall concentration of oxylipins in the ULF. Analysis of endometrial transcriptome between HF and SF heifers revealed important differences in the expression of genes associated with cell signaling, metabolism, attachment, and migration, with a large representation of genes encoding extracellular matrix proteins. In general, differentially expressed genes were expected to be downregulated by IFN-τ but seemed to fail to be downregulated in SF heifers, resulting in higher expression in SF compared with HF heifers. Subfertile heifers had lower concentrations of glycerol and an altered profile of oxylipins in the ULF, with a lower abundance of oxylipins derived from arachidonic acid and dihomo-γ-linolenic acid, and a greater abundance of oxylipins derived from linoleic acid. Measurements of ovarian function did not differ between groups and, therefore, did not influence the observed results in uterine biology. Overall, the endometrial responsiveness to IFN-τ is variable among individuals and associated with subsequent fertility of heifers, indicating that communication between conceptus and endometrium is critical for the uterine receptivity and survival of pregnancy.

Minimally Invasive Approaches to Diagnose and Monitor Eosinophilic GI Diseases.

This review seeks to understand novel avenues for eosinophilic GI disease management. Biomarkers offer a unique and non-invasive approach to tracking EoE disease progression. While no biomarkers have definitively met the diagnostic criteria for eosinophilic GI diseases, some biomarkers have been shown to be associated with disease activity. Here, we examine the potential of recently studied biomarkers. Current research shows advancements in blood, luminal fluid, and breath testing. Particular areas of interest include mRNA analyses, protein fingerprinting, amplicon sequence variants (ASVs), T cells and IgE receptors, eosinophilic cationic proteins, cytokines, and nitric oxide exhalation. Preliminary results showed that mucosal biomarkers, directly captured from the esophagus, may reflect the best representation of biopsy-based results, in contrast to biomarkers obtained from indirect or peripheral (blood, breath) methods. However, this is based on limited clinical studies without sufficient numbers to evaluate true diagnostic accuracy. Large-scale randomized trials are needed to fully ascertain both the optimal sampling technique and the specific biomarkers that reflect diagnostic status of the disease.

Metastasis-associated protein 1 participates in regulating luminal acidification of the epididymis via repressing estrogen receptor alpha transcription.

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.

102 Estradiol and follicular growth are associated with estrous behavior monitored by an accelerometer in grazing beef cows

Abstract In beef cattle, 40% of cows that display estrus do not remain pregnant after embryo transfer (ET). Estrus is a physiological behavior influenced by the concentration of sex-steroids. The use of accelerometer devices for estrus detection is a common practice in dairy operations. It remains unclear if this same artificial intelligence software can accurately measure reproductive behavior in beef females. We hypothesized that estrous variables detected by an accelerometer are associated with ovarian, hormonal, pregnancy outcome, and fertility-related variables measured in the cow. Fifty Bos indicus-influenced, cyclic, primiparous cows were fitted with an accelerometer collar (SCR Engineers, Israel) and a breeding indicator (Estrotectâ) to detect estrus after a synchronization protocol in five subsequent experimental replicates (n = 50*5 = 250). Only cows that showed estrus (d 0) remained in the study. The following variables were used from the accelerometer software: Activity Peak, Rumination Nadir, Estrus Duration, and Heat Index, calculated by proprietary algorithms. Dominant follicle (DF) diameter and plasma concentration of estradiol were measured on d -2 (day of progesterone device removal) and on d 0. On d -1 and d 4, endometrial thickness and luminal fluid accumulation were measured by ultrasonography. On d 7, two embryos were transferred/cow. Pregnancies were diagnosed on d 45 and terminated. After 30 d, a new replicate started. Cows included in at least two replicates that remained pregnant 100%, 99-50%, or less than 50% were classified as fertile, sub fertile, and infertile, respectively. Estrus display was recorded for 166 cows (66%). Estrus detection by accelerometer and Estrotect had excellent agreement rate (kappa coefficient 0.76). Cows detected in estrus only by Estrotect had 66% less pregnancy/ET than cows in estrus by both (9/21 vs. 97/145; P = 0.01). The DF diameter on d 0 was not associated with accelerometer variables. Cows with greater and moderate Activity Peak had a greater growth rate of the DF from d -2 to d 0 than lower Activity Peak cows (interaction day*Activity Peak; P = 0.06). Estradiol on d -1 was positively associated with the Activity Peak (P < 0.0001). Cows with lower Activity Peak had a thicker endometrium on both d -1 and d 4 (P=0.02), and greater uterine luminal fluid score on d -1 (P=0.03). Fertility status was not associated with any accelerometer variables. In conclusion, the Activity Peak is associated positively with estradiol concentration and DF growth, however no association was found with pregnancy outcome and fertility. Therefore, the main advantage of accelerometers in beef ranches is the ability to detect estrus at a similar accuracy as breeding indicators, using less labor. Future enhancements in hardware and software to allow prediction of pregnancy outcomes are expected to increase reproductive efficiency in cow-calf operations.

A revised conceptual framework for mouse vomeronasal pumping and stimulus sampling

The physiological performance of any sensory organ is determined by its anatomy and physical properties. Consequently, complex sensory structures with elaborate features have evolved to optimize stimulus detection. Understanding these structures and their physical nature forms the basis for mechanistic insights into sensory function. Despite its crucial role as a sensor for pheromones and other behaviorally instructive chemical cues, the vomeronasal organ (VNO) remains a poorly characterized mammalian sensory structure. Fundamental principles of its physico-mechanical function, including basic aspects of stimulus sampling, remain poorly explored. Here, we revisit the classical vasomotor pump hypothesis of vomeronasal stimulus uptake. Using advanced anatomical, histological, and physiological methods, we demonstrate that large parts of the lateral mouse VNO are composed of smooth muscle. Vomeronasal smooth muscle tissue comprises two subsets of fibers with distinct topography, structure, excitation-contraction coupling, and, ultimately, contractile properties. Specifically, contractions of a large population of noradrenaline-sensitive cells mediate both transverse and longitudinal lumen expansion, whereas cholinergic stimulation targets an adluminal group of smooth muscle fibers. The latter run parallel to the VNO's rostro-caudal axis and are ideally situated to mediate antagonistic longitudinal constriction of the lumen. This newly discovered arrangement implies a novel mode of function. Single-cell transcriptomics and pharmacological profiling reveal the receptor subtypes involved. Finally, 2D/3D tomography provides non-invasive insight into the intact VNO's anatomy and mechanics, enables measurement of luminal fluid volume, and allows an assessment of relative volume change upon noradrenergic stimulation. Together, we propose a revised conceptual framework for mouse vomeronasal pumping and, thus, stimulus sampling.

The multifaceted links between hearing loss and chronic kidney disease.

Hearing loss affects nearly 1.6 billion people and is the third-leading cause of disability worldwide. Chronic kidney disease (CKD) is also a common condition that is associated with adverse clinical outcomes and high health-care costs. From a developmental perspective, the structures responsible for hearing have a common morphogenetic origin with the kidney, and genetic abnormalities that cause familial forms of hearing loss can also lead to kidney disease. On a cellular level, normal kidney and cochlea function both depend on cilial activities at the apical surface, and kidney tubular cells and sensory epithelial cells of the inner ear use similar transport mechanisms to modify luminal fluid. The two organs also share the same collagen IV basement membrane network. Thus, strong developmental and physiological links exist between hearing and kidney function. These theoretical considerations are supported by epidemiological data demonstrating that CKD is associated with a graded and independent excess risk of sensorineural hearing loss. In addition to developmental and physiological links between kidney and cochlear function, hearing loss in patients with CKD may be driven by specific medications or treatments, including haemodialysis. The associations between these two common conditions are not commonly appreciated, yet have important implications for research and clinical practice.

Select amino acids recover cytokine-altered ENaC function in human bronchial epithelial cells.

The airway epithelium plays a pivotal role in regulating mucosal immunity and inflammation. Epithelial barrier function, homeostasis of luminal fluid, and mucociliary clearance are major components of mucosal defense mechanisms. The epithelial sodium channel (ENaC) is one of the key players in controlling airway fluid volume and composition, and characteristic cytokines cause ENaC and barrier dysfunctions following pulmonary infections or allergic reactions. Given the limited understanding of the requisite duration and magnitude of cytokines to affect ENaC and barrier function, available treatment options for restoring normal ENaC activity are limited. Previous studies have demonstrated that distinct amino acids can modulate epithelial ion channel activities and barrier function in intestines and airways. Here, we have investigated the time- and concentration-dependent effect of representative cytokines for Th1- (IFN-γ and TNF-α), Th2- (IL-4 and IL-13), and Treg-mediated (TGF-β1) immune responses on ENaC activity and barrier function in human bronchial epithelial cells. When cells were exposed to Th1 and Treg cytokines, ENaC activity decreased gradually while barrier function remained largely unaffected. In contrast, Th2 cytokines had an immediate and profound inhibitory effect on ENaC activity that was subsequently followed by epithelial barrier disruption. These functional changes were associated with decreased membrane protein expression of α-, β-, and γ-ENaC, and decreased mRNA levels of β- and γ-ENaC. A proprietary blend of amino acids was developed based on their ability to prevent Th2 cytokine-induced ENaC dysfunction. Exposure to the select amino acids reversed the inhibitory effect of IL-13 on ENaC activity by increasing mRNA levels of β- and γ-ENaC, and protein expression of γ-ENaC. This study indicates the beneficial effect of select amino acids on ENaC activity in an in vitro setting of Th2-mediated inflammation suggesting these amino acids as a novel therapeutic approach for correcting this condition.

A nanoporous hydrogel-based model to study chemokine gradient-driven angiogenesis under luminal flow.

The growth of new blood vessels through angiogenesis is a highly coordinated process, which is initiated by chemokine gradients that activate endothelial cells within a perfused parent vessel to sprout into the surrounding 3D tissue matrix. While both biochemical signals from pro-angiogenic factors, as well as mechanical cues originating from luminal fluid flow that exerts shear stress on the vessel wall, have individually been identified as major regulators of endothelial cell sprouting, it remains unclear whether and how both types of cues synergize. To fill this knowledge gap, here, we created a 3D biomimetic model of chemokine gradient-driven angiogenic sprouting, in which a micromolded tube inside a hydrogel matrix is seeded with endothelial cells and connected to a perfusion system to control fluid flow rates and resulting shear forces on the vessel wall. To allow for the formation of chemokine gradients despite the presence of luminal flow, a nanoporous synthetic hydrogel that supports angiogenesis but limits the interstitial flow proved crucial. Using this system, we find that luminal flow and resulting shear stress is a major regulator of the speed and morphogenesis of angiogenic sprouting, whose action is mediated through changes in vascular permeability.