Brush Border Research Articles

A Screening Study on Effects Varying Dietary Macronutrient Composition on Gut Functions in Lumpfish (Cyclopterus lumpus)

Cultivation of lumpfish (Cyclopterus lumpus) as lice cleaner fish for salmon is now expanding. For successful cultivation of a new species, understanding the basic biology of digestive functions is vital to facilitate and optimize diet formulation. This paper presents results from two experiments conducted to deepen our knowledge on lumpfish intestine physiology. Experiment 1 was a 42-day feeding trial in which lumpfish were fed twelve different diets in the following ranges of macronutrients: Protein 43-68%, lipid 4-17%, and carbohydrate 6-17%. Intestinal tissue, gut content and liver were sampled from 6 fish per tank. The results showed that with increasing lipid level and corresponding decrease in protein level, there was a linear decrease in several of the observed biomarkers, including activity of brush border membrane digestive enzymes, expression of genes related to nutrient digestion and transport, ion exchange, immune regulation, and cell remodeling. Increased intracellular accumulation of lipid (steatosis) was observed in gut and liver with increasing dietary lipid level. Fewer effects were observed for increased dietary carbohydrate and corresponding decreased protein level. Experiment 2 was a two-week feeding trial for estimation of macronutrient digestibility in which lumpfish were fed three diets, all containing 55% crude protein, with lipid to carbohydrate ratio of the low lipid diet of 7.5%/18.3%, the medium lipid diet of 13.8%/14.6%, and high lipid diet of 18.1%/9.5%. Fecal samples were collected as pooled samples per tank. These results showed that fatty acid digestibility’s increased as dietary lipid level increased. Of note, starch digestibility decreased greatly as starch level increased, whereas protein digestibility did not change as lipid or starch level varied. Taken together, the present studies indicated that increasing lipid level in the diet with corresponding decrease in protein level affects digestion, absorption, and immune responses in the lumpfish intestine. Variation in dietary carbohydrate to protein level showed less effects, possibly due to low starch digestibility which makes the variation in available carbohydrates much less than the variation in the analysed level of dietary carbohydrates.

Protective effect of Astragalus membranaceus and Astragaloside IV in sepsis-induced acute kidney injury.

Background: Acute kidney injury (AKI) is the most common target organ damage in sepsis. Sepsis-associated AKI (SA-AKI) may be characterized by damage to the renal tubular epithelium. In this study, the pharmacological mechanisms of Astragalus membranaceus and its active monomer Astragaloside IV (AS-IV) were predicted based on a network pharmacology approach and validated both in vitro and in vivo using the SA-AKI model.Method: We constructed an in vivo sepsis model using a mouse cecum ligation puncture (CLP) and HK-2 cells were treated with lipopolysaccharide (LPS) to mimic Gram (–) induced sepsis to assess the renal-protective efficacy of Astragalus membranaceus and AS-IV.Results: The findings demonstrated that Astragalus membranaceus and AS-IV attenuate renal tubular injury in mice with polymicrobial sepsis, including vacuolization, loss of brush border, mitochondrial ultrastructural changes, and increased staining of kidney injury molecule-1 (KIM-1). AS-IV protected human proximal tubular epithelial (HK-2) cells against LPS induced cell viability loss. Both Astragalus membranaceus and AS-IV activated the PI3K/AKT pathway both in vitro and in vivo, as shown by Western blot and immunohistochemistry analysis.Conclusion: The findings demonstrate that Astragalus membranaceus and AS-IV protect against sepsis-induced kidney tubular injury by activating the PI3K/AKT pathway.

Condensed tannins increased intestinal permeability of Chinese seabass (Lateolabrax maculatus) based on microbiome-metabolomics analysis

Condensed tannins (CT) have been reported to induce intestinal injury of fish, whereas the injury mechanism is unclear. This study evaluated the effects of dietary grape seed CT on growth, intestinal permeability, tight junction structure, intestinal digestive and brush border enzyme activity, bacterial community and intestinal metabolites of Chinese seabass (Lateolabrax maculatus). A total of 640 fish were randomly distributed into 16 tanks with 4 tanks per group and 40 fish per tank. Four diets were formulated to contain 0 (CT0), 1 (CT1), 2 (CT2), and 2 g/kg of CT plus 4 g/kg of polyethylene glycol (PEG) (CT2 + PEG). PEG specifically binds with CT to neutralize CT activity. The experiment was arranged as complete randomized design. Fish were hand-fed to apparent satiation twice daily and the feeding trial lasted for 63 days. Fish fed CT2 had lower (P < 0.05) final body weight, weight gain rate, specific growth rate and feed intake, but higher (P < 0.05) serum diamine oxidase activity and endotoxin level than other diets. Tight junction structures in CT1 and CT2 showed varying degrees of ambiguity and damage. The trypsin activity ranked as CT0 > CT1 > CT2 (P < 0.05). Fish fed CT2 had lower (P < 0.05) activities of lipase, amylase and creatine kinase than other diets. Na+, K+-ATPase activity was lower (P < 0.05) in CT1 and CT2 than in CT0. All parameters were similar (P > 0.05) between CT0 and CT2 + PEG. Intestinal microbiota analysis showed that the abundances of Bacillus, Acinetobacter and Pseudomonas were decreased (P < 0.05) but Clostridium was increased (P < 0.05) as dietary CT increasing. Fish fed CT2 + PEG had higher (P < 0.05) abundances of Bacillus, Acinetobacter and Pseudomonas but lower (P < 0.05) Clostridium than those of fish fed CT2. Bacterial diversity measurements were not affected (P > 0.05) by dietary treatments. Metabolomics analysis indicated that CT altered the intestinal metabolomics profile of L. maculatus. The valeric acid was decreased (P < 0.05), but phenylacetic acid was increased (P < 0.05) as dietary CT increasing. Compared to CT0, CT2 had lower (P < 0.05) isobutyric acid but higher (P < 0.05) butyric acid and isovaleric acid. In conclusion, this study suggests association between reduced growth and increased intestinal permeability and varying intestinal bacterial microbiota and metabolites of L. maculatus. This study warns that dietary CT at 2 g/kg could induce intestinal injury, microecological imbalance and metabolic disorder of fish, and the resulting adverse effects may neutralize by applying polyethylene glycol in fish diets.

Fipronil exposure compromises respiration and damages the Malpighian tubules of the stingless bee Partamona helleri Friese (Hymenoptera: Apidae).

Fipronil has been widely used in agriculture and forestry in Brazil to control several pests. However, this insecticide may be hazardous to non-target organisms, including stingless bees, which are essential pollinators of crops and natural environments. Here, we investigated the effect of 24-h acute oral exposure to LC50 of fipronil on the Malpighian tubules of the stingless bee Partamona helleri (Friese). Insecticide exposure decreases the respiration rate of forager bees, and the Malpighian tubules are severely affected, as shown by the epithelial architecture disorganization, loss of cytoplasmic content, degradation of the brush border, and nuclear pyknosis. In addition, fipronil ingestion increases the number of Malpighian cells positive for peroxidase, LC3, cleaved caspase-3, and JNK. However, Notch and ERK1/2-positive cells decrease in the exposed bees. These changes in the signaling proteins indicate an increase in oxidative stress, autophagy and apoptosis, and impairment of cell recovery. Overall, our results demonstrate the toxicological effects of fipronil on a stingless bee, which compromises the physiology of this important pollinator.

Exploratory phosphoproteomics profiling of Aedes aegypti Malpighian tubules during blood meal processing reveals dramatic transition in function.

Malpighian tubules, the renal organs of mosquitoes, facilitate the rapid dehydration of blood meals through aquaporin-mediated osmosis. We performed phosphoproteomics analysis of three Malpighian tubule protein-libraries (1000 tubules/sample) from unfed female mosquitoes as well as one and 24 hours after a blood meal. We identified 4663 putative phosphorylation sites in 1955 different proteins. Our exploratory dataset reveals blood meal-induced changes in phosphorylation patterns in many subunits of V-ATPase, proteins of the target of rapamycin signaling pathway, vesicle-mediated protein transport proteins, proteins involved in monocarboxylate transport, and aquaporins. Our phosphoproteomics data suggest the involvement of a variety of new pathways including nutrient-signaling, membrane protein shuttling, and paracellular water flow in the regulation of urine excretion. Our results support a model in which aquaporin channels translocate from intracellular vesicles to the cell membrane of stellate cells and the brush border membrane of principal cells upon blood feeding.

Effect of Excessive Serotonin on Pharmacokinetics of Cephalexin after Oral Administration: Studies with Serotonin-Excessive Model Rats.

Serotonin (5-HT) is important for gastrointestinal functions, but its role in drug absorption remains to be clarified. Therefore, the pharmacokinetics and oral absorption of cephalexin (CEX) were examined under 5-HT-excessive condition to understand the role of 5-HT. 5-HT-excessive rats were prepared by multiple intraperitoneal dosing of 5-HT and clorgyline, an inhibitor for 5-HT metabolism, and utilized to examine the pharmacokinetics, absorption behavior and the intestinal permeability for CEX. Higher levels of 5-HT in brain, plasma and small intestines were recognized in 5-HT-excessive rats, where the oral bioavailability of CEX was significantly enhanced. The intestinal mucosal transport via passive diffusion of CEX was significantly increased, while its transport via PEPT1 was markedly decreased specifically in the jejunal segment, which was supported by the decrease in PEPT1 expression on brush border membrane (BBM) of intestinal epithelial cells. Since no change in antipyrine permeability and significant increase in FITC dextran-4 permeability were observed in 5-HT-excessive rats, the enhanced permeability for CEX would be attributed to the opening of tight junction, which was supported by the significant decrease in transmucosal electrical resistance. In 5-HT-excessive rats, furthermore, total body clearance of CEX tended to be larger and the decrease in PEPT2 expression on BBM in kidneys was suggested to be one of the reasons for it. 5-HT-excessive condition enhanced the oral bioavailability of CEX in rats, which would be attributed to the enhanced permeability across the intestinal mucosa via passive diffusion through the paracellular route even though the transport via PEPT1 was decreased.

Effects of Different Marine-Based Lipid Sources on Growth Performance, Activities of Digestive Enzyme, Antioxidant Responses, and Lipid Metabolism of Large Yellow Croaker (Larimichthys crocea) Larvae

The study of lipid nutrition is an important guarantee for the development of high-efficiency artificial microdiet for fish larvae. Existing studies on lipid nutrition of larvae mainly focus on nutrient requirements and metabolism through a single lipid source. However, there are few reports on the effects of different marine-based lipid sources on fish larvae. In this study, a 30-day feeding experiment was conducted to evaluate the effects of different dietary marine-based lipid sources on survival, growth performance, appetite gene expression, activities of digestive enzyme, antioxidant responses, and lipid metabolism of large yellow croaker (Larimichthys crocea) larvae (initial weight 4.71 ± 0.21 mg ). Four isonitrogenous (520 g/kg crude protein) and isolipidic (190 g/kg crude lipid) diets were formulated to contain fish oil (FO), krill oil (KO), squid viscera oil (SVO), and Schizochytrium sp. oil (SSO), respectively. Results showed that larvae fed with the SSO diet had significantly higher survival rate (SR) than those fed with dietary FO and SVO ( P &lt; 0.05 ). However, larvae fed with the SSO diet had significantly lower final body weight (FBW) and specific growth rate (SGR) than those fed with other diets ( P &lt; 0.05 ). Furthermore, larvae fed with the SSO diet had significantly higher percentage of total n-3 long-chain polyunsaturated fatty acids (LC-PUFA) than those fed with other diets, followed by KO and SVO diets ( P &lt; 0.05 ). Larvae fed with the SSO diet had significantly lower mRNA expression of orexigenic genes (npy and ghrelin) and significantly higher mRNA expression of anorexigenic gene (cart) than those fed with dietary FO ( P &lt; 0.05 ). Meanwhile, larvae fed with the SSO diet had significantly higher activity of alkaline phosphatase (AKP) in intestinal segments (IS) and brush border membrane (BBM) than those fed with dietary FO and SVO ( P &lt; 0.05 ). Larvae fed with the SSO diet had significantly higher activity of total antioxidant capacity (T-AOC) than those fed with dietary FO ( P &lt; 0.05 ). The TG content and mRNA expression of lipogenesis genes (srebp-1c, fas, and scd1) were markedly lower in larvae fed with SSO, KO, and SVO diets than in those fed with dietary FO ( P &lt; 0.05 ). Meanwhile, larvae fed with KO and SSO diets had significantly higher mRNA expression of the lipolysis gene (cpt-1) than those fed with other diets ( P &lt; 0.05 ). In conclusion, results of the present study showed that FO can be completely replaced in large yellow croaker larvae feed with KO and SVO without negative effects. Moreover, the complete replacement of FO by SSO can improve survival, activities of digestive enzyme, antioxidant responses, and lipid metabolism, despite inhibiting the appetite and growth of large yellow croaker larvae.

A Novel Soluble ACE2 Protein Provides Lung and Kidney Protection in Mice Susceptible to Lethal SARS-CoV-2 Infection.

Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) uses full-length angiotensin converting enzyme 2 (ACE2) as a main receptor to enter target cells. The goal of this study was to demonstrate the preclinical efficacy of a novel soluble ACE2 protein with increased duration of action and binding capacity in a lethal mouse model of COVID-19. A human soluble ACE2 variant fused with an albumin binding domain (ABD) was linked via a dimerization motif hinge-like 4-cysteine dodecapeptide (DDC) to improve binding capacity to SARS-CoV-2. This novel soluble ACE2 protein (ACE2-1-618-DDC-ABD) was then administered intranasally and intraperitoneally to mice before intranasal inoculation of SARS-CoV-2 and then for two additional days post viral inoculation. Untreated animals became severely ill, and all had to be humanely euthanized by day 6 or 7 and had pulmonary alveolar hemorrhage with mononuclear infiltrates. In contrast, all but one mouse infected with a lethal dose of SARS-CoV-2 that received ACE2-1-618-DDC-ABD survived. In the animals inoculated with SARS-CoV-2 that were untreated, viral titers were high in the lungs and brain, but viral titers were absent in the kidneys. Some untreated animals, however, had variable degrees of kidney proximal tubular injury as shown by attenuation of the proximal tubular brush border and increased NGAL and TUNEL staining. Viral titers in the lung and brain were reduced or nondetectable in mice that received ACE2-1-618-DDC-ABD, and the animals developed only moderate disease as assessed by a near-normal clinical score, minimal weight loss, and improved lung and kidney injury. This study demonstrates the preclinical efficacy of a novel soluble ACE2 protein, termed ACE2-1-618-DDC-ABD, in a lethal mouse model of SARS-CoV-2 infection that develops severe lung injury and variable degrees of moderate kidney proximal tubular injury.

Growth and Welfare of Rainbow Trout (Oncorhynchus mykiss) in Response to Graded Levels of Insect and Poultry By-Product Meals in Fishmeal-Free Diets.

Simple SummaryProcessed animal proteins, such as poultry by-product meal (PBM) and insect meal from black soldier fly (BSFM), are receiving growing interest as alternative or complementary protein sources for carnivorous farmed fish diets, due to their high nutritional value and low environmental footprint. The present study aimed to evaluate the effects of PBM and BSFM as partial substitutes for vegetable protein in fishmeal-free diets on growth, whole-body composition, nutrient-energy mass balance and retention, digestive functions, stress, metabolic status, innate immunity, and liver health in rainbow trout (Oncorhynchus mykiss). The overall results showed that fishmeal-free diets, including high levels of both PBM and BSFM, either singly or in combination, improved growth, dietary nutrient and energy utilization, and gut function relative to a plant-protein control diet or a fishmeal-based one, with no detrimental effects on fish welfare.This study compared the nutrient-energy retention, digestive function, growth performance, and welfare of rainbow trout (ibw 54 g) fed isoproteic (42%), isolipidic (24%), fishmeal-free diets (CV) over 13 weeks. The diets consisted of plant-protein replacement with graded levels (10, 30, 60%) of protein from poultry by-product (PBM) and black soldier fly H. illucens pupae (BSFM) meals, either singly or in combination. A fishmeal-based diet was also tested (CF). Nitrogen retention improved with moderate or high levels of dietary PBM and BSFM relative to CV (p < 0.05). Gut brush border enzyme activity was poorly affected by the diets. Gastric chitinase was up-regulated after high BSFM feeding (p < 0.05). The gut peptide and amino acid transport genes were differently regulated by protein source and level. Serum cortisol was unaffected, and the changes in metabolites stayed within the physiological range. High PBM and high BSFM lowered the leukocyte respiratory burst activity and increased the lysozyme activity compared to CV (p < 0.05). The BSFM and PBM both significantly changed the relative percentage of lymphocytes and monocytes (p < 0.05). In conclusion, moderate to high PBM and BSFM inclusions in fishmeal-free diets, either singly or in combination, improved gut function and nutrient retention, resulting in better growth performance and the good welfare of the rainbow trout.

Lipid Messenger Phosphatidylinositol-4,5-Bisphosphate Is Increased by Both PPARα Activators and Inhibitors: Relevance for Intestinal Cell Differentiation.

Simple SummaryFibrates, such as fenofibrate, are widely used drugs for dyslipidaemia treatment. It is known that they activate peroxisome proliferator-activated receptor α (PPARα) which serves as a lipid sensor in the organism. This article addresses how activators and inhibitor of the PPARα could affect differentiation of intestinal cells. Carcinogenesis is a disruption of normal differentiation process and colorectal carcinoma is the third most common cancer in terms of incidence, but the secondp in terms of mortality. One of the important signalling pathways in intestinal cell differentiation as well as carcinogenesis is PI3K/Akt/PTEN. We showed that PPARα activators as well as inhibitor affected the levels of one member of this pathway called phosphatidylinositol-4,5-bisphosphate. This molecule is important for formation of microvilli, the essential structures of fully differentiated intestinal cells.We investigated the effects of PPARα activators fenofibrate and WY-14643 as well as the PPARα inhibitor GW6471 on the PI3K/Akt/PTEN pathway of intestinal cell differentiation. Our previous study showed that all these compounds increased the expression of villin, a specific marker of intestinal cell differentiation in HT-29 and Caco2 cells. Our current results confirmed the central role of lipid messenger phosphatidylinositol-4,5-bisphosphate (PIP2), a known player in brush border formation, in mediating the effects of tested PPARα ligands. Although all tested compounds increased its levels, surprisingly, each of them affected different PIP2-metabolizing enzymes, especially the levels of PIP5K1C and PTEN. Moreover, we found a positive relationship between the expression of PPARα itself and PIP2 as well as PIP5K1C. By contrast, PPARα was negatively correlated with PTEN. However, the expression of antigens of interest was independent of PPARα subcellular localization, suggesting that it is not directly involved in their regulation. In colorectal carcinoma tissues we found a decrease in PTEN expression, which was accompanied by a change in its subcellular localization. This change was also observed for the regulatory subunit of PI3K. Taken together, our data revealed that fenofibrate, WY-14643, and GW6471 affected different members of the PI3K/Akt/PTEN pathway. However, these effects were PPARα-independent.

Oral administration of pentachlorophenol impairs antioxidant system, inhibits enzymes of brush border membrane, causes DNA damage and histological changes in rat intestine.

Pentachlorophenol (PCP) is a broad spectrum biocide that has many domestic and industrial applications. PCP enters the environment due to its wide use, especially as a wood preservative. Human exposure to PCP is through contaminated water and adulterated food products. PCP is highly toxic and is classified as class 2B or probable human carcinogen. In this study, we explored the effect of PCP on rat intestine. Adult rats were orally given different doses of PCP (25-150-mg/kg body weight/day) in corn oil for 5 days, whereas controls were given similar amount of corn oil. The rats were sacrificed 24h after the last treatment. A marked increase in lipid peroxidation, carbonyl content, and hydrogen peroxide level was seen. The glutathione and sulfhydryl group content was decreased in all PCP treated groups. This strongly suggests the generation of reactive oxygen species (ROS) in the intestine. PCP administration suppressed carbohydrate metabolism, inhibited enzymes of brush border membrane (BBM), and antioxidant defense system. It also led to increase in DNA damage, which was evident from comet assay, DNA-protein cross-linking, and DNA fragmentation. Histological studies supported the biochemical results showing marked dose-dependent tissue damage in intestines from PCP treated animals. This study reports for the first time that oral administration of PCP induces ROS, impairs the antioxidant system, damages DNA, and alters the enzyme activities of BBM and metabolic pathways in rat intestine.

Therapeutic effects of sulforaphane in ulcerative colitis: effect on antioxidant activity, mitochondrial biogenesis and DNA polymerization

ABSTRACT Objectives Ulcerative colitis (UC), an inflammatory bowel disease, affects mucosal lining of colon leading to inflammation and ulcers. Sulforaphane is a natural compound obtained from cruciferous vegetables. We aimed to investigate potential therapeutic effects of sulforaphane in experimentally induced UC in rats through affection antioxidant activity, mitochondrial biogenesis and DNA polymerization. Methods UC was induced in rats via an intracolonic single administration of 2 ml of 4% acetic acid. UC rats were treated with 15 mg/kg sulforaphane. Samples of colon were used to investigate gene expression and protein levels of peroxisome proliferator-activated receptor-gamma coactivator (PGC-1), mitochondrial transcription factor A (TFAM), mammalian target of rapamycin (mTOR), cyclin D1, nuclear factor erythroid 2-related factor-2 (Nrf2), heme Oxygenase-1 (HO-1) and proliferating cell nuclear antigen (PCNA). Results UC showed dark distorted Goblet cell nucleus with disarranged mucus granules and no distinct brush border with atypical microvilli. All morphological changes were improved by treating with sulforaphane. Finally, treatment with sulforaphane significantly increased expression of PGC-1, TFAM, Nrf2 and HO-1 associated with reduction in expression of mTOR, cyclin D1 and PCNA. Conclusion Sulforaphane could cure UC in rats. The protective activity can be explained by enhancing antioxidant activity, elevating mitochondrial biogenesis and inhibiting DNA polymerization.

Structure of the Harmonin PDZ2 and coiled-coil domains in a complex with CDHR2 tail and its implications.

Harmonin is a protein containing multiple PDZ domains and is required for the development and maintenance of hair cell stereocilia and brush border microvilli. Mutations in the USH1C gene can cause Usher syndrome type 1C, a severe inheritable disease characterized by the loss of hearing and vision. Here, by solving the high-resolution crystal structure of Harmonin PDZ2 and coiled-coil domains in a complex with the tail of cadherin-related family member 2, we demonstrated that mutations located in the Harmonin PDZ2 domain and found in patients could affect its stability, and thus, the target binding capability. The structure also implies that the coiled-coil domain could form antiparallel dimers under high concentrations, possibly when Harmonin underwent liquid-liquid phase separation in the upper tip-link density in hair cell stereocilia or microvilli of enterocytes of the intestinal epithelium. The crystal structure, together with the biochemical analysis, provided mechanistic implications for Harmonin mutations causing Usher syndrome, non-syndromic deafness, or enteropathy.

High phosphate-induced progressive proximal tubular injury is associated with the activation of Stat3/Kim-1 signaling pathway and macrophage recruitment.

Dietary phosphate intake in the Western population greatly exceeds the recommended dietary allowance and is linked to enhanced cardiovascular and all-cause mortality. It is unclear whether a chronic high phosphate diet (HPD) causes kidney injury in healthy individuals. Here, we show that feeding a 2% HPD in C57BL/6N mice for one up to six months resulted in hyperphosphatemia, hyperphosphaturia, increased plasma levels of fibroblast growth factor (FGF) 23, and parathyroid hormone (PTH) compared to mice on a 0.8% phosphate diet. Kidney injury was already noted after two months of HPD characterized by loss of proximal tubular (PT) cell polarity, flattened epithelia, disruption of brush border membranes, vacuolization, increased PT cell proliferation, marked interstitial mononuclear infiltration, and progressive accumulation of collagen fibers. HPD increased Stat3 activation and Kim-1 expression in PT epithelial cells and enhanced renal synthesis of chemokines recruiting monocytes and macrophages as well as macrophage related factors. Enhanced recruitment of F4/80+ macrophages around injured PT lesions was timely associated with increased Kim-1 synthesis, tubular MCP-1 expression, and degree of PT injury score. Likewise, tubulointerstitial fibrosis was associated with activation of Stat3/Kim-1 signaling pathway. The stimulation of human proximal tubular cells with high phosphate activated Stat3 signaling and induced HAVCR1 and CCL2 expression. We conclude that high phosphate results in progressive proximal tubular injury, indicating that high dietary phosphate intake may affect kidney health and therefore represents an underestimated health problem for the general population.

Design, Synthesis, and Biological Evaluation of a Novel Series of 4-Guanidinobenzoate Derivatives as Enteropeptidase Inhibitors with Low Systemic Exposure for the Treatment of Obesity.

To discover a novel series of potent inhibitors of enteropeptidase, a membrane-bound serine protease localized to the duodenal brush border, 4-guanidinobenzoate derivatives were evaluated with minimal systemic exposure. The 1c docking model enabled the installation of an additional carboxylic acid moiety to obtain an extra interaction with enteropeptidase, yielding 2a. The oral administration of 2a significantly elevated the fecal protein output, a pharmacodynamic marker, in diet-induced obese (DIO) mice, whereas subcutaneous administration did not change this parameter. Thus, systemic exposure of 2a was not required for its pharmacological effects. Further optimization focusing on the in vitro IC50 value and T1/2, an indicator of dissociation time, followed by enhanced in vivo pharmacological activity based on the ester stability of the compounds, revealed two series of potent enteropeptidase inhibitors, a dihydrobenzofuran analogue ((S)-5b, SCO-792) and phenylisoxazoline (6b), which exhibited potent anti-obesity effects despite their low systemic exposure following their oral administration to DIO rats.

ACE2 Promoted by STAT3 Activation Has a Protective Role in Early-Stage Acute Kidney Injury of Murine Sepsis.

Sepsis-induced AKI (SIAKI) is the most common complication with unacceptable mortality in hospitalized and critically ill patients. The pathophysiology of the development of SIAKI is still poorly understood. Our recent work has demonstrated the role of signal transducer and activator of transcription 3 (STAT3) pathways in regulating inflammation and coagulation in sepsis. We hypothesized that STAT3 activation has a critical role in early-stage SIAKI. The early-stage SIAKI model was established in cecal ligation and puncture (CLP) mice, which recapitulates the clinical and renal pathological features of early-stage AKI patients. Brush border loss (BBL) was the specific pathological feature of acute tubular injury in early-stage AKI. The role of STAT3 signaling and angiotension system in early-stage SIAKI was evaluated. The STAT3 activation (increased pSTAT3) and increased angiotensin-converting enzyme 2 (ACE2) expressions were observed in CLP mice. The low responsive expressions of pSTAT3 and ACE2 to septic inflammation in CLP AKI mice were associated with BBL. Correlation analysis of proteins' expressions showed pSTAT3 expression was significantly positively related to ACE2 expression in CLP mice. Reduced pSTAT3 after S3I201 intervention, which blocked STAT3 phosphorylation, decreased ACE2 expression, and exacerbated tubular injury in early-stage SIAKI. Our data indicate that endogenous increase of ACE2 expression upregulated by STAT3 activation in early-stage SIAKI play protective role against acute tubular injury.

Villin expression in human tumours: a tissue microarray study on 14,398 tumours

Background Villin is a protein of the brush border of epithelial cells which is used as an immunohistochemical marker for colorectal and gastrointestinal neoplasms. However, other tumor entities can also express villin. Methods To comprehensively determine villin expression, tissue microarrays containing 14,398 samples from 118 different tumor types as well as 608 samples of 76 different normal tissues were analyzed by immunohistochemistry. Results Villin was found in 54 of 118 tumor categories, including 36 tumor categories with strong staining. Villin expression was frequent in colorectal, upper gastrointestinal tract, pancreatobiliary, and renal tumors as well as in mucinous ovarian cancers, yolk sac tumors and in neuroendocrine neoplasms. Reduced villin expression was linked to advanced pT stage, lymph vessel invasion and microsatellite instability (p≤0.0006) in colorectal adenocarcinoma. In summary, our data demonstrate that villin expression is most common in gastrointestinal, pancreatobiliary, and neuroendocrine neoplasms, yolk sac tumors and mucinous ovarian cancers. Conclusion Our data support a high utility of villin immunohistochemistry for the identification of tumors with gastrointestinal, pancreatobiliary, and yolk sac tumor origin. However, considering that at least a weak villin positivity in some tumor cells occurred in 54 different tumor categories, villin immunohistochemistry should be applied as a part of a marker panel rather than as a stand-alone marker.

Effect of feeding calcium gluconate embedded in a hydrogenated fat matrix on feed intake, gastrointestinal fermentation and morphology, intestinal brush border enzyme activity and blood metabolites in growing lambs.

Gluconate salts have been identified as a butyrate precursor when fed to non-ruminant species and may increase the butyrate concentration in the large intestine supporting gastrointestinal health and development. The objective of this study was to evaluate the dose response of hydrogenated fat-embedded calcium gluconate (HFCG) on performance and gastrointestinal tract (GIT) development in growing lambs. Thirty-two wether lambs were used in a randomized complete block design and assigned to 1 of 4 treatments differing in the inclusion of HFCG: 0.0% (CON), 0.075% (LOW), 0.30% (MED), and 0.60% of the diet (HIGH). Lambs were allocated into individual pens and fed ad libitum with feed delivered twice daily. Feed intake was recorded daily, and body weight (BW) was assessed at the beginning and the end of the 29-d period. Blood was sampled on day 21, prior to feeding and 6 h post-feeding to evaluate changes in β-hydroxybutyrate, glucose, and insulin concentrations. Total fecal collection was conducted during days 25 to 28 to assess apparent total tract digestibility. On day 29, lambs were slaughtered, and the entire GIT was separated by region to enable sampling of tissue and digesta. Data were analyzed to assess linear, quadratic, and cubic effects of HFCG dose. Final BW, average daily gain, and dry matter intake decreased linearly (P ≤ 0.02) with increasing HFCG. Increasing inclusion of HFCG linearly decreased (P = 0.01) the thickness of the stratum corneum in ruminal papillae but did not affect other strata (P ≥ 0.34). Omasal digesta weight linearly decreased (P = 0.01) as the concentration of HFCG increased and abomasal digesta weight was cubically affected (P = 0.03) the increasing dose of HFCG. Short-chain fatty acid concentration in the cecum was cubically affected (P < 0.01) with increasing dose of HFCG where low dose had the greatest concentration. Moreover, increasing the dietary supply of HFCG linearly increased the proportion of acetate (P = 0.04) in the cecum and linearly decreased the proportion of propionate in the digesta of both the cecum (P < 0.01) and colon (P = 0.01). Colon crypt depth was quadratically (P = 0.03) affected with the increasing dose of HFCG, where lambs fed MED had greatest crypt depth. We conclude that feeding HFCG to growing lambs did not increase butyrate concentration in the large intestine and consequently does not increase the absorptive surface area of the whole tract, the size of the GIT, or the functionality of the intestine.

Evaluating the Effects of an Indian Medicinal Plant, Salacia oblonga on Intestinal Morphology, Functionality and the Microbiome In Vivo (Gallus gallus)

ObjectivesTo assess the changes associated with the dietary consumption of alpha-glucosidase and alpha-pancreatic amylase inhibitor, Salacia oblonga water extract (SOWE) on intestinal health in-vivo. MethodsWe utilized the intra-amniotic in-vivo (Gallus gallus) method to meet the objective. Five treatment groups viz. No Injection, 18 MΩ H2O, 5% Inulin, 2% SOWE, and 5% SOWE were created with n∼10 for each group. On day 17 of embryonic development 1 mL of the treatments were injected into the amnion of the eggs. On day 21 (upon hatch), blood and tissue samples were collected. Hemoglobin level, mRNA expression and morphological examination of the duodenum and the cecal microbiome were conducted in the different groups. ResultsThe histomorphology analysis revealed a significant decrease in villi surface area, goblet cell (number and diameter) and a reduction in Paneth cell number indicating negative side effects of the extract. There was a significant reduction in inflammatory proteins at 2% SOWE but not 5% SOWE. Finally, the 16s rRNA analysis revealed significant changes in alpha diversity between the groups. ConclusionsThis is the first study that assessed the effects of undigested complex carbohydrates on brush border membrane morphology, functionality, and the microbiome. The study reveals potential side effects of consuming carbohydrate enzyme inhibitors such as Salacia. Funding SourcesN/A.

Alterations in Intestinal Functionality and Bacterial Populations Following Intra-amniotic Administration (Gallus gallus) of Nicotinamide Riboside and Its Derivatives

ObjectivesThe primary aim of this study was to assess the effects of intra-amniotic administration of NRCl and two new derivatives (NRTBCl and NRTOCl) on BBM functionality via the evaluation of duodenal gene expression of BBM biomarkers, specifically essential digestive, absorptive and immune proteins, and the effects on cecal microbiome. MethodsWe synthesized and characterized nicotinamide riboside tributyrate chloride (NRTBCl, water-soluble) and nicotinamide riboside trioleate chloride (NRTOCl) as two new ester derivatives of nicotinamide riboside chloride (NRCl). NRCl and its derivatives were assessed in vivo (Gallus gallus), via intra-amniotic administration (day 17 of embryonic incubation), with the following treatment groups: (1) non-injected (control); and injection of (2) deionized H2O (control); (3) NRCl (30 mg/mL dose); (4) NRTBCl (30 mg/mL dose); and (5) NRTOCl (30 mg/mL dose). Upon injection, the effects on physiological markers involved in brush border membrane morphology, intestinal microbiota, and duodenal gene expression of key proteins were investigated. ResultsNRTBCl exposure significantly increased (p < 0.05) average cecum weight compared to other NR treatments. NR treatments significantly increased (p < 0.05) populations of Clostridium and the NRCl treatment demonstrated significant increased (p < 0.05) populations of Bifidobacterium, Lactobacillus, compared to the control groups (Non-injected and H2O). Duodenal gene expression analysis revealed that NRCl, NRTBCl, and NRTOCl treatments significantly upregulated (p < 0.05) the expression of ZnT1, MUC2, and IL6 compared to the controls, which suggests alterations in brush border membrane functionality. The administration of NRCl and its derivatives appears to trigger increased expression of digestive proteins and on the composition and function of cecal microbial populations. ConclusionsThis study is the first to demonstrate alterations in duodenal gene expression and cecal microbial profiles after the intra-amniotic administration of NR and its derivatives, which have been shown to improve microbial profiles through antimicrobial effects. Thus, the present study presents strong evidence that NR and its derivatives positively modulate the intestinal microbial profile, composition, and function. Funding SourcesNo external funding was utilized for the study.