Mouse Stomach Research Articles

PAN-selective inhibition of cAMP-phosphodiesterase 4 (PDE4) induces gastroparesis in mice.

Inhibitors of cAMP-phosphodiesterase 4 (PDE4) exert a number of promising therapeutic benefits, but adverse effects, in particular emesis and nausea, have curbed their clinical utility. Here, we show that PAN-selective inhibition of PDE4, but not inhibition of PDE3, causes a time- and dose-dependent accumulation of chow in the stomachs of mice fed ad libitum without changing the animals' food intake or the weight of their intestines, suggesting that PDE4 inhibition impairs gastric emptying. Indeed, PDE4 inhibition induced gastric retention in an acute model of gastric motility that traces the passage of a food bolus through the stomach over a 30minutes time period. In humans, abnormal gastric retention of food is known as gastroparesis, a syndrome predominated by nausea (>90% of cases) and vomiting (>80% of cases). We thus explored the abnormal gastric retention induced by PDE4 inhibition in mice under the premise that it may represent a useful correlate of emesis and nausea. Delayed gastric emptying was produced by structurally distinct PAN-PDE4 inhibitors including Rolipram, Piclamilast, Roflumilast, and RS25344, suggesting that it is a class effect. PDE4 inhibitors induced gastric retention at similar or below doses commonly used to induce therapeutic benefits (e.g., 0.04mg/kg Rolipram), thus mirroring the narrow therapeutic window of PDE4 inhibitors in humans. YM976, a PAN-PDE4 inhibitor that does not efficiently cross the blood-brain barrier, induced gastroparesis only at significantly higher doses (≥1mg/kg). This suggests that PDE4 inhibition may act in part through effects on the autonomic nervous system regulation of gastric emptying and that PDE4 inhibitors that are not brain-penetrant may have an improved safety profile. The PDE4 family comprises four subtypes, PDE4A, B, C, and D. Selective ablation of any of these subtypes in mice did not induce gastroparesis per se, nor did it protect from PAN-PDE4 inhibitor-induced gastroparesis, indicating that gastric retention may result from the concurrent inhibition of multiple PDE4s. Thus, potentially, any of the four PDE4 subtypes may be targeted individually for therapeutic benefits without inducing nausea or emesis. Acute gastric retention induced by PDE4 inhibition is alleviated by treatment with the widely used prokinetic Metoclopramide, suggesting a potential of this drug to alleviate the side effects of PDE4 inhibitors. Finally, given that the cause of gastroparesis remains largely idiopathic, our findings open the possibility that a physiologic or pathophysiologic downregulation of PDE4 activity/expression may be causative in a subset of patients.

Curcumin induces expression of 15-hydroxyprostaglandin dehydrogenase in gastric mucosal cells and mouse stomach in vivo: AP-1 as a potential target

15-Hydroxyprostaglandin dehydrogenase (15-PGDH) catalyzes the conversion of oncogenic prostaglandin E2 to non-tumerigenic 15-keto prostaglandin E2. In the present study, we found that curcumin, a yellow coloring agent present in the rhizome of Curcuma longa Linn (Zingiberaceae), induced expression of 15-PGDH at the both transcriptional and translational levels in normal rat gastric mucosal cells. By using deletion constructs of 15-PGDH promoter, we were able to demonstrate that activator protein-1 (AP-1) is the principal transcription factor responsible for regulating curcumin-induced 15-PGDH expression. Curcumin enhanced the expression of c-Jun and c-Fos that are functional subunits of AP-1, in the nuclear fraction of cells. Silencing of c-Jun suppressed curcumin-induced expression of 15-PGDH. Moreover, the chromatin immunoprecipitation assay revealed curcumin-induced binding of c-Jun to the AP-1 consensus sequence present in the 15-PGDH promoter. Curcumin increased phosphorylation of ERK1/2 and JNK, and pharmacologic inhibition of these kinases abrogated the curcumin-induced phosphorylation of c-Jun and 15-PGDH expression. In contrast, tetrahydrocurcumin which lacks the α,β-unsaturated carbonyl group failed to induce 15-PGDH expression, suggesting that the electrophilic carbonyl group of curcumin is essential for its induction of 15-PGDH expression. Curcumin restored the expression of 15-PGDH which is down-regulated by Helicobacter pylori through suppression of DNA methyltransferase 1. In addition, oral administration of curcumin increased the expression of 15-PGDH and its regulators such as p-ERK1/2, p-JNK, and c-Jun in the mouse stomach. Taken together, these findings suggest that curcumin-induced upregulation of 15-PGDH may contribute to chemopreventive effects of this phytochemical on inflammation-associated gastric carcinogenesis.

Polysaccharides from acerola, cashew apple, pineapple, mango and passion fruit co-products: Structure, cytotoxicity and gastroprotective effects

This study aimed to characterize the chemical structure of water-soluble polysaccharide fractions extracted from co-products of acerola, cashew apple, pineapple, mango and passion fruit, evaluating their gastroprotective effect in vivo associated to their antioxidant activity. The results from the structural analysis revealed that all fractions extracted from the co-products were heteropolysaccharides composed by the monosaccharides glucose, arabinose, galactose and d-galacturonic acid, and presented a pectic nature, which was evidenced by the predominant presence of arabinogalactan in their constitutions. The polysaccharides obtained from acerola, cashew apple and pineapple exhibited similarity in their morphology compared to those from mango and passion fruit. The fractions did not show cytotoxic effects for epithelial culture IEC-6A, and stimulated cell growth as visualized by the increase in cellular viability. Notable antioxidant activities were found for cashew apple (147 μM Trolox.g−1), pineapple (80.0 μM Trolox.g−1) and acerola (79.0 μM Trolox.g−1). These same polysaccharides promoted higher preservation of glutathione levels in mice stomach tissue, resulting in its effective protection from the damage caused by ethanol. On the other hand, high malondialdehyde levels were observed in the stomach tissues of the animals treated with the polysaccharides from mango and passion fruit. The co-products from acerola and cashew were able to protect the gastric mucosa of mice from the lesions induced by ethanol. The results found in this research introduce polysaccharide fractions extracted from acerola, cashew apple and pineapple co-products as safe and natural antioxidants, with a high potential to be applied in different industry fields.

Nutrient-sensing components of the mouse stomach and the gastric ghrelin cell.

The ability of the gut to detect nutrients is critical to the regulation of gut hormone secretion, food intake, and postprandial blood glucose control. Ingested nutrients are detected by specific gut chemosensors. However, knowledge of these chemosensors has primarily been derived from the intestine, while available information on gastric chemosensors is limited. This study aimed to investigate the nutrient-sensing repertoire of the mouse stomach with particular emphasis on ghrelin cells. Quantitative RT-PCR was used to determine mRNA levels of nutrient chemosensors (protein: G protein-coupled receptor 93 [GPR93], calcium-sensing receptor [CaSR], metabotropic glutamate receptor type 4 [mGluR4]; fatty acids: CD36, FFAR2&4; sweet/umami taste: T1R3), taste transduction components (TRPM5, GNAT2&3), and ghrelin and ghrelin-processing enzymes (PC1/3, ghrelin O-acyltransferase [GOAT]) in the gastric corpus and antrum of adult male C57BL/6 mice. Immunohistochemistry was performed to assess protein expression of chemosensors (GPR93, T1R3, CD36, and FFAR4) and their co-localization with ghrelin. Most nutrient chemosensors had higher mRNA levels in the antrum compared to the corpus, except for CD36, GNAT2, ghrelin, and GOAT. Similar regional distribution was observed at the protein level. At least 60% of ghrelin-positive cells expressed T1R3 and FFAR4, and over 80% expressed GPR93 and CD36. The cellular mechanisms for the detection of nutrients are expressed in a region-specific manner in the mouse stomach and gastric ghrelin cells. These gastric nutrient chemosensors may play a role modulating gastrointestinal responses, such as the inhibition of ghrelin secretion following food intake.

Histological Effect of Aspirin on the Stomach of Male Albino Swiss Mice (Mus musculus)

The present study aimed to determine the impacts of Aspirin drug on the stomach of albino Swiss mice. The study sample included 10 male mice divided into 2 groups. The first group was orally administered with 0.1 mL of 0.75 mg/kg aspirin once daily, whereas the second group, the control, was treated with similar doses of distilled water. Following 60 days of successive treatment, a number of parameters was studied including difference in body weight and histopathological changes in the stomach as diagnosed after histological preparation. The results showed a significant decrease (p<0.05) in body weight average of the treated mice compared with the control group. The results also revealed the occurrence of several histopathological changes in the stomach of treated animals, including raptures in the epithelium lining, congestion in the muscular layer, and vasodilation. In addition, alterations such as hemorrhage, pyknosis in muscle cells, sloughing of the lining of the muscular layer, and external bleeding were observed. It can be concluded from these results that orally administered aspirin exerts negative effects on the stomach in mice.

Salvia miltiorrhiza-Derived Sal-miR-58 Induces Autophagy and Attenuates Inflammation in Vascular Smooth Muscle Cells

Autophagy is associated with the cytoprotection of physiological processes against inflammation and oxidative stress. Salvia miltiorrhiza possesses cardiovascular protective actions and has powerful anti-oxidative and anti-inflammatory effects; however, whether and how Salvia miltiorrhiza-derived microRNAs (miRNAs) protect vascular smooth muscle cells (VSMCs) by inducing autophagy across species are unknown. We first screened and identified Sal-miR-58 from Salvia miltiorrhiza as a natural autophagy inducer. Synthetic Sal-miR-58 suppresses chronic angiotensin II (Ang II) infusion-induced abdominal aortic aneurysm (AAA) formation in mice, as well as induces autophagy in VSMCs and attenuates the inflammatory response elicited by Ang II in vivo and in vitro. Mechanistically, Sal-miR-58 downregulates Krüppel-like factor 3 (KLF3) expression through direct binding to the 3′ UTR of KLF3, which in turn relieves KLF3 repression of E3 ubiquitin ligase neural precursor cell-expressed developmentally downregulated 4-like (NEDD4L) expression, whereas NEDD4L upregulation increases the ubiquitination and degradation of the platelet isoform of phosphofructokinase (PFKP), subsequently leading to a decrease in the activation of Akt/mammalian target of rapamycin (mTOR) signaling and facilitating VSMC autophagy induced by Sal-miR-58 in the context of chronic Ang II stimulation and aneurysm formation. Our results provide the first evidence that plant-derived Sal-miR-58 induces autophagy and attenuates inflammation in VSMCs through cross-species modulation of the KLF3/NEDD4L/PFKP regulatory pathway.

Comparative acute toxicity of supramolecular complexes of Fenbendazole with Different Polymers used for targeted Delivery

The purpose of the research is studying acute toxicity of supramolecular complexes of fenbendazole with different polymers obtained by mechanochemical technology as compared to the substance of the drug when administered to the stomach of white mice. Materials and methods. The acute toxicity parameters of supramolecular complexes of fenbendazole (SMCF) with different polymers were studied under the “Guide to experimental (preclinical) study of new pharmacological substances” and other rules. The drugs were administered once to the stomach of white mice of both genders with a body weight of 16–18 g at doses of 7,000, 10,000, 15,000 and 20,000 mg/kg by the drug. Each dose was tested on 10 white mice. For 14 days, we observed the general condition and behavior of animals, symptoms of intoxication and possible death. Postmortem examination was performed for the dead animals. Acute toxicity was determined by Litchfield and Wilcoxon (M. D. Belenky, 1963). Results and discussion. LD50 of the SMCF and the major drug of fenbendazole could not be established due to low toxicity. LD50 of the SMCF administered to the stomach of white mice was 20 000 mg/kg.

Transcriptomic landscape profiling of metformin-treated healthy mice: Implication for potential hypertension risk when prophylactically used.

Recently, the first‐line anti‐diabetic drug metformin shows versatile protective effects against several diseases and is potentially prescribed to healthy individual for prophylactic use against ageing or other pathophysiological processes. However, for healthy individuals, it remains unclear what effects metformin treatment will induce on their bodies. A systematic profiling of the molecular landscape of metformin treatment is expected to provide crucial implications for this issue. Here, we delineated the first transcriptomic landscape induced by metformin in 10 tissues (aorta, brown adipose, brain, eye, heart, liver, kidney, skeletal muscle, stomach and testis) of healthy mice by using RNA‐sequencing technique. A comprehensive computational analysis was performed. The overrepresentation of cardiovascular disease‐related gene sets, positive correlation with hypertension‐related transcriptomic signatures and the associations of drugs with hypertensive side effect together indicate that although metformin does exert various beneficial effects, it would also increase the risk of hypertension in healthy mice. This prediction was experimentally validated by an independent animal experiments. Together, this study provided important resource necessary for investigating metformin's beneficial/deleterious effects on various healthy tissues, when it is potentially prescribed to healthy individual for prophylactic use.

Sorbaria kirilowii Ethanol Extract Exerts Anti-Inflammatory Effects In Vitro and In Vivo by Targeting Src/Nuclear Factor (NF)-κB.

Inflammation is a fundamental process for defending against foreign antigens that involves various transcriptional regulatory processes as well as molecular signaling pathways. Despite its protective roles in the human body, the activation of inflammation may also convey various diseases including autoimmune disease and cancer. Sorbaria kirilowii is a plant originating from Asia, with no anti-inflammatory activity reported. In this paper, we discovered an anti-inflammatory effect of S. kirilowii ethanol extract (Sk-EE) both in vivo and in vitro. In vitro effects of Sk-EE were determined with lipopolysaccharide (LPS)-stimulated RAW264.7 cells, while ex vivo analysis was performed using peritoneal macrophages of thioglycollate (TG)-induced mice. Sk-EE significantly reduced the nitric oxide (NO) production of induced macrophages and inhibited the expression of inflammation-related cytokines and the activation of transcription factors. Moreover, treatment with Sk-EE also decreased the activation of proteins involved in nuclear factor (NF)-κB signaling cascade; among them, Src was a prime target of Sk-EE. For in vivo assessment of the anti-inflammatory effect of Sk-EE, HCl/EtOH was given by the oral route to mice for gastritis induction. Sk-EE injection dose-dependently reduced the inflammatory lesion area of the stomach in gastritis-induced mice. Taking these results together, Sk-EE exerts its anti-inflammatory activity by regulating intracellular NF-κB signaling pathways and also shows an authentic effect on reducing gastric inflammation.

Chronic Restraint Stress Induces Gastric Mucosal Inflammation with Enhanced Oxidative Stress in a Murine Model.

BackgroundAlthough the underlying mechanisms of chronic stress are still unknown, this condition has been related to the pathophysiology of gastric mucosal inflammation, whose development is accelerated by oxidative stress. The present study investigates how chronic stress influences gastric mucosal oxidative stress and inflammation.MethodsEight-week-old C57BL/6J male mice were subjected to two-week intermittent restraint stress. The expressions of CD11b (a specific for monocyte/macrophage), monocyte/macrophage cell surface markers (CD68 and F4/80), NADPH oxidase-4 (Nox-4) and 8-hydroxy-2’-deoxyguanosine (8-OHdG, a sensitive biomarker of oxidative stress) were determined using immunohistochemistry, RT-PCR, and enzyme-linked immunosorbent assay, respectively. The expressions of antioxidant enzymes, such as superoxide dismutase, catalase, and glutathione peroxidase, were examined by RT-PCR and Western blotting. The expressions of proinflammatory cytokines, including monocyte chemoattractant protein-1 (MCP-1), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α), were determined using immunohistochemistry and RT-PCR, respectively.ResultsChronic stress increased the lymphocytic infiltration and inflammation within the gastric mucosa of mice. Stress remarkably increased the expression levels of CD11b and mRNA expression levels of CD68 and F4/80 in the mucosa of the stomach of stressed mice. Stress remarkably increased both mRNA and plasma concentrations of Nox-4 and 8-OHdG; and markedly reduced gastric mRNA and protein expression levels of antioxidant enzymes such as superoxide dismutase, catalase, and glutathione peroxidase. The expressions of proinflammatory cytokines (MCP-1, IL-1β, and TNF-α) were predominantly observed in the gastric mucosal layers of the stressed mice. Furthermore, stress remarkably elevated the gastric mucosal mRNA expression levels of MCP-1, IL-1β, and TNF-α.ConclusionTwo weeks of restraint stress induced gastric inflammation in the murine model with enhanced oxidative stress and reduced anti-oxidative system.

Optimizing the 1525 Nm Emission of Er3+-Sensitized Nanoprobe for In Vivo NIR-IIb Luminescence Bioimaging

Introduction Fluorophores emitting in the second near-infrared window (NIR-II, 900-1700 nm)[1] have exhibited significant improvements in detection depth and resolution owing to low tissue autofluorescence and up to a 1000-fold reduction in scattering losses compared to those for visible (400-780 nm)[2] and NIR-I regions (780-900 nm)[3]. Especially, the range of 1500-1700 nm is denoted as the “NIR-IIb” region, which provides the lowest photon scattering in the NIR-II region, thus allowing for the most desirable imaging clarity and deep penetration using existing InGaAs detectors [4]. Unfortunately, NIR-IIb fluorescent probes with sufficient brightness are scarce. Herein, we present an efficient NIR-IIb imaging agent NaErF4:0.5%Tm3+@NaLuF4 with a shell thickness of 4.30 nm. The influence of the types of epitaxial heterogeneous shells, the doping effect and optimal doping concentration of Tm3+ ions, as well as the critical shell thickness for obtaining the surface quenching-assisted downshifting were systematically investigated to acquire efficient 1525 nm emission of Er3+ ions under 800 nm excitation. The quantum yield in 1500-1700 nm region reached to 13.92%, enabling high-resolution through-skull microscopic cerebrovascular imaging and large-depth in vivo gastrointestinal tract imaging with extremely low excitation powder density of 35mW/cm2. Method The 793 nm laser beam emitted by the semiconductor laser (Suzhou Rugkuta Optoelectronics Co., Ltd., China) was collimated and expanded through the lens so that the sample can be uniformly illuminated. The emitting fluorescent signal was collected onto the InGaAs camera (Tekwin, China) through a prime lens (focal length: 50 mm, Edmund Optics). A 1450 nm long-pass filter (Thorlabs) was placed in front of the detector to extract the pure NIR-IIb signals. After anesthesia, the mice were placed on the imaging platform. The PEGylated β-NaErF4:0.5%Tm@4.30nm NaLuF4 nanoparticles (200 μL, 5 mg/mL) was injected into the tail vein, then the laser was turned on immediately and the image was recorded at different time points.The β-NaErF4:0.5%Tm@4.30nm NaLuF4 nanoparticles (200 μL, 5 mg/mL) were perfused into the stomach of nude mice. The images of the mice were recorded using the above-mentioned NIR-IIb fluorescence macroscopic imaging system. Results and Conclusions The brightest β-NaErF4:0.5%Tm@4.30nm NaLuF4 nanoparticels (NPs) were used for the in vivo bio-imaging. Firstly, 200 μL NPs (PEGylated β-NaErF4:0.5%Tm@4.30nm NaLuF4, 5mg/mL) were intravenously injected into the mouse for the whole body imaging (Fig.1), which verified the feasibility of the probe for NIR-IIb in vivo imaging. Under the excitation of a 793 nm laser (~35 mW/cm2), a bright, clear vascular network in the nearly “transparent” body was presented. Within 5 min post-injection, it could be seen that the liver has been gradually brighter, for which the NPs were captured by the liver (Fig.1B). Thanks to the low autofluorescence and suppressed tissue scattering in NIR-IIb region, a clear outline of liver was distinguished. It is worth noticing that, owing to the excellent luminescence properties of the probe, the excitation power density at the imaging plane could be as low as ~35 mW/cm2, which is much smaller than that in previously reported work.[5] The deep-tissue penetration in the NIR-IIb region was well validated in Fig. 2. After anesthesia, a mouse with a cranial window and a mouse with intact skull were injected with PEGylated β-NaErF4:0.5%Tm@4.30nm NaLuF4 NPs (200 μL, 100 mg/mL) respectively and fixed in the imaging system for microscopic cerebrovascular imaging. The 793 nm laser beam was expanded, reflected, and uniformly illuminated in the region of interest. The NIR-IIb fluorescence signals emitted by the NPs in the blood vessel were collected by the objective lens and focused by the tube lens onto the InGaAs detector (Tekwin, China). In order to visually show the structure of the blood vessels in the mouse brain, we stitched several images recorded by a scan lens (LSM03, Thorlabs) into an image with a large field of view. Compared to open-skull brain imaging (Fig.2A), NIR-IIb through-skull imaging (Fig.2B) also provided fairly rich details, and a small blood vessel with the diameter of 18.609 μm was easily identified.

Helicobacter suis Infection in Mouse Induced not Only Gastric, but Hepatic and Pulmonary MALT Lymphoma: Relation to Substance P.

The hepatic and pulmonary MALT lymphoma (mucosa-associated lymphoid tissue lymphoma) is clinically occasionally observed but its pathogenesis is unknown and thought to be important to establish the treatment strategy. The present study was designed to clarify the characteristics of these lymphomas and the effect of the Helicobacter eradication regimen and substance P antagonist. After the long term infection of Helicobacter suis to the C57BL/6 mice stomach, the whole organ was surveyed pathologically. Histochemical characteristics of the lesion and the localization of bacteria were observed. In addition, the effect of the administration of antibiotics and a proton pump inhibitor or the substance P antagonist was investigated. We have detected the hepatic and pulmonary MALT lymphoma after the long term infection. In situ hybridization study revealed the positive reaction of Helicobacter suis in the hepatic and pulmonary MALT lymphoma. After the administration of antibiotics and a proton pump inhibitor, the bacterial number has significantly decreased and the tumor size in the fundus, liver and lung markedly reduced. Substance P immunoreactivity was clearly shown in the lymphoma cells in the liver and lung, and the spantide II administration induced the marked decrease in the size of tumors. By our experiments using the long term infection of Helicobacter suis to the C57BL/6 mice, we have detected the liver and pulmonary MALT lymphoma. In situ hybridization study suggested the direct interaction of this bacterium to the etiology of these lesions. Substance P within the lymphoma cells was suggested to work on the maintenance of the extragastric MALT lymphoma.

Pan-enteric neuropathy and dysmotility are present in a mouse model of short-segment Hirschsprung disease and may contribute to post-pullthrough morbidity

PurposeHirschsprung disease (HSCR) is characterized by distal intestinal aganglionosis. While surgery is lifesaving, gastrointestinal (GI) motility disorders persist in many patients. Our objective was to determine whether enteric nervous system (ENS) abnormalities exist in the ganglionated portions of the GI tract far proximal to the aganglionic region and whether these are associated with GI dysmotility. MethodsUsing Ednrb-null mice, a model of HSCR, immunohistochemical analysis was performed to evaluate quantitatively ENS structure in proximal colon, small intestine, and stomach. Gastric emptying and intestinal transit were measured in vivo and small and large bowel contractility was assessed by spatiotemporal mapping ex vivo. ResultsProximal colon of HSCR mice had smaller ganglia and decreased neuronal fiber density, along with a marked reduction in migrating motor complexes. The distal small intestine exhibited significantly fewer ganglia and decreased neuronal fiber density, and this was associated with delayed small intestinal transit time. Finally, in the stomach of HSCR mice, enteric neuronal packing density was increased and gastric emptying was faster. ConclusionsENS abnormalities and motility defects are present throughout the ganglionated portions of the GI tract in Ednrb-deficient mice. This may explain the GI morbidity that often occurs following pull-through surgery for HSCR.

Sex‐Differences in Gastric Smooth Muscle Function: Role of G Protein‐Coupled Estrogen Receptor

The regulation of gastrointestinal (GI) motility by estrogen and its receptors has been a topic of great interest owing to sexual dimorphism that exists in motility disorders. Scientific and clinical evidence establishes the fact of GI motility disturbances during estrous cycle but, there is a gap in the knowledge and understanding of estrogen and its receptors role in GI motility physiology during these cycles. Estrogen activates classical nuclear receptors ERα, ERβ and atypical 7‐transmembrane G protein‐coupled estrogen receptor (GPER or GPR30) to initiate both genomic and non‐genomic effects. GPER is involved in the several rapid cellular signaling events. The pleiotropic effects manifested by GPER action include regulation of body weight, glucose homeostasis, anti‐inflammation, anti‐nociception, and vascular muscle relaxation. Ablation of GPER leads to increase in mean arterial pressure in female mice, whereas treatment with GPER‐specific agonist G‐1 lowers arterial blood pressure in hypertensive and ovariectomized female rats. The role of GPER in mediating estrogen effects on GI smooth muscle is unknown.AimTo understand the role of GPER in gastric smooth muscle function in male and female mice.MethodsMuscle strips and muscle cells were isolated from the stomachs of control and GPER knockout mice. Expression of GPER was analyzed by qRT‐PCR and western blot. cAMP levels in response to estrogen and G‐1 was measured by radioimmunoassay. Relaxation was measured in organ bath studies in muscle strips and by scanning micrometry in isolated muscle cells.ResultsExpression of GPER (mRNA and protein) in the gastric smooth muscle was higher in female mice compared to male mice. 17β‐estradiol and G‐1 caused an increase in cAMP levels and inhibited acetylcholine‐induced muscle contraction (i.e., caused muscle relaxation) in gastric muscle strips and isolated muscle cells from both female and male mice, however, the increase in cAMP levels and the amount of relaxation was significantly greater in gastric muscle from female mice compared to male mice. The effect of 17β‐estradiol and G‐1 on muscle relaxation was concentration‐dependent. Relaxation was blocked by the GPER specific antagonist G15 (10 μM) in gastric muscle from control mice and absent in gastric muscle from GPER knockout mice. Relaxation was higher in proestrus and estrus phases compared to diestrus phase in female mice and this is consistent with higher expression of GPER in proestrus and estrus phases compared to diestrus phase.ConclusionActivation of GPER causes relaxation via cAMP/PKA pathway in gastric smooth muscle from female and male mice. Greater relaxation in gastric smooth muscle from female mice is due to higher expression of GPER in female mice compared to male mice. Pilot studies suggests that higher expression of GPER in the stomach of female mice is due to differential regulation of GPER expression by epigenetic mechanisms.Support or Funding InformationSupported by grants from VETAR (SM); IACUC approval AD10001201

Nonselective Inhibition of PDE4 Induces Gastroparesis in Mice

PurposeNon/PAN‐selective inhibitors of Type 4 phosphodiesterases (PDE4s), a group of isoenzymes that hydrolyze the second messenger cAMP, exert promising therapeutic benefits, such as anti‐inflammatory as well as cognition‐ and memory‐enhancing effects. However, these drugs also induce several side effects, in particular emesis and nausea, curbing their clinical use. The PDE4 family consists of four subtypes/genes (PDE4A to D). As each plays unique and non‐overlapping roles in the body, targeting individual PDE4 subtypes is a promising approach to separate the therapeutic benefits from the side effects of current PDE4 inhibitors. To this end, this study aimed to identify the specific PDE4s involved in emesis and nausea. While mice are anatomically unable to vomit, PDE4 inhibition in mice induces gastroparesis, an abnormal retention of food in the stomach, which is a well‐known cause of nausea and vomiting in humans and was thus used as a correlate.MethodsGastroparesis in knock‐out mice deficient in individual PDE4 subtypes and in mice treated with PAN‐PDE4 inhibitors was compared to their respective controls. We used a long term model, in which food accumulated in the stomach of mice fed ad libitum for 3 days, as well as an acute model, in which fasted mice were given a FITC‐labeled food bolus by oral gavage, euthanized 30 min later, and FITC/food‐retention in the stomach was then measured.ResultsTreatment of wildtype mice with PAN‐PDE4 inhibitors increased stomach sizes to twice that of controls without changing their food intake or the weight of their intestines, suggesting a specific effect of PDE4 inhibition in restricting gastric emptying. Structurally distinct PDE4 inhibitors all induced gastroparesis, indicating it is a class effect. Conversely, YM976, an inhibitor which does not cross the blood‐brain‐barrier, did not induce gastroparesis suggesting that PDE4 inhibitor‐induced gastroparesis results from altered neuronal regulation, rather than from PDE4 inhibition in the stomach/periphery. Ablation of PDE4A, B, C or D in the respective KO mice did not induce, nor did it protect the animals from PAN‐PDE4 inhibitor‐induced gastroparesis, implying that it results from the concurrent inhibition of multiple PDE4s.ConclusionsSelective ablation of individual PDE4 subtypes does not induce gastroparesis in mice. Thus, potentially, any of the four PDE4 subtypes may be targeted individually for therapeutic benefits without inducing nausea or emesis. In addition, PDE4 inhibitors that do not cross the blood‐brain barrier may also be free of these side effects.Support or Funding InformationSupported by grants from the NIH (HL76125, HL141473, HL066299).

Dynamic characterization of intestinal metaplasia in the gastric corpus mucosa of Atp4a-deficient mice.

Parietal cells of the gastric mucosa contain a complex and extensive secretory membrane system that harbors gastric H+, K+-adenosine triphosphatase (ATPase), the enzyme primarily responsible for gastric lumen acidification. Here, we describe the characterization of mice deficient in the H+, K+-ATPase α subunit (Atp4a−/−) to determine the role of this protein in the biosynthesis of this membrane system and the biology of the gastric mucosa. Atp4a−/− mice were produced by gene targeting. Wild-type (WT) and Atp4a−/− mice, paired for age, were examined at 10, 12, 14 and 16 weeks for histopathology, and the expression of mucin 2 (MUC2), α-methylacyl-CoA racemase (AMACR), Ki-67 and p53 proteins was analyzed by immunohistochemistry. For further information, phosphoinositide 3-kinase (PI3K), phosphorylated-protein kinase B (p-AKT), mechanistic target of rapamycin (mTOR), hypoxia-inducible factor 1α (HIF-1α), lactate dehydrogenase A (LDHA) and sirtuin 6 (SIRT6) were detected by Western blotting. Compared with the WT mice, hypochlorhydric Atp4a−/− mice developed parietal cell atrophy and significant antral inflammation (lymphocyte infiltration) and intestinal metaplasia (IM) with elevated MUC2 expression. Areas of dysplasia in the Atp4a−/− mouse stomach showed increased AMACR and Ki-67 expression. Consistent with elevated antral proliferation, tissue isolated from Atp4a−/− mice showed elevated p53 expression. Next, we examined the mechanism by which the deficiency of the H+, K+-ATPase α subunit has an effect on the gastric mucosa. We found that the expression of phosphorylated-PI3K, p-AKT, phosphorylated-mTOR, HIF-1α, LDHA and SIRT6 was significantly higher in tissue from the Atp4a−/− mice compared with the WT mice (P<0.05). The H+, K+-ATPase α subunit is required for acid-secretory activity of parietal cells in vivo, the normal development and cellular homeostasis of the gastric mucosa, and attainment of the normal structure of the secretory membranes. Chronic achlorhydria and hypergastrinemia in aged Atp4a−/− mice produced progressive hyperplasia and mucolytic and IM, and activated the Warburg effect via PI3K/AKT/mTOR signaling.

High-Salt Diet-Induced Gastritis in C57BL/6 Mice is Associated with Microbial Dysbiosis and Alleviated by a Buckwheat Diet.

A high-salt diet is a cause of gastritis, but the associated mechanism remains unclear. Recent studies have shown that gastric flora is associated with a variety of stomach diseases, but it is not known whether gastric flora is involved in gastritis induced by a high-salt diet. Gastritis is successfully induced in C57BL/6 mice fed a high-salt diet (salt: 5% NaCl) for four weeks. Through 16S rRNA gene sequencing, the composition of the stomach microbiota of mice fed normal and high-salt diets are compared, the results of which show that the high-salt diet induces significant changes in the gastric flora. Phylogenetic investigation of communities by reconstruction of unobserved states (PICRUSt) is used to predict the function of the microbiota in the stomach of mice, and the results indicate that a high-salt diet leads to a decrease in the ability of the gastric microbiota to metabolize polysaccharides and vitamins. A buckwheat diet is used to treat gastritis. The results show gastritis induced by the high-salt diet is significantly alleviated, and the dysbiosis in the stomach also improved. Buckwheat diet may be one of the ways to prevent and treat gastritis caused by a high-salt diet.

Distinct Cell Types With the Bitter Receptor Tas2r126 in Different Compartments of the Stomach.

Cells expressing bitter taste receptors (T2Rs or Tas2rs) in extraoral tissues are considered to be chemosensory cells mediating protective responses to potentially harmful or even antiinflammatory or antimicrobial compounds. In a previous study the activity of the Tas2R143/Tas2R135/Tas2r126 cluster promoter in the stomach was monitored using a Cre-reporter mouse line. Reporter gene expression and Tas2r126 mRNA were found in brush cells located at the distal wall of the gastric groove. In this study, we explored whether brush cells and epithelial cells of the stomach in fact contain the Tas2r126 receptor protein. Using immunohistochemistry, we demonstrate the presence of Tas2r126 immunoreactivity in different cell populations in the glandular stomach, in a subset of brush cells at the gastric groove and in unique glandular units as well as in certain enteroendocrine cells. In brush cells at the gastric groove, a strong immunofluorescence signal for the Tas2r126 receptor was observed at the most apical region of the cells, i.e., the microvillar tuft. In addition, we found a high density of Tas2r126-positive brush cells in the unique glandular units. These invaginations are located distally to the groove, open directly into the furrow and are enwrapped by smoothelin-immunoreactive muscles. In the corpus, Tas2r126 immunoreactivity was found in histamine-producing ECL cells and in ghrelin-producing X/A-like cells, the main enteroendcrine cells of this compartment. In the antrum, Tas2r126 labeling was observed in serotonin-storing EC cells and ghrelin cells, both representing only minor populations of enteroendocrine cells in this compartment. In conclusion, our data provide evidence for the presence of the Tas2r126 receptor protein in distinct cell types in the epithelium lining the mouse stomach which render the stomach responsive to agonists for bitter receptors.

The Effects of Newcastle virus Histologically Suspension on Some Organs of White Mice (Balb /c)

Newcastle disease is a contagious bird disease affecting wild avian species and many domestic,it may be transmissible to humans. Detection of Newcasstle virus confirmed by rapid test technique(Immunochromatography) . ten sample (10% ) out of 100 fasces sample collected from chickens were posative. Twelve mice used in this study devided into two groups ,first group consist of six mice induced with 0.2 ml from one positive sample of Newcasttle virus suspension to evaluate some histopathological changes caused by Newcasstle virus in small intestine and stomach of mice,second group induced with 0.2 ml from phosphate buffer saline only. The results revealed that histopathological changes in liver induced by using 0.2 ml from the positive Newcasstle virus suspension as hyperpalsia in kufper cells. Lung of mice treated with this concentration of Newcasstle virus has partial pnemonia and thicking the intra alveolar walls. While brain of mice treated with same concentration suffering from increase in number of glial cells ( glialosis ) and apoptosis in brain cells.

Molecular Alterations in the Stomach of Tff1-Deficient Mice: Early Steps in Antral Carcinogenesis.

TFF1 is a peptide of the gastric mucosa co-secreted with the mucin MUC5AC. It plays a key role in gastric mucosal protection and repair. Tff1-deficient (Tff1KO) mice obligatorily develop antropyloric adenoma and about 30% progress to carcinomas. Thus, these mice represent a model for gastric tumorigenesis. Here, we compared the expression of selected genes in Tff1KO mice and the corresponding wild-type animals (RT-PCR analyses). Furthermore, we systematically investigated the different molecular forms of Tff1 and its heterodimer partner gastrokine-2 (Gkn2) in the stomach (Western blot analyses). As a hallmark, a large portion of murine Tff1 occurs in a monomeric form. This is unexpected because of its odd number of seven cysteine residues. Probably the three conserved acid amino acid residues (EEE) flanking the 7th cysteine residue allow monomeric secretion. As a consequence, the free thiol of monomeric Tff1 could have a protective scavenger function, e.g., for reactive oxygen/nitrogen species. Furthermore, a minor subset of Tff1 forms a disulfide-linked heterodimer with IgG Fc binding protein (Fcgbp). Of special note, in Tff1KO animals a homodimeric form of Gkn2 was observed. In addition, Tff1KO animals showed strongly reduced Tff2 transcript and protein levels, which might explain their increased sensitivity to Helicobacter pylori infection.