Gastric Smooth Muscle Cells Research Articles

Rotigaptide inhibits spontaneous contractions of gastric smooth muscle in diabetic rats via the PKCα-Cx43 pathway.

The study aimed to investigate the effect of rotigaptide (ZP123) on spontaneous contractions of gastric smooth muscle in diabetic rats and explore the underlying mechanisms. Twelve rats were randomly divided into model and normal control groups. Changes in gastric smooth muscle spontaneous contractions in each group were observed. Western blot analysis was performed to detect Cx43 and PKCα expression. Rat gastric smooth muscle cells were cultured in vitro and divided into normal glucose, high glucose and high glucose+rotigaptide group. The intracellular Ca2+ content was observed by immunofluorescence. The amplitude and frequency of gastric smooth muscle spontaneous contractions were reduced in the model group than the normal control group (all p < .01), which were reduced after rotigatide treatment than before treatment in the model group (all p < .01). The model+rotigaptide group showed decreased membrane expression of Cx43, increased cytoplasmic expression of Cx43, increased membrane expression of p-PKCα Thr497 and lower membrane/cytoplasm ratio of Cx43 expression compared with the model group (all p < .01). The intracellular Ca2+ content was increased in the high glucose group than the normal glucose group (p < .01), while no significant difference was observed between the high glucose+rotigaptide and high glucose groups. Our findings suggest that rotigatide can stabilize the intracellular Ca2+ concentration in gastric smooth muscle cells under high glucose condition by upregulating PKCα activity and downregulating the number of GJs and the opening rate of GJ hemichannels through the PKCα-Cx43 pathway, thus inhibiting spontaneous contractions of gastric smooth muscle in diabetic rats.

Low-intensity pulsed ultrasound combined with ST36 modulate gastric smooth muscle contractile marker expression via RhoA/Rock and MALAT1/miR-449a/DLL1 signaling in diabetic rats.

Low-intensity pulsed ultrasound (LIPUS) combined with acupoint can promote gastric motility of diabetic rats. The switch of gastric smooth muscle cell (GSMCs) phenotype was related to the diabetes-induced gastric dysfunction, but the mechanism is not clearly elucidated. This study was aimed at exploring the underlying mechanism of LIPUS stimulation application in diabetic gastroparesis rats. In this study, Sprague-Dawley male rats were divided into three groups: control group (CON), diabetic gastroparesis group (DGP), and LIPUS-treated group (LIPUS). LIPUS irradiation was performed bilaterally at ST36 for 20 min per day for 4 weeks. The gastric emptying rate was measured by ultrasound examination. Contraction ability of GSMCs was assessed by muscle strip experiment. The expression of related proteins or mRNAs including α-SMA, SM22α, MHC, RhoA, Rock2, p-MYPT1, MYPT1, p-MLC, MLC, MALAT1, miR-449a, and DLL1 was detected by different methods such as western blotting, RT-qPCR, immunohistochemistry, and immunofluorescence staining, as appropriate. (a) LIPUS stimulation at ST36 could improve the gastric motility dysfunction of diabetic rats. (b) LIPUS increased RhoA, Rock2, p-MYPT1, and p-MLC expression level. (c) MALAT1 and DLL1 contents were decreased, but the level of miR-449a was increased in the LIPUS group. LIPUS may affect the contractile marker expression of gastric smooth muscle through the RhoA/Rock and MALAT1/miR-449a/DLL1 pathway to ameliorate DGP.

Involvement of oxidative stress-AMPK-Cx43-NLRP3 pathway in extracellular matrix remodeling of gastric smooth muscle cells in rats with diabetic gastroparesis.

This study aimed to investigate the changes in oxidative stress, adenosine monophosphate-activated protein kinase (AMPK), connexin43 (Cx43), nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) expression, and extracellular matrix (ECM) in the gastric smooth muscle tissues of rats with diabetic gastroparesis (DGP) and high glucose-cultured gastric smooth muscle cells, determine the existence of oxidative stress-AMPK-Cx43-NLRP3 pathway under high glucose condition, and the involvement of this pathway in ECM remodeling in DGP rats. The results showed that with increasing duration of diabetes, oxidation stress levels gradually increased, the AMPK activity decreased first and then increased, NLRP3, CX43 expression, and membrane/cytoplasmratio of Cx43 expression were increased in the gastric smooth muscle tissues of diabetic rats. Changes in ECM of gastric smooth muscle cells wereobserved in DGP rats. The DGP group showed higher collagen type I content, increased expression of Caspase-1, transforming growth factor-beta 3 (TGF-β3), and matrix metalloproteinase-2 (MMP-2), decreased tissue inhibitor of metalloproteinase-1 (TIMP-1) expression, and higher interleukin-1betacontent when compared with the control group. For gastric smooth muscle cells cultured under higher glucose, the MMP-2 and TGF-β3 expression wasdecreased, TGF-β1 and TIMP-1 expression was increased, the interleukin-1betacontent was decreased in cells after inhibition of NLRP3 expression; the NLRP3 and Caspase-1expression was decreased, and adenosine triphosphate content was lower after inhibition of Cx43; the expression of NLRP3, Caspase-1, P2X7, and the membrane/cytoplasm ratio of CX43 expression was decreased in cells after inhibition of AMPK and oxidative stress, the phospho-AMPK expression was also decreased after suppressing oxidative stress. Our findings suggest that high glucose induced the activation of the AMPK-Cx43-NLRP3 pathway through oxidative stress, and this pathway was involved in the ECM remodeling of gastric smooth muscles in DGP rats by regulating the biological functions of TGF-β3, TGF-β1, MMP-2, and TIMP-1.

Preparation of the microcapsules of Atractylodes chinensis volatile oil and its effect on gastric smooth muscle cell contraction

Abstract Atractylodes chinensis volatile oil (ACVO) microcapsules were prepared with chitosan as parietal material. The optimal conditions for encapsulation were investigated by the Response Surface Method (RSM). ACVO microcapsules were characterized by scanning electron microscopy (SEM), particle size analyzer and Fourier Transform infrared spectroscopy (FTIR), respectively. In vitro release properties of ACVO microcapsules were investigated at pH 1.5 and 7.4, respectively. The effects of ACVO microcapsules on the length of rat gastric smooth muscle cells (GSMC), mRNA and protein expression of calmodulin (CaM) and myosin light chain kinase (MLCK) were investigated. Results showed that under optimal conditions, encapsulation efficiency (EE) was 82.19 %. ACVO microcapsules were spherical with a particle size of 1100 nm. ACVO microcapsules had a good release profile at pH 1.5, and the cumulative release within 72 h reached 85.32 %. Cell experiments showed that ACVO microcapsules (0.02, 0.04 μg/mL) had no effect on cell activity, while induced GSMC to contract, and improved the mRNA, protein expressions of CaM and MLCK in GSMC.

Neural regulation of slow waves and phasic contractions in the distal stomach: a mathematical model

Objective. Neural regulation of gastric motility occurs partly through the regulation of gastric bioelectrical slow waves (SWs) and phasic contractions. The interaction of the tissues and organs involved in this regulatory process is complex. We sought to infer the relative importance of cellular mechanisms in inhibitory neural regulation of the stomach by enteric neurons and the interaction of inhibitory and excitatory electrical field stimulation. Approach. A novel mathematical model of gastric motility regulation by enteric neurons was developed and scenarios were simulated to determine the mechanisms through which enteric neural influence is exerted. This model was coupled to revised and extended electrophysiological models of gastric SWs and smooth muscle cells (SMCs). Main results. The mathematical model predicted that regulation of contractile apparatus sensitivity to intracellular calcium in the SMC was the major inhibition mechanism of active tension development, and that the effect on SW amplitude depended on the inhibition of non-specific cation currents more than the inhibition of calcium-activated chloride current (kiNSCC = 0.77 vs kiAno1 = 0.33). The model predicted that the interaction between inhibitory and excitatory neural regulation, when applied with simultaneous and equal intensity, resulted in an inhibition of contraction amplitude almost equivalent to that of inhibitory stimulation (79% vs 77% decrease), while the effect on frequency was overall excitatory, though less than excitatory stimulation alone (66% vs 47% increase). Significance. The mathematical model predicts the effects of inhibitory and excitatory enteric neural stimulation on gastric motility function, as well as the effects when inhibitory and excitatory enteric neural stimulation interact. Incorporation of the model into organ-level simulations will provide insights regarding pathological mechanisms that underpin gastric functional disorders, and allow for in silico testing of the effects of clinical neuromodulation protocols for the treatment of these disorders.

The effect of calcium channels blockade on slow-wave distribution in the electrophysiological model of human gastric wall smooth muscle cells

The effect of calcium channels blockade on slow-wave distribution in the electrophysiological model of human gastric wall smooth muscle cells

Prokinetic effects of Citrus reticulata and Citrus aurantium extract with/without Bupleurum chinense using multistress-induced delayed gastric emptying models

Context Citrus aurantium L (Rutaceae) (Au) and Citrus reticulata Blanco (Rutaceae) (Ci) are commonly used as couplet prokinetics and Bupleurum chinense DC. (Umbelliferae) (Bup) is an herbal antidepressant in traditional Chinese medicine. Objective This study evaluates the synergistic prokinetic effects of Bup with Au and Ci in mice suffering from multistress-induced delayed gastric emptying (DGE). Materials and methods Kunming mice were divided into four groups: control, DGE, AuCi and AuCiBup. Mice were gavaged with AuCi (14.25 g/kg) or AuCiBup (22.13 g/kg) extract for 12 days. Gastric reminder rate, intestinal driving ratio, sucrose preference and open field test were examined, and serotonin (5-HT), motilin (MTL), substance P (SP), 5-HT4R and c-kit were assayed. Intracellular Ca2+ levels in primary cultured gastric smooth muscle cells (GSMCs) were determined. Results Both AuCi and AuCiBup treatment significantly reduced gastric residual rate (39.5% and 67.7%, p < 0.01). Higher serum levels of 5-HT, MTL and SP were observed in treatment groups (AuCi: 0.060 mg/L, AuCiBup: 0.089 mg/L, DGE: 0.025 mg/L, p < 0.01). The expression of 5-HT4R and c-kit in the antrum and duodenum was upregulated after treatment (AuCi and AuCiBup, 4.3-times, 2.8-times to DGE, p < 0.01). Medicated serums of AuCi and AuCiBup effectively increased the influx of Ca2+ into GSMCs in vitro (1.8-times, p < 0.01). In terms of 5-HT4R expression, circulatory contents of 5-HT and SP and Ca2+ influx, AuCiBup demonstrated better prokinetic effects than AuCi. Conclusions These findings indicate the potential for developing combination therapy with antidepressants and prokinetics in gastrointestinal dysmotility management.

Bioactive Metabolites from the Fruiting Body and Mycelia of Newly-Isolated Oyster Mushroom and Their Effect on Smooth Muscle Contractile Activity.

Higher basidiomycetes are recognized as functional foods due to their bioactive compound content, which exerts various beneficial effects on human health, and which have been used as sources for the development of natural medicines and nutraceuticals for centuries. The aim of this study was to evaluate and compare the biological potential of basidiocarp and mycelial biomass produced by submerged cultivation of a new regionally isolated oyster mushroom. The strain was identified with a high percentage of confidence (99.30%) as Pleurotus ostreatus and was deposited in the GenBank under accession number MW 996755. The β-glucan content in the basidiocarp and the obtained mycelial biomass was 31.66% and 12.04%, respectively. Three mycelial biomass and basidiocarp extracts were prepared, and the highest total polyphenol content (5.68 ± 0.15 mg GAE/g DW and 3.20 ± 0.04 mg GAE/g DW) was found in the water extract for both the fruiting body and the mycelium biomass. The in vitro antioxidant activity of the extracts was investigated, and it was determined that the water extracts exhibited the most potent radical scavenging activity. The potential ability of this new fungal isolate to affect the contractile activity (CA) of dissected smooth muscle preparations (SMP) was examined for the first time. It was found that oyster mushrooms likely exhibit indirect contractile effects on the gastric smooth muscle (SM) cells.

Insulin-like growth factor-1 inhibits apoptosis of rat gastric smooth muscle cells under high glucose condition via adenosine monophosphate-activated protein kinase (AMPK) pathway.

Diabetic gastroparesis (DGP) is a common chronic complication of diabetes characterized by decreased gastric motility, and an effective number of gastric smooth muscle cells (GSMCs) ensures gastric motility. A previous study documented that apoptosis was present in gastric smooth muscles in rats with DGP and adenosine monophosphate-activated protein kinase (AMPK) was an important factor of apoptosis of rat GSMCs cultured under high glucose conditions. This study aimed to explore the effect of insulin-like growth factor-1 (IGF-1) on apoptosis of high glucose cultured rat GSMCs after silencing of AMPK and elucidate the underlying mechanism. A total of 120 rats were divided into normal control (NC, n = 20), diabetic gastroparesis (DGP, n = 50) and DGP + IGF-1 (n = 50) groups. After establishing the rat model of DGP, rats in the DGP+IGF-1 group received an intraperitoneal injection of IGF-1 at a dose of 1.5 μg/kg/d for 10 weeks. The level of AMPK activity, liver kinase B1 (LKB1) activity, and calcium/calmodulin-dependent protein kinase b (CaMKKb) expression in rat gastric smooth muscle tissues was detected by Western blot analysis. Apoptosis in rat gastric smooth muscle tissues was detected by TUNEL assay. We also cultured rat GSMCs in vitro under high glucose (HG) condition (35 mM), incubated cells with IGF-1, and silenced AMPK with siRNA. The cells were divided into HG, HG + IGF-1, HG + siRNA, and HG + siRNA + IGF-1 groups. The apoptosis rates of rat GSMCs after silencing AMPK were detected by TUNEL assay and flow cytometry, and apoptosis-related protein expression in rat GSMCs was detected by Western blot. IGF-1 decreased LKB1 activity, CaMKKb expression, AMPK activity, and inhibited apoptosis in rat gastric smooth muscle tissues. Compared with rat GSMCs cultured in vitro under HG conditions, apoptosis rates were reduced after treatment with IGF-1 and AMPK silencing (both p < 0.01). Apoptosis rates were higher in the HG + siRNA group compared with the HG + IGF-1 group (p < 0.05). IGF-1 down-regulated the expression of calcium/calmodulin-dependent kinase II (CaMKII) and p53, up-regulated the expression of p21, PLC-b3, PI3K p110 Ser1070, and the activities of Akt, p70S6K, mTORC1, and mTORC2. IGF-1 also up-regulated Bcl-2 expression and down-regulated the expression of BAX and Caspase-3. IGF-1 can inhibit the apoptosis of rat GSMCs under high glucose conditions, its mechanism may be related to the regulation of expression and activity of p53, PI3K, TSC-2, Akt, mTOR, 4E-BP1, p70S6K, p21, CaMKII, and PLC-b3 in rat GSMCs acting through AMPK pathway.

The Role of Potassium Channel Gates in the Electrophysiology of the Human Gastric Smooth Muscle Cell

Background: The cell membrane acts as a filter, allowing ions to enter and leave the cell. Ionic channels are responsible for passing ions. This task is the responsibility of the ion channel gates, and ion transfer generates the action potential. Potassium channels play a prominent role in gastrointestinal smooth muscle cells and slow-wave production. Potassium channels are involved in acid secretion and gastric contraction. Gastric functional problems such as reflux disease and motility disorder are classified as electrophysiological disorders. Objectives: This study aimed to investigate the effect of potassium channel gates on the electrophysiology of the human gastric smooth muscle cells. Methods: Three states were considered for the potassium channels gate (Including physiological state, 50% blockage, and 90% blockage) to investigate the effect of the status of the gates, and a slow-wave diagram was obtained in these three states. Then, the value and the time of action potential were compared at five indicator points (initial potential, maximum spike potential, minimum valley potential, maximum plateau potential, and resting potential) in slow-wave. Results: The results showed that the maximum effect of the activation parameter of the potassium channel gate (τd,Kni) was in 90% blockage compared to the physiological state, so that the maximum spike potential decreases by 2.43%. Also, a 90% blockage in the fast potassium channel gate inactivation parameter (τf,Kfi) increased the maximum spike potential by 12.6% compared to the physiological state, while the minimum valley potential increased by 3%. In addition, the τf,Kfi parameter reduced the time of occurrence of the maximum plateau potential by 7.9%. Conclusions: Potassium channels affect the slow-wave of the human gastric smooth muscle cell in spike, valley, and plateau phases. Using this method and blocking ion channels by pharmacological agents, the effect of ions in different phases of the slow-wave can be investigated. Also, it can help improve the contractile and motility disorders of the smooth muscles of the gastrointestinal tract.

Pharmacological effects of Pugionium cornutum (L.) Gaertn. extracts on gastrointestinal motility are partially mediated by quercetin

BackgroundThe majority of global population suffer from various functional gastrointestinal disorders. Pugionium cornutum (L.) Gaertn. (PCG) is used to relieve indigestive symptoms in traditional Chinese medicine. However, little is known about the effects of bioactive components from PCG extracts on gastrointestinal motility.MethodsCrude ethanol extract of PCG (EEP) was prepared from Pugionium cornutum (L.) Gaertn. Different solvents were used to prepare fine extracts from EEP, including water extract of PCG (WEP), petroleum ether extract of PCG (PEEP), dichloromethane extract of PCG (DEP) and ethyl acetate extract of PCG (EAEP). Smooth muscle cell model and colonic smooth muscle stripe model were used to test the bioactive effects and mechanisms of different PCG extracts on contraction and relaxation. Diverse chromatographic methods were used to identify bioactive substances from PCG extracts.ResultsEEP was found to promote the relaxation of gastric smooth muscle cell and inhibit the contraction of colonic smooth muscle strip. Among the fractions of EEP, EAEP mainly mediated the relaxation effect by stimulating intracellular calcium influx. Further evidences revealed that EAEP was antagonistic to acetylcholine. In addition, COX and NO-GC-PKC pathways may be also involved in EAEP-mediated relaxation effect. Quercetin was identified as a bioactive compound from PCG extract for the relaxation effect.ConclusionOur research supports the notion that PCG extracts promote relaxation and inhibits contraction of gastrointestinal smooth muscle at least partially through the effect from quercetin.

MALAT1: A Pivotal lncRNA in the Phenotypic Switch of Gastric Smooth Muscle Cells via the Targeting of the miR-449a/DLL1 Axis in Diabetic Gastroparesis.

Diabetic gastroparesis (DGP) is a common complication of diabetes mellitus (DM). Our previous study suggested that the expression of the long non-coding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is closely related to DGP. However, the role of MALAT1 in DGP pathogenesis remains unclear. Here, we aim to characterize the role of MALAT1 in DGP. First, we analyzed the lncRNA expression profiles through lncRNA sequencing. Next, we detected MALAT1 expression in the stomach tissues of DGP model mice and diabetic patients. Then, we investigated the role and mechanisms of MALAT1 in the proliferation, migration, phenotypic switch, and carbachol-induced intracellular Ca2+ changes in human gastric smooth muscle cells (HGSMCs) under high glucose (HG) conditions, using short hairpin RNA technology, RNA immunoprecipitation, and dual-luciferase reporter assays. We show that MALAT1 expression was upregulated in the gastric tissues of DGP model mice, the adjacent healthy tissues collected from diabetic gastric cancer patients with DGP symptoms, and in HGSMCs cultured under HG conditions. Functionally, MALAT1 knockdown in vitro impacted the viability, proliferation, migration and promoted the phenotypic switch of HGSMCs under HG conditions. Additionally, we show that MALAT1 sponged miR-449a, regulating Delta-like ligand 1 (DLL1) expression in HGSMCs; any disturbance of the MALAT1/miR-449a/DLL1 pathway affects the proliferation, migration, phenotypic switch, and carbachol-induced Ca2+ transient signals in HGSMCs under HG conditions. Collectively, our data highlight a novel regulatory signaling pathway, the MALAT1/miR-449a/DLL1 axis, in the context of DGP.

Molecular mechanism of mice gastric oxidative damage induced by nanoparticulate titanium dioxide.

Nanoparticulate titanium dioxide (Nano-TiO2) has been widely used in food industry, and it has been demonstrated to have adverse effects on mice and human stomach, but its mechanism is rarely concerned. The aim of this study is to determine the effects of nano-TiO2 on the stomach and confirm the role of oxidative stress and apoptosis in the mice gastric damage caused by nano-TiO2, as well as its molecular mechanisms. Mice were continuously exposed to nano-TiO2 with 1.25, 2.5 and 5mg/kg bw by intragastric administration for 9 months in the present study. The ultrastructure, levels of reactive oxygen species (ROS) and peroxides, activities of antioxidant enzymes and mitochondria-related enzymes, ATP contents as well as apoptosis-related factors expression in mice stomach were examined. Oxidative stress, apoptosis and nano-TiO2 aggregation were found in gastric mucosal smooth muscle cells after nano-TiO2 exposure. Nano-TiO2 exposure also resulted in the over-production of ROS and peroxides, decrease of ATP production and activities of antioxidant enzymes and mitochondria-related ATPases, upregulation of apoptosis-related factors including γH2AX, Cyt c, caspase 3, and p-JNK expression, and down-regulation of Bcl-2 expression in mice stomach. The gastric toxicity of mice induced by chronic exposure to low dose nano-TiO2 may be associated with oxidative stress and mitochondria-mediated apoptosis in mice.

Preliminary engineering for in situ in vivo bioprinting: a novel micro bioprinting platform for in situ in vivo bioprinting at a gastric wound site

We first proposed the concept of in situ in vivo bioprinting in order to address the existing deficiencies in conventional bioprinting. Herein we verified this concept taking the case of the treatment for gastric wall injury and presented this work as a preliminary step towards a new method in the field of bioprinting. In this study, a micro bioprinting platform which can be installed to an endoscope was developed to enter the human body and process bioprinting. Printed circuit micro-electro-mechanical-system techniques were used in the design and fabrication of the platform. Control system with high accuracy was developed and performance tests were carried out to verify the feasibility of the platform. The 2-layer tissue scaffolds were printed in a stomach model. Gelatin–alginate hydrogels with human gastric epithelial cells and human gastric smooth muscle cells were used as bioinks to mimic the anatomical structure of a stomach. A 10 d cell culture showed that printed cells remained a high viability and a steady proliferation, which indicated good biological function of cells in printed tissue scaffolds. This work presents an innovative advance not only in the field of bioprinting but also in the clinical sciences.

Gastric smooth muscle cells manifest an abnormal phenotype in Parkinson's disease rats with gastric dysmotility.

Gastroparesis is a common symptom in Parkinson's disease (PD) and whether any change occurs in gastric smooth muscle cells (SMCs) of PD patients is unclear. We previously reported that rats with bilateral substantia nigra lesions induced by 6-hydroxydopamine (6-OHDA), referred to as 6-OHDA rats, manifest typical gastroparesis. In the present study, we further investigate the underlying mechanism. By means of an organ bath system and an implantable radiotelemetry system, both a weakened contractile force of gastric circular smooth muscle and gastric myoelectric activity were detected in the 6-OHDA rats and phasic and tonic contractions elicited by carbachol or high concentration of potassium were significantly reduced in gastric circular muscle strips. A thickened smooth muscle layer was observed under a light microscope and an ultrastructure of hypertrophic SMCs, with increased caveolae and decreased dense bodies, was observed under transmission electron microscope. Furthermore, the mRNA and protein expression levels of contractile markers (myosin light chain 20, myosin heavy chain 11 and α-smooth muscle actin) and the transcription factor serum response factor (SRF) were significantly decreased, while the TNFα and IL-1β content was increased in the 6-OHDA rats. These results suggest that the decreased contractile force in 6-OHDA rats may be associated with the phenotypic abnormality observed in SMCs, which is due to downregulated contractile proteins induced by decreased SRF expression in the inflammatory muscular microenvironment.

Identification of the Uptake Transporter Responsible for Distribution of Acotiamide into Stomach Tissue.

The acetylcholinesterase inhibitor, acotiamide, improves gastric motility and is clinically used to treat functional dyspepsia. The present study aimed to identify the transporters involved in the distribution of acotiamide in stomach tissue. Acotiamide uptake by the gastric cancer-derived model cell line, Hs746 T, was Na+- and pH-independent. The initial uptake velocity of acotiamide was saturable with increasing concentrations of acotiamide and was inhibited by selective serotonin reuptake inhibitors, which are potent inhibitors of the plasma membrane monoamine transporter (PMAT). The uptake of acotiamide by PMAT gene-transfected HEK293 cells was saturable, with similar Km (197.9 μM) values to those of uptake by Hs 746T cells (106 μM). Moreover, immunoreactivity of PMAT was found in the gastric smooth muscle and vascular endothelial cells. These results suggest that PMAT contributes to the distribution of acotiamide in the stomach, where it exerts its pharmacological effects.

Localization of cannabinoid and cannabinoid related receptors in the cat gastrointestinal tract

A growing body of literature indicates that activation of cannabinoid receptors may exert beneficial effects on gastrointestinal inflammation and visceral hypersensitivity. The present study aimed to immunohistochemically investigate the distribution of the canonical cannabinoid receptors CB1 (CB1R) and CB2 (CB2R) and the putative cannabinoid receptors G protein-coupled receptor 55 (GPR55), nuclear peroxisome proliferator-activated receptor alpha (PPARα), transient receptor potential ankyrin 1 (TRPA1), and serotonin receptor 5-HT1a 5-HT1aR) in tissue samples of the gastrointestinal tract of the cat. CB1R-immunoreactivity (CB1R-IR) was observed in gastric epithelial cells, intestinal enteroendocrine cells (EECs) and goblet cells, lamina propria mast cells (MCs), and enteric neurons. CB2R-IR was expressed by EECs, enterocytes, and macrophages. GPR55-IR was expressed by EECs, macrophages, immunocytes, and MP neurons. PPARα-IR was expressed by immunocytes, smooth muscle cells, and enteroglial cells. TRPA1-IR was expressed by enteric neurons and intestinal goblet cells. 5-HT1a receptor-IR was expressed by gastrointestinal epithelial cells and gastric smooth muscle cells. Cannabinoid receptors showed a wide distribution in the feline gastrointestinal tract layers. Although not yet confirmed/supported by functional evidences, the present research might represent an anatomical substrate potentially useful to support, in feline species, the therapeutic use of cannabinoids during gastrointestinal inflammatory diseases.

Effects of Xiangsha Liujunzi decoction drug serum on gastric antrum smooth muscle cells from rats with functional dyspepsia by regulating gastrointestinal hormones

Abstract Objective To observe the effect and mechanism of Xiangsha Liujunzi decoction (XSLJZD) drug serum on gastric antrum smooth muscle cells (SMCs) in rats with functional dyspepsia (FD). Methods Gastric antrum SMCs from rats with FD were isolated, cultured, and then divided into six groups as follows: control, model, domperidone, low-dose XSLJZD (LXSLJZD), medium-dose XSLJZD (MXSLJZD), and high-dose XSLJZD (HXSLJZD). Each group was administered the corresponding drug serum for intervention. Drug serum intervention conditions and proliferative activity of SMCs were tested by cholecystokinin octapeptide. Ghrelin, gastrin, somatostatin, and substance P (SP) levels were measured by ELISA. Somatostatin and SP mRNA expression was measured by real-time PCR. Results A concentration of 10% drug serum for 24 h was decided to be the best intervention condition for later study. The mean optical density value in the model group was lower than that in the control group (P = .001). Optical density values in the domperidone and HXSLJZD groups were higher than those in the model group (P = .025, P = .032, respectively). Gastrin, SP, and ghrelin levels in the model group were lower (P = .007, P = .037, P = .005, respectively), but somatostatin levels were higher, compared with those in the control group (P = .031). Gastrin, SP, and ghrelin levels in the domperidone, MXSLJZD, and HXSLJZD groups were higher than those in the model group (all P Conclusions XSLJZD and domperidone drug serum effectively promote proliferative activity of gastric antrum SMCs in an FD model. The mechanism of this activity may be regulated by gastrointestinal hormones.

Mechanism of AMPK-mediated apoptosis of rat gastric smooth muscle cells under high glucose condition.

To observe changes in AMP-activated protein kinase (AMPK) activity and phosphorylation changes in AMPK signaling pathway in gastric smooth muscle cells of rats with diabetic gastroparesis (DGP), investigate the effect of AMPK on apoptosis and explore the underlying mechanism. After establishing rat model of DGP, rats were divided into normal control (NC) and DGP groups. The phosphorylation changes in AMPK pathway were detected by AMPK Signaling Phospho-Antibody Array, and the apoptosis-related proteins were determined. Rat gastric smooth muscle cells were cultured in vitro under different glucose conditions, and divided into normal and high glucose groups. The AMPK activity and intracellular Ca2+ changes in cells were observed. After AMPK silencing, cells were divided into high glucose-24h, high glucose-48h and high glucose-48h+siRNA groups. Changes in expression of apoptosis-related proteins were observed. AMPK activity and apoptosis rates were both increased in gastric smooth muscle tissues in DGP rats (P<0.05, P<0.001, respectively). A total of 14 apoptosis-related differentially phosphorylated proteins were identified. Under high-glucose condition, AMPK activity and intracellular Ca2+ concentrations in rat gastric smooth muscle cells were increased (both P<0.05). After AMPK silencing, p53 expression was decreased, Akt and p70 S6 ribosomal protein kinase (p70S6K) activities were were increased, Bcl-2 expression was increased, CaMKII activity was decreased in the high glucose-48h group. Under high-glucose condition, activated AMPK can directly or indirectly promote cells apoptosis by regulating the expression and activity of p53, Akt, p70S6K, Protein kinase A (PKA), Phospholipidol C (PLC)-β3, CaMKII, CaMKIV and eukaryotic translation initiation factor 4E binding protein1 (4E-BP1) in rat gastric smooth muscle cells.

Expression and function of umami receptors T1R1/T1R3 in gastric smooth muscle.

l-amino acids, such as monosodium glutamate (MSG), activate the umami receptor T1R1/T1R3. We previously showed increased peristalsis in response to activation of T1R1/T1R3 by MSG in mouse colon. However, the expression and function of these receptors in the different regions of the stomach are not clear. Mouse gastric smooth muscle cells (SMCs) were isolated and cultured in Dulbecco's Modified Eagle Medium. Expression of T1R1 and T1R3 was measured by RT-PCR and Western blot. The effect of MSG with and without inosine monophosphate (IMP, an allosteric activator of T1R1/T1R3) on acetylcholine (ACh)-induced contraction was measured in muscle strips and isolated SMCs by scanning micrometry. The effect of MSG with or without IMP on activation of G proteins and ACh-induced Ca2+ release was measured in SMCs. Monosodium glutamate inhibited ACh-induced contractions in muscle strips from both antrum and fundus and the effect of MSG was augmented by IMP; the effects were concentration-dependent and not affected by the nitric oxide synthase inhibitor, L-NNA, or tetrodotoxin suggesting a direct effect on SMCs. In isolated gastric SMCs, T1R1 and T1R3 transcripts and protein were identified. Addition of MSG with or without IMP inhibited ACh-induced Ca2+ release and muscle contraction; the effect on contraction was blocked by pertussis toxin suggesting activation of Gαi proteins. MSG in the presence of IMP selectively activated Gαi2 . Umami receptors (T1R1/T1R3) are present on SMCs of the stomach, and activation of these receptors induces muscle relaxation by decreasing [Ca2+ ]i via Gαi2 .