Role of Activated Protein C in Helicobacter pylori-Associated Gastritis

ABSTRACT The protein C (PC) pathway has recently been suggested to play a role in the regulation of the inflammatory response. To further extend the anti-inflammatory effect of activated PC (APC) in vivo, particularly its biological relevance to human disease, the activity of APC in the mucosa of patients with Helicobacter pylori-associated gastritis and the effect of vacuolating cytotoxin (VacA), cytotoxin-associated antigen (CagA), andH. pylori lipopolysaccharide (LPS) on PC activation were evaluated. This study comprised 35 patients with chronic gastritis. There were 20 patients with and 15 without H. pylori infection. The levels of PC and APC-PC inhibitor (PCI) complex were measured by immunoassays. The level of PC was significantly decreased and the level of APC-PCI complex was significantly increased in biopsy specimens from gastric corpus and antrum in patients with H. pylori-associated gastritis as compared to H. pylori-negative subjects. The concentrations of VacA, CagA, and LPS were significantly correlated with those of the APC-PCI complex in biopsy mucosal specimens from the gastric corpus and antrum. H. pylori LPS, VacA, and CagA induced a dose-dependent activation of PC on the surface of monocytic cells. APC inhibited the secretion of tumor necrosis factor alpha (TNF-α) induced by H. pylori LPS. Overall, these results suggest that H. pylori infection is associated with increased APC generation in the gastric mucosa. The inhibitory activity of APC on TNF-α secretion may serve to protect H. pylori-induced gastric mucosal damage.

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Therapeutic effects of Zuojin Pill on Helicobacter pylori-induced chronic atrophic gastritis through JMJD2B/COX-2/VEGF signaling axis

10.21203/rs.3.rs-20652/v2 ◽

2020 ◽

Author(s):

Shihua Wu ◽

Chunmei Bao ◽

Ruilin Wang ◽

Xiaomei Zhang ◽

Sijia Gao ◽

...

Keyword(s):

Helicobacter Pylori ◽

Cell Viability ◽

Atrophic Gastritis ◽

Chronic Atrophic Gastritis ◽

Gastric Mucosal Damage ◽

Therapeutic Effects ◽

Cox 2 ◽

H Pylori

Abstract Background: Zuojin Pill (ZJP), a famous Chinese medicinal formula, widely accepted for treatment of chronic atrophic gastritis (CAG) in China. This study aimed to explore the therapeutic effects and mechanisms of ZJP in Helicobacter pylori (H. pylori) - induced chronic atrophic gastritis (CAG) in vivo and in vitro. Methods: CAG rat model was induced by H. pylori. ZJP (0.63, 1.26, and 2.52 g/kg, respectively) was administered orally for four weeks. Therapeutic effects of ZJP were identified by H&E staining and serum indices. In addition, cell viability, morphology and proliferation were detected by cell counting kit-8 (CCK8) and high-content screening assay (HCS), respectively. Moreover, relative mRNA expression and protein expression related to JMJD2B/COX-2/VEGF axis was detected to investigate the potential mechanisms of ZJP in CAG. Results: Results showed the symptoms (weight loss and gastric mucosa damage) of CAG were alleviated, and the contents of TNF-α in serum was markedly decreased after treating with ZJP. Moreover, cell viability, proliferation and morphology changes of GES-1 cells were ameliorated by ZJP intervention. In addition, proinflammatory genes and JMJD2B/COX-2/VEGF axis related genes were suppressed by ZJP administration in vitro and in vivo. Meanwhile, immunohistochemistry (IHC) and western blot confirmed down-regulation of these genes by ZJP intervention. Conclusion: ZJP treatment can alleviate gastric mucosal damage induced by H. pylori via JMJD2B/COX-2/VEGF axis.

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Activated protein C targets immune cells and rheumatoid synovial fibroblasts to prevent inflammatory arthritis in mice

Rheumatology ◽

10.1093/rheumatology/key429 ◽

2019 ◽

Vol 58 (10) ◽

pp. 1850-1860 ◽

Cited By ~ 4

Author(s):

Meilang Xue ◽

Suat Dervish ◽

Kelly J McKelvey ◽

Lyn March ◽

Fang Wang ◽

...

Keyword(s):

Protein C ◽

Activated Protein C ◽

Synovial Fibroblasts ◽

Endothelial Protein C Receptor ◽

Mouse Thymus ◽

Tnf Α ◽

Thymus Cells ◽

Proliferation And Invasion

Abstract Objectives To investigate whether activated protein C (APC), a physiological anticoagulant can inhibit the inflammatory/invasive properties of immune cells and rheumatoid arthritis synovial fibroblasts (RASFs) in vitro and prevent inflammatory arthritis in murine antigen-induced arthritis (AIA) and CIA models. Methods RASFs isolated from synovial tissues of patients with RA, human peripheral blood mononuclear cells (PBMCs) and mouse thymus cells were treated with APC or TNF-α/IL-17 and the following assays were performed: RASF proliferation and invasion by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and cell invasion assays, respectively; cytokines and signalling molecules using ELISA or western blot; Th1 and Th17 phenotypes in human PBMCs or mouse thymus cells by flow cytometry. The in vivo effect of APC was evaluated in AIA and CIA models. Results In vitro, APC inhibited IL-1β, IL-17 and TNF-α production, IL-17-stimulated cell proliferation and invasion and p21 and nuclear factor κB activation in RASFs. In mouse thymus cells and human PBMCs, APC suppressed Th1 and Th17 phenotypes. In vivo, APC inhibited pannus formation, cartilage destruction and arthritis incidence/severity in both CIA and AIA models. In CIA, serum levels of IL-1β, IL-6, IL-17, TNF-α and soluble endothelial protein C receptor were significantly reduced by APC treatment. Blocking endothelial protein C receptor, the specific receptor for APC, abolished the early or preventative effect of APC in AIA. Conclusion APC prevents the onset and development of arthritis in CIA and AIA models via suppressing inflammation, Th1/Th17 phenotypes and RASF invasion, which is likely mediated via endothelial protein C receptor.

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Follow-up analysis and histopathological study of gastric mucosa in patients with Helicobacter pylori infection

Journal of International Medical Research ◽

10.1177/03000605211055397 ◽

2021 ◽

Vol 49 (12) ◽

pp. 030006052110553

Author(s):

Guang Zhao ◽

Zhishang Zhang ◽

Baohui Li ◽

Silin Huang ◽

Wensi Li ◽

...

Keyword(s):

Helicobacter Pylori ◽

Gastric Mucosa ◽

Intraepithelial Neoplasia ◽

Mucosal Damage ◽

Helicobacter Pylori Infection ◽

Gastric Mucosal Damage ◽

Histopathological Study ◽

Significant Difference ◽

H Pylori

Objective To investigate the histomorphological characteristics of the gastric mucosa and the prognosis in patients with Helicobacter pylori infection. Methods Progressive damage to the gastric mucosa was examined by immunohistochemistry in 2294 patients with H. pylori infection and follow-up information was analyzed. Results H. pylori initially colonized the mucus layer covered by the gastric mucosa epithelium, then selectively adhered to and destroyed the surface mucus cells causing intra-gastric and extra-gastric lesions. Gastric mucosal damage induced by H. pylori was divided into five stages according to the depth of H. pylori invasion and degree of lesion deterioration: mucilaginous, surface mucocellular, lamina propria lesion, mucosal atrophy, and intraepithelial neoplasia stages. Morphological follow-up analysis revealed no significant difference in 6-month curative effects between stage I and stage II, but significant differences were found between stages II and III, stages III and IV, and between stages IV and stage V, respectively. Conclusions This novel staging strategy may be a valuable tool for diagnosing and predicting the results of gastric mucosal damage induced by H. pylori infection.

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In Vitro and In Vivo Antibacterial Activities of Patchouli Alcohol, a Naturally Occurring Tricyclic Sesquiterpene, against Helicobacter pylori Infection

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00122-17 ◽

2017 ◽

Vol 61 (6) ◽

Cited By ~ 16

Author(s):

Y. F. Xu ◽

D. W. Lian ◽

Y. Q. Chen ◽

Y. F. Cai ◽

Y. F. Zheng ◽

...

Keyword(s):

Helicobacter Pylori ◽

Bacterial Resistance ◽

Clinical Isolates ◽

Potential Candidate ◽

Necrosis Factor Alpha ◽

Content Type ◽

Patchouli Alcohol ◽

H Pylori

ABSTRACT This study further evaluated the in vitro and in vivo anti-Helicobacter pylori activities and potential underlying mechanism of patchouli alcohol (PA), a tricyclic sesquiterpene. In the in vitro assay, the capacities of PA to inhibit and kill H. pylori were tested on three standard strains at different pH values and on 12 clinical isolates. The effects of PA on H. pylori adhesion (and its alpA, alpB, and babA genes), motility (and its flaA and flaB genes), ultrastructure, and flagellation were investigated. Moreover, the H. pylori resistance to and postantibiotic effect (PAE) of PA were determined. Furthermore, the in vivo effects of PA on H. pylori eradication and gastritis were examined. Results showed that MICs of PA against three standard strains (pH 5.3 to 9) and 12 clinical isolates were 25 to 75 and 12.5 to 50 μg/ml, respectively. The killing kinetics of PA were time and concentration dependent, and its minimal bactericidal concentrations (MBCs) were 25 to 75 μg/ml. In addition, H. pylori adhesion, motility, ultrastructure, and flagellation were significantly suppressed. PA also remarkably inhibited the expression of adhesion genes (alpA and alpB) and motility genes (flaA and flaB). Furthermore, PA treatment caused a longer PAE and less bacterial resistance than clarithromycin and metronidazole. The in vivo study showed that PA can effectively eradicate H. pylori, inhibit gastritis, and suppress the expression of inflammatory mediators (COX-2, interleukin 1β, tumor necrosis factor alpha, and inducible nitric oxide synthase [iNOS]). In conclusion, PA can efficiently kill H. pylori, interfere with its infection process, and attenuate gastritis with less bacterial resistance, making it a potential candidate for new drug development.

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Gastroprotective effects of Nelumbinis Rhizomatis Nodus carbonisata-derived carbon dots on ethanol-induced gastric ulcers in rats

10.21203/rs.3.rs-101143/v1 ◽

2020 ◽

Author(s):

Juan Luo ◽

Jie Hu ◽

Meiling Zhang ◽

Yue Zhang ◽

Jiashu Wu ◽

...

Keyword(s):

Carbon Dots ◽

Average Diameter ◽

Mucosal Damage ◽

In Vitro Cytotoxicity ◽

Gastric Mucosal Damage ◽

Gastric Ulcers ◽

Necrosis Factor Alpha ◽

In Vivo Experiments

Abstract BackgroundGastric ulcers is a common gastrointestinal digestive system disease. Considering the frequency of human gastric ulcers, the side effects and cost of some existing synthetic drugs, the use of natural products is an important choice for many people. The aim of present study was to explore gastroprotective effects of nelumbinis rhizomatis nodus carbonisata carbon dots (NRNC-CDs) on ethanol-induced gastric ulcers in rats.MethodsThe NRNC-CDs were synthesized via high temperature calcinations treatment at 350 ℃ for 1 h were characterized by various spectroscopic and electron microscopy techniques for their structural, morphological, and optical properties. In vitro cytotoxicity of CDs for the human gastric epithelial cells line (GES-1 cells) was assessed by the CCK-8 assay. Furthermore, the study evaluated gastroprotective effects of NRNC-CDs on ethanol-induced gastric ulcers in rats, followed by a preliminary study on the possible mechanisms of gastroprotection.ResultesNRNC-CDs with a quantum yield of 1.38% have an average diameter of 2.89±0.82nm and the lattice spacing of 0.29 nm , and exerted low toxicity to GES-1 cells by CCK-8 test. In vivo experiments showed that NRNC-CDs remarkably reduced gastric mucosal damage and significantly increased the levels of superoxide dismutase (SOD), catalase (CAT), glutathione (GSH) and glutathione peroxidase (GSH-Px). In addition, NRNC-CDs also significantly inhibited tumor necrosis factor-alpha (TNF-a) and pro-inflammatory interleukin-6 (IL-6) level in gastric tissues. Histological findings demonstrated that NRNC-CDs exhibited a protective effect against tissue alterations in response to the ethanol-induced ulcer.ConclussionThe potent gastroprotective effect of NRNC-CDs were thus attributed to its anti-inflammatory and antioxidant effects. This discovery provides guidance for further research the effect of CDs in gastrointestinal digestive diseases.

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Complex T Cell Interactions Contribute to Helicobacter pylori Gastritis in Mice

Infection and Immunity ◽

10.1128/iai.01269-12 ◽

2012 ◽

Vol 81 (3) ◽

pp. 740-752 ◽

Cited By ~ 31

Author(s):

Brian M. Gray ◽

Clinton A. Fontaine ◽

Sara A. Poe ◽

Kathryn A. Eaton

Keyword(s):

T Cells ◽

Helicobacter Pylori ◽

T Cell ◽

Published Data ◽

Proinflammatory Response ◽

Necrosis Factor Alpha ◽

Content Type ◽

Tnf Α ◽

Ifn Γ ◽

H Pylori

ABSTRACTDisease due to the gastric pathogenHelicobacter pylorivaries in severity from asymptomatic to peptic ulcer disease and cancer. Accumulating evidence suggests that one source of this variation is an abnormal host response. The goal of this study was to use a mouse model ofH. pylorigastritis to investigate the roles of regulatory T cells (Treg) as well as proinflammatory T cells (Th1 and Th17) in gastritis, gastric T cell engraftment, and gastric cytokine production. Our results support published data indicating that severe gastritis in T cell recipient mice is due to failure of Treg engraftment, that Treg ameliorate gastritis, and that the proinflammatory response is attributable to interactions between several cell subsets and cytokines. We confirmed that gamma interferon (IFN-γ) is essential for induction of gastritis but showed that IFN-γ-producing CD4 T cells are not necessary. Interleukin 17A (IL-17A) also contributed to gastritis, but to a lesser extent than IFN-γ. Tumor necrosis factor alpha (TNF-α) and IL-17F were also elevated in association with disease. These results indicate that whileH. pylori-specific CD4+T cells and IFN-γ are both essential for induction of gastritis due toH. pylori, IFN-γ production by T cells is not essential. It is likely that other proinflammatory cytokines, such as IL-17F and TNF-α, shown to be elevated in this model, also contribute to the induction of disease. We suggest that gastritis due toH. pyloriis associated with loss of immunoregulation and alteration of several cytokines and cell subsets and cannot be attributed to a single immune pathway.

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Expression of Protein S in the Murine Heart and Cultured Mouse Cardiomyocytes Is Down-regulated by Cytokines

Thrombosis and Haemostasis ◽

10.1055/s-0037-1616096 ◽

2001 ◽

Vol 86 (08) ◽

pp. 623-629 ◽

Cited By ~ 5

Author(s):

Takayoshi Shimokawa ◽

Eriko Yamafuji ◽

Tetsuhito Kojima ◽

Hidehiko Saito ◽

Koji Yamamoto

Keyword(s):

Protein C ◽

Interleukin 1 ◽

Activated Protein C ◽

Protein S ◽

Suppressive Effect ◽

Inflammatory Conditions ◽

Tnf Α ◽

Factor Α ◽

Necrosis Factor

SummaryProtein S (PS), a co-factor of activated protein C, is a vitamin K-dependent anticoagulant protein and is known to be produced extrahepatically. In the present study, the concentration of PS mRNA was determined tissue by tissue in the mouse, and it was high in lung, adrenal and heart as well as in liver. We further investigated the effects of lipopolysaccharide (LPS), tumor necrosis factor-α (TNF-α), and interleukin-1 (IL-1) on the PS mRNA expression in murine tissues in vivo. Although LPS and TNF-α significantly decreased the expression level of PS mRNA in all tissues examined (e.g., lung, liver, heart, and kidney) and the PS antigen level in plasma, the suppressive effect of IL-1 on PS gene expression was limited to heart. More specifically, considerable amounts of PS mRNA and antigen were expressed in a cultured mouse cardiomyocyte cell line, and again, treatment with IL-1 decreased the PS expression in these cells. These observations raise a possibility that the expression of cardiac PS may contribute to the regional anticoagulant potential in heart, and suggest that the decreased PS expression by cytokines may result in an increase in the systemic and/or regional prothrombotic potential under inflammatory conditions.

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Therapeutic effect of Zuojin Pill on chronic atrophic gastritis induced by Helicobacter pylori through JMJD2B/COX-2/VEGF

10.21203/rs.3.rs-20652/v3 ◽

2020 ◽

Author(s):

Shihua Wu ◽

Chunmei Bao ◽

Ruilin Wang ◽

Xiaomei Zhang ◽

Sijia Gao ◽

...

Keyword(s):

Helicobacter Pylori ◽

Cell Viability ◽

Atrophic Gastritis ◽

Chronic Atrophic Gastritis ◽

Gastric Mucosal Damage ◽

Therapeutic Effects ◽

Cox 2 ◽

H Pylori

Abstract Background: Zuojin Pill (ZJP), a famous Chinese medicinal formula, is widely accepted for treatment of chronic atrophic gastritis (CAG) in China. This study aimed to explore the therapeutic effects and mechanisms of ZJP in Helicobacter pylori (H. pylori) - induced chronic atrophic gastritis (CAG) in vivo and in vitro. Methods: CAG rat model was induced by H. pylori. ZJP (0.63, 1.26, and 2.52 g/kg, respectively) was administered orally for four weeks. Therapeutic effects of ZJP were identified by H&E staining and serum indices. In addition, cell viability, morphology and proliferation were detected by cell counting kit-8 (CCK8) and high-content screening assay (HCS), respectively. Moreover, relative mRNA expression and protein expression related to JMJD2B/COX-2/VEGF axis was detected to investigate the potential mechanisms of ZJP in CAG.Results: Results showed the symptoms (weight loss and gastric mucosa damage) of CAG were alleviated, and the contents of TNF-α in serum was markedly decreased after treating with ZJP. Moreover, cell viability, proliferation and morphology changes of GES-1 cells were ameliorated by ZJP intervention. In addition, proinflammatory genes and JMJD2B/COX-2/VEGF axis related genes were suppressed by ZJP administration in vitro and in vivo. Meanwhile, immunohistochemistry (IHC) and western blot confirmed down-regulation of these genes by ZJP intervention. Conclusion: ZJP treatment can alleviate gastric mucosal damage induced by H. pylori via JMJD2B/COX-2/VEGF axis.

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Role of Outer Membrane Vesicles From Helicobacter pylori in Atherosclerosis

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.673993 ◽

2021 ◽

Vol 9 ◽

Author(s):

Na Wang ◽

Faying Zhou ◽

Caiyu Chen ◽

Hao Luo ◽

Jingwen Guo ◽

...

Keyword(s):

Helicobacter Pylori ◽

Outer Membrane ◽

Umbilical Vein ◽

Membrane Vesicles ◽

Plaque Formation ◽

Outer Membrane Vesicles ◽

Inflammatory Factors ◽

Necrosis Factor Alpha ◽

H Pylori

Infection is thought to be involved in the pathogenesis of atherosclerosis. Studies have shown the association between helicobacter pylori (H. pylori) and coronary artery disease. It is interesting to find H. pylori DNA and cytotoxin-associated gene A (CagA) protein in atherosclerotic plaque. Outer membrane vesicles (OMVs), secreted by H. pylori, exert effects in the distant organ or tissue. However, whether or not OMVs from H. pylori are involved in the pathogenesis of atherosclerosis remains unknown. Our present study found that treatment with OMVs from CagA-positive H. pylori accelerated atherosclerosis plaque formation in ApoE–/– mice. H. pylori-derived OMVs inhibited proliferation and promoted apoptosis of human umbilical vein endothelial cells (HUVECs), which was also reflected in in vivo studies. These effects were normalized to some degree after treatment with lipopolysaccharide (LPS)-depleted CagA-positive OMVs or CagA-negative OMVs. Treatment with H. pylori-derived OMVs increased reactive oxygen species (ROS) levels and enhanced the activation of nuclear factor-κB (NF-κB) in HUVECs, which were reversed to some degree in the presence of a superoxide dismutase mimetic TEMPOL and a NF-κB inhibitor BAY11-7082. Expressions of interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α), two inflammatory factors, were augmented after treatment with OMVs from H. pylori. These suggest that H. pylori-derived OMVs accelerate atherosclerosis plaque formation via endothelium injury. CagA and LPS from H. pylori-OMVs, at least in part, participate in these processes, which may be involved with the activation of ROS/NF-κB signaling pathway. These may provide a novel strategy to reduce the incidence and development of atherosclerosis.

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A Sensitive and Facile Assay for the Measurement of Activated Protein C Activity Levels in Vivo

Thrombosis and Haemostasis ◽

10.1055/s-0038-1651630 ◽

1993 ◽

Vol 69 (05) ◽

pp. 441-447 ◽

Cited By ~ 16

Author(s):

Carolyn L Orthner ◽

Billy Kolen ◽

William N Drohan

Keyword(s):

Protein C ◽

Plasma Concentrations ◽

Limited Proteolysis ◽

Activated Protein C ◽

Microtiter Plate ◽

Reversible Inhibitor ◽

Activity Levels ◽

Standard Curve ◽

Agarose Beads

SummaryActivated protein C (APC) is a serine protease which plays an important role as a naturally occurring antithrombotic enzyme. APC, which is formed by thrombin-catalyzed limited proteolysis of the zymogen protein C, functions as an anticoagulant by proteolytic inactivation of the coagulation cofactors VIIIa and Va. APC is inhibited by several members of the serpin family as well a by α2-macroglobulin. APC is being developed as a therapeutic for the prevention and treatment of thrombosis. We have developed an assay to quantify circulating levels of enzymatically active APC during its administration to patients, in healthy individuals, and in various disease states. This assay utilizes an EDTA-dependent anti-protein C monoclonal antibody (Mab) 7D7B10 to capture both APC and protein C from plasma, prepared from blood collected in an anticoagulant supplemented with the reversible inhibitor p-aminobenzamidine. Mab 7D7B10-derivatized agarose beads are added to the wells of a 96-well filtration plate, equilibrated with Tris-buffered saline, and incubated for 10 min with 200 μl of plasma. After washing, APC and protein C are eluted from the immunosorbent beads with a calcium-containing buffer into the wells of a 96-well microtiter plate containing antithrombin III (ATIII) and heparin. The amidolytic activity of APC is then measured on a kinetic plate reader following the addition of L-pyroglutamyl-L-prolyl-L-arginine-p-nitroanilide (S-2366) substrate.The rate of substrate hydrolysis was proportional to APC concentration over a 200-fold concentration range (5.0 to 1,000 ng/ml) when measured continuously over a 15 to 30 min time period. The coefficient of variation was 5.9% at 35 ng/ml and 8.8% at 350 ng/ml APC. The sensitivity of the assay could be increased by measuring the amount of color produced after longer incubation times in the endpoint mode. The measured APC activity levels were little affected by varying protein C or prothrombin over the extremes of 0 to 150% of normal plasma concentrations. By constructing the standard curve in protein C-deficient plasma, the concentration of APC activity in normal pooled plasma was determined to be 2.8 ng/ml (45 pM), which represents 0.08% of the protein C concentration. The assay was approximately 50-fold more sensitive than the identical assay, but using Mab-coated microtiter wells rather than immunosorbent beads as the capture step.

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