Gracjana Krzysiek-Maczka

The Impact of Asymmetric Dimethylarginine (ADAMA), the Endogenous Nitric Oxide (NO) Synthase Inhibitor, to the Pathogenesis of Gastric Mucosal Damage

This review was designed to provide an update on the role of asymmetric arginine (ADMA), the endogenous inhibitor of nitric oxide (NO) synthase in the pathophysiology of the upper gastrointestinal (GI) tract. Numerous studies in the past confirmed that NO is a multifunctional endogenous gas molecule involved in most of the body organs functional and metabolic processes including the regulation of gastrointestinal (GI) secretory functions, motility, maintenance of GI integrity, gastroprotection and ulcer healing. NO is metabolized from L-arginine by enzymatic reaction in the presence of constitutive NO synthase. In upper GI tract, NO acts as a potent vasodilator known to increase gastric mucosa blood flow, regulates the secretion of mucus and bicarbonate, inhibits the gastric secretion and protects the gastric mucosa against the damage induced by a variety of damaging agents and corrosive substances. In contrast, ADMA first time described by Vallance and coworkers in 1992, is synthesized by the hydrolysis of proteins containing methylated arginine amino acids located predominantly within the nucleus of cells. This molecule has been shown to competitively inhibit NO synthase suggesting its regulatory role in the functions of vascular endothelial cells and systemic circulation in humans and experimental animals. Nowadays, ADMA is a potentially important risk factor for coronary artery diseases and a marker of cardiovascular risk. Increased plasma levels of ADMA have been documented in several conditions that are characterized by endothelial dysfunction, including hypertension, hypercholesterolemia, hyperglycemia, renal failure and tobacco exposure. The role of ADMA in other systems including GI-tract has been so far less documented. Nevertheless, ADMA was shown to directly induce oxidative stress and cell apoptosis in gastric mucosal cells in vitro and to contribute to the inflammatory reaction associated with major human pathogen to gastric mucosa, Helicobacter pylori (H.pylori). Infection of gastric mucosa with this germ or H. pylori water extract led to marked increase in the plasma concentration of ADMA and significantly inhibited bicarbonate secretion, considered as one of the important components of upper GI-tract defense system. When administered to rodents, ADMA aggravated gastric mucosal lesions injury induced by cold stress, ethanol and indomethacin and this worsening effect on gastric lesions was accompanied by the significant increase in the plasma level of ADMA. This exaggeration of gastric lesions by ADMA was coincided with the inhibition of NO, the suppression of gastric blood flow and excessive release of proinflammatory cytokine TNF-α. This metabolic analog of L-arginine applied to rats was exposed to water immersion and restraint stress and ischemia-reperfusion, causing an elevation of plasma levels of ADMA and gastric MDA content, which is the marker of lipid peroxidation. These effects, including the rise in the plasma levels of ADMA in rats with stress and ischemia-reperfusion-induced gastric lesions, were attenuated by concomitant treatment with L-arginine, the substrate for NO-synthase, and superoxide dismutase (SOD), a reactive oxygen metabolite scavenger added to ADMA. We conclude that ADMA could be considered as an important factor contributing to the pathogenesis of gastric mucosal damage and inflammatory reaction in H. pylori-infected stomach due to inhibition of NO, suppression of GI microcirculation, and the proinflammatory and proapoptotic actions of this arginine analog.

Moderate exercise training attenuates the severity of experimental rodent colitis: the importance of crosstalk between adipose tissue and skeletal muscles.

Although progress has been recently made in understanding of inflammatory bowel diseases (IBD), their etiology is unknown apart from several factors from adipose tissue and skeletal muscles such as cytokines, adipokines, and myokines were implicated in the pathogenesis of ulcerative colitis. We studied the effect high-fat diet (HFD; cholesterol up to 70%), low-fat diet (LFD; cholesterol up to 10%), and the normal diet (total fat up to 5%) in rats with TNBS colitis forced to treadmill running exercise (5 days/week) for 6 weeks. In nonexercising HFD rats, the area of colonic damage, colonic tissue weight, the plasma IL-1β, TNF-α, TWEAK, and leptin levels, and the expression of IL-1β-, TNF-α-, and Hif1α mRNAs were significantly increased and a significant fall in plasma adiponectin and irisin levels was observed as compared to LFD rats. In HFD animals, the exercise significantly accelerated the healing of colitis, raised the plasma levels of IL-6 and irisin, downregulated the expression of IL-1β, TNF-α, and Hif1α, and significantly decreased the plasma IL-1β, TNF α, TWEAK, and leptin levels. We conclude that HFD delays the healing of colitis in trained rats via decrease in CBF and plasma IL-1β, TNF-α, TWEAK, and leptin levels and the release of protective irisin.

Exogenous Asymmetric Dimethylarginine (ADMA) in Pathogenesis of Ischemia-Reperfusion-Induced Gastric Lesions: Interaction with Protective Nitric Oxide (NO) and Calcitonin Gene-Related Peptide (CGRP)

Asymmetric dimethylarginine (ADMA) is an endogenous nitric oxide (NO) synthesis inhibitor and pro-inflammatory factor. We investigated the role of ADMA in rat gastric mucosa compromised through 30 min of gastric ischemia (I) and 3 h of reperfusion (R). These I/R animals were pretreated with ADMA with or without the combination of l-arginine, calcitonin gene-related peptide (CGRP) or a small dose of capsaicin, all of which are known to afford protection against gastric lesions, or with a farnesoid X receptor (FXR) agonist, GW 4064, to increase the metabolism of ADMA. In the second series, ADMA was administered to capsaicin-denervated rats. The area of gastric damage was measured with planimetry, gastric blood flow (GBF) was determined by H2-gas clearance, and plasma ADMA and CGRP levels were determined using ELISA and RIA. ADMA significantly increased I/R-induced gastric injury while significantly decreasing GBF, the luminal NO content, and the plasma level of CGRP. This effect of ADMA was significantly attenuated by pretreatment with CGRP, l-arginine, capsaicin, or a PGE2 analogue. In GW4064 pretreated animals, the I/R injury was significantly reduced and this effect was abolished by co-treatment with ADMA. I/R damage potentiated by ADMA was exacerbated in capsaicin-denervated animals with a further reduction of CGRP. Plasma levels of IL-10 were significantly decreased while malonylodialdehyde (MDA) and plasma TNF-α contents were significantly increased by ADMA. In conclusion, ADMA aggravates I/R-induced gastric lesions due to a decrease of GBF, which is mediated by a fall in NO and CGRP release, and the enhancement of lipid peroxidation and its pro-inflammatory properties.

New satiety hormone nesfatin-1 protects gastric mucosa against stress-induced injury: Mechanistic roles of prostaglandins, nitric oxide, sensory nerves and vanilloid receptors

Nesfatin-1 belongs to a family of anorexigenic peptides, which are responsible for satiety and are identified in the neurons and endocrine cells within the gut. These peptides have been implicated in the control of food intake; however, very little is known concerning its contribution to gastric secretion and gastric mucosal integrity. In this study the effects of nesfatin-1 on gastric secretion and gastric lesions induced in rats by 3.5h of water immersion and restraint stress (WRS) were determined. Exogenous nesfatin-1 (5-40μg/kg i.p.) significantly decreased gastric acid secretion and attenuated gastric lesions induced by WRS, and this was accompanied by a significant rise in plasma NUCB2/nefatin-1 levels, the gastric mucosal blood flow (GBF), luminal NO concentration, generation of PGE2 in the gastric mucosa, an overexpression of mRNA for NUBC2 and cNOS, as well as a suppression of iNOS and proinflammatory cytokine IL-1β and TNF-α mRNAs. Nesfatin-1-induced protection was attenuated by suppression of COX-1 and COX-2 activity, the inhibition of NOS with L-NNA, the deactivation of afferent nerves with neurotoxic doses of capsaicin, and the pretreatment with capsazepine to inhibit vanilloid VR1 receptors. This study shows for the first time that nesfatin-1 exerts a potent protective action in the stomach of rats exposed to WRS and these effects depend upon decrease in gastric secretion, hyperemia mediated by COX-PG and NOS-NO systems, the activation of vagal and sensory nerves and vanilloid receptors.

Role of angiotensin-(1-7) in gastroprotection against stress-induced ulcerogenesis. The involvement of Mas receptor, nitric oxide, prostaglandins and sensory neuropeptides

Angiotensin-(1–7) [Ang-(1–7)] is a major vasoactive metabolite of angiotensin I (Ang I), both being important components of the renin-angiotensin system (RAS). Ang-(1–7) acting via Mas receptor was documented in kidneys, heart, brain, and gastrointestinal (GI)-tract. We studied the gastroprotective activity of exogenous Ang-(1–7) in rats exposed to water immersion and restraint stress (WRS) without or with A-779 [d-Ala7-Ang-(1–7), an antagonist of Ang-(1–7) Mas receptors], AVE 0991 (5-formyl-4-methoxy-2-phenyl-1[[4-[2-(ethylaminocarbonylsulfonamido)-5-isobutyl-3-thienyl]-phenyl]-methyl]-imidazole), the agonist of Ang-(1–7) receptor, as well as the inhibition of nitric-oxide (NO) synthase, the suppression of cyclo-oxygenase (COX)-1 (indomethacin, SC-560 [5-(4-chloro-phenyl)-1-(4-methoxyphenyl)-3-trifluoromethyl-pyrazole]), the activity COX-2 (rofecoxib), and denervation with capsaicin. The mRNA expression of constitutively expressed nitric-oxide synthase (cNOS), inducible nitric-oxide synthase (iNOS), interleukin (IL)-1β, and tumor necrosis factor (TNF)-α was analyzed by reverse transcription polymerase chain reaction. The WRS lesions were dose-dependently reduced by pretreatment with Ang-(1–7), which also caused an increase in gastric blood flow (GBF) and luminal content of NO. COX-1 and COX-2 inhibitors or L-NNA (N5-[imino(nitroamino)methyl]-L-ornithine) reversed the reduction in lesion number and the rise in GBF evoked by Ang-(1–7). Ang II augmented the WRS lesions, decreased GBF and increased the plasma IL-1β and TNF-α levels. Capsaicin denervation attenuated the reduction of Ang-(1–7)-induced gastric lesions and the rise in GBF; these effects were restored by supplementation with calcitonin gene–related peptide (CGRP). The cNOS mRNA was upregulated while iNOS, IL-1β and TNF-α mRNAs were downregulated in Ang-(1–7)-pretreated rats. We conclude that Ang-(1–7), in contrast to Ang II, which worsened WRS ulcerogenesis, affords potent gastroprotection against WRS ulcerogenesis via an increase in GBF mediated by NO, endogenous prostaglandins, sensory neuropeptides, and anti-inflammatory action involving the inhibition of proinflammatory markers iNOS, IL-1β, and TNF-α.

Role of Helicobacter pylori infection in cancer-associated fibroblast-induced epithelial-mesenchymal transition in vitro

Major human gastrointestinal pathogen Helicobacter pylori (H. pylori) colonizes the gastric mucosa causing inflammation and severe complications including cancer, but the involvement of fibroblasts in the pathogenesis of these disorders in H. pylori‐infected stomach has been little studied. Normal stroma contains few fibroblasts, especially myofibroblasts. Their number rapidly increases in the reactive stroma surrounding inflammatory region and neoplastic tissue; however, the interaction between H. pylori and fibroblasts remains unknown. We determined the effect of coincubation of normal rat gastric fibroblasts with alive H. pylori (cagA+vacA+) and H. pylori (cagA−vacA−) strains on the differentiation of these fibroblasts into cells possessing characteristics of cancer‐associated fibroblasts (CAFs) able to induce epithelial‐mesenchymal transition (EMT) of normal rat gastric epithelial cells (RGM‐1).

279 Exogenous and Endogenous Hydrogen Sulfide (H2S) in Gastroprotection Against Ischemia/Reperfusion Lesions Progressing Into Chronic Ulcers. Role of Endogenous Prostaglandins, Sensory Neuropeptides and Vanilloid Receptors

and, 3) human esophageal epithelial HET-1A cell line, which has many features of EPC. Organ cultures and HET-1A cells were treated with vehicle or HTL (1-5 mg/ml) for 1-4 hrs; HET-1A cells were also treated with either: 1 μg/ml misoprostol (PGE1 analog); 1 μg/ ml 16,16 dmPGE2; 0.4 mM protein kinase A (PKA) inhibitor, Rp-cAMP; or, 50 ng/ml KGF. Studies: 1) epithelial integrity using confocal microscopy; 2) expression of COX2, EP1-4 receptors, KGF and KGFR using immunostaining and immunoblotting; 3) In HET-1A cells we quantified: a) expression of COX2, EP receptors1-4, KGF and KGFR; b) [i]cAMP (using Biotrack EIA); cAMP response element-binding protein (CREB); and, c) cell proliferation (BrdU assay) prior to and after treatment with PGEs or KGF. Results: In normal esophagus, EPC expressed COX2, EP receptor2 and KGFR. Stromal cells of submucosa expressed KGF. In esophageal explants, HTL treatment increased COX2 and KGFR expression in EPC by 69±5% and 72±6% (both p 50% (both p 163-fold, CREB phosphorylation by 62% and cell proliferation by 64% (both p<0.01). These effects were abolished by pretreatment with PKA inhibitor. KGF stimulated HET-1A cell proliferation by 68% (p< 0.01). Conclusions: 1) This is the first demonstration that COX2 and EP-2 receptor are expressed and co-localized in esophageal EPC indicating that COX2 generated PGEs play an autocrine regulatory role in EPC proliferation and survival via the c-AMP/CREB/PKA signaling pathway. 2) KGFR is expressed in EPC while KGF is expressed in stromal cells reflecting local mesenchymal-EPC interactions. 3) HTL treatment significantly upregulates expression of COX2 and KGFR in EPC. 4) These findings elucidate new mechanisms regulating EPC, esophageal mucosal defense and provide novel insight into the protective and healing actions of hydrotalcite.

Su1755 Hydrogen Sulfide Exhibits a Potent Gastroprotective Action Against Stress-Induced Gastric Lesions. Involvement of Prostaglandins and Sensory Afferents

G A A b st ra ct s of the oxyntic mucosa, increased apoptosis, oxidative stress, and surface layer erosion. Inflammation was also observed in the mucosa as shown by the activation of the NFkB pathway and increases in TFF1, TIMP1 and Cox-2 expression. The pit region of the mucosa was affected by changes in cell interactions due to changes in beta-catenin and E-cadherin expression, loss of interaction in adherent junctions and activation of the beta-catenin pathway. Many of these changes, already detectable in 20-day-old fetuses, were specific of the offspring and were not observed in dams. All pups were refed a normal diet after weaning. We evaluated the persistence of stomach changes by comparing adult rats with MMD during gestation and suckling period and normal diet thereafter with control rats. The gastric mucosa thickness remained smaller in than that of control rats. SOD expression, NFkB activity, and apoptotic cell loss remained increased. CONCLUSIONS: These results indicate that the deleterious effects of MDD during gestation and lactation were accompanied by inflammatory and ulcerative states that were not reversed by the ingestion of a normal diet after weaning.