Portal hypertension predisposes the gastric mucosa to hemorrhagic shock/reperfusion injury

We have previously demonstrated that the portal hypertensive (PHT) gastric mucosa has unique functional and morphologic features which predispose it to increased damage by noxious agents such as aspirin or alcohol. In this study we examine the PHT gastric mucosa following hemorrhagic shock and reperfusion. Portal hypertension was produced in 12 rats using staged portal vein occlusion, and seven animals (controls) underwent sham procedures. All rats received 2 ml 0.1 N HCl intragastrically. Shock was then induced by withdrawing blood to reduce systemic arterial pressures to approximately 30 mm Hg for 20 min. Blood was then reinfused and stomachs were excised 20 min later for gross and microscopic quantitation of injury. We found that gross damage to gastric glandular mucosa was more than doubled in PHT rats over sham-operated controls. In PHT gastric mucosa, deep histologic necrosis involved 22.5 +/- 3.8% of mucosal section lengths compared with 6.9 +/- 2.0% in control mucosa (P less than 0.005). Histologically, mucosal injury patterns were predominantly ischemic, with deep necrosis and disintegration of glands and microvessels, all prominently more extensive in PHT mucosa. We conclude that PHT gastric mucosa has significantly increased susceptibility to damage following hemorrhagic shock/reperfusion compared with portal normotensive mucosa. Since hemorrhagic shock is a frequent complication of portal hypertension, this study suggests clinical investigations of gastric mucosal function and pathophysiology in PHT patients following hemorrhage and resuscitation.     

Epidermal growth factor protects portal hypertensive gastric mucosa in ischemia/reperfusion: the role of capillary endothelia and prostaglandins.

BACKGROUND. Epidermal growth factor (EGF) protects gastric mucosa against a variety of injurious agents, but the mechanism is unclear. Because the abnormal microvasculature of portal hypertensive (PHT) gastric mucosa is a major target of ischemia/reperfusion (I/R) injury, we used this model to assess EGF's protective role at the microvascular level. METHODS. Rats with PHT (staged portal vein ligation) received either EGF, 20 micrograms/kg, or saline solution intravenously, with or without indomethacin pretreatment (20 mg/kg subcutaneously). I/R was produced by withdrawing blood to systemic pressures of 30 mm Hg for 20 minutes and reinfusing it. Stomachs were excised 20 minutes later and evaluated for gross and histologic necrosis, microvascular permeability, mucosal ultrastructure and vimentin, and cyclooxygenase immunofluorescence. RESULTS. In saline-treated rats, gross and histologic damage involved 46% +/- 3% of glandular mucosa and 23% +/- 3% of mucosal sections, respectively. Microvascular permeability was increased 43-fold over that of normal control rats. Vimentin immunofluorescence intensity was reduced to 36% +/- 4% that of normal control rats. EGF pretreatment reduced histologic necrosis to 2% +/- 1% (p less than 0.01). Microvascular permeability and vimentin intensity were almost normalized. Indomethacin partially reversed the mucosal protection induced by EGF. CONCLUSIONS. EGF significantly reduces I/R injury to PHT gastric mucosa. Microvascular endothelia of PHT gastric mucosa are the major target of I/R injury and the site of EGF's protective action. Prostaglandins in part mediate EGF's protective action.     

Efficacy of HSP72 induction in rat liver by orally administered geranylgeranylacetone

Abstract It is well known that heat‐shock proteins (HSPs) have a cyto‐protective function as “molecular chaperones” when cells are exposed to several stress conditions. Geranylgeranylacetone (GGA) is an antiulcer drug that was developed in Japan and it has recently been reported to induce HSP72 in rat gastric mucosa. In this experiment, we investigated the induction of HSP72 in rat liver in response to oral administration of GGA and assessed its ability to induce tolerance to warm ischemic injury by this approach. We prepared donor rats by orally administering GGA to them and compared HSP72 expression in graft liver, survival rates, and serum TNF‐α concentrations after liver transplantation with the findings in controls. The survival rates were significantly increased when the livers were obtained from donor rats given GGA. Western blotting revealed expression of HSP72 in graft livers given GGA, and the serum TNF‐α levels were significantly suppressed in the rats given GGA. Oral administration of GGA induced HSP72 in graft livers, and they were better able to tolerate warm ischemic injury. Oral administration of GGA appears to provide a promising new strategy for preventing ischemia‐reperfusion injury.     

New insights into impairment of mucosal defense in portal hypertensive gastric mucosa

Portal hypertension (PHT) increases susceptibility of the gastric mucosa to injury. The aim of this study was to investigate whether PHT affects rat gastric mucosal defense mechanisms in vivo at the preepithelial, epithelial, and/or post-epithelial levels. PHT was produced in rats by staged portal vein ligation and sham-operated (SO) rats served as controls. The gastric mucosa was exposed, chambered, and continuously superfused with buffers under in vivo microscopy. We measured gastric mucosal gel layer thickness, surface epithelial cell intracellular pH (pH₁), mucosal blood flow, and mucosal/serosal oxygenation. In PHT rats, gastric mucosal gel layer thickness was significantly reduced (88 ±16 μm in PHT rats vs. 135 ±25 μm in SO rats; P <0.0001), and the surface epithelial cell pH₁ was significantly decreased (6.80 ± 0.11 in PHT rats vs. 7.09 ± 0.21 in SO rats; P <0.01). Although total gastric mucosal blood flow was significantly increased in PHT rats by 72 % (P <0.05), the oxygenation of the gastric mucosal surface was decreased by 42 % (P <0.05) compared with SO rats. PHT impairs pre-epithelial (mucosal gel layer thickness), epithelial (pH₁), and post-epithelial (maldistribution of blood flow) components of the gastric mucosal barrier. These findings can explain the increased susceptibility of portal hypertensive gastric mucosa to injury. Supported by Veterans Affairs Medical Research Service Merit Review Awards (J.D.K., I.J.S., and AS.T.) and a REAP award. Presented at the Fortieth Annual Meeting of The Society for Surgery of the Alimentary Tract, Orlando, Fla., May 16–19, 1999.     

Overexpression of inducible nitric oxide synthase in gastric mucosa of rats with portal hypertension: correlation with gastric mucosal damage

BACKGROUND: An increased biosynthesis of nitric oxide (NO) has been implicated in the hyperdynamic circulation and development of collaterals of portal hypertension (PHT) because of its potent vasodilatory effects. NO is synthesized from L-arginine by three different isozymes of nitric oxide synthase (nNOS, iNOS and eNOS). Thus, the expression of inducible NOS (iNOS) might account for NO overproduction in PHT. However, in previous investigations, the role of iNOS in the pathogenesis of PHT gastropathy remained controversial. Our current study was in both molecular and protein levels to determine whether the expression of iNOS is responsible for PHT gastropathy. MATERIALS AND METHODS: PHT was induced experimentally by partial ligation of the portal vein. Fourteen days after partial ligation of the portal vein, the rats were randomly assigned to receive either vehicle or L-NAME (NOS inhibitor) at doses of 5 mg/kg/day, 10 mg/kg/day, or 25 mg/kg/day by gastric lavage twice a day for 1 week. Sham operated rats served as controls. Northern hybridization and in situ hybridization are used to compare the expression of gastric mucosa iNOS mRNA in the PHT rats and the controls. NO was measured by the Griess method after reduction of nitrate to nitrite with nitrate reductase. Immunohistochemical staining was carried out to detect the iNOS protein. In addition, the severity of gross gastric mucosal lesions was evaluated macroscopically by a gross ulcer index. RESULTS: The iNOS expression at both mRNA and protein was prominently increased in PHT rats, accompanied with the enhanced NO production. The gastric mucosa iNOS mRNA and serum NO levels were significantly decreased after L-NAME administration (P < 0.05). However, the markedly reduced gastric mucosal damage in PHT rats was observed only at high does of L-NAME (25 mg/kg/day) administration. CONCLUSION: PHT triggers overexpression of iNOS mRNA and proteins in rat gastric mucosa, but that this alone does not account for PHT gastropathy.     

Bile acid-induced gastric mucosal injury: significance of portal hypertension and mucosal capillary permeability

Rats with portal vein occlusion (PVO) were studied to determine severity of taurocholate-induced gastric mucosal injury during portal hypertension and following its resolution by collaterals. During the portal hypertensive state, after intragastric taurocholate, PVO rats compared with sham-operated controls had significantly greater macroscopic damage (14 +/- 1 vs 3 +/- 1% of total mucosa) and histologic deep necrosis (25 +/- 4 vs 3 +/- 1% of mucosal length). Gastric mucosal capillary permeability was also significantly greater in PVO rats, as assessed by increased appearance of intravenous Evans blue dye in gastric wall and contents. After 21 days of PVO portal pressures returned to normal. At this time taurocholate-induced gastric mucosal damage and capillary leak became similar in PVO and sham-operated rats. It is concluded that the portal hypertensive state alone predisposes to severe gastric mucosal damage and capillary leak.     

The effect of portal hypertension on the glycoprotein biosynthesis of rat gastric mucosa.

The aim of this study was to assess the influence of epidermal growth factor (EGF) on glycoprotein biosynthesis in portal hypertensive (PHT) gastric mucosa. Portal-vein ligation (PVL) for a period of 4 weeks was applied to 40 male Wistar rats to produce experimental portal hypertension. The rats were subdivided into four groups. Human EGF was administrated to these four groups of animals at a does of 0, 10, 25, and 50 microg/kg/day for 7 days. An additional group of 10 rats without PVL and EGF pretreatment was employed as a control. The severity of gross gastric mucosal lesions was evaluated macroscopically by a gross ulcer index. Glycoprotein biosynthesis of the gastric mucosa was determined by the incorporation rate of [(3)H]glucosamine. Quantitative changes of gastric mucosal hexosamines were also used for mucosal glycoproteins analyses. The gross mucosal damage was considerably greater in the PVL group without EGF pretreatment than in the EGF-pretreated groups (p <.05). The incorporation rate of [(3)H]glucosamine was significantly higher in the control group and the EGF-pretreated groups than in the PVL group without EGF pretreatment (p <.05). Moreover, the incorporation rate of [(3)H]glucosamine and the gastric mucosal hexosamine content were closely relevant to administration does of human EGF (p <.001). In addition, the reduction of glycoprotein biosynthesis was closely related to the increase in portal pressure (p =.001) and the severity of portal hypertensive gastropathy (p <.001). Our current study shows that the rate of incorporation of glucosamine is decreased in the PHT gastric mucosa and that EGF significantly stimulated glycoprotein synthesis in the PHT gastric mucosa. Accordingly, these findings may be helpful to explain the protective effect of EGF on the PHT gastric mucosa via increased glycoprotein biosynthesis in the stomach.     

替普瑞酮对大鼠肾脏缺血再灌注损伤的保护作用

目的 探讨替普瑞酮对肾脏缺血再灌注损伤的保护作用和可能机制。 方法 应用替普瑞酮(400 mg/kg)诱导雄性SD大鼠肾脏高表达热休克蛋白72（HSP72）。以钳夹大鼠左肾蒂45 min后，松开血管夹并切除右肾，建立大鼠缺血再灌注肾脏损伤模型。假手术组为打开腹腔，分离肾血管周围组织，但不钳夹血管。模型建立后24 h处死大鼠，留取血清测血肌酐（Scr）和尿素氮（BUN）。肾组织石蜡切片行PAS染色，以损伤肾小管所占百分比评分法评估肾组织肾小管损伤程度。TUNEL法检测缺血再灌注损伤时肾脏细胞凋亡的发生情况。Western印迹检测X连锁凋亡抑制蛋白（XIAP）的水平。 结果 缺血再灌注损伤可导致急性肾衰竭，表现为血Scr、BUN明显升高（P < 0.01）；PAS染色显示外髓部有大片肾小管坏死，甚至出现基底膜裸露；TUNEL染色中肾小管上皮细胞TUNEL阳性细胞数明显增多（P < 0.01）；Western印迹结果显示，肾组织XIAP蛋白水平明显降低（P < 0.01）。替普瑞酮处理后，肾组织HSP72表达水平明显增高（P < 0.01）；缺血再灌注所致的肾脏损伤明显改善，包括肾小管的损伤、细胞凋亡以及肾功能。此外，替普瑞酮可稳定肾组织XIAP的蛋白水平（P < 0.05）。 结论 替普瑞酮可诱导肾脏高表达HSP72。替普瑞酮可能通过减少肾脏XIAP蛋白的降解，抑制细胞凋亡，减轻缺血再灌注的肾脏损伤。     

门脉高压性大鼠胃粘膜细胞动力学放射自显影研究

为探讨门脉高压性胃病(PHG)发病机理,用放射自显影~3H-TdR两次脉冲标记法研究大鼠门脉高压时胃粘膜细胞动力学改变,并尝试用降钙素基因相关肽(CGRP)保护胃粘膜。结果表明:大鼠在门脉高压时胃粘膜细胞D?NA合成期、细胞周期延长(P<0.01),标记指数下降(P<0.01);应用CGRP后,DNA合成期、细胞周期缩短,标记指数回升。提示胃粘膜细胞自身代谢障碍,防护能力削弱是PHG细胞动力学机制,CGRP可改善粘膜细胞增殖受抑状态,发挥保护作用。     

The portal hypertensive gastric mucosa: histologic, ultrastructural, and functional analysis after aspirin-induced damage

We assessed macroscopic, histologic, ultrastructural, and functional features of aspirin-induced gastric mucosal injury in portal hypertensive and sham-operated rats. Portal hypertension was produced by staged portal vein ligation. Four hours after intragastric acidified aspirin administration, intraluminal pH in portal hypertensive rats was 6.6 +/- 0.2 and 4.3 +/- 0.5 in sham-operated controls (p less than 0.01). Gross mucosal damage was significantly greater in portal hypertensive rats compared with controls (18 +/- 2 versus 7 +/- 1% of total mucosal area). Histologic deep necrosis involved 22 +/- 2% of mucosal section lengths in portal hypertensive rats compared with 7 +/- 1% in sham-operated rats (p less than 0.01). In portal hypertensive rats, histologic and ultrastructural evaluation demonstrated capillary endothelial abnormalities, arterialization of submucosal veins, and markedly greater severity of microvascular damage than in sham-operated controls. Neutralized aspirin (pH, 7.0) did not produce any significant damage detectable grossly, histologically, or by transmission electron microscopy in portal hypertensive rats. We conclude that acid-dependent aspirin-induced gastric mucosal damage is significantly increased in portal hypertension.     

Acceleration of gastric ulcer healing by omeprazole in portal hypertensive rats. Is its action mediated by gastrin release and the stimulation of epithelial proliferation?

BACKGROUND: Gastric ulcer healing is delayed in patients with portal hypertension (PHT) and often responds poorly to histamine H(2) blockers. Although proton pump inhibitors are more effective anti-ulcer agents, there is little information regarding their efficacy for gastric ulcer in cases of PHT. Therefore, we investigated the effects of a proton pump inhibitor, omeprazole, on the healing of acetic-acid-induced gastric ulcer in PHT rats. METHODS: Animals studied were 80 male Sprague-Dawley rats aged 7 weeks, of which half underwent two-staged portal vein ligation (PHT rats) and half underwent a sham operation (SO rats). Gastric ulcers were induced by acetic acid. Starting from day 4 after ulcer induction, rats received omeprazole or vehicle orally (50 mg/kg) for 5 or 10 days. Ulcer area, proliferating cell nuclear antigen labelling index (PCNA LI), and serum gastrin levels were recorded. RESULTS: PHT significantly inhibited epithelial cell proliferation and delayed gastric ulcer healing as indicated by a decreased PCNA LI at the ulcer margin and almost 2-fold larger ulcer area in PHT versus SO rats 14 days after ulcer induction. Ten-day treatment with omeprazole (vs. vehicle) significantly accelerated ulcer healing to a similar extent in both PHT and SO rats. Serum gastrin levels were significantly higher in PHT rats than in SO rats following treatment with omeprazole. Omeprazole (vs. vehicle) restored the decreased PCNA LI at the ulcer margin in PHT rats to that noted in SO rats. CONCLUSIONS: In PHT rats, omeprazole accelerates gastric ulcer healing, stimulates epithelial cell proliferation at the ulcer margin, and increases serum gastrin levels. Since gastrin is a potent stimulator of gastric epithelial cell proliferation, increased serum gastrin levels may be an important factor in omeprazole-induced stimulation of epithelial cell proliferation and acceleration of gastric ulcer healing in conditions of PHT.     

亚砷酸钠诱导HSP 72在大鼠肝脏热缺血–再灌注损伤中的保护作用

目的:观察亚砷酸钠(SA)诱导大鼠肝脏产生热休克蛋白72(HSP72)的过程及其抗肝脏热缺血-再灌注损伤的作用。材料与方法:将24只SD大鼠随机分成SA组(预先给予SA6mg/kg)和对照组(预先给予生理盐水)。24h后,两组均阻断肝脏中、左叶血供60min,再灌注3、6h;分别用Westernblot检测肝脏中的HSP72表达;外周血测定ALT、AST、LDH;组织学作HE染色、免疫组化测定HSP72。结果:注射SA6mg/kg后12h肝脏开始产生HSP72,24h达到高峰,并持续至60h。肝脏热缺血60min再灌注3、6h后,SA组HSP72显著表达,血清ALT、AST、LDH显著下降(P<0.05),组织学检查显示肝脏损伤明显减轻。结论:SA可诱导大鼠肝脏产生HSP72;SA预先诱导肝脏产生对HSP72可使肝脏抵御热缺血鄄再灌注损伤。     

Topical isoproterenol protects the rat gastric mucosa from ethanol-induced injury

This study evaluated the effects of topical isoproterenol, a beta-adrenergic agonist, on the morphologic damage produced in the gastric mucosa by ethanol. The orogastric instillation of 100% ethanol in rats resulted in gross lesion formation and deep histologic injury in the gastric mucosa. Animals pretreated with oral isoproterenol (50 micrograms/kg, 500 micrograms/kg, 50 mg/kg) showed dose-dependent protection from both the gross and the histologic mucosal injury (P less than 0.01, ANOVA). Pretreatment with propranolol (2 mg/kg/sec) but not indomethacin (5 mg/kg/sec) blocked this protective effect. Isoproterenol had no effect on ethanol-induced mast cell degranulation as both mucosal and submucosal mast cell counts were significantly and equally decreased in all groups treated with 100% ethanol (P less than 0.05). These findings show that topical isoproterenol protects the rat gastric mucosa from both the gross and the histologic injury caused by 100% ethanol. This protection is mediated by a beta-adrenergic receptor mechanism as it can be blocked by prior treatment with propranolol, but does not involve stabilization of mucosal or submucosal mast cell membranes.     

肝硬变门脉高压休克再灌注胃粘膜损伤的实验研究

本实验应用电子自旋共振（ＥＳＲ）技术及自旋捕捉剂ＰＢＮ直接测定肝硬变门脉高压（ＰＨＴ）大鼠胃粘膜在休克再灌注前后及应用超氧化物歧化酶（ＳＯＤ）、丹参（ＲＳＭ）治疗后氧自由基（ＯＦＲ）的动态变化，同时检测胃粘膜中ＳＯＤ活性，并观察同期胃粘膜的光镜、电镜病理改变。结果：ＰＨＴ胃粘膜在休克再灌注过程中有大量ＯＦＲ产生，粘膜损伤的严重程度与ＯＦＲ含量及ＳＯＤ活性密切相关；ＰＨＴ胃粘膜更易受休克再灌注时ＯＦＲ的损伤，早期应用抗氧化剂ＳＯＤ及ＲＳＭ，可通过不同机理减轻胃粘膜再灌注损伤。     

Geranylgeranylacetone suppresses inflammatory responses and improves survival after massive hepatectomy in rats

Overproduction of heat shock protein 70 (HSP70) in the liver protects hepatocytes under various pathologic conditions. In this study we examined the effects of a nontoxic HSP70 inducer, geranylgeranylacetone (GGA), on acute hepatic failure after 95% hepatectomy in rats. When GGA (100 mg/kg) or vehicle was intragastrically administered to rats 4 hours before 95% hepatectomy, all 25 rats pretreated with vehicle died within 60 hours after the operation, whereas 10 of 25 rats pretreated with GGA survived. During the 24-hour postoperative period, GGA significantly suppressed the release of aspartate or alanine aminotransferase and elevation of the serum interleukin-6 level, and completely inhibited an increase in the serum level of tumor necrosis factor-alpha. Histologic examinations showed that GGA prevented hemorrhagic necrosis, which was observed in vehicle-treated livers more than 12 hours after the operation. During the 24-hour postoperative period, HSP70 induction was absent in remnant livers of vehicletreated rats. In contrast, GGA stimulated the HSP70 mRNA expression and HSP70 accumulation within 4 hours, and viable hepatocytes contained abundant HSP70 in their nuclei. Our results suggest that GGA may prevent acute liver failure after massive hepatectomy, at least in part, by enhancing HSP70 induction in the remnant liver. Presented at the Forty-Second Annual Meeting of The Society for Surgery of the Alimentary Tract, Atlanta, Georgia, May 20–23, 2001 (oral presentation). Supported by a Grant-in-Aid for Scientific Research (No. 12557105) from the Japanese Ministry of Education, Science and Culture (K.R.).     

The physiology of postvagotomy duodenal ulcer healing: a prostaglandin connection

This study evaluated the type and mode of action exerted by truncal vagotomy on mucosal prostaglandin content in the rat. Ninety-six rats were equally divided into sham-operated, vagotomy, atropine-treated, and cimetidine-treated groups. Each group was subdivided into stressed (cold-restraint) and nonstressed cells. Gastric mucosal injury was graded, and duodenal mucosa was processed for determination of prostaglandin content. Results included: (1) significant increases in duodenal mucosal prostaglandins in all experimental groups compared with sham-operated controls, (2) decreases in duodenal mucosal prostaglandin content associated with stress in all groups, and (3) significantly less gastric mucosal injury in all stressed, experimentally treated rats compared with sham-operated animals. These results demonstrate that attenuation of gastric acid-secretion, achieved either surgically or pharmacologically and proved by decrease in stress gastric injury, is associated with an increase in the content of duodenal mucosal prostaglandins. Such augmentation of mucosal prostaglandins could account for, along with the direct acid-lowering effect of vagotomy, the success of truncal vagotomy in the surgical cure of duodenal ulcer disease.     

Protective effect of pentoxifylline on gastric mucosa

Pentoxifylline (PF) has been shown to increase tissue oxygen tension. This study was performed to determine if PF has a protective effect on the gastric mucosa against alcohol (EtOH)-induced injury. Fasted Sprague-Dawley rats were pretreated with randomized test solution (control, normal saline, or PF, 75 mg/kg) intraperitoneally (ip). At 30 min, 100% EtOH (pH 8.5) was given intragastric. At 90 min, laparotomy was performed and gastric serosal stomach surface oxygen tensions (pO2) were measured. Stomachs were excised and opened and pH was measured. Photographs were taken and sections were obtained for histologic analysis to determine mucosal injury. The PF-pretreated rats had significantly higher serosal pO2 and significantly lower intragastric pH than control animals. There was significantly less gross and histologic mucosal injury in PF-treated animals. We conclude that PF is protective against EtOH gastric mucosal injury. This effect correlates with increased gastric serosal pO2 and is likely due to improved microcirculatory blood flow following PF administration.     

Injury to the Gastric Mucosa and Cellular Dynamics in a Rat Model of Duodenogastric Reflux: The Possible Significance of Gastrin Induction and a Heat Shock Protein

Injury to the gastric mucosa caused by duodenogastric reflux (DGR) is often encountered after gastrectomy or truncal vagotomy (V) with pyloroplasty. This study was designed to investigate the histological features of the gastric mucosa under such conditions. A rat model of DGR and DGR+V was established and the thickness of the oxyntic mucosa was measured. Cellular dynamics in the presence of injury to the gastric mucosa caused by DGR were investigated by the immunohistochemical staining of bromodeoxyuridine (BrdU) and heat shock protein 70 (HSP70). The relationship between persistent hypergastrinemia and mucosal injury was also studied. Duodenogastric reflux activated the intracellular induction of HSP70 in our rat model of DGR. Hypergastrinemia was noted in the V group. Compared with values from the DGR group, the numbers of BrdU-labeled cells increased, the glandular proliferation zone expanded, and the thickness of the oxyntic mucosa was significantly higher in the DGR+V group. Compared with the DGR group, there was greater induction of HSP in the DGR+V group during the acute stage. This finding suggests that denervation of the gastric mucosa and hypergastrinemia after vagotomy may be associated with the expression of HSP. Received: May 17, 1999 / Accepted: May 30, 2000