Defective mitogen-activated protein kinase (ERK2) signaling in gastric mucosa of portal hypertensive rats: potential therapeutic implications.

Portal hypertensive (PHT) gastropathy is a frequent, serious complication of liver cirrhosis. PHT gastric mucosa has numerous abnormalities such as reduced mucosal potential differences, reduced surface oxygenation, and increased susceptibility to injury caused by alcohol, aspirin, and other noxious factors. Because such mucosal injury is initially mediated by oxygen free radicals, and because mitogen-activated protein (MAP) kinase (ERK2) protects against cellular stress and induces cell proliferation, we postulated that oxidative stress-induced ERK2 activation is defective in PHT gastric mucosa. Here we show that in PHT gastric mucosa, ERK2 activation by oxidative stress is impaired. This impairment is mediated by overexpression of MAP kinase phosphatase-1 (MKP-1), which results from the underlying and continual oxidative state associated with portal hypertension, and is ameliorated by inhibiting MKP-1. Furthermore, we found that supplementing vitamin E, a free radical scavenger, reduces the oxidative state in PHT gastric mucosa, normalizes MKP-1 expression, and thereby reverses impairment of oxidative stress-induced ERK2 activation. Finally, we show that orally administered vitamin E completely reverses the increased susceptibility of PHT gastric mucosa to alcohol injury. Our findings point to a new molecular and mechanistic basis for PHT gastropathy and provide a new therapeutic modality for protection of PHT gastric mucosa.

Gene therapy for gastric ulcers with single local injection of naked DNA encoding VEGF and angiopoietin-1.

BACKGROUND & AIMS: Angiogenesis, formation of new capillary blood vessels, is crucial for gastroduodenal ulcer healing because it enables delivery of oxygen and nutrients to the healing site. Because angiogenesis is stimulated by vascular endothelial growth factor (VEGF) and angiopoietin-1 (Ang1), we studied whether local gene therapy with nonviral DNA encoding VEGF and/or Ang1 into the ulcer base could accelerate ulcer healing through enhanced angiogenesis. METHODS: Gastric ulcers were induced in rats by acetic acid applied to the serosal surface of the stomach, and the site around the ulcer was injected with nonviral plasmid-encoding full-length complementary DNA (cDNA) of human recombinant (rh) VEGF165, rhAng1, or their combination. For some studies, neutralizing anti-VEGF antibody was administered. RESULTS: Single local injection of plasmids encoding VEGF165 and Ang1 significantly increased neovascularization and accelerated ulcer healing. A neutralizing anti-VEGF antibody significantly reduced the acceleration of ulcer healing resulting from the treatment. Coinjection of both plasmids encoding rhVEGF165 and rhAng1 resulted in formation of more mature vessels and to more complete restoration of gastric glandular structures within the ulcer scar. However, this did not result in further reduction of ulcer size. CONCLUSIONS: VEGF and Ang1 gene therapy, with limited duration of target gene expression, significantly accelerates gastric ulcer healing. Coinjection of both plasmids leads to more complete structural restoration. Inhibition of accelerated healing by a neutralizing anti-VEGF antibody indicates an essential role for VEGF and enhanced angiogenesis in ulcer healing.

Portal hypertensive gastric mucosa has reduced activation of MAP kinase (ERK2) in response to alcohol injury: a key to impaired healing?

Portal hypertensive (PHT) gastric mucosa has increased susceptibility to injury and impaired mucosal healing. Because our previous study showed that ulcer-induced activation of mitogen-activated protein (MAP) kinase (ERK) plays a pivotal role in gastric mucosal healing, we investigated whether ERK activation is altered in PHT gastric mucosa following alcohol injury. We studied ERK2 phosphorylation and activity and expression of MAP kinase phosphatase-1 (MKP-1) in gastric mucosa of PHT and sham-operated (SO) normal rats both at baseline and following alcohol injury. In SO gastric mucosa, ERK2 phosphorylation and activity were significantly increased time-dependently following alcohol injury: by 221% and 137%, respectively at 24 h vs. baseline. In contrast, in PHT gastric mucosa following alcohol injury, neither ERK2 phosphorylation nor activity was increased versus baseline. In PHT gastric mucosa, MKP-1 mRNA and protein expression were increased at baseline versus SO rats and were increased further following alcohol injury with values higher by 20%-40% at each study time versus SO rats. Because ERK2 is crucial for mucosal healing, reduced ERK2 activation resulting from the overexpression of MKP-1 might be the basis for the impaired mucosal healing in PHT gastric mucosa.

Despite activation of EGF-receptor-ERK signaling pathway, epithelial proliferation is impaired in portal hypertensive gastric mucosa: relevance of MKP-1, c-fos, c-myc, and cyclin D1 expression.

Portal hypertensive (PHT) gastric mucosa has increased susceptibility to injury and impaired mucosal healing. Our previous study demonstrated increased ERK activation and MAP kinase phosphatase-1 (MKP-1) overexpression in PHT gastric mucosa. However, it remains unknown which tyrosine kinase receptors are involved in ERK activation and whether ERK activation results in increased cell proliferation. We examined whether EGF receptor (EGF-R) is involved in ERK activation and whether ERK activation triggers epithelial proliferation in PHT gastric mucosa. In gastric mucosa of PHT and sham-operated (SO) rats we studied: (1) EGF-R mRNA and protein expression as well as phosphorylation and membrane protein tyrosine kinase (PTK) activity; (2) ERK2 phosphorylation and activity; (3) MKP-1 mRNA and protein; (4) c-fos, c-myc and cyclin D1 mRNAs, and gastric epithelial proliferation. In PHT gastric mucosa: (1) EGF-R mRNA, protein and phosphorylation and membrane PTK activity were all significantly increased by 38%, 49%, 43% and 49%, respectively; (2) ERK2 phosphorylation and activity were significantly increased by 40% and 50 %, respectively; (3) MKP-1 mRNA and protein expression were significantly increased by 27% and 34%, respectively. In contrast, (4) c-fos, c-myc, and cyclin D1 mRNAs expression were all significantly decreased in PHT gastric mucosa by 36%, 33%, and 49%, respectively, and cell proliferation was significantly lower that in SO rats (11% in PHT vs. 18% in SO). These results suggest that in PHT gastric mucosa, ERK activation is mediated through EGF-R upregulation, but the gastric epithelial proliferation is impaired, possibly by MKP-1 overexpression, leading to reduction of c-fos, c-myc and cyclin D1.

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 pre-epithelial, 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 (pHi), mucosal blood flow, and mucosal/serosal oxygenation. In PHT rats, gastric mucosal gel layer thickness was significantly reduced (88 +/- 16 microm in PHT rats vs. 135 +/- 25 microm in SO rats; P <0.0001), and the surface epithelial cell pHi 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 (pHi), 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.

Antacid talcid activates in gastric mucosa genes encoding for EGF and its receptor. The molecular basis for its ulcer healing action.

In previous studies [Gut 35 (1994) 896-904], we demonstrated that antacid talcid (TAL) accelerates gastric ulcer healing and provides better quality of mucosal restoration within the scar than the omeprazole (OME). However, the mechanisms of TAL-induced ulcer healing are not clear. Since growth factors promote cell proliferation, re-epithelization, angiogenesis and ulcer healing, we studied whether TAL and/or OME affect expression of epidermal growth factor (EGF) and its receptors (EGF-R) in both normal and ulcerated gastric mucosae. Rats with or without acetic acid-induced gastric ulcers (n = 64) received i.g. twice daily 1 mL of either: A) placebo (PLA); B) TAL 100 mg; or C) OME 50 mg x kg(-1) for 14 d. Studies of gastric specimens: 1) ulcer size; 2) quantitative histology; 3) expression of EGF mRNAs was determined by RT/PCR; 4) gastric sections were immunostained with antibodies against EGF and its receptors. In non-ulcerated gastric mucosa of placebo or omeprazole treated group, EGF expression was minimal, while EGF-R was localized to few cells in the mucosal proliferative zone. Gastric ulceration triggered overexpression of EGF and its receptor in epithelial cells of the ulcer margin and scar. In ulcerated gastric mucosa TAL treatment significantly enhanced (versus PLA and omeprazole) expression of EGF and EGF-R. OME treatment reduced expression of EGF in ulcerated mucosa by 55 +/- 2% (P < 0.01). It is concluded that: 1) treatment with TAL activates genes for EGF and its receptor in normal and ulcerated gastric mucosae; 2) since EGF promotes growth of epithelial cells and their proliferation and migration, the above actions of TAL provide the mechanism for its ulcer healing action and improved (versus OME) quality of mucosal restoration.

Sequential expression of adrenomedullin and its receptor during gastric ulcer healing in rats.

Adrenomedullin (AM) is a potent vasodilatory peptide. While its growth-regulating action in some cultured cells has been recognized, expression of AM and its receptor during gastric ulcer healing has not been explored. Specimens of gastric walls from control rats or gastric ulcers were obtained at 1, 3, 7, and 14 days after gastric ulcer induction. AM and its receptor mRNAs expression were determined by reverse transcription-polymerase chain reaction (RT-PCR) and in situ hybridization. By RT-PCR, AM mRNA was increased by 167% at three days, while AM receptor mRNA was increased by 234% at seven days (both P < 0.05). By in situ hybridization, AM and AM receptor mRNAs were increased at ulcer margin from three days after ulcer induction. By immunohistochemistry, AM was increased in the ulcer margin at three and seven days. In separate in vitro studies using a rat gastric epithelial (RGM1) cell line, AM treatment significantly increased transforming growth factor-alpha mRNA expression and cell proliferation.

Expression of endothelin-1, and endothelin A and B receptors in portal hypertensive esophagus of rats.

Nitric oxide synthase is overexpressed in the portal hypertensive (PHT) esophagus, suggesting that expression of other vasoactive mediatora could also be affected. Therefore, in the present study we determined the expression of endothelin-1 (ET-1) and endothelin receptors, which could contribute to the regulation of the vascular tone in PHT esophagus. In esophageal specimens of PHT and sham operated rats, expression of ET-1 and its receptors A and B (ET(A)R and ET(B)R) mRNAs was studied by reverse transcription-polymerase chain reactions. ET-1 protein expression was assessed by immunostaining and enzyme immunoassay. In PHT esophagus, expression of ET-1, ET(A)R and ET(B)R mRNAs was significantly increased by 2.2-, 2.5- and 1.5-fold, respectively, compared with sham operated. The ET-1 protein was significantly increased by 2.2-fold vs. controls as measured by enzyme immunoassay. ET-1 protein was predominantly localized to endothelia of submucosal veins. Thus, portal hypertension induces over-expression of ET-1 in endothelia of esophageal submucosal vessels. Since ET-1 and its receptors could promote vascular proliferation and induce mucosal damage, the overexpressed ET-1 may play an important role in the development and rupture of esophageal varices in portal hypertension.

Isolation of morphologically and functionally intact gastric mucosal microvessels rapid communication.

Gastric mucosal microvessels were isolated after arterial perfusion of the rat stomach with magnetized iron oxide suspension. After homogenization of scrapped gastric mucosa, microvessels were initially separated with a high power magnet and further separated and purified by using a nylon sieve. Aliquots of purified microvessels were assessed for viability, histologic appearance, ultrastructure and generation of prostacyclin. Microvessels were plated on Matrigel and cultured in DMEM with high glucose and 10% FBS for 1, 3 or 5 days. After 1, 3 and 5 days of culturing, endothelial viability was assessed with Fast green exclusion, and the basal and stimulated (with calcium ionophore) generation of prostacyclin was determined by assaying aliquots of the incubating medium for 6-keto PGF(1alpha). At 1 and 3 hrs after isolation, microvessels demonstrated intact morphologic structures as reflected by transmission EM and 92+/-4% of viable endothelial cells. The microvessels plated on Matrigel maintained good viability for at least 5 days and generated prostacyclin at the baseline and following ionophore stimulation. These data demonstrate that isolated microvessels cultured under optimal conditions are fully viable and functional.

ATP-induced CA2+-signaling enhances rat gastric microvascular endothelial cell migration.

The effects of exogenous ATP on Ca2+ signaling and wound healing were investigated in rat gastric microvascular endothelial cells (RGMEC). ATP (10 microM) triggered a significant rise in intracellular Ca2+ concentration ([Ca2+]i) from 46+/-2 nM at baseline to peak values averaging 283+/-31 nM (n = 5 experiments, 132 cells). Return to the basal [Ca2+]i was delayed by slowly declining plateau phase that persisted for 200+/-30 s. Removal of extracellular Ca2+ did not significantly affect the peak rise in [Ca2+]i, but reduced the plateau. ATP (10 microM) also significantly increased the migration of RGMEC in a wounded monolayer. Addition of the non-subtype selective purinergic receptor antagonist, suramin, abrogated the effects of ATP on [Ca2+]i and migration. We conclude that local elevation of ATP acting through purinergic receptors induce Ca2+ signals in RGMEC and may contribute to endothelial cell migration.

Isolation and characterization of rat gastric microvascular endothelial cells as a model for studying gastric angiogenesis in vitro.

We have previously characterized morphologic features of wounding-induced angiogenesis that occurs in response to acute and chronic gastric mucosal injury. As a means of investigating the molecular mechanisms underlying gastric angiogenesis, microvascular endothelial cells were isolated from stomachs of normal (non-injured) rats. The isolation procedure adapted and combined aspects of previous methods and employed positive selection using magnetic beads coated with monoclonal antibody specific for rat CD31 (PECAM-1), a cell surface marker restricted to platelets, monocytes, T lymphocytes and endothelial cells. The isolated microvascular endothelial cells expressed vascular endothelium-specific antigen and the endothelial-specific receptors, Tie2 and flt-1 (VEGFR1). When plated on growth factor-reduced matrigel, the isolated microvascular endothelial cells formed capillary-like structures reflecting in vitro angiogenesis. These cells were also responsive to vascular endothelial growth factor, VEGF, further verifying their endothelial nature. The rat microvascular endothelial cells isolated by this procedure should be useful in delineating molecular mechanisms and regulation of the angiogenesis that is essential for the healing of acute and chronic gastric injury.

Inhibition of angiogenesis by nonsteroidal anti-inflammatory drugs: insight into mechanisms and implications for cancer growth and ulcer healing.

Angiogenesis, the formation of new capillary blood vessels, is essential not only for the growth and metastasis of solid tumors, but also for wound and ulcer healing, because without the restoration of blood flow, oxygen and nutrients cannot be delivered to the healing site. Nonsteroidal anti-inflammatory drugs (NSAIDs) such as aspirin, indomethacin and ibuprofen are the most widely used drugs for pain, arthritis, cardiovascular diseases and, more recently, the prevention of colon cancer and Alzheimer disease. However, NSAIDs produce gastroduodenal ulcers in about 25% of users (often with bleeding and/or perforations) and delay ulcer healing, presumably by blocking prostaglandin synthesis from cyclooxygenase (COX)-1 and COX-2 (ref. 10). The hypothesis that the gastrointestinal side effects of NSAIDs result from inhibition of COX-1, but not COX-2 (ref. 11), prompted the development of NSAIDs that selectively inhibit only COX-2 (such as celecoxib and rofecoxib). Our study demonstrates that both selective and nonselective NSAIDs inhibit angiogenesis through direct effects on endothelial cells. We also show that this action involves inhibition of mitogen-activated protein (MAP) kinase (ERK2) activity, interference with ERK nuclear translocation, is independent of protein kinase C and has prostaglandin-dependent and prostaglandin-independent components. Finally, we show that both COX-1 and COX-2 are important for the regulation of angiogenesis. These findings challenge the premise that selective COX-2 inhibitors will not affect the gastrointestinal tract and ulcer/wound healing.

Basic fibroblast growth factor stimulates repair of wounded hepatocyte monolayer: modulatory role of protein kinase A and extracellular matrix.

The two important repair mechanisms after hepatocyte injury are proliferation and migration of the nearby healthy hepatocytes. Although previous studies have shown that basic fibroblast growth factor (bFGF) levels are markedly elevated after liver injury, the role of bFGF in the repair of the wounded hepatocytes is not well understood. The aim of this study was to delineate the role of bFGF in the repair of the wounded hepatocyte monolayers. Specifically, we examined the role of bFGF in cellular proliferation and migration of hepatocytes with an in vitro wound model. Standardized excisional wounds were created in clone 9 rat hepatocyte monolayers by a razor blade, and the extent of epithelial proliferation and migration was measured. After wound formation, bFGF (30 ng/mL) significantly stimulated proliferation of hepatocytes at the wound margin. bFGF also stimulated the migration of hepatocytes at the wound front. bFGF stimulation of hepatocyte migration correlated with increased formation of actin stress fibers and bFGF-receptor protein level. The bFGF stimulation of hepatocyte migration was abolished by various protein kinase A activating agents including 3-isobutyl-1-methylxanthine, 8-bromoadenosine-3', 5'-cyclic monophosphate, forskolin, and cholera toxin. In addition, protein kinase A activating agents almost completely prevented bFGF-induced actin stress fiber formation in the cells at the wound front. Varying the basement membrane composition of the extracellular matrix had a selective enhancing effect on the basal rates of hepatocyte migration (collagen IV > or = laminin > collagen I > fibronectin > control (plastic)). bFGF treatment resulted in a similar additive increase in hepatocyte migration across all coated surfaces studied. We conclude that bFGF promotes hepatocyte wound repair by stimulating both proliferation and migration of the hepatocyte at the margin of the wound.

Ethanol injury triggers activation of adrenomedullin and its receptor genes in gastric mucosa.

Adrenomedullin (AM) is a potent vasodilatory peptide, which is present in the stomach. However, its precise function in the gastric mucosa is unknown. The expression and localization of AM and its receptor in gastric mucosa injured by ethanol also have not been explored, forming the basis for this study. Gastric samples of rats were obtained at 0 and 8 hr and 1, 2, and 4 days after intragastric administration of 100% ethanol. By RT-PCR, AM mRNA expression in gastric mucosa at 8 and 24 hr following ethanol injury was increased by 2-fold and by 2.5-fold (both P<0.01), respectively, and returned to normal at two days. AM receptor mRNA expression was increased by 2.7-fold, 2.3-fold, and 2.4-fold at 8, 24, and 48 hr, respectively (all P<0.01), and returned to normal at four days. By in situ hybridization, AM and AM receptor mRNAs were present in normal gastric mucosa and up-regulated in gastric mucosa following ethanol injury. The immunohistochemical signal for AM was significantly increased in the mucosa bordering erosion sites. We conclude that ethanol injury up-regulates the expression of both AM and AM receptor in gastric mucosa.

Reduced adrenomedullin expression in gastric mucosa of portal hypertensive rats after ethanol-induced injury.

OBJECTIVE: To determine the expression and localization of adrenomedullin (AM) and its receptor (AM-R) in portal hypertensive (PHT) gastric mucosa after intragastric ethanol administration. SUMMARY BACKGROUND DATA: The repair of gastric mucosal injury requires reestablishment of the microvascular network. The authors previously demonstrated impaired angiogenesis of PHT gastric mucosa after ethanol-induced injury. Because AM, a potent vasodilatory peptide, is also a novel growth and angiogenic factor, the authors hypothesized that AM is involved in the impaired repair of PHT gastric mucosa and its microvasculature after damage. METHODS: Either PHT or sham-operated rats received intragastrically 100% ethanol, and the stomachs were excised at 1, 6, and 24 hours later. Expression and localization of AM and AM-R mRNA were examined by competitive reverse transcription-polymerase chain reaction and in situ hybridization. AM protein expression was examined by Western blot analysis. RESULTS: One hour after ethanol administration, AM mRNA expression in PHT gastric mucosa was significantly decreased by 81%, especially in the superficial mucosa, compared with the gastric mucosa in sham-operated rats. The significant decrease lasted for 24 hours. AM protein expression was significantly decreased by 43% compared with the sham-operated gastric mucosa. Although AM-R mRNA expression in both groups was significantly increased 1 hour after ethanol administration and lasted for 24 hours compared with baseline, there were no differences between the two groups. CONCLUSIONS: The expression of AM in PHT gastric mucosa after ethanol-induced injury is significantly decreased compared with controls. This finding could explain one mechanism for the impaired angiogenesis after injury of PHT gastric mucosa.

Activation of VEGF and Ras genes in gastric mucosa during angiogenic response to ethanol injury.

Our previous studies demonstrated that ethanol injury triggers the angiogenic response in gastric mucosa bordering necrosis. The present study was aimed to determine whether vascular endothelial growth factor (VEGF) (a potent angiogenic peptide selectively acting on endothelial cells) and Ras (a mediator of cell proliferation and a putative regulator of VEGF expression) are involved in gastric angiogenesis after ethanol injury. We studied the angiogenic response and expression of VEGF and Ras in gastric mucosa after ethanol injury. Ethanol damage triggered 1) angiogenesis in the gastric mucosa bordering necrosis, 2) significant increases in VEGF mRNA and protein expression, and 3) significant increases in the expression of Ki-ras mRNA and Ras proteins. Neutralizing anti-VEGF antibody significantly reduced (by greater than threefold) the angiogenic response to ethanol-induced injury. Moreover, mevastatin, an inhibitor of Ras activation, completely blocked the induction of VEGF expression in cultured primary endothelial cells. Because, in other tissues, VEGF is one of the most potent angiogenic factors and VEGF expression is dependent on Ras, our data indicate that Ras and VEGF are involved in gastric mucosal angiogenesis after ethanol injury.

Induction of in vitro angiogenesis in the endothelial-derived cell line, EA hy926, by ethanol is mediated through PKC and MAPK.

We have previously shown that ethanol-induced injury to the gastric mucosa triggers increased expression of the angiogenic factors, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) and angiogenesis. To further investigate ethanol-induced angiogenesis, we used an in vitro angiogenesis model which employs the ability of an endothelial-derived cell line (EA hy926) to form tubelike structures resembling capillaries when plated on the matrix material, Matrigel. We report that serum-starved EA hy926 cells, incubated for as little as 5 minutes with ethanol concentrations of 1.0-2.5%, formed tubelike structures reflecting in vitro angiogenesis. Control cells, not incubated with ethanol, did not form tubelike structures. Incubation for 5 minutes with 2.5% ethanol resulted in increased activities of PKC and MAP kinase (ERK2) by 1.6-fold (p < 0.05) and 2.3-fold (P < 0.001), respectively. Furthermore, inhibitors of the MAPK kinase, MEK (PD98059) and PKC (GF 109203X) prevented the induction of in vitro angiogenesis by ethanol.

Expression of adrenomedullin in portal hypertensive gastric mucosa of rats.

We examined the expression and localization of adrenomedullin (AM) mRNA, AM receptor (AM-R) mRNA and AM protein in normal and portal hypertensive (PHT) rat gastric mucosa. Methods included competitive reverse transcription-polymerase chain reaction (RT/PCR), in situ hybridization and Western blot analysis. Both AM mRNA and AM-R mRNA were strongly expressed not only in normal rat gastric mucosa but also PHT rat gastric mucosa. While total AM and AM-R mRNA abundance was similar in both normal and PHT stomachs, in the superficial mucosa-where major defense mechanisms are located-their expression was significantly reduced in PHT rats by 33% (AM mRNA) and 25% (AM-R mRNA) (both p .0.05). AM protein was also expressed similarly in both normal and PHT gastric mucosa. We conclude that AM and its receptor play a role in regulation of gastric mucosal microcirculation and thus gastric mucosal defense.

Partial portacaval shunt for variceal hemorrhage: longitudinal analysis of effectiveness.

OBJECTIVE: To determine rates of survival, long-term patency, and recurrent variceal hemorrhage among patients with alcoholic cirrhosis treated by partial portacaval shunt. DESIGN: Single-institution cohort follow-up study of 72 consecutive patients who underwent small-diameter portacaval H-graft shunt with collateral ablation during a 10-year period (1981 through 1990). Subjects were enrolled and followed up for up to 15 years. Shunt patency was assessed by portography and/or ultrasonography. We performed 7-year Kaplan-Meier analyses of survival (in 65 patients in Child classes A and B), shunt patency, and absence of variceal bleeding. SETTING: Tertiary academic referral center of the US Department of Veterans Affairs. PATIENTS: Patients with alcoholic cirrhosis were considered for operation after at least 1 proven episode of variceal hemorrhage. Patients with portal vein thrombosis were excluded; patients in Child class C underwent operation only for compelling indications. Of the 72 who underwent partial shunting, 38 were in Child class A, 27 were in class B, and 7 were in class C. INTERVENTIONS: Partial portacaval shunt (6-, 8- or 10-mm polytetrafluoroethylene H-graft with collateral ablation) and serial follow-up. MAIN OUTCOME MEASURES: Study end points were death, recurrent variceal hemorrhage, and unavailability for follow-up. Other measures included graft patency and nonvariceal rebleeding. RESULTS: Cumulative probability of 7-year patency for grafts at risk was 95%. The 7-year probability for absence of variceal bleeding in patients at risk was 92%. In 65 patients in Child classes A and B, operative mortality was 7.7% and the cumulative probability of 7-year survival was 54%. CONCLUSION: For variceal bleeding associated with alcoholic cirrhosis, the small-diameter polytetrafluoroethylene portacaval H-graft with collateral ablation affords durable patency and protection against variceal rebleeding.

Induction of mitogen-activated protein kinase signal transduction pathway during gastric ulcer healing in rats.

BACKGROUND & AIMS: Previous studies have shown that gastric ulceration stimulates epithelial cell proliferation and overexpression of epidermal growth factor (EGF) and EGF receptor (EGF-R) in the mucosa bordering necrosis. The aim of this study was to investigate whether extracellular signal-regulated kinase (ERK) cascade is involved in the healing of experimental gastric ulcers. METHODS: We studied EGF-R levels, EGF-R phosphorylation levels, and ERK1 and ERK2 activity in normal and ulcerated rat gastric mucosa. We also examined the effect of Tyrphostin A46 (potent inhibitor of EGF-R and EGF-R kinase-dependent proliferation) on the above parameters. RESULTS: During the initial stages of healing (3 and 7 days), ulcerated mucosa showed significant increase (vs. controls) in protein tyrosine kinase activity, EGF-R levels (510% and 550%), EGF-R phosphorylation levels, ERK1 activity (430% and 880%), and ERK2 activity (550% and 990%). Tyrphostin A46 treatment significantly inhibited ulcer healing and reduced EGF-R levels, EGF-R phosphorylation, and ERK1 and ERK2 activity. CONCLUSIONS: These findings indicate that experimental gastric ulcer healing involves activation of EGF-R-ERK signal transduction pathway.