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Pepsinogen C gene product is a possible growth factor during gastric mucosal healing
A888 AGA ABSTRACTS
• G3639 EXPRESSION AND LOCALIZATION OF ERBB4 AND ITS LIGAND, NEUREGULIN, IN HUMAN GASTROINTESTINAL MUCOSA. H.Kataoka TJoh, K.Tsuchida, K.Seno, Y.Yokoyama1, T.Kato2, M.ItohL llst Dept. Int. Med. and 2Dept. Bioregulation Res., Nagoya City University Medical School, Nagoya, Japan. It is well known that epidermal growth factor (EGF) and heparin-binding EGF-like growth factor (HB-EGF) bind to EGF receptor (ErB1) and regulate mucosal cell growth in human gastrointestinal (GI) mucosa. ErbB4, a relatively new member of ErbB family, is directly activated by neuregulin (NRG). ErbB4 is also activated by EGF through transphosphorylation of ErbB1-ErbB4 heterodimer complex. Furthermore, HB-EGF, betacellulin, and epiregulin were recently shown to directly bind and activate ErbB4. These things suggest that ErbB4 may play a significant role in GI mucosa. However, expression and localization of NRG and ErbB4 have not been clearly defined in human GI mucosa, forming the basis for this study. Methods.; 1. NRG and ErbB4 protein expression was examined by Western-blotting using cell membranes prepared from surgical specimens. 2. Localization was investigated by immunohistochemistry using three kinds of tissue sections; formalin, acetone, and frozen. Immunoreactivity was confirmed by competition assays. 3. Expression of mRNA was quantified by combination of multi-target RT-PCR and laser-induced fluorescence linked capillary-gel electrophoresis (LIF-CGE). Total RNA was extracted from 40 endoscopic biopsy specimens and multi-target RT-PCR was performed including amplifying GAPDH as an internal standard in the same tube. LIF-CGE allowed for high precision detection of RT-PCR-amplified multi-target cDNA. Results: 1. NRG appeared as a protein band of 60 kDa, corresponding to the transmembrane precursor form, and was detected in all histologically normal esophageal, gastric and duodenal mucosa. 2. ErbB4 was detected as a protein band of 180 kDa. Immuno-staining of ErbB4 was detected in esophageal squamous cells, and gastric surface epithelial cells, but not in gastric fundic and pyloric glandular cells. ErbB4 was also found in duodenal absorptive cells, but not in goblet cells and Brnnner's glandular cells. 3. NRG and ErbB4 mRNA expression was detected in all samples, and was significantly increased in duodenum (p<0.05). 4. In gastric cancer samples, although the increase in expression of NRG mRNA was not significant, mRNA for ErbB4 was significantly overexpressed (p<0.05). Strong expression of the ErbB4 protein in gastric cancer tissue and cells was also detected by Western-blotting and immunohistochemistry, respectively. Conclusions: These results suggest that ErbB4 may play an important role in normal esophageal squamous epithelium, gastric epithelium in surface area, and duodenal absorptive cells. ErbB4 may also be important for malignant cell growth of the stomach. • G3640 PORTOSYSTEMIC SHUNTING IN CHILDREN WITH PORTAL HYPERTENSION: ITS ROLE IN THE ERA OF ENDOSCOPIC THERAPY. T.Kato, R.Romero, R.Koutouby, J.Thompson, C.L.Schleien, A.G.Tzakis Division of Transplantation and Department of Pediatrics, University of Miami School of Medicine, Miami, Florida. Objective Description of recent experiences with portosystemic shunting (PS) in children. Background The use of endoscopic therapy (ET) has dramatically decreased the role of surgery in the management of portal hypertension in children. Reports suggest that surgical treatments are associated with more complications and, therefore, should be avoided. However, ET normally requires multiple follow-up procedures and may not be always beneficial to children. Patients and Methods 12 children (age 1 to 16 year-old, mean age of 7.0 yearold) underwent surgical PS at the University of Miami between October 1994 and October 1997. The etiologies of portal hypertension were: extrahepatic portal vein obstruction (n=6), portal vein thrombosis after orthotopic liver transplantation (OLT) (n=l), congenital hepatic fibrosis (n=2), methotrexate toxicity (n=l), and hepatic cirrhosis (n=2). All but one had history of GI bleeding. In one patient, the procedure was indicated for hypersplenism and chronic abdominal pain from splenomegaly. None of the patients were immediate candidates for OLT as they had normal or good synthetic reserve in their liver function. Types of shunt included: distal splenorenal shunt (n=10), portocaval shunt (n=l) and other (n=l). Follow-up was ranged from 2 to 26 months (median 12 months). Results All patients are currently alive and well with patent shunts. Mean hospital stay for the procedure was 8 days. Early postoperative complications included line related infection in two children. Two patients required readmission, for small bowel obstruction in one and shunt stenosis in another. The patient with shunt stenosis required reconstruction. There has been no episode of rebleeding. Mild portosystemic encephalopathy was seen in one child controlled with lactulose and antibiotics. A trend toward improvement in growth parameters has seen in the youngest patients with the longest follow-up periods. Conclusions Children with good liver synthetic function and portal hypertension tolerated surgery well, have short hospital stays, and have low rate of postoperative complications. Patients are not subjected to repetitive procedures and have rapid and effective control of recurrent bleeding. Improvement in growth parameters as a result of PS may be seen.
GASTROENTEROLOGY Vol. 114, No. 4 G3641 THE EFFECT OF GLUCOCORTICOID ON RAT INTESTINAL ENZYME ACTIVITIES DURING EARLY POSTNATAL PERIOD. T. H. Kim, D. K. Chang, C. H. Lee, H. C. Jung, L S. Song, C, Y. Kim. Department of Internal Medicine and Liver Research Institute, Seoul National University College of Medicine, Seoul, Korea. Backgrounds/Aims: In rats, the high intestinal lactase activity that is found during the first 2 week after birth decreases abruptly late in the third week. Concomitantly, sucrase activity appears and increases. The role of glucocorticoid hormone and thyroid hormone on the regulation of rat intestinal disaccharidases in early postnatal period has been studied. We performed this study to examine the effect of dexamethasone(DEX) and propylthiouracil(PTU) on the developmental changes of rat intestinal disaccharidases activities. Methods : Sprugue-Dawley rats were divided into 4 groups, control (0.9%NaCI 0.01ml/g body wt/d), DEX, PTU, and DEX+PTU groups. Dexamethasone(0.4ug/g body wt/d, day 8-day 22) was injected intraperitonally once a day. Propylthiouracil(0.01g/100ml) was administered to infant rats via drinking water of the dam(beginnig gestational day 15). Rats were sacrificed on days 9, 12, 15, 18, and 23. Lactase and sucrase activites were measured in the proximal and distal part of small intestine. Results: Dexamethasone precociously induced sucrase activity from day 12 and enhanced activity thereafter until day 23 in both proximal and distal small intestine in DEX group compared with control group. In PTU group, PTU blocked normal increase of sucrase activities until day 18. In DEX plus PTU group, sucrase activities showed similar pattern to DEX group. For the lactase activity, DEX did not elicit precocious maturation in proximal small intestine. On the contrary, DEX increased lactase activities in the distal small intestine on days 12 and days 18. Conclusions: 1)Dexamethasone induce precocious maturation of sucrase activity during early postnatal period, but it does not elicit precocious decline of lactase activity. 2)These effects of dexamethasone in the maturation of rat intestinal enzymes seem to be independent of thyroid hormone effect. G3642
PEPSINOGEN C GENE PRODUCT IS A POSSIBLE G R O W T H FACTOR DURING GASTRIC MUCOSAL HEALING. K.Kishi, Y.Kinoshita, Y.Matsushima, A.Okada, T.Maekawa, C.Kawanami, N.Watanabe, T.Chiba. Die. Gastroenterology & Hepatology, Dept. Medicine, Postgraduate Sch. Medicine, Kyoto Univ., Kyoto 606, & 2nd Dept. Medicine, Shimane Med. Univ., Shimane 693, Japan. Pepsinogens (PG's) are mainly synthesized and secreted by the gastric mucnsal cells. Although PG's play important roles in nutrient digestion in the stomach, whether they have other physiological roles is unknown. Recently, the serum PG concentrations of patients with Helicobacter pylori-induced gastritis were found to be elevated. Whether this elevation is due to leakage from damaged cells or enhancement of PG synthesis by mucosal cells of the inflamed tissues is unclear, but this finding indicates that PG's may play a role in inflammatory processes in the gastric mucosa. In this study, as part of our search for new growth factor genes in the stomach, we cloned a PGC gene fragment, cDNAs were obtained from the rat gastric wall with acetic acid-induced ulcers by subtracting those from the non-ulcerated wall and transfected into COS-7 cells. We selected the transfected COS-7 cells with conditioned media showing potent growth stimulating effects on the normal rat gastric mucosal cell line RGM1 and successfully isolated several eDNA clones. DNA sequencing of these clones revealed that the DNA sequence of one was identical to that of a part of the rat PGC gene (between the 968th and 1179th base pairs). Furthermore, in another experiment, we selected the directly transfected RGM1 cells that grew in the low-serum medium containing G418 and isolated several eDNA clones, one of which encodes the same part of the PGC gene obtained above. In order to ensure mRNA expression was augmented in gastric ulcer tissue, Northern blot analysis was performed using the cloned PGC eDNA. PGC gene expression was enhanced not only in acetic acid-induced gastric ulcers but also in ind0methacininduced mucosal lesions. It was also increased in the Helicobacterfelis-infected stomachs. These data indicate that PGC gene expression is enhanced in damaged gastric tissue and this gene may play a role in gastric epithelial cell growth during gastric mucosal healing. G3643 TOTAL ANTIOXIDANT CAPACITY OF HUMAN COLON. TR Koch, L Yuan, West Virginia University, Morgantown, WV; GL Telford, Medical College of Wisconsin; SJ Stryker, Northwestern University Medical Center; E Abdel-Rahman, EC Opara, Duke University Med Center, Durham, NC. Previous studies have shown that Crnhn's disease responds to nutritional therapy, although the mechanism of this improvement is not clear. In a rodent model, acute depletion of the antioxidant, glutathione, induces marked enlargement of intestinal lymphoid follicles. We recently described reduced glutathione levels in the mucosal-submucosal (MS) and muscularis exterua (ME) layers from ulcerative colitis (UC) and Crohn's colitis (CC). Since it is not clear whether glutathione depletion in UC and CC is related to decreased bioavailability of antioxidants or to their increased utilization, this study was designed to examine the total antioxidant capacity of human colon and
• G3639 EXPRESSION AND LOCALIZATION OF ERBB4 AND ITS LIGAND, NEUREGULIN, IN HUMAN GASTROINTESTINAL MUCOSA. H.Kataoka TJoh, K.Tsuchida, K.Seno, Y.Yokoyama1, T.Kato2, M.ItohL llst Dept. Int. Med. and 2Dept. Bioregulation Res., Nagoya City University Medical School, Nagoya, Japan. It is well known that epidermal growth factor (EGF) and heparin-binding EGF-like growth factor (HB-EGF) bind to EGF receptor (ErB1) and regulate mucosal cell growth in human gastrointestinal (GI) mucosa. ErbB4, a relatively new member of ErbB family, is directly activated by neuregulin (NRG). ErbB4 is also activated by EGF through transphosphorylation of ErbB1-ErbB4 heterodimer complex. Furthermore, HB-EGF, betacellulin, and epiregulin were recently shown to directly bind and activate ErbB4. These things suggest that ErbB4 may play a significant role in GI mucosa. However, expression and localization of NRG and ErbB4 have not been clearly defined in human GI mucosa, forming the basis for this study. Methods.; 1. NRG and ErbB4 protein expression was examined by Western-blotting using cell membranes prepared from surgical specimens. 2. Localization was investigated by immunohistochemistry using three kinds of tissue sections; formalin, acetone, and frozen. Immunoreactivity was confirmed by competition assays. 3. Expression of mRNA was quantified by combination of multi-target RT-PCR and laser-induced fluorescence linked capillary-gel electrophoresis (LIF-CGE). Total RNA was extracted from 40 endoscopic biopsy specimens and multi-target RT-PCR was performed including amplifying GAPDH as an internal standard in the same tube. LIF-CGE allowed for high precision detection of RT-PCR-amplified multi-target cDNA. Results: 1. NRG appeared as a protein band of 60 kDa, corresponding to the transmembrane precursor form, and was detected in all histologically normal esophageal, gastric and duodenal mucosa. 2. ErbB4 was detected as a protein band of 180 kDa. Immuno-staining of ErbB4 was detected in esophageal squamous cells, and gastric surface epithelial cells, but not in gastric fundic and pyloric glandular cells. ErbB4 was also found in duodenal absorptive cells, but not in goblet cells and Brnnner's glandular cells. 3. NRG and ErbB4 mRNA expression was detected in all samples, and was significantly increased in duodenum (p<0.05). 4. In gastric cancer samples, although the increase in expression of NRG mRNA was not significant, mRNA for ErbB4 was significantly overexpressed (p<0.05). Strong expression of the ErbB4 protein in gastric cancer tissue and cells was also detected by Western-blotting and immunohistochemistry, respectively. Conclusions: These results suggest that ErbB4 may play an important role in normal esophageal squamous epithelium, gastric epithelium in surface area, and duodenal absorptive cells. ErbB4 may also be important for malignant cell growth of the stomach. • G3640 PORTOSYSTEMIC SHUNTING IN CHILDREN WITH PORTAL HYPERTENSION: ITS ROLE IN THE ERA OF ENDOSCOPIC THERAPY. T.Kato, R.Romero, R.Koutouby, J.Thompson, C.L.Schleien, A.G.Tzakis Division of Transplantation and Department of Pediatrics, University of Miami School of Medicine, Miami, Florida. Objective Description of recent experiences with portosystemic shunting (PS) in children. Background The use of endoscopic therapy (ET) has dramatically decreased the role of surgery in the management of portal hypertension in children. Reports suggest that surgical treatments are associated with more complications and, therefore, should be avoided. However, ET normally requires multiple follow-up procedures and may not be always beneficial to children. Patients and Methods 12 children (age 1 to 16 year-old, mean age of 7.0 yearold) underwent surgical PS at the University of Miami between October 1994 and October 1997. The etiologies of portal hypertension were: extrahepatic portal vein obstruction (n=6), portal vein thrombosis after orthotopic liver transplantation (OLT) (n=l), congenital hepatic fibrosis (n=2), methotrexate toxicity (n=l), and hepatic cirrhosis (n=2). All but one had history of GI bleeding. In one patient, the procedure was indicated for hypersplenism and chronic abdominal pain from splenomegaly. None of the patients were immediate candidates for OLT as they had normal or good synthetic reserve in their liver function. Types of shunt included: distal splenorenal shunt (n=10), portocaval shunt (n=l) and other (n=l). Follow-up was ranged from 2 to 26 months (median 12 months). Results All patients are currently alive and well with patent shunts. Mean hospital stay for the procedure was 8 days. Early postoperative complications included line related infection in two children. Two patients required readmission, for small bowel obstruction in one and shunt stenosis in another. The patient with shunt stenosis required reconstruction. There has been no episode of rebleeding. Mild portosystemic encephalopathy was seen in one child controlled with lactulose and antibiotics. A trend toward improvement in growth parameters has seen in the youngest patients with the longest follow-up periods. Conclusions Children with good liver synthetic function and portal hypertension tolerated surgery well, have short hospital stays, and have low rate of postoperative complications. Patients are not subjected to repetitive procedures and have rapid and effective control of recurrent bleeding. Improvement in growth parameters as a result of PS may be seen.
GASTROENTEROLOGY Vol. 114, No. 4 G3641 THE EFFECT OF GLUCOCORTICOID ON RAT INTESTINAL ENZYME ACTIVITIES DURING EARLY POSTNATAL PERIOD. T. H. Kim, D. K. Chang, C. H. Lee, H. C. Jung, L S. Song, C, Y. Kim. Department of Internal Medicine and Liver Research Institute, Seoul National University College of Medicine, Seoul, Korea. Backgrounds/Aims: In rats, the high intestinal lactase activity that is found during the first 2 week after birth decreases abruptly late in the third week. Concomitantly, sucrase activity appears and increases. The role of glucocorticoid hormone and thyroid hormone on the regulation of rat intestinal disaccharidases in early postnatal period has been studied. We performed this study to examine the effect of dexamethasone(DEX) and propylthiouracil(PTU) on the developmental changes of rat intestinal disaccharidases activities. Methods : Sprugue-Dawley rats were divided into 4 groups, control (0.9%NaCI 0.01ml/g body wt/d), DEX, PTU, and DEX+PTU groups. Dexamethasone(0.4ug/g body wt/d, day 8-day 22) was injected intraperitonally once a day. Propylthiouracil(0.01g/100ml) was administered to infant rats via drinking water of the dam(beginnig gestational day 15). Rats were sacrificed on days 9, 12, 15, 18, and 23. Lactase and sucrase activites were measured in the proximal and distal part of small intestine. Results: Dexamethasone precociously induced sucrase activity from day 12 and enhanced activity thereafter until day 23 in both proximal and distal small intestine in DEX group compared with control group. In PTU group, PTU blocked normal increase of sucrase activities until day 18. In DEX plus PTU group, sucrase activities showed similar pattern to DEX group. For the lactase activity, DEX did not elicit precocious maturation in proximal small intestine. On the contrary, DEX increased lactase activities in the distal small intestine on days 12 and days 18. Conclusions: 1)Dexamethasone induce precocious maturation of sucrase activity during early postnatal period, but it does not elicit precocious decline of lactase activity. 2)These effects of dexamethasone in the maturation of rat intestinal enzymes seem to be independent of thyroid hormone effect. G3642
PEPSINOGEN C GENE PRODUCT IS A POSSIBLE G R O W T H FACTOR DURING GASTRIC MUCOSAL HEALING. K.Kishi, Y.Kinoshita, Y.Matsushima, A.Okada, T.Maekawa, C.Kawanami, N.Watanabe, T.Chiba. Die. Gastroenterology & Hepatology, Dept. Medicine, Postgraduate Sch. Medicine, Kyoto Univ., Kyoto 606, & 2nd Dept. Medicine, Shimane Med. Univ., Shimane 693, Japan. Pepsinogens (PG's) are mainly synthesized and secreted by the gastric mucnsal cells. Although PG's play important roles in nutrient digestion in the stomach, whether they have other physiological roles is unknown. Recently, the serum PG concentrations of patients with Helicobacter pylori-induced gastritis were found to be elevated. Whether this elevation is due to leakage from damaged cells or enhancement of PG synthesis by mucosal cells of the inflamed tissues is unclear, but this finding indicates that PG's may play a role in inflammatory processes in the gastric mucosa. In this study, as part of our search for new growth factor genes in the stomach, we cloned a PGC gene fragment, cDNAs were obtained from the rat gastric wall with acetic acid-induced ulcers by subtracting those from the non-ulcerated wall and transfected into COS-7 cells. We selected the transfected COS-7 cells with conditioned media showing potent growth stimulating effects on the normal rat gastric mucosal cell line RGM1 and successfully isolated several eDNA clones. DNA sequencing of these clones revealed that the DNA sequence of one was identical to that of a part of the rat PGC gene (between the 968th and 1179th base pairs). Furthermore, in another experiment, we selected the directly transfected RGM1 cells that grew in the low-serum medium containing G418 and isolated several eDNA clones, one of which encodes the same part of the PGC gene obtained above. In order to ensure mRNA expression was augmented in gastric ulcer tissue, Northern blot analysis was performed using the cloned PGC eDNA. PGC gene expression was enhanced not only in acetic acid-induced gastric ulcers but also in ind0methacininduced mucosal lesions. It was also increased in the Helicobacterfelis-infected stomachs. These data indicate that PGC gene expression is enhanced in damaged gastric tissue and this gene may play a role in gastric epithelial cell growth during gastric mucosal healing. G3643 TOTAL ANTIOXIDANT CAPACITY OF HUMAN COLON. TR Koch, L Yuan, West Virginia University, Morgantown, WV; GL Telford, Medical College of Wisconsin; SJ Stryker, Northwestern University Medical Center; E Abdel-Rahman, EC Opara, Duke University Med Center, Durham, NC. Previous studies have shown that Crnhn's disease responds to nutritional therapy, although the mechanism of this improvement is not clear. In a rodent model, acute depletion of the antioxidant, glutathione, induces marked enlargement of intestinal lymphoid follicles. We recently described reduced glutathione levels in the mucosal-submucosal (MS) and muscularis exterua (ME) layers from ulcerative colitis (UC) and Crohn's colitis (CC). Since it is not clear whether glutathione depletion in UC and CC is related to decreased bioavailability of antioxidants or to their increased utilization, this study was designed to examine the total antioxidant capacity of human colon and