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The augmenter of liver regeneration enhances the mitochondrial oxidative phosphorylation of rat liver
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15 CORTICOSTEROIDS STUnnATE BICARBONATE EXCRETION IN CHOLANGIOCYTES BY EMIANCING THE ACTIVITY AND EXPRESSION OF Ua’lH- AND CI/HCO;EXCHANGERS Alvaro D Maruccl L Gislioza A, Barbaro B , Montembbianesl R, Mmetoia L , Alpm G , Mancmo M Capocacaa L , and Benedem A Dw Gastro tiluv Rome “La Saplenza” and Ancona, and Texas A&M Uoiv Background/Aim Cortlcostermds are currently under evaluation for the treatment of cholan~opathies However, theu effects on the functions of the ~ntrahepatx blliary epithelium are unknown We mvesrigated both m wvo and in vmo the expression of glucocomcold receptors (GcR) m the iat mtrahepaoc blhaty eplthelium and the role of corticosteroids in the regulation of biliary epithelium secretory act&es Methods: The expression of GcR was studied m srtu by Immuno~stochemistry and in isolated bile ducts (IBDU) by immune-bistochemistry and RT-PCR The acute and chronic effects of comcosterotds [two days treatment with dexamethasone (0 3 mg, tld) or hudesomde (0 05 mg lid)] on bile flow, txarbonate biliary excretmn and rhe acfivmes of Na’/‘tl+ CI’/HCOx. exchangers and Na‘ HCO; cotranmorter >n IBDU were evaluated Protem exores~ton of the Na-IH’ exchanger lsoform I (NHEl) and CIDHCOIexchanger m cholaogiocytes was evaluated by western-blot Results: We found that GcR were expressed m rat cholanglocyies and were markedly increased (p< 0 02) after choianglocyte prohferation induced by bde duct obstruction Cholanglocytes expressed GcR at a hrgher level when compared with hepatocytes Acute adrmmstratlon of dexametbasone (5 uM) or budesomde (1 uM) for 40-60 mm faded to miluence bxarbonate b;diruy secretloo m bile Bstula rats and K/HCOa transport processes m IBDU However, treatment for 2 days wth dexamethasone or budesomde induced a significant (p
EPITHELIAL AND LYMPHOCYTE APOPTOSIS IN CROHN’S DISEASE Di Sabatino A, Parroni R, Ciccocioppo R, D’AId S, Perego M, Alvisi C, Villani L. Luinetli 0, Ricevuti L. Tinozzi S. Cifone MG. Cow GR. Gastroenterology Unit, Depts of Medicine, Pathology and Skgely, IRCCS Policlinico S. Matteo, University of Pavia: Dept of Experimental Medicine, University of L’Aquila, Italy. Backmound & Aims: Since apoptosis plays a crucial role in the conbul of intestinal epithelial cell rumover and in tbe regulation of inflmmnawry response, we studied epithelial and lamina prop& mononuclear cell (LPMC) apoptosis in C&n’s disease (CD). Patients & Methods: Endoscopic ileal and coIonic biopsy specimens were collected fmm macroscopically involved areas of I2 CD patients (mean age 34.8 years, range 2&57), and from I2 patients (mean age 37.7 years, range 1868) undergoing endoswpy for investigation of persistenr diarrhoea and imitable bowel syndrome. The clinical severity of disease was assessed by the evaluation of the C&n’s disease activity index (CDAI) (active disease >150). For the “in situ” detection of apoptotic cells, terminal deoxynucleotidyl waasferase-mediated digoxigenindeoxyu&ne triphospbate nick end labelling (TUNEL) was used, by a pemxidase ApopTag Kit (Oncor). Counts were performed in a blind fashion by an emat observer, by a differential count of at least 2000 cells in the ep&elium and in;he Iamine pmpria and the results expressed as the number of positive cells/1000 cells. m: The proportions of TUNEL+ epithelial cells were increased in active CD (35.4 f 9.6%) in comparison 1” quiescent CD (7.8 f 1.1%) and controls (8.2 + O.B%o).No significant difference was found between inactive CD patients and biopsied contmls. In active CD a direct correlation between the proporiion of TUNEL’ eoithelial cells and the CDAI was found. The ~~rotwrtions of TUNEL’ L:PMCs were decreased in active CD (4.4 f 0,3k) in comparison to contmls,(8.5 f 0.4?&),Tbe pmpatioF,of TUNEL’ LPMCs were bigha in quiescent CD patients (13.6 f 1.9%) compared to controls. In active CD ah inverse correlation between the proportions oFTWNEL+ LPMCs and the CDAl was found. Conclusions: Epithelial apoptosis is increased in intestinal infIammed areas of active CD and falls lo nornml level dining remission. 11is likely that this increase is secondary to the higher mobility of cells which cover the areas of epithelial loss to assist regeneration. Decreased LPMC apaptosis in active CD may aggravate end perpetuate the abnormal mucosal inilammatory response in this condition. Increased LPMC apoptosis in quiescent CD could be a homeostatic mechanism able to downregulate the lymphocyte proliferation and activation.
14 THE AUGMENTER OF LIVER REGENERATION ENHANCES THE MITOCHONDRIAL OXIDATIVE PHOSPHORYLATION OF RAT LIVER. Polimeno L, OCapuanoF, Mamngi LC, Margiotta M, Francavilla A. Chair of Gastroenterology and Digestive Endoscopy, Dept of Emergency and Organ Transplantation (D.E.T.O.) and OInstitute of Medical Biochemistry end Chemistry, University of Bari, Bari, Italy. Introduction. The proliferative activity of mammalian Augmenter of Liver Regeneration (ALR) in liver regeneration of rats and dogs has been extensively defined, nevertheless the mechanism through which ALR exeTts its activity is not yet fully understood. One possible suggestion comes from the proven 50% homology that ALR protein shows to the product of the dual function nuclear Saccharomyces Cerevisiae gene ERVI, which is essential for oxidative phosphorylation, growth and life of the yeast. To verify if also ALR could represent an important factor regulating the mitochondrial biogenesis, tbe phosphorilative capacity of liver mitochondria isolated from rats treated with ALR was studied. Animals. F344 male rats, 180-200 g of body weight, were injected i.p. with a single dose of ALRp (500 &rat at 8.00 a.m.) and sacrificed 24 hours thereafter the injection. Control animals were treated with saline and sacrificed at the same time. Experimental. Mitochondria freshly isolated from treated and control rats were assayed for respiratory activity and ATP synthesis. By using three different oxidizable substrates, the oxygen consumption in the state IV and in the state III coupled and uncoupled respiration was measured. The ATP synthase activity was determined in samples of mitochondrial suspensions respiring in the coupled state III. Aliquats of mitochondria were utilised for spectrophotometric determination of cytochromes content. Results. Both in control and treated liver mitochondria respiration with all the three substrates was contmlled bv ADP (rrlus Pi) and further stimulated by the uncoupler addition. These measurements show that both types of mitochondria were well coupled, i.e. respiration was tightly controlled by the protonmotive force Ap. Treated mitochondria exhibit, however, an higher respiratory activity with respect to the control mitochondria in all conditions examined and in particular with succinate as substrate. In these mitochondria, moreover, the increase of respiratory activity was accomoanied bv. the increase of bath the rate of ATP synthesis and . cytocbromes content. Conclusions. These findings suggest that ALR may affect the expression of the gene involved in the &dative phospborylation. This could be part of the complex mechanism that regulates hepatocytes proliferation.
CHEMOKINE EXPRESSION I-N POUCHITIS Rizzello F, Uguccioni M*, Amadini C, Venturi A, Morselli C, Salvatori S, Romboli E, Baggiolini MS and Campieri M. Department of Internal Medicine and Gastroenterology, University of Bologna, Italy; *Theodor Kocher Institute, University of Bern, Switzerland Background & Aim: Chemokines are thought to be important for the recruitment of inflammatory cells to the site of inflammation. We have studied the expression of CC-chemokines Eotaxin, interferon-y inducible protein-10 (IP-IO), monocyte chemoattractant protein-l (MCP-I) and -3 (MCP-3) and the CXC-chemokine interleukin-8 (IL8) in pouchitis - the major long-term complication after pouch surgery for ulcerative colitis - and non-inflamed pouch to assessthe role of these chemokines in the pathogenesis of pouchitis. Materials & Methods: Biopsies of 8 patients with non-inflamed pouch and 10 patients with pouch&, before and after antibiotic treatment, were studied. Biopsies were taken endoscopically, frozen immediately and subsequently stained for IP-10, MCP-I, MCPJ, Eotaxin and IL-8 in serial sections. Chemokine-expressing cells were quantified by image analysis and subsequently statistical analysis was performed. Results: Cells infiltrating the lamina propria expressed the analysed chemokines. The expression of MCP-I, MCP-3 and IL-8 was significantly enhanced in pouchitis versus normal pouch (~0.005, l%O.OS and p
15 CORTICOSTEROIDS STUnnATE BICARBONATE EXCRETION IN CHOLANGIOCYTES BY EMIANCING THE ACTIVITY AND EXPRESSION OF Ua’lH- AND CI/HCO;EXCHANGERS Alvaro D Maruccl L Gislioza A, Barbaro B , Montembbianesl R, Mmetoia L , Alpm G , Mancmo M Capocacaa L , and Benedem A Dw Gastro tiluv Rome “La Saplenza” and Ancona, and Texas A&M Uoiv Background/Aim Cortlcostermds are currently under evaluation for the treatment of cholan~opathies However, theu effects on the functions of the ~ntrahepatx blliary epithelium are unknown We mvesrigated both m wvo and in vmo the expression of glucocomcold receptors (GcR) m the iat mtrahepaoc blhaty eplthelium and the role of corticosteroids in the regulation of biliary epithelium secretory act&es Methods: The expression of GcR was studied m srtu by Immuno~stochemistry and in isolated bile ducts (IBDU) by immune-bistochemistry and RT-PCR The acute and chronic effects of comcosterotds [two days treatment with dexamethasone (0 3 mg, tld) or hudesomde (0 05 mg lid)] on bile flow, txarbonate biliary excretmn and rhe acfivmes of Na’/‘tl+ CI’/HCOx. exchangers and Na‘ HCO; cotranmorter >n IBDU were evaluated Protem exores~ton of the Na-IH’ exchanger lsoform I (NHEl) and CIDHCOIexchanger m cholaogiocytes was evaluated by western-blot Results: We found that GcR were expressed m rat cholanglocyies and were markedly increased (p< 0 02) after choianglocyte prohferation induced by bde duct obstruction Cholanglocytes expressed GcR at a hrgher level when compared with hepatocytes Acute adrmmstratlon of dexametbasone (5 uM) or budesomde (1 uM) for 40-60 mm faded to miluence bxarbonate b;diruy secretloo m bile Bstula rats and K/HCOa transport processes m IBDU However, treatment for 2 days wth dexamethasone or budesomde induced a significant (p
EPITHELIAL AND LYMPHOCYTE APOPTOSIS IN CROHN’S DISEASE Di Sabatino A, Parroni R, Ciccocioppo R, D’AId S, Perego M, Alvisi C, Villani L. Luinetli 0, Ricevuti L. Tinozzi S. Cifone MG. Cow GR. Gastroenterology Unit, Depts of Medicine, Pathology and Skgely, IRCCS Policlinico S. Matteo, University of Pavia: Dept of Experimental Medicine, University of L’Aquila, Italy. Backmound & Aims: Since apoptosis plays a crucial role in the conbul of intestinal epithelial cell rumover and in tbe regulation of inflmmnawry response, we studied epithelial and lamina prop& mononuclear cell (LPMC) apoptosis in C&n’s disease (CD). Patients & Methods: Endoscopic ileal and coIonic biopsy specimens were collected fmm macroscopically involved areas of I2 CD patients (mean age 34.8 years, range 2&57), and from I2 patients (mean age 37.7 years, range 1868) undergoing endoswpy for investigation of persistenr diarrhoea and imitable bowel syndrome. The clinical severity of disease was assessed by the evaluation of the C&n’s disease activity index (CDAI) (active disease >150). For the “in situ” detection of apoptotic cells, terminal deoxynucleotidyl waasferase-mediated digoxigenindeoxyu&ne triphospbate nick end labelling (TUNEL) was used, by a pemxidase ApopTag Kit (Oncor). Counts were performed in a blind fashion by an emat observer, by a differential count of at least 2000 cells in the ep&elium and in;he Iamine pmpria and the results expressed as the number of positive cells/1000 cells. m: The proportions of TUNEL+ epithelial cells were increased in active CD (35.4 f 9.6%) in comparison 1” quiescent CD (7.8 f 1.1%) and controls (8.2 + O.B%o).No significant difference was found between inactive CD patients and biopsied contmls. In active CD a direct correlation between the proporiion of TUNEL’ eoithelial cells and the CDAI was found. The ~~rotwrtions of TUNEL’ L:PMCs were decreased in active CD (4.4 f 0,3k) in comparison to contmls,(8.5 f 0.4?&),Tbe pmpatioF,of TUNEL’ LPMCs were bigha in quiescent CD patients (13.6 f 1.9%) compared to controls. In active CD ah inverse correlation between the proportions oFTWNEL+ LPMCs and the CDAl was found. Conclusions: Epithelial apoptosis is increased in intestinal infIammed areas of active CD and falls lo nornml level dining remission. 11is likely that this increase is secondary to the higher mobility of cells which cover the areas of epithelial loss to assist regeneration. Decreased LPMC apaptosis in active CD may aggravate end perpetuate the abnormal mucosal inilammatory response in this condition. Increased LPMC apoptosis in quiescent CD could be a homeostatic mechanism able to downregulate the lymphocyte proliferation and activation.
14 THE AUGMENTER OF LIVER REGENERATION ENHANCES THE MITOCHONDRIAL OXIDATIVE PHOSPHORYLATION OF RAT LIVER. Polimeno L, OCapuanoF, Mamngi LC, Margiotta M, Francavilla A. Chair of Gastroenterology and Digestive Endoscopy, Dept of Emergency and Organ Transplantation (D.E.T.O.) and OInstitute of Medical Biochemistry end Chemistry, University of Bari, Bari, Italy. Introduction. The proliferative activity of mammalian Augmenter of Liver Regeneration (ALR) in liver regeneration of rats and dogs has been extensively defined, nevertheless the mechanism through which ALR exeTts its activity is not yet fully understood. One possible suggestion comes from the proven 50% homology that ALR protein shows to the product of the dual function nuclear Saccharomyces Cerevisiae gene ERVI, which is essential for oxidative phosphorylation, growth and life of the yeast. To verify if also ALR could represent an important factor regulating the mitochondrial biogenesis, tbe phosphorilative capacity of liver mitochondria isolated from rats treated with ALR was studied. Animals. F344 male rats, 180-200 g of body weight, were injected i.p. with a single dose of ALRp (500 &rat at 8.00 a.m.) and sacrificed 24 hours thereafter the injection. Control animals were treated with saline and sacrificed at the same time. Experimental. Mitochondria freshly isolated from treated and control rats were assayed for respiratory activity and ATP synthesis. By using three different oxidizable substrates, the oxygen consumption in the state IV and in the state III coupled and uncoupled respiration was measured. The ATP synthase activity was determined in samples of mitochondrial suspensions respiring in the coupled state III. Aliquats of mitochondria were utilised for spectrophotometric determination of cytochromes content. Results. Both in control and treated liver mitochondria respiration with all the three substrates was contmlled bv ADP (rrlus Pi) and further stimulated by the uncoupler addition. These measurements show that both types of mitochondria were well coupled, i.e. respiration was tightly controlled by the protonmotive force Ap. Treated mitochondria exhibit, however, an higher respiratory activity with respect to the control mitochondria in all conditions examined and in particular with succinate as substrate. In these mitochondria, moreover, the increase of respiratory activity was accomoanied bv. the increase of bath the rate of ATP synthesis and . cytocbromes content. Conclusions. These findings suggest that ALR may affect the expression of the gene involved in the &dative phospborylation. This could be part of the complex mechanism that regulates hepatocytes proliferation.
CHEMOKINE EXPRESSION I-N POUCHITIS Rizzello F, Uguccioni M*, Amadini C, Venturi A, Morselli C, Salvatori S, Romboli E, Baggiolini MS and Campieri M. Department of Internal Medicine and Gastroenterology, University of Bologna, Italy; *Theodor Kocher Institute, University of Bern, Switzerland Background & Aim: Chemokines are thought to be important for the recruitment of inflammatory cells to the site of inflammation. We have studied the expression of CC-chemokines Eotaxin, interferon-y inducible protein-10 (IP-IO), monocyte chemoattractant protein-l (MCP-I) and -3 (MCP-3) and the CXC-chemokine interleukin-8 (IL8) in pouchitis - the major long-term complication after pouch surgery for ulcerative colitis - and non-inflamed pouch to assessthe role of these chemokines in the pathogenesis of pouchitis. Materials & Methods: Biopsies of 8 patients with non-inflamed pouch and 10 patients with pouch&, before and after antibiotic treatment, were studied. Biopsies were taken endoscopically, frozen immediately and subsequently stained for IP-10, MCP-I, MCPJ, Eotaxin and IL-8 in serial sections. Chemokine-expressing cells were quantified by image analysis and subsequently statistical analysis was performed. Results: Cells infiltrating the lamina propria expressed the analysed chemokines. The expression of MCP-I, MCP-3 and IL-8 was significantly enhanced in pouchitis versus normal pouch (~0.005, l%O.OS and p