- Email: [email protected]
763 THE HEPATIC ACUTE PHASE RESPONSE CONTROLS INFLAMMATION BY PROMOTING MYELOID DERIVED SUPPRESSOR CELL FUNCTIONS IN SEPSIS
S280
Poster Session − Saturday, April 25
Conclusions: Indole correlates with HE, has a significant intestinal production and hepatic extraction, increases after TIPS and is related to the modification of psychometric performance. These data suggest that indole may be involved in the pathophysiology of HE. 762 ROLE OF HOMOCYSTEINE IN THE PLATELET FUNCTION OF BILE DUCT LIGATED RATS P. Romecin, J.E. Mill´an, F. Gragnolini, E. Garc´ıa, C. Ortiz, N. Atucha, J. Garc´ıa-Esta˜n. Physiology, Universidad de Murcia, Murcia, Spain E-mail: [email protected] Introduction: Previously, we have found that intracellular Ca2+ homeostasis ([Ca2+ ]i ) is altered in platelets of an experimental model of cholestasis, the bile-duct-ligated (BDL) rat. Different studies indicate that cholestatic diseases are associated with hyperhomocysteinemia and alterations in platelet function, compatible with a platelet activation status. We have hypotethized that a folic acid deficiency in cirrhosis may contribute to those platelet alterations, due to its involvement in the metabolism of homocysteine (Hcy). Objective: The present study investigated the effects of acute Hcy (10 to 100 uM) and chronic folic acid treatment on platelet aggregation and [Ca2+ ]i homeostasis. Methods: Cholestasis was induced in rats by bile duct ligation and shamoperated rats were used as controls. The animals were studied 21 days after the operation. Platelet rich plasma (PRP) aggregation in response to ADP (with or without Hcy) was analyzed using a lumiaggregometer. To analyze the [Ca2+ ] by fluorescence spectroscopy, platelet suspensions were washed and [Ca2+ ]i with fura-2. In one control and one BDL group, folic acid (8 mg/kg) was administered in the drinking water starting 1−2 days before cholestasis induction. Results: Acute treatment with Hcy increased the aggregation response to ADP and [Ca2+ ]i responses to thrombin in BDL and control rats. Capacitative Ca2+ entry, as evaluated by inhibition of SERCA with thapsigargin, was not altered. Chronic treatment with folic acid decreased the aggregation response in control and BDL rats, but this decrease was greater in BDL rats. Folic acid treatment decreased thrombin-induced Ca2+ entry and release in control rats but did not change it in BDL rats; Capacitative Ca2+ entry, however, was decreased in both control and BDL rats; total calcium stored was not altered. Conclusion: Homocysteine plays a role in the enhanced platelet aggregation response of BDL rats, however, the altered Ca2+ signalling in platelets from BDL rats can not be completely explained by increased homocysteine plasma levels. 763 THE HEPATIC ACUTE PHASE RESPONSE CONTROLS INFLAMMATION BY PROMOTING MYELOID DERIVED SUPPRESSOR CELL FUNCTIONS IN SEPSIS L. Sander1,2 , S. Dutton Sackett1 , F. Tacke1 , C. Trautwein1 . 1 Department of Medicine III, RWTH University Hospital, Aachen, Germany; 2 Immunology Institute, Mount Sinai School of Medicine, New York, NY, USA E-mail: [email protected] Sepsis is a major cause of mortality worldwide characterized by a dysregulated inflammatory response to infection. Patients with chronic liver diseases have significantly increased risk of acquiring sepsis and its accompanying complications, and thus have higher sepsis related mortality. The underlying mechanisms are only partially understood. The liver is the major source of APPs which are regarded as important components of the innate immune response. However, given the large diversity of APPs with pro- and anti-inflammatory functions, their overall role in infections is not well defined. Using a murine model for polymicrobial sepsis we show here that hepatocyte-specific deficiency in common interleukin-6 (IL-6) family cytokine receptor gp130 (gp130Dhepa) or gp130/signal transducer and activator of transcription (STAT) 1/3 signaling (gp130StatDhepa) abolished
APP production and enhanced mortality despite normal bacterial clearance and inflammation. Surprisingly, gp130Dhepa mice failed to downmodulate the inflammatory response. This was associated with a lack of myeloid derived suppressor cell (MDSC) accumulation in the spleen, a population of cells known mainly for its immunosuppressive functions in cancers, like hepatocellular carcinoma. MDSCs were critical for inhibiting inflammatory cytokine secretion, and their adoptive transfer protected gp130Dhepa mice from sepsis associated mortality. Transcriptional analysis identified a specific gp130 dependent genetic programm that was initiated in hepatocytes in response to sepsis. APPs and certain chemokines showed a particularly strong gp130 dependent regulation. Administration of the APP serum amyloid A (SAA) and the chemokine KC, both strongly upregulated in sepsis in the livers of control but not gp130Dhepa mice, reversed the detrimental effects of gp130 deficiency and restored mobilization, survival and splenic accumulation of MDSCs. Our results demonstrate that the hepatic acute phase response plays an important negative regulatory role during innate immunity to infection mediated through a novel gp130dependent pathway of mobilizing immunosuppressive MDSCs. 764 ABNORMAL SYSTEMIC BIOMOLECULAR EXPRESSION OF CALCIUM-SENSING RECEPTORS (CARS) IN CIRRHOTIC RATS: A NEW MECHANISM OF DISEASE IN PORTAL HYPERTENSION G. Sansoe’1 , M. Aragno2 , C. Tomasinelli2 , L. Valfre’ di Bonzo2 , F. Rosina1 , A. Smedile3 , M. Rizzetto3 , M. Parola2 . 1 Division of Gastroenterology, Gradenigo Hospital, 2 Dpt. of Experimental Medicine and Oncology, 3 Dpt. of Gastroenterology, University of Torino, Torino, Italy E-mail: [email protected] Extracellular calcium (Ca++ ) may cause vasodilatation through stimulation of membrane-bound Ca++ -sensing receptors (CaRs), which mediate intracellular generation of PGE2 and decreased production of cAMP in vascular smooth muscle cells and several epithelia [1]. On the other hand, in activated hepatic stellate cells (HSCs) from cirrhotic livers L-type voltageoperated Ca++ channels (L-VOCCs) are over-expressed and responsible for Ca++ -mediated HSC contraction leading to increased resistance to portal flow [2]. CaRs expression and immuno-location in liver, kidneys and mesenteric intestine of cirrhotic rats have never been investigated. Aim and Methods: To address this issue in advanced experimental cirrhosis, we evaluated biomolecular expression of CaRs through Western blot analysis in tissue samples of kidney, liver and small intestine of control and cirrhotic rats. Slices of these organs were also submitted to determination of CaRs immuno-location through indirect immunofluorescence after tissue staining with anti-CaRs, anti-Flk-1 (an endothelial antigen), and anti-nucleus (DAPI blue) monoclonal antibodies. 8 control rats and 8 rats with ascitic cirrhosis induced by 12-week CCl4 intoxication were studied. Results: Compared to controls, hepatic, renal and intestinal CaR protein content was uniformly reduced by at least 30% in cirrhotic rats (all P < 0.05). In cirrhotic rat tissue slices, CaRs were found to be strictly immuno-located in sub-endothelial layers of renal medullary arteries and intrahepatic portal venules. Submucosal expression of CaRs in small bowel was restricted to the tip of villi and no significant association of these receptors with blood vessels was seen. Conclusions: Due to their localization in sub-endothelial layers of kidney arteries and intrahepatic portal venules, without any concurrent expression in smooth muscle layers of mesenteric arteries, CaRs should be targeted by specific pharmacological agonists (i.e. Poly-L-Arginine or Poly-LLysine) [3] in order to manage the complications of portal hypertension without aggravating hyperdynamic circulation in liver cirrhosis. References [1] Am J Physiol 283: F963−20, 2002. [2] Hepatology 33: 1007−8, 2001. [3] Am J Physiol 273: C1168−75, 1997.
Poster Session − Saturday, April 25
Conclusions: Indole correlates with HE, has a significant intestinal production and hepatic extraction, increases after TIPS and is related to the modification of psychometric performance. These data suggest that indole may be involved in the pathophysiology of HE. 762 ROLE OF HOMOCYSTEINE IN THE PLATELET FUNCTION OF BILE DUCT LIGATED RATS P. Romecin, J.E. Mill´an, F. Gragnolini, E. Garc´ıa, C. Ortiz, N. Atucha, J. Garc´ıa-Esta˜n. Physiology, Universidad de Murcia, Murcia, Spain E-mail: [email protected] Introduction: Previously, we have found that intracellular Ca2+ homeostasis ([Ca2+ ]i ) is altered in platelets of an experimental model of cholestasis, the bile-duct-ligated (BDL) rat. Different studies indicate that cholestatic diseases are associated with hyperhomocysteinemia and alterations in platelet function, compatible with a platelet activation status. We have hypotethized that a folic acid deficiency in cirrhosis may contribute to those platelet alterations, due to its involvement in the metabolism of homocysteine (Hcy). Objective: The present study investigated the effects of acute Hcy (10 to 100 uM) and chronic folic acid treatment on platelet aggregation and [Ca2+ ]i homeostasis. Methods: Cholestasis was induced in rats by bile duct ligation and shamoperated rats were used as controls. The animals were studied 21 days after the operation. Platelet rich plasma (PRP) aggregation in response to ADP (with or without Hcy) was analyzed using a lumiaggregometer. To analyze the [Ca2+ ] by fluorescence spectroscopy, platelet suspensions were washed and [Ca2+ ]i with fura-2. In one control and one BDL group, folic acid (8 mg/kg) was administered in the drinking water starting 1−2 days before cholestasis induction. Results: Acute treatment with Hcy increased the aggregation response to ADP and [Ca2+ ]i responses to thrombin in BDL and control rats. Capacitative Ca2+ entry, as evaluated by inhibition of SERCA with thapsigargin, was not altered. Chronic treatment with folic acid decreased the aggregation response in control and BDL rats, but this decrease was greater in BDL rats. Folic acid treatment decreased thrombin-induced Ca2+ entry and release in control rats but did not change it in BDL rats; Capacitative Ca2+ entry, however, was decreased in both control and BDL rats; total calcium stored was not altered. Conclusion: Homocysteine plays a role in the enhanced platelet aggregation response of BDL rats, however, the altered Ca2+ signalling in platelets from BDL rats can not be completely explained by increased homocysteine plasma levels. 763 THE HEPATIC ACUTE PHASE RESPONSE CONTROLS INFLAMMATION BY PROMOTING MYELOID DERIVED SUPPRESSOR CELL FUNCTIONS IN SEPSIS L. Sander1,2 , S. Dutton Sackett1 , F. Tacke1 , C. Trautwein1 . 1 Department of Medicine III, RWTH University Hospital, Aachen, Germany; 2 Immunology Institute, Mount Sinai School of Medicine, New York, NY, USA E-mail: [email protected] Sepsis is a major cause of mortality worldwide characterized by a dysregulated inflammatory response to infection. Patients with chronic liver diseases have significantly increased risk of acquiring sepsis and its accompanying complications, and thus have higher sepsis related mortality. The underlying mechanisms are only partially understood. The liver is the major source of APPs which are regarded as important components of the innate immune response. However, given the large diversity of APPs with pro- and anti-inflammatory functions, their overall role in infections is not well defined. Using a murine model for polymicrobial sepsis we show here that hepatocyte-specific deficiency in common interleukin-6 (IL-6) family cytokine receptor gp130 (gp130Dhepa) or gp130/signal transducer and activator of transcription (STAT) 1/3 signaling (gp130StatDhepa) abolished
APP production and enhanced mortality despite normal bacterial clearance and inflammation. Surprisingly, gp130Dhepa mice failed to downmodulate the inflammatory response. This was associated with a lack of myeloid derived suppressor cell (MDSC) accumulation in the spleen, a population of cells known mainly for its immunosuppressive functions in cancers, like hepatocellular carcinoma. MDSCs were critical for inhibiting inflammatory cytokine secretion, and their adoptive transfer protected gp130Dhepa mice from sepsis associated mortality. Transcriptional analysis identified a specific gp130 dependent genetic programm that was initiated in hepatocytes in response to sepsis. APPs and certain chemokines showed a particularly strong gp130 dependent regulation. Administration of the APP serum amyloid A (SAA) and the chemokine KC, both strongly upregulated in sepsis in the livers of control but not gp130Dhepa mice, reversed the detrimental effects of gp130 deficiency and restored mobilization, survival and splenic accumulation of MDSCs. Our results demonstrate that the hepatic acute phase response plays an important negative regulatory role during innate immunity to infection mediated through a novel gp130dependent pathway of mobilizing immunosuppressive MDSCs. 764 ABNORMAL SYSTEMIC BIOMOLECULAR EXPRESSION OF CALCIUM-SENSING RECEPTORS (CARS) IN CIRRHOTIC RATS: A NEW MECHANISM OF DISEASE IN PORTAL HYPERTENSION G. Sansoe’1 , M. Aragno2 , C. Tomasinelli2 , L. Valfre’ di Bonzo2 , F. Rosina1 , A. Smedile3 , M. Rizzetto3 , M. Parola2 . 1 Division of Gastroenterology, Gradenigo Hospital, 2 Dpt. of Experimental Medicine and Oncology, 3 Dpt. of Gastroenterology, University of Torino, Torino, Italy E-mail: [email protected] Extracellular calcium (Ca++ ) may cause vasodilatation through stimulation of membrane-bound Ca++ -sensing receptors (CaRs), which mediate intracellular generation of PGE2 and decreased production of cAMP in vascular smooth muscle cells and several epithelia [1]. On the other hand, in activated hepatic stellate cells (HSCs) from cirrhotic livers L-type voltageoperated Ca++ channels (L-VOCCs) are over-expressed and responsible for Ca++ -mediated HSC contraction leading to increased resistance to portal flow [2]. CaRs expression and immuno-location in liver, kidneys and mesenteric intestine of cirrhotic rats have never been investigated. Aim and Methods: To address this issue in advanced experimental cirrhosis, we evaluated biomolecular expression of CaRs through Western blot analysis in tissue samples of kidney, liver and small intestine of control and cirrhotic rats. Slices of these organs were also submitted to determination of CaRs immuno-location through indirect immunofluorescence after tissue staining with anti-CaRs, anti-Flk-1 (an endothelial antigen), and anti-nucleus (DAPI blue) monoclonal antibodies. 8 control rats and 8 rats with ascitic cirrhosis induced by 12-week CCl4 intoxication were studied. Results: Compared to controls, hepatic, renal and intestinal CaR protein content was uniformly reduced by at least 30% in cirrhotic rats (all P < 0.05). In cirrhotic rat tissue slices, CaRs were found to be strictly immuno-located in sub-endothelial layers of renal medullary arteries and intrahepatic portal venules. Submucosal expression of CaRs in small bowel was restricted to the tip of villi and no significant association of these receptors with blood vessels was seen. Conclusions: Due to their localization in sub-endothelial layers of kidney arteries and intrahepatic portal venules, without any concurrent expression in smooth muscle layers of mesenteric arteries, CaRs should be targeted by specific pharmacological agonists (i.e. Poly-L-Arginine or Poly-LLysine) [3] in order to manage the complications of portal hypertension without aggravating hyperdynamic circulation in liver cirrhosis. References [1] Am J Physiol 283: F963−20, 2002. [2] Hepatology 33: 1007−8, 2001. [3] Am J Physiol 273: C1168−75, 1997.