New Strategy for Liver Failure by Regulating Hepatic Extracellular Matrix

(1)肝功能衰竭患者血清IV型胶原蛋白水平与肝组织显微变化的意义：在有和无慢性肝损伤患者围手术期测定血清中肝纤维化标志物之一IV型胶原蛋白，探讨该参数是否随肝脏急性应激而改变，并预测肝切除术的手术风险。肝硬化患者血清ⅳ型胶原蛋白水平明显升高。在术后肝功能衰竭的病例中，血清IV型胶原蛋白水平在术前和术后均明显高于正常病例。尸检所得的肝脏显微镜检查结果显示，病变间隙有明显的IV型胶原沉积。在几项术前肝功能检查中，IV型胶原蛋白显示为术后肝衰竭的独立危险因素。因此，围手术期血清IV型胶原蛋白水平的测定似乎有助于预测慢性肝损伤患者肝切除术的风险。(2)体外共培养系统中肝细胞与星状细胞相互作用的研究：采用共培养模型和条件培养基研究了星状细胞对肝细胞增殖和白蛋白合成的影响。当与星状细胞共培养时，即使在无血清条件下，肝细胞DNA合成也得到增强。条件培养基中HGF浓度低于6 ng/ml，刺激肝细胞增殖。肝细胞和星状细胞之间的细胞接触可能是肝细胞DNA合成的另一个因素。共培养不影响白蛋白的产生。另一方面，即使在无血清状态下，与肝细胞共培养也能增强星状细胞的增殖。肝细胞条件培养基和共培养刺激星状细胞DNA合成。因此，肝细胞和星状细胞的DNA合成可能受到来自单个细胞的一些因素的调节，肝细胞和星状细胞之间的细胞间接触可能在它们的增殖和分化中起作用。(3) NAC （n -乙酰半胱氨酸）抑制肝星状细胞增殖和肝纤维化的作用：n -乙酰半胱氨酸剂量依赖性阻断了原代培养的肝星状细胞中血小板源性生长因子诱导的DNA合成和信号转导，包括MAPK和Akt。多种蛋白酶抑制剂的体外降解实验表明，n -乙酰- l-半胱氨酸等还原剂增强了组织蛋白酶B的催化活性，并引发血小板源性生长因子受体的细胞外蛋白水解，从而导致细胞对血小板源性生长因子- bb脱敏。western blot结果显示，肝星状细胞向培养基中分泌成熟组织蛋白酶B，而血管平滑肌细胞不分泌组织蛋白酶B。这一机制在体内肝纤维化模型中得到证实。少

(1) SIGNIFICANCE OF SERUM LEVELS OF TYPE IV COLLAGEN AND MICROSCOPICAL CHANGE IN LIVER TISSUE IN PATIENT WITH LIVER FAILURE : Type IV collagen, one of serum markers for hepatic fibrosis, was measured perioperatively in patients with and without chronic liver damage in order to investigate whether this parameter changes responding to acute stress to the liver and can predict surgical risk of hepatic resection. The serum type IV collagen level was significantly elevated in cases with liver cirrhosis. In cases with postoperative liver failure, serum type IV collagen levels were significantly higher both pre-and postoperatively as compared to those of uneventful cases. Microscopic findings of the liver obtained from autopsy indicated marked amount of deposition of type IV collagem in Disse space. Among several preoperative liver function tests, type IV collagen revealed to be an independent risk factor for postoperative liver failure. Thus, perioperative measurement of the serum type IV co … More llagen levels seemed to be useful for the prediction of the risk of hepatic resection in cases with chronic liver damage.(2)INVESTIGATION OF INTERACTION WITH HEPATOCYTES AND STELLATE CELLS USING IN VITRO CO-CULTURE SYSTEM : Effect of stellate cells on proliferation and albumin synthesis of hepatocytes was investigated using co-culture models and conditioned mediums. When co-cultured with stellate cells, hepatocyte DNA synthesis was enhanced even under a serum-free condition. Conditioned medium, in which HGF concentration was below 6 ng/ml, stimulated the proliferation of hepatocytes. Cell contact between hepatocytes and stellate cells may be an additional factor for DNA synthesis of hepatocytes. Albumin production was not affected by co-culture. On the other hand, stellate cell proliferation was enhanced by co-culture with hepatocytes even under a serum-free status. Conditioned medium of hepatocytes and co-culture stimulated DNA synthesis of stellate cells. Such a mitogenic activity of conditioned medium was partially attenuated by anti-IGF-1 neutralizing antibody Thus, DNA synthesis of hepatocytes and stellate cells may be regulated by some factors derived from individual cells and cell-cell contact between hepatocytes and stellate cells may play a role in their proliferation and differentiation.(3)ACTION OF NAC (N-ACETYL CYSTEINE) TO SUPPRESS PROLIFERATION OF HEPATIC STELLATE CELL AND LIVER FIBROSIS : N-acetyl-L-cysteine dose-dependently blocked platelet-derived growth factor-induced DNA synthesis and signal transduction, including MAPK and Akt, inprimary-cultured hepatic stellate cell. In vitro degradation assay with various protease inhibitors showed that reducing agents, including N-acetyl-L-cysteine, enhanced the catalytical activity of cathepsin B and triggered extracellular proteolysis of platelet-derived growth factor receptor, thereby leading to desensitization of the cell to platelet-derived growth factor-BB.Moreover, western blot showed that hepatic stellate cell secreted mature-cathepsin B into the culture medium, while vascular smooth muscle cell did not secrete cathepsin B.This mechanism was demonstrated in in vivo liver fibrosis models. Less