Effects of Smad4 on liver fibrosis and hepatocarcinogenesis in mice treated with CCl4/ethanol / 中华肝脏病杂志

To study the effects of Smad4 on liver fibrosis and hepatocarcinogenesis in mice treated with CCl(4)/ethanol. The wild-type mice (Smad4 +/+) and the Smad4 knockout mice (Smad4 +/-) were injected subcutaneously with carbon tetrachloride(CCl(4))/ethanol twice a week for twenty weeks. The expression of Smad4, TGFbeta1, Smad2, Smad3, Smad6, TIMP1, MMP2 and MMP9 was detected by RT-PCR. In the cirrhotic liver, the expression of Smad4 mRNA was significantly higher than that in the normal liver. Comparing with wild-type mice (Smad4 +/+), the TGFbeta1-Smad4 signaling was markedly attenuated in the Smad4 knockout mice (Smad4 +/-). After induction by CCl(4)/ethanol, the hepatic fibrosis in the Smad4 knockout mice (Smad4 +/-) was obviously alleviated compared with the wild-type mice (Smad4 +/+), and the incidence rate of hepatocarcinogenesis of the former was also lower than that of the latter(32.0% vs 41.9%). These results indicate that knocking out Smad4 can delay the progression of liver fibrosis and liver cancer.

Surgical treatment for hilar cholangiocarcinoma of Bismuth-Corlette type IV / 中华外科杂志

<p><b>OBJECTIVE</b>To investigate the surgical treatment for hilar cholangiocarcinoma(HCCA) of Bismuth-Corlette type IV.</p><p><b>METHODS</b>The clinical data of 73 HCCA patients admitted in Southwest Hospital, the Third Military Medical University from January 2002 to December 2008 were analyzed retrospectively. There were 41 males and 32 females, aged from 30 to 84 years old (averaged, 56.8 years old). All patients were diagnosed as hilar mass with hilar biliary obstruction by B-ultrasound, CT, magnetic resonance imaging, endoscopic retrograde cholangiopancreatography, magnetic resonance cholangiopancreatography or percutaneous transhepatic cholangiography, and confirmed by pathological examination in intra-operation and post-operation. Diagnosis of all patients were according to Bismuth-Corlette type IV.</p><p><b>RESULTS</b>Fifteen cases underwent percutaneous transhepatic cholangial drainage, stents were implanted in 8 patients. Simple internal drainage was performed on 25 of the 73 patients and 4 with palliative resection. Radical resection was performed on 19 patients. The radical resection rate of HCCA were 26.0%. The 1, 3 years of survival rates were 36.8%, 10.5% respectively. The 1 year survival rate was 6.3% in drainage group.</p><p><b>CONCLUSION</b>Radical resection is the potentially curative treatment for HCCA, a sufficient, reasonable use of internal and external drainage would improve the patient's quality of life.</p>

Experimental study on tolerance time limits from warm ischemia to cold preservation of liver grafts / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the tolerance time limits from warm ischemia to cold preservation of liver grafts.</p><p><b>METHODS</b>Orthotopic liver transplantations (OLTs) were performed on Bama miniature swine. Morphological and functional changes of the liver grafts and biliary tracts after 10 minutes of warm ischemia followed by different durations of cold preservation and its reversibility were investigated.</p><p><b>RESULTS</b>When the grafts were subjected to 10 minutes of warm ischemia followed by less than 16 hours of cold preservation, all animals could survive 1 week and there was no animal death from biliary necrosis. However, when the cold preservation time exceeded 16 hours, the incidence of biliary necrosis was significantly increased (P<0.05), and recipient death from bile leaks occurred. With further prolongation of the cold preservation time, primary graft nonfunction and intraoperative or early postoperative deaths occurred and the living animals all developed biliary necrosis. When compared with the less than 16 hours cold preservation group, the morphological scores and apoptosis index of the epithelial cells of bile ducts in grafts after reperfusion were significantly elevated in the more than 16 hours cold preservation group (P<0.05) and the activity of Na+-K+-ATPase and Ca2+-ATPase of bile ducts in grafts were also significantly reduced (P<0.05). Liver function tests showed that the recoveries of AST, AST, GGT and ALP were quicker in the 16 hours cold preservation group then those over 16 hour preservation ones. Correlation analysis revealed that the incidence of biliary necrosis was significantly correlated with the morphological score (r = 0.972) and with the apoptosis index of the epithelial cells of bile ducts in grafts after reperfusion (r = 0.931) and also correlated negatively (P<0.01) with the activity of Na+-K+-ATPase (r = -0.973) and Ca2+-ATPase (r = -0.973).</p><p><b>CONCLUSIONS</b>It is concluded that with 10 minutes of warm ischemia, cold preservation of the grafts should not be longer than 16 hours in order to avoid early biliary necrosis, and the corresponding tolerance time limit of the livers to the cold preservation was less than 20 hours.</p>

Effect of proto-oncogene protein c-met ribozyme transfection on biological behavior of gastric carcinoma cells / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To explore the effect of c-met ribozyme transfection on biological behavior of gastric carcinoma cells.</p><p><b>METHODS</b>U1/met292 plasmid containing c-met ribozyme gene was transfected into L2 subline of SGC-7901 gastric cell line, and the proliferative ability, distribution of cell cycle, protein expression of VEGF and c-met, as well as the potential of liver metastasis of the transfected subline were determined.</p><p><b>RESULTS</b>There were no significant difference in proliferative ability, distribution of cell cycle between the transfected cells and the control cells. The protein expression of VEGF and c-met, as well as the liver metastatic potential significantly decreased in the transfected cells than those in the control cells (P< 0.05).</p><p><b>CONCLUSION</b>The liver metastatic potential of c-met positive gastric cancer cells may be prevented by inhibiting c-met expression.</p>

Effects of portaazygous disconnection, portocaval shunt and selective shunts on experimental rat liver cirrhosis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To evaluate the effects of portaazygous disconnection (PAD), portacaval shunt (PCS) and distal splenocaval shunt (DSCS) on the portosytemic shunting (PSS), hepatic function (HF), hepatic mitochondrial respiratory function (HMRF), oral glucose tolerance test (OGTT) and arterial ketone body ratio (KBR) in order to provide a sound basis for selecting suitable operations for patients.</p><p><b>METHODS</b>Using a cirrhotic portal hypertensive model induced by CCl4/ethanol in Wistar rats, the PSS, HF, HMRF, OGTT and KBR were determined three weeks after PCS, DSCS and PAD.</p><p><b>RESULTS</b>It was revealed that: (1) In the cirrhotic portal hypertension rats, the PSS increased significantly, HMRF and hepatic reserve function (HRF) decreased significantly when compared with the control rats. (2) At the time of first postoperative week, the mean blood glucose value in the 120-minute OGTT in each PAD, PCS and DSCS groups had significant differences compared with the cirrhotic control group. But during the second and third postoperative weeks, the mean blood glucose values in the 120-minute OGTT in both PAD and DSCS groups had no significant differences compared with the cirrhotic control group except for the PCS group. The values of KBR in the three operative groups decreased significantly compared with the cirrhotic control group during the two postoperative weeks. In the third postoperative week, only the values of KBR in the PCS group had a significant difference compared with the cirrhotic control group. (3) After PCS, the PSS was further increased; HF and HMRF were significantly decreased. Little improvement was found in the third postoperative week. (4) After DSCS and PAD, the above mentioned indices were less influenced, and they were restored more quickly than those in the PCS group.</p><p><b>CONCLUSION</b>We found that PAD and DSCS are more desirable than PCS.</p>

Inhibitory effect of retroviral vector containing anti-sense Smad4 gene on Ito cell line, LI90 / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Transforming growth factor-beta1 (TGF-beta1) exerts strong fibrogenic potential in culture-activated HSCs. Smad4 is a key intracellular mediator for the transforming growth factor-beta (TGF-beta) superfamily of growth factors. The aim of this study was to assess the effects of the antisense Smad4 gene on Ito cell line, LI90.</p><p><b>METHODS</b>The recombinant retroviral vector pLXSN-Smad4 was constructed by cloning the rat antisense Smad4 cDNA into the retroviral vector pLXSN. Retroviruses with or without the antisense gene were obtained by transfecting pLXSN-Smad4 and pLXSN vectors into PA317 cells. Human hepatic stellate cells (HSCs) LI90 were infected with these retroviruses followed by selection with G418. The expression of Smad4 was detected by Northern and Western blots. Cell biological characteristics, including cell growth curve, 3H-TdR and 3H-proline uptake by HSCs and the production of extracellular matrix were assessed.</p><p><b>RESULTS</b>mRNA and protein expressions of Smad4 in LI90 cells transfected with retrovirus containing the antisense Smad4 gene were much lower than those in LI90 cells transfected with empty vector or parental LI90 cells. Cells hypoexpressing the Smad4 gene exhibited a slower rate of growth, a lower uptake of 3H-TdR and 3H-proline (P < 0.01), and smaller production of th extracellular matrix, compared with parental LI90 cells and cells transfected with empty retrovirus.</p><p><b>CONCLUSIONS</b>The antisense Smad4 gene can suppress the expression of the Smad4 gene, reduce endogenous production of Smad4 mRNA and protein, block TGF-beta1 signaling pathway, inhibit activation of Ito cells, obstruct the growth of Ito cells, decrease the production of the extracellular matrix (ECM). Our results may provide a basis for the development of antifibrotic gene therapy.</p>

Obstruction of TGF-beta1 signal transduction by anti-Smad4 gene can therapy experimental liver fibrosis in the rat / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the therapeutic effects to block the TGF-beta1 (transforming growth factor beta1) signal transduction by antisense Smad4 gene on experimental fibrotic liver.</p><p><b>METHODS</b>Using the rat model of liver fibrosis induced by Carbon Tetrachloride (CCl4)/ethanol, we transfected antisense Smad4 gene mediated by adenovirus via portal vein infusion into the liver, and observed the expression of Smad4 by Retro-Polymerase Chain Reaction (RT-PCR) and Western Blot. We also investigated the pathologic features and collagen expression.</p><p><b>RESULTS</b>In the non-therapeutic cirrhotic liver, the expression of Smad4 mRNA was significantly increased than normal liver, and so was the collagen I. After antisense Smad4 gene being transfected, the expression of Smad4 mRNA and that of collagen I in the therapeutic liver was significantly decreased, compared with the non-therapeutic cirrhotic liver. The fibrous degree of therapeutic liver was also reduced compared with the non-therapeutic fibrous liver.</p><p><b>CONCLUSION</b>These results indicate that because antisense Smad4 gene could block TGF-beta1 signal transduction by reducing the expression of Smad4, so it could inhibit the production of extracellular matrix (ECM) and improve hepatic fibrosis.</p>

The feasibility and safety of radiofrequency ablation for secondary splenomegaly and hypersplenism in dogs / 中华外科杂志

<p><b>OBJECTIVE</b>To assess the feasibility and safety of radiofrequency ablation (RFA) in spleen to treat secondary splenomegaly and hypersplenism in dogs.</p><p><b>METHODS</b>Fourteen healthy mongrel dogs were randomly divided into two groups: group A (n = 4) and group B (n = 10) Both groups underwent ligation of the splenic vein and its collateral branches to induce congestive splenomegaly. At the end of the 3rd week, radiofrequency thermal ablation of the spleen was performed in the group B via laparotomy. After splenic RFA, the procedure-related complications were observed, CT scan was performed and the spleens were obtained according to schedule. The radiofrequency (RF) thermal lesions and its histo- pathological changes of the spleen were examined regularly.</p><p><b>RESULTS</b>There were no morbidity and mortality in the experimental dogs. CT findings revealed that splenomegaly could sustained over 2 months after ligation of the splenic vein. The segmental RF lesions included hyperintense zone of coagulative necrosis and more extensively peripheral hypo-intense infarcted zone. The latter was called as "bystander effect". The infarcted zone would be absorbed and subsequently disappeared between 4 and 6 weeks after RFA, and the size of the remnant spleen shrunk, but the lesion of coagulative necrosis hardly altered. The fundamental histopathological changes of splenic lesions caused by RF thermal energy included local coagulative necrosis and the peripheral thrombotic infarction zone. Subsequently, tissue absorption and fibrosis occurred in the zone of thrombotic infarction. Simultaneously occluded vessels, fibrin deposition, and disappearance of normal splenic sinuses resulted in the condensed structure of the viable remnant spleen, which were the pathological basis responsible for the shrunk spleen.</p><p><b>CONCLUSIONS</b>It is feasible and safe to perform RFA in the spleen to treat experimental splenomegaly and hypersplenism. The RFA technique could be safely performed clinically via laparotomy or laparoscopic procedure to strictly isolate the spleen from the surrounding organs.</p>

Determination of telomerase activity in carcinoma of bile duct and its clinical significance / 第三军医大学学报

Objective　To explore the relationship between te lomerase activity and bile duct carcinoma and its significance in clinical diag nosis. Methods　Telomerase activities were examined in 23 cases of bile duct carcinoma, 5 cases of carcinoma adjacent tissues and 5 cases of nor mal bile duct tissues respectively with telomerase PCR-ELISA method. Results　Telomerase activity was detected in 18 of 23 cases (78.3%) of b ile duct carcinoma, and was not found in 5 cases of carcinoma adjacent tissues a nd 5 cases of normal bile duct tissues. The detection rate of telomerase activit y had no correlation with patients' age, sex, tumor site and size but significan tly correlated to metastasis of tumor (P<0.05). Conclusion　The level of telomerase activity was significantly higher in bile duct carcino ma and may be served as one of the clinical marker for malignant neoplasm becaus e of its spsecificity.

Effects of sympathetic denervation on liver regeneration after partial hepatic resection / 第三军医大学学报

Objective　To establish the animal model of denervation of sympathetic nerve and to explore the effects of denervation of the sympathetic nerve on liver regeneration after partial resection. Methods　The animal model of denervation of sympathetic nerve was made with 6-OHDA. A total of thirty male Wistar rats were divided equally into experimental and control group. The left and middle lobe of liver were resected with improved Higgins and Anderson's method. Meanwhile, denervation was made in the experimental group. All the rats were killed by haemospasia on the 7 th day after operation. HMI, RLR and MI were measured. The rates of DNA synthesis were detected by 3H-TdR method. Results　The concentration of NE decreased extremely on day 3 to day 14 after administration of 6-ONDA. No death happened in all the rats 7 days after liver resection. HMI, RLR, MI and 3H-TdR incorporation significantly decreased in experimental group compared with that in control (P<0.01). Conclusion　The chemical denervation of sympathetic nerve can be aroused by administration of 6-OHDA. Regeneration of the liver is inhibited by the denervation of sympathetic nerves.

Effects of phenylephedrine on free calcium distribution in hepatocytes / 第三军医大学学报

Objective　To investigate the changes of free calcium distribution in hepatocytes after administration of phenylephedrine. Methods The changes of fluorescence intensity were observed by confocal laser scanning microscopy after administration of phenylephedrine alone or pretreated with phentolamine before phenylephedrine administration. Results　The fluorescence intensity increased rapidly after administration of phenylephedrine to hepatocytes. When liver cells were pretreated by phentolamine before phenylephedrine administration, the changes of fluorescence intensity not obvious. Meanwhile, the inconformity of the fluorescence intensity in hepatocytes suggested the existence of the second subarea of free calcium distribution. Conclusion　Ca2+ signal can be arisen by phenylephedrine via the α-receptor in hepatocytes in vitro. The distribution and dynamic changes of free calcium in hepatocytes display some characteristics.

Determination of telomerase activity in carcinoma of bile duct and its clinical significance / 第三军医大学学报

Objective　To explore the relationship between te lomerase activity and bile duct carcinoma and its significance in clinical diag nosis. Methods　Telomerase activities were examined in 23 cases of bile duct carcinoma, 5 cases of carcinoma adjacent tissues and 5 cases of nor mal bile duct tissues respectively with telomerase PCR-ELISA method. Results　Telomerase activity was detected in 18 of 23 cases (78.3%) of b ile duct carcinoma, and was not found in 5 cases of carcinoma adjacent tissues a nd 5 cases of normal bile duct tissues. The detection rate of telomerase activit y had no correlation with patients' age, sex, tumor site and size but significan tly correlated to metastasis of tumor (P<0.05). Conclusion　The level of telomerase activity was significantly higher in bile duct carcino ma and may be served as one of the clinical marker for malignant neoplasm becaus e of its spsecificity.

Effects of sympathetic denervation on liver regeneration after partial hepatic resection / 第三军医大学学报

Objective　To establish the animal model of denervation of sympathetic nerve and to explore the effects of denervation of the sympathetic nerve on liver regeneration after partial resection. Methods　The animal model of denervation of sympathetic nerve was made with 6-OHDA. A total of thirty male Wistar rats were divided equally into experimental and control group. The left and middle lobe of liver were resected with improved Higgins and Anderson's method. Meanwhile, denervation was made in the experimental group. All the rats were killed by haemospasia on the 7 th day after operation. HMI, RLR and MI were measured. The rates of DNA synthesis were detected by 3H-TdR method. Results　The concentration of NE decreased extremely on day 3 to day 14 after administration of 6-ONDA. No death happened in all the rats 7 days after liver resection. HMI, RLR, MI and 3H-TdR incorporation significantly decreased in experimental group compared with that in control (P<0.01). Conclusion　The chemical denervation of sympathetic nerve can be aroused by administration of 6-OHDA. Regeneration of the liver is inhibited by the denervation of sympathetic nerves.

Effects of phenylephedrine on free calcium distribution in hepatocytes / 第三军医大学学报

Objective　To investigate the changes of free calcium distribution in hepatocytes after administration of phenylephedrine. Methods The changes of fluorescence intensity were observed by confocal laser scanning microscopy after administration of phenylephedrine alone or pretreated with phentolamine before phenylephedrine administration. Results　The fluorescence intensity increased rapidly after administration of phenylephedrine to hepatocytes. When liver cells were pretreated by phentolamine before phenylephedrine administration, the changes of fluorescence intensity not obvious. Meanwhile, the inconformity of the fluorescence intensity in hepatocytes suggested the existence of the second subarea of free calcium distribution. Conclusion　Ca2+ signal can be arisen by phenylephedrine via the α-receptor in hepatocytes in vitro. The distribution and dynamic changes of free calcium in hepatocytes display some characteristics.