Effects of mitofusin-2 gene on cell proliferation and chemotherapy sensitivity of MCF-7 / 华中科技大学学报（医学）（英德文版）

In order to evaluate the effect of mitofusin-2 gene (mfn2) on proliferation and chemotherapy sensitivity of human breast carcinoma cell line MCF-7 in vitro, pEGFPmfn2 plasmid carrying full length of mitofusin-2 gene was transfected, by using sofast, into MCF-7 cells. Mitofusin-2 gene expression in MCF-7 cells transfected by sofast after 48 h was detected by PCR and Western blotting, and the stable expression of GFP protein in MCF-7 cells by Western blot analysis. The proliferation of MCF-7 cells was assayed by MTT and cell counting. By using PI method, the effects of mfn2 on the cell cycle distribution of MCF-7 were measured. Annexin-V/PI double labeling method was employed to detect the changes in apoptosis induced by chemotherapeutics before and after transfection. The results showed that the MCF-7 cells transfected with mfn2 gene could stably and highly express GFP protein. MTT assay revealed that after transfection of mfn2 cDNA, the proliferation of MCF-7 cells was significantly inhibited. DNA histogram showed that cells arrested in S phase, and the percentage of S phase cells was 42.7, 17.2 and 19.6 in mfn2 cDNA transfection group, blank plasmid transfection group and blank control group, respectively (P<0.05). The apoptosis ratio of the cells transfected with mfn2 gene was increased from 3.56% to 15.95%, that of the cells treated with camptothecin (CAMP) followed by mfn2 gene transfection was 69.6%, and that in blank plasmid transfection group and blank control group was 31.0% and 23.4% respectively (P<0.05). It was suggested that transfection of mfn2 gene could significantly inhibit the proliferation of MCF-7 cells and promote their sensitivity to CAMP with a synergic effect.

The Protective Effects of In Vitro Cultivated Calculus Bovis on the Cerebral and Myocardial Cells in Hypoxic Mice / 华中科技大学学报（医学）（英德文版）

The protective effects of in vitro cultivated calculus bovis (ICCB) on the cerebral and myocardial cells in hypoxic mice and the mechanism were examined. In one group, mice were intra-gastrically (i.g.) given ICCB for 15 days and then they were subjected to acute cerebral ischemia by decapitation, and then the panting time was recorded. In the other group, 12 min after exposure to hypoxia, mice was administered the ICCB i.g. for 5 days, and then the blood serum and tissues of brain,heart, liver were harvested and examined for SOD, GSH-px and T-AOC activity and content of MDA. The tissues of brain and heart were observed electron-microscopically for ultrastructural changes. The corpus striatum and hippocampus of brain were collected and examined for content of dopamine (DA) and norepinephrine (NE). The ultrastrural examination showed that the pathological change in brain and heart in the ICCB group was very slight, while abnormal changes in the control group were obviously more serious. ICCB significantly prolonged the panting time of the hypoxic mice (P<0.001), increased the activity of SOD, GSH-px, T-AOC in serum and tissues of brain, liver,heart and elevated the content of DA and NE. ICCB also pronouncedly reduced content of MDA in serum and tissues of brain, heart and liver. Significant differences in these parameters were noted between ICCB group and controls. It is concluded that ICCB can exert protective effect on the cells of brain and myocardium by enhancing the tolerance of the tissues to hypoxia and the body's ability to remove free radicals and regulating the neurotransmitters.

Effects of 2-APB on store-operated Ca2+ channel currents of hepatocytes after hepatic ischemia/reperfusion injury in rats / 华中科技大学学报（医学）（英德文版）

The effects of hepatic ischemia/reperfusion (1/R) injuries on hepatocellular viability and store-operated calcium current (Isoc) in isolated rat hepatocytes and the effects of 2-APB on store-operated calcium current (Isoc) in isolated rat hepatocytes after hepatic ischemia/reperfusion injuries were studied. Hepatic ischemia and reperfusion injury model was established and whole cell patch-clamp techniques were used to investigate the effects of 2-APB on Isoc. The results showed that ischemia/reperfusion injuries could significantly reduce hepatocellular viability and further increase Isoc in hepatocytes and 2-APB (20, 40, 60, 80, 100 micromol/L,) produced a concentration-dependent decrease of Isoc with IC50 value of 64. 63 +/- 10.56 micromol/L, (n = 8). It was concluded that ischemia/reperfusion injuries could reduce hepatocellular viability, probably through increased Isoc in hepatocytes and 2-APB had a protective effect on ischemia/reperfusion-induced liver injury, probably though inhibiting Isoc.

Effects of 2-APB on Store-operated Ca2+ Channel Currents of Hepatocytes after Hepatic Ischemia/Reperfusion Injury in Rats / 华中科技大学学报（医学）（英德文版）

The effects of hepatic ischemia/reperfusion (I/R) injuries on hepatocellular viability and store-operated calcium current (Isoc) in isolated rat hepatocytes and the effects of 2-APB on storeoperated calcium current (Isoc) in isolated rat hepatocytes after hepaticischemia/reperfusion injuries were studied. Hepatic ischemia and reperfusion injury model was established and whole cell patch-clamp techniques were used to investigate the effects of 2-APB on Isoc. The results showed that ischemia/reperfusion injuries could significantly reduce hepatocellular viability and further increase Isoc in hepatocytes and 2-APB (20, 40, 60, 80, 100 μmol/L) produced a concentration-dependent decrease of Isoc with IC50 value of 64.63±10.56 μmol/L (n= 8). It was concluded that ischemia/reperfusion injuries could reduce hepatocellular viability, probably through increased Isoc in hepatocytes and 2-APB had a protective effect on ischemia/reperfusion-induced liver injury, probably though inhibiting Isoc.

KLF6mRNA expression in primary hepatocellular carcinoma / 华中科技大学学报（医学）（英德文版）

To investigate the expression of KLF6mRNA in primary hepatocellular carcinoma (HCC), nomal liver tissues and the tissues adjacent to the cancers, reverse-transcription polymerase chain reaction (RT-PCR) was employed to investigate the expression of the KLF6 gene in HCC, the corresponding adjacent non-cancerous tissues and normal liver tissue. Our results showed that an amplified fragment of 427 bp DNA was detected in 18 of 19 (94.7%) adjacent non-cancerous tissues and normal liver tissue, and in 12 (85.7%) of 14 HCC. There were no significant differences in the levels of KLF6 mRNA between normal liver and liver tumors (P>0.05). It is concluded that KLF6 mRNA is generally expressed in HCC.

Effects of dendritic cells transfected with full length wild type P53 and modified by gastric cancer lysate on immune response / 华中科技大学学报（医学）（英德文版）

To investigate the effects of dendritic cells (DCs) transfected with full length wild type p53 and modified by gastric cancer lysates on immune response, the wild type P53 was transducted to DCs with adenovirus, and the DCs were modified by gastric cancer lysates (Lywt-P53DC). The concentration of the surface molecules (B7-1, B7-2, MHC-I , MHC-II) of all DCs was determined by FACS, and the ability of the DCs to induce efficient and specific immunological response in anti-51Cr-labeled target cells studied. BALB/c mice model infected with DCs and Mk28 was established. CTL response in mice immunized with Lywt-p53DC and the effectiveness of Lywt-p53DC in the treatment of tumor-bearing mice was assayed. FACS revealed that the surface molecules of Ly-wt-P53 DC had a high expression: for B7-1 86.70% +/- 0.07%, B7-2 18.77% +/- 0.08%, MHC-I 87.20% +/- 0.05%, MHC-II 56.70%+/-0.07%; The T lymphocytes had a specific CTL lysing ability induced by Lywt-P53DC with the CTL lysis rate being 81%. The immune protective effect of Lywt-p53DC group was more obvious than any other groups (P<0.05). The tumor diameter in Lywt-p53DC group was 3.10+/-0.31 mm, 2.73+/-0.23 mm, 3.70+/-0.07 mm on the day 13, 16 and 19, smaller than DC, wtp53DC and LyDC groups (P<0.05). On the other hand, the growth rate of tumor in Lywt-p53DC group was slower than any other groups (P<0.05). It was suggested that DCs transfected with wild type P53 and modified by gastric cancer lysates had specific CTL killing capability.

KLF6mRNA expression in primary hepatocellular carcinoma / 华中科技大学学报（医学）（英德文版）

To investigate the expression of KLF6mRNA in primary hepatocellular carcinoma (HCC), nomal liver tissues and the tissues adjacent to the cancers, reverse-transcription polymerase chain reaction (RT-PCR) was employed to investigate the expression of the KLF6 gene in HCC, the corresponding adjacent non-cancerous tissues and normal liver tissue. Our results showed that an amplified fragment of 427 bp DNA was detected in 18 of 19 (94.7%) adjacent non-cancerous tissues and normal liver tissue, and in 12 (85.7%) of 14 HCC. There were no significant differences in the levels of KLF6 mRNA between normal liver and liver tumors (P>0.05). It is concluded that KLF6 mRNA is generally expressed in HCC.

An antisense plasmid targeting survivin expression induces apoptosis and sensitizes hepatocarcinoma cells to chemotherapy / 华中科技大学学报（医学）（英德文版）

To explore the change of sensitivity to chemotherapy of antisense RNA targeting survivin on hepatocarcinoma carcinoma cells in vitro. Survivin mRNA structure region was amplified by RT-PCR and inserted inversely into eukaryotic expression vector pcDNA3. The antisense expression plasmid pcDNA3/survivin was transfected into HepG2 with lipofectAMINE 2000 (LF2000), with low concentration of 5-fluorouracil (5-Fu) added. Survivin protein was detected by Western-blot, the growth activity was measured by MTT, and apoptosis was detected by Flow Cytometry 12 h, 24 h, 48 h after transfection. The activity of caspase-3 was found by quantitative assay 48 h after transfection. The construction of antisense RNA vector pcDNA3/survivin was verified by restricted endonuclease digestion and nucleotide sequencing. Compared with normal group, 5-Fu and antisense survivin group, the cells growth inhibition, apoptosis index, and caspase-3 activity were increased in antisense survivin transfected + 5-Fu group. The threshold of apoptosis was decreased after survivin was silenced, and the sensitivity to chemotherapy was increased. These findings suggest the existence of a potential new target for gene therapy.

Pathological morphology alteration of the splanchnic vascular wall in portal hypertensive patients / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To investigate the pathological morphology alteration of the splanchnic vascular wall in portal hypertensive patients.</p><p><b>METHODS</b>Splenic arteries, veins and gastric coronary veins from portal hypertensive patients (n = 50) were removed during esophagogastric devascularization with splenectomy and were observed under optic and electron microscopes. The expression of iNOS in the splenic artery wall was analysed with immunohistochemistry.</p><p><b>RESULTS</b>The internal elastic membrane and medial elastic fibers of the splenic artery wall were broken and degenerated. Atrophy, apoptosis and phenotypic changes were seen in smooth muscle cells of splenic arteries. Positive staining for iNOS was seen in the cytoplasm of smooth muscle cells and iNOS activity was elevated compared with the non-cirrhotic patients (P < 0.01). In the splenic and gastric coronary veins of cirrhotic patients, we found proliferative intima, extensive thrombi adhering to the venous wall, mimicked arteriosclerosis plaques accompanied with hypertrophy of smooth muscle cells, and thickened muscle fibers of veins with increase in extracellular matrix.</p><p><b>CONCLUSION</b>Portal hypertension may be complicated by splanchnic arterial and venous vasculopathy. There may be an interactive relationship among portal hypertension, splanchnic hyperdynamic disturbances and splanchnic vasculopathy in the pathogenesis of portal hypertension.</p>

Pathogenesis of hilar cholangiocarcinoma and infection of hepatitis virus / 中华外科杂志

OBJECTIVE To study the correlation between hepatic hilar cholangiocarcinoma and the infection of HB (C)V. METHODS Combined with clinical data, immunohistochemistry was used to detect HBXAg antigen and HCV-C protein in 68 formalin-fixed and paraffin-embedded samples of hilar cholangiocarcinoma. RESULTS Six cases(8.8%) were positive to HBXAg antigen and 24 cases (35.0%) to HCV-C protein respectively. One case was positive to both HBXAg antigen and HCV-C protein. There were statistical differences in differentiation and invasion, lymph node metastasis, treatment between hilar cholangiocarcinomas infected HB(C)V and those non-infected. CONCLUSIONS HBXAg antigen and HCV-C protein may play an important role in the pathogenesis of hilar cholangiocarcinoma. Hilar cholangiocarcinoma infected HB(C)V may have a high malignancy and more poor prognosis.

Influence of spleen on cell cycle and expression of proliferating cell nuclear antigen of the liver during diethylnitrosamine-induced rat hepatocarcinogenesis / 中华外科杂志

<p><b>OBJECTIVE</b>To investigate the role of the spleen on hepatocyte proliferative activity during experimental hepatocarcinogenesis in rats.</p><p><b>METHODS</b>Cell-cycle and expression of proliferating cell nuclear antigen (PCNA) of hepatocytes were studied by immunohistochemical and flow cytometric technique in two groups: splenectomy (45 rats) and sham-operation with laparotomy (45 rats).</p><p><b>RESULTS</b>(1) Hepatocirrhosis was formed in group A earlier than in group B. Marked degenerative changes of the liver parachyma showing vacuolization of hepatocytes were observed in hepatic lobules. (2) The proliferative level of hepatocytes increased with the progression of hepatocarcinogenesis, and topped at the 18 th week. (3) The proliferative level of the splenectomy group was lower than that of the sham-operation group in the mid-stage of carcinogenesis (t = 4.76, P < 0.05). After stopping feeding diethylnitrosamine (DENA), however, no difference was found in the hepatocyte proliferative activity between the two groups at the 18th week.</p><p><b>CONCLUSIONS</b>There is a close relationship between hepatic proliferative activity and spleen, and the spleen may play an important role in facilitating hepatic proliferation.</p>

Frequency of loss expression of the DPC4 protein in various locations of biliary tract carcinoma / 中华预防医学杂志

<p><b>OBJECTIVE</b>To clarify the relationship between the loss of expression of the deleted in pancreatic carcinoma locus 4 (DPC4) proteins and the pathogenesis of biliary tract carcinoma.</p><p><b>METHODS</b>71 primary biliary tract carcinoma (BTCa), including 38 common bile duct (CBD) carcinomas, 18 gallbladder carcinomas, 15 hilar bile ducts (HBD) carcinomas were examined by immunohistochemical staining. In addition, the CBD carcinomas were divided into two groups: tumors with metastasis (M(+) group, 27 cases) and tumors without metastasis (M(-) group, 11 cases).</p><p><b>RESULTS</b>The frequency of loss of the expression of DPC4 protein was 32.8% in BTCa, 47.3% in CBD carcinoma, 11% in gallbladder carcinoma, 13% in HBD carcinoma. Comparison of the frequency of loss expression of DPC4 was significant statistical difference in CBD carcinoma versus gallbladder carcinoma and HBD carcinoma (P < 0.01). The frequency of loss expression of DPC4 was 48.1% in the M(+) group and 45.4% in the M(-) group.</p><p><b>CONCLUSION</b>There are a close relationship between pathogenesis of BTCa and inactivation of DPC4 and different frequencies of DPC4 gene alternation in various locations of the biliary tract, which are not significantly increased with tumor metastasis in BTCa.</p>

NF-kappa B expression in cholangiocarcinoma transfected with hepatitis C virus core gene / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the role of hepatitis C virus (HCV) in the development of cholangiocarcinoma.</p><p><b>METHODS</b>Recombinant plasmid of HCV-C gene constructed by molecular cloning technique was identified with restricting enzyme map. Then, it was transfected into QBC939 cells with lipofectin. After selection with G418, the resistant colonies were obtained and analysed by immunocytochemistry and Western blotting. Their morphology was observed by transmission electron microscopy (TEM). The expression of NF-kappa B was detected by immunocytochemistry.</p><p><b>RESULTS</b>The results suggested that the recombinant plasmid was proved to carry the target gene by restricting enzyme map. Moreover, it could express HCV-C protein efficiently in QBC939 cells. The HCV-like particles were found in the cytoplasm by TEM, which were spherical with diameter of 50-80 nm possessing an outer membrane. Moreover, NF-kappa B activation was shown in HCV core gene-transfected cells.</p><p><b>CONCLUSION</b>Because HCV-C gene could express steadily in cholangiocarcinoma cells, the transfected tumor cells (QBC939-HCVc) are an experimental model for studying the effect of HCV on the development of cholangiocarcinoma. The activation of NF-kappa B may be related to escape from immune surveillance and carcinogenesis of cholangiocarcinoma.</p>

Expression of nuclear factor kappa B in hepatitis C virus core gene transfected cholangiocarcinoma cells / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To establish an experimental model for exploring the role of hepatitis C virus (HCV) in the development of cholangiocarcinoma.</p><p><b>METHODS</b>Recombinant plasmids of HCV-C gene were constructed by molecular cloning techniques and identified by PCR and restriction enzyme mapping.The plasmids were then transfected into QBC939 cells (a cholangiocarcinoma cell line) by Lipofection. After selection with G418, resistant colonies were obtained and analyzed by immunocytochemistry and Western blotting. The morphology was observed by trans mission electron microscopy (TEM). The expression of NF-(k)B was detected by immunocytochemistry.</p><p><b>RESULTS</b>Recombinant plasmid was shown by PCR and restriction enzyme mapping to carry the target gene. Moreover, it could efficiently express HCV-C protein in QBC939 cells. HCV-like particles were found in the cytoplasm by TEM, which were spherical with a diameter of 50-80 nm and possessed an outer membrane. Moreover, NF-(k)B activation could be shown in HCV core-transfected cells.</p><p><b>CONCLUSION</b>Expression of the HCV-C gene in cholangiocarcinoma cells was achieved. Transfected tumor cells (QBC939-HCVc) could be used as a model to study the effect of HCV on the development of cholangiocarcinoma.</p>

Expression of TREM-1 in patients with biliary infection / 中国普通外科杂志

Objective To investigate the expression and significance of human TREM-1 mRNA in patients with (biliary) infection. Methods Peripheral blood of 32 patients with biliary infection and 7 healthy volunteers were (collected). TREM-1 mRNA was determined by semi-quantitative RT-PCR. TNF-? was determined by ELISA method. Results The values of TREM-1/?-actin of control group was 0.48?0.072, while those of biliary infection group in 1d, 2d, 3d, 7d were 0.93?0.070,0.90?0.060,0.82?0.092,0.66?0.062 respectively (P

Proliferative activity for a cholangiocarcinoma cell line of bile from patients with choledocholithiasis / 中国普通外科杂志

Objective To explore the effects of bile from patients with choledocholithiasis on the growth of human cholangiocarcinoma cell line QBC939 and the potential relation between choledocholithiasis and cholangiocarcinoma. MethodsWT5”BZ Choledocholithiasis bile (CB) and normal bile (NB) specimen were used for this study. The proliferative effects of bile were measured by methabenzthiazuron (MTT) assay and cell cycle and apoptosis were analyzed by flow cytometry. KG2Results CB significantly promoted the proliferation of QBC939 cells, QBC939 proliferative index increased significantly after treated with 1% CB for 48 h ( P

Effect of integrin-?1 antisense oligodeoxynucleotides on human pancreatic cancinoma transplanted subcutaneously in nude mice / 中国普通外科杂志

Objective To investigate the effect of integrin-?1 antisense oligodeoxynucleotide(ASODN) on(human) pancreatic cancinoma transplanted subcutaneously in nude mice.Methods The models of human(pancreatic) cancinoma transplanted subcutaneously were established in nude mice,then divided randomly into 3 groups and different treatment was given respectively(control group,random oligodeoxynucleotide group and ASODN group).After treatment,the weight of nude mice and tumor volume were observed,and the tumor growth inhibitory rate and the tumor response rate were calculated.The expressions of integrin-?1 mRNA and protein in tumor tissue were determined by RT-PCR and Western-blot.Results The tumor growth inhibitory rate in the random oligodexynucleotide group and the ASODN group was 4.75% and 72.70%,respectively.The tumor decrease rate of the ASODN group was 10.91%.The expression level of integrin-?1 mRNA and protein was decreased in the ASODN group compared with other 2 control groups. Conclusions Our findings suggest that integrin-?1 antisense oligodeoxynucleotides result in marked inhibition of human pancreatic(cancinoma) growth in nude mice.It may be a novel treatment approach for human pancreatic carcinoma.

Proliferative activity of bile of patients undergoing transduodenal sphincteroplasty / 中国普通外科杂志

Objective To explore the effects of bile in patients undergoing transduodenal sphincteroplasty (TSB) on the growth of human cholangiocarcinoma cell line QBC939 and the potential relation between transduodenal sphincteroplasty and cholangiocarcinoma. Methods TSB and normal bile (NB) sample were used for this study. The proliferative effect of bile was measured by using methabenzthiazuron (MTT) assay, and cell cycle and apoptosis were analyzed by using flow cytometry. Results Compared with NB,TSB significantly promote the proliferation of human cholangiocarcinoma QBC939 cells (P

An experimental study on inhibition of apoptosis by HBx to human hepatic carcinoma cells in a nude mouse model / 中国普通外科杂志

Objective To investigate the effects of HBx on proliferation and apoptosis of human hepatocellular carcinoma cell line HepGz in vitro and in vivo. Methods The expression plasmids of pHA-HBx encoding full length of HBx gene was transfected into HepG2 cells with Lipofectamine 2000. The proliferation activity of transformed cells was measured by calculating the growth curve,population doubling time and by the 3H-TdR incorporation rate assay. The nude mouse model of HepG2 expressing HBX gene was established and the apoptosis rate of tumor cells was detected with TUNEL assay. Results RT-PCR indicated that the HBx gene was integrated into the genome DNA of HepG2 cells and transcripted. The growth curve and population doubling time manifested that the proliferation activity of transformed cells was much higher than that of control group cells. The apoptosis rate of tumor cells expressing HBx gene was much lower than that of control group cells. ConclusionHBx facilitates the proliferation activity of hepatoma cells in vitro and in vivo and inhibits the apoptosis of hepatoma cells induced by adriamycin in nude mice.

Effect of octreotide on pancreatic cancer cells transfected with SST 2 gene / 中国普通外科杂志

Objective To observe the effect of octreotide on human pancreatic cancer cells (PC-3) apoptosis after PC-3 transfected with somatostatin receptor type 2 (SST 2) gene. Methods SST 2 was transfected into PC-3 by liposome,the result of transfection was detected by immunohistochemistry. The apoptosis of PC-3 induced by using different dosage of octreotide were detected by MTT assay and flow cytometry. Results The effects of killing PC-3 by different dosage (0.2,0.4 and 0.8?g/ml ) of octreotide in transfected groups were significantly stronger than those in non-transfected groups(P