Octarepeat peptides of prion are essential for multidrug resistance in gastric cancer cells.

OBJECTIVE: In previous studies cellular prion protein (PrPc) is confirmed to be involved in multidrug resistance (MDR) of gastric cancer. Although octarepeat peptides are important functional domains of PrPc and are closely related to the transport of Cu2+/Zn2+ and antioxidative function, the significance in MDR remains unknown. We aimed to investigate the role of octarepeat peptides in gastric cancer MDR. METHODS: Small interfering RNA (siRNA) against PrPc were transfected into adriamycin-resistant gastric cancer cell lines to inhibit the expression of wild type PrPc, and then constructs encoding PrPc without octarepeat peptides and PrPc without the fifth repeat peptide were transfected, respectively, to establish the cell models. In vitro drug sensitivity, cell apoptosis, measurement of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and glutathione (GSH), as well as changes in glutathione S-transferase (GST) were detected. RESULTS: In vitro drug sensitivity test showed that octarepeat peptides could modulate the drug resistance of gastric cancer cells, but the deletion of the fifth repeat peptide had no effect. Specifically, the anti-apoptotic capacity of gastric cancer cells decreased significantly when the octarepeat peptides of PrPc was absent. Moreover, the activities of total SOD, Cu2+/Zn2+-SOD, GSH-Px, GSH, and GST detected in different stressing periods revealed that cells lacking octarepeat peptides of PrPc exhibited weakened responses to stress. However, absence of the fifth repeat peptide did not exert any effect on stress response. CONCLUSION: The octarepeat peptides of prion is responsible for MDR in gastric cancer cells while the fifth repeat peptide is not.

Dynamic changes and surveillance function of prion protein expression in gastric cancer drug resistance.

AIM: To explore the dynamic changes of prion protein (PrPc) in the process of gastric cancer drug resistance and the role of PrPc expression in the prognosis of gastric cancer patients receiving chemotherapy. METHODS: A series of gastric cancer cell lines resistant to different concentrations of adriamycin was established, and the expression of PrPc, Bcl-2 and Bax was detected in these cells. Apoptosis was determined using Annexin V staining. Western blotting and immunohistochemistry were performed to detect the expression of PrPc in patients receiving chemotherapy and to explore the role of PrPc expression in predicting the chemosensitivity and the outcome of gastric cancer patients receiving chemotherapy. Follow-up was performed for 2 years. RESULTS: PrPc expression was increased with the increase in drug resistance. Bcl-2, together with PrPc, increased the level of anti-apoptosis of cancer cells. Increased PrPc expression predicted the enhanced level of anti-apoptosis and resistance to anticancer drugs. PrPc expression could be used as a marker for predicting the efficacy of chemotherapy and the prognosis of gastric cancer. Increased PrPc expression predicted both poor chemosensitivity and a low 2-year survival rate. Contrarily, low PrPc expression predicted favorable chemosensitivity and a relatively high 2-year survival rate. CONCLUSION: PrPc expression is associated with histological types and differentiation of gastric cancer cells; The PrPc expression level might be a valuable marker in predicting the efficacy of chemotherapy and the prognosis of gastric cancer patients receiving chemotherapy.

[Effect of a hypothetical gene Af116609 on multi-drug resistance of gastric cancer cells].

OBJECTIVE: To investigate the effect of gene Af116609 on gastric cancer multi-drug resistance (MDR) by introducing it into gastric cancer multi-drug resistant (MDR) cell line SGC7901/VCR. METHODS: Gene Af116609 was cloned from SGC7901/VCR by RT-PCR and its differential expression between gastric cancer MDR cells and its parental cells was displayed by Northern blot. The gene was introduced to gastric cancer cells by transfection of recombinant eukaryotic expression vector by electroporation. MTT assay in vitro was applied to investigate its effect on multi-drug resistance phenotype of gastric cancer cells. RESULTS: The full length CDS of gene Af116609, as long as 327 bp, was cloned from gastric cancer MDR cell line SGC7901/VCR and its sequence was coincident with the hypothetical gene Af116609 in GenBank. It was overexpressed in MDR cells than its parental cells at mRNA level. In the MTT assay in vitro, the drug sensitive cells transfected with sense eukaryotic expression vector showed upregulated targeted gene, with increased resistance to vincristine, 5-fliorouracil and arabinoside, and decreased resistance to adriamycin, but no influence on resistance to methotrexate. However, the drug resistant cells transfected with anti-sense eukaryotic expression vector, showed down regulated targeted gene, with less resistance to all the five anticancer drugs to different degrees. CONCLUSION: Gene Af116609 is related to MDR phenotype of gastric cancer cells and may become a candidate molecular target to reverse the MDR of gastric cancer.

[The expression and possible function of RhoA in human gastric cancer cell lines].

OBJECTIVE: To study the expression and possible function of RhoA in human gastric cancer cell lines. METHODS: The expression of RhoA in human gastrointestinal cancer cell lines was detected by Western blot. Antisense plasmid of RhoA was constructed by pGEFL and transferred into gastric cancer cell line AGS by lipofectamine. Cell survival was examined by MTT assays, and cell cycle was detected by flow cytometry. RESULTS: The expression of RhoA protein in 10 different kinds of human cancer cell lines was much higher than that in immortalized human intestinal epithelial cell line. After being transfected with antisense RhoA, with the decrease in RhoA protein expression, the growth rate of AGS was inhibited, and the number of cells in S phase was increased by 14%. CONCLUSION: RhoA is overexpressed in many human cancer cell lines. Some of the malignant characteristics of a gastric cancer cell line can be partially reversed by inhibiting RhoA expression.

[The overexpression and significance of MKP-1 in oxygen-deprived gastric cancer cell line SGC7901].

OBJECTIVE: To investigate the expression status of Mitogen-activated Protein Kinase Phosphatase-1 in oxygen-deprived gastric cancer cell line SGC7901 and its role in HIF-1 regulation. METHODS: The expression of MKP-1 in gastric cell line SGC7901 was detected with Western blot and semiquantity RT-PCR; Eukaryotic sense expression vector was constructed based on DNA recombination technology. Transfections of SGC7901 were performed using liposome; The luciferase activity was determined using Dual Luciferase Reporter System and the levels of VEGF in SGC7901cells under normoxia and hypoxia were measured by ELISA. RESULTS: (1) Semiquantity RT-PCR and Western blot suggested that the expression of MKP-1 was elevated in oxygen-deprived gastric cancer cell line SGC7901; (2) 48 hours after transfection, the phosphorylated form of HIF-1alpha in cell line transfected with recombinant plasmids was lower compared with that in cell line transfected with empty vectors after 12 hours of exposure to hypoxia; (3) There was very low luciferase activity under nomoxia while under hypoxia luciferase activity increased in a time-dependent manner and at all time points there was significant lower luciferase activity in SGC7901 cells than in cells transfected with empty plasmids. (4) At different points of time course, the expression of VEGF in SGC7901 was significantly higher under hypoxia than that in SGC7901 under nomorxia, while under hypoxia, the expression of VEGF in SGC7901 transfected with recombinant plasmids was significantly lower than that in SGC7901 transfected with empty vectors at all time points as indicated. CONCLUSION: The expression of MKP-1 in SGC7901 was elevated under hypoxia, which could downregulate the HIF-1 trans-activition activity thereby repressing the expression of downstream target gene VEGF.

[Clinical application of ventriculoperitoneal shunt assisted by laparoscope].

OBJECTIVE: To compare the clinical effectiveness of ventriculoperitoneal shunt (VPS) assisted by laparoscope versus the clinical effectiveness of VPS via traditional laparotomy in treating hydrocephalus. METHODS: The clinical data on 26 cases of VPS with laparoscope assistance and on 234 cases of VPS via traditional laparotomy were summerized and analyzed retrospectively. The operation efficacy and the complication rate were compared between the two groups. RESULTS: In the traditional laparotomy group, there were 6 cases of peritoneal catheter obstruction or displacement, 5 cases of postoperative infection, and 2 childhood cases where the peritoneal catheter made a way into the rectum and through the anus. But in the laparoscope group, there was only one case of postoperative infection. Postoperatively the laparoscope group was better than the traditional laparotomy group in respect to the postoperative outcome, complication rate and the clinical improvement. Thirteen cases were treated by the VPS using right liver-diaphragm interspace drainage and another 13 cases were treated by the VPS using pelvis cavity drainage in the laparoscope group, respectively. CONCLUSION: The technique of VPS assisted by laparoscopies safe, reliable, and has the advantages of accurate positioning, minimal injury and less postoperative complications; it is worthy of clinical application and popularization. Positioning the peritoneal catheter to the liver-diaphragm interspace can avoid the complications caused by perforation of the cavity organs.

[Protective effect of selective cyclooxygenase-2 inhibitor on alcohol-induced liver injury in rats].

OBJECTIVES: To investigate the effect of selective cyclooxygenase-2 (COX-2) inhibitor on alcohol-induced liver injury in rats. METHODS: 58 male Wistar rats were randomly divided into three groups: control group treated with dextrose and corn oil, model group with ethanol and corn oil, treatment group with corn oil and ethanol plus a selective COX-2 inhibitor celecoxib. All treatments were injected into stomach through intragastric tubes. Liver samples were analyzed for histopathology with light microscope (LM) and transmission electron microscope (TEM), and the expression of COX-2 with western blotting. Levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in serum, levels of 6-Keto-prostaglandin F1 alpha (6-k-PGF1a) and thromboxane B2 (TXB2) in liver, and activity of glutathione s-transferase (GST) both in liver tissue and in plasma were measured. RESULTS: LM and TEM indicated hepatocytes were injured obviously in the model group and slightly in the treatment group. The levels of AST and ALT in serum, TXB2 in liver and the activity of GST in plasma increased significantly in the model group (t> or =2.294, P<0.05), but the activity of GST in liver decreased significantly (t=8.856, P<0.01) compared with those in the control group. To compare with the model group, the levels of AST and TXB2 decreased significantly (t=4.305, P<0.01; t=2.799, P<0.01), meanwhile the activity of GST increased significantly (t=10.134, P<0.01) in the treatment group. COX-2 expression in liver by western blotting increased significantly in the model group, compared with the control group (t=4.067, P<0.01) and the treatment group (t=2.251, P<0.05). Exceptionally, the level of 6-k-PGF1a decreased significantly (t=2.284, P<0.05) in the model group. CONCLUSION: COX-2 has involved in the alcohol-induced liver injury, and its inhibitor can diminish alcohol-induced liver injury in rats through decreasing TXB2 level

[Rac subfamily expression and activity in gastrointestinal cancer cell lines].

OBJECTIVE: To investigate the significance of Rac subfamily members in the gastrointestinal carcinogenesis and progression. METHODS: The mRNA expression of Rac1, Rac2 and Rac3 in 12 kinds of gastrointestinal cancer cell lines was examined by semi-quantitative RT-PCR. The activities of Rac1 protein in 5 kinds of gastric cancer cell lines were tested by pull-down assay. RESULTS: Compared with the normal gastric mucosa and intestinal epithelial cell line, the mRNA expression of Rac1 and Rac3 was up-regulated in most of gastrointestinal cancer cell lines. The activities of Rac1 protein increased markedly in gastric cancer cell lines. CONCLUSION: The increased mRNA expression of Rac1 and Rac3 in gastrointestinal cancer cell lines and the abnormal activation of Rac1 protein in gastric cancer cell lines might be correlated with the carcinogenesis of gastrointestinal cancer.

[The overexpression of prion protein in drug resistant gastric cancer cell line SGC7901/ADR and its significance].

OBJECTIVE: To investigate the overexpression of prion protein (PrP) in drug-resistant gastric cancer cell line SGC7901/ADR and its role in multidrug resistance in gastric cancer. METHODS: (1) The expression of PrP in SGC7901/ADR, SGC790/VCR and their parental cell line SGC7901 was detected with Northern and Western blot at the mRNA and protein level. (2) Eukaryotic sense and antisense expression vector were constructed based on DNA recombination technology and (3) introduced into SGC7901 and SGC7901/ADR cell lines through electroporation. (4) The accumulation and retention of ADR in transiently transfected cells were detected by flow cytometry. RESULTS: (1) Northern and western blot suggested significantly higher expression of PrP in SGC7901/ADR and SGC7901/VCR than that in SGC7901. (2) 48 hours after the vectors transfection, the average fluorescence intensity of Adr in transfected cells were detected. The accumulation intensity were 8.9 +/- 0.7 in BS, 6.6 +/- 0.3 in PS and 7.5 +/- 0.6 in PA. The retention intensity were 9.3 +/- 0.6 in SGC7901, 5.9 +/- 0.5 in PS and 7.1 +/- 0.5 in PA. There were significant difference between PS and BS with P < 0.01, as well as RA and BA with P < 0.01. These data suggested that PrP gene could affect the drug accumulation in gastric cancer cells after its transfected into cells. CONCLUSION: PrP was highly expressed in gastric cancer cell lines SGC7901/ADR and SGC7901/VCR. Overexpression of PrP had certain effect on drug accumulation in gastric cancer cells.

[Differential expression of RPL6/Taxreb107 in drug resistant gastric cancer cell line SGC7901/ADR and its correlation with multiple-drug resistance].

OBJECTIVE: To investigate the differential expression of RPL6/Taxreb107 between drug-resistant gastric cancer cell line SGC7901/ADR and gastric cancer cell line SGC7901 as well as its correlation with multiple-drug resistance (MDR) in gastric cancer cells. METHODS: Total RNA was extracted from SGC7901 and SGC77901/ADR, with internal control RT-PCR, Northern blot, gene cloning and expression, construction of eukaryotic expression vector, gene transfection by electroporation. The accumulation and retention of ADR in transiently transfected cell was detected by flow cytometry. RESULTS: The internal control RT-PCR and Northern blot showed high RPL6/Taxreb107 expression in SGC7901/ADR cell line. Sense and antisense eukaryonic expression vectors demonstrated by double enzyme digestion were successfully transfected into gastric cancer cell line SGC7901 and SGC7901/ADR respectively by electroporation. The accumulation and retention of ADR detected 48 hours after transfection showed that RPL6 gene had shown effect on drug resistance in gastric cancer cell. CONCLUSION: The high expression of RPL6/Taxreb107 in drug resistant gastric cancer cell shows its correlation with multiple-drug resistance in gastric cancer.

[Effects of different temperature and time on the period of validity and quality in blood preservation].

AIM: To examine the effects of different temperature protection on measures on preservation damages in liquid blood and explore the corresponding. METHODS: Take equal half blood samples from 10 healthy blood donors and divided each sample into two groups, put the fresh blood into CP2D-A solution at 0 degrees C and 4 degrees C, respectively and take the samples 21 days and 42 days, later and then measured the contents of membrane phospholipids with shafig-UR-rehman method, CaM with purification PED test, LPO with spectrophotometry. RESULTS: At the same temperature, when the preservation time was prolonged, peroxidation was increased, the preservation damages were also augmented; the damages were declined when the temperature was lower during the same period, the aging of blood was more evident at 4 degrees C. CONCLUSION: Blood peroxidation temperature is lower. The author pointed out the questions and prospects of blood preservation.

[Effects of different temperature and time on the quality of blood preservation in liquid blood].

AIM: To explore the effects of different temperature and time on preservation damages in liquid blood. METHODS: Take blood sample from 10 healthy blood donors, put the fresh blood into CP2D-A liquid at 0 degrees C and 4 degrees C, and take the samples after 1 week, 2 weeks and 3 weeks, and then measured the contents of GSH-Px, TSH, LPO, the contracting protein of RBC, and membrane fluidity. RESULTS: At the same temperature, when the preservation time is prolonged, peroxidation is increased, the preservation damages are also augmented; the damages are declined when the temperature is lower during the same period, the aging of blood was more evident at 4 degrees C. CONCLUSION: Blood peroxidation temperature is lower. The 0 degrees C group is better than 4 degrees C group.

Effect of apolipoprotein E gene Hha I restricting fragment length polymorphism on serum lipids in cholecystolithiasis.

AIM:To investigate the role of apolipoprotein E (apoE) polymorphism in the lithogenesis of gallstone and the hereditary pathogenesis of the disease.METHODS: Polymerase chain reaction (PCR) was used to study apoE phenotypes and allele frequencies in patients with gallstones and control, and the fasting serum lipids of subjects were also measured by enzymatic methods.RESULTS:The levels of triglyceride (TG) and very low density lipoprotein cholesterol (VLDL-C) were much higher in E(2/3) patients than that in E(2/3) control. E(3/3) patients were accompanied with remarkably low levels of high density lipoprotein cholesterol (HDL-C) and its subforms.But in E(3/4) patients there were only slight changes in levels of VLDL-C and low density lipoprotein cholesterol (LDL-C).CONCLUSION:Different apoE phenotype patients with gallstones have different cheracteristics of dyslipidemia and the average level of serum lipids in patients with gallstones are higher than subjects without gallstones in the same apoE gene phenotype.epsilon2 allele is possibly one of the dangerous factors in the lithogenesis of cholecystolithiasis.