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Objective To investigate the effects of Pulsatilla saponin D and sorafenib on the metastasis of human hepa?toma cell line. Methods The human hepatoma cell line BEL-7402 cells were divided into Pulsatilla saponin D group (con?centration of 11.9 mg/L), sorafenib group (concentration of 2.15μmol/L), the combined group (Pulsatilla saponin D 11.9 mg/L+Sorafenib 2.15μmol/L) and the control group (ordinary broth). The inhibition effects of Pulsatilla saponin D and sorafenib monotherapy and combination therapy on BEL-7402 cell migration were detected by MTT assay, Transwell chamber experi?ment and cell scratch experiment. Western blot assay was used to detect the expression levels of matrix metalloproteinase (MMP)-2 and MMP-9 gene protein. Results MTT assay showed that Pulsatilla saponin D (11.9 mg/L), sorafenib (2.15μmol/L) monotherapy and combination therapy had inhibitory effects on BEL-7402 cell proliferation, and the 24-h inhibi?tion rate was<15%. Results of Transwell chamber experiment and cell scratch test showed that the migration inhibitory rate was significantly higher in combination group than that of monotherapy group (P<0.01). The combined effect of madicine was the addition (0.85≤Q≤1.15). Western blot detection showed that there was a higher effect of down-regulation on MMP-2 and MMP-9 in combined group than that of monotherapy group. Conclusion Pulsatilla saponin D and sorafenib synergis?tically inhibit the metastasis of BEL-7402 cells. The joint effects are superior to monotherapy.
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Objective We examined the Grp78, ERK1/2 and phospho-ERK1/2 expressions in hepatocellular carcinoma(HCC) tissue samples in vitro, we interfered the expression of Grp78 in SMMC-7721 cells to explore whether Grp78 is involved in ERK1/2 signal pathway. Methods The Grp78, ERK1/2 and phospho-ERK1/2 expressions were detected by immunohistochemistry and confirmed by Western blotting in 47 HCC tissue samples. The Grp78 expression in SMMC-7721 cells was interfered by plasmid transfection and siRNA, ERK1/2 phosphorylation and expression were determined by Western blotting. Results The Grp78 expression was significantly correlated with ERK1/2 and phospho-ERK1/2 in HCC tissue samples. Overexpression of Grp78 promoted ERK1/2 phosphorylation in SMMC-7721 cells and the increased ERK1/2 phosphorylation was inhibited by Grp78 knockdown. Conclusion Grp78 is involved in the regulation of ERK1/2 signal pathway and might be a potential target for the comprehensive therapy of HCC.
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BACKGROUND: It has been confirmed that some casting alloy can inhibit the activity of human gingival fibroblasts in vitro, which can be used for inducing apoptosis of human gingival fibroblasts (HGFs). However, the mechanism of this effect is poorly understood.OBJECTIVE: To investigate the effect of Ni-Cr, Co-Cr, Au and pure Ti ceramic alloys on the expression of P70S6k by HGFs in vitro.DESIGN, TIME AND SETTING: Grouping controlled experiments with the expression of P70s6k by HGFs in each group as observation objects. Experiments were performed at the Institute of Immunohistochemistry and Western blotting, Technology Building of Liaoning Medical University from July to October 2009.MATERIALS: Ni-Cr, Co-Cr, Au and pure Ti ceramic alloys were processed by Shenzhen Mecodent Dental Laboratory Co., Ltd. The gingival from gingival neck of the first premolar (which goes to be extracted due to the need of orthodontic treatment) of teenager who was15 years old without any clinical disease was obtained, followed by the primary culture of HGFs. The fifth passage of cells in logarithmic phase were used to measure indexes.METHODS: Ni-Cr, Co-Cr, Au and pure Ti ceramic alloys were incubated in DMEM to prepare alloy leaching liquor, which were then added in HGF medium at 10% concentration. DMEM containing 10% fetal bovine serum was used in negative controls. MAIN OUTCOME MEASURES: The expression of P70S6k in each group was measured by Western Blotting following 72 hours of intervention with alloy leaching liquor.RESULTS: Western-blot results showed that there was no significant difference in the average gray value of P70S6k expression in Au, pure Ti ceramic alloys and control groups (P > 0.05). There was also no significant difference between Au ceramic alloys and pure Ti ceramic alloys groups or between Ni-Cr ceramic alloys and Co-Cr ceramic alloys groups. (P > 0.05), but Ni-Cr ceramic alloys group or Co-Cr ceramic alloys group showed significant difference compared with the Au, pure Ti ceramic alloys and control groups (P < 0.01).CONCLUSION: Au ceramic alloys and pure Ti ceramic alloys show that they have no obvious effect on the proliferation activity of HGFs. Ni-Cr and Co-Cr ceramic alloys showed an inhibitory effect on the proliferation activity of HGFs.
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BACKGROUND: Polymorphism in promoter region can change the expression of genes, which may be associated with susceptivity of diseases. Gene polymorphism of interleukin-6 (IL-6) promoter is associated with nationality and many diseases. Different nationalities often display different characteristics of gene polymorphism.OBJECTIVE: To observe the distribution of polymorphism at sites -597 and -572 in IL-6 promoter region in Tibet Tibetan population and to provide the theoretical data for Tibetan population genetics and background of immunity.DESIGN: Randomized investigation.SETTING: Institute of Anthropology, Jinzhou Medical College.PARTICIPANTS: Totally 108 healthy Tibetan teenagers were selected from Lasa and Naqu region in Tibet autonomous region from October 2003 to July 2004, including 60 males and 48 females, aged from 14-21 years. Inclusive criteria:The parents of the volunteers were healthy Tibetans after body examination. The volunteers knew the fact, agreed to participate into the trail and signed the informed consent.METHODS: 5 mL peripheral vein blood was collected from 108 Tibetan teenagers. DNA from human leucocytes was extracted by salt fractionation. IL-6 promoter including -597 and -572 fragments was amplified by polymerase chain reaction (PCR). Representative fragments were cloned then sequenced after restriction fragment length polymorphism (RFLP).MAIN OUTCOME MEASURES: Distribution of polymorphism in Tibet Tibetan population; Results after comparison with those of other nationalities including Han population.RESULTS: Data of 108 Tibetan teenagers were involved in the result analysis. ①Distribution of polymorphism on -572C/G site of IL-6 promoters in population of either sex: There were no GA and AA genotypes at site -597, but only GG genotype appeared. There were CC, CG and GG genotypes at site -572, and the frequencies were 0.63, 0.35 and 0.02 in order. Distribution of genotype with representativeness met the Hardy-Weinberg equilibrium. Allele frequencies were 0.81 and 0.19, respectively. There was no significant difference of either sex in genotype and allele frequency (P ＞ 0.05). ②Distribution of polymorphism on -597 G/A and -572 C/G in different nationalities: GG, GA, AA genotypes appeared on -597 site in England and France, and G and A allele frequencies were 0.60 and 0.40,respectively. It was significantly different from that of Tibetan in Tibet. Furthermore, Japanese had no polymorphism,which was similar to that of Hans in China (P ＞ 0.05). ③Genotype of different straps and results of DNA sequencing:Only GG genotype was found on -597 site (without the restriction site, one fragment after restriction, PCR amplification products), no GA and AA genotypes. CC, CG and GG genotypes appeared at site -572, and frequencies were 0.64,0.35 and 0.01, respectively. Distribution of genotype with representativeness met the Hardy-Weinberg equilibrium (P ＞0.05). Allele frequencies were 0.81 and 0.19, respectively. There was no significant difference of either sex in genotype and allele frequency. Distribution of gene frequency and allele frequency in IL-6 were similar between Tibetan and that of Hans, but it was significantly different from that of population in England, France and America.CONCLUSION: There are nationality differences of IL-6 gene polymorphism at sites -597 and -572. No polymorphism is found at site -597 in Tibetan. Race differences are seen at site -572, having CC, CG and GG genotypes and G allele is rate. Compared with white population, there is significant difference in genotype and allele frequency at site -572. Their characteristics are close to Hah population and Japanese, which may be associated with genetic gene of persons living in plateau.