高表达蛋白hSav1对人胚肾HEK293增殖能力的影响

Objective To elucidate the effects of human Salvador 1 (hSav1 ) on cell proliferation of human embryonic kidney cell line HEK293. Methods The plasmid CFP-N1-hSav1 was constructed and transfected into HEK293 cells with lipofectamine 2000. The transfection efficiency was detected by fluorescent microscopy. The effects of hSav1 on cell proliferation were measured by MTT and BrdU incorporation. Results The transfection efficiency was about 70% -80%. The MTT results showed that the inhibition rate of cell proliferation in transfected group at the 12th, 24th, 36th, and 48th h was 2% , 5% , 15% , and 23% , respectively;while that in the control group was 2% , 3% , 2% , and 2% respectively. The BrdU incorporation revealed that the BrdU incorporation rate in transfected group (12. 9 ±5. 3)% was significantly lower than in control group (27.3±3.8)% (P<0.05). Conclusion hSav1 is a newly identified protein that can cause cell proliferation inhibition in HEK293 cells.     

La核糖核蛋白6对人血管内皮细胞增殖与凋亡的作用

Objective To investigate regulatory effect of Acheron (Achn) on proliferation and apoptosis of human vascular endothelial cell. Methods ( 1 ) Eahy926 cells were cultured in serum-free DMEM medium (96-well plates) and were divided into Achn inhibition group (transfected with plasmid psi-Achn), psi4.1 group (transfected with psi4. 1 empty vector), Achn induction group (transfected with pcDNA-Achn), pcDNA3.1 group (transfected with pcDNA3.1 empty vector), cotransfection group [cotransfected with pcDNA-Achn + psi-calcium/calmodulin-dependent serine protein kinase (CASK)] , blank control group (treated with PBS) according to the random number table (the same method below). The cell proliferation was determined by MTT assay at post transfection hour (PTH) 1, 24, 48, 72, with expression of absorbance value. (2) Total protein of Eahy926 cells were extracted and quantitated by BCA assay, and then they were divided into Achn antibody precipitation group (100 μg protein) , CASK antibody precipitation group ( 100 μg protein), IgG antibody group ( 100 μg protein), Western blot group (20 μg protein).Achn and CASK protein levels were determined by immunoprecipitation and Western blot. (3) Synchronously cultured Eahy926 cells were divided into LPS induction group (treated with 5 mol/L LPS), Achn transfection group (transfected with pcDNA-Achn), cotransfection group (cotransfected with psi-CASK and pcDNA-Achn) , KCl group (treated with 5 mol/L KCl), and blank control group (treated with 5 mol/LPBS). Cells in transfection groups were stimulated by LPS for 12 hours after PTH 24. Caspase-3 protein level was detected by immunohistochemistry. (4) Synchronously cultured Eahy926 cells were divided into Achn inhibition group (transfected with psi-Achn vector), Achn induction group ( transfected with pcDNA-Achn vector), and blank control group ( treated with PBS). Apoptosis rate was determined by FITC/PI with flow cytometry. Data were processed with one-way analysis of variance and t test. Results ( 1 ) The cell proliferation in Achn inhibition group was lower than that in psi4.1 group from PTH 24, and the differences were statistically significant at PTH 48, 72 (with t value respectively 10. 777, 6.112, P values all below 0. 05 ).The cell proliferation in Achn induction group during PTH 24-72 were higher that in pcDNA3. 1 group (with t value respectively 5. 367, 6. 053, 9. 831, P values all below 0.05 ). The cell proliferation in cotransfection group at PTH 48, 72 were significantly lower than that in Achn induction group ( with t value respectively 5.481, 9. 517, P values all below 0. 05). (2) Achn protein was detected in CASK antibody precipitation group while CASK protein was also detected in Achn antibody precipitation group. (3) Caspase-3 level in Achn transfection group was lower [( 15.6 ± 0. 5 ) %] as compared with that in LPS induction group [(32. 8 ±2.6)%, t = 10. 083, P ＜ 0. 05], and that in cotransfection group showed further inhibition [(7.0 ±2.0)%,t =9.827, P ＜0.01]. (4) Apoptosis rate in Achn inhibition group[(45.6 ± 10.9)%] was higher than that in blank control group [(13.2±4.3) %, t =7.043, P ＜0.05]; while that in Achn inductiongroup [(5.3 ±2.9)%] was lower than that in blank control group ( t =6.499, P ＜0.05).Conclusions Achn can promote human vascular endothelial cell proliferation, and inhibit its apoptosis induced by LPS or burn serum, and the effect is related to CASK.     

La核糖核蛋白6对人血管内皮细胞增殖与凋亡的作用

Objective To investigate regulatory effect of Acheron (Achn) on proliferation and apoptosis of human vascular endothelial cell. Methods ( 1 ) Eahy926 cells were cultured in serum-free DMEM medium (96-well plates) and were divided into Achn inhibition group (transfected with plasmid psi-Achn), psi4.1 group (transfected with psi4. 1 empty vector), Achn induction group (transfected with pcDNA-Achn), pcDNA3.1 group (transfected with pcDNA3.1 empty vector), cotransfection group [cotransfected with pcDNA-Achn + psi-calcium/calmodulin-dependent serine protein kinase (CASK)] , blank control group (treated with PBS) according to the random number table (the same method below). The cell proliferation was determined by MTT assay at post transfection hour (PTH) 1, 24, 48, 72, with expression of absorbance value. (2) Total protein of Eahy926 cells were extracted and quantitated by BCA assay, and then they were divided into Achn antibody precipitation group (100 μg protein) , CASK antibody precipitation group ( 100 μg protein), IgG antibody group ( 100 μg protein), Western blot group (20 μg protein).Achn and CASK protein levels were determined by immunoprecipitation and Western blot. (3) Synchronously cultured Eahy926 cells were divided into LPS induction group (treated with 5 mol/L LPS), Achn transfection group (transfected with pcDNA-Achn), cotransfection group (cotransfected with psi-CASK and pcDNA-Achn) , KCl group (treated with 5 mol/L KCl), and blank control group (treated with 5 mol/LPBS). Cells in transfection groups were stimulated by LPS for 12 hours after PTH 24. Caspase-3 protein level was detected by immunohistochemistry. (4) Synchronously cultured Eahy926 cells were divided into Achn inhibition group (transfected with psi-Achn vector), Achn induction group ( transfected with pcDNA-Achn vector), and blank control group ( treated with PBS). Apoptosis rate was determined by FITC/PI with flow cytometry. Data were processed with one-way analysis of variance and t test. Results ( 1 ) The cell proliferation in Achn inhibition group was lower than that in psi4.1 group from PTH 24, and the differences were statistically significant at PTH 48, 72 (with t value respectively 10. 777, 6.112, P values all below 0. 05 ).The cell proliferation in Achn induction group during PTH 24-72 were higher that in pcDNA3. 1 group (with t value respectively 5. 367, 6. 053, 9. 831, P values all below 0.05 ). The cell proliferation in cotransfection group at PTH 48, 72 were significantly lower than that in Achn induction group ( with t value respectively 5.481, 9. 517, P values all below 0. 05). (2) Achn protein was detected in CASK antibody precipitation group while CASK protein was also detected in Achn antibody precipitation group. (3) Caspase-3 level in Achn transfection group was lower [( 15.6 ± 0. 5 ) %] as compared with that in LPS induction group [(32. 8 ±2.6)%, t = 10. 083, P ＜ 0. 05], and that in cotransfection group showed further inhibition [(7.0 ±2.0)%,t =9.827, P ＜0.01]. (4) Apoptosis rate in Achn inhibition group[(45.6 ± 10.9)%] was higher than that in blank control group [(13.2±4.3) %, t =7.043, P ＜0.05]; while that in Achn inductiongroup [(5.3 ±2.9)%] was lower than that in blank control group ( t =6.499, P ＜0.05).Conclusions Achn can promote human vascular endothelial cell proliferation, and inhibit its apoptosis induced by LPS or burn serum, and the effect is related to CASK.     

干扰Livin基因对人胆管癌QBC939细胞增殖影响的研究

目的 研究转染Livin反义寡核苷酸(antisense oligodeoxynucleotide,ASODN)对人胆管癌细胞株QBC939细胞Livin mRNA及Livin蛋白表达以及细胞增殖的影响.方法 设计合成全硫代磷酸化修饰并5'端FITC荧光标记的Livin ASODN.用脂质体介导Livin ASODN转染QBC939细胞,荧光显微镜观察24 h Livin ASODN转染人细胞的情况,并计算转染率.MTT法检测Livin反义寡核苷酸对QBC939细胞增殖的抑制作用.RT-PCR和细胞免疫荧光化学分别检测转染后48 h Livin mRNA表达及Livin蛋白的变化.结果 500 mol/L ASODN转染后24 h可达到最佳的转染效果;转染后60 h,能明显抑制胆管癌细胞的增殖.转染60 h后,RT-PCR及细胞免疫荧光化学检测分别显示Livin mRNA及Livin蛋白表达水平明显低于对照组(P<0.05).结论 脂质体介导转染Livin ASODN能特异性地抑制QBC939细胞中的Livin基因及Livin蛋白表达,抑制胆管癌QBC939细胞的增殖,降低癌细胞活力.

Abstract：

Objective To study the effect of transfection of livin antisense oligodeoxynucleotide (Livin ASODN) on Livin mRNA and Livin protein expression and proliferation of QBC939 cells.Methods Livin ASODN was transfected into cell line QBC939 by LipofectamineTM 2000. Fluorescence microscopy was used to observe the ASODN transfected cells and to calculate the rate of transfection.to measure Livin mRNA and Livin protein expression by RT-PCR and immunohistochemistry and con-focal laser scanning microscopy after the transfection. Changes in cell proliferation were detected by MTT. Results The highest efficiency was at 24 hours after 500 nmol/L Livin ASODN transfection.The results of MTT showed that the inhibition of cell proliferation of QBC939 cells was most obvious at 60 hours after Livin ASODN transfection (P<0. 05). The level of Livin mRNA and Livin protein expression in the ASODN group was obviously lower than that in the control group (P<0. 05).Conclusion The transfection of Livin ASODN inhibited Livin gene and Livin protein expression, and obviously inhibited the proliferation and depressed the vitality of QBC939 cells.     

RhoA基因沉默对肝癌HepG2细胞增殖和迁移能力的影响

Objective To construct a RhoA-siRNA expression vector and determine its role on the malig-nant behavior of HepG2 cells.Methods A RhoA-siRNA DNA fragment was synthesized and cloned into the expression vector of pGenesil-1.The constructed Rhon-siRNA DNA plasmid was stably transfected into HerG2 cells by lipofectamine,and then HepG2 cells were divided into the HepG2/RhoA-siRNA group (HepG2 cells were transfected with pGenesil-1-RhoA-siRNA),HepG2/control group(HepG2 cells were transfected with control plasmid) and HepG2 group (without plasmid transfection).The inbibitory effect of RhoA-siRNA on RhoA protein expression was shown by Western blot.The proliferation,migration,growth potentiality and cell cycle of transfected HepG2 cells were evaluated by MTT assay,wounded healing,the plate cloning formation test and flow cytometry,respectively.All data were analyzed by one-way analysis of variance (ANOVA) and chi-square test.Results The expression of RhoA protein in the HepG2/RhoA-siRNA group was,significantly decreased compared with that in the other two groups (F=178.19,P<0.05).Scratched cells were healed within 48 hours in the HepG2/control group and HepG2 group,but not in the HepG2/RhoA-siRNA group.The clone formation rates in the HepG2/RhoA-siRNA group,HepG2 group and HepG2/control group were 39%±3%,67%±5%and 70%±6%,respectively,with a significant difference among the three groups(χ2=33.34,38.69,P<0.05).Flow cytometry showed that the number of cells transfected with RhoA-siRNA was highest in the G0/G1 phase and lowest in the S phase(F=70.46,76.57.P<0.05).Conclusion The RhoA-siRNA expression vector can effectively suppress the proliferation and migration of HepG2 cells,which may provide a novel gene therapy for hepatocellular carcinoma.     

HCCR-2反义核酸对肝癌HepG2细胞的作用

Objective To investigate the effects of antisense recombinant euraryotic expression vector of HCCR-2 on the proliferation and apoptosis of HepG2. Methods The antisense recombinant eukaryotic expression vector of HCCR-2 was constructed. The vector was stably transfected to the HepG2 cells, and positive clones were selected by G418 (antiseuse vector group), pIRES2-EGFP vector was transfected into the HepG2 cells in the same way (pIRES2-EGFP group). The conditions of the nontransfected HepG2 cells were used as control (HepG2 group). Changes in cell growth curve, cell cycle, cell apoptosis and morphology of HepG2 cells after the transfec-tion were detected by MTT method, flow cytometry and transmission electron microscopy, respectively. All the data were analyzed by one-way ANOVA and chi-square test. Results The expression level of HCCR-2 mRNA was down-regulated to 0.39±0.04 in antisense vector group, and the expression level of HCCR-2 mRNA in pIRES2-EGFP group and HepG2 group were 0.62±0.06 and 0.72±0.03, respectively, with significant difference among the 3 groups (F=43.701, P<0.05). The apoptotic rate of HepG2 cells in antisense vector group, pIRES2-EGFP grop and HepG2 group were 13.30%, 2.51% and 2.07%, respectively, with significant difference among the 3 group (χ2=6.793, 8.721, P<0.05). The growth of HepG2 cells in antisense vector group was retarded, and was blocked in G0/G1 stage. Conclusions The HCCR-2 antisense recombinant eukaryotic expression vector can inhibit the mRNA expression of HCCR-2 and promote the apoptosis of cells. HCCR-2 may be involved in cell regulation and the proliferation of hepatocellular carcinoma cells.     

大鼠肾上腺细胞经胶原凝胶三维培养后进行同种移植的实验研究

Objective To investigate a novel method of adrenocotical cells transplantation. Methods Adrenal glands of neonate rats were dissociated into adrenal cortical cells. Cells were cultured in self-made collagen type Ⅰ gel for one week, and then transplanted under the renal capsule of bilateral adrenalectomy mate rats. Blood samples were collected per week after surgery. Animals were sacrificed at the 8th week, and histological characteristics of the allografts, proliferation of transplanted cells, CYP11B1 and CYP11B2 expression as well as plasma aldosterone and corticosterone were observed. Results There were totally 15 rats receiving collagen gel transplantation, and 12 survived 8 weeks post-operation. The plasma cortocosterone level in collagen gel transplantation group was significantly higher than in adrenocortical cell transplantation group, and reached the normal level from 6th to 8th week, but the change in plasma aldosterone level in collagen gel transplantation group was similar to that of adrenocortical cell transplantation group. The adrenocortical cells cultured in gel grew well, and had a high proliferation rate. 95% of them were fasciculata cells which expressed CYP11B1, and the rest were glomerulosa cells which expressed CYP11B2. No inflammatory cell infiltration was observed. Conclusion The proliferation and function of adrenocortical cells could be promoted when they were cultured in collagen gel.     

ShRNA-mediated gene silencing of β-catenin inhibits growth of human colon cancer cell Colo205 in vitro

Objective To observe the gene silencing and disruption of WNT pathway mediated by the specific shRNA targeted against β-catenin and its effect on cell proliferation of the human colon cancer cell line Colo205. Methods The shRNA plasmid vectors against β-catenin were constructed and transfected into Colo205 ceils with LipofectamineTM 2000. The expression of β-catenin was detected by RT-PCR and Western blot. Immunofluorescence staining was also performed to detect the β-catenin protein expression in cells. The cell proliferation inhibition was determined by MTT assay and soft agar colony formation assay. Results The shRNA vectors targeted against β-catenin were successfully constructed and efficiently suppressed the expression of β-catenin mRNA and protein (P<0.05). The expression inhibition rates were 47.89% and 45.26% at the mRNA and protein level respectively. Immunofluorescence microscopy also demonstrated the inhibition of β-catenin protein induced by these specific shRNAs. The MTT assay indicated that the specific shRNA resulted in significant inhibition of cell growth on the culture plates in time-dependent manner. At 72 h post-transfection, the cell viability of CAT group was 48.5%, which was significantly different as compared with that of blank control group's 91.3%(P<0.05). In the soft agar, there were 9, 46, 43 cell colonies in the CAT, blank, and negative control groups respectively, which were significantly different (P<0.05). Conclusions The specific shRNAs targeted against β-catenin has a gene silencing effect and blocks the WNT signaling pathway, which can inhibit the growth of Colo205 cells.     

ShRNA-mediated gene silencing of β-catenin inhibits growth of human colon cancer cell Colo205 in vitro

Objective To observe the gene silencing and disruption of WNT pathway mediated by the specific shRNA targeted against β-catenin and its effect on cell proliferation of the human colon cancer cell line Colo205. Methods The shRNA plasmid vectors against β-catenin were constructed and transfected into Colo205 ceils with LipofectamineTM 2000. The expression of β-catenin was detected by RT-PCR and Western blot. Immunofluorescence staining was also performed to detect the β-catenin protein expression in cells. The cell proliferation inhibition was determined by MTT assay and soft agar colony formation assay. Results The shRNA vectors targeted against β-catenin were successfully constructed and efficiently suppressed the expression of β-catenin mRNA and protein (P<0.05). The expression inhibition rates were 47.89% and 45.26% at the mRNA and protein level respectively. Immunofluorescence microscopy also demonstrated the inhibition of β-catenin protein induced by these specific shRNAs. The MTT assay indicated that the specific shRNA resulted in significant inhibition of cell growth on the culture plates in time-dependent manner. At 72 h post-transfection, the cell viability of CAT group was 48.5%, which was significantly different as compared with that of blank control group's 91.3%(P<0.05). In the soft agar, there were 9, 46, 43 cell colonies in the CAT, blank, and negative control groups respectively, which were significantly different (P<0.05). Conclusions The specific shRNAs targeted against β-catenin has a gene silencing effect and blocks the WNT signaling pathway, which can inhibit the growth of Colo205 cells.     

高表达蛋白hSav1对人胚肾HEK293增殖能力的影响

目的 观察高表达人Salvadorl(hSav1)蛋白对人胚肾293(HEK293)细胞增殖的影响.方法 构建含有绿色荧光蛋白(GFP)的hSav1质粒GFP-N1-hSav1;将构建好的GFP-N1-hSav1质粒在脂质体Lipofectamine 2000的介导下转染HEK293细胞,荧光显微镜下观察质粒转染效率;分别在转染后12、24、36、48 h通过噻唑蓝(MTT)比色法计算细胞增殖抑制率,抑制率=[(对照-空白)-(给药-空白)]/(对照-空白)×100%;转染后36 h加入5-溴-2'-脱氧尿苷(BrdU),通过其掺入比率来显示hSav1对细胞增殖的影响.结果 质粒GFP-N1-hSav1构建成功,转染效率为70%-80%左右,蛋白hSav1明显高表达;MTT结果显示转染质粒后12、24、36、48 h,高表达蛋白hSav1组的细胞增殖抑制率分别为2%、5%、15%、23%,而阴性对照分别为2%、3%、2%、2%;BrdU结果显示,在HEK293细胞中高表达蛋白hSav1后细胞增殖抑制率为(27.3±3.8)%,阴性对照组的细胞增殖抑制率为(12.9±5.3)%,差异有统计学意义(P<0.05).结论 在人胚肾HEK293中高表达蛋白hSav1可明显抑制细胞增殖能力.     

Loss of BAP1 protein expression in the first metastatic site predicts prognosis in patients with clear cell renal cell carcinoma

Objectives

To investigate the intratumoral heterogeneity of BAP1 and PBRM1 expression at the primary site and metastatic sites and to evaluate whether BAP1 and PBRM1 expression in metastatic sites of clear cell renal cell carcinoma (ccRCC) has prognostic value.

内皮素对人肾小球系膜细胞表达细胞粘附分子的影响

目的 探讨内皮素-1(ET1-1)对体外培养的人肾小球系膜细胞表达细胞间粘附分子-1(ICAM-1)和血管细胞粘附分子-1(VCAM-1)的调节作用。方法 利用Northern杂交技术和细胞ELISA方法分别从mRNA和蛋白质两个水平观察ICAW-1和VCAM-1的变化。结果 在正常培养条件下,人肾小球系膜细胞仅低水平表达一定量的ICAM-1、VCAM-1的mRNA和蛋白质;ET-1(10~(-7)mol)刺激后,这两种细胞粘附分子的mRNA表达迅速上调,且随后其相应蛋白质表达也显著增加并呈时间依赖性。结论 提示ET-1对粘附分子表达的促进作用可能在传小球疾病的发生中具有重要意义。     

ICE在肾癌中的表达及与细胞凋亡率的关系

目的探讨白细胞介素１β转换酶（ＩＣＥ）在肾癌中的表达及与细胞凋亡的关系。方法用免疫组化及流式细胞技术，测定５２例肾癌和１４例癌旁正常肾组织ＩＣＥ的表达及细胞凋亡率。结果ＩＣＥ在肾癌细胞中的表达主要位于胞浆，阳性率为８９％，高于正常肾组织的３５％，Ｐ＜００１；阳性细胞含量为（７９４±１５５）％，高于正常肾组织的（３２７±９７）％，Ｐ＜００１，相对蛋白含量（ＦＩ值）为１５９±０１６，高于正常肾组织的０５６±００８，Ｐ＜００１，肾癌细胞凋亡率为（４２±１９）％，正常肾细胞凋亡率为（９２±１６）％；ＩＣＥ表达的ＦＩ值与肾癌细胞凋亡率呈正相关（ｒ＝０４６７，Ｐ＜００５）。结论ＩＣＥ的超表达使肾癌细胞具有更易于凋亡的趋势，是肾癌发生、发展的负调控因子，但未引起肾癌细胞凋亡率的明显改变，推测在肾癌中可能有ＩＣＥ的抑制因子存在或ＩＣＥ结构和功能的变异。     

DNA hypermethylation in papillary renal cell carcinoma

What’s known on the subject? and What does the study add? Epigenetic alterations play an essential role during carcinogenesis, and earlier studies suggested that global histone modification levels are predictive for patients’ outcome in various tumour entities (e.g. prostate, lung, breast and kidney cancer). So far, only smaller studies on histone H3K9 acetylation in patients with papillary urothelial neoplasm of low malignant potential (PUNLMP) and histone H2AX phosphorylation in non‐muscle invasive bladder cancer exist; both studies indicate prognostic relevance for bladder cancer. We demonstrate that histone methylation levels (H3K4, H4K20) decrease from normal tissue over non‐muscle invasive and muscle invasive bladder cancer to bladder cancer metastases. Histone modifications are correlated to advanced pathological stage in non‐muscle and muscle invasive bladder cancer. Furthermore, H4K20me3 appeared to be predictive for bladder cancer specific survival in patients with muscle‐invasive bladder cancer undergoing radical cystectomy.

OBJECTIVE

To investigate the pattern of DNA CpG island hypermethylation in papillary renal cell carcinoma (pRCC).

MATERIAL AND METHODS

DNA from pRCC (n= 32) and adjacent normal tissue (n= 15) was isolated. A quantitative methylation‐specific PCR was performed to analyse the methylation pattern at APC (actin beta), CDH1 (E‐cadherin), GSTP1 (glutathione S‐transferase pi 1), RASSF1A (Ras association domain family member 1A) and TIMP3 (TIMP metallopeptidase inhibitor 3); a sequence of ACTB without CpG was used to normalize for DNA input and to calculate the relative amount of methylated DNA (normalized index of methylation, NIM).

RESULTS

RASSF1A hypermethylation was observed in most pRCC and normal samples (100 vs 94.4%), but the median NIM was significantly higher in pRCC samples (2.11 vs 0.61; P < 0.001). RASSF1A hypermethylation allowed discrimination of pRCC and normal tissue with a sensitivity of 87.5% and a specificity of 73.3% as determined via receiver operator characteristic analysis (area under curve = 0.814). Hypermethylation at APC (3.0 vs 6.7%), CDH1 (15.6 vs 0%), GSTP1 (21.9 vs 6.7%) and TIMP3 (6.3 vs 0%) was infrequent in pRCC and normal tissue. CDH1 was significantly correlated with pathological stage (P= 0.015), and patients with methylated CDH1 methylation showed a trend towards shorter recurrence‐free survival (log‐rank P= 0.057). The number of methylated gene sites was correlated with pathological stage (P= 0.007) and lymph node metastasis (P= 0.008).

CONCLUSIONS

DNA hypermethylation at RASSF1A is common in pRCC tissue irrespective of the histological subtype, but also frequently seen at lower levels in normal adjacent tissue. Aberrant hypermethylation could be a prognostic marker for pRCC.     

Serum cell adhesion molecules in patients with colorectal cancer

(Received for publication on Oct. 17, 1996; accepted on July 8, 1997)     

Dapper1在胃癌组织中的表达及其对survivin介导的细胞凋亡的调控

目的 研究Dapper1(Dpr1)在胃癌组织中的表达及其对survivin介导的细胞凋亡的调控.方法 用RT-PCR方法检测30例患者胃癌及癌旁正常组织中Dpr1 mRNA的表达情况;用RT-PCR检测SGC7901细胞转染pcDNA3.1-Dpr1质粒前后Dpr1和survivin mRNA表达的变化;用Western blot检测转染前后Dvl-2、β-catenin及survivin蛋白表达的变化;用流式细胞术检测转染后SGC7901细胞凋亡率的变化. 结果本组30例胃癌组织中17例Dapperl mRNA表达下调,发生率为57%,且与胃癌的浸润深度和分化程度相关(P＜0.05);转染pcDNA3.1-Dpr1后SGC7901细胞中Dpr1 mRNA表达水平上调,survivin mRNA表达水平下调;Dvl-2、β-catenin以及Survivin蛋白表达量降低;转染pcDNA3.1-Dpr1质粒后SGC7901细胞的凋亡率升高. 结论 Dpr1能够通过经典WNT通路抑制凋亡相关蛋白survivin的表达,在胃癌细胞的凋亡中发挥重要作用.     

Prognostic value of PD-1 and PD-L1 expression in patients with metastatic clear cell renal cell carcinoma

Objective

In renal cell carcinoma (RCC), several prognostic biomarkers have been identified and are under investigation. Several reports have shown that the expression of programmed death 1 (PD-1) and its ligand PD-L1 is associated with poor outcome for patients with RCC. The present study is aimed at evaluating the expression of PD-1 and PD-L1 and to investigate their clinical and prognostic significance in patients with clear cell RCC (CCRCC) having received molecular targeted therapies. In addition, we also evaluated the relationship between the expression of PD-1 and PD-L1 and intratumoral tumor infiltrating lymphocytes (TILs).

人肝硬化组织中DMBT1的表达与卵圆细胞增生的关系

目的 探讨DMBTl在人类肝硬化组织中的表达及其与卵圆细胞增殖的关系.方法 对22例人肝硬化标本进行CK19、Hep和DMBT1免疫组化染色,用激光显微切割方法研究DMBT1在增生的胆小(PBD)中的缺失(LOH),同时对其中5例标本进行电镜观察·以10例正常肝脏作对照.结果 光镜下,所有肝硬化组织中均可找到少量卵圆细胞,其数量与炎症程度成正相关,电镜下证实卵圆细胞位于PBD中.DMBT1在卵圆中细胞也有表达,其在PBD中的阳性率为45.5%.LOH分析发现DMBT1在PBD中的发生率为45.5%,与周围肝细胞相比有显著性差异(P<0.05).而正常肝组织中未找到双向表达的卵圆细胞,仅少量大胆管有DMBTl的表达.结论 人肝硬化组织中存在双向分化的卵圆细胞,位于PBD中,DMBT1在此类细胞中高表达,DMBT1 LOH较特异的出现在PBD中,并与蛋白表达呈现相对一致性,说明DMBT1可能参与卵圆细胞的分化增殖,在肝硬化的发生与发展中可能起到调控作用.