Correlation between expression of gastrin, somatostatin and cell apoptosis regulation gene bcl-2/bax in large intestine carcinoma

AIM: To explore the correlation between expression of somatostatin (SS), gastrin (GAS) and cell apoptosis regulation gene bcl-2/bax in large intestine carcinoma.METHODS: Sixty-two large intestine cancer tissue samples were randomly and retrospectively selected from patients with large intestine carcinoma. Immunohistochemical staining for bcl-2, bax, GAS, SS was performed according to the standard streptavidin-biotin-peroxidase (S-P) method.According to the semi-quantitative integral evaluation, SS and GAS were divided into three groups as follows. Scores1-3 were defined as the low expression group, 4-8 as the intermediate expression group, 9-16 as the high expression group. Bax and bcl-2 protein expressions in different GAS and SS expression groups of large intestine carcinoma were assessed.RESULTS: The positive expression rate of bax had a prominent difference between SS and GAS high, intermediate and low expression groups (P＜0.05, x2ss = 9.246; P＜0.05,x2GAS = 6.981). The positive expression rate of bax in SS high (80.0%, 8/10) and intermediate (76.5%, 13/17)expression groups was higher than that in low expression group (40.0%, 14/35) (P＜0.05, x2high vs low = 5.242; P＜0.05,x2middle vs low = 6.097). The positive expression rate of bax in GAS high expression group (27.3%, 3/8) was lower than that in low expression group (69.4%, 25/36) (P＜0.05,x2 = 4.594). However, bax expression in GAS intermediate expression group (46.7%, 7/15) was lower than that in low expression group, but not statistically significant. The positive expression rate of bcl-2 had a prominent difference between SS and GAS high, intermediate and low expression groups (P＜0.05, x2ss = 7.178; P＜0.05, x2GAS = 13.831). The positive expression rate of bcl-2 in GAS high (90.9%, 10/11)and intermediate (86.7%, 13/15) expression groups was higher than that in low expression group (44.4%, 16/36)(P＜0.05,x2high vs low = 5.600; P＜0.05, x2 middle vs low = 7.695).However, the positive expression rate of bcl-2 in SS high (40.0%, 4/10) and intermediate (47.1%, 8/9) expression groups was lower than that in low expression group (77.1%, 27/35)(P＜0.05, x2 high vs low = 4.710; P＜0.05, x2 middle vs low = 4.706).There was a significant positive correlation between the integral ratio of GAS to SS and the integral of bcl-2 (P＜0.01,r=0.340). However, there was a negative correlation between the integral ratio of GAS to the SS and bax the integral of (P＜0.05, r = -0.299).CONCLUSION: The regulation and control of gastrin,somatostatin in cell apoptosis of large intestine carcinoma may be directly related to the abnormal expression of bcl-2, bax.

Bcl-2 GENE REARRANGEMENT DETERMINED BY PCR AS AMEAN TO DETECT MINIMAL RESIDUAL DISEASE INMALIGNANT LYMPHOMAS

Objective: To develop a sensitive method to detect minimal residual disease and to elucidate the significance of bcl-2 gene rearrangement in diagnosis and treatment of malignant lymphoma. Methods: Using polymerase chain reaction (PCR) to detect bcl-2 gene rearrangement and using serial dilution method to define the sensitivity of PCR. Results: In 9 different malignant lymphoma cell lines, Su-DHL-4 and Su-DHL-6 were shown bcl-2(MBR)/JH rearrangement, the sensitivity of PCR was 1:105. In 16 patients with follicular lymphoma, the peripheral blood and bone marrow were PCR positive in 4 cases both at initial diagnosis and after complete remission. Conclusion: Detection of bcl-2 gene rearrangement by PCR provides a sensitive and specific assay of minimal residual disease. It is helpful to improve staging of disease, prognosis and evaluation of the treatment results.

Study on the Regulation of Bcl-2 Gene on Rat Spermatogenic Cells Apoptosis in Transcription Level

Objective To detect the change of Bcl 2 gene expression in the apopototic process of spermatogenic cells in rat with vasoligation and vasostomy, and to find out the relationship between the transcription of Bcl 2 and the apoptosis of spermatognic cells Materials & Methods Sixty adult male Sprague Dawley rats in 3 groups were operated with vasoligation and vasostomy. Then hybridization in situ with hypersensitive Bcl 2 RNA probe was used to detect the change of Bcl 2 mRNA. Results The transcription of Bcl 2 gene in spermatogenic cells was obviously inhibited in the vasoligation group compared with that in the control group (P<0.05), and the transcription in the vasostomy group showed no difference from that of the control group. Conclusion Bcl 2 gene has an anti apoptotic effect in rats with vasostomy, and there was a transcriptional regulation of Bcl 2 gene in rat spermatogenic cell during the period of pre vasoligation to post vasoligation and to post vasosotomy.

DETECTION OF BCL-2 GENE MAJOR BREAKPOINT REGION REARRANGEMENT IN HUMAN B-CELL LYMPHOMAS

Objective To investigate the frequency of t(14; 18) in different subtypes of B-cell lymphomas and the ability or the polymerase chain reaction(PCR) to detect this rearrangement in frozen samples. Methods 1o7 cases of B-cell lymphomas were studied uslng DNA extracted from rresh-frozen tissues. The DNA samples were amplified by PCR for bcl-2 MBR/JH. The products of bcl-2/JH rearrangement were hybridized with an internal olignucleotide probe or bcl-2 MBR. Results The rearranged bcl-2MBR/JH gene was detected in 13 of the 25(52. o% ) follicular center lymphomas, according to REAL classification: 8 of 11 (72. 7%) grade 1, 2 of 5(40. 0%) grade I, and 3 of 90 (33. 3%) grade, 17 of 82(2o. 8%) cases or difruse large B-cell lymphomas were found to have detectable bel-2 MBR/J. rearrangement- Conclusion The rrequency or bcl-2 MBR/JH rearrangement in diffuse large B-cell lymphomas is significantly lower than those in follicular center lympkomas(X2= 9. 28, P <o. oo5), suggesting that bcl2/JH rearrangements occur mainly in follicular center lymphomas. in addition, the result of reconstruction experiments suggest that amplification or bcl-2 MBR/JH rearrangements by PCR is both sensitive and specific for detection of t (14; 18 ) translocation.

Silencing of Bcl-2 gene expression by siRNA transfection in- hibits the protective effect of fluvastatin against cell apoptosis in human aortic endothelial cells

Objective To study the protective effect of fluvastatin,one of the HMG-CoA reductase inhibitors (statins),against oxygen radical-induced oxidative damages in human aortic endothelial cell,and the role of Bcl-2 in this protection.Methods Human aortic endothelial cells with or without Bcl-2 siRNA transfection were subjected to 1-100 nM of fluvastatin and 100 la hydrogen peroxide for 24 hours.Bcl-2 mRNA and protein expression were measured by Taqman quantitative PCR and Western blotting.Cell apoptosis was measured by normal and fluorescent microscopy and Cell Death Detection ELISA.Results In the Bcl-2-expressed cells,fluvastatin significantly reversed hydrogen peroxide-induced microscopic apoptosis and apoptotic DNA fragmentation,which were accompanied by a markedly upregulation of Bcl-2 expression by fluvastatin.However,the endothelial protection by fluvastatin was completely lost in Bcl-2 siRNA transfected cells.Conclusion Fluvastatin protects human endothelial cells against oxygen radical-induced cell apoptosis in vitro,and this protection seemed to be mediated in a Bcl-2 dependent pathway.(J Geriatr Cardil 12008;5:33-38)

THE EXPRESSION OF BCL-2 GENE AFTER TRANSIENT FOCAL ISCHAEMIA AND THE EFFECT OF MK-801

in order to clarify the pesible role of the bcl-2 gene imolved in the cell death Program,and the relatiouship of glutamate receptors with bcl-2 gene expressin, this study examied the expression of bcl-2 gene protein, the neuronal status of apoptosis and the effects of MK-801 using immunohistochemistry and in situ terminal.labelling methods after 30 min of.middle cerebral artery(MCA) occlusion and followed by 24 h of reperfusion. The presence of bcl-2 gene protein increased in the ipeilateral hemisphere of ischaemis espeially in the MCA territory MK-801 enhanced the expresion of the bcl-2 gene protein. No DNA fragmentation was detected in this experiment. In conclusion. bcl-2 gene activity increased during transient focal ischaemia, and was potentiated by MK MK801, which may be an endogenous protective mechanism .against ischaemic apoptosis. Apoptosis wasnot detected after tranient focal ischoemia. for 30 min rollowed by 24 h of reperfusiou.

Bcl-2/Fas与子宫内膜异位症发病关系的研究

目的研究Bcl-2/Fas基因蛋白与子宫内膜异位症的发病关系。方法用免疫组化ABC法检测了40例正常子宫内膜和46例子宫内膜异位症患者。结果子宫内膜异位症患者的内膜组织中的表达评分(172.4±87.6)明显高于正常子宫内膜中的表达评分(106.2±118.1),有显著性差异(P<0.05);Fas基因蛋白在子宫内膜异位患者异位内膜细胞中的表达评分(207.6±100.2)与正常子宫内膜上皮中的表达评分(119.3±91.3)无显著性差异(P>0.05)。结论Bcl-2基因与子宫内膜异位症的发生有一定关系,Fas基因对子宫内膜异位症的发生关系有待于进一步研究。

Bcl-2基因反义寡核苷酸对乳腺癌细胞株药物敏感性的影响

目的:探讨bcl-2基因反义寡核苷酸对人乳腺癌细胞株MCF-7的bcl-2蛋白的表达和药物敏感性的影响。方法:用人工合成bcl-2基因反义寡核苷酸预培养人乳腺癌细胞株MCF-7,以免疫组化法检测细胞bcl-2蛋白的表达水平;以MTT法检测bcl-2反义寡核苷酸、阿霉素单独及联合应用对MCF-7细胞生长的抑制情况,比较bcl-2蛋白表达与肿瘤细胞药物敏感性之间的相关性。结果:联合应用bcl-2反义寡核苷酸与阿霉素能够明显增加MCF-7细胞对阿霉素的敏感性,细胞存活率明显下降。免疫组化结果显示MCF-7细胞bcl-2表达水平与细胞药物敏感性有相关性。结论:bcl-2反义寡核苷酸能够抑制MCF-7乳腺癌细胞bcl-2基因的表达,提高MCF-7细胞对阿霉素的敏感性。

抗凋亡基因bcl-2 cDNA表达型质粒的构建及序列分析

目的 :克隆抗凋亡基因bcl-2,并构建其表达型质粒 ;进行序列分析 ,为基因转染作准备。方法 :应用PCR定向克隆技术 ,克隆SD大鼠鼠脑bcl-2全编码cDNA序列 ;应用双脱氧末端终止法进行重组质粒中bcl-2的cDNA序列分析 ;利用基因重组技术 ,将bcl-2全编码cDNA序列 ,导入pCMV -Tag -1表达型质粒。结果 :成功地克隆SD大鼠bcl-2全编码cDNA序列 ,并将其成功地导入pCMV -Tag -1表达型质粒 ;经序列分析 ,发现bcl-2全编码序列与GeneBank中的相应序列相比 ,有3个碱基出现差别 ,分别为171位a→g,233位a→c ,530位 g→a,均为同义突变。结论

Selection of Effective Bcl-2 Antisense Oligodeoxynucleotides with Computer Software and Experimental Assay

Objective： To explore and investigate the selection of effective antisense oligodeoxynuleotides with the help of computer and RNAstructure folding software. Methods： Bcl-2 gene was used as the target gene and five antisense oligodeoxynuleotides were designed to be bound to Bcl-2 mRNA optimal secondary structure regions that were predicted free from intramolecular fold or instability of free energy. The five antisense oligodeoxynucleotides were studied with experimental assay of leukemia cells, including cell grow assay with tropan blue exclusion, expression of Bcl-2 protein detected with immunochemistry and flowcytometry, Bcl-2 mRNA content detected with RT-PCR technique, as well as apoptosis observed and determined with morphonological method, electrophoresis and flowcytometry. Results： The results showed that two of the five antisense oligodeoxynucleotides were effective antisense oligodeoxynucleotides, which were able to inhibit cell growth in leukemia, to decrease the level of Bcl-2 mRNA and protein, to induce apoptosis of leukemia cells significantly. Conclusion： The computational prediction of antisense efficacy is faster than other methods and more efficient, which can potentially speed the development of sequences for both research and clinical applications.

急性髓性白血病bcl-2mRNA表达与抗癌药敏感性及临床疗效关系

目的:探讨急性髓性白血病ｂｃｌ－２ｍＲＮＡ表达与抗癌药物敏感性及临床疗效的关系。方法:用半定量ＲＴ－ＰＣＲ方法检测２９例急性髓性白血病ｂｃｌ—２ｍＲＮＡ表达,并用ＭＴＴ法测定１０种抗癌药的抑制率,以正常健康人ｂｃｌ－２ｍＲ－ＮＡ表达水平均值作为分界点,分成阴性组和阳性组,统计比较抗癌药在两组中的抑制率并观察临床疗效。结果:ｂｃｌ—２ｍＲ－ＮＡ表达阴性组抗癌药抑制率大于阳性组,其中氮芥、阿糖胞苷、鬼臼乙叉甙存在显著差异(Ｐ＜０．０５),甲氨蝶呤、环磷酰胺存在极显著差异(Ｐ＜０．０１)。ｂｃｌ－２ ｍＲＮＡ阴性组化疗缓解率高于阳性组,两组具统计学意义(Ｐ＜００５)。结论:ｂｃｌ－２ｍＲＮＡ在急性髓性白血病表达可作为一种多药抵抗分子来预测体外药物的敏感性,ｂｃｌ－２ｍＲＮＡ低表达者化疗疗效好。

bcl-2小分子干扰RNA对人子宫内膜癌RL-952细胞阿霉素敏感性的影响

目的研究靶向bcl-2小分子干扰RNA(siRNA)干扰序列对人子宫内膜癌RL-952细胞阿霉素(DOX)敏感性的影响及相关作用机制。方法 2014年5—8月,将siRNA干扰序列转染人子宫内膜癌RL-952细胞,设3个实验组:空白对照组、bcl-2 siRNA干扰组(转染bcl-2 siRNA干扰序列)及阴性对照组(转染siRNA空载序列),检测bcl-2 siRNA干扰组的转染效率和3组bcl-2表达水平。将细胞随机分成5组,分别为空白对照组、阴性对照组(转染siRNA空载序列)、干扰组(转染bcl-2 siRNA干扰序列)、DOX组(加入5μg/ml DOX)、联合组(转染siRNA干扰序列并加入5μg/ml DOX),检测细胞活力、细胞凋亡率、半胱氨酸蛋白酶(Caspase)-3、Caspase-9活性及细胞色素C水平。结果 siRNA干扰序列转染人子宫内膜癌RL-952细胞的转染效率为(94.6±12.5)%。3组bcl-2表达水平比较,差异有统计学意义(F=8.75,P<0.05)。bcl-2 siRNA干扰组bcl-2表达水平低于空白对照组和阴性对照组(q=3.99、2.87,P<0.01)。5组细胞活力比较,差异有统计学意义(F=42.52,P<0.05)。干扰组、DOX组、联合组细胞活力低于空白对照组(q=7.66、8.64、6.89,P<0.05);联合组细胞活力低于DOX组(q=6.76,P<0.01)。5组细胞凋亡率比较,差异有统计学意义(F=112.34,P<0.05)。干扰组、DOX组、联合组细胞凋亡率高于空白对照组(q=5.78、4.99、6.21,P<0.05);联合组细胞凋亡率高于DOX组(q=4.89,P<0.01)。5组Caspase-3、Caspase-9活性及细胞色素C水平比较,差异有统计学意义(P<0.05)。干扰组、DOX组、联合组Caspase-3、Caspase-9活性及细胞色素C水平高于空白对照组(P<0.05);DOX组、联合组Caspase-3活性高于阴性对照组,干扰组、DOX组、联合组Caspase-9活性高于阴性对照组(P<0.05);联合组Caspase-3活性高于干扰组,DOX组、联合组Caspase-9活性高于干扰组(P<0.05);联合组Caspase-3、Caspase-9活性及细胞色素C水平高于DOX组(P<0.05)。结论靶向bcl-2 siRNA干扰序列可以增强人子宫内膜癌RL-952细胞的DOX敏感性,bcl-2可作为人子宫内膜癌基因治疗的候选靶点。

P53和Bcl-2蛋白在基底细胞癌中的表达及临床意义

目的探讨P53、B细胞淋巴瘤/白血病-2基因(bcl-2)蛋白在基底细胞癌(BCC)组织中的表达及临床意义。方法用免疫组化链霉素抗生素蛋白-过氧化物酶连结(SP)法检测P53、Bcl-2蛋白在40例BCC(BCC组)组织中的表达,以20例正常皮肤组织作为对照组。结果 P53、Bcl-2蛋白在BCC组表达阳性率分别为55%(22/40)、85%(34/40),在对照组表达阳性率分别为15%(3/20)、10%(2/20)。P53、Bcl-2蛋白在BCC组中的表达明显强于对照组,差异有统计学意义(χ2=6.35、7.45,P<0.01)。结论 P53、Bcl-2蛋白的异常表达与BCC相关,说明其可能在BCC的发生及发展过程中起重要作用。

Intravenous acid fibroblast growth factor protects intestinal mucosal cells against ischemia-reperfusion injury via regulating Bcl-2/Bax expression

AIM: To detect the effect of acid fibroblast growth factor (aFGF) on apoptosis and gene expression of bax and bcl-2 gene in rat intestine after ischemia/reperfusion (I/R) injury, and to explore the protective mechanisms of aFGF.METHODS: One hundred and eight Wistar rats were randomly divided into sham-operated control group (C)(n = 6), intestinal ischemia group (I) (n = 6), aFGF treatment group (A) (n = 48) and intestinal ischemia reperfusion group (R) (n = 48). In group I, the animals were killed after 45 min of superior mesenteric artery (SMA) occlusion, while in groups R and A, the rats sustained 45 min of SMA occlusion and were then treated with normal saline and aFGF, respectively, sustained 15 min, 30 min, 1, 2, 6, 12, 24, or 48 h of reperfusion, respectively. In group C, SMA was separated, but without occlusion. Apoptosis in intestinal villus was determined with terminal deoxynucleotidyl transferase mediated dUTP-biotin nickend labeling technique (TUNEL). Intestinal tissue samples were taken not only for detection of bax and bcl-2 gene expression by RT-PCR, but also for detection of bax and bcl 2 protein expression and distribution by immunohistochemical analysis.RESULTS: The rat survival rates in aFGF treated group were higher than group R (P＜0.05) and the improvement of intestinal histological structures was observed at 2, 6, and 12 h after the reperfusion in group A compared with group R. The apoptotic rates were (41.17±3.49)%, (42.83±5.23)% and (53.33±6.92)% at 2, 6 and 12 h after reperfusion, respectively in group A, apparently less than those of group R at matched time points (50.67±6.95, 54.17±7.86, 64.33±6.47, respectively) (P＜0.05). The bax gene transcription and translation were significantly decreased in group A vs group R, while mRNA and protein contents of Bcl-2 in group A were obviously higher than those in groupR during 2-12 h period after reperfusion.CONCLUSION: The changes in histological structure and the increment of apoptotic rate indicated that the intestinal barrier was damaged after intestinal I/R injury, whilst intravenous aFGF could alleviate apoptosis induced by ischemia and reperfusion in rat intestinal tissues, in which genes of bax and bcl-2 might play important roles.

Relationship between expression and distribution of cyclooxygenase-2 and bcl-2 in human gastric adenocarcinoma

AIM: To explore expression and distribution features of COX-2 and bcl-2 in human gastric adenocarcinoma tissues and to study its biological significance.METHODS: Totally 36 human gastric carcinoma samples were enrolled in this study (cardiac adenocarcinoma 16 cases, distal gastric adenocarcinoma 20 cases). The expressions of COX-2 and bcl-2 in cancerous tissues and corresponding para-cancerous tissues were investigated by immunohistochemistry using COX-2 polyclonal antibody and bcl-2 monoclonal antibody. The normal gastric mucosa tissues were used as control.RESULTS: The expressions of COX-2 and bcl-2 in gastric carcinoma were significantly higher than that in the paracancerous tissues (77.8% vs 47.2%, P＜0.01, 80.56% vs 58.33%, P＜0.05). The expression of COX-2 in cardiac adenocarcinoma was remarkably higher than that in the distal gastric carcinoma (93.8% vs 65.0%, P＜0.01). The expression of COX-2 was mainly localized in the cytoplasm of tumor cells and partly in the nucleus. There is a transition of the COX-2 cytoplasmic positivity to nucleic in tumor cells with the increase of gastric carcinoma pathological grade. Interstitial macrophages, fibroblasts and vascular endothelial cells also expressed COX-2. The tissues with higher expression of COX-2 also expressed high level of bcl-2 protein.CONCLUSION: Abnormal expression pattern of COX-2within the tissues of human gastric cancer is correlated with tumor location and lymph node metastasis. COX-2may regulate expression of apoptosis suppressor gene (bcl-2) through interaction of tumor cells and stromal cells and play an important role in the generation and development of tumors, which will be of great help in developing new methods for antitumor therapy.

EFFECT OF TWO NEW BCL-2 ANTISENSES ON DRUG-SENSITIVITY OF CELLS FROMN LEUKEMIA PATIENTS

Objective: To investigate the effect of two antisense oligonucleotides on cell surviving, bcl-2 expression and apoptosis of leukemia cells. Methods: The experimental assays were performed with cell culture, immunochemistry and flowcytometry. Results: The two antisense oligodeoxynucleotides, combined with Vp16 or Ara-c or DNR, were able to decline the survival rate of myeleukemic cells, downregulate bcl-2 gene expression and induce apoptosis of leukemic cells significantly, as compared with Vp16 or Ara-c or DNR alone. Conclusion: It is possible for the two new bcl-2 antisenses to be developed into clinical trials for leukemia and tumor with bcl-2 gene overexpression. Key words Leukemia - Antisense oligodeoxynucleotides - bcl-2 gene - drug-resistance CLC number R733.7 Foundation item: this work was supported by the Key Foundation of Science & Technology Program of Guangzhou (No. 2001-Z-037-01); and the Nature Science key Foundation of Guangdong Province(No. 021195).Biography: LEI Xiao-yong(1970–), male, associate professor, doctor of medicine, Institute of Pharmacy and Pharmacology, Nanhua University, majors in tumor pharmacology.

The Variety Selection and Testing for Effect on HepG-2 Cell Line of Dioscorea membranacea Pierre （Hua Khao Yen Tai） Extract Using Molecular Genetics

In this research, Dioscorea membranacea Pierre was studied by using 30 samples from Khao Ruak Sub-district, Chai Badan District, Lop Buri Province, in Thailand. In this research, some morphology was studied including shapes, leaves, stem colors, epidermal cells, stomata sizes and stipules. To study the genetic relationships, the AFLP technique and computer program were used. The Dioscorea membranacea Pierre was classified into 2 groups according to its phylogenetic type： the first group was ＂Hua Khao Yen Tai-Nuea＂ （Smilax corbularia Kunth）, and the second group included 30 further samples of Hua Khan Yen Tai （Dioscorea membranacea Pierre）. The ethanolic crude extract was also applied to test the anti-proliferative activity in the liver hepatocellular carcinoma （HepG2） cell lines which illustrates the characteristics of apoptosis： cell shrinkage, membrane blebbing and nuclear condensation. The expression ofBax gene is increased more than that of the control group while Bcl-2 gene which is anti-apoptotic is decreased. Furthermore, the result of western blot analysis reveals the up-regulation of Bax protein and down-regulation of Bcl-2 protein when compared with untreated cells. This might indicate that ethanolic crude extracts of Hua Khao Yen Tai could induce apoptosis and anti-proliferative on HepG2 cell lines, The results also revealed that some morphology cannot be used to predict which Dioscorea membranacea Pierre plants would be most effective.

^(103)Pd-induced apoptosis of proliferative smooth muscle cells in bile ducts of dogs:significance and effects on related genes

BACKGROUND:With the objective of developing a locally- produced radioactive stent,the present study used in vivo animal experiments to explore apoptosis of proliferative smooth muscle cells resulting from facilitation of the expression of genes caused byγ-radiation in order to prevent bile duct restenosis.We therefore explored the effects and significance ofγ-radiation on the activity of caspase-3,Fas and Bcl-2 genes in apoptosis of proliferative smooth muscle cells in the bile duct walls of dogs. METHODS:Twelve dogs were randomly divided into 2 groups(6 in each group).A postinjury bile duct stenosis model was established and radioactive 103 Pd( 103 palladium) or ordinary bile duct stents were implanted into the bile ducts.HE staining,RT-PCR and immunohistochemistry were used to detect the proliferation and apoptosis of bile duct smooth muscle cells in proliferative endomembrane and the expression of related caspase-3,Bcl-2 and Fas genes. RESULTS:The expression of caspase-3 and Fas genes in the bile duct tissues of dogs with radioactive stents was higher than that of dogs with ordinary stents.There was significant apoptosis of proliferative smooth muscle cells in the bile ducts.The expression of the Bcl-2 gene in the bile duct tissues of dogs with radioactive stents was lower than that in those with ordinary stents.There was significant apoptosis of proliferative smooth muscle cells in the dogs with low Bcl-2 gene expression. CONCLUSIONS:Radiation increases the activity of caspase-3 and Fas genes and is associated with apoptosis. The radioactive 103 Pd stent may facilitate apoptosis of proliferative smooth muscle cells in the bile ducts of dogs by activating these genes.The Bcl-2 gene expression level is correlated with the occurrence of apoptosis and the radiosusceptibility of cells.

Apoptosis of human colon carcinoma HT-29 cells induced by ceramide

AIM： To investigate the effect of exogenous ceramideinduced apoptosis on human colon carcinoma HT-29 cells. METHODS： Light microscope, transmission electron microscope and fluorescence microscope were used to observe the morphology change of apoptosis in HT-29 cells. Agarose gel electrophoresis was performed to detect the DNA fragment. Mitochondrial function was detected by MTT assay, mRNA expression of Bcl-2 family gene members was determined by reverse transcription polymerase chain reaction （RT-PCR） assay. RESULTS： After C2-ceramide treatment, typical characteristics of apoptosis, such as nuclear chromatin breakage, apoptotic body and DNA ladder, could be observed. After exposure to 50μmol/L C2-ceramide for 12 and 24 h, cell apoptosis was 64.1% and 81.3% respectively, which had a time-and dose-effect relationship. Mitochondrial function started to decrease from 6 h after exposure to ceramide. Meanwhile, ceramide up-regulated or down-regulated the mRNA expression of Bcl-2 family gene members. CONCLUSION： Ceramide induces apoptosis of human colon carcinoma HT-29 cells by affecting the expression of Bcl-2 family gene members and impacting the mitochondrial function.

Prognostic role of sensitive-to-apoptosis gene expression in rectal cancer

AIM:To investigate the association between prognosis of rectal cancer treated with chemoradiotherapy(CRT) and expression of sensitive-to-apoptosis(SAG),B-cell lymphoma-extra large(Bcl-X L) and Bcl-2 homologous antagonist/killer(Bak).METHODS:Real-time quantitative polymerase chain reaction was used to determine the expression of proteins of interest,namely SAG,Bcl-X L,Bak and β-actin,in rectal carcinoma patients who had a follow-up period of 3 years after CRT.Biopsy specimens were excised from the rectal tumor preceding CRT.RESULTS:SAG,Bcl-X L and Bak proteins showed significant correlations with each other.In multivariate analysis,patients with high vs low SAG expression showed a statistically significant difference in 2-year survival rates:56% vs 73%,respectively(P = 0.056).On the other hand,there were no significant correlations between the expression levels of all three genes and metastatic rates or tumor responses to CRT.Mean overall survival in the patients with elevated SAG expression was 27.1 mo ± 3.9 mo [95% confidence interval(CI):19.3-34.9],and in patients with reduced expression,it was 32.1 mo ± 2.5 mo(95% CI:27.3-36.9).The corresponding values for Bcl-X L were 28.0 mo ± 4.1 mo(95% CI:19.9-36.1) and 31.7 mo ± 2.9 mo(95% CI:26.0-37.5),and those for Bak were 29.8 mo ± 3.7 mo(95% CI:22.5-37.2) and 30.6 mo ± 2.4 mo(95% CI:25.5-35.0),respectively.CONCLUSION:Two-year survival rates significantly correlated with low SAG expression,and SAG may be a candidate gene for good prognosis,independent of therapeutic response of different individuals.