Livin Gene Research Articles

Study of transfection of Livin antisense oligodeoxynucleotide on apoptosis of QBC939 cells and its mechanism

Objective To study the effect of transfection of Livin antisense oligodeoxynucleotide (Livin ASODN) on Livin mRNA and Livin protein expression and apoptosis of QBC939 cells.Methods Livin ASODN were transfected into cell line QBC939 byLipofectamine TM 2000.60 h After transfection,to measure Livin mRNA expression by reverse transcriptase polymerase chain reaction (RT-PCR) and Livin protein expression by immunohistochemistry and confocal laser scanning microscopy ; the changes of cell apoptotic were detected by methyl thiazol tetrazolium (MTT).and the apoptosis of QBC939 cell was detected by Annexin V-PE flow cytometry (FCM) after the transfection.Results The mRNA and protein expression level of Livin in ASODN group was significantly lower than that in control group (P ＜0.05) (54.96 ± 3.00,88.31 ± 2.50,85.90 ± 0.62,90.36 ± 2.43) ; the results of MTT showed that the rate of cell apoptosis of QBC939 cells were the most obvious at 60 hours after Livin ASODN transfection (46.41 ± 1.13,3.10 ±0.44,2.66 ± 0.26,0) (P ＜ 0.05).the apoptosis of QBC939 cell was higher than those in control group after Livin ASODN transfection [(36.98 ±4.09)％,(3.77 ±0.25)％,(7.51 ±0.3)％,(6.83 ±0.23) ％].Conclusion The transfection of Livin ASODN inhibited Livin gene and protein expression,and Obviously induced apoptosis of Bile duct cancer QBC939 cells,which may be the new target for gene therapy in Bile duct cancer. Key words: Livin; Antisense oligodeoxynucleotide; Bile duct cancer; Cell apoptosis

Expression and prognostic significance of Livin in gastric cancer

Livin is one of the most important members of the inhibitor of apoptosis protein family. It is overexpressed in several types of tumors and may have prognostic significance. The present study investigated the biological role of Livin in the oncogenic behavior of gastric cancer cells, the expression of Livin in gastric cancer tissue and the relationship of its expression with various clinicopathological parameters and patient survival. Small interfering RNA blocked Livin gene expression in AGS and SNU638 human gastric cancer cell lines. The expression of Livin was investigated in gastric cancer tissues by RT-PCR, western blotting and immunohistochemistry. The associations with various clinicopathological parameters and survival were analyzed. Livin knockdown inhibited tumor cell migration, invasion and proliferation in AGS and SNU638 cells. Livin knockdown induced apoptosis by activating caspase-3, caspase-7 and PARP. Livin knockdown induced cell cycle arrest by a decrease in cyclin D1, cyclin-dependent kinase 4 and 6 and an increase in expression of p21 and p27. The ERK1/2 and JNK signaling pathways were inhibited by Livin knockdown. Livin expression was upregulated in gastric cancer tissues at the mRNA and protein levels. However, no significant correlation was found between Livin expression and various clinicopathological parameters including survival. In conclusion, Livin expression may be important in the alteration of invasive and oncogenic phenotypes of gastric cancer cells. The prognostic relevance of Livin remains unclear.

The antitumor effect of human cord blood-derived dendritic cells modified by the livin α gene in lung cancer cell lines

The growth of malignant tumors is associated with mechanisms of immune escape and inhibition of apoptosis. Livin is a novel member of the inhibitors of apoptosis (IAP) protein family that inhibits cell apoptosis. Livin is specifically expressed by the majority of tumor cells, but it is not expressed in normal adult tissues. In this study, we used umbilical cord blood (UCB)-derived dendritic cells (DCs) infected with a recombinant adenovirus encoding the livin gene as a vaccine to activate effector cells such as cytotoxic T lymphocytes (CTLs) to recognize and kill livin-expressing cancer cells in vitro as an improved strategy for overcoming the ability of these cancer cells to escape apoptosis and antitumor immune responses. We employed interferon-γ (IFN-γ) enzyme-linked immunospot assays to confirm that our immunization strategy induced an antigen-specific reaction to livin and flow cytometric analysis of staining with Annexin V and PI to measure the cytotoxic activity of the effector cells against the livin-expressing lung cancer cell lines A549 and H460. Our results show that the recombinant adenovirus was able to promote the maturation of the UCB-derived DCs. This DC vaccine could activate antigen-specific T cells to produce IFN-γ upon recognition of livin peptide in the context of the appropriate HLA molecule. The antigen-specific T cells mediate significant cytotoxicity against the cancer cells, but are unlikely to cause an autoimmune reaction against the human bronchial epithelia cells (HBE), which do not express livin.

Research progress of Livin gene in tumors

Livin,a novel member of human inhibitor of apoptosis protein family,is highly expressed in various malignant tumors. It plays an important role in the initiation,development,treatment and prognosis of tumors.Many studies have proved that down-regulation of Livin expression using RNA interference technique can reduce tumor growth,increase the apoptotic rate,and sensitize tumor cells to radiotherapy and chemo therapy.Full-length Livin is cleaved by the effector caspases to produce a truncated form (tLivin) with apoptosis-promotion activity.tLivin provides a new direction for the treatment of the malignant tumors. Key words: Gene; Apoptosis; Neoplasms

Expression and role of the inhibitor of apoptosis protein livin in chemotherapy sensitivity of ovarian carcinoma

Ovarian cancer has the highest mortality rate of all gynecological malignancies. Livin, a novel member of the inhibitor of apoptosis protein family, has been found to be expressed in various malignancies and is suggested to have poor prognostic significance. However, no data are available concerning the significance of livin in ovarian cancer. In the present study, we detected the expression of livin isoforms in human epithelial ovarian cancer (EOC) tissues using semi-quantitative RT-PCR and western blot assays. The data indicated that livin expression positive ratio was much higher in cancer tissues compared to those in benign ovarian tumors and normal ovarian specimens. To determine the role of livin in the process of ovarian cancer growth, RNA interference mediated by recombinant lentivirus vectors expressing livin shRNA was applied to induce a long-lasting downregulation of livin gene expression in SKOV3 human ovarian cancer cell line. Cell apoptosis and chemosensitivity were evaluated by MTT assay and flow cytometry, respectively, following downregulation of livin expression, and the cleavage of molecular markers of the mitochondrial apoptotic signaling pathway was investigated by immunoblotting. Livin knockdown with siRNA enhanced spontaneous and drug-induced apoptosis in SKOV3 cells. The inhibition of livin resulted in profound pro-apoptotic and antiproliferative effects, and was associated with the activation of caspase signaling. In conclusion, these data suggested that livin plays an important role in inhibiting the apoptosis of ovarian cancer cells. Specific silencing of livin expression could promote cell apoptosis, enhance chemotherapy sensitivity and may be a promising target for further research in clinical chemotherapy of epitheliod ovarian cancer.

Livin mediates tumor cell invasion in the DU-145 cell line via NF-κB

Livin is a member of the family of inhibitors of apoptosis proteins (IAPs) and tumor cell invasion is a general property of multiple IAPs. Livin is highly expressed in prostate cancer (PCa) tissues. Livin overexpressing cells are more resistant to apoptotic stimuli than normal cells. Thus, aberrantly increased cell survival is an invariable requirement of metastasis. In this study, we investigated whether livin signaling affects metastasis by transfecting siRNA targeting livin into the DU-145 prostate cancer cell line to confirm the anti-invasion effect and blockade of the livin gene. We found that livin knockdown inhibited DU-145 prostate carcinoma cell invasion. We investigated how livin promotes tumor cell invasion, and found that livin induction of fibronectin contributed to tumor cell invasion. In addition, we found that livin induction of fibronectin regulates tumor cell invasion via nuclear factor κB (NF-κB) signaling. These data showed that livin, as a gene directly promoting metastasis, can be useful for therapeutic intervention against advanced and disseminated PCa.

Research of interference Livin gene inhibits growth of QBC939 cell

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. Key words: Livin; Gene interference; Bile duct cancer; Cell proliferation

Livin gene in cancer diagnosis and treatment

Livin, a novel human inhibitor of apoptosis protein family member, participates in the regulation of apoptosis through inhibition of caspases and activation of mitogen activated protein kinase ( MAPK )and other pathways. Livin is over-expressed in many tumors and plays an important role in the initiation and development of tumor, providing a promising new field of cancer research. Key words: Neoplasms; Apoptosis; Genes

Effects of human Livin gene down-regulation by RNA interference on xenograft of transplantation tumor induced by SPC-A1 in nude mice

To explore the in vivo inhibitory effect of Livin gene silencing by RNA interference on xenograft of lung adenocarcinoma SPC-A-1 cells in BALB/C nude mice. Three different BALB/C nude mice models were established by subcutaneously inoculating differently treated SPC-A-1 cells into 3 nude mice groups: the blank control group was inoculated with blank SPC-A-1 cells, while the negative group was inoculated with cells transfected with lentivirus-delivered negative shRNA, the experimental group was inoculated with cells with lentivirus-delivered Livin shRNA. Then the growth of tumors was observed, and the volume and weight of the tumors were measured at different time points. The curve of tumor growth was then described, and the inhibition rate was calculated. Livin gene expression in the tumor tissues was determined by RT-PCR and Immunohistochemistry. Cell apoptosis of tumor tissues was detected by TUNEL. Slower tumor growth, smaller tumor volume and lighter tumor weight were observed in the experimental group as compared to the blank and negative groups (F = 70.509, P < 0.01; F = 12.821, P < 0.01). The inhibition rate of tumor volume was (59.5 ± 3.4)%, and the inhibition rate of tumor weight was (71.1 ± 5.6)%. Livinα mRNA and Livinβ mRNA expressions in the experimental group were significantly lower than the 2 control groups [(37.2 ± 1.6)% versus (63.3 ± 3.8)%, (66.1 ± 2.6)%; (29.4 ± 1.1)% versus (53.2 ± 3.4)%, (52.3 ± 3.1)% (F(α) = 45.309, P < 0.01; F(β) = 30.076, P < 0.01)]. Livin protein expression level was also significantly lower than the blank and the negative groups [(15.3 ± 2.8)% versus (51.3 ± 2.1)%, (52.5 ± 2.5)%, F = 78.92, P < 0.01]. The apoptosis rate in the experimental group was significantly higher than that in the 2 control groups [(35.4 ± 3.2)% versus (5.4 ± 1.3)%, (8.6 ± 1.5)%, F = 14.509, P < 0.01]. The lentivirus-delivered Livin shRNA was shown to inhibit the proliferation of transplantation tumor of lung carcinoma effectively, and Livin may be a target for gene therapy in lung cancer.

SiRNA targeting Livin decreases tumor in a xenograft model for colon cancer

To evaluate the effect of silencing Livin gene expression with siRNA to apoptosis and proliferation in a colon cancer cell line. To investigate the anticancer effect of silencing Livin gene expression, we established an siRNA transfected cell line using the HCT116 colon cancer cell line. After confirming the successful transfection, MTT assay, flow cytometry and annexin V staining were employed to evaluate the antiapoptotic effect. To confirm the in vivo effect of Livin-siRNA, different doses of Livin-siRNA were injected into xenografted tumors in BALB/c nude mice model. Livin expression was dramatically decreased after siRNA transfection, especially at 25 μmol/L of siRNA, but this suppression was not dose-dependent. The cell count at 18 h after transfection was significantly reduced as compared with controls (P < 0.01), but tended not to decrease proportionally depending on transfected dose or time. MTT assay revealed that silencing the Livin gene suppressed cellular proliferation at 18 h after transfection (P = 0.04); however, the inhibitory effect disappeared thereafter. Also, there was no significant difference in cellular proliferation depending on siRNA dose. The rate of apoptosis also increased with silencing of the Livin gene. In vivo, the tumor size significantly decreased after Livin-siRNA injection at 20 μmol/L concentration (P = 0.03). There were no significant body weight changes of mice after siRNA injection. Histologic examination revealed no significant toxic reaction in kidney, liver and brain of mice. siRNA-mediated downregulation of Livin expression can induce apoptosis in colon cancer in vitro and in vivo, which suggests the possibility of new cancer therapeutics using siRNA.

Livin‐α promotes cell proliferation by regulating G1‐S cell cycle transition in prostate cancer

Prostate cancer is the third most common cancer and the second leading cause of cancer death for males in US. Livin has recently been described as a cancer-associated member of inhibitor of apoptosis proteins family, highly expressed in prostate cancer. Livin gene encodes two splicing variants, termed Livin-α and Livin-β. We hypothesized that deregulation of proliferation could be due in part to Livin expression. Pathological analysis of Livin was performed in 20 prostate cancer tissues and 5 benign prostatic hyperplasia tissues. The expression of Livin isoforms was also investigated by Western blot in prostate cancer cell lines LNCaP and PC3. The role of Livin-α in vitro was further studied. Using Livin-α knockdown and overexpression models, cell cycle analysis, Ki-67 immunocytostaining, and MTT assay were performed respectively. Livin expression positive ratio was shown to be 5.4%, 23.6%, 52.4%, 73.4% in benign prostatic hyperplasia, low, medium, and high grade of prostate cancer respectively, and Livin was positively correlated with clinical pathological grades of prostate cancer. Livin-α was expressed in both LNCaP and PC3; meanwhile; Livin-β was only detected in the PC3. Livin-α siRNA not only resulted in G(1)-S cell cycle arrest, but also strongly correlated with the descended proliferation index and survival rate in LNCaP. In comparison, overexpression of Livin-α resulted in an accelerated S phase entry combined with elevated proliferation index and survival in LNCaP. Livin-α may promote cell proliferation by regulating G(1)-S cell cycle transition and possibly play an important part in initiation of prostate cancer.

Construction and identification of gene vector expressing PTEN while simultaneously silencing Livin

To study the effect of simultaneously increasing PTEN gene expression and inhibiting Livin gene expression on the gastric carcinoma cell line (BGC823) and construct a recombinant vector expressing PTEN while simultaneously silencing Livin. The siRNA expression unit against Livin gene (siLivin) was cleaved from pRNAT-U6.1-Livin vector and then inserted into pCL-neo-PTE to construct the recombinant vector pCL-neo-PTEN-siLivin. Then pCL-neo-PTEN-siLivinp, pCL-neo-PTEN, pRNAT-U6.1-Livin, pCL-neo and pRNAT-U6.1 were respectively transfected into the gastric carcinoma cell line (BGC823) with LipofectAMINE(TM) 2000. The mRNA and protein expression level of PTEN and Livin genes in each cell group was detected by fluorescent quantitative RT-PCR and Western blot. Recombinant vectors of pCL-neo-PTEN, pRNAT-U6.1-Livin and pCL-neo-PTEN-siLivin were constructed successfully. After transfection with pCL-neo-PTEN-siLivin, the mRNA and protein expression level of PTEN (0.897±0.112) rose in BGC823 cells while Livin gene became silenced. And the characterization of regulated cell bioactivity improved. There were significant differences between transfected and control groups (P<0.05). And the inhibiting effect on the proliferation and metastasis of BGC823 cell by increasing PTEN expression and silencing Livin simultaneously was better than that only by regulating PTEN genes or Livin genes alternatively. The recombinant vector of expressing PTEN and silencing Livin gene simultaneously is successfully constructed.

The studies on regulating the expression of PTEN and Livin genes in gastric carcinoma cell line BGC823 and characterization of their bioactivity

Objective To study the effect of simultaneously increasing PTEN gene expression and inhibiting Livin expression on the proliferation, apoptosis, invasion and metastasis of gastric carcinoma cell line (BGC823) in vitro. Methods To construct the eukaryotic expression vector pCL-neo-PTEN and siRNA vector pRNAT-U6, 1-Livin. The siRNA expression unit against Livin gene (siLivin) was cut from pRNAT-U6, 1-Livin vctor, and then inserted into pCL-neo-PTE to construct the recombinant vector pCL-neo-PTEN-siLivin. Then, pCL-neo-PTEN-siLivinp, pCL-neo-PTEN, pRNAT-U6, 1-Livin, pCL-neo,pRNAT-U6. 1 were respectively transfected into the gastric carcinoma cell hne (BGC823) with LipofectAMINE 2000TM. The mRNA and protein expression level of PTEN and Livin genes in each group cells was detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting, Cell proliferation curve, the activity of Caspase-3 and 9, cell apoptosis, ability of invasion and metastasis were detected in vitro. Results Recombants vector of pCL-neo-PTEN, pRNAT-U6, l-Livin and pCL-neo-PTEN-siLivin were constructed successfully. After transfected with pCL-neo-PTEN-siLivin, the mRNA (0. 897±0. 112) and protein expression level of PTEN in BGC823 cells were rised,meanwhile, Livin genes were silenced (0. 071 ±0. 009). And characterization of the regulated cells bioactivity has improved, such as the reduced proliferation ,higher apoptosis ( 16. 72±1.84) ,more effective activity of Caspase-3 and 9 ( 1.88 ±O. 21,1.07 ± 0. 18 ), fewer cells penetrating the Matrigel (23. 1 ± 3. 5 ). There were significant differences between transfected groups and control groups ( P < 0. 05 ). And the inhibiting effect on the BGC823 cells' proliferation and metastasis by increasing PTEN expression and silencing Livin simultaneously was better than that only by regulating PTEN genes or Livin genes alternatively. Conclusion The PTEN gene upregulated and the Livin gene silenced at same time could contribute to reduce the proliferation and metastasis in gastric carcinoma cell line BGC823. Key words: PTEN; Livin; Gastric carcinoma cell line BGC823; Regulating; Growth inhibition

Inhibition of tumorigenesis and invasion of hepatocellular carcinoma by siRNA-mediated silencing of the livin gene

Livin, a member of the inhibitor of apoptosis protein family, is overexpressed in a variety of solid tumors and cancer cell lines. Livin overexpression has been reported in hepatocellular carcinoma (HCC), suggesting the biological significance of livin in HCC progression. However, the mechanisms of livin and the consequence of its down-regulation in HCC have not been fully elucidated. A small interfering RNA (siRNA) eukaryotic expression vector specific to livin was constructed by gene recombination, and the nucleic acid was sequenced. It was then transfected into the human HCC cell line SMMC-7721. RT-PCR and Western blotting were used to validate the gene-silencing efficiency of livin in SMMC-7721 cells. Stable clones were obtained by G418 screening. Using multiple cellular and molecular approaches, such as an apoptosis assay, MTT assay, flow cytometry, Western blotting and a migration assay, the effects of livin inhibition on cell growth, migration and the induction of apoptosis in SMMC-7721 cells were observed. The siRNA eukaryotic expression vector specific to livin was constructed by gene recombination, and efficiently decreased the mRNA and protein expression of livin. The targeted inhibition of livin strongly sensitized SMMC-7721 cells to pro-apoptotic stimuli, and was associated with caspase-3 activation. In addition, the MMT assay indicated that the silencing of livin inhibited the cell growth of SMMC-7721 cells by specifically inhibiting cell mitosis. The results also showed that the silencing of livin strongly reduced the invasive capacity of SMMC-7721 cells. The findings suggest that livin expression not only provides HCC cells with increased resistance to apoptotic stimuli, but also contributes significantly to the proliferation and invasive capacity of HCC cells. Inhibition of livin may be a potential targeted approach for the treatment of HCC.

Suppression of Livin Gene Expression by siRNA Leads to Growth Inhibition and Apoptosis Induction in Human Bladder Cancer T24 Cells

Apoptosis deficiency is a hallmark of many cancer cells. Functional suppression of specific antiapoptotic factors might provide a feasible strategy in cancer gene therapy. Livin, the latest found inhibitor of apoptosis protein (IAP) family member, plays important role in cell growth and apoptosis. It has been reported that Livin is highly expressed in bladder cancer tissues. In this study, we found that, unlike other cancer cell lines, there was only Livin-alpha not Livin-beta expression in bladder cancer cell lines. We further investigated the effects of Livin knockdown on human bladder cancer T24 cell growth and apoptosis. We found that small interfering RNA (siRNA) mediated Livin suppression significantly inhibited T24 cell proliferation and colony formation ability. Livin knockdown dramatically increased the T24 cell apoptotic rate in response to different proapoptotic stimuli, such as Mitomycin and TNF-alpha, and this was associated with caspase-3 and caspase-9 activation. These results suggest that Livin knockdown can inhibit cell growth and increase sensitivity to apoptotic stimuli, and might serve as a potent target in bladder cancer gene therapy.

Livin gene plays a role in drug resistance of colon cancer cells

Objective The objective of this study was to investigate the effect of knockdown of Livin expression on reversing drug resistance phenotype of colon cancer HCT-8/V cells. Design and methods Specific short hairpin RNA (shRNA) was chosen and transfected in human colon cancer HCT-8/V cell line. Cell apoptosis and chemosensitivity were evaluated following downregulation of Livin expression. Results In the current study, Livin was found to be highly expressed in the HCT-8/V colon cancer cells, which were resistant to several anti-tumor drugs. Knocking down of Livin expression in HCT-8/V cells by specific RNAi facilitated the apoptosis of HCT-8/V cells in response to vincristine (VCR), etoposide (VP-16), and 5-flourouracil (5-FU). Chemosensitivity assay confirmed the results and demonstrated the reversal of drug resistance phenotype of HCT-8/V cells. Conclusion These data suggest that specific silencing of Livin gene expression could be a promising target for further research in clinical chemotherapy of colon cancer.

The expression and significance of Smac, Livin and Caspase-3 gene in osteosarcoma

Objective To explore the expression of Samc, Livin and Caspase-3 genes in osteosar-coma,and its effect on biological behaviors of osteosarcoma. Methods Using immunohistochemistry meth-od (SABC), the expression of Smac, Livin and Caspase-3 protein was detected in 46 cases of osteosarco-mas. The relationship between the expression of Smac and Livin genes with clinicopathological parameters of osteosarcomas was analyzed. Results There was expression of Smac, Livin and Caspase-3 genes in 29 cases (63.0%), 30 cases (65.2%) and 32 cases (72.4%) in osteosarcomas, respectively. The expres-sion rate of Smac and Livin had no relationship with histologic staging and WHO types of osteosarcoma, but correlated with metastasis (P<0.05). Smac had a positive correlation with Livin expression (r=0.639,P<0.01), but had no relationship with Caspase-3 expression in osteosarcomas. Conclusion Apoptosis correlated factors, Smac, Livin and Caspase-3 were all highly expressed in osteosarcoma, and maybe they arc involved in the occurrence and development of osteosarcoma by modulating apoptosis. Key words: Osteosarcoma; Caspase-3; Apoptosis

Expression of livin in gastric cancer and induction of apoptosis in SGC-7901 cells by shRNA-mediated silencing of livin gene

Background Because of increased resistance to apoptosis in tumor cells, inhibition of specific anti-apoptotic factors may provide a rational approach for the development of novel therapeutic strategies. Livin, a novel inhibitor of apoptosis protein family, has been found to be expressed in various malignancies and is suggested to have poorly prognostic significance. However, no data are available concerning the significance of livin in gastric cancer. In this study, we detected the expression of livin in human gastric carcinoma and investigated the apoptotic susceptibility of SGC – 7901 cell by shRNA-mediated silencing of the livin gene. Methods The mRNA and protein expression of livin were analyzed by RT-PCR and western blot assay. The relationship between livin expression and clinical pathologic parameters was investigated. The small interfering RNA eukaryotic expression vector specific to livin was constructed by gene recombination, and the nucleic acid was sequenced. Then it was transfected into SGC-7901 cells by Lipofectamin 2000. RT-PCR and Western blot assay were used to validate gene-silencing efficiency of livin in SGC-7901 cells. Stable clones were obtained by G418 screening. The cell apoptosis was assessed by flow cytometry (FCM). Cell growth state and 50 % inhibition concentration (IC50) of 5-FU and cisplatin was determined by MTT method. Results The expression of livin mRNA and protein were detected in 19 of 40 gastric carcinoma cases (47.5%) and SGC-7901 cells. No expression of livin was detected in tumor adjacent tissues and benign gastric lesion. The positive correlation was found between livin expression and poor differentiation of tumors as well as lymph node metastases ( P < 0.05). Four small interfering RNA eukaryotic expression vector specific to livin were constructed by gene recombination. And one of them can efficiently decrease the expression of livin, the inhibition of the gene was not less than 70% ( P < 0.01). The recombinated plasmids were extracted and transfected gastric cancer cells. The stable clones were obtained by G418 screening, and were amplified and cultured. When livin gene was silenced, the reproductive activity of the gastric cancer cells was significantly lower than the control groups( P < 0.05). The study also showed that IC50 of 5-Fu and cisplatin on gastric cancer cells treated by shRNA was decreased and the cells were more susceptible to proapoptotic stimuli (5-Fu and cisplatin) ( P < 0.01). Conclusions Livin is overexpressed in gastric carcinoma with a relationship to tumor differentiation and lymph node metastases, which is suggested to be one of the molecular prognostic factors for some cases of gastric cancer. ShRNA can inhibit livin expression in SGC-7901 cells and induce cell apoptosis. Livin may serve as a new target for apoptosis-inducing therapy of gastric cancer.

Effects of Livin gene RNA interference on apoptosis of cervical cancer Hela cells and enhanced sensitivity to cisplatin

The recombinant plasmids pGenesil-1-BIRC71 and pGenesil-1-BIRC72 were transfected into HeLa cells and cisplatin was added with different concentrations in order to study the inhibitory effects of Livin gene, increase the apoptosis induced by cisplatin, and detect the expression of Bcl-2, Bax, caspase-3, and survivin genes. The pGenesil-1-BIRC71 and pGenesil-1-BIRC72 were transfected into HeLa cells, and the expression levels of Livin, Bcl-2, Bax, caspase-3, and survivin genes were detected by using fluorescence quantitative real-time PCR. Then cisplatin at different concentrations (3.0, 6.0 and 9.9 microg/mL) was added into the transfected HeLa cells, and 24, and 48 h later, the apoptosis rate was measured by flow cytometry. After transfection of pGenesil-1-BIRC71 and pGenesil-1-BIRC72 into HeLa cells, the expression level of Livin gene was obviously reduced, and the apoptosis rate was significantly increased in transfection group as compared with control group (P<0.05). Cisplatin could increase the apoptosis rate in a dose- and time-dependent manner. After cisplatin was added, the expression levels of Bcl-2 mRNA were reduced, and those of Bax, caspase-3, and survivin mRNA were increased in transfection group as compared with those in control group (P<0.05). It was concluded that shRNA expression vector targeting Livin gene could inhibit the expression of Livin gene in HeLa cells and enhance the apoptosis induced by cisplatin, which was related to the decreased expression of Bcl-2 and activation of Bax and caspase-3. Survivin might play an important role as an antagonist in the process of apoptosis induction.

The effect of enhancement chemotherapy sensitivity by combining siRNA recombinant expression vector targeting Livin gene with 5-Fu To human colon cancer cells

Objective To construct SiRNA recombinant expression vector targeting Livin gene and research the effect of it in inhibition of human colon cancer cells proliferation and enhancement of chemotherapy sensitivity.Methods SiRNA recombinant expression vector targeting Livin gone Was constructed and transfected into human colon cancer cell.The effect of SiRNA recombinant expression vector was detected by RT-PCR and Western blot.Using auto-enzyme-labeled reading device with the 490nm wave,the value of A in each foramen was respectively measured after 24,48,72 hr of transfection by MTT reduction assay,and then calculated the inhibitor rate of tumor cell.After 48 hours of transfection,the change of the apoptosis rate was detected under the 488nm wave by flow cytometry.Results It was confirmed by restriction endonuclease and sequence analysis that SiRNA recombinant expression vector targeting Livin gene was constructed successfully.Inhibition ratio of Livin SiRNA at mRNA and protein levels were 30.18% and 28.88%.It could enhance chemotherapy sensitivity by combining 5-Fu.Conclusion The SiRNA recombinant expression vector targeting Livin gene has been constructed successfully and can inhibite the expression of Livin gene.It also can enhance chemotherapy sensitivity and apoptosis induction effect by combining 5-Fu in human colon cancer cells significantly. Key words: Livin; RNA interference; Gene therapy; Chemotherapy