Survivin Gene Expression Research Articles

Characterization of Modified Antisense Oligonucleotides inXenopus laevisEmbryos

A wide variety of modified oligonucleotides have been tested as antisense agents. Each chemical modification produces a distinct profile of potency, toxicity, and specificity. Novel cationic phosphoramidate-modified antisense oligonucleotides have been developed recently that have unique and interesting properties. We compared the relative potency and specificity of a variety of established antisense oligonucleotides, including phosphorothioates (PS), 2'-O-methyl (2'OMe) RNAs, locked nucleic acids (LNAs), and neutral methoxyethyl (MEA) phosphoramidates with new cationic N,N-dimethylethylenediamine (DMED) phosphoramidate-modified antisense oligonucleotides. A series of oligonucleotides was synthesized that targeted two sites in the Xenopus laevis survivin gene and were introduced into Xenopus embryos by microinjection. Effects on survivin gene expression were examined using quantitative real-time PCR. Of the various modified oligonucleotide designs tested, LNA/PS chimeras (which showed the highest melting temperature) and DMED/phosphodiester chimeras (which showed protection of neighboring phosphate bonds) were potent in reducing gene expression. At 40 nM, overall specificity was superior for the LNA/PS-modified compounds compared with the DMED-modified oligonucleotides. However, at 400 nM, both of these compounds led to significant degradation of survivin mRNA, even when up to three mismatches were present in the heteroduplex.

Effects of human papillomavirus type 16 oncoproteins on survivin gene expression

Survivin has recently been identified as a novel member of the inhibitor of apoptosis (IAP) gene family. The product of this gene not only suppresses apoptosis but also controls cell division. Survivin is undetectable in most terminally differentiated normal tissues but is expressed in embryonic and fetal organs and is present in most malignant tumours. Human papillomaviruses (HPV) are thought to play an important role in the development of cervical cancer. By interfering in the cell cycle, the viral oncoproteins (E6 and E7) can induce the immortalization of the host cell. The transcriptional effects of the HPV-16 E6 and E7 proteins on the survivin promoter in transiently transfected cell lines using luciferase tests were examined. HPV-16 E6, but not E7, was found to significantly transactivate the survivin promoter. Experiments performed in different cancer cell lines and with different E6 mutants indicated that the effect of E6 on the survivin promoter is largely dependent on p53 status. In accordance with this, the p53 tumour suppressor protein downregulated the expression of survivin. As E6 is able to interact with p53 and induces its ubiquitin-dependent degradation, it appears that the transactivation effect of E6 on survivin is mediated by the p53 degradation pathway. Transduction of HPV-16 E6 and E7 into human embryonic fibroblast cells showed that the HPV oncoproteins can upregulate endogenous survivin mRNA. Importantly, cell cycle synchronization experiments showed that the effect of HPV-16 E6 on survivin transcription is independent of the cell cycle.

Delivery of different length poly( l-lysine)-conjugated ODN to HepG2 cells using N-stearyllactobionamide-modified liposomes and their enhanced cellular biological effects

Short (14–20-mer range) synthetic oligodeoxynucleotides (ODNs) allow specific modulation of cellular gene expression at various stages, thus providing a versatile tool for fundamental studies and a rational approach to anticancer chemotherapy. However, several problems, such as metabolic stability, efficient cell internalization of ODNs and their efficient entrapment into liposomes continue to markedly limit this approach. To improve the target specificity and biological activity of ODN, three different length of poly( l-lysine) (PLL) were conjugated to ODN and these conjugates were encapsulated in N-stearyllactobionamide (N-SLBA)-modified liposomes, N-SLBA is a ligand for the asialoglycoprotein receptor. Then, we investigated their effects on cell cycle and survivin protein levels of HepG2 cells. The results showed that the encapsulation efficiency was improved because the polycationic charges of PLL neutralized the polyanionic charges of ODN. Among them, PLL ( M W 2000 and 10,000)-conjugated ODN encapsulated in N-SLBA liposomes induced apoptosis of HepG2 cells and highly inhibited survivin gene expression.

Expression of TR6 and survivin gene in the tissue of cardiac cancer and its clinical significance

OBJECTIVE:To explore the expression of Survivin gene inhibiting apoptosis in the tissue of cardiac cancer and its correlation with clinicopathological features. METHODS: Two-step immunohistochemistry technology (Envision) was adopted to detect the expression of Survivin gene in 66 cases of cardiac cancer, and the relationship between the expression and clinicopathological features of cardiac cancer was investigated. RESULTS: In cancer tissue and tissues surrounding cancer, the positive expression rates of Survivin were 62.1% (41/66) and 24.2% (16/66) respectively,no Survivin expression was found in normal gastric mucosa, and the positive rates among these three groups had a statistical difference,P0.05. Survivin protein positive expression in low differentiation gastric cardiac carcinoma was 73.8% (31/42), and compared with high and moderate differentiation carcinoma positive rate 41.7% (10/24) there was a statistical difference,χ~2=5.409 4,P0.05. Surrivin protein positive expression rate was 73.3% (33/45) in those patients with lymph node metastasis,while only 38.1% (8/21) in those patients without lymph node metastasis, and a statistical difference was found, χ~2=6.132 4,P0.05. Positive Survivin expression rate was 74.4% in those patients with serosal infiltration while only 44.4% (12/27) in those patients without serosal infiltration,also there was a significant difference between them, χ~2=4.862 8,P0.05. TNM classification stage Ⅲ gastric cardiac carcinoma’s positive Survivin expression rate was 95% (19/20) and it was significantly higher than that of stage Ⅰ and Ⅱ, 47.8% (22/46), χ~2=11.254 4,P0.05. The expression of Survivin gene had a positive correlation with gastric cardiac carcinoma differentiation, clinical staging and lymph node metastasis, but it had no correlation with ages, sex and tumor size of cardiac carcinoma. CONCLUSIONS: Survivina inhibitor of apoptosis may correlate with the carcinogenesis and progression of cardiac carcinoma. The detection of Survivin can be worthwhile to judge the malignance and progression of cardiac cancer.

Persistent Activation of Stat3 Signaling Induces Survivin Gene Expression and Confers Resistance to Apoptosis in Human Breast Cancer Cells

Signal transducer and activator of transcription 3 (Stat3) protein is persistently activated in breast cancer and promotes tumor cell survival. To gain a better understanding of the role of constitutive Stat3 signaling in breast cancer progression, we evaluated the expression profile of potential Stat3-regulated genes that may confer resistance to apoptosis. Stat3 signaling was blocked with antisense oligonucleotides in human MDA-MB-435s breast cancer cells and Affymetrix GeneChip microarray analysis was done. The candidate Stat3 target gene Survivin was further evaluated in molecular assays using cultured breast cancer cells and immunohistochemistry of breast tumor specimens. Survivin, a member of the inhibitor of apoptosis protein family, was identified as a potential Stat3-regulated gene by microarray analysis. This was confirmed in Survivin gene promoter studies and chromatin immunoprecipitation assays showing that Stat3 directly binds to and regulates the Survivin promoter. Furthermore, direct inhibition of Stat3 signaling blocked the expression of Survivin protein and induced apoptosis in breast cancer cells. Direct inhibition of Survivin expression also induced apoptosis. Increased Survivin protein expression correlates significantly (P = 0.001) with elevated Stat3 activity in primary breast tumor specimens from high-risk patients who were resistant to chemotherapy treatment. We identify Survivin as a direct downstream target gene of Stat3 in human breast cancer cells that is critical for their survival in culture. Our findings suggest that activated Stat3 signaling contributes to breast cancer progression and resistance to chemotherapy by, at least in part, inducing expression of the antiapoptotic protein, Survivin.

Expression of Survivin and p53 gene and its relationship in primary hepatocellular carcinoma(HCC)

目的 探讨Survivin在原发性肝细胞癌(HCC)组织中的表达及与p53的关系.方法采用Western blot和RT-PCR检测43例肝癌组织、20例癌旁组织和11例正常肝组织中Survivin蛋白和mRNA的表达情况.免疫组化(SP法)检测Survivin和p53蛋白的表达情况.结果 43例HCC标本中分别有30例表达Survivin(69.8%),而除1例癌旁组织外,其他癌旁组织和正常肝组织中未见Survivin表达.Survivin表达与肝癌组织的病理分级、临床分期等无明显相关.Survivin表达与p53表达亦无明显相关.结论 Survivin在肝癌组织中高表达,Survivin的表达与p53基因的突变无明显关系。

The Research of Specific Inhibition of Survivin Expression To Reverse Drug Resistance with RNA Interference Technology.

Objective:Use RNA interference technology to down-regulate the expression of survivin gene in KM3 cells, observe its induction of apoptosis of KM3 cells, and if it can increase chemotherapy sensitivity of KM3 cells to adriamycin.Methods: SiRNA designed and in-vitro chemosynthesized aiming to survivin was transfected into human myeloma cell line KM3 mediated by liposome. Survivin mRNA transcription and protein expression of KM3 cells were then detected using RT-PCR and Western-blot 24, 48 and 72 hours after transfection, so as to invest the silencing effect. Besides, the apoptosis of the cell line were observed under fluorescence microscopy before and after transfection. Furthermore, cytotoxic analysis was used to compare the sensitivity variation of KM3 cells to adriamycin before and after transfection.Results: Down-regulations of survivin mRNA transcription and protein expression of KM3 cells could be observed 24 hours after siRNA tranfection, which were (12±2.3)% and (9.4±1.8)% respectively. After 48 hours, they were (61.4±7.9)% and (38.6±6.7)% respectively, while the survivin mRNA transcription and protein expression was increased quickly 72 hours later. Under fluorescence microscopy, the apoptosis rate of KM3 cells transfected with survivin siRNA 48 hours later was 28±7%, which was significantly higher than the control cells. (P<0.05). The IC50 value of KM3 cells to adriamycin decreased from 1.12±0.14uM to 0.21±0.03uM, which means the sensitivity of KM3 cells to adriamycin was 5.3 fold increased.Conclusions: SiRNA which was designed and synthesized to target survivin could effectively down-regulate survivin mRNA transcription and protein expression in KM3 cells, induce KM3 cells to apoptosis, increase its sensitivity to adriamycin after down-regulation, and effectively reverse drug-resistance of MM cells.

Survivin gene expression is negatively regulated by the p53 tumor suppressor gene in non-small cell lung cancer

Survivin is considered to be associated with tumorigenesis by regulating apoptosis and cell proliferation. Recent experimental studies reported survivin gene expression to be negatively regulated by wild-type p53. We investigated resected tumor specimens from 140 non-small cell lung cancer (NSCLC) patients. Quantitative reverse-transcription PCR analysis was performed to evaluate survivin gene expression. PCR-single strand conformation polymorphism following sequencing was performed to investigate mutations of p53. The apoptotic index and the Ki-67 proliferation index were also evaluated according to the survivin expression. The survivin expression was low in normal lung tissue. In contrast, the survivin expression varied greatly among tumor tissues. The survivin expression in squamous cell carcinomas was significantly higher than that in adenocarcinomas (P=0.0109). The survivin expression in moderately or poorly differentiated tumors was significantly higher than that in well-differentiated tumors (P=0.0334). Furthermore, the survivin expression in tumors with mutant p53 was significantly higher than that in tumors with wild-type p53 (P=0.0026). In addition, the apoptotic index was significantly lower in high-survivin tumors than in low-survivin tumors (P<0.0001). The Ki-67 proliferation index was significantly higher in high-survivin tumors than in low-survivin tumors (P<0.0047). This study indicated survivin gene expression to be negatively regulated by p53 in NSCLC, and that survivin expression could inhibit apoptosis and accelerate tumor cell proliferation to produce more aggressive carcinomas.

Survivin, a potential biomarker in the development of Barrett's adenocarcinoma

Survivin, a member of the inhibitor-of-apoptosis family, is reported to be overexpressed in esophageal cancer but no data are available about its status in the metaplastic/dysplastic sequence. The aim of this study was to measure survivin gene expression in normal squamous/columnar epithelium and in the various stages of development of Barrett's adenocarcinoma. Endoscopic biopsy or operative specimen samples from 5 tissue types were analyzed: (1) squamous epithelium from 3 cm above the gastroesophageal junction in patients with a negative pH study and no histologic injury (n = 17, pH- control); (2) antral tissue from patients with no evidence of Barrett's, dysplasia, or cancer (n = 29, antral control); (3) specialized intestinal metaplasia from patients with Barrett's esophagus (n = 16; Barrett's group); (4) low- or high-grade dysplasia (n = 12, dysplasia group), and (5) adenocarcinoma (n = 45 cancer group). After laser-capture microdissection cellular RNA was extracted from each tissue and reverse transcribed to complementary DNA. Expression levels of survivin were measured by reverse-transcription polymerase chain reaction. Survivin gene expression was greater in columnar (antral) compared with squamous (pH-) control tissues (P = .03). Expression in quiescent Barrett's epithelium was similar to both control tissues. Expression levels in dysplastic epithelium were greater than in squamous control (P = .01) and Barrett's tissues (P = .04), but not higher than columnar control tissues, whereas expression in adenocarcinoma was greater than all tissues except dysplasia (P < .001). Survivin expression may be a biomarker in the development of Barrett's adenocarcinoma that is able to distinguish between quiescent Barrett's, dysplastic Barrett's, and Barrett's adenocarcinoma.

Comparison of extracellular matrix and apoptotic markers between benign lesions and carcinomas in human breast

The expression of the extracellular matrix-related genes, such as fibronectin, laminin and tenascin C, and apoptosis-related genes, such as bax, bcl2 and survivin, was evaluated by reverse transcription-polymerase chain reaction (RT-PCR) and by immunohistochemistry in normal breast tissue and benign and malignant breast tumors and then correlated to several clinical parameters: estrogen and progesterone receptors, Ki67, ErbB2, tumor size, lymph node status and grading. Seventy-three breast tissue samples were examined. After RNA extraction, an RT-PCR was performed to detect fibronectin, laminin, tenascin C, bax, bcl2 and survivin gene expression. Thirty-two samples were evaluated also by immunohistochemistry at the protein level to detect fibronectin, laminin, tenascin C, bax and survivin. We found a significant correlation (P=0.025) between fibronectin gene expression and lymph node status, and a significant negative correlation (P=0.049) between laminin gene expression and Ki67. In addition, we found a statistically significant increase in survivin transcription in malign tumors compared to fibroadenomas (P=0.024). The negative correlation between laminin transcription and Ki67 could suggest that laminin impacts negatively on tumor proliferation, and the positive correlation between fibronectin and lymph node status may lead to consider fibronectin as predictive of long distance metastasis.

Antisense RNA targeting survivin enhances the chemosensitivity of LOVO/Adr cells to taxotere

To investigate the ability of antisense RNA eukaryotic expression plasmid pcDNA3.0/survivin targeting survivin gene to inhibit survivin expression and enhance the sensitivity to taxotere in multidrug resistant colon carcinoma cell line LOVO/Adr. The antisense RNA eukaryotic plasmid pcDNA3.0/survivin was transfected into LOVO/Adr cells by lipofectamine. The expression of survivin mRNA was measured using RT-PCR. After treated with taxotere, MTT assay and flow cytometry were used to evaluate the proliferation inhibition and apoptosis of LOVO/Adr cells. The expression of survivin mRNA in LOVO/Adr cells transfected with pcDNA3.0/survivin was down-regulated in a time- dependent manner. The inhibitory rate of taxotere (0.5 micromol/L) was (37.3 +/- 2.9)% in pcDNA3.0/survivin transfected cells, significantly higher than (21.9 +/- 2.3)% and (21.1 +/- 1.9)% in pcDNA3.0 transfected and untransfected control cells respectively (P< 0.01). The apoptosis rate of taxotere was (28.7 +/- 1.7)% in pcDNA3.0/survivin transfected cells,significantly higher than (13.4 +/- 1.6)% and (14.3 +/- 1.8)% in pcDNA3.0 transfected and untransfected cells respectively. The antisense RNA eukaryotic expression plasmid pcDNA3.0/survivin could down-regulate the expression of survivin gene and enhance the chemosensitivity of LOVO/Adr cells to taxotere, which may provide a novel therapy for colon carcinoma.

Research on relationship of survivin gene expression with malignant proliferation and apoptosis of brain glioma

To investigate the expression level of inhibitor of apoptosis protein survivin gene in human brain glioma and its role in malignant proliferation and antiapoptosis of brain glioma. Eighty-three cases of brain glioma specimen was chosen, protein expression of survivin and proliferating cell nuclear antigen (PCNA) was investigated by immunohistochemistry streptavidin-biotin complex (SABC) method, the immunoreactivity score (IRS) of survivin and the proliferative index (PI) were counted. Apoptotic cells were screened by TdT-mediated dUTP-biotin nick-end labeling (TUNEL) method, and the apoptotic index (AI) of brain glioma was calculated. The survivin IRS, PI and AI of brain glioma were 3.8 +/- 3.9, (28.4 +/- 19.5)% and (1.0 +/- 0.8)% respectively, and all of them were elevated with the increase of pathological grade of brain glioma (P < 0.01 for all). PI in survivin positive group was significantly higher than that in survivin negative group (P < 0.01), and PI was positively correlated with survivin IRS (r = 0.740, P < 0.01). There was no significant difference between AI in survivin positive group and that in survivin negative group (P > 0.05), however, AI was negatively correlated with survivin IRS (r = -0.307, P < 0.01). Survivin is overexpressed in brain glioma, and which may play important roles in malignant proliferation and antiapoptosis of brain glioma.

Systemic Treatment with Tetra-O-Methyl Nordihydroguaiaretic Acid Suppresses the Growth of Human Xenograft Tumors

We have previously shown that the transcriptional inhibitor tetra-O-methyl nordihydroguaiaretic acid (M4N) induces growth arrest in tumor cells and exhibits tumoricidal activity when injected intratumorally into tumor cell explants in mice. The experiments reported here were designed to determine whether M(4)N can be given systemically and inhibit the growth of five different human xenograft tumors. Nude (nu/nu) mice bearing xenografts of each of five human tumor types (i.e., hepatocellular carcinoma, Hep 3B; prostate carcinoma, LNCaP; colorectal carcinoma, HT-29; breast carcinoma, MCF7; and erythroleukemia, K-562) were treated with M4N given i.v. or i.p. in a Cremophor EL-based solvent system or orally in a corn oil based diet. Tumors from the treated animals were measured weekly and analyzed for the expression of the Cdc2 and survivin genes, both previously shown to be down-regulated by M4N. Systemic M4N treatment suppressed the in vivo growth of xenografts in each of the five human tumor types. Four of the five tumor models were particularly sensitive to M4N with tumor growth inhibitions (T/C values) of < or = 42%, whereas the fifth, HT-29, responded to a lesser extent (48.3%). Growth arrest and apoptosis in both the xenograft tumors and in the tumor cells grown in culture were accompanied by reductions in both Cdc2 and tumor-specific survivin gene expression. Pharmacokinetic analysis following oral and i.v. administration to ICR mice indicated an absolute bioavailability for oral M4N of approximately 88%. Minimal drug-related toxicity was observed. These preclinical studies establish that when given systemically, M4N can safely and effectively inhibit the growth of human tumors in nude mice.

Expression of Survivin Gene and Its Relationship with Clinical Multidrug Resistance in Osteosarcoma

Objective: To study the expression of survivin and its relationship with clinical multidrug resistance in osteosarcoma. Methods: By using immunohistochemistry (S-P) method, the expression of Survivin in osteosarcoma, osteochondroma and normal osseous tissue, and the expression of P-glycoprotein in osteosarcoma was detected. Results: Survivin positive expression rate was 65.71% in osteosarcoma, but no expression of Survivin was detectable in osteochondroma and normal osseous tissue. The positive expression rate of Survivin was significantly associated with Enneking clinical stages and histological typing (WHO), but no relationship was found among Survivin expression and age, sex and tumor location. The positive expression rate of P-glycoprotein was 45.71%. There was a significant correlation between Survivin and p-glycoprotein. Conclusion: Survivin overexpression was significantly associated with clinical multidrug resistance in osteosarcoma. It could be a potential target for treatment of osteosarcoma.

Differential roles for the coactivators CBP and p300 on TCF/β-catenin-mediated survivin gene expression

The inhibitor of apoptosis (IAP) protein survivin is highly expressed in cancers, but not in normal differentiated tissues. TCF/beta-catenin signaling has been reported to participate in the regulation of survivin transcription in colon cancer. We have recently characterized ICG-001, a small molecule specific inhibitor of the beta-catenin/Creb-binding protein (CBP) interaction. Inhibition of the beta-catenin/CBP interaction represses a subset of TCF/beta-catenin-mediated transcription. ICG-001 potently inhibits survivin gene transcription and expression. ICG-001-mediated downregulation of survivin expression enhanced caspase-3 activity and apoptosis, which was rescued by overexpression of wild type but not mutant (C84A) survivin. Small interfering RNA and genetic reduction of CBP also decreased survivin expression. Chromatin immunoprecipitation assay confirmed that CBP is the crucial coactivator for TCF/beta-catenin-mediated survivin transcription. Furthermore, ICG-001-induced recruitment of p300 to the survivin promoter led to concomitant recruitment of SUMO-1, HDAC6 and PML proteins, which have been associated with transcriptional repression. These findings demonstrate that CBP and p300 play very distinct roles in survivin gene transcription.

Molecular Mechanism of Inhibition of Survivin Transcription by the GC-rich Sequence-selective DNA Binding Antitumor Agent, Hedamycin: EVIDENCE OF SURVIVIN DOWN-REGULATION ASSOCIATED WITH DRUG SENSITIVITY

Expression of the antiapoptotic protein survivin is associated with cancer cell viability and drug resistance. Thus, control of its expression in cancer cells has significant consequences for cancer therapeutics. Here we have shown that hedamycin, a GC-rich DNA binding drug, down-regulated survivin expression. Using a series of survivin promoter-luciferase constructs, we have identified an 86-bp GC-rich DNA element (-124 to -39) that mediates the ability of hedamycin to down-regulate survivin expression. Furthermore, both in vivo foot-printing and in vitro gel mobility shift experiments revealed that hedamycin bound to a 21-bp GC-rich DNA element (-115 to -95) in the survivin promoter. This drug-DNA interaction abrogated the binding of Sp-1 or Sp1-like proteins to the 21-bp cis-acting DNA element, and mutagenesis of this region consistently diminished survivin promoter activity. Finally, down-regulation of survivin transcription by hedamycin modulated the viability of cancer cells. These data suggest that abrogation of Sp-1 or Sp1-like protein binding to the 21-bp DNA element in the survivin promoter contributes at least in part to the inhibitory effect of hedamycin on survivin gene transcription. Drug-induced modulation of survivin gene expression may provide novel approaches for cancer therapeutics.

Real-time Detection of Gene Expression in Cancer Cells Using Molecular Beacon Imaging: New Strategies for Cancer Research

Development of novel approaches for quantitative analysis of gene expression in intact tumor cells should provide new means for cancer detection and for studying the response of cancer cells to biological and therapeutic reagents. We developed procedures for detecting the levels of expression of multiple genes in fixed as well as viable cells using molecular beacon imaging technology. We found that simultaneous delivery of molecular beacons targeting survivin and cyclin D1 mRNAs produced strong fluorescence in breast cancer but not in normal breast cells. Importantly, fluorescence intensity correlated well with the level of gene expression in the cells detected by real-time reverse transcription-PCR or Western blot analysis. We further show that molecular beacons can detect changes of survivin gene expression in viable cancer cells following epidermal growth factor stimulation, docetaxel treatment, and overexpression of p53 gene. Thus, molecular beacon imaging is a simple and specific method for detecting gene expression in cancer cells. It has great potential for cancer detection and drug development.

Expression of survivin gene in pancreatic carcinoma

To determine the expression status of survivin gene in pancreatic carcinoma. Expression of survivin gene was evaluated by immunohistochemistry, Western Blot and RT-PCR in 59 cases of pancreatic carcinoma along with their corresponding adjacent benign tissues, 11 cases of chronic pancreatitis, and 7 pancreatic carcinoma cell lines. The positive expression rate of survivin in pancreatic carcinoma was 72.8% (43/59). There was no relationship between the expression of survivin and tumor stage and differentiation. No expression of survivin was detected in benign tissue adjacent to the tumors as well as in samples of chronic pancreatitis. All 7 pancreatic carcinoma cell lines showed a positive expression of survivin mRNA and protein. The expression of survivin appears to be tumor specific to some extent in our pancreatic carcinoma samples. Survivin may be an ideal target for therapy against pancreatic carcinoma.

In vivo expression patterns of survivin and its splicing variants in chronic liver disease and hepatocellular carcinoma

Our aim was to clarify the significance of expression levels and post-transcriptional splicing patterns of survivin during multistep hepatocarcinogenesis and tumor progression. Using immunohistochemistry, we first elucidated the expression of survivin protein in tissues of hepatocellular carcinoma (HCC) and the adjacent non-cancerous tissues. Furthermore, we investigated survivin gene expression patterns in these tissues. Survivin protein was expressed not only in most HCC tissues but also in some cirrhotic nodules. In non-cancerous regions, the levels of survivin mRNA increased in proportion to their stage of progression. Survivin protein was expressed mainly in periportal areas, where proliferating cells were localized. In HCC, mRNA levels of survivin and survivin Delta Ex3 correlated with high proliferative activity, whereas the levels of surviving 2B did not. These findings of mRNA and protein expressions of survivin in chronically injured avers indicate that it has an important role in hepatocarcinogenesis. A lack of correlation between proliferative activity and survivin 2B mRNA levels in HCC is suggestive of a previous hypothesis that this variant decreases sruvivin function in a dominant negative manner. Thus, our data suggest different functions of these splicing variants and their important roles in tumor progression.

Knockdown of survivin gene expression by RNAi induces apoptosis in human hepatocellular carcinoma cell line SMMC-7721

To investigate the survivin gene expression in human hepatocellular carcinoma cell line SMMC-7721 and the effects of survivin gene RNA interference (RNAi) on cell apoptosis and biological behaviors of SMMC-7721 cells. Eukaryotic expression vector of survivin gene RNAi and recombinant plasmid pSuppressorNeo-survivin (pSuNeo-SVV), were constructed by ligating into the vector, pSuppressorNeo (pSuNeo) digested with restriction enzymes Xba I and Sal I and the designed double-chain RNAi primers. A cell model of SMMC-7721 after treatment with RNAi was prepared by transfecting SMMC-7721 cells with the lipofectin transfection method. Strept-avidin-biotin-complex (SABC) immunohistochemical staining and RT-PCR were used to detect survivin gene expressions in SMMC-7721 cells. Flow cytometry was used for the cell cycle analysis. Transmission electron microscopy was performed to determine whether RNAi induced cell apoptosis, and the method of measuring the cell growth curve was utilized to study the growth of SMMC-7721 cells before and after treatment with RNAi. The eukaryotic expression vector of survivin gene RNAi and pSuNeo-SVV, were constructed successfully. The expression level of survivin gene in SMMC-7721 cells was observed. After the treatment of RNAi, the expression of survivin gene in SMMC-7721 cells was almost absent, apoptosis index was increased by 15.6%, and the number of cells was decreased in G2/M phase and the cell growth was inhibited. RNAi can exert a knockdown of survivin gene expression in SMMC-7721 cells, and induce apoptosis and inhibit the growth of carcinoma cells.