| Background Renal cell carcinoma (RCC) is one of the most common cancer in urinary system. The incidence and mortality is the second after bladder cancer. Recent epidemiological investigations have show the incidence of renal cell carcinoma has been increasing worldwide. The treatment of advanced stage, recurrence and metastasis RCC were the difficult problem in clinic. RCC is highly resistant to systemic chemotherapy, and no agent should be considered the standard for treating metastatic RCC..The development of novel and effective therapeutic strategies for metastatic RCC is urgently needed. Previous studies have shown that BTV induces apoptosis in infected several cancer cells and it has the potential of becoming an oncolytic virus. However the efficacy of BTV in renal cancer cells has not been fully determined and whether the death pathways of the different cells were the same were not determined. In the present study, we investigated the anticancer activity of BTV against RCC using primary human renal tubular epithelial cells (RTEc) and two RCC cell lines (A498 and Renca). We created up the animal model of subcutaneous RCC using the BALB/c mice in order to verity its selective infection to the RCC in vivo. We also explored the molecular mechanisms that might be involved in the anticancer activity of BTV in RCC cells.Objective To study the different characteristic of BTV-HbC3 infected primary human renal tubular epithelial cells(RTECs), mouse renal carcinoma cells (Renca) and human renal carcinoma cells(A498). To verity its selectively infection to the tumor cells in vivo. To investigate the mechanism of Bluetongue virus-HbC3 induced A498 and Renca cell death.Methods Primary human RTEc were established and identified by Transmission electron microscopy. RTECs, A498 and Renca cells were infected with BTV-HbC3 at a multiplicity of infection (MOI) of 1. Observe the cytopathic effects (CPE) by microscopy and the ultra-structural changes by transmission electron. The presence of BTV genome was also detected by agarose gel electrophoresis. MTT assay was used to comparative analyses of differential survival rates using different Multiplicities of Infection after the three cells were infected 24h,36h, and 48h. The subcutaneous tumor-bearing mice model of renca cells were created up successfully using the BALB/c mice, and the mice was intratumorally injected with BTV-HbC3. Observed the body weight, tumor growth and the pathological and immunohistochemical of the tumor tissue. We used TCID50 assay to measure the Virus multiplication after infected A498 and Renca cells in 60h. The early apoptosis of the three cells which were induced by BTV at 1MOI were detected with Annexin V-FITC/PI assay. The DNA fragments of the post infection A498 and Renca cell were also measured. FCM analyzed each group's cell-cycle changes. The changes of nuclear and endoplasmic reticulum of the two post infection cancer cell were detected by fluorescence microscopyResults Primary RTEc were established and identified successfully. Over 80% CPE was observed at the postinfection A498 and Renca cell. The electron microscopy examination showed characteristics of necrocytosis and lots of BTV-HbC3 appeared intra-cellular cytoplasm. But the characteristic of the primary RTEC was not changed by phase contrast microscope and Transmission electron microscopy examination. BTV viral genomic ds-RNA fragments could be found in BTV infected A498 and Renca cells through gel electrophoresis. The survival rate of three cells which infected with BTV at 10 MOI were 99.25±3.27%,25.69±3.50% and 22.63±5.85% respectively. Compared with the RTEc, the survival rates of the A498 and renca cell were significant difference when infected with BTV at MOI of 1 in same postinfection time. It showed that there was a dose-effect and time-effect relationship between them. The tumor volume reduced dramatically in contrast with the control group. After BTV treatment Subsequent pathological and immunohistochemical analysis also demonstrated its distinct infection restricted in the Renca cells. Apoptotic DNA fragments were detected in A498 cells and Renca cells. Comparing with control cells, cell numbers of postinfection in S stages increased. There were apoptotic body formation, chromatin condensation Vacuoles were derived predominantly from the endoplasmic reticulum in postinfection A498 and Renca cells, the florescence of the normal control cells and infected cell were 56.396±7.658,33.497±5.836(P<0.01) in A498cells and 70.146±4.754,42.291±6.893(P<0.01)respectively in Renca cells.Conclusion BTV-HbC3 could not infect the primary human RTECs instead selectively degradated A498 and Renca cells. It showed that there was a dose-effect and time-effect relationship between them. Bluetongue virus-HbC3 could also anti-tumor in vivo. It could induce A498 and Renca cells death by apoptotic pathways. BTV has the potential of becoming a new oncolytic virus.
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