CAJ | 학술논문

目的观察蓝舌病毒湖北株(BTV-HBC3)对小鼠肾癌细胞株Renca体内外的杀伤效应.方法接种1 MOI的BTV-HBC3于Renca细胞,观察Renca感染后的细胞病变效应(CPE),应用扫描电镜观察感染48 h后的超微结构,以噻唑蓝(MTT)比色法检测病毒分别以不同感染复数(MOI)感染Renca细胞后24、36、48 h的细胞存活率,并应用琼脂糖凝胶电泳分析感染后48 h的Renca细胞及其对照组病毒基因组.同时制备BALB/C小鼠Renca细胞荷瘤动物模型,观察应用BTV-HBC3处理后皮下荷瘤生长,并进行苏木素-伊红(HE)染色病理分析,应用免疫组织化学检测病毒蛋白的表达.结果 Renca细胞感染BTV-HBC348 h后,CPE程度大于90%,透射电镜下,胞质内可见增殖的病毒颗粒,呈晶格状排列.Renca细胞接种1 MOI的BTV-HBC3 24、36、48 h后细胞的存活率分别为(44.45±5.26)%、(30.47±4.77)%、(20.34±5.26)%,且该病毒对Renca细胞的杀伤作用呈现一定的量效关系;感染后细胞进行琼脂糖电泳可以检测到特异性长(L)、中(M)、短(S)3组分片段的病毒的dsRNA基因组;体内实验表明BTV-HBC3也明显抑制肿瘤生长(P<0.01);组织病理学观察中肿瘤组织未见坏死区,可见大片透明空泡样组织,肿瘤组织内可见腺体样结构,而小鼠其他各脏器组织切片中未见病毒感染,治疗组肿瘤组织可见病毒蛋白的阳性染色,其余组织器官经免疫组织化学分析未见病毒蛋白表达.结论蓝舌病毒湖北株对小鼠肾癌细胞体内外具有显著的靶向杀伤效应.
목적 관찰람설병독호북주(BTV-HBC3)대소서신암세포주Renca체내외적살상효응.방법 접충1 MOI적BTV-HBC3우Renca세포,관찰Renca감염후적세포병변효응(CPE),응용소묘전경관찰감염48 h후적초미결구,이새서람(MTT)비색법검측병독분별이불동감염복수(MOI)감염Renca세포후24、36、48 h적세포존활솔,병응용경지당응효전영분석감염후48 h적Renca세포급기대조조병독기인조.동시제비BALB/C소서Renca세포하류동물모형,관찰응용BTV-HBC3처리후피하하류생장,병진행소목소-이홍(HE)염색병리분석,응용면역조직화학검측병독단백적표체.결과 Renca세포감염BTV-HBC348 h후,CPE정도대우90%,투사전경하,포질내가견증식적병독과립,정정격상배렬.Renca세포접충1 MOI적BTV-HBC3 24、36、48 h후세포적존활솔분별위(44.45±5.26)%、(30.47±4.77)%、(20.34±5.26)%,차해병독대Renca세포적살상작용정현일정적량효관계;감염후세포진행경지당전영가이검측도특이성장(L)、중(M)、단(S)3조분편단적병독적dsRNA기인조;체내실험표명BTV-HBC3야명현억제종류생장(P<0.01);조직병이학관찰중종류조직미견배사구,가견대편투명공포양조직,종류조직내가견선체양결구,이소서기타각장기조직절편중미견병독감염,치료조종류조직가견병독단백적양성염색,기여조직기관경면역조직화학분석미견병독단백표체.결론 람설병독호북주대소서신암세포체내외구유현저적파향살상효응.
Objective To investigate the anti-tumor effect of Bluetongue virus-HBC3 on murine renal cell carcinoma cell ( Renca) in vitro and in vivo. Methods The renal cells were inoculated with BTVHBC3 at MOI of 1. Observed the cytopathic effects (CPE) by microscopy and the ultra-structural changes by transmission electron. MTT assay was used to comparative analyses of differential survival rates using different Multiplicities of Infection ( MOI) after the renal cells were infected 24,36,48 h. The presence of BTV genome was also detected by agarose gel electrophoresis. 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 tumor growth, and the pathological and immunohistochemical of the tumor tissue. We also detected the serum IFN-α of the mice. Results Over 90% cytopathic effects (CPE) were readily observed and detected in renca cells infected with BTV 48h postinfection. In electron microscopy (EM), BTV virions could be seen amplified efficiently in renca cells. The survival rate of the renca cell were respectively (44.45 ± 5. 26) % , (30. 47 ± 4. 77)% , ( 20. 34 ± 5. 26 ) % when infected with BTV at MOI of 1 24,36,48 h postinfection. It showed that there was a dose-effect relationship between them. BTV viral genomic ds-RNA fragments could be found in BTV infected Renca cells through gel electrophoresis. 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 Rencacells. Conclusion BTV-HBC3 could selectively degradated Renca cells.