蓝藻抗病毒蛋白N衍生物的细胞毒性及抗病毒活性检测

目的]研究蓝藻抗病毒蛋白N(CVN)衍生物(LCVN)对永生化上皮角质形成细胞(HaCaT)的毒性及LCVN凝胶的制备和体外活性检测。方法]用MTT法测定LCVN对HaCaT细胞株的半数毒性浓度(CC50)以及LCVN对流感病毒毒株A/HK/8/68(H3N2)的半数抑制浓度(IC50);用流式细胞检测法检测LCVN对HaCaT细胞周期与细胞凋亡的影响;用酶联免疫吸附法(ELISA)检测LCVN凝胶与HIV包膜蛋白GP120的相互作用。结果]培养24和48 h后,LCVN对HaCaT的CC50分别为(9.25±1.21)与(1.94±0.12)μmol/L,将LCVN做成凝胶后,与HIV外膜蛋白GP120的结合在一定浓度范围内具有剂量关系,LCVN凝胶对A/HK/8/68(H3N2)毒株的IC50为(98.20±0.03)nmol/L。结论]LCVN对HaCaT细胞的毒性明显小于CVN,当做成凝胶以后LCVN对流感病毒A/HK/8/68(H3N2)具明显抑制作用,并能与HIV包膜蛋白GP120相互作用。

Assay of Cytotoxicity and Antiviral Activity of Cyanovirin-N Derivative

Objective] To study the cytotoxicity of LCVN,the derivative of Cyanovirin N,to the immortal human keratinocyte cell line(HaCat),conduct preparation and antiviral assay of LCVN gel.Method] The 50% cell cytotoxicity concentration(CC50) of LCVN on HaCaT cells and the 50% inhibitory concentration(IC50) of LCVN against influenza virus A/HK/8/68(H3N2) infection were determined by MTT assay.The effect of LCVN on the cell cycle of HaCaT cells was examined by flow cytometry.The interaction between LCVN and the envelopment protein GP120 of HIV was evaluated by enzyme linked immunosorbent assay(ELISA).Result] The CC50 values of LCVN on HaCaT cells were(9.25±1.21) and(1.94±0.12) μmol/L,following 24 and 48 h treatment respectively.After prepared as a gel form,LCVN interacted with GP120 of HIV in a dose-response relationship in certain concentration ranges.The IC50 value of LCVN against influenza virus A/HK/8/68(H3N2) was(98.20±0.03) nmol/L.Conclusion] The cytotoxicity of LCVN on HaCaT cells is evidently lower than that of CVN,and the LCVN gel possesses anti-influenza A/HK/8/68(H3N2) activity and can interact with GP120 of HIV.