核酸适配体对岩沙海葵毒素细胞毒性保护作用

目的 核酸适配体（Aptamer）是利用指数富集的配基系统进化技术（SELEX）筛选得到的单链DNA或RNA片段，能高亲和、高特异性地与靶标分子结合。岩沙海葵毒素（Palytoxin，PLTX）是目前已知的最具毒性的非蛋白质类天然海洋毒素之一。基于核酸适配体对PLTX的特异性靶向功能，探究核酸适配体对细胞毒性的保护作用。方法 采用细胞毒性实验分别检测比较了PLTX对NIH-3T3、CHO-K1、HACAT三种不同细胞的毒性作用，以及适配体拮抗PLTX的细胞毒作用。采用红细胞溶血实验检测PLTX对人红细胞的溶血作用，以及适配体抑制PLTX的溶血活性作用。结果 三种细胞毒性曲线结果显示，HACAT细胞的ID50约为3.42ng/ml，CHO-K1细胞的ID50约为1.06×10-2ng/ml，NIH-3T3细胞的ID50约为1.39×10-2ng/ml。在不同浓度下，PLTX组与PLTX:aptamer=1:1组进行比较，结果显示适配体抑制PLTX对HACAT细胞的毒性作用（P<0.05）。在50nM PLTX浓度下，PLTX组与PLTX:aptamer=1:1组进行比较，结果显示适配体抑制PLTX对人红细胞溶血活性作用（P<0.05）。结论 实验结果表明HACAT细胞对2.5×10-3~2.5×102ng/ml浓度范围的PLTX毒性作用较敏感；核酸适配体对HACAT细胞具有细胞毒性保护作用。在50nM PLTX浓度下，相应浓度适配体能够有效抑制PLTX对人红细胞的溶血活性，起到显著细胞保护作用。

Protective effect of Nucleic acid aptamers on cytotoxicity of palytoxin

Objective Nucleic acid Aptamers are single-stranded DNA or RNA fragments screened by Systematic Evolution of Ligands by Exponential Enrichmenty (SELEX), which can bind to target molecules with high affinity and specificity. Palytoxin (PLTX) is one of the most toxic non-protein natural marine toxins. Based on the specific targeting function of aptamer to PLTX, the antagonistic effects of aptamer to PLTX were explored. Methods The cytotoxicity of PLTX on NIH-3T3, CHO-K1 and HACAT cells and the antagonistic effecst of aptamer to PLTX were detected and compared by cytotoxicity experiment. The hemolytic effect of PLTX on human erythrocytes and the inhibitory effect of aptamer on the hemolytic activity of PLTX were detected by erythrocyte hemolysis test. Results The results of three cytotoxicity curves showed that the ID50 of HACAT cells was about 3.42ng/ml, that of CHO-K1 cells was about 1.06×10-2ng/ml, and that of NIH-3T3 cells was about 1.39×10-2ng/ml. At different concentrations, PLTX group was compared with PLTX: aptamer = 1:1 group. The results showed that aptamer inhibited the toxic effect of PLTX on HACAT cells (P < 0.05). At a concentration of 50 nM PLTX, the PLTX group was compared with the PLTX:aptamer=1:1 group, and the results showed that the aptamer inhibited the hemolytic activity of PLTX on human erythrocytes (P<0.05).Conclusion The experimental results show that HACAT cells are more sensitive to PLTX toxicity in the concentration range of 2.5×10-3~2.5×102ng/ml and the aptamer has a cytotoxic protective effect on HACAT cells. At a concentration of 50 nM PLTX, the corresponding concentration of aptamer can effectively inhibit the hemolytic activity of PLTX on human erythrocytes and play a significant cytoprotective effect.