Song Kyoung Ju, Ahn Hye-Jin, Nam Ho-Woo
The Catholic Institute of Parasitic Diseases, College of Medicine, The Catholic University of Korea, Seoul 137-701, Korea.
Korean J Parasitol. 2012 Mar;50(1):1-6. doi: 10.3347/kjp.2012.50.1.1. Epub 2012 Mar 6.
Toxoplasma gondii penetrates all kinds of nucleated eukaryotic cells but modulates host cells differently for its intracellular survival. In a previous study, we found out that serine protease inhibitors B3 and B4 (SERPIN B3/B4 because of their very high homology) were significantly induced in THP-1-derived macrophages infected with T. gondii through activation of STAT6. In this study, to evaluate the effects of the induced SERPIN B3/B4 on the apoptosis of T. gondii-infected THP-1 cells, we designed and tested various small interfering (si-) RNAs of SERPIN B3 or B4 in staurosporine-induced apoptosis of THP-1 cells. Anti-apoptotic characteristics of THP-1 cells after infection with T. gondii disappeared when SERPIN B3/B4 were knock-downed with gene specific si-RNAs transfected into THP-1 cells as detected by the cleaved caspase 3, poly-ADP ribose polymerase and DNA fragmentation. This anti-apoptotic effect was confirmed in SERPIN B3/B4 overexpressed HeLa cells. We also investigated whether inhibition of STAT6 affects the function of SERPIN B3/B4, and vice versa. Inhibition of SERPIN B3/B4 did not influence STAT6 expression but SERPIN B3/B4 expression was inhibited by STAT6 si-RNA transfection, which confirmed that SERPIN B3/B4 was induced under the control of STAT6 activation. These results suggest that T. gondii induces SERPIN B3/B4 expression via STAT6 activation to inhibit the apoptosis of infected THP-1 cells for longer survival of the intracellular parasites themselves.
刚地弓形虫可侵入各种有核真核细胞，但为在细胞内存活，它对宿主细胞的调控方式有所不同。在之前的一项研究中，我们发现丝氨酸蛋白酶抑制剂B3和B4（因其高度同源，故称为丝氨酸蛋白酶抑制剂B3/B4）在受刚地弓形虫感染的THP-1衍生巨噬细胞中通过激活信号转导和转录激活因子6（STAT6）而显著上调。在本研究中，为评估诱导产生的丝氨酸蛋白酶抑制剂B3/B4对刚地弓形虫感染的THP-1细胞凋亡的影响，我们设计并测试了针对丝氨酸蛋白酶抑制剂B3或B4的各种小干扰RNA（si-RNA），用于检测其在星形孢菌素诱导的THP-1细胞凋亡中的作用。当用基因特异性si-RNA转染THP-1细胞敲低丝氨酸蛋白酶抑制剂B3/B4后，通过检测裂解的半胱天冬酶3、聚ADP核糖聚合酶和DNA片段化发现，刚地弓形虫感染后THP-细胞的抗凋亡特性消失。在过表达丝氨酸蛋白酶抑制剂B3/B4的HeLa细胞中也证实了这种抗凋亡作用。我们还研究了抑制STAT6是否会影响丝氨酸蛋白酶抑制剂B3/B4的功能，反之亦然。抑制丝氨酸蛋白酶抑制剂B3/B4并不影响STAT6的表达，但转染STAT6 si-RNA可抑制丝氨酸蛋白酶抑制剂B3/B4的表达，这证实了丝氨酸蛋白酶抑制剂B3/B4是在STAT6激活的调控下被诱导产生的。这些结果表明，刚地弓形虫通过激活STAT6诱导丝氨酸蛋白酶抑制剂B3/B4的表达，以抑制被感染的THP-1细胞凋亡，从而使细胞内寄生虫自身能够存活更长时间。
