Data from: Cruzipain activates latent TGF-β from host cells during T. cruzi invasion

Several studies indicate that the activity of cruzipain, the main lysosomal cysteine peptidase of Trypanosoma cruzi, contributes to parasite infectivity. In addition, the parasitic invasion process of mammalian host cells is described to be dependent on the activation of the host TGF-β signaling pathway by T. cruzi. Here, we tested the hypothesis that cruzipain could be an important activator of latent TGF-β and thereby trigger TGF-β-mediated events crucial for the development of Chagas disease. We found that live epimastigotes of T. cruzi, parasite lysates and purified cruzipain were able to activate latent TGF-β in vitro. This activation could be inhibited by the cysteine peptidase inhibitor Z-Phe-Ala-FMK. Moreover, transfected parasites overexpressing chagasin, a potent endogenous cruzipain inhibitor, prevented latent TGF-β activation. We also observed that T. cruzi invasion, as well as parasite intracellular growth, were inhibited by the administration of Z-Phe-Ala-FMK or anti-TGF-β neutralizing antibody to Vero cell cultures. We further demonstrated that addition of purified cruzipain enhanced the invasive activity of trypomastigotes and that this effect could be completely inhibited by addition of a neutralizing anti-TGF-β antibody. Taken together, these results demonstrate that the activities of cruzipain and TGF-β in the process of cell invasion are functionally linked. Our data suggest that cruzipain inhibition is an interesting chemotherapeutic approach for Chagas disease not only because of its trypanocidal activity, but also due to the inhibitory effect on TGF-β activation.多项研究表明，克鲁兹因（cruzipain）——克氏锥虫（Trypanosoma cruzi）的主要溶酶体半胱氨酸肽酶——的活性有助于该寄生虫的感染性。此外，已有研究证实，哺乳动物宿主细胞的寄生虫侵袭过程依赖于克氏锥虫对宿主转化生长因子-β（TGF-β）信号通路的激活。本研究旨在验证如下假说：克鲁兹因可作为潜在转化生长因子-β的重要激活因子，进而触发恰加斯病（Chagas disease）发生发展过程中至关重要的TGF-β介导的生物学事件。实验结果显示，活的克氏锥虫上鞭毛体（epimastigotes）、寄生虫裂解液以及纯化的克鲁兹因均可在体外激活潜在TGF-β。该激活过程可被半胱氨酸肽酶抑制剂Z-Phe-Ala-FMK所抑制。此外，过表达查加斯素（chagasin）——一种强效内源性克鲁兹因抑制剂——的转染寄生虫，可阻断潜在TGF-β的激活。我们还观察到，向Vero细胞培养体系中添加Z-Phe-Ala-FMK或抗TGF-β中和抗体，可抑制克氏锥虫的细胞侵袭能力以及寄生虫的胞内增殖过程。进一步实验证实，添加纯化的克鲁兹因可增强锥鞭毛体（trypomastigotes）的侵袭活性，且该效应可通过加入抗TGF-β中和抗体完全阻断。综上，上述结果表明，克鲁兹因与TGF-β在细胞侵袭过程中的活性存在功能关联。本研究数据提示，抑制克鲁兹因有望成为治疗恰加斯病的潜在化疗策略，这不仅因其具备杀锥虫活性，还因其可阻断TGF-β的激活过程。