Division of Molecular Medicine, Bose Institute, P-1/12, CIT Scheme VII- M, Kolkata, 700 054, West Bengal, India.
Division of Molecular Medicine, Bose Institute, P-1/12, CIT Scheme VII- M, Kolkata, 700 054, West Bengal, India.
Exp Parasitol. 2020 Oct;217:107948. doi: 10.1016/j.exppara.2020.107948. Epub 2020 Jul 19.
Immunomodulation is an emerging concept to combat infection in recent years. Immunomodulators like arabinosylated-lipoarabinomannan (Ara-LAM) and glycyrrhizic-acid (GA) possess anti-leishmanial property, whereas sodium-antimony-gluconate (SAG) is still considered as the first choice for chemotherapy against leishmaniasis. During infection, invasion of Leishmania donovani needs the potential requirement of Ca, which is further responsible for apoptosis in intracellular amastigotes. However, suppression of elevated intracellular calcium by the activation of plasma-membrane-calcium-ATPase (PMCA4) facilitates survival of L. donovani in the host. In the present study, SAG, Ara-LAM, and GA were found to evoke significant increase in intracellular Ca in L. donovani infected macrophages by inhibiting PMCA4. Moreover, PMCA4 inhibition by TFP or PMCA4 siRNA elevated the level of PKCβ, whereas calcium-independent upregulation of PKCζ remained unchanged in infected macrophages. Furthermore, application of immunomodulators in infected macrophages resulted in down-regulation of PKCζ, conversion of anti-inflammatory to pro-inflammatory cytokines and inhibition of PMCA4. Plasma membrane-associated ceramide which is known to be elevated during leishmaniasis, triggered upregulation of PMCA4 via PKCζ activation. Interestingly, immunomodulators attenuated ceramide generation, which resulted into reduced PKCζ activation leading to the decreased PMCA expression in infected macrophages. Therefore, our study elucidated the efficacy of SAG, Ara-LAM, and GA in the reduction of parasite burden in macrophages by suppressing PMCA activation through inhibition of ceramide mediated upregulation of PKCζ.
免疫调节是近年来对抗感染的一个新兴概念。免疫调节剂，如阿拉伯糖基化-脂阿拉伯甘露聚糖（Ara-LAM）和甘草酸（GA），具有抗利什曼原虫的特性，而葡萄糖酸锑钠（SAG）仍然被认为是治疗利什曼病的首选化疗药物。在感染过程中，利什曼原虫的入侵需要钙的潜在需求，这进一步导致细胞内无鞭毛体的细胞凋亡。然而，通过激活质膜钙 ATP 酶（PMCA4）抑制细胞内钙的升高，有利于利什曼原虫在宿主体内的存活。在本研究中，发现 SAG、Ara-LAM 和 GA 通过抑制 PMCA4 可引起感染巨噬细胞内 Ca 显著增加。此外，TFP 或 PMCA4 siRNA 抑制 PMCA4 可提高 PKCβ 的水平，而感染巨噬细胞中钙非依赖性的 PKCζ 的上调保持不变。此外，在感染的巨噬细胞中应用免疫调节剂可导致 PKCζ 的下调，将抗炎细胞因子转化为促炎细胞因子，并抑制 PMCA4。已知在利什曼病期间会升高的质膜相关神经酰胺，通过 PKCζ 的激活触发 PMCA4 的上调。有趣的是，免疫调节剂可减轻神经酰胺的生成，从而导致 PKCζ 的激活减少，从而导致感染的巨噬细胞中 PMCA 表达减少。因此，我们的研究表明，SAG、Ara-LAM 和 GA 通过抑制神经酰胺介导的 PKCζ 上调来抑制 PMCA 激活，从而降低巨噬细胞中的寄生虫负荷，从而发挥其疗效。
