Department of Medicine, Uniformed Services University of Health Sciences, Bethesda, Maryland
Department of Medicine, Uniformed Services University of Health Sciences, Bethesda, Maryland.
Am J Physiol Renal Physiol. 2014 Sep 1;307(5):F516-24. doi: 10.1152/ajprenal.00006.2014. Epub 2014 Jul 2.
Visceral leishmaniasis patients have been reported to have a urine concentration defect. Concentration of urine by the renal inner medulla is essentially dependent on a transcription factor, NFAT5/TonEBP, because it activates expression of osmoprotective genes betaine/glycine transporter 1 (BGT1) and sodium/myo-inositol transporter (SMIT), and water channel aquaporin-2, all of which are imperative for concentrating urine. Leishmania parasites evade macrophage immune defenses by activating protein tyrosine phosphatases, among which SHP-1 is critical. We previously demonstrated that SHP-1 inhibits tonicity-dependent activation of NFAT5/TonEBP in HEK293 cells through screening a genome-wide small interfering (si) RNA library against phosphatases (Zhou X, Gallazzini M, Burg MB, Ferraris JD. Proc Natl Acad Sci USA 107: 7072-7077, 2010). We sought to examine whether Leishmania can activate SHP-1 and inhibit NFAT5/TonEBP activity in the renal inner medulla in a murine model of visceral leishmaniasis by injection of female BALB/c mice with a single intravenous dose of 5 × 10(5) L. chagasi metacyclic promastigotes. We found that SHP-1 is expressed in the kidney inner medulla. L. chagasi activates SHP-1 with an increase in stimulatory phosphorylation of SHP-1-Y536 in the region. L. chagasi reduces expression of NFAT5/TonEBP mRNA and protein as well as expression of its targeted genes: BGT1, SMIT, and aquaporin-2. The culture supernatant from L. chagasi metacyclic promastigotes increases SHP-1 protein abundance and potently inhibits NFAT5 transcriptional activity in mIMCD3 cells. However, L. chagasi in our animal model has no significant effect on urinary concentration. We conclude that L. chagasi, most likely through its secreted virulence factors, activates SHP-1 and reduces NFAT5/TonEBP gene expression, which leads to reduced NFAT5/TonEBP transcriptional activity in the kidney inner medulla.
内脏利什曼病患者被报道存在尿液浓缩缺陷。肾髓质的尿液浓缩主要依赖于转录因子 NFAT5/TonEBP，因为它激活了甜菜碱/甘氨酸转运体 1（BGT1）和钠/肌醇转运体（SMIT）以及水通道 aquaporin-2 的表达，所有这些都是浓缩尿液所必需的。利什曼原虫通过激活蛋白酪氨酸磷酸酶来逃避巨噬细胞的免疫防御，其中 SHP-1 至关重要。我们之前通过筛选针对磷酸酶的全基因组小干扰（si）RNA 文库，证明 SHP-1 通过抑制 HEK293 细胞中 NFAT5/TonEBP 的渗透压依赖性激活来抑制蛋白酪氨酸磷酸酶（Zhou X、Gallazzini M、Burg MB、Ferraris JD.Proc Natl Acad Sci USA 107: 7072-7077, 2010）。我们试图通过向雌性 BALB/c 小鼠单次静脉注射 5×10（5）个 L. chagasi 动合子前鞭毛体来研究内脏利什曼病小鼠模型中内脏利什曼原虫是否可以激活 SHP-1 并抑制肾髓质中的 NFAT5/TonEBP 活性。我们发现 SHP-1 在肾脏髓质中表达。L. chagasi 通过增加 SHP-1-Y536 区域的刺激性磷酸化来激活 SHP-1。L. chagasi 减少 NFAT5/TonEBP mRNA 和蛋白及其靶向基因 BGT1、SMIT 和 aquaporin-2 的表达。L. chagasi 动合子前鞭毛体的培养上清液增加了 mIMCD3 细胞中 SHP-1 蛋白的丰度，并强烈抑制 NFAT5 转录活性。然而，在我们的动物模型中，L. chagasi 对尿液浓缩没有显著影响。我们得出结论，L. chagasi 很可能通过其分泌的毒力因子激活 SHP-1 并降低 NFAT5/TonEBP 基因表达，从而导致肾髓质中 NFAT5/TonEBP 转录活性降低。
