| Preeclampsia (PE) is a common and serious pregnancy-specific syndrome. Recent studies proof that immune system disorder in maternal-fetal interface has important role in PE as a result of placental ischemia. Uterine decidual natural killer (uNK) cells are the main lymphocytes in early gestation. Thus uNK cells have been thought to play a role in PE, although the precise functions of uNK cells in vivo are still unknown. As a result we successfully established a phosphatidylserine (PS)/phosphatidylcholine (PC) micro vesicles-induced PE-like model in mice. The number and phenotype of NK cells in the peripheral blood, spleen and uterus as well as cytokines levels in the peripheral blood were analysised. The number of NK cells in the uterus showed a sudden increase on gestation day (gd) 11.5 and then gradually reduced. But the mice injected with PS/PC showed a significant reduction of NK cells number in the uterus on gd 14.5 and 17.5 compared with controls. PS/PC treated mice got higher CD244, CD94 and NKG2D expression in the peripheral blood natural killer (pNK) cells, while lower CD244, CD94 and NKG2D expression in dNK cells. Lower CD 122 expression was both found in pNK and uNK cells. Compared with controls, the mice injected with PS/PC had significantly increased TGF-βcontent in peripheral blood but TNF-αand IFN-γreminded unchanged. These results suggest that the reduced number of uNK cells as well as the reduced expressions of CD244, CD94, CD 122 and NKG2D may be related to maternal-fetal immune tolerance. TGF-βincrease in PS/PC treated mice may also have impact on NK cell activation and the expression of surface molecules, thereby affecting the immune balance in maternal-fetal interface. So we deduced that PS/PC induced preeclampsia-like mice may cause by the alteration of phenotype and function of pNK and uNK cells. These changes may affect the NK cells ability to regulate trophoblast invasion resulting in insufficient uterine spiral arteries remodeling and placental ischemia.In order to clarify the effects of 17β-estradiol (E2) on natural killer (NK) cells and the possibly regulatory mechanisms, we obtained highly purified and viable NK cells from C57BL/6J mouse spleen by a magnetic cell sorter (MACS). These cells were treated with E2 and then their cytotoxicity and proliferative capacity were examined. To further investigate the mechanisms on the effect of E2 on NK cells, expressions of activation-associated markers (CD69, CD 122) and inhibitory receptors (CD94, Ly49), and intracellular cytokine production were analyzed. At last, we performed the cDNA microarray to explore the possible involved genes. We found that E2 could suppress NK cell cytotoxicity and proliferative capacity in vitro. E2 reduced NK cell cytotoxicity and proliferative capacity, which may be through influencing the phenotypes and cytokine expression of NK cells, mainly involving CD94 and IFN-gamma. Furthermore, regulation of Stat4, Fyn, Sh2d1a, Eat2, Cd244, Irfl, Runx1, Irf7, Irf5, Esrra and Nr5al genes may be related to the cytotoxicity, proliferation and cytokine production of E2-mediated purified NK cells.We also investigate the effects of estrogen on the number and cytotoxic activity of NK cells in vivo. The number and cytotoxicity of NK cells in four groups (control, sham+vehicle, Ovx+vehicle and Ovx+E2) were determined. The results showed that 17(3-estradiol (E2) increased the number of NK cells, but reduced their cytotoxicity. The increase of NK cells proportions by E2 may be mediated by upregulating the expression of MCM7 and MCM10 proteins, which are required for DNA replication licensing in cell proliferation. The suppressed cytotoxicity of splenic NK cells by E2 may be attributable to the down-regulation of NK cells activating receptors—CD69, NKp46, NKG2DLand 2B4 (CD244), which directly inhibitedNKcell activation, resulting in the reduced secretion of the soluble factors—granzyme B and FasL. INF-y might also act as a negative regulator in the low cytotoxicity of NK cells. In addition, the number of the NK cells is not parallel to their cytotoxicity with a long-term exposure to E2 in vivo. These results suggest that E2-mediated low cytotoxicity of NK cells may regulate host immune response in pregnancy and some female predominant diseases.Proliferation and expansion of interstitial fibroblasts are predominant features of progressive chronic kidney diseases. However, how interstitial fibroblast proliferation is controlled remains ambiguous. Here we show that tissue-type plasminogen activator (tPA) is a potent mitogen that promotes interstitial fibroblast proliferation through a cascade of signaling events. In vitro, tPA promoted cell proliferation of rat kidney fibroblasts (NRK-49F), as assessed by cell counting, MTT assay and BrdU labeling. tPA also accelerated NRK-49F cell cycle progression. Fibroblast proliferation induced by tPA was associated with an increased expression of numerous proliferation-related genes, including c-fos, c-myc, proliferating cell nuclear antigen and cyclin D1. The mitogenic effect of tPA was independent of its protease activity, but requires LDL receptor-related protein 1 (LRP-1). Interestingly, inhibition ofβ1 integrin signaling prevented tPA-mediated fibroblast proliferation. tPA rapidly induced tyrosine phosphorylation of focal adhesion kinase (FAK), which led to activation of its downstream mitogen-activated protein kinase signaling. Blockade of FAK, but not integrin-linked kinase, abolished the tPA-triggered ERK1/2 activation, proliferation-related gene induction and fibroblast proliferation. Conversely, ectopic expression of FAK induced proliferation-related gene expression. In vivo, proliferation of interstitial fibroblasts in tPA null mice was attenuated after obstructive injury, compared with the wild-type controls. These studies illustrate that tPA is a potent mitogen that promotes renal interstitial fibroblast proliferation through LRP-1-mediatedβ1 integrin and FAK signaling.Renal fibrosis is a devastating condition of progressive kidney scarring and functional impairment. Because aberrant activation of Wnt/p-catenin signaling occurs in the fibrotic kidneys, this pathway might be exploited as a potential target for anti-fibrosis therapy. In this study, we examined the effects ofβ-catenin activation on tubular epithelial-mesenchymal transition (EMT) in vitro and evaluated the therapeutic efficacy of a novel small molecule inhibitor (ICG-001) that specifically disrupts P-catenin-mediated gene transcription in obstructive nephropathy. In vitro, ectopic expression of constitutively activeβ-catenin in tubular epithelial cells (HKC-8) suppressed E-cadherin and induced Snaill, fibrobnecin and plasminogen activator inhibitor-1 (PAI-1) expression. Incubation of HKC-8 cells with ICG-001 dose-dependently suppressed the p-catenin-driven gene transcription in a Topflash luciferase reporter assay. ICG-001 also abolished the expression of Snaill, PAI-1, collagen I, fibronectin andα-smooth muscle actin (α-SMA) in HKC-8 cells induced by TGF-β1. This anti-fibrotic effect of ICG-001 was clearly independent of a potential disruption of Smad signaling. In vivo, administration of ICG-001 through daily intraperitoneal injections attenuated renal fibrosis in obstructive nephropathy. ICG-001 significantly mitigated fibrotic lesions and suppressed the expression of fibronectin, collagen I, collagen III, a-SMA and PAI-1, as well as fibroblast-specific protein-1 in the obstructed kidneys. ICG-001 also repressed renal expression of Snail 1 and Snail2/Slug, key transcriptional factors that regulate EMT. These results suggest that targeting Wnt/β-catenin signaling by small molecule inhibitor could be a novel and effective approach for the treatment of fibrotic kidney diseases.
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