Immunomodulatory effect of vitamin D on the activation of inflammasomes in placental tissue from pregnant women with pre-eclampsia

Abstract

Pregnancies complicated by Pre-Eclampsia (PE) are associated with placental ischaemia/hypoxia, oxidative stress, and elevated levels of proinflammatory cytokines. These physiological changes can trigger cell death by apoptosis, an essential process for the maintenance of homeostasis in human tissues, including the placenta and considered an intermediate event in the generation of pregnancy syndromes, such as PE. In addition to this cellular process, the activation of NLRP3 inflammasome, represented by increased gene and protein expression of components of this inflammation-related complex such as caspase-1, interleukin-1 beta (IL-1²) and high-mobility group box-1 (HMGB1) was described in placenta of preeclamptic pregnant women, when compared to normotensive pregnant women. The hyperactivation of NLRP3 inflammasome and the triggering of apoptosis in placental tissues may be involved in the exacerbated systemic inflammatory state in PE. The present project aims to evaluate the immunomodulatory effect of vitamin D on the inflammassomes NLRP1 and NLRP3 and on apoptosis in placental explants of pregnant women with PE and in the BeWo trophoblastic cell line. It will be evaluated 45 pregnant women distributed as follows: 15 women with early-onset PE (gestational age < 34 weeks), 15 with late-onset PE (gestational age e 34 weeks) and 15 normotensive pregnant women. After caesarean section, fragments of approximately 10 mg will be immediately removed from the central region of the placenta, constituting samples of the villous cytotrophoblast and the region of syncytiotrophoblast in contact with the maternal face. BeWo cells and explants will be cultured in the presence or absence of Monosodium Urate (MSU), hydrogen peroxide (H2O2) and Tumor Necrosis Factor (TNF-±) and treated with vitamin D. After 4 and 18 hours of culture the explants and the cells will submitted for evaluation of the gene and protein expression of NLRP1, NLRP3, caspase-1, IL-1², IL-18, TNF-±, HMGB1, tumor necrosis factor related apopotosis-inducing ligant (TRAIL) and Vitamin D Receptor (VDR) by RT-qPCR, ELISA and Western Blot. The results will be evaluated using parametric or non-parametric tests depending of the variability of the results. (AU)