Modulating effect of tumor necrosis factor (TNF) and vitamin D on TNFR2 receptor expression in monocytes and lymphocytes from pregnant women with preeclampsia and on the differentiation of regulatory T cells.

• Grant number: 24/14678-3
• Support Opportunities: Regular Research Grants
• Start date: April 01, 2025
• End date: March 31, 2028
• Field of knowledge: Biological Sciences - Immunology - Applied Immunology
• Principal Investigator: Maria Terezinha Serrão Peraçoli
• Grantee: Maria Terezinha Serrão Peraçoli
• Host Institution: Instituto de Biociências (IBB). Universidade Estadual Paulista (UNESP). Campus de Botucatu. Botucatu , SP, Brazil
• Associated researchers: Jose Carlos Peraçoli ; Mariana Romão Veiga ; Vanessa Rocha Ribeiro Vasques

Abstract

Introduction: Preeclampsia (PE) is a pregnancy-specific syndrome characterized by the activation of monocytes with an inflammatory (M1) profile, as opposed to the anti-inflammatory M2 profile, leading to elevated levels of pro-inflammatory cytokines, such as tumor necrosis factor (TNF). TNF is initially produced as a transmembrane protein (mTNF) and can be cleaved by TNF converting enzyme (TACE) to form soluble TNF (sTNF). The binding of TNF to TNFR1 and TNFR2 receptors activates distinct signaling pathways in innate and adaptive immune cells. TNFR1 can be activated by both mTNF and sTNF, while TNFR2 is primarily activated by mTNF. TNFR2 is predominantly expressed by CD4+/Foxp3+ regulatory T cells (Tregs), and co-expression of CD25 with TNFR2 identifies cells with enhanced suppressive capacity. In PE, the balance between Tregs and Th17 cells is disrupted, showing a polarization towards increased Th17 cells and decreased Tregs. Immune checkpoint inhibitors, or biomolecules with negative regulatory activity, are expressed by Tregs and are considered crucial for balancing pro- and anti-inflammatory signals at the maternal-fetal interface, ensuring maternal tolerance and successful pregnancy. Consequently, using inflammatory response modulators such as vitamin D in the treatment of pregnant women with PE may be significant for improving systemic inflammatory responses by activating M2 and Treg cells capable of modulating and suppressing the inflammation associated with this pregnancy syndrome. Objective: To understand the role of TNF receptors (TNFR1 and TNFR2) in subpopulations of monocytes and lymphocytes involved in the systemic inflammatory response of pregnant women with preeclampsia and to use vitamin D to modulate the inflammatory profile of these cells, establishing the balance between innate and adaptive immunity in PE. Subjects and Methods: Peripheral blood will be collected from 20 pregnant women with PE, 20 normotensive pregnant women, and 16 healthy non-pregnant women. Mononuclear cells will be cultured in vitro in the presence or absence of TNF or vitamin D for 4, 18, or 24 hours. The expression of receptors CD14, TLR4, CD64, CD163, CD206, TNFR1, TNFR2, and mTNF on monocytes, and the expression of regulatory T cell markers CD3, CD4, CD25, TNFR1, TNFR2, CD155, CTLA-4, PD-1, TIGIT, GITR, FoxP3, and ROR³t will be determined by flow cytometry. Cytokine concentrations of TNF, IL-10, IL-17A, TGF-², IL-35, IL-6, and soluble receptors TNFR1, and TNFR2 will be measured in plasma or supernatant before and after culture with TNF and vitamin D using enzyme-linked immunosorbent assay (ELISA). Results will be analyzed using parametric or non-parametric tests with a significance level of 5%. (AU)