Gene and protein expression of Toll-like receptor-1, -2, -4, and -6 in maternal neutrophils and monocytes and activation of these cells in normal and complicated pregnancies by RPM-PT or PPT in the presence or absence of chorioamnionitis

• Grant number: 11/14195-2
• Support Opportunities: Scholarships in Brazil - Doctorate
• Start date: November 01, 2011
• End date: February 28, 2015
• Field of knowledge: Health Sciences - Medicine - Maternal and Child Health
• Principal Investigator: Márcia Guimarães da Silva
• Grantee: Natália Prearo Moço
• Host Institution: Faculdade de Medicina (FMB). Universidade Estadual Paulista (UNESP). Campus de Botucatu. Botucatu , SP, Brazil

Abstract

Introduction: Toll-like receptors (TLRs) are an important part of the innate immune system, and they are expressed in various human tissues. Few studies have focused on the role played by TLRs in pregnancy, and scarce data are available in relation to the expression of such receptors in maternal neutrophils and monocytes in pregnancies complicated by the preterm premature rupture of membranes (PPROM) and preterm labor (PTL). Objective: To outline the gene and protein expression of TLR-1, -2, -4 and -6 in neutrophils and monocytes from peripheral maternal blood as well as to evaluate the state of activation of such cells during normal pregnancy and in pregnancies complicated by PPROM and PTL in the presence or not of histologic chorioamnionitis. Patients and methods: A longitudinal study will be performed. It will include 15 TLR expression measurements in maternal neutrophils by gestational week from the 5th gestational week on, including the moment of delivery. The control group will consist of 50 non-pregnant women who must show similar age and parity to those in the study group. Concurrently, a cross-sectional study will be conducted, and it will include 45 pregnant women with PPROM and 45 pregnant women in PTL with an intact amniotic sac, in the presence of histologic chorioamnionitis or not. The control group will consist of pregnant women at term without obstetric complications, belonging to the longitudinal study. 35 mL of peripheral blood will be collected for the isolation and culture of neutrophils and monocytes, and 5 mL for total RNA extraction. At the moment of pregnancy resolution, fragments of the chorioamniotic membranes will be fixed, dehydrated in alcohol, diaphanized in xylol and then included into paraffin blocks. 5-mm sections will be stained by the Hematoxylin-Eosin (HE) method. Messenger RNA from TLR-1,-2,-4 and -6 in maternal neutrophils will be detected by the real-time Polymerase-Chain Reaction (PCR) method; the protein detection of TLR2 and TLR4 in neutrophils will be evaluated by flow cytometry, and the chorioamnionitis study will be performed by histopathologic analysis. The evaluation of the activation of neutrophils and monocytes stimulated by specific PAMPs and for the TLRs of interest will be performed by dosing cytokines IL-1, IL-6, IL-8 and TNF-a and by the ELISA method, in addition to the dosage of hydrogen peroxide production. Statistical analysis will be performed by respecting the presuppositions determined by the results. The level of significance adopted will be of 5%, and the SigmaStat 9.0 (Jandel Corporation) will be used.

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

MOCO, NATALIA PREARO; CAMARGO BATISTA, RENATA APARECIDA; MARTIN, LAURA FERNANDES; DE OLIVEIRA, LEANDRO GUSTAVO; GARCIA DE LIMA PARADA, CRISTINA MARIA; DIAS-MELICIO, LUCIANE ALARCAO; GOLIM, MARJORIE DE ASSIS; DA SILVA, MARCIA GUIMARAES. Toll-Like Receptor-2 and-4 Expression by Maternal Neutrophils in Preterm Labor. GYNECOLOGIC AND OBSTETRIC INVESTIGATION, v. 83, n. 1, p. 1-8, 2018. (13/12624-9, 11/14195-2)

Academic Publications

(References retrieved automatically from State of São Paulo Research Institutions)

MOÇO, Natália Prearo. Expressão gênica e protéica de receptores Toll-like em células do sangue periférico materno de gestações normais e complicadas por prematuridade. 2015. Doctoral Thesis - Universidade Estadual Paulista (Unesp). Faculdade de Medicina. Botucatu Botucatu.