Abstract
BackgroundAn appropriate transcriptional profile in the placenta and fetal membranes is required for successful pregnancy; any variations may lead to inappropriate timing of birth. Epigenetic regulation through reversible modification of chromatin has emerged as a fundamental mechanism for the control of gene expression in a range of biological systems and can be modified by pharmacological intervention, thus providing novel therapeutic avenues. TIMP-1 is an endogenous inhibitor of MMPs, and hence is intimately involved in maintaining the integrity of the fetal membranes until labor.Objective and MethodsTo determine if TIMP-1 is regulated by DNA methylation in gestational tissues we employed an in vitro model in which gestational tissue explants were treated with demethylating agent 5-aza-2'-deoxycytidine (AZA) and lipopolysaccharide (LPS).ResultsQuantitative Real-Time PCR (qRT-PCR) revealed that TIMP-1 transcription was significantly increased by combined treatment of AZA and LPS, but not LPS alone, in villous, amnion and choriodecidua explants after 24 and 48 hrs, whilst western blotting showed protein production was stimulated after 24 hrs only. Upon interrogation of the TIMP-1 promoter using Sequenom EpiTyper MassARRAY, we discovered sex-specific differential methylation, in part explained by x-linked methylation in females. Increased TIMP-1 in the presence of LPS was potentiated by AZA treatment, signifying that a change in chromatin structure, but not in DNA methylation at the promoter region, is required for transcriptional activators to access the promoter region of TIMP-1.ConclusionsCollectively, these observations support a potential role for pharmacological agents that modify chromatin structure to be utilized in the therapeutic targeting of TIMP-1 to prevent premature rupture of the fetal membranes in an infectious setting.
Highlights
An appropriate transcriptional profile in the placenta and fetal membranes is required for successful pregnancy; any variations may lead to inappropriate timing of birth
Quantitative Real-Time PCR revealed that tissue inhibitors of metalloproteinases (TIMPs)-1 transcription was significantly increased by combined treatment of AZA and LPS, but not LPS alone, in villous, amnion and choriodecidua explants after 24 and 48 hrs, whilst western blotting showed protein production was stimulated after 24 hrs only
Upon interrogation of the TIMP-1 promoter using Sequenom EpiTyper MassARRAY, we discovered sex-specific differential methylation, in part explained by x-linked methylation in females
Summary
An appropriate transcriptional profile in the placenta and fetal membranes is required for successful pregnancy; any variations may lead to inappropriate timing of birth. The end of human gestation is signified by the physiological hallmarks of uterine contractions and rupture of the fetal membranes (amnion and chorion) [1] These two processes are essential for the advancement of labor, and inappropriate timing of these processes may lead to Matrix metalloproteinase (MMP) mediated extracellular matrix (ECM) degradation plays a key role in normal growth and remodelling of fetal membranes throughout gestation and at term, and are critically involved in the weakening and subsequent rupture of the fetal membranes at the time of labor [3]. Increased MMPs, MMP-9, in fetal membrane rupture at both preterm and term labor is well documented [6, 7, 9,10,11,12,13] and the MMP-9:TIMP-1 ratio in fetal membranes at term gestation correlates with their tensile strength [14]. This altered MMP-9:TIMP-1 ratio is exacerbated in the presence of infection [7, 15,16,17,18,19]
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