The Function And Mechanism Study Of The Correlation Between Long Noncoding RNA INHBA-AS1 And Preeclampsia

Preeclampsia(PE)is a pregnancy-specific syndrome characterized by new onset of hypertension and proteinuria after 20 weeks of gestation.At present,the pathogenesis is still unclear,which may be related to the impaired uterine spiral artery remodeling.Other possible mechanisms include overactivation of inflammatory immunity,impaired vascular endothelium,genetic factors,nutritional deficiency,insulin resistance and so on.The main pathological manifestations of preeclampsia placenta are that trophoblasts invade the endometrium too shallowly,accompanied by physiological vascular remodeling disorder of uterine spiral artery,decrease and low density of villus area.The physiological vascular remodeling disorder is the characteristic pathological change of preeclampsia.Long noncoding RNA(lncRNA)refers to an RNA molecule with a transcript of more than 200nt in length.A growing number of studies have found that lncRNAs are closely linked to many life activities.Previous studies have found that lncRNA is abnormally expressed in preeclampsia placenta compared with normal pregnant women,and affects trophoblast proliferation,migration,invasion,apoptosis and other functions.In the preliminary study of our research group,placental tissue samples from 9 cases of early-onset severe preeclampsia(EOSPE)and 32 cases of normal pregnant women delivered by cesarean section in Nanfang Hospital were collected for the second generation sequencing.The results showed that lncRNA INHBA-AS1(INHBA antisense RNA 1)was highly expressed in EOSPE.From December 2016 to May 2018,20 cases of PE and 20 cases of normal pregnant placenta samples from the obstetrics of Southern Hospital were tested by quantitative real-time PCR(qRT-PCR),which also showed that ESHBA-AS1 was significantly up-regulated in PE.Then the biological function and possible mechanism of INHBA-AS1 were studied in trophoblastic cell line HTR-8/Svneo.The experimental results suggest that the cell invasion and migration are inhibited after overexpression of INHBA-AS1.Conversely,knocking out INHBA-AS1 with CRISPR/Cas9 technology promoted cell invasion and migration.We verified that the lncRNA INHBA-AS1 is mainly located in the nucleus by,fluorescence in situ hybridization(FISH)and nuclear separation experiments,indicating that it may affect the cellular transcription process.By mass spectrometry of RNA-pulldown proteins,INHBA-AS1 binding proteins may be associated with nuclear complexes and INHBA-AS1 may bind to some nuclear transcription factors.The target genes of transcription factor CENPB(centromere protein B),TRAF1(TNF receptor associated factor 1),were regulated by INHBA-AS1.Furthermore,CHIP and luciferase reporter gene assays demonstrated that INHBA-AS1 binds to CENPB to affect the transcription of the TRAF1.Meanwhile,TRAF1 could affect invasion/migration related pathway proteins MMP2,MMP3 and Vimentin.Therefore,we believe that INHBA-AS1 may be related to the pathogenesis of preeclampsia.By inhibiting the invasion and migration of trophoblasts,the invasion of endometrium is too shallow,resulting in shallow placental implantation,which leads to the occurrence and development of PE.In summary,INHBA-AS1 may be a potential biomarker and therapeutic target for PE.