Gut Microbiota-derived Endotoxin Enhanced The Incidence Of Cardia Bifida During Cardiogenesis

AimsCytotoxicity and inflammation-associated toxic responses could be induced by bacterial lipopolysaccharides(LPS)in vitro and in vivo,respectively.However,the mechanism involved in LPS-induced cardiac malformation in prenatal fetus is still unknown.We exposed the cellular and molecular mechanisms of the abnormal development of heart tube induced by LPS in the early chick embryo.MethodsFirstly,the endotoxin level was detected after the intestinal flora of human and mouse were disturbed.After treated LPS,We observed chick embryos' development situation in HH10.Phase in HH7,HH10 materials,use immunofluorescence,in situ hybridization,Q-PCR,Western blot,electroporation transfection and transplantation,observe how LPS influence the expression of transcription factors,myocardial cell migration,differentiation,proliferation apoptosis of myocardial cells and ROS level change in the development of early embryonic heart tube,to explore the effect of LPS on cardiovascular tube development mechanism.In addition,combined with in vitro primary culture and H9c2 cells,we explored the mechanism of LPS induced ROS level to participate in cardiac malformation.ResultsIn this study,it is demonstrated that LPS was induced in gut microbiota imbalance human and mice.LPS exposure during gastrulation in the chick embryo increased the incidence of cardia bifida.Gene transfection and tissue transplantation trajectory indicated that LPS exposure restricted the cell migration of cardiac progenitors to primary heart field in gastrula chick embryos.In vitro explant allograft of GFP-labeled anterior primitive streak demonstrated that LPS treatments could inhibit cell migration.A similar observation was obtained from cell migration assays of scratch wounds using primary culture of cardiomyocytes or H9c2 cells.In embryos exposed to LPS,expressions of Nkx2.5 and GATA5 were disturbed.These genes are associated with cardiomyocyte differentiation when heart tube fusion occurs.Furthermore,pHIS3,cCaspase3 immunohistological staining indicated that cell proliferation decreased and cell apoptosis increased in the heart tube of chick embryos.In vivo,pHIS3 immunohistological staining and Hochest/PI staining produced similar conclusions.LPS exposure also led to production of excess ROS,which might damage the cardiac precursor cells of developing embryos.Finally,it was demonstrated that LPS-induced cardia bifida could be partially reversed through addition of antioxidants.ConclusionsTogether,these results reveal that excess ROS generation is involved in the LPSinduced defects in heart tube during chick embryo development.