Effect Of Chromogranin A Dereived Peptide CHR On Hyperpermeability Of Blood Brain Barrier And The Molecular Mechanism In Sepsis

Objective To explore the effect of Chromogranin A derived peptide CGA47-66(Chromfungin,CHR)on lipopolysaccharide(LPS)induced high permeability of brain microvascular endothelial cells and its molecular mechanism.Methods(1)Human brain microvascular endothelial cells(HBMEC)exposed to different concentration of LPS(0.01/0.05/0.1/0.5/1/5、10/50μg/ml).Cell counting kit-8(CCK8)method was used to detect the viability of cells and select the optimum stimulate concentration of LPS;(2)HBMEC exposed to LPS,CHR and CGA4-16 respectively.The expresion of the tight junction protein ZO-1 and Claudin-5 in different times were detected by western blot,then selected the optimum stimulate time.(3)HBMEC exposed to LPS,CHR and CGA4-16 respectively.The relative permeability of HBMEC cells was performed by a transwell chamber system and epithelial voltmeter(EVOM)method;the expresion of the tight junction protein ZO-1 and Claudin-5 was detected by western blot and quantitative real-time polymerase chain reaction.Results(1)Compared with the control group,the viability of HBMEC cells was significantly inhibited(all P<0.05)with the increase concentration of LPS,and the cells inhibition ratio is about 50%when LPS concentration is 5μg/ml;(2)The expression of tight junction proteins ZO-1 and Claudin-5 were down-regulated gradually after LPS treatment,and they decreased to the lowest level after 6 hours(P<0.05),and there was no significant difference in the expression after 12h or 24 h compared with 6h(all P>0.05).(3)After incubation with LPS,the transendothelial electrical resistance of HBMEC cells was significantly decreased(t=-18.214,P<0.001)and the permeability of the cells was significantly increased(t=6.083,P=0.004),and the expression of tight junctions ZO-1 and Claudin-5 were significantly decreased(all P<0.05),however,CHR can significantly attenuate the above changes.Conclusion Chromogranin A dereived peptide CHR inhibited the hyperperneability of human brain microvascular endothelial cells induced by LPS,And this inhibitory effect of CHR may be related to the incresed expression of tight junction proteins.