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Aim:To design and screen antisense oligodeoxynucleotides(ASODNs),whichinhibit type-1 plasminogen activator inhibitor(PAI-1)expression in human umbili-cal vein endothelial cells(HUVEC)in vitro.Methods:Twenty seven ASODNsagainst different sites of PAI-1 mRNA were designed and transfected to HUVECby lipofectin in vitro.The effects of ASODNs on PAI-1 antigen,PAI-1 activityand PAI-1 mRNA expression were detected by ELISA,amidolytical assay and RT-PCR,respectively.Results:Transforming growth factor β_1(TGF-β_1)-treatedHUVEC increased the expression of PAI-1 compared with the normal HUVEC.Five among twenty seven designed ASODNs were effective in inhibiting the in-crease in PAI-1 antigen and PAI-1 activity in a dose-dependent manner after 48-htransfection.In particular,ASODN 14(AO14)exhibited the best inhibitory effect.The control sequences of AO14,including sense,scramble,and mismatchsequences,did not significantly inhibit PAI-1 activity.It was revealed that theinhibitory efficacy of AO14 was in a sequence-specific manner.RT-PCR showedthat ASODN 1,7,8,14,and 15 decreased PAI-1 mRNA expression induced byTGF-β_1 and AO14 showed the best inhibitory effect.Conclusion:ASODN 1,7,8,14,and 15,among twenty seven designed ASODNs against PAI-1 mRNA,signifi-cantly decreased PAI-1 antigen and PAI-1 activity induced by TGF-β_1x in a dose-dependent manner in HUVEC in vitro.AO14 showed the best inhibitory effect onPAl-1 expression in a sequence-specific manner.The results of RT-PCR indicatedthat inhibitory effects of ASODNs on PAI-1 biosynthesis occurred at the mRNAlevel.Four among five effective target sites of ASODNs located at the translationinitiation Site or within the translation area of PAI-1 mRNA,suggesting that thesesites may be promising sites for the design of effective ASODNs.
Aim: To design and screen antisense oligodeoxynucleotides (ASODNs), which inhibitor of type 1 plasminogen activator inhibitor (PAI-1) expression in human umbili-cal vein endothelial cells (HUVEC) in vitro. Methods: Twenty seven ASODNsagainst different sites of PAI-1 mRNA were designed and transfected to HUVECby lipofectin in vitro.The effects of ASODNs on PAI-1 antigen, PAI-1 activity and PAI-1 mRNA expression were detected by ELISA, amidolytic assay and RT-PCR, respectively.Results: Transforming growth factor β_1 (TGF-β1) -treated HUVEC increased the expression of PAI-1 compared with the normal HUVEC. Five of the seven destined ASODNs were effective in inhibiting the in-crease in PAI-1 antigen and PAI-1 activity in a dose-dependent manner after 48-htransfection.In particular, ASODN 14 (AO14) exhibited the best inhibitory effect. The control sequences of AO14, including sense, scramble, and mismatch sequences, did not significantly inhibit PAI-1 activity. It was revealed that the inhibitory effect of AO14 w PAI-1 mRNA expression induced by TGF-β_1 and AO14 showed the best inhibitory effect. Conlusion: ASODN 1,7,8, 14, and 15, among twenty seven designed ASODNs against PAI-1 mRNA, signifi-cantly decreased PAI-1 antigen and PAI-1 activity induced by TGF-β_1x in a dose-dependent manner in HUVEC in vitro. effect onPAl-1 expression in a sequence-specific manner. The results of RT-PCR indicated that inhibitory effects of ASODNs on PAI-1 biosynthesis occurred at the mRNA level. Flow among five effective target sites of ASODNs located at the translation initiation site or within the translation area of PAI-1 mRNA, suggesting that thesesites may be promising sites for the design of effective ASODNs.