The Interference Effect Of TGF-β1 Small Interference RNA On Human Lung Fibroblast

Objectives:Idiopathic pulmonary fibrosis is a disease which has a high incidence and mortality rate. Actual research data has indicated that cellulate factor and its network play significant roll in the disease .So, TGF-β1 is an important factor about Human pulmonary fibrosis, TGF-β1 is secreted by lymphocyte and monocyte,It has many bioactivities such as:It can provoked immature lung fibroblasts to grow and proliferate,stimulate cells to differentiate and matrixes to synthetize, increase congestion of ECM but inhibit it,s degradation. TGF-β1 can effect transcription and contranslation on the processes of collagen, encourage formation and deposition of collagen protein. Accordingly, In the process of pulmonary fibrosis ,TGF-β1 is an important factor,So inhibiting the expression of TGF-β1 is so much helpful to delay it, but the effect is discontented when researchers have applicated the conventional drugs of anti-fibrosis to inhibit the expression of TGF-β1.Recently, with the discovery and development of the RNA interference(RNAi) technique, RNAi technique has been widely application in the research of genic regulation and control.RNAi is the most transcription gene silence induced by dsRNA,which can silence gene specifically and result in gene functional incapacitation or reduction.In my study, I designed and composed shRNA which had aimed directly at TGF-β1 mRNA of human, transfected into the cultural LFb of human by lipoplast, observed and analysised the inhibition effect on the genetic expression of TGF-β1 and to provide the new methods to prevention and cure ILD.Methods:One 19-nucleotides siRNA targeting the TGF-β1 mRNA sequence 952-970 and constructing the green fluorescence plasmid DNA of expressing TGF-β1 siRNA were transfected into the cultural LFb of human by lipoplast. AT 24,48 and 72 hours, the preliminary effect was evaluated with green fluorescence microscope, and the TGF-β1 mRNA expression was detected by Real-time PCR and the TGF-β1 level was quantitatively by ELISA. Results: Obtaining the human TGF-β1 mRNA sequence from GenBank, we choosed one gene locus and designed one sequence of siRNA by the software of Block-iTTM RNAi Designer, hand the work to jingsai company; 24,48,72 hours after transfection, observing by the green fluorescence microscope, Fluorescence was observed in the TGF-β1 siRNA plasmid groups ,Fluorescence was not observed in the null plsmid groups; at differently time-points of the transfection, the TGF-β1 mRNA in the siRNA group were down regulated and the TGF-β1 protein in the siRNA group was down regulated respectively.Conclusions: Our data suggested that : the expression of TGF-β1 in siRNA group was visibly lower than control group, it was proved that specific siRNA could efficiently inhibit the expression of target gene.