摘要
AIM: To test the hypothesis that introduction of antisense TβR Ⅰ and TβR Ⅱ eukaryotic expressing plasmids into a rat model of immunologically induced liver fibrosis might block the action of TGF-β1 and halt the progression of liver fibrosis. METHODS: RT-Nest-PCR and gene recombination techniques were used to construct rat antisense TβR Ⅰ and TβR Ⅱ recombinant plasmids which could be expressed in eukaryotic cells. The recombinant plasmids and empty vector (pcDNA3) were encapsulated by glycosyl-poly-L-lysine and then transducted into rats of pig serum-induced liver fibrosis model. Expression of exogenously transfected gene was assessed by Northern blot, and hepatic expressions of TβR Ⅰ and TβR Ⅱ were evaluated by RT-PCR and Western blot.We also performed ELISA for serum TGF-β1, hydroxyproline of hepatic tissues, immunohistochemistry for collagen types Ⅰ and Ⅲ, and VG staining for pathological study of the liver tissues. RESULTS: The exogenous antisense TβR Ⅰ and TβR Ⅱ plasmids could be well expressed in vivo, and block mRNA and protein expression of TβR Ⅰ and TβR Ⅱ in the fibrotic liver at the level of mRNA respectively. These exogenous plasmid expressions reduced the level of TGF-β1 (antisense TβR Ⅰ group 23.998+3.045 ng/mL, antisense TβR Ⅱ group 23.156+3.131 ng/mL, disease control group 32.960+3.789 ng/mL; F-=-38.19, 36.73, P<0.01). Compared with disease control group, the contents of hepatic hydroxyproline (antisense TβR Ⅰ group 0.169+0.015 mg/g liver, antisense TβR Ⅱ group 0.167+0.009 mg/g liver, disease control group 0.296+0.026 mg/g liver; F=14.39, 15.48, P<0.01) and the deposition of collagen types Ⅰ and Ⅲ decreased in the two antisense treatment groups(antisense TβR Ⅰ group, collagen type Ⅰ 669.90+50.67, collagen type Ⅲ 657.29+49.48; antisense TβR Ⅱ group, collagen type Ⅰ 650.26+51.51, collagen type Ⅲ 661.58+55.28; disease control group, collagen type I 1209.44+116.60, collagen type Ⅲ 1175.14+121.44; F=15.48 to 74.89, P<0.01). Their expres
AIM: To test the hypothesis that introduction of antisense TβR Ⅰ and TβR Ⅱ eukaryotic expressing plasmids into a rat model of immunologically induced liver fibrosis might block the action of TGF-β_1 and halt the progression of liver fibrosis. METHODS: RT-Nest-PCR and gene recombination techniques were used to construct rat antisense TβR Ⅰ and TβR Ⅱ recombinant plasmids which could be expressed in eukaryotic cells.The recombinant plasmids and empty vector (pcDNA3) were encapsulated by glycosyl-poly-L- lysine and then transducted into rats of pig serum-induced liver fibrosis model.Expression of exogenously transfected gene was assessed by Northern blot,and hepatic expressions of TβR Ⅰ and TβR Ⅱ were evaluated by RT- PCR and Western blot.We also performed ELISA for serum TGF-β_1,hydroxyproline of hepatic tissues,immunohistoche- mistry for collagen types Ⅰ and Ⅲ,and VG staining for pathological study of the liver tissues. RESULTS: The exogenous antisense TβR Ⅰ and TβR Ⅱ plasmids could be well expressed in vivo,and block mRNA and protein expression of TβR Ⅰ and TβR Ⅱ in the fibrotic liver at the level of mRNA respectively.These exogenous plasmid expressions reduced the level of TGF-β_1 (antisense TβR Ⅰ group 23.998±3.045 ng/mL,antisense TβR Ⅱ group 23.156±3.131 ng/mL,disease control group 32.960±3.789 ng/mL; F=38.19,36.73,P<0.01).Compared with disease control group,the contents of hepatic hydroxyproline (antisense TβR Ⅰ group 0.169±0.015 mg/g liver,antisense TβR Ⅱ group 0.167±0.009 mg/g liver, disease control group 0.296±0.026 mg/g liver; F=14.39, 15.48,P<0.01) and the deposition of collagen types Ⅰ and Ⅲ decreased in the two antisense treatment groups (antisense TβR Ⅰ group,collagen type Ⅰ 669.90±50.67, collagen type Ⅲ 657.29±49.48; antisense TβR Ⅱ group, collagen type Ⅰ 650.26±51.51,collagen type Ⅲ 661.58±55.28; disease control group,collagen type Ⅰ 1209.44±116.60, collagen type Ⅲ 1175.14±121.44; F=15.48 to 74.89,P<0.01). Their expres
