BACKGROUND: Previous studies have shown that transforming growth factor-beta 1 (TGF-beta 1) is the most potent means of stimulating liver fibrogenesis by myofibroblast-like cells derived from hepatic stellate cells. T...BACKGROUND: Previous studies have shown that transforming growth factor-beta 1 (TGF-beta 1) is the most potent means of stimulating liver fibrogenesis by myofibroblast-like cells derived from hepatic stellate cells. Thus, TGF-beta 1 could be a target for treating hepatic fibrosis. This study aimed to investigate the inhibitory effects of specific TGF-beta 1 small interference RNA (siRNA) on immune hepatic fibrosis induced by Concanavalin A (Con A) in mice. METHODS: Three short hairpin RNAs targeting different positions of TGF-beta 1 were designed and cloned to the plasmid pGenesil-1 to obtain three recombinant expression vectors (pGenesil-TGF-beta 1-ml, pGenesil-TGF-beta 1-m2 and pGenesil-TGF-beta 1-m3). Thirty male Kunming mice were randomly divided into 6 groups: normal, model, control, and three treatment groups. The immune hepatic fibrosis models were constructed by injecting Con A via the tail vein at 8 mg/kg per week for 6 weeks. At weeks 2, 4 and 6, pGenesil-TGF-beta 1-ml, pGenesil-TGF-beta 1-m2 or pGenesi1-TGF-beta 1-m3 was injected by a hydrodynamics-based transfection method via the tail vein at 0.8 ml/10 g within 24 hours after injection of Con A in each of the three treatment groups. The mice in the control group were injected with control plasmid pGenesil-HK at the same dose. All mice were sacrificed at week 7. The levels of hydroxyproline in liver tissue were determined by biochemistry. Liver histopathology was assessed by Van Gieson staining. The expression levels and localization of TGF-beta 1, Smad3, and Smad7 in liver tissue were detected by immunohistochemistry. The expression of TGF-beta 1, Smad3, Smad7 and alpha-smooth muscle actin (alpha-SMA) mRNAs in the liver were assessed by semi-quantitative RT-PCR. RESULTS: The levels of hydroxyproline in the liver tissue of the treatment groups were lower than those of the model group (P<0.01). Histopathologic assay showed that liver fibrogenesis was clearly improved in the treatment groups compared with the model group. The expression levels of TGF-be展开更多
Background Signal transducer and activator of transcription 3 （STAT3） is usually constitutively activated in a variety of malignancies. It directly contributes to tumorigenesis, invasion, and metastasis. The surgica...Background Signal transducer and activator of transcription 3 （STAT3） is usually constitutively activated in a variety of malignancies. It directly contributes to tumorigenesis, invasion, and metastasis. The surgical treatment of breast cancer has made no breakthroughs in terms of treatment effect, in spite of its long history. Current biotherapies bring a note of optimism to breast cancer treatment. To explore the possibility of a siRNA targeted STAT3 blocking treatment for over-activated tumor cells, we evaluated the efficacy of a STAT3 siRNA on human breast cancer cells in vitro and in vivo. Methods Three MCF-7 human breast cancer cell lines were tested： control MCF-7 cells, non-specific siRNA transfected MCF-7 cells and STAT3 siRNA transfected MCF-7 cells. Expression of STAT3 in MCF-7 cells was inhibited by RNA interference （RNAi）. The STAT3 mRNA and protein levels were detected by semi-quantity RT-PCR and Western blotting. Cell proliferation and apoptosis were determined by MTT method and flow cytometry. The three groups of MCF-7 cells mentioned above were transplanted subcutanuously into nude mice and their tumorgenic ability observed. The STAT3 mRNA and protein levels of the samples from tumors in different groups were determined by semi-quantity RT-PCR and Western blotting and compared. Results In STAT3 siRNA transfected MCF-7 cells, the expressions （STAT3/13-actin） of STAT3 mRNA （0.327±0.020） and protein （0.153±0.006） were significantly lower than that in control MCF-7 cells （mRNA 1.093±0.018, protein 1.374±0.022） and non-specific siRNA transfected MCF-7 cells （mRNA 1.035_±0.050, protein 1.320±0.033） （P 〈0.05）. MTT showed that cell proliferation was significantly reduced and the cell growth inhibition ratio in the STAT3-siRNA group was （44.00±5.10）%, significantly higher than that in non-specific siRNA transfected MCF-7 cells （（16.10_±1.05）%, P 〈0.05）. Flow cytometry results showed that more apoptosis was observed in the STAT3-siRNA group. The rat展开更多
文摘BACKGROUND: Previous studies have shown that transforming growth factor-beta 1 (TGF-beta 1) is the most potent means of stimulating liver fibrogenesis by myofibroblast-like cells derived from hepatic stellate cells. Thus, TGF-beta 1 could be a target for treating hepatic fibrosis. This study aimed to investigate the inhibitory effects of specific TGF-beta 1 small interference RNA (siRNA) on immune hepatic fibrosis induced by Concanavalin A (Con A) in mice. METHODS: Three short hairpin RNAs targeting different positions of TGF-beta 1 were designed and cloned to the plasmid pGenesil-1 to obtain three recombinant expression vectors (pGenesil-TGF-beta 1-ml, pGenesil-TGF-beta 1-m2 and pGenesil-TGF-beta 1-m3). Thirty male Kunming mice were randomly divided into 6 groups: normal, model, control, and three treatment groups. The immune hepatic fibrosis models were constructed by injecting Con A via the tail vein at 8 mg/kg per week for 6 weeks. At weeks 2, 4 and 6, pGenesil-TGF-beta 1-ml, pGenesil-TGF-beta 1-m2 or pGenesi1-TGF-beta 1-m3 was injected by a hydrodynamics-based transfection method via the tail vein at 0.8 ml/10 g within 24 hours after injection of Con A in each of the three treatment groups. The mice in the control group were injected with control plasmid pGenesil-HK at the same dose. All mice were sacrificed at week 7. The levels of hydroxyproline in liver tissue were determined by biochemistry. Liver histopathology was assessed by Van Gieson staining. The expression levels and localization of TGF-beta 1, Smad3, and Smad7 in liver tissue were detected by immunohistochemistry. The expression of TGF-beta 1, Smad3, Smad7 and alpha-smooth muscle actin (alpha-SMA) mRNAs in the liver were assessed by semi-quantitative RT-PCR. RESULTS: The levels of hydroxyproline in the liver tissue of the treatment groups were lower than those of the model group (P<0.01). Histopathologic assay showed that liver fibrogenesis was clearly improved in the treatment groups compared with the model group. The expression levels of TGF-be
文摘Background Signal transducer and activator of transcription 3 （STAT3） is usually constitutively activated in a variety of malignancies. It directly contributes to tumorigenesis, invasion, and metastasis. The surgical treatment of breast cancer has made no breakthroughs in terms of treatment effect, in spite of its long history. Current biotherapies bring a note of optimism to breast cancer treatment. To explore the possibility of a siRNA targeted STAT3 blocking treatment for over-activated tumor cells, we evaluated the efficacy of a STAT3 siRNA on human breast cancer cells in vitro and in vivo. Methods Three MCF-7 human breast cancer cell lines were tested： control MCF-7 cells, non-specific siRNA transfected MCF-7 cells and STAT3 siRNA transfected MCF-7 cells. Expression of STAT3 in MCF-7 cells was inhibited by RNA interference （RNAi）. The STAT3 mRNA and protein levels were detected by semi-quantity RT-PCR and Western blotting. Cell proliferation and apoptosis were determined by MTT method and flow cytometry. The three groups of MCF-7 cells mentioned above were transplanted subcutanuously into nude mice and their tumorgenic ability observed. The STAT3 mRNA and protein levels of the samples from tumors in different groups were determined by semi-quantity RT-PCR and Western blotting and compared. Results In STAT3 siRNA transfected MCF-7 cells, the expressions （STAT3/13-actin） of STAT3 mRNA （0.327±0.020） and protein （0.153±0.006） were significantly lower than that in control MCF-7 cells （mRNA 1.093±0.018, protein 1.374±0.022） and non-specific siRNA transfected MCF-7 cells （mRNA 1.035_±0.050, protein 1.320±0.033） （P 〈0.05）. MTT showed that cell proliferation was significantly reduced and the cell growth inhibition ratio in the STAT3-siRNA group was （44.00±5.10）%, significantly higher than that in non-specific siRNA transfected MCF-7 cells （（16.10_±1.05）%, P 〈0.05）. Flow cytometry results showed that more apoptosis was observed in the STAT3-siRNA group. The rat
