Objective To explore the effect of TGF-a1 on the mRNA and prot ei n expression of connective tissue growth factor (CTGF) in human peritoneal mesot helial cells (HPMCs) and the probable mechanism. Methods Greater oment...Objective To explore the effect of TGF-a1 on the mRNA and prot ei n expression of connective tissue growth factor (CTGF) in human peritoneal mesot helial cells (HPMCs) and the probable mechanism. Methods Greater omenta from hea lthy adults were used for primary culture of HPMCs. The third generation cells w ere stimulated by 5 ng/ml TGF-a1. Immunohistochemistry, Western blot, ELISA an d RT-PCR were employed to examine the follows:the mRNA and protein expression o f CTGF, the mRNA and protein expression of fibronectin (FN) and collagen type Ⅰ (ColⅠ), the protein expression of p-Smad2/3 and its migration in HPMCs. Result s (1)The mRNA expression of CTGF in the treatment groups increased obviously a s compared to the control group, and peaked at 48 h. (2)The protein expression of intracellular CTGF in the control group was very little, whereas it increase d markedly 24 h after TGF-a1 stimulation, and peaked at 48 h. The mRNA express ion of intracellular FN, and the protein expression of supernatent FN, as well a s the mRNA and protein expression of intracellular ColⅠin the treatment group u p-regulated significantly in a time dependent manner as compared to those in th e control group. (3)p-Smad2/3 in HPMCs hardly expressed in the control group( 3%p-Smad2/3-positive cells), and remarkably expressed 15 min after TGF-a1 s timulation(29%), peaked at 1 h (84%) and then decreased after 2 h(37%). Meanw hile, p-Smad2/3 mainly distributed in cytoplasm at 15 min, concentrated in cell nucleus and peri-nucleus at 1 h, and distributed in cytoplasm again at 2 h. Co nclusions TGF-a1 can induce the mRNA and protein expression of CTGF in the pro gression of human peritoneal fibrosis. The probably mechanism is that signaling through Smad pathway in HPMCs can be activated by TGF-a1 stimulation.展开更多
文摘Objective To explore the effect of TGF-a1 on the mRNA and prot ei n expression of connective tissue growth factor (CTGF) in human peritoneal mesot helial cells (HPMCs) and the probable mechanism. Methods Greater omenta from hea lthy adults were used for primary culture of HPMCs. The third generation cells w ere stimulated by 5 ng/ml TGF-a1. Immunohistochemistry, Western blot, ELISA an d RT-PCR were employed to examine the follows:the mRNA and protein expression o f CTGF, the mRNA and protein expression of fibronectin (FN) and collagen type Ⅰ (ColⅠ), the protein expression of p-Smad2/3 and its migration in HPMCs. Result s (1)The mRNA expression of CTGF in the treatment groups increased obviously a s compared to the control group, and peaked at 48 h. (2)The protein expression of intracellular CTGF in the control group was very little, whereas it increase d markedly 24 h after TGF-a1 stimulation, and peaked at 48 h. The mRNA express ion of intracellular FN, and the protein expression of supernatent FN, as well a s the mRNA and protein expression of intracellular ColⅠin the treatment group u p-regulated significantly in a time dependent manner as compared to those in th e control group. (3)p-Smad2/3 in HPMCs hardly expressed in the control group( 3%p-Smad2/3-positive cells), and remarkably expressed 15 min after TGF-a1 s timulation(29%), peaked at 1 h (84%) and then decreased after 2 h(37%). Meanw hile, p-Smad2/3 mainly distributed in cytoplasm at 15 min, concentrated in cell nucleus and peri-nucleus at 1 h, and distributed in cytoplasm again at 2 h. Co nclusions TGF-a1 can induce the mRNA and protein expression of CTGF in the pro gression of human peritoneal fibrosis. The probably mechanism is that signaling through Smad pathway in HPMCs can be activated by TGF-a1 stimulation.
