Objective. To elucidate the role of transforming growth factor-beta (TGF-beta)/Smad signalling in the increased expression of the collagen gene in systemic sclerosis (SSc) fibroblasts. Methods. Dermal fibroblasts from seven patients with diffuse SSc of recent onset and from seven healthy individuals were studied. The expression levels of Smad2, Smad3 and Smad4 proteins were determined by immunoblotting. Smad3 phosphorylation and the interaction of Smad3 with Sp1 or p300 were analysed using immunoprecipitation. The effects of overexpression of Smad proteins or Sp1 on the human alpha 2(I) collagen gene transcription were investigated with chloramphenicol acetyltransferase (CAT) assays using the -772 COL1A2/CAT construct. Results. Constitutive increased Smad3 phosphorylation was detected in SSc fibroblasts compared with normal fibroblasts. Increased interaction of Smad3 with Sp1 as well as p300 was also detected in SSc fibroblasts. The overexpression of Smad3 caused an increase of up to 5-fold in COL1A2 promoter activity in normal fibroblasts, while Smad3 caused a small increase in COL1A2 promoter activity in SSc fibroblasts. However, neither Smad2 nor Smad4 caused significant effects in COL1A2 promoter activity in normal fibroblasts or SSc fibroblasts. The overexpression of Sp1 caused further increase in COL1A2 promoter activity stimulated by TGF-beta in normal fibroblasts, but did not change COL1A2 promoter activity in the presence of TGF-beta in SSc fibroblasts. The combined overexpression of Smad3 and Sp1 significantly enhanced TGF-beta response in normal fibroblasts, but less markedly in SSc fibroblasts. Conclusions. These results suggested that SSc fibroblasts are less sensitive to exogenous TGF-beta stimulation because they are already activated by the autocrine TGF-beta loop.
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Yu, Shenghuan
Zeng, Long
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Zeng, Long
Rao, Fang
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Res Ctr Med Sci, Guangzhou, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Rao, Fang
Deng, Chunyu
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Res Ctr Med Sci, Guangzhou, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Deng, Chunyu
Zhang, Mengzhen
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Res Ctr Med Sci, Guangzhou, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Zhang, Mengzhen
Xiao, Haiyin
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Res Ctr Med Sci, Guangzhou, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Xiao, Haiyin
Xiao, Feifei
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Xiao, Feifei
Xue, Yumei
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Cardiovasc Inst, Guangzhou, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Xue, Yumei
Wu, Shulin
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Cardiovasc Inst, Guangzhou, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Wu, Shulin
Du, Zhimin
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Dongguan Tungwah Songshan Lake Hosp, Dongguan, Peoples R China
Dongguan Tungwah Songshan Lake Hosp, Dongguan 523820, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China
Du, Zhimin
Wei, Wei
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Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Cardiovasc Inst, Guangzhou, Peoples R China
Guangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Cardiovasc Inst, Guangzhou 510080, Peoples R ChinaGuangdong Acad Med Sci, Guangdong Prov Peoples Hosp, Guangdong Prov Key Lab Clin Pharmacol, Guangzhou, Peoples R China