OBJECTIVETo explore the inhibition effect of N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) on myofibroblast differentiation of MRC-5 human fetal lung fibroblasts induced by angiotensin (Ang) II.

METHODSThe study was divided into 2 step: (1) MRC-5 human fetal lung fibroblasts was induced for 48 h at different dose of Ang II and at different time point by 100 nmol/L Ang II. Then the expression of collagen type I and α-smooth muscle actin (α-SMA) were mesaured by western blot. (2) MRC-5 human fetal lung fibroblasts were divided into 4 group: (1) control, (2) Ang II, (3) Ang II+Ac-SDKP, (4) Ang II+8-Me-cAMP (a specific activator of Epac). The α-SMA expression was observed by immnocytochemical stain. The protein expression of collagen type I, α-SMA, serum response factor (SRF), myocardin-related transcription factor (MRTF)-A, exchange protein directly activated by cAMP (Epac) 1, 2 were measured by Westen blot.

RESULTSMyofibroblast differentiation could be induced by Ang II from MRC-5 cells with a dose- and time-dependent manner. The up-regulation of SRF and MRTF-A were observed in MRC-5 cells induced by Ang II and accompanied with collagen I and α-SMA increased. Pre-treatment with 8-Me-cAMP or Ac-SDKP could attenuated all this changes induced by Ang II, and promoted the expression of Epac1.

CONCLUSIONAc-SDKP can inhibit the myofibroblast differentiation of MRC-5 cells induced by Ang II via Epac1 activating.

Actins ; Angiotensin II ; Cell Differentiation ; drug effects ; Collagen ; Collagen Type I ; Cyclic AMP ; analogs & derivatives ; Fetus ; cytology ; Fibroblasts ; cytology ; Guanine Nucleotide Exchange Factors ; Humans ; Lung ; cytology ; Myofibroblasts ; drug effects ; Oligopeptides ; pharmacology ; Serum Response Factor ; Trans-Activators

OBJECTIVETo perform a comparative proteomic analysis for identification of pulmonary proteins related to the progression of silicosis and anti-fibrotic effect of N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP).

METHODSBronchial instillation of SiO₂powder (for 4 or 8 weeks) was applied in rats to establish a silicosis model. Ac-SDKP treatment was performed before (prevention group) or after (treatment group) SiO₂instillation. The control group was treated by bronchial instillation of sodium chloride solution of the same volume as SiO₂powder for 4 or 8 weeks. Proteins in lung tissue were separated by two-dimensional gel electrophoresis and stained with colloidal Coomassie brilliant blue. The gel images were scanned with the Lab Scan III system and analyzed with Imagemaster 6.0. The protein spots with significant differences between two groups (i.e., P value was less than 0.05 in One-way ANOVA) and with a change in volume over 30% were defined as differential proteins. Comparison was performed between the silicosis group and control group after 4 or 8 weeks, between the Ac-SDKP treatment group and silicosis group after 8 weeks, and between the Ac-SDKP prevention group and silicosis group after 8 weeks. The differentially expressed proteins were subjected to in-gel digestion with trypsin and MALDI-TOF-MS and Mascot search engine analysis to identify these proteins.

RESULTSThirty-three differential proteins were identified. In comparison with the control group (4 weeks), the silicosis group (4 weeks) had 17 up-regulated proteins and 11 down-regulated proteins. In comparison with the control group (8 weeks), the silicosis group (8 weeks) had 16 up-regulated proteins and 12 down-regulated proteins. In comparison with the silicosis group (8 weeks), the Ac-SDKP treatment group had 5 up-regulated proteins and 6 down-regulated proteins, and the Ac-SDKP prevention group had 8 up-regulated proteins and 10 down-regulated proteins.

CONCLUSIONCritical regulatory proteins related to silicotic fibrosis and anti-silicotic effect of Ac-SDKP have been identified. These proteins may play an important role in proliferation, apoptosis, inflammation, epithelial-mesenchymal transition, and signal transduction in silicosis.

Animals ; Disease Models, Animal ; Lung ; metabolism ; Male ; Oligopeptides ; therapeutic use ; Proteome ; metabolism ; Rats ; Rats, Wistar ; Silicosis ; drug therapy ; metabolism