AIM: To detect whether N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) inhibits epithelial-mes-enchymal transition in A549 cells induced by TGF-β1 through suppressing the expression of heat shock protein 27 (HSP27) and zinc finger proteins Snail (including SNAI1and SNAI2) which ultimately inhibited the deposition of type I and type III collagens.METHODS:The colocalizations of HSP27 and SNAI1/SNAI2 respectively on A549 alveolar epi-thelial cells induced by TGF-β1 were measured by confocal microscopy .The expression of HSP27, SNAI1 and SNAI2 at mRNA level was detected by real-time PCR.Western blotting analysis was used to detect the expression of HSP 27, SNAI1 and SNAI2 on epithelial-mesenchymal transition in A549 cells induced by TGF-β1 and also the deposition of type I and type III collagens in A549 cells transfected with HSP27shRNA prior to TGF-β1 stimulation.RESULTS: Compared with control group, TGF-β1 increased the expression of HSP27, SNAI1, SNAI2, type I and type III collagen, which decreased significantly followed by Ac-SDKP intervention.The expression of SNAI1, type I and type III collagen decreased signifi-cantly after transfected with HSP27shRNA in A549 cells, which had the similar effect on Ac-SDKP intervention.CON-CLUSION:Ac-SDKP inhibits the transition of cultured A 549 cells to myofibroblasts and attenuates collagen synthesis by suppressing the expression of HSP 27 and zinc finger proteins SNAI 1 and SNAI2.

[Abstract ] Objective Silicosis is one of the most serious occupational diseases in China .In this study,we explored the reg -ulatory effect of N-acetyl-seryl-aspartyl-lysyl-proline ( Ac-SDKP ) on angiotensin ( Ang ) Ⅱ-induced extracellular signal-regulated ki-nase ( ERK1/2) and Jun N-terminal kinase ( JNK) signals and its inhibitory effect on the differentiation of human embryonic lung MRC-5 fibroblasts to myofibroblasts via Ang Ⅱ-induced ERK1/2 and JNK signals . Methods Human embryonic lung MRC-5 fibro-blasts were induced by Ang Ⅱand pre-treated with the JNK signal inhibitor ( SP600125 ) , the ERK1/2 signal inhibitor ( PD98059 ) or Ac-SDKP.The proliferation of the cells was measured by MTT assay .The expressions of αS-MA, SRF, p-ERK1/2 and p-JNK were determined by immunocytochemical staining , and the expression levels of these proteins and collagen Ⅰwere detected by Western blot .Results The A value of Ang Ⅱ group (0.56 ±0.08) measured by MMT assay was 2.07 fold as control group ( 0.27 ±0.05 ). Pretreatment with SP600125 , PD98059 and Ac-SDKP, the A value were (0.39 ±0.02), (0.40 ±0.03) and (0.36 ±0 0.5) that had a statistical significance with Ang Ⅱgroup.The up-regulation of colla-gen type Ⅰ,α-SMA, SRF were induced by Ang Ⅱ by 4.50, 3.50 and 3.00 fold compared with control group.Moreover, the expression of p-ERK1/2 and p-JNK were increased as 6.71 and 7.90 fold as control. Pre-treatment with Ac-SDKP could inhibit p-JNK and p-ERK1/2 to 29.79% and 46.84% compared with AngⅡ group. Conclusion Ac -SDKP can inhibit the differentiation of human embryonic lung MRC-5 fibroblasts to myofibroblasts by regulating AngⅡ-induced JNK and ERK1/2 signals.

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

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