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Fibroblasts sources, in addition to the traditional organizational sources, the peripheral blood sources has also been confirmed. Cycle into fiber cells (circulating fibrocytes, cFb) can express hematopoietic cell surface signs and synthesis of a variety of extracellular matrix proteins, type I collagen (Collagen Type I Collagen-Ⅰ), Ⅲ type collagen (Collagen Type Ⅲ Collagen-Ⅲ), α-smooth muscle actin (α-smooth muscle actin, α-SMA) as the main identification signs, it has the synthesis of a variety of extracellular matrix proteins, cytokines, and antigen presenting contraction of the wound, the ability to promote the formation of new blood vessels to participate in promote healing and the role of local tissue fibrosis. It has been confirmed that the blood circulation system exists the small cFb, the free Si02 dust on the research cycle fibroblasts has not been reported. Therefore, the experimental application of reverse transcriptase PCR assay Collagen-Ⅰ, Collagen-Ⅲ, α-SMA mRNA expression levels by ELISA Determination of Collagen-Ⅰ, Collagen-Ⅲ, α-SMA protein expression level, preliminary study in vitro SiO2 dust on cycle fibroblasts. Objective: To investigate the role of free Si02 dust of circulating fibroblasts, provide clues to cycle further explore the mechanism of action of pneumoconiosis in fibroblasts and experimental basis. Methods: A rat peripheral blood cFb intraperitoneal AM in vitro culture model, the MTT method to measure cell growth curve, immunohistochemical method identified by the supernatant stimulation but without Si02 stimulus and non supernatant stimulation cFb, stimulated with different concentrations (0,20,40,60,80,100,120 μg / ml) free Si02 dust in macrophages 24 h, centrifuged culture supernatant, and supernatant the stimulus cFb 24 h. 2. Of different Si02 dust concentration stimulate different training days cFb, relative expression levels of type I collagen type III, α-SMA mRNA by reverse transcriptase PCR assay, immunohistochemistry and ELISA method Ⅰ, Ⅲ collagen protein, α-SMA expression level. Results: 1.cFb is not the supernatant stimulus and the supernatant stimulation (the supernatant without Si02 stimulus) are cultured 6 d begin to express Collagen-Ⅰ, Collagen-Ⅲ, 19 d, the expression of α -SMA, cFb by supernatant stimulation (supernatant by Si02 stimulus) 3 d culture began to express different concentrations of the Si02 dust stimulate Collagen-Ⅰ, Collagen-Ⅲ, 13 d begin to express α-SMA.2. AM supernatant stimulus cFbCollagen-Ⅰ, Collagen-Ⅲ, α-SMA mRNA and protein expression levels: compared with supernatant group without, as 20, 40, 60 and 80 μg / ml concentration group difference was statistically significance (P lt; 0.05), expression gradually increased from 0 to 80μg/ml concentration group. Stimulate AM supernatant 3.80μg/ml SiO2 stimulate different culture days cFb ① Collagen-Ⅰ, Collagen-Ⅲ mRNA expression: 3 d compared 9,12,15,18 d difference was statistically significant (P lt ; 0.05), from the first 9 d with culture days increased expression increased gradually. ② α-SMA mRNA expression level: comparison and 13 d, 19,22,25,28 d difference was statistically significant (P lt; 0.05) increase in the number of days from 19 d with culture expression gradually increased. (3) Collagen-I and Collagen-III protein expression: 3 d compared to the first 9,12,15,18 d, the difference was statistically significant (P lt; 0.05), and increased expression with culture days from 9 d. ④ α-SMA protein expression: comparison with the first 13 d, 22,25,28 d difference was statistically significant (P lt; 0.05) increased expression with culture the number of days from the first 19 d. Conclusion: 1. Culture supernatants of AM stimulated by free SiO2 dust can cause blood lymphocyte advance time to cFb transformation. Free SiO2 dust stimulate the AM culture supernatant stimulation cFb with SiO2 concentration increased presence of Collagen-Ⅰ, Collagen-Ⅲ, α-SMA mRNA and protein expression increased.
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