| Objective:The aim of the experiment is to evaluate the role of matrine and asiaticoside in prevention and treatment of COPD and airway remodeling through exploring the regulative effects of matrine and asiaticoside on viabiling of HFL-1stimulated by inflammative and non-inflammative mediators.Methord:The HLF-1cultured in vitro was used in the experiment. Matrine(0.1、0.2、0.4、0.8、1.6、3.2、6.4mmol/L)andasiaticoside(6.25、12.5、25、50、100、200、400ug/ml)were used at various concentration in culture medium. The quantity of HLF-1was measured with CCK-8. The mRNA of TGF-β1、CTGF and Type I Collagen of HLF-1induced by Angiotensin Ⅱ10-7mol/L were measured by Realtime PCR after interference with matrine and asiaticoside. The method of Realtime PCR was also used to measure the mRNA of HIF-la and VEGF induced by hypoxia(2%O2) with the interference of matrine and asiaticoside.Results:(1)The CCK-8Test: After intervene of matrine with various concentration, the OD value of HLF-1in control group was0.81±0.06; the diversity was not significant at concentration of0.1、0.2、0.4mmol/L (P>0.05); the concentration above0.8mmol/L could suppress the proliferation of HLF-1significantly (P<0.01)After intervene of matrine with various concentration, the OD value of HLF-1in control group was0.82±0.07, the concentration from6.25to200ug/ml did not have the effect of suppressing the proliferation of HLF-1(P>0.05), while the400ug/ml had thie effect (P<0.01)(2)After being induced by Angiotensin Ⅱ10-7mol/L for24hours, the TGF-β1mRNA of HFL-1was up regulated to206.37±129.83(P<0.05), the CTGF to21.92±7.58(P<0.01), while Type I Collagen mRNA to368.64±192.78(P<0.01), all of which had significant contrast comparing to the control group.On the base of Angiotensin Ⅱ inducing, matrine of0.8mmol/L concentration was added. As a result, the mRNA of TGF-β1up-regulated by Angiotensin Ⅱ was depressed to1.86±0.10(P<0.05), the CTGF to0.0014±0.00071(P<0.01), and Type Ⅰ Collagen mRNA expression decreased to0.46±0.36(P<0.01), all of which had significant contrast comparing to the Angiotensin II induced group.On the base of Angiotensin II inducing, asiaticoside of400ug/ml concentration was added. As a result, the mRNA of TGF-β1up regulated by Angiotensin II was depressed to2.31±1.39(P<0.05), the CTGF to0.0017±0.00163(P<0.01), and Type I Collagen mRNA expression decreased to0.51±0.32(P<0.01), all of which had significant contrast comparing to the Angiotensin Ⅱ induced group.(3)After being induced by2%O2for24hours, the mRNA of HIF-la was up regulated to19.50±11.54(P<0.01), while the VEGF mRNA increased to157.46±112.30(P<0.01), both of which had significant contrast comparing to the control group.On the base of2%O2inducing, matrine of0.8mmol/L concentration was added. As a result, the mRNA of HIF-1α up regulated by hypoxia was depressed to0.78±0.74(P<0.01), and the VEGF mRNA expression decreased to5.53±5.89(P<0.05), both of which had significant contrast comparing to the hypoxia group.On the base of2%O2inducing, asiaticoside of400ug/ml concentration was added. As a result, the mRNA of HIF-1α up regulated by hypoxia was depressed to0.14±0.08(P<0.01), and the VEGF mRNA expression decreased to3.56±3.42(P<0.05), both of which had significant contrast comparing to the hypoxia group.Conclusions:(1)In the normal condition, the HLF-1can express the mRNA of TGF-β1、CTGF、 Type Ⅰ Collagen、HIF-1α and VEGF in a small amount.(2)After being induced by Angiotensin Ⅱ, the TGF-β1、CTGF and Type Ⅰ Collagen mRNAexpression of HLF-1were up regulated, which means Angiotensin Ⅱ is an inducer in the proliferation of HLF-1.(3)After induced by hypoxia, the HIF-1α and VEGF mRNA expression of HLF-1were also up regulated, which means hypoxia may play an inprotant role in the process of COPD and airway remodeling.(4)The proliferation of HLF-1in matrine group was reduced markedly in aconcentration dependent manner in center concentration extent. The medicine also could down regulate the mRNA expression of TGF-β1、 CTGF and Type Ⅰ Collagen of HLF-1after the stimulus of Angiotensin Ⅱ. Meanwhile, it could down regulate the mRNA expression of HIF-la and VEGF induced by hypoxia at the same time.(5)The low concentration of asiaticoside did not reduce the proliferation of HLF-1, which may because of its low cell toxicity. But it could reduce the TGF-β1、CTGF and Type I Collagen mRNA expression of Angiotensin II stimulated cells and HIF-1α and VEGF mRNA expression of hypoxia induced cells.Therefore, matrine and asiaticoside can depress the proliferation of HLF-1and down regulate the mRNA expression of inflammatory factors. Thus, the drugs may postpone the proceeding of airway remodeling in cell level. |
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