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Objective: To study the effect and mechanism of tanshinone IIA on human skin fibroblasts cell (HSF). Methods: CCK- 8 method was used to determine the effect of different concentrations of TSA on the proliferation of HSF induced by TGF-β1. The plate cloning ability of HSF treated with different concentrations of TSA (2.5, 5, 10 and 20 μmol/L) for 48 h were analyzed by plate clonogenesis assay. The protein expression of TGF-β1/Smad signaling pathway proteins and α-SMA, VEGFA and COL I were further measured by Western blotting. Results: CCK-8 and plate clonognesis assay results showed that TSA significantly inhibited the proliferation and colony forming efficiency of HSF (P < 0.01). Western blotting results revealed that each concentration group of TSA significantly inhibited the protein levels of p-Smad2 and p-Smad3, and down-regulated the ratio of p-Smad2/Smad2 (P < 0.01). The ratio of p-Smad3/Smad3 was significantly decreased in 5, 10 and 20 μmol/L TSA groups. Additionally, the expression levels of α-SMA, VEGFA and COL I in HSF decreased significantly with the increase of TSA concentration (P < 0.01). Conclusion: TSA exhibits the inhibitory effect on proliferation of HSF, and its mechanism may be related to TGF-β1/Smads signaling pathway.
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Objective: To optimize the extraction technology of immune active glycoproteins AmPR10-16kD and HQGP-2 from Astragalus membranaceus var. mongholicus (AMM). Methods: The optimized extraction temperature conditions were investigated by circular dichroism of water-soluble protein involving in AmPR10-16kD and HQGP-2 with secondary structure from AMM. The optimized extraction technology was investigated using single factor test and orthogonal test with gray value of water-soluble protein AmPR10-16kD and HQGP-2 as the index which was determined by Image of gel graphical analysis software. In this study, the effects of temperature, solid-liquid ratio, time, solvent, granularity, and times on gray value were investigated, for which the inhibitory effect of water-soluble protein was determined as an evidence by CCK-8 method, and the content of water-soluble protein is determined as an evidence by BCA method. Results: The optimized extraction technique for proteins AmPR10-16kD and HQGP-2 in AMM was established, that was 5.0 g powder of AMM over the No.4 sieve, olvent Tris-HCl, solid-liquid ratio 1:16 and 60 min for extraction at the temperature of 40 ℃ and being mixed under 100 r/min. The water-soluble protein extract rate in the orthogonal test analysis was 65 mg/g, of which inhibitory effect was 90.90% at a concentration of 90 μg/mL. Conclusion: The optimal extraction conditions could accurately reflect the relative amounts of AmPR10-16kD and HQGP-2 maximum extraction rate, providing a stable, reasonable, and feasible extraction process for further study of the bioactive substance of AMM.
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Objective:To prepare a kind of titanium implant doped with cobalt and to study its cytotoxicity.Methods:The surface of the titanium was anodized to form TiO2 nanotube arrays.Different amount of cobalt was doped by hydrothermal treatment,which was controlled by tuning the hydrothermal treatment duration.The cytotoxicity of the cobalt doped nanotubular implant coating on bone marrow stromal cells (BMSCs)was measured by CCK-8.Results:The nanotubular implant coating with different amount of cobalt was fabricated.The proliferation of BMSCs was inhibited by the nanotubular morphology and cobalt doping.Samples formed by hydro-thermal treatment in 0.1 M cobalt acetate showed significantly cytotoxicity.Conclusion:Hydrothermal treatment of anodized titanium is an effective way for developing novel cobalt doped nanotubular implant coating.The proper dose of cobalt doping needs to be further investigated.