ABSTRACT
Objective@#To investigate the effect of brucea javanica oil emulsion (BJOE) on the proliferation, collagen synthesis and ERK1/2 MAPK singaling pathway of hypertrophic scar fibroblasts (HSFs).@*Methods@#The human hypertrophic scar tissues were collected, and HSFs were isolated, cultured and identified in vitro. Cells were treated with BJOE under the concentration of 0.5, 1, 5, 10, 20 and 50 mg/ml for 24 h respectively; the half inhibitory concentration(IC50)of 24 h was calculated. The experimental groups were treated for 24, 48, 72 h by BJOE at the IC50, the control group was conventionally cultured without BJOE. Cell proliferation was detected by CCK-8 assay and cell cycle was determined by flow cytometry (FCM). The morphological changes of cells treated with BJOE at a concentration of IC50 for 24 h were observed. The protein expression levels of collagen Ⅰ, vimentin, ERK1/2 and p-ERK1/2 were measured by Western blot.@*Results@#The IC50 of BJOE was 1.176 mg/ml, so the concentration of 1 mg/ml was selected for the subsequent experiment. After 24, 48, 72 h treated with BJOE at a concentration of 1 mg/ml, the inhibition rates of BJOE on HSFs were (53.13±2.40)%, (75.07±2.67)%and (88.65±0.32)%, respectively. The difference was significant (P<0.05). Under the effect of BJOE, the number of HSFs significantly decreased, the interval between cells widened, cell protrusions were significantly shortened or even disappeared, and the cells became round. The percentage of cells decreased in S phase. Expression of type Ⅰ collagen, vimentin and p-ERK1/2 in HSF was significantly inhibited (P<0.05), while there was no significant difference in the expression of ERK1/2(P>0.05).@*Conclusions@#BJOE inhibites the proliferation of HSF and the synthesis of type Ⅰ collagen, and its mechanism is related to the inhibition of ERK1/2 phosphorylation and vimentin expression.
ABSTRACT
Objective: To screen the components with potent antiproliferative effects on human hypertrophic scar fibroblasts (HSF) in Panax notoginseng and to explore their mechanisms. Methods: The Soxhlet extraction method was used to obtain the different extracts from P. notoginseng by solvents with different polarities. MTS method was used to screen the ingredients with antiproliferative activity on HSF and flow cytometry was used to detect their influence on cell cycle. Then, spectrum-activity relationship of the active ingredients was analyzed by HPLC. UPLC-Q/TOF-MS was used to identify the ingredients with obvious activity. The antiproliferative mechanism was predicted by reverse docking. Results: The ethyl acetate extract of P. notoginseng showed higher antiproliferative activity (P < 0.01), significantly increased the proportion of cells in G0/G1 phase (P < 0.01), and reduced the proliferation index (PI) (P < 0.01). The main active components were saponins. The result of confirming experiment showed that ginsenosides Rh1 and Rg1 could inhibit the cell proliferation in a dose-dependent manner. Results of reverse docking indicated the antiproliferative effects might be related to the regulation of some target proteins such as MAP2K, MAPK14, and HRAS, as well as the related pathways. Conclusion: The ethyl acetate extract of P. notoginseng shows the antiproliferative activity on HSF, and the antiproliferative ingredients are saponins. The underlying mechanisms might be related with the regulation of MAPK and focal adhesion signaling pathways.
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Objective To investigate the apoptotic effects of hypertrophic scar fibroblast (HSF) induced by HMME-PDT.Methods Fibroblasts were cultured from nontreated hypertrophic scars,and cells at passages 4-6 were used for the experiments (photosensitizer dose 4 μg/ml,λ630 nm,pow er density 10 mw/cm2,energy fluence 2.5 J/cm2).Morphological and biochemical changes in fibroblasts were assessed by Hoechst 33258 staining and fluorescence microscopy.The rate of apoptotic or necrotic cells was detected by flow cytometry (FCM) through double staining of Annexin V -FITC and popodium iodide (PI),respectively.Results Marked morphological features of cell apoptosis were viewed under the fluorescent microscope through Hoechst 33258 staining.The analysis of FCM indica ted that the apoptotic rate was significantly increased after HMME PDT [(34.82 ± I.42) ％ vs (3.12±0.28) ％,P＜0.05],and apoptotic rate was higher than necrosis rate [(14.65±1.02) ％ vs (34.82±1.42) ％,P＜0.05].Conclusions Low level exposure to 630 nm PDT mediated by HMME appears to induce fibroblast apoptosis.