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目的研究Ca2+诱导分化的人角质形成细胞(HKC)上清液对人成纤维细胞(HFB)增殖及Ⅰ,Ⅲ型胶原蛋白合成的影响。方法分别用0.09,1.0和1.5mmol/L Ca2+的K-SFM培养液培养HKC 3d,分化标记物K10染色鉴定HKC的分化程度;采用HKC上清液与DMEM 1∶1比例的混合液培养HFB 2d,以DMEM培养的HFB为对照组;MTT法测定HFB增殖;ELISA法测定HFB上清液中Ⅰ,Ⅲ型胶原含量。结果 1.0或1.5mmol/L Ca2+的K-SFM培养HKC 1d时,HKC出现复层生长,3d时K10的表达均较0.09mmol/L Ca2+组的增强(P<0.05);相较于DMEM培养HFB组,0.09mmol/L Ca2+HKC上清液与DMEM混合液促进HFB增殖及Ⅰ,Ⅲ型胶原的合成,1.0和1.5mmol/L Ca2+HKC上清液分别与DMEM混合均抑制HFB增殖及Ⅰ,Ⅲ型胶原蛋白的合成(P<0.05)。结论不同分化程度的HKC上清液可以调控HFB的增殖及Ⅰ,Ⅲ型胶原蛋白合成。
Objective To investigate the effects of supernatant of human keratinocytes (HKC) induced by Ca2 + on the proliferation of human fibroblasts (HFB) and the synthesis of collagen type Ⅰ and Ⅲ. Methods HKC 3d was cultured with 0.09, 1.0 and 1.5 mmol / L Ca2 + in K-SFM medium, and the differentiation of HKC was identified by K10 differentiation marker. The culture of HKB was induced by a 1: 1 mixture of HKC supernatant and DMEM HFB cultured in DMEM was used as a control group. The proliferation of HFB was measured by MTT assay. The contents of collagen type Ⅰ and Ⅲ in HFB supernatant were determined by ELISA. RESULTS: KCs cultured in HKC at 1.0 or 1.5 mmol / L Ca2 + for 1 d showed that HKC appeared to be stratum proliferatum, and the expression of K10 was enhanced at 3 d compared with 0.09 mmol / L Ca2 + group (P <0.05) Group, 0.09 mmol / L Ca2 + HKC supernatant and DMEM mixture promoted the proliferation of HFB and the synthesis of type Ⅰ and type Ⅲ collagen. The 1.0 and 1.5 mmol / L Ca2 + HKC supernatant mixed with DMEM respectively inhibited the proliferation of HFB and Ⅰ , Collagen type Ⅲ synthesis (P <0.05). Conclusions HKC supernatants with different degrees of differentiation can regulate HFB proliferation and type Ⅰ and type Ⅲ collagen synthesis.