杜仲总提物对大鼠成骨细胞增殖、分化及Ⅰ型胶原蛋白表达的影响

目的观察杜仲总提取物(DZCE)对新生大鼠颅盖骨成骨细胞增殖、分化及分泌Ⅰ型胶原能力的影响。方法体外分离培养大鼠成骨细胞,分为对照组、地塞米松(DEX)10-8mol/L组、杜仲总提取物不同剂量组(10μg/L、1μg/L、0.1μg/L)共5个组,CCK-8(cell counting Kit-8)检测细胞增殖情况。由Pierce BCA蛋白分析试剂盒进行蛋白定量,然后由碱性磷酸酶(ALP)活性测定试剂盒检测成骨细胞的分化情况。用In cell western方法测定成骨细胞中Ⅰ型胶原蛋白(Col1αI)分泌随时间的变化情况。各组干预24 h后,实时荧光定量法检测成骨细胞Col1αI mRNA的表达情况。结果 1杜仲总提取物各剂量均能刺激成骨细胞的细胞增殖(P0.01),同时也都能使成骨细胞的分化能力增强(P0.05)。2药物干预后,Ⅰ型胶原蛋白的分泌随时间的变化情况是第1天的高中剂量,第3天的中低剂量能使成骨细胞的I型胶原蛋白表达提高(P0.05),第5天、第7天、第9天,各组Ⅰ型胶原蛋白的分泌量均增加,但杜仲总提取物各剂量组与空白组差异无统计学意义(P0.05)。3PCR检测发现,杜仲总提取物各剂量组的I型胶原基因的表达增加,而且这种促进I型胶原分泌的能力有量效趋势。结论杜仲总提取物能显著提高成骨细胞增殖、分化和Col1αⅠ的表达。     

Novel type II collagen reporter mice: New tool for assessing collagen 2α1 expression in vivo and in vitro

We report the generation of a new mouse strain harboring a Col2‐pd2EGFP reporter transgene; pd2EGFP has a much shorter half‐life than EGFP, making it a near real‐time reporter for Col2α1 expression in vivo and in vitro. In the post‐natal growth plate, pd2EGFP fluorescence was expressed in almost all proliferative chondrocytes and in some hypertrophic chondrocytes based on localization with type X collagen. In articular cartilage, pd2EGFP fluorescence diminished over time, nicely illustrating the decrease of type II collagen synthesis in articular chondrocytes during growth. Monolayers of FACS‐sorted chondrocytes from P1‐2 mice showed faster loss of pd2EGFP compared to EGFP, reflecting rapid chondrocyte de‐differentiation. High‐density culture of FACS‐pd2EGFP‐ growth plate chondrocytes revealed the typical temporal expression pattern in which type II collagen preceded type X collagen matrix deposition. The Col2‐pd2EGFP reporter mouse will be a valuable tool for studies of growth plate chondrocyte biology. Developmental Dynamics 240:663–673, 2011. © 2011 Wiley‐Liss, Inc.