细胞外基质对人毛乳头细胞特性的影响  

Effect of extracellular matrix on the dermal papilla cells in vitro

作　　者：洪伟晋 苗勇[1] 樊哲祥 冯传波[1] 杜丽娟[1] 刘秉承[1] 胡志奇[1] 

机构地区：[1]南方医科大学南方医院整形美容外科,广州510515

出　　处：《中华实验外科杂志》2016年第2期342-346,共5页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金（81471900）;广东省自然科学基金（S2012010009454）;高等学校博士学科点专项科研基金（201244331100）;南方医科大学科研启动计划（PY2013N040）;南方医院院长基金（2013C009）

摘　　要：目的观察不同细胞外基质（ECM）对体外培养的人毛乳头（DP）细胞特性的影响。方法将新鲜分离的人DP接种至已预先分别被Ⅳ型胶原蛋白、纤粘连蛋白、基质胶（Matrigel）或透明质酸钠凝胶包被的培养板，显微镜下观察DP细胞的迁出和生长；采用同样方法进行第4代DP细胞的培养，采用噻唑蓝（MTT）法检测各组细胞的增殖活性，实时定量聚合酶链反应（Real—timePCR）和Western blot法检测不同ECM对DP细胞相对特殊标志物表达的影响。结果Ⅳ型胶原蛋白和纤粘连蛋白组中的DP在接种36h即有细胞迁出，Matrigel组则至少需4d，透明质酸钠凝胶组中一直未见细胞迁出。Ⅳ型胶原蛋白和纤粘连蛋白组与对照组类似，细胞均町呈现出聚集性生长趋势；DP细胞在Matrigel包被的培养孔中央呈贴壁生长，在周围则聚集形成球样结构；细胞无法在透明质酸钠凝胶表面贴擘生长。细胞培养5d后，Ⅳ型胶原和纤粘连蛋白组中DP细胞吸光度（A）值（1．36±0．28和1．31±0．18）明显高于对照组（1．21±0．12），差异有统计学意义（P〈0．05）；Matrigel和透明质酸钠凝胶两组中DP细胞A值（1．08±0．21和0．26±0．05）明显低于对照组，差异有统计学意义（P〈0．05）。Real—time PCR结果显示：与对照组比较，Matrigel组中仅神经细胞黏附分子（NCAM）和α-平滑肌动蛋白（α—SMA）基因的表达显著增加，而多能蛋白聚糖（Versican）的表达无变化。透明质酸钠凝胶组中Versican、NCAM和α—SMA基因的表达均显著降低，而Ⅳ型胶原和纤粘连蛋白组中Versican、NCAM和α-SMA蛋白的表达无明显变化。Westernblot结果显示：与对照组比较，各组DP细胞内Versiean、NCAM和α—SMA蛋白的表达差异无统计学意义（P〉0．05）。结论在平面培养条件下，外源性的ECM（Matrigel、1V型胶原、透明质酸钠凝胶和纤粘连蛋白）虽然可改变DP�Objective To investigate the effects of extraccllular matrix （ECM） on the dermal papilla （DP） cells. Methods The human DP obtained with enzyme digestion method were randomly seeded onto type Ⅳ collagen, fibronectin, matrigel or hyaluronic acid （HA） pre -coated culture plates. We observed the migration and morphology of DP cells under the microscope. The fourth passage of DP cells was randomly seeded onto type Ⅳ collagen, fibronectin, matrigel or HA pre - coated culture plates. The proliferative effect of ECM on DP cells was measured using methyl thiazol tetrazolium （MTT） assay. To understand the influence of ECM on the markers of DP cells, real - time quantitative polymerase chain reaction （ Real - time PCR） and Western blotting were conducted. Results Compared with control group （48 h）, cells can migrate fornl DP after 36 h within collagen type Ⅳ and fibronectin groups; however, some of DP cells and none of DP cells can migrated from Matrigel and HA groups, respectively. When DP cells were seeded onto type IV collagen and fibronectin - coated culture plate, the morphology and growth of cells were similar to the control group; DP cells were adherent growth in the central wells, but they were spherical growth in the edge of wells in the group of Matrigel. There are significant difference in the A value of DP cells compared with type Ⅳ collagen （ 1.36 ±0. 28） and fibronectin （ 1.31 ±0. 18） with control （ 1.21 ± 0. 12） after cultured for 5 days （P 〈0. 05）. There are also significant difference in the A value of DP cells compared with Matrigel （ 1.08 ±0. 21 ） and HA （ 1.31 ±0. 18 ） with control （ 1.21 ±0. 12, P 〈 0. 05 ）. Only the genes expression level of neural cell adhesion molecule （NCAM） and α - smooth muscle actin （α - SMA） in group of Matrigel are significantly higher than control group （P 〈 0. 05 ）. On the contrast, the expression of Versican, NCAM and α- SMA in group of HA are all significantly lower than control group �

关 键 词：胶原蛋白 基质胶 毛乳头细胞 纤粘连蛋白 透明质酸 

分 类 号：R575.2[医药卫生—消化系统]