同种异体富血小板血浆对创面胶原合成的作用及其机制研究

Effects and mechanism of allogeneic platelet rich plasma on collagen synthesis in wound healing

目的 探讨同种异体来源的富血小板血浆（PRP）促进创面胶原合成的作用及机制.方法 采用7周龄雄性SD清洁级大鼠50只,10只大鼠用来制备富血小板血浆,其余40只大鼠随机分为PRP组和对照组,每组各20只,PRP组大鼠创面涂抹0.1 ml同种异体的PRP,对照组涂抹同等剂量的生理盐水,观察创面再生愈合情况,采用Masson染色分析细胞及组织形态学变化,免疫印迹法检测Ⅰ型和Ⅲ型胶原的蛋白表达情况;采用实时定量PCR检测Ⅰ型和Ⅲ型胶原组织mRNA的表达.计量资料的比较采用t检验,组间比较采用单因素方差分析.结果 创面形成后第3、6、10、15天,PRP组创面愈合率明显高于对照组（30.33±3.35比18.35±2.04,55.5l±2.74比36.83±2.34,79.64±1.40比56.92± 1.44,86.88±2.12比65.80± 1.76）,差异有统计学意义（t=13.66 ～ 50.48,P值均＜0.05）;Masson染色显示PRP组的创面胶原形成快、纤维粗壮、排列致密;与对照组相比,PRP组细胞Ⅰ型胶原蛋白（1.92±0.09比1.18±0.11）和Ⅲ型胶原蛋白（1.16±0.05比0.74±0.11）表达明显上调,差异有统计学意义（t=22.99,P＜0.01;t=17.62,P＜0.05）;与对照组相比,PRP组在创面形成后第3、6、10、15天的Ⅰ型胶原蛋白（5.17±0.11比1.79±0.18,6.97±0.09比1.96±0.08,6.00±0.26比2.10±0.05,4.95±0.11比3.58±0.09）和Ⅲ型胶原蛋白（2.35 ±0.08比1.44±0.05,3.08±0.05比1.84±0.06,3.48±0.07比2.36±0.09,4.42±0.07比2.77±0.10）的mRNA均表达上调,差异有统计学意义（t=43.37～ 188.37,P值均＜0.01）.结论 同种异体富血小板血浆可通过促进Ⅰ型和Ⅲ型胶原蛋白合成加速创面愈合.

Objective To investigate the effects and mechanism of allogeneic platelet rich plasma （PRP） on collagen in wound surface at different time.Methods A total of 50 clean 7-week rats were selected for this study,including 10 rats for platelet-rich blood plasma preparation,20 rats for PRP group and 20 rats for control group,0.1 ml allogenic PRP and 0.1 ml saline were smeared respectively on wound surfaces of PRP and control group,wound regeneration and healing were examined.Cellular and histological morphology alteration was observed via Masson staining,type Ⅰ and type Ⅲ collagen protein and mRNA expression level were detected by Western blot and real-time PCR.T test was applied for comparison between two samples and one-way ANOVA was utilized for comparison between two groups.Results The wound healing rate of PRP group was higher than that of control group on 3rd,6th,10th and 15th day （30.33±3.35 vs.18.35 ± 2.04,55.51 ± 2.74 vs.36.83 ± 2.34,79.64 ± 1.40 vs.56.92 ± 1.44,86.88 ±2.12 vs.65.80±1.76） after wound surface formation,there were statistic differences （t=13.66-50.48,all P〈0.05）.The wound collagen of PRP group form faster and coarser,and the fibers arrayed more densely in Masson staining.The protein expression of type Ⅰ collagen （1.92±0.09 vs.1.18± 0.11） and type Ⅲ collagen（1.16±0.05 vs.0.74±0.11） of PRP group were higher than that of control group （t =22.99,P〈 0.01;t =17.62,P 〈 0.05）;the mRNA expression of type Ⅰ collagen （5.17± 0.11 vs.1.79 ± 0.18,6.97 ± 0.09 vs.1.96±0.08,6.00±0.26 vs.2.10±0.05,4.95±0.11 vs.3.58±0.09） and type Ⅲ collagen（2.35± 0.08 vs.1.44±0.05,3.08±0.05 vs.1.84±0.06,3.48±0.07 vs.2.36±0.09,4.42±0.07 vs.2.77±0.10） were higher than that of control group on 3rd,6th,10th and 15th day after wound surface formation,there were significant differences （t =43.37-188.37,all P〈0.05）.Conclusion The allogeneic platelet rich plasma may promote fibroblasts secreted collagen by activated and releasing all kinds of growth factors,especially type Ⅰ and type Ⅲ collagen to accelerate the wound healing.