改良保存液处理红细胞对糖尿病小鼠创面愈合的影响

目的研究改良处理红细胞回输对糖尿病小鼠伤口愈合的影响,并探讨其可能的作用机制。方法建立糖尿病小鼠模型,随机分为献血组和实验组。采集献血组小鼠血液,分别用标准和改良保存液保存,在7、14、21和28 d检测红细胞(RBC)携氧能力及变形能力。在实验组小鼠背部制作伤口愈合模型,并将保存7 d的血液回输给实验组小鼠,再分为标准组及改良组,每组6只,在0、1、4、7和14 d观察小鼠伤口愈合情况,第14天处死小鼠,取创面皮肤组织进行HE、Masson染色,免疫组化方法对伤口组织的纤维化程度进行分析,Western blot检测伤口组织PI3K/AKT、MEK/ERK及其磷酸化蛋白水平。结果与标准组比较,改良组游离血红蛋白(fHb)含量降低,而P50、pH、2,3-二磷酸甘油酸(2,3-DPG)和红细胞变形性(RCD)升高(P<0.05);改良组创面面积显著减小、皮肤组织和血管面积增加、胶原纤维的表达和纤维化程度升高(P<0.05)。小鼠伤口组织中HIF-1α、VEGF和EGF浓度水平增加(P<0.05)。PI3K/AKT、MEK/ERK表达增加,PI3K/AKT和MEK/ERK磷酸化显著增强(P<0.05)。结论改良保存液处理的红细胞可能通过激活HIF-1α信号通路促进糖尿病小鼠的伤口愈合。

Influences on wound healing in diabetic mice by blood transfusion treated with improved blood preservation solution

Objective To study whether blood transfusion with improved blood preservation solution can improve wound healing in diabetic mice,and to investigate its possible mechanism.Methods Kunming mice were used to establish diabetic mice model.They were randomly divided into blood donation group and the experimental group.The blood was placed in sterile centrifugal tube containing anticoagulant and then preserved for 7,14,21 and 28 days with improved and standard blood preservation solutions,respectively.The oxygen carrying capacity and deformability of erythrocytes in each preservation period were tested,then the improved and standard autologous blood were retransfused with 7 days of preservation.The experimental group was randomly divided into the standard group and the improved group with 6 mice in each group.The wound healing of two groups of mice was observed on 0,1,4,7 and 14 days,and the mice were killed on 14 days.The degree of fibrosis was analyzed by means of HE,Masson staining and immunohistochemistry methods.Phosphorylation of PI3K,AKT,MEK and ERK were analyzed byWestern blot analysis.Results Compared with the standard group,the improvement group exhibited decreased fHb content,and increased P50 oxygen pressure,blood pH,2,3-Diphosphoglycerate(2,3-DPG)and red blood cell deformability(RCD)(P<0.05).HE staining results revealed that compared with the standard group,the area of granular tissue on the skin and the vascular area were increased on the 14th day after operation in the improvement group.Masson staining revealed that,the expression of collagen fibers and the degree of fibrosis in the mice skin of the improvement group were elevated at 14 days after the operation,when compared with the standard group.Immunohistochemistry results indicated that in comparison to standard group,levels of HIF-1α,VEGF,and EGF in skin tissues of the improvement group increased(P<0.05).The protein levels of PI3K,AKT,MEK,ERK and its phosphorylation increased significantly in the improvement group(P<0.05).Conclusions Red blood cell treated with improved blood preservation solution may promote wound healing in diabetic mice by activating HIF-1αpathway.