包载碱性成纤维细胞生长因子纳米粒结合超声微泡爆破技术预防糖尿病心肌病

Preventing diabetic cardiomyopathy in diabetic rats by combined therapy of basic fibroblast growth factor-loaded nanoparticles and ultrasound-targeted microbubble destruction technique

目的研究包载碱性成纤维细胞生长因子（bFGF）纳米粒联合超声微泡靶向爆破（UTMD）技术靶向递送bFGF对于糖尿病心肌病（OCM）的预防作用。方法采用冷冻干燥技术制备bFGF载药纳米粒。75只SD大鼠适应性喂养1周后采用随机数字表法选择15只作为正常对照组，其余60只采用腹腔注射链脲佐菌素（70mg／kg）建立大鼠1型糖尿病动物模型，共分为4组每组15只：糖尿病模型组、bFGF溶液治疗组、bFGF纳米粒治疗组、bFGF纳米粒＋UTMD治疗组。药物干预前以及药物干预12周后均用常规B超检测左心室舒张末内径（LVIDd）、左心室后壁厚度（LVPW）及左室短轴缩短率（LVFS）。处死大鼠后取出心肌组织，通过Masson胶原染色以及CD31免疫组化染色分别测定心肌的胶原分数以及微血管密度。多组均数间比较采用单因素方差分析，采用LSD．t检验进行两两比较。结果bFGF纳米粒形态圆整，包封率高达（84．3±2．8）％且稳定性好。体内动物实验证明，经药物干预12周之后，相比糖尿病组以及bFGF溶液治疗组、bFGF纳米粒治疗组大鼠，bFGF纳米粒结合UTMD技术干预组大鼠心肌LVIDd[分别为（7．37±0．43）比（5．82±0．24）、（6．65±0．28）、（6．73±0．25）mm；t=8．09、4．83、3．51，均P〈0．051、LVFS[分别为（50＋4）比（38±4）、（44±4）、（43±4）mm，t=6．79、3．93、3．54，均P〈0．05]显著升高；而LVPW[分别为（1．59±0．08）比（1．86±0．09）、（1．72±0．11）、（1．73±0．08）mm，t=5．42、3．34、3．12，均P〈0．05]显著下降。另外，心肌Masson胶原染色以及CD31免疫组化染色证实：12周后，bFGF纳米粒结合UTMD技术干预组大鼠心肌胶原分数显著低于糖尿病组及其他各干预组（t=23．50、8．86、3．34，均P〈0．05）；而心肌微血管密度显著高于糖尿病组以及其他治疗组（t=7．30、5．18、3．45，均QP〈0．05）。结论bFGF纳米粒联合UTMD技术能够有效预防糖尿病引起的心肌结构以及功能的病变。

Objective To investigate the preventive effect of basic fibroblast growth factor （bFGF） on diabetic cardiomyopathy （DCM） using nanoparticles combined with ultrasound-targeted microbubble destruction technique （UTMD）. Method The bFGF-loaded nanoparticles （bFGF-NPs） was prepared by lyophilization technique. Among 75 SD rats, 15 were assigned to control group using random number table after adaptive feeding for 1 week. The rest 60 rats deyeloped type 1 diabetes with intraperitoneal injection of streptozotocin （STZ, 70 mg/kg）. The diabetic rats were divided to 4 groups （15 rats per group）： diabetic model, bFGF solution treatment, bFGF nanopartielcs treatment, bFGF nanopartieles ＋ UTMD treatment. Before and twelve weeks after intervention, all rats underwent the transthoracic echocardiography. The left ventricular end-diastolic diameter （LVIDd）, left ventrieular posterior wall （LVPW） and left ventrieular fraction shortening （LVFS） were measured. At last, the rats were sacrificed and myocardial tissue of all rats were stained with masson staining and CD31 immunohistochemistry staining to quantify myocardial collagen fraction （CVF） and myocardial microvascular density （MVD）. One-way ANOVA was used for multiple comparison. LSD-t test were used for comparison between two groups. Results bFGF-NPs showed good morphology and high encapsulation efficiency （84.3%±2.8%） with high stability. The animal experiments demonstrated that after 12-week treatment, LVIDd （（7.37±0.43） vs （5.82±0.24）, （6.65±0.28）, （6.73±0.25） mm; t=8.09, 4.83, 3.51, all P〈0.05） and LVFS （（50±4） vs （38±4）, （44±4）, （43±4） mm, t=6.79, 3.93, 3.54, all P〈0.05） increased, and LVPW （（1.59±0.08） vs （1.86±0.09）, （1.72±0.11）, （1.73 ±0.08） mm, t=5.42, 3.34, 3.12, all P〈0.05） decreased significantly in bFGF-NP ＋ UTMD group, compared with diabetic group, bFGF solution goups and bFGF nanoparticles group. In addition, after 12-week treatment myocardial masson staining and CD31 immunohistochemistry staining showed that CVF （t=23.5, 8.86, 3.34, all P〈0.05） in bFGF-NP ＋ UTMD group was lower than diabetic group and other intervene groups; MVD （t=7.30, 5.18, 3.45, all P〈0.05） was higher than diabetic group and other intervene groups. Conclusion bFGF-NPs combined with UTMD technique can improve myocardial structure and fianction in diabetes model. It might provide a novel technique for DCM prevention.