以水凝胶微球装载罗哌卡因改善切口痛的有效性及安全性

Efficacy and safety of hydrogel microspheres-loaded ropivacaine for improving incision pain

目的:评价以微流控法制备的明胶水凝胶微球用于装载罗哌卡因治疗切口痛的有效性及安全性。方法:通过微流控技术制备水凝胶微球,利用共轴剪切作用获取甲基丙烯酰化明胶(GelMA)液滴,待循环冷冻后经紫外光固化得到水凝胶微球。通过共孵育将罗哌卡因装载入水凝胶微球,用光学显微镜及扫描电子显微镜观察微球表面特征,以透析法检测其体外释放速率。将L-929和SH-SY5Y细胞分别分为0.9%氯化钠溶液对照组(Ctr组)、水凝胶微球组(GelMA组)、罗哌卡因组(R组)、水凝胶微球罗哌卡因组(Gel-R组),用CCK8法和Calcein/PI活/死染色法分别检测细胞的增殖和存活情况。将SD大鼠随机分为空白对照2组(Ctr 2组)、微球2组(GelMA 2组)、罗哌卡因2组(R 2组)、水凝胶微球罗哌卡因2组(Gel-R 2组),行为学采用切口痛模型,以vonfrey法测定大鼠机械痛阈值。取大鼠足底切口处及心、肝、肾组织,以HE染色法检测组织相容性。结果:以微流控技术制备的水凝胶微球有形状圆润、粒径分布集中的特点,且具有疏松多孔的结构,具备装载并缓慢释放罗哌卡因的功能。细胞实验中,CCK8法结果提示Gel-R 2组毒性低于R组(P<0.05)。活/死染色结果显示,在培养的5 d时间里,R组较其他组出现更多死亡细胞。行为学结果提示,Gel-R组在整个切口愈合过程中均表现出镇痛作用。HE染色法显示,各切口处组织没有明显组织损伤及炎症反应,脏器未见明显病理表现。结论:以微流控法制备的明胶水凝胶微球具有形貌整齐、结构完整的属性,并在动物模型和细胞实验中验证了其可以延长罗哌卡因作用时间,且具有良好的生物相容性。

Objective To evaluate the efficacy and safety of gelatin hydrogel microspheres prepared by microfluidic technique for loading ropivacaine in the treatment of incision pain.Methods The hydrogel microspheres were prepared by microfluidic technique,the methacryloyl gelatin(GelMA)droplets were obtained by coaxial shearing,and then the hydrogel microspheres were obtained by UV curing after cyclic freezing.Ropivacaine was loaded into hydrogel microspheres through coincubation,the surface characteristics of the microspheres were observed with optical microscope and scanning electron microscope,and the release rate in vitro was measured by dialysis.L-929 and SH-SY5Y cells were divided into 0.9%sodium chloride solution control group(Ctr group),hydrogel microsphere group(GelMA group),ropivacaine group(R group),hydrogel microsphere ropivacaine group(Gel-R group),respectively.Cell proliferation and survival were detected by CCK8 method and Calcein/PI live/dead staining method.SD rats were randomly divided into blank control group 2(Ctr group 2),microsphere group 2(GelMA group 2),ropivacaine group 2(R group 2),hydrogel microsphere ropivacaine group 2(Gel-R group 2).The mechanical pain threshold in behavior was determined with vonfrey method in the incision pain rat models.The incisional tissues of the rat's foot,and hearts,livers,kidneys were harvested to determine histocompatibility by HE staining.Results The hydrogel microspheres prepared by microfluidic technique had the characteristics of rounded shape,concentrated particle size distribution,and loose and porous structure,which possessed the function of loading and slowly releasing ropivacaine.In the cell experiment,the results of CCK8 method showed that the toxicity of the Gel-R group was lower than that of the group R(P<0.05).The live/dead staining results showed that during the 5-day culture period,compared with other groups,more dead cells appeared in the R group.Behavior results suggested that the Gel-R group 2 exhibited analgesic effects throughout the entire incision healing process.The HE staining indicated that there was no significant tissue damage or inflammatory response in the tissues at each incision,and no obvious pathological manifestations were observed in the organs.Conclusions The gelatin hydrogel microspheres prepared by microfluidic technique have the properties of regular morphology and complete structure,and they can prolong the action time of ropivacaine and have good biocompatibility in animal models and cell experiments.