间充质干细胞注射液关节腔及静脉单次注射给药急性毒性及局部刺激性

Local irritation and acute toxicities of mesenchymal stem cell injection following single intra-articular injection and intravenous injection

目的 选取兔和大鼠,通过关节腔及静脉单次注射评价间充质干细胞(MSC)注射液的急性毒性及刺激性,以期为其临床安全用药提供参考依据。方法 (1)急性毒性试验:SD大鼠,关节腔注射和静脉注射MSC注射液(给药剂量分别为5.0×107和1.0×108kg^(-1)),单次给药,同时设正常对照组,每组10只,雌雄各半,给药后观察30 d。试验期间每日进行1次临床观察,给药前(d 0)及给药后d 1(给药当天),3,7,14,21和28测体重,恢复期结束次日(d 31)解剖,对体表及皮下、头部、颈部、胸腔及胸腔脏器、盆腔及盆腔脏器、腹腔及腹腔脏器及给药部位等采用肉眼逐项进行详细大体观察。(2)刺激性试验:4只日本大耳白家兔,雌雄各半,采用同体左右侧对比法分为正常对照侧和给药侧,用关节腔注射及静脉注射(给药剂量分别为4.0×106和6.0×106kg^(-1))单次给药,给药后观察14 d。试验期间每日观察全身及注射部位,d 3和d 14进行解剖检查及组织病理学检查。结果 (1)急性毒性:与正常对照组相比,静脉注射给药组SD大鼠临床观察和体重均未见明显异常;关节腔注射给药组出现给药侧后肢蜷缩,呈跛行,并给药部位逐渐出现膝关节肿胀,恢复期后可陆续恢复;关节腔注射给药组雄性大鼠给药后d 1和d 3体重显著降低(P<0.05),其余各检查时间点平均体重无明显差异,雌性大鼠体重无差异。试验期间无动物死亡;静脉注射及关节腔注射组动物解剖检查均无异常。(2)刺激性试验:与正常对照侧比较,关节腔注射给药家兔膝关节周围组织可见明显的充血、红肿,经恢复期可陆续恢复;3 d解剖检查可见关节腔内有淡粉色浓稠积液溢出,组织病理学检查可见滑膜中度病变,对局部组织有一定的刺激性;14 d,病变程度明显减轻,但未完全恢复正常。静脉注射给药组家兔临床观察及组织病理学检查均未见明显异常。结论 大鼠单次关节腔及静脉注射给予MSC注射液未出现明显毒性。兔单次关节腔注射给予MSC注射液对注射部位关节腔局部组织有一定刺激性,停药后有明显的恢复趋势。兔单次静脉注射给予MSC注射液对注射部位静脉血管无刺激性。

OBJECTIVE To evaluate the acute toxicity(in rats)and irritation(in rabbits)of mesenchymal stem cell injection by single intra-articular injection and intravenous injection so as to provide reference for clinical safety use.METHODS①Acute toxicity test:SD rats were administered by intraarticular injection and intravenous injection(the doses were 5.0×107 kg-1 and 1.0×108 kg-1,respectively).SD rats were randomly divided into 4 groups including two blank control groups with different routes of administration(n=10 in each group).SD rats were allowed to recover for 30 d after administration of a single dose.Clinical observation was conducted once a day during the trial.On the day before administration(d 0)and 1,3,7,14,21 and 28 d after administration,the body mass was recorded,anatomy was performed the next day after recovery(d 31),and detailed gross observation was carried out of the body surface and subcutaneous tissues,head,neck,chest and thoracic organs,pelvic and pelvic organs,abdominal cavity and abdominal organs,and drug administration sites with naked eyes.②Irritation test:using isomorphism(left-right)contrast method,four Japanese white rabbits(half male and half female)were divided into two groups(the control group and the administration group).Both were administered by intra-articular injection and intravenous injection(the doses were 4.0×106 kg-1 and 6.0×106 kg-1,respectively).Rabbits were allowed to recover for 14 d after administration of a single dose.The whole body and the injection sites were observed once a day during the test.Anatomic and histopathological examinations were performed at the end of 72 h and 14 d restoration.RESULTS①Acute toxicity:No obvious abnormalities were observed during body mass examination and clinical observation of SD rats with intravenous injection.In the intra-articular injection group,the hind limbs of the administration side were cured up and limped,and knee joint swelling at the administration site gradually appeared,which recovered in succession during recovery.The body mass of male rats in the intra-articular injection group decreased significantly on the 1st and 3rd day after administration(P<0.05),but there was no significant difference in the average body mass at other points.No abnormality was detected in the body mass of female animals.No animals died during the entire period.The anatomical examination was normal.②Irritation:Obvious hyperemia,redness and swelling could be seen in the tissues around the knee joints of the animals in the intra-articular injection group,which recovered in succession during recovery.72 h after administration,anatomical examination found that there was a pale pink thick effusion in the joint cavity of the intra-articular injection group.And histopathological examination showed moderate synovial lesions,with irritation to local tissues.14 d after administration,the lesions were significantly reduced,but did not completely return to normal.Clinical observation and histopathological examination of Japanese white rabbits administered intravenously showed no obvious abnormality.CONCLUSION No significant toxicity is observed after single intra-articular injection and intravenous injection with mesenchymal stem cell injection in rats.Irritation to local tissues is observed after single intra-articular injection with mesenchymal stem cell injection(4.0×106 kg-1)in rabbits,but might recover gradually after drug withdrawal.No irritation to vein vessels at injection sites is observed after single intravenous injection with mesenchymal stem cell injection(6.0×106 kg-1)in rabbits.