Cox A16型手足口病BALB/c乳鼠模型的建立

Establishment of a BALB/c sulking mouse model of coxsackievirus A16 infected hand,foot and mouth disease

目的建立3日龄BALB/c乳鼠Cox A16型手足口病动物模型,并进行免疫、病理学研究。方法3日龄BALB/c乳鼠腹腔注射感染Cox A16型毒株,测定其50%致死剂量(LD50),观察其14 d内存活天数、临床症状打分。采用3LD50病毒攻毒,并观察其14 d内感染程度、体重变化、死亡数,计算平均存活天数、死亡率,至感染第6天发病达到高峰时取材,采用CBA法检测血清中MCP-1、MIP-1α、G-CSF含量,采用实时荧光定量RT-PCR法检测后肢肌肉组织、心脏、脑、肠病毒载量,采用HE染色观察后肢肌肉、脑病理病变情况。结果Cox A16毒株感染3日龄BALB/c乳鼠,出现活动减少、后肢麻痹瘫痪、体重减轻、死亡,其病毒毒力为每毫升56 LD50。感染后14 d内,与正常对照组比较,模型组乳鼠生存天数、死亡率、临床感染评分、体重,均具有显著性差异(P<0.01)。感染后6d,血清中MCP-1、G-CSF含量均显著性升高(P<0.01)。后肢肌肉组织、心脏、脑、肠病毒载量较正常组均显著升高(P<0.01),其中后肢肌肉组织病毒载量最高。HE染色病理观察表明,后肢肌肉组织有大面积萎缩、炎症,脑组织神经细胞有不同程度的萎缩。结论成功构建了3日龄BALB/c乳鼠Cox A16型手足口病动物模型,为COX A16型手足口病药物评价及机制研究奠定基础。

Objective To establish a HFMD BALB/c sulking mouse model infected by coxsackievirus A16,and evaluate immunological and pathological characteristics of this model.Methods In this study,3-day-old BALB/c suckling mice were infected with 100μL coxsackievirus A16 via intraperitoneal injection.The survival rate,clinical score and the50%lethal dose(LD50)were monitored daily 14 days after infection.Challenge with the coxsackievirus A16 with a dose of 3 LD50,3-day-old BALB/c suckling mice were monitored daily the survival rate,clinical score and weight within 14 d.6 d after infection,the concentration of MCP-1,MIP-1 and G-CSF in serum was determined by cytometric bead array(CBA)method;viral loads of the hind limb muscle,heart,brain,intestine were determined by the real-time RT-PCR;histological change of muscle and brain were determined.Results After different concentrations of virus infection,3-dayold BALB/c suckling mice appeared inactivity,hind limb weakness,paralysis and even death,and the virulence of coxsackievirus A16 for 3-day-old BALB/c suckling mice was 56 LD50/mL.Within 14 days after infection,compared with the control group,there were significant differences in the survival days,mortality,clinical score,and body weight of suckling mice in the model group(P<0.01).6 d post infection,the concentrations of MCP-1 and G-CSF in serum are significantly increased(P<0.01)compared with the control group.The viral loads of hindlimb muscle,heart,brain and intestine was significantly higher than that of the control group(P<0.01),and the viral loads of hindlimb muscle was the highest.Pathological change showed that there were large area of atrophy and inflammation in the hind limbs muscle and atrophy in the nerve cells of the brain.Conclusions A Cox A16 infected 3-day-old BALB/c suckling mouse model was successfully established,which can serve for medicine evaluation and mechanism.