可分泌表达神经保护肽的重组慢病毒对闭合性脑损伤小鼠的神经保护作用

目的：通过滴鼻给药途径，给予闭合性脑损伤小鼠可分泌表达神经保护肽NAP的重组慢病毒，观察该重组病毒经鼻-脑通路对中枢神经系统的保护作用，为携带可分泌表达NAP的重组慢病毒治疗神经系统退行性疾病提供理论依据。 方法：选用8~12周成年雄性昆明小鼠54只，随机分为4组：空白对照组10只（Control组）、重锤加害组20只（CHI组）、重组慢病毒rLent/NT4-NAP保护组20只（rLent组）和重组慢病毒rLent/GFP组4只（GFP组）。观察各组小鼠的死亡率、神经功能损伤（NSS）评分、脑水肿含量和病理改变情况。应用共聚焦显微镜观测GFP组小鼠鼻黏膜、嗅神经和脑组织内绿色荧光表达情况。 结果：rLent组小鼠死亡率（15％）明显低于CHI组小鼠（55％）；rLent组小鼠NSS评分在创伤后1、3、5和7 d均显著低于CHI组（P<0.01）； rLent组小鼠创伤后24 h的脑水肿含量明显低于CHI组（P<0.01）；HE染色显示rLent组小鼠病理改变明显轻于CHI组；GFP组仅在鼻黏膜处可见呈条索样的绿色荧光，而在嗅神经及大脑则未发现绿色荧光。 结论：分泌表达NAP的重组慢病毒可感染鼻黏膜细胞，分泌表达的短肽NAP能够改善闭合性脑损伤小鼠的神经功能；未加装分泌表达元件的重组病毒rLent/GFP仅能感染鼻黏膜细胞，不能通过血脑屏障进入中枢神经系统。

Neuroprotective effect of lentiviral vector secreting neuroprotective peptide on closed head injury in mice

Objective To observe the neuoprotective effect of lentiviral vector expressing and secreting neuroprotective peptide(NAP)on closed head injury in mice via nose-brain pathway and provide theoretical basis for treatment of neurodegeneration diseases by lentiviral vector secreting NAP.Methods 54 adult male Kunming mice were divided into four groups:control group(n=10),closed head jnjury group(CHI,n=20),rLent/NT4-NAP group(rLent,n=20),and rLent/GFP group(GFP,n=4).The mortality,NSS and brain edema in various groups were observed.The pathological changes in brain injury region of various groups stained by HE were observed.The expressions of GFP in nasal epithelium olfactory nerve and brain tissue in GFP group were detected under laser confocal microscope.Results The mortality of mice in rLent group(15%)was significantly lower than that in CHI group(55%).The NSS in rLent group were lower than those in CHI groups at 1,3,5 and 7 d after brain injury(P<0.01).The percentage of H2O in rLent group was lower than that in CHI group at 24 h after brain injury(P<0.01).HE staining result showed that the pathological changes in rLent group were fewer than those in CHI group.There was significant GFP in the nasal epithelium,but no fluorescence in olfactory nerve and brain tissue.Conclusion The recombinant rLent/NT4-NAP could efficietnly infect nasal epithelium in vivo.NAP provides significant neuroprotective effect from the complex array of injuries elicited by head trauma.The recombinant rLent/GFP could only infect nasal epithelium,but not go through blood-brain barrier(BBB)to central nervous system(CNS).