Effects of direct transduction of neuronal nitric oxide synthase protein into the nucleus tractus solitarius on circulation

背景：孤立束核（NTS）被认为是外周气压感受器传入纤维的主要终止部位。据报道，双侧显微注射一种反义神经元一氧化氮合酶（nNOS）。寡核苷酸进入NTS增加动脉血压（ABP）。在之前的研究中，我们证明了通过微注射含有日本包膜血凝病毒（HVJ-E）载体的β-半乳糖苷酶（β-gal），可以在大鼠体内将蛋白质直接转导到特定的限制性脑区。然而，目前尚不清楚转导蛋白是否实现其功能。在本研究中，我们研究了体内nNOS蛋白直接转导到NTS对循环的影响。方法：雄性Wistar大鼠，体重250 ~ 350 g，用1.45 g/kg, i.p的氨基甲酸乙酯麻醉。连续测定ABP、平均ABP （mABP）和心率（HR）。将携带HVJ-E载体的nNOS双侧微注射至NTS。同样，将β-gal蛋白与HVJ-E载体一起微注射作为对照。实验结束后，取大鼠延髓，行免疫组化染色。结果：nNOS与HVJ-E载体在NTS中微注射后1h ~ 3h， mABP较对照组显著降低。(1h: nNOS， Oh值的94%，n=6； 3 -gal值的106%，n=9; p<0.05。2h: nNOS, 89%；/3 -gal, 106%, n=9；p < 0.01。3h: nNOS, 86%；3 -gal, 106%, n=9；p < 0.01)。结果：nNOS与HVJ-E载体在NTS中微注射后1h ~ 3h， mABP较对照组显著降低。(1h: nNOS， Oh值的94%，n=6； 3 -gal值的106%，n=9; p<0.05。2h: nNOS, 89%；/3 -gal, 106%, n=9；p < 0.01。3h: nNOS, 86%；3 -gal, 106%, n=9；p < 0.01)。结果：nNOS与HVJ-E载体在NTS中微注射，在h - 3h时，mABP较对照组显著降低。(1h:nNOS， Oh值的94%，n=6； 3 -gal值的106%，n=9; p<0.05。2h:nNOS, 89%；/3 -gal, 106%, n=9；p < 0.01。3h:nNOS, 86%；3 -gal, 106%, n=9；p < 0.01)。结论：我们在这里证明了转导的蛋白实现了它的功能。我们认为，我们在这里提出的靶向递送系统类型可能在中枢神经系统治疗蛋白的管理中具有广泛的应用。少

BACKGROUND: The nucleus tractus solitarii (NTS) is known to be the principal termination site of the primary afferent fibers from peripheral barorecepters. It has been reported that bilateral microinjection of an antisense neuronal nitric oxide synthase (nNOS). oligonucleotide into the NTS increases in arterial blood pressure (ABP). In the previous study, we demonstrated that direct in vivo protein transduction into a specific restricted brain area is possible in rats by microinjecting β-galactosidase (β-gal) with hemagglutinnating virus of Japan envelope (HVJ-E) vector. However, it is not known whether the transducted protein fulfills its function. In the present study, we examined the effect of direct in vivo nNOS protein transduction into the NTS on circulation.METHODS: Male Wistar rats, weighing 250-350 g, were anesthetized with urethane (1.45 g/kg, i.p.). The ABP, mean ABP (mABP), and heart rate (HR) was measured continuously. The nNOS with HVJ-E vector was microinjected bilateral … More ly into the NTS. As a control β-gal protein with HVJ-E vector was microinjected similarly. After the experiment, the medulla oblongata was removed and the immunohistochemical staining was performed.RESULTS: Microinjection of nNOS with HVJ-E vector into the NTS produced a significant decrease in mABP compared with the control at 1h - 3h. (1h: nNOS, 94% of the value at Oh, n=6; 3 -gal, 106%, n=9; p<0.05. 2h: nNOS, 89%; /3 -gal, 106%, n=9; p<0.01. 3h: nNOS, 86%; 3 -gal, 106%, n=9; p<0.01.)RESULTS: Microinjection of nNOS with HVJ-E vector into the NTS produced a significant decrease in mABP compared with the control at 1h - 3h. (1h: nNOS, 94% of the value at Oh, n=6; 3 -gal, 106%, n=9; p<0.05. 2h: nNOS, 89%; /3 -gal, 106%, n=9; p<0.01. 3h: nNOS, 86%; 3 -gal, 106%, n=9; p<0.01.)RESULTS : Microinjection of nNOS with HVJ-E vector into the NTS produced a significant decrease in mABP compared with the control at lh - 3h. (1h:nNOS, 94% of the value at Oh, n=6 ; 3 -gal, 106%, n=9 ; p<0.05. 2h:nNOS, 89% ; /3 -gal, 106%, n=9 ; p<0.01. 3h:nNOS, 86% ; 3 -gal, 106%, n=9 ; p<0.01.)CONCLUSION : We demonstrated here that the transducted protein fulfills its function. We suggested that the type of targeted delivery system we present here may have wide applications in the administration of therapeutic proteins to the central nervous system. Less