Department of Urology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA.
Gene Ther. 2009 May;16(5):660-8. doi: 10.1038/gt.2009.5. Epub 2009 Feb 19.
We examined whether replication-defective herpes simplex virus (HSV) vectors encoding the 67 kDa form of the glutamic acid decarboxylase (GAD(67)) gene product, the gamma-aminobutyric acid (GABA) synthesis enzyme, can suppress detrusor overactivity (DO) in rats with spinal cord injury (SCI). One week after spinalization, HSV vectors expressing GAD and green fluorescent protein (GFP) (HSV-GAD) were injected into the bladder wall. Rats with SCI without HSV injection (HSV-untreated) and those injected with lacZ-encoding reporter gene HSV vectors (HSV-LacZ) were used as controls. Three weeks after viral injection, continuous cystometry was performed under awake conditions in all three groups. In the HSV-GAD group, the number and amplitude of non-voiding contractions (NVCs) were significantly decreased (40-45% and 38-40%, respectively) along with an increase in voiding efficiency, compared with HSV-untreated and HSV-LacZ groups, but micturition pressure was not different among the three groups. Intrathecal application of bicuculline partly reversed the decreased number and amplitude of NVCs, and decreased voiding efficiency in the HSV-GAD group. In the HSV-GAD group, GAD(67) mRNA and protein levels were significantly increased in the L6-S1 dorsal root ganglia (DRG) compared with the HSV-LacZ group, while 57% of DRG cells were GFP-positive, and these neurons showed increased GAD(67)-like immunoreactivity compared with the HSV-LacZ group. These results indicate that GAD gene therapy effectively suppresses DO after SCI predominantly through the activation of spinal GABA(A) receptors. Thus, HSV-based GAD gene transfer to bladder afferent pathways may represent a novel approach for treatment of neurogenic DO.
我们研究了复制缺陷型单纯疱疹病毒（HSV）载体能否抑制脊髓损伤（SCI）大鼠逼尿肌过度活动（DO），这些载体编码谷氨酸脱羧酶（GAD）67 kDa 形式的基因产物，即 GABA 合成酶。在 SCI 后 1 周，将表达 GAD 和绿色荧光蛋白（GFP）的 HSV 载体（HSV-GAD）注入膀胱壁。未注射 HSV 的 SCI 大鼠（HSV-未处理）和注射编码报告基因 HSV 载体（HSV-LacZ）的大鼠作为对照。病毒注射后 3 周，三组大鼠在清醒状态下进行连续膀胱测压。与 HSV-未处理组和 HSV-LacZ 组相比，HSV-GAD 组的非排尿收缩（NVC）次数和幅度明显减少（分别减少 40-45%和 38-40%），同时排尿效率增加，但三组间排尿压力无差异。鞘内应用荷包牡丹碱部分逆转了 HSV-GAD 组 NVC 次数和幅度的减少，以及排尿效率的降低。在 HSV-GAD 组，L6-S1 背根神经节（DRG）中 GAD（67）mRNA 和蛋白水平明显高于 HSV-LacZ 组，而 57%的 DRG 细胞为 GFP 阳性，这些神经元的 GAD（67）样免疫反应性较 HSV-LacZ 组增强。这些结果表明，GAD 基因治疗通过激活脊髓 GABA(A)受体有效抑制 SCI 后的 DO。因此，将 HSV 为基础的 GAD 基因转移到膀胱传入途径可能为治疗神经源性 DO 提供一种新方法。
