以瑞特氏症模式小鼠研究運動障礙之療癒

Abstract

瑞特氏症 (Rett Syndrome, RTT) 是一種主要發生在女性幼童的神經發育疾病，罹 患率大約為每10,000~15,000個新生兒會有一例。幾乎所有RTT患者都會表現出手部扭 絞的刻板行為並無法行走，這些運動障礙通常會隨著年齡惡化，而在疾病末期呈現類似 巴金森氏症的特徵。這些RTT運動障礙之成因目前仍屬未知。遺傳上導致RTT的主要原 因為X染色體上製造第二型甲基CpG結合蛋白(methyl-CpG binding protein 2, MeCP2)之基因發生突變。過去的研究顯示，在MeCP2缺失的小鼠可重現許多類似RTT 的症狀，包括活動力降低、四肢交握的刻板行為、以及運動協調及運動學習的障礙，但 並無報導指出MeCP2的缺失如何導致RTT的運動障礙。紋狀體(Striatum)為基底核的主 要輸入神經核，負責運動控制之外，也與某些會表現出心理運動障礙的神經發育疾病如 自閉症的致病機轉有關。我們先前的研究發現，MeCP2的存在對於紋狀體正常調控活 動力與運動學習不可或缺，顯示RTT的運動障礙至少有一部分是因為紋狀體的MeCP2 缺失所造成(國科會計畫編號NSC99-2320-B-004-001-MY2)。值得注意的是，雖然最 近的研究顯示MeCP2缺失小鼠的某些症狀可被回復，但尚未有報導指出是否將MeCP2 重新表現在紋狀體中即足以反轉或回復MeCP2缺失小鼠上RTT相關的運動障礙。本計畫 預定利用兩組實驗來檢驗這個可能性。首先我們將利用兩種品系(floxed-STOP-Mecp2 及Dlx5/6-Cre)的基因轉殖鼠互交，繁殖出特定在紋狀體恢復MeCP2表現的「條件回復 小鼠」(conditional rescued mice, cRes)，並測量其運動行為及分析紋狀體之表型特徵 (Aim 1)。為了更進一步得知紋狀體重新表現MeCP2的回復效果是否有特定區域及特定 時間點的差異，我們將利用線相關病毒(adeno-associated virus, AAV)載體，在特定發 育階段將MeCP2基因在紋狀體中進行區域專一性的重新表現，並測量其所導致之運動 功能與紋狀體特徵之回復(Aim 2)。本研究將不只有助於找出RTT模式小鼠中治療運動障 礙之最佳腦區與發育時間窗，也對紋狀體神經迴路對於心理運動功能之調控有更進一步 的了解。

Rett Syndrome (RTT) is a neurodevelopmental disorder primarily affect female with an estimated prevalence rate of one in 10,000~15,000 births. Almost all RTT patients develop stereotypical hand wringing and walking disabilities which usually exacerbate with age and resemble Parkinson’s features late in life. The etiology of psychomotor deficits in RTT remains largely unknown. Mutations of the X-linked gene encoding methyl-CpG binding protein 2 (MeCP2) have been identified as the major cause of RTT. Mice with deficient MeCP2 recapitulate many RTT-like symptoms including hypoactivity, stereotypic clasping, and deficits in motor coordination and motor skill learning. However, no report demonstrates how loss of MeCP2 leads to motor deficits in RTT. The striatum, a major input nucleus of the basal ganglia, has been implicated in motor control and various developmental disorders such as autism that involves psychomotor deficits. Our previous findings supported that MeCP2 is indispensible for the striatum to control normal motor activities and motor learning, suggesting the motor dysfunction in RTT is at least partly a consequence of MeCP2 deficiency in the striatum (supported by NSC-99-2320-B-004-001 -MY2). Notably, although recent studies reported that RTT-like phenotypes in Mecp2-null mice are reversible, it has yet to be determined whether restoration of MeCP2 expression in the striatum is sufficient to reverse psychomotor deficits associated with RTT. In this proposal, we plan to test the possibility that the motor dysfunction in Mecp2 deficient mice can be rescued by re-introducing MeCP2 in the striatum. We will begin the experiments by crossing floxed-STOP-Mecp2 mice to Dlx5/6-Cre mice to generate the conditional rescued (cRes) mice and measure their motor behaviors and the striatal phenotypes (Aim 1). To gain additional spatial and temporal control and to determine when and where re-expression of MeCP2 is sufficient to rescue motor deficits in RTT mice, we will then employ an adeno-associated virus (AAV)-Cre mediated approach to restore MeCP2 expression in subregions of the striatum at different developmental time points, and test the recovery of motor function and striatal organization (Aim 2). Our study will not only help to pinpoint the brain region and the developmental time window to ameliorate motor deficits in mouse models of RTT, but also gain insight into the role of the striatal circuits in psychomotor control.