透過脊神經注射達成非病毒載體之大鼠背根神經節基因轉殖

Abstract

對背根神經節進行基因轉殖是可以被應用於治療周邊神經病變的一個有力的治療方式，然而過去的進行方式如背根神經節內注射或脊髓穿刺都存在有許多缺點，包括有容易造成神邊神經損傷或轉殖效益低落等缺點。所以在我們的研究中採用聚乙烯亞胺(PEI)混合可表現綠色螢光(GFP)之DNA植體的方式並透過脊神經注射來達成背根神經節之基因轉殖。在轉殖後一週，我們可發現在脊神經注射組有 82.8% ± 1.70% 的神經細胞試轉殖成功比對背根神經節內注射組別也有(81.3% ± 5.1%, p = 0.82)相較於脊髓穿刺組別有非常高之基因轉殖成功率。此外GFP 在背根神經節內的大神經細胞與小神經細胞的轉殖效益也是相同的。另外在安全性測試中，我們也證實透過脊神經注射不會引起神經損傷、發炎、行為改變等變化。我們的結果證明透過脊神注射聚乙烯亞胺(PEI)混合DNA植體來進行背根神經節的基因轉殖是一個安全且有效的方法。

Delivering gene constructs into the dorsal root ganglia (DRG) is a powerful but challenging therapeutic strategy for sensory disorders affecting the DRG and their peripheral processes. The current delivery methods of direct intra-DRG injection and intrathecal injection have several disadvantages, including potential injury to DRG neurons and low transfection efficiency, respectively. This study aimed to develop a spinal nerve injection strategy to deliver polyethylenimine mixed with plasmid (PEI/DNA polyplexes) containing green fluorescent protein (GFP). Using this spinal nerve injection approach, PEI/DNA polyplexes were delivered to DRG neurons without nerve injury. Within one week of the delivery, GFP expression was detected in 82.8% ± 1.70% of DRG neurons, comparable to the levels obtained by intra-DRG injection (81.3% ± 5.1%, p = 0.82) but much higher than those obtained by intrathecal injection. The degree of GFP expression by neurofilament(+) and peripherin(+) DRG neurons was similar. The safety of this approach was documented by the absence of injury marker expression, including activation transcription factor 3 and ionized calcium binding adaptor molecule 1 for neurons and glia, respectively, as well as the absence of behavioral changes. These results demonstrated the efficacy and safety of delivering PEI/DNA polyplexes to DRG neurons via spinal nerve injection.