Dong Kwang Seo1, Jeong Hoon Kim1, Joongkee Min1, Hyung Ho Yoon1, Eun-Sil Shin1, Seong Who Kim2, Sang Ryong Jeon3. 1. Department of Neurological Surgery, Asan Medical Center, University of Ulsan College of Medicine, 88, Olympic-ro 43-gil, Songpa-gu, Seoul, 05505, South Korea. 2. Department of Biochemistry and Molecular Biology, University of Ulsan College of Medicine, Seoul, South Korea. 3. Department of Neurological Surgery, Asan Medical Center, University of Ulsan College of Medicine, 88, Olympic-ro 43-gil, Songpa-gu, Seoul, 05505, South Korea. srjeon@amc.seoul.kr.
Abstract
BACKGROUND: While pure mesenchymal stem cell (MSC) treatment for spinal cord injury (SCI) is known to be safe, its efficacy is insufficient. Therefore, gene-modified stem cells are being developed to enhance the effect of pure MSCs. We investigated the effect of stem cell therapy through the transfection of a Wnt3a-producing gene that stimulates axonal regeneration. METHOD: MSCs obtained from the human umbilical cord blood (hMSCs) were multiplied, cultivated, and transfected with the pLenti-Wnt3a-GFP viral vector to produce Wnt3a-secreting hMSCs. A total of 50 rats were injured with an Infinite Horizon impactor at the level of the T7-8 vertebrae. Rats were divided into five groups according to the transplanted material: (1) phosphate-buffered saline injection group (sham group, n = 10); (Pertz et al. Proc Natl Acad Sci USA 105:1931-1936, 39) Wnt3a protein injection group (Wnt3a protein group, n = 10); (3) hMSC transplantation group (MSC group, n = 10); (4) hMSCs transfected with the pLenti vector transplantation group (pLenti-MSC group, n = 10); (5) hMSCs transfected with the pLenti+Wnt3a vector transplantation group (Wnt3a-MSC group, n = 10). Behavioral tests were performed daily for the first 3 days after injury and then weekly for 8 weeks. The injured spinal cords were extracted, and axonal regeneration markers including choline acetyltransferase (ChAT), growth-associated protein 43 (GAP43), and microtubule-associated protein 2 (MAP2) were investigated by immunofluorescence, RT-PCR, and western blotting. RESULTS: Seven weeks after the transplantation (8 weeks after SCI), rats in the Wnt3a-MSC group achieved significantly higher average scores in the motor behavior tests than those in the other groups (p < 0.05). Immunofluorescent stains showed greater immunoreactivity of ChAT, GAP43, and MAP2 in the Wnt3a-MSC group than in the other groups. RT-PCR and western blots revealed greater expression of these proteins in the Wnt3a-MSC group than in the other groups (p < 0.05). CONCLUSIONS: Wnt3a-secreting hMSC transplantation considerably improved neurological recovery and axonal regeneration in a rat SCI model.
BACKGROUND: While pure mesenchymal stem cell (MSC) treatment for spinal cord injury (SCI) is known to be safe, its efficacy is insufficient. Therefore, gene-modified stem cells are being developed to enhance the effect of pure MSCs. We investigated the effect of stem cell therapy through the transfection of a Wnt3a-producing gene that stimulates axonal regeneration. METHOD: MSCs obtained from the human umbilical cord blood (hMSCs) were multiplied, cultivated, and transfected with the pLenti-Wnt3a-GFP viral vector to produce Wnt3a-secreting hMSCs. A total of 50 rats were injured with an Infinite Horizon impactor at the level of the T7-8 vertebrae. Rats were divided into five groups according to the transplanted material: (1) phosphate-buffered saline injection group (sham group, n = 10); (Pertz et al. Proc Natl Acad Sci USA 105:1931-1936, 39) Wnt3a protein injection group (Wnt3a protein group, n = 10); (3) hMSC transplantation group (MSC group, n = 10); (4) hMSCs transfected with the pLenti vector transplantation group (pLenti-MSC group, n = 10); (5) hMSCs transfected with the pLenti+Wnt3a vector transplantation group (Wnt3a-MSC group, n = 10). Behavioral tests were performed daily for the first 3 days after injury and then weekly for 8 weeks. The injured spinal cords were extracted, and axonal regeneration markers including choline acetyltransferase (ChAT), growth-associated protein 43 (GAP43), and microtubule-associated protein 2 (MAP2) were investigated by immunofluorescence, RT-PCR, and western blotting. RESULTS: Seven weeks after the transplantation (8 weeks after SCI), rats in the Wnt3a-MSC group achieved significantly higher average scores in the motor behavior tests than those in the other groups (p < 0.05). Immunofluorescent stains showed greater immunoreactivity of ChAT, GAP43, and MAP2 in the Wnt3a-MSC group than in the other groups. RT-PCR and western blots revealed greater expression of these proteins in the Wnt3a-MSC group than in the other groups (p < 0.05). CONCLUSIONS:Wnt3a-secreting hMSC transplantation considerably improved neurological recovery and axonal regeneration in a rat SCI model.