Neuroprotective effect of grafting GDNF gene-modified neural stem cells on cerebral ischemia in rats.

Abstract

Previous studies indicated the beneficial effects of glial cell line-derived neurotrophic factor (GDNF) and transplanted neural stem cells (NSCs) on stroke. Here, we explored whether transplantation of neural stem cells (NSCs) modified by GDNF gene provides a better therapeutic effect than native NSCs after stroke. Primary rat NSCs were transfected with GDNF plasmid (GDNF/NSCs, labeled by green fluorescent protein from AdEasy-1, GFP). Adult rats were subjected to two-hour middle cerebral artery occlusion and reperfusion, followed by infusion of NSCs (labeled with5-bromo-2'-deoxyuridine before infusion, BrdU), GDNF/NSCs and saline at 3 days after reperfusion (NSCs group, GDNF/NSCs group, control group), respectively. All rats were sacrificed at 1, 2, 3, 5, and 7 weeks after reperfusion. Modified Neurological Severity Scores (mNSS) test and H and E staining were respectively performed to evaluate neurological function and lesion volume. Immunohistochemistry was used to identify implanted cells and observe the expressions of Synaptophysin (Syp) and postsynaptic density-95 (PSD-95) and caspase-3. TdT-mediated dUTP-biotin nick-end labeling (TUNEL) was employed to observe apoptotic cells. Western blotting was used to detect brain-derived neurotrophic factor (BDNF) and NT-3 protein expression. Significant recovery of mNSS was found in GDNF/NSCs rats at 2 and 3 weeks after reperfusion compared with NSCs rats. Lesion volume in the NSCs and GDNF/NSCs groups was reduced significantly compared with control group. The number of NSCs in the GDNF/NSCs group was significantly increased in comparison with NSCs group. Moreover, Syp-immunoreactive product at 2 and 3 weeks after reperfusion and PSD-95 immunoreactive product in the GDNF/NSCs group were significantly increased compared with NSCs group. In contrast, caspase-3 positive cells and TUNEL-positive cells in the GDNF/NSCs group were significantly decreased compared with NSCs group. Significant increase of BDNF protein in the GDNF/NSCs and NSCs groups was observed compared to the control group at different time points of reperfusion, and GDNF/NSCs grafting significantly increased BDNF protein expression compared to NSCs grafting. In addition, significant increase of NT-3 protein in GDNF/NSCs and NSCs groups was detected only at 1 week of reperfusion compared to control group. The results demonstrate that grafting NSCs modified by GDNF gene provides better neuroprotection for stroke than NSCs grafting alone.

• Authors: B Chen, X-Q Gao, C-X Yang, S-K Tan, Z-L Sun, N-H Yan, Y-G Pang, M Yuan, G-J Chen, G-T Xu, K Zhang, Q L Yuan
• Journal: Brain research (Brain Res) Vol. 1284 Pg. 1-11 (Aug 11 2009) ISSN: 1872-6240 [Electronic] Netherlands
• PMID: 19520066 (Publication Type: Journal Article)

Chemical References

• Brain-Derived Neurotrophic Factor
• Disks Large Homolog 4 Protein
• Dlg4 protein, rat
• Glial Cell Line-Derived Neurotrophic Factor
• Intracellular Signaling Peptides and Proteins
• Membrane Proteins
• Synaptophysin
• Nerve Growth Factor

Topics

• Animals
• Blotting, Western
• Brain Ischemia (metabolism, pathology, therapy)
• Brain-Derived Neurotrophic Factor (biosynthesis)
• Disks Large Homolog 4 Protein
• Female
• Genetic Therapy (methods)
• Glial Cell Line-Derived Neurotrophic Factor (genetics)
• Immunohistochemistry
• In Situ Nick-End Labeling
• Infarction, Middle Cerebral Artery (therapy)
• Intracellular Signaling Peptides and Proteins
• Male
• Membrane Proteins (biosynthesis)
• Nerve Growth Factor (biosynthesis)
• Neurons (transplantation)
• Rats
• Rats, Wistar
• Recovery of Function
• Stem Cell Transplantation (methods)
• Synaptophysin (biosynthesis)
• Transfection