Overexpression of CNTF in Mesenchymal Stem Cells reduces demyelination and induces clinical recovery in experimental autoimmune encephalomyelitis mice.
Human Mesenchymal Stem Cells (MSCs) were previously reported to ameliorate neuronal functional deficits in the MOG35-55-induced experimental autoimmune encephalomyelitis (EAE) mice by inducing T cell anergy. Human Ciliary neurotrophic factor (CNTF) recently was found to promote myelogenesis and reduce inflammation in CNTF-deficient EAE mice. We ectopically overexpressed CNTF in human MSCs to investigate its potential role in promoting remyelination and improving functional recovery in EAE mice. MSCs transfected by Ad-CNTF-IRES-EGFP (MSC-CNTF) were injected intravenously into EAE mice 10 days after the immunization. Neurological functional tests were scored daily by grading clinical signs (score 0-6). Immunofluorescence microscopy was used to detect MSC-CNTF in spinal cord. Expression of NG2, CNTF, and cleaved caspase-3 was measured by immunohistochemistry. CNTF expression was also analyzed by Western blot. Myelin was detected by Solochrome Cyanin staining. Our results found that CNTF concentration in MSC-CNTF cells was 20-fold higher than that in either MSC or Ad-EGFP-transfected MSCs (MSC-EGFP) in vitro. Mice receiving MSC-CNTF cells showed remarkable neuronal functional recovery: the cumulative clinical scores were significantly decreased, and the disease onset was statistically delayed. Mice receiving MSC-CNTF cells showed reduced TNF-alpha, IFN-gamma and increased the level of cytokine IL-10 in peripheral blood and a large number of MSC-CNTF cells were detected in the spleen, but were not detected in other organs such as lung, liver and kidney. In the lesions of these mice, 1) the number of cleaved caspase3-positive cells was significantly reduced; 2) MSC-CNTF- and NG2-positive cells were significantly increased; and 3) the expression of CNTF was dramatically increased. In addition, demyelination was significantly reduced in MSC-CNTF mice. These data indicated that MSC-CNTF may improve functional recovery in EAE mice, possibly by exerting their immunoregulatory activity, inhibiting inflammation, homing MSC-CNTF cells to the lesions, elevating CNTF expression, reducing demyelination, and stimulating oligodendrogenesis.
Lu
Zhengqi
Z
Department of Neurology, the Third Affiliated Hospital of Sun Yat-Sen University, 600 Tianhe Road, Guangzhou, Guangdong 510630, China.
Hu
Xueqiang
X
Zhu
Cansheng
C
Wang
Dunjing
D
Zheng
Xueping
X
Liu
Quentin
Q
D000256:Adenoviridae / Q000235:genetics; D000596:Amino Acids / Q000187:drug effects; D000704:Analysis of Variance; D000818:Animals; D000941:Antigens / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D002452:Cell Count; D020934:Ciliary Neurotrophic Factor / Q000172:deficiency / Q000235:genetics / Q000627:therapeutic use*; D016207:Cytokines / Q000097:blood; D003711:Demyelinating Diseases / Q000628:therapy*; D004195:Disease Models, Animal; D004681:Encephalomyelitis, Autoimmune, Experimental / Q000139:chemically induced / Q000188:drug therapy* / Q000473:pathology / Q000503:physiopathology; D005260:Female; D005822:Genetic Vectors / Q000502:physiology; D006023:Glycoproteins; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D045164:Mesenchymal Stem Cell Transplantation / Q000379:methods; D059630:Mesenchymal Stem Cells / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D063308:Myelin-Oligodendrocyte Glycoprotein; D010446:Peptide Fragments; D011509:Proteoglycans / Q000378:metabolism; D020127:Recovery of Function / Q000235:genetics / Q000502:physiology*; D018709:Statistics, Nonparametric; D013997:Time Factors; D000099216:Chondroitin Sulfate Proteoglycan 4
D000596:Amino Acids; D000941:Antigens; D053148:Caspase 3; D020934:Ciliary Neurotrophic Factor; D016207:Cytokines; D006023:Glycoproteins; D049452:Green Fluorescent Proteins; D063308:Myelin-Oligodendrocyte Glycoprotein; D010446:Peptide Fragments; D011509:Proteoglycans; D000099216:Chondroitin Sulfate Proteoglycan 4; C098463:dolaisoleucine; C485184:enhanced green fluorescent protein; C445081:myelin oligodendrocyte glycoprotein (35-55)