TY - JOUR
T1 - Brain-derived neurotrophic factor released from engineered mesenchymal stem cells attenuates glutamate- and hydrogen peroxide-mediated death of staurosporine-differentiated RGC-5 cells
AU - Harper, Matthew M.
AU - Adamson, Laura
AU - Blits, Bas
AU - Bunge, Mary Bartlett
AU - Grozdanic, Sinisa D.
AU - Sakaguchi, Donald S.
N1 - Funding Information:
The authors would like to acknowledge Drs. Neeraj Agarwal (University of North Texas Health Sciences Center, for the kind gift of the RGC-5s) and Virginia Lee (University of Pennsylvania, for the gift of the anti-neurofilament antibody). We would also like to thank Roxanne Reger and Darwin Prockop (Center For Gene Therapy, Tulane University) for their help and advice with the mesenchymal stem cells. The MSCs employed in this work were provided by the Tulane Center for Gene Therapy through a grant from NCRR of the NIH, Grant # P40RR017447. Finally, we would like to thank members of the Sakaguchi lab for their help with the preparation of this manuscript.
Funding Information:
Funding: NIGMS RO1 GM072005-01; The Glaucoma Foundation, New York; Fight for Sight, The Stem Cell Research Fund, Biosciences Alliance Initiative from the State of Iowa, The Miami Project to Cure Paralysis.
PY - 2009/10
Y1 - 2009/10
N2 - The purpose of this study was to determine the viability of cell-based delivery of brain-derived neurotrophic factor (BDNF) from genetically modified mesenchymal stem cells (MSCs) for neuroprotection of RGC-5 cells. RGC-5 cells were differentiated with the protein kinase inhibitor staurosporine (SS) and exposed to the cellular stressors glutamate or H2O2. As a neuroprotective strategy, these cells were then co-cultured across a membrane insert with mesenchymal stem cells (MSCs) engineered with a lentiviral vector for production of BDNF (BDNF-MSCs). As a positive control, recombinant human BDNF (rhBDNF) was added to stressed RGC-5 cells. After SS-differentiation RGC-5s developed neuronal-like morphologies, and a significant increase in the proportion of RGC-5s immunoreactive for TuJ-1 and Brn3a was observed. Differentiated RGC-5s also had prominent TrkB staining, demonstrating expression of the high-affinity BDNF receptor. Treatment of SS-differentiated RGC-5s with glutamate or H2O2, produced significant cell death (56.0 ± 7.02 and 48.90 ± 4.58% of control cells, respectively) compared to carrier-solution treated cells. BDNF-delivery from MSCs preserved more RGC-5 cells after treatment with glutamate (80.0 ± 5.40% cells remaining) than control GFP expressing MSCs (GFP-MSCs, 57.29 ± 1.89%, p < 0.01). BDNF-MSCs also protected more RGC-5s after treatment with H2O2 (65.6 ± 3.47%) than GFP-MSCs (46.0 ± 4.20%, p < 0.01). We have shown survival of differentiated RGC-5s is reduced by the cellular stressors glutamate and H2O2. Additionally, our results demonstrate that genetically modified BDNF-producing MSCs can enhance survival of stressed RGC-5 cells and therefore, may be effective vehicles to deliver BDNF to retinal ganglion cells affected by disease.
AB - The purpose of this study was to determine the viability of cell-based delivery of brain-derived neurotrophic factor (BDNF) from genetically modified mesenchymal stem cells (MSCs) for neuroprotection of RGC-5 cells. RGC-5 cells were differentiated with the protein kinase inhibitor staurosporine (SS) and exposed to the cellular stressors glutamate or H2O2. As a neuroprotective strategy, these cells were then co-cultured across a membrane insert with mesenchymal stem cells (MSCs) engineered with a lentiviral vector for production of BDNF (BDNF-MSCs). As a positive control, recombinant human BDNF (rhBDNF) was added to stressed RGC-5 cells. After SS-differentiation RGC-5s developed neuronal-like morphologies, and a significant increase in the proportion of RGC-5s immunoreactive for TuJ-1 and Brn3a was observed. Differentiated RGC-5s also had prominent TrkB staining, demonstrating expression of the high-affinity BDNF receptor. Treatment of SS-differentiated RGC-5s with glutamate or H2O2, produced significant cell death (56.0 ± 7.02 and 48.90 ± 4.58% of control cells, respectively) compared to carrier-solution treated cells. BDNF-delivery from MSCs preserved more RGC-5 cells after treatment with glutamate (80.0 ± 5.40% cells remaining) than control GFP expressing MSCs (GFP-MSCs, 57.29 ± 1.89%, p < 0.01). BDNF-MSCs also protected more RGC-5s after treatment with H2O2 (65.6 ± 3.47%) than GFP-MSCs (46.0 ± 4.20%, p < 0.01). We have shown survival of differentiated RGC-5s is reduced by the cellular stressors glutamate and H2O2. Additionally, our results demonstrate that genetically modified BDNF-producing MSCs can enhance survival of stressed RGC-5 cells and therefore, may be effective vehicles to deliver BDNF to retinal ganglion cells affected by disease.
KW - BDNF
KW - ganglion cells
KW - glaucoma
KW - mesenchymal stem cells
KW - neuroprotection
KW - RGC-5
UR - http://www.scopus.com/inward/record.url?scp=69749118894&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=69749118894&partnerID=8YFLogxK
U2 - 10.1016/j.exer.2009.05.013
DO - 10.1016/j.exer.2009.05.013
M3 - Article
C2 - 19524566
AN - SCOPUS:69749118894
VL - 89
SP - 538
EP - 548
JO - Experimental Eye Research
JF - Experimental Eye Research
SN - 0014-4835
IS - 4
ER -
