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Non-invasive stem cell therapy in a rat model for retinal degeneration and vascular pathology.

Wang S, Lu B, Girman S, Duan J, McFarland T, Zhang QS, Grompe M, Adamus G, Appukuttan B, Lund R - PLoS ONE (2010)

Bottom Line: There is an urgent need to develop therapies that offer generic neuro-and vascular-protective effects with non-invasive intervention.Animals received syngeneic MSCs (1x10(6) cells) by tail vein at an age before major photoreceptor loss. both rod and cone photoreceptors were preserved (5-6 cells thick) at the time when control animal has a single layer of photoreceptors remained; Visual function was significantly preserved compared with controls as determined by visual acuity and luminance threshold recording from the superior colliculus; The number of pathological vascular complexes (abnormal vessels associated with migrating pigment epithelium cells) and area of vascular leakage that would ordinarily develop were dramatically reduced; Semi-quantitative RT-PCR analysis indicated there was upregulation of growth factors and immunohistochemistry revealed that there was an increase in neurotrophic factors within eyes of animals that received MSCs.These results underscore the potential application of MSCs in treating retinal degeneration.

View Article: PubMed Central - PubMed

Affiliation: Casey Eye Institute, Oregon Health & Science University, Portland, Oregon, United States of America. wangsha@ohsu.edu

ABSTRACT

Background: Retinitis pigmentosa (RP) is characterized by progressive night blindness, visual field loss, altered vascular permeability and loss of central vision. Currently there is no effective treatment available except gene replacement therapy has shown promise in a few patients with specific gene defects. There is an urgent need to develop therapies that offer generic neuro-and vascular-protective effects with non-invasive intervention. Here we explored the potential of systemic administration of pluripotent bone marrow-derived mesenchymal stem cells (MSCs) to rescue vision and associated vascular pathology in the Royal College Surgeons (RCS) rat, a well-established animal model for RP.

Methodology/principal findings: Animals received syngeneic MSCs (1x10(6) cells) by tail vein at an age before major photoreceptor loss.

Principal results: both rod and cone photoreceptors were preserved (5-6 cells thick) at the time when control animal has a single layer of photoreceptors remained; Visual function was significantly preserved compared with controls as determined by visual acuity and luminance threshold recording from the superior colliculus; The number of pathological vascular complexes (abnormal vessels associated with migrating pigment epithelium cells) and area of vascular leakage that would ordinarily develop were dramatically reduced; Semi-quantitative RT-PCR analysis indicated there was upregulation of growth factors and immunohistochemistry revealed that there was an increase in neurotrophic factors within eyes of animals that received MSCs.

Conclusions/significance: These results underscore the potential application of MSCs in treating retinal degeneration. The advantages of this non-invasive cell-based therapy are: cells are easily isolated and can be expanded in large quantity for autologous graft; hypoimmunogenic nature as allogeneic donors; less controversial in nature than other stem cells; can be readministered with minor discomfort. Therefore, MSCs may prove to be the ideal cell source for auto-cell therapy for retinal degeneration and other ocular vascular diseases.

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Upregulation of trophic factors.A. Semi-quantitative RT-PCR for CNTF, bFGF, BDNF and beta actin. Lane 1: RNA isolated from MSC prior to injection; Lane 2–4: RNA isolated from retinas treated with MSC; Lane 5–7: RNA isolated from non-treated control retinas. B. Densitometry analysis of CNTF, BDNF and bFGF in treated versus untreated samples. Beta actin was used to normalize the data for comparison. Level of CNTF and BDNF in the treated retinas were significantly higher than non-treated controls (p<0.05), while the level of bFGF in MSC treated retina did not increase significantly. C–J: confocal images of retinal sections double stained with antibodies to CNTF (green) and GFAP (red), counterstained with DAPI (blue in C and G) from MSC treated and controls. Strong CNTF staining in MSC treated retina (D) compared with untreated control (H); E&I: retinal sections stained with GFAP (red) showing upregulation of GFAP in Müller glia in both MSC treated and untreated control; F&J: merged images showing colocalization of CNTF and GFAP in MSC treated retina (F), which was not observed in untreated control (J) (Scale bar equals 50 µm).
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pone-0009200-g004: Upregulation of trophic factors.A. Semi-quantitative RT-PCR for CNTF, bFGF, BDNF and beta actin. Lane 1: RNA isolated from MSC prior to injection; Lane 2–4: RNA isolated from retinas treated with MSC; Lane 5–7: RNA isolated from non-treated control retinas. B. Densitometry analysis of CNTF, BDNF and bFGF in treated versus untreated samples. Beta actin was used to normalize the data for comparison. Level of CNTF and BDNF in the treated retinas were significantly higher than non-treated controls (p<0.05), while the level of bFGF in MSC treated retina did not increase significantly. C–J: confocal images of retinal sections double stained with antibodies to CNTF (green) and GFAP (red), counterstained with DAPI (blue in C and G) from MSC treated and controls. Strong CNTF staining in MSC treated retina (D) compared with untreated control (H); E&I: retinal sections stained with GFAP (red) showing upregulation of GFAP in Müller glia in both MSC treated and untreated control; F&J: merged images showing colocalization of CNTF and GFAP in MSC treated retina (F), which was not observed in untreated control (J) (Scale bar equals 50 µm).

Mentions: We hypothesized that the neuro-vascular protection afforded by the introduction of MSCs was achieved by the increase in production of neurotrophic growth factors within the retina. To investigate this theory we performed semi-quantitative RT-PCR from retinal tissue isolated from animals at P90. We found that growth factors including ciliary neurotrophic factor (CNTF), basic fibroblast growth factor (bFGF), and brain derived neurotrhophic factor were upregulated in MSC treated eyes (n = 3) compared with control eyes (n = 3) (Figure 4A&B). However, only CNTF and BDNF were significantly increased over controls as determined by densitometry analysis. To determine the cells responsible for the increase in this growth factor production in the retina, antibodies against CNTF, bFGF and BDNF were applied to retinal sections of MSC-injected (Figure 4C) and control (Figure 4G). Strong staining of CNTF was found in MSC-injected retina compared with controls (Figure 4D vs.H). There were no obvious difference observed for the other proteins between MSC treated and controls (data not shown). The retinal sections were double stained with glial fibrillary acidic protein (GFAP) (Figure 4E&I) for Müller cells and CNTF, which revealed their co-localization in MSC-injected retina (Figure 4F), not in control (Figure 4J), suggesting that Müller glia cells upregulate expression of CNTF in response to the presence of MSC within the eye.


Non-invasive stem cell therapy in a rat model for retinal degeneration and vascular pathology.

Wang S, Lu B, Girman S, Duan J, McFarland T, Zhang QS, Grompe M, Adamus G, Appukuttan B, Lund R - PLoS ONE (2010)

Upregulation of trophic factors.A. Semi-quantitative RT-PCR for CNTF, bFGF, BDNF and beta actin. Lane 1: RNA isolated from MSC prior to injection; Lane 2–4: RNA isolated from retinas treated with MSC; Lane 5–7: RNA isolated from non-treated control retinas. B. Densitometry analysis of CNTF, BDNF and bFGF in treated versus untreated samples. Beta actin was used to normalize the data for comparison. Level of CNTF and BDNF in the treated retinas were significantly higher than non-treated controls (p<0.05), while the level of bFGF in MSC treated retina did not increase significantly. C–J: confocal images of retinal sections double stained with antibodies to CNTF (green) and GFAP (red), counterstained with DAPI (blue in C and G) from MSC treated and controls. Strong CNTF staining in MSC treated retina (D) compared with untreated control (H); E&I: retinal sections stained with GFAP (red) showing upregulation of GFAP in Müller glia in both MSC treated and untreated control; F&J: merged images showing colocalization of CNTF and GFAP in MSC treated retina (F), which was not observed in untreated control (J) (Scale bar equals 50 µm).
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pone-0009200-g004: Upregulation of trophic factors.A. Semi-quantitative RT-PCR for CNTF, bFGF, BDNF and beta actin. Lane 1: RNA isolated from MSC prior to injection; Lane 2–4: RNA isolated from retinas treated with MSC; Lane 5–7: RNA isolated from non-treated control retinas. B. Densitometry analysis of CNTF, BDNF and bFGF in treated versus untreated samples. Beta actin was used to normalize the data for comparison. Level of CNTF and BDNF in the treated retinas were significantly higher than non-treated controls (p<0.05), while the level of bFGF in MSC treated retina did not increase significantly. C–J: confocal images of retinal sections double stained with antibodies to CNTF (green) and GFAP (red), counterstained with DAPI (blue in C and G) from MSC treated and controls. Strong CNTF staining in MSC treated retina (D) compared with untreated control (H); E&I: retinal sections stained with GFAP (red) showing upregulation of GFAP in Müller glia in both MSC treated and untreated control; F&J: merged images showing colocalization of CNTF and GFAP in MSC treated retina (F), which was not observed in untreated control (J) (Scale bar equals 50 µm).
Mentions: We hypothesized that the neuro-vascular protection afforded by the introduction of MSCs was achieved by the increase in production of neurotrophic growth factors within the retina. To investigate this theory we performed semi-quantitative RT-PCR from retinal tissue isolated from animals at P90. We found that growth factors including ciliary neurotrophic factor (CNTF), basic fibroblast growth factor (bFGF), and brain derived neurotrhophic factor were upregulated in MSC treated eyes (n = 3) compared with control eyes (n = 3) (Figure 4A&B). However, only CNTF and BDNF were significantly increased over controls as determined by densitometry analysis. To determine the cells responsible for the increase in this growth factor production in the retina, antibodies against CNTF, bFGF and BDNF were applied to retinal sections of MSC-injected (Figure 4C) and control (Figure 4G). Strong staining of CNTF was found in MSC-injected retina compared with controls (Figure 4D vs.H). There were no obvious difference observed for the other proteins between MSC treated and controls (data not shown). The retinal sections were double stained with glial fibrillary acidic protein (GFAP) (Figure 4E&I) for Müller cells and CNTF, which revealed their co-localization in MSC-injected retina (Figure 4F), not in control (Figure 4J), suggesting that Müller glia cells upregulate expression of CNTF in response to the presence of MSC within the eye.

Bottom Line: There is an urgent need to develop therapies that offer generic neuro-and vascular-protective effects with non-invasive intervention.Animals received syngeneic MSCs (1x10(6) cells) by tail vein at an age before major photoreceptor loss. both rod and cone photoreceptors were preserved (5-6 cells thick) at the time when control animal has a single layer of photoreceptors remained; Visual function was significantly preserved compared with controls as determined by visual acuity and luminance threshold recording from the superior colliculus; The number of pathological vascular complexes (abnormal vessels associated with migrating pigment epithelium cells) and area of vascular leakage that would ordinarily develop were dramatically reduced; Semi-quantitative RT-PCR analysis indicated there was upregulation of growth factors and immunohistochemistry revealed that there was an increase in neurotrophic factors within eyes of animals that received MSCs.These results underscore the potential application of MSCs in treating retinal degeneration.

View Article: PubMed Central - PubMed

Affiliation: Casey Eye Institute, Oregon Health & Science University, Portland, Oregon, United States of America. wangsha@ohsu.edu

ABSTRACT

Background: Retinitis pigmentosa (RP) is characterized by progressive night blindness, visual field loss, altered vascular permeability and loss of central vision. Currently there is no effective treatment available except gene replacement therapy has shown promise in a few patients with specific gene defects. There is an urgent need to develop therapies that offer generic neuro-and vascular-protective effects with non-invasive intervention. Here we explored the potential of systemic administration of pluripotent bone marrow-derived mesenchymal stem cells (MSCs) to rescue vision and associated vascular pathology in the Royal College Surgeons (RCS) rat, a well-established animal model for RP.

Methodology/principal findings: Animals received syngeneic MSCs (1x10(6) cells) by tail vein at an age before major photoreceptor loss.

Principal results: both rod and cone photoreceptors were preserved (5-6 cells thick) at the time when control animal has a single layer of photoreceptors remained; Visual function was significantly preserved compared with controls as determined by visual acuity and luminance threshold recording from the superior colliculus; The number of pathological vascular complexes (abnormal vessels associated with migrating pigment epithelium cells) and area of vascular leakage that would ordinarily develop were dramatically reduced; Semi-quantitative RT-PCR analysis indicated there was upregulation of growth factors and immunohistochemistry revealed that there was an increase in neurotrophic factors within eyes of animals that received MSCs.

Conclusions/significance: These results underscore the potential application of MSCs in treating retinal degeneration. The advantages of this non-invasive cell-based therapy are: cells are easily isolated and can be expanded in large quantity for autologous graft; hypoimmunogenic nature as allogeneic donors; less controversial in nature than other stem cells; can be readministered with minor discomfort. Therefore, MSCs may prove to be the ideal cell source for auto-cell therapy for retinal degeneration and other ocular vascular diseases.

Show MeSH
Related in: MedlinePlus