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Jerry R Mendell - One of the best experts on this subject based on the ideXlab platform.
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microRNA 29 overexpression by adeno associated virus suppresses fibrosis and restores muscle function in combination with micro dystrophin
JCI insight, 2017Co-Authors: Jerry R Mendell, Kristin N Heller, Louise R RodinoklapacAbstract:: Duchenne muscular dystrophy (DMD) is caused by dystrophin deficiency resulting in progressive muscle weakness and fibrotic scarring. Muscle fibrosis impairs blood flow, hampering muscle repair and regeneration. Irrespective of the success of gene restoration, functional improvement is limited without reducing fibrosis. The levels of miR-29c, a known regulator of collagen, are reduced in DMD. Our goal is to develop translational, antifibrotic therapy by overexpressing miR-29c. We injected the gastrocnemius muscle with either self-complementary AAV.CMV.miR-29c or single-stranded AAV.MCK.micro-dystrophin alone or in combination in the mdx/utrn+/- mouse, a DMD mouse model. Treatment of 3-month-old mdx/utrn+/- mice with AAV.miR-29c showed a reduction in collagen and increased absolute and specific force compared with untreated animals, but neither parameter reached WT levels. Combinatorial gene delivery in 3-month-old mdx/utrn+/- mice further decreased fibrosis, and showed a reduction of transcript levels for Col1A, Col3A, fibronectin, and Tgfb1. In addition, absolute and specific force was normalized and equivalent to WT. However, protection against eccentric contraction fell short of WT levels at this time point. When this same mouse model was treated with miR-29c/micro-dystrophin combinatorial therapy at 1 month of age, there was complete normalization of specific and absolute force and protection against eccentric contraction-induced injury was comparable to WT. These findings highlight the potential for miR-29c as an important addition to the armamentarium for translational gene therapy, especially when used in combination with micro-dystrophin in DMD.
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379. microRNA-29 and Micro-Dystrophin Combinatorial Therapy Suppresses Fibrosis and Restores Function to mdx/utrn+/− Mice
Molecular Therapy, 2016Co-Authors: Kristin N Heller, Eric Meadows, Jerry R Mendell, Sarah Lewis, Louise R. Rodino-klapacAbstract:Duchenne muscular dystrophy (DMD) is caused by dystrophin deficiency resulting in muscle loss and progressive muscle weakness and fibrotic scarring. Muscle fibrosis impairs blood flow and excludes endomysial derived constituents hampering muscle repair and regeneration. Irrespective of the success of gene restoration (molecular or pharmacologic) functional improvement is limited without reduction of muscle fibrosis. miR-29c regulates collagen levels making it an ideal candidate for decreasing muscle fibrosis. miR-29c levels are reduced in DMD and our goal is to develop an anti-fibrotic therapy by overexpressing miR-29c with adeno-associated virus (AAV) mediated delivery in combination with micro-dystrophin to improve membrane stability. We injected scAAVrh.74.CMV. miR-29c alone, co-delivered with rAAVrh.74.MCK. micro-dystrophin, and rAAVrh.74.MCK. micro-dystrophin alone by intramuscular injection (IM) into the left gastrocnemius (GAS) muscle of 3 month old mdx/utrn+/- mice, a DMD mouse model. GAS muscle was analyzed 3 months post-injection to assess collagen accumulation by Sirius Red staining and subsequent quantification with ImageJ. Additional outcomes included miR-29c and collagen transcript levels, force measurements in the GAS muscle, fiber diameter measurements and western blot analysis for proteins involved in muscle regeneration (MyoD, Myogenin). Analogous to DMD tissue, we demonstrated a significant reduction in miR-29c levels in mdx/utrn+/− muscle correlated with increased fibrosis measured by Sirius red staining. Following 3 months of treatment with scAAV. miR-29c alone, there was a significant reduction in fibrosis (treated-23.5%±1.3 vs. untreated-27.8% ±0.6, p
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379 microRNA 29 and micro dystrophin combinatorial therapy suppresses fibrosis and restores function to mdx utrn mice
Molecular Therapy, 2016Co-Authors: Kristin N Heller, Eric Meadows, Jerry R Mendell, Sarah Lewis, Louise R RodinoklapacAbstract:Duchenne muscular dystrophy (DMD) is caused by dystrophin deficiency resulting in muscle loss and progressive muscle weakness and fibrotic scarring. Muscle fibrosis impairs blood flow and excludes endomysial derived constituents hampering muscle repair and regeneration. Irrespective of the success of gene restoration (molecular or pharmacologic) functional improvement is limited without reduction of muscle fibrosis. miR-29c regulates collagen levels making it an ideal candidate for decreasing muscle fibrosis. miR-29c levels are reduced in DMD and our goal is to develop an anti-fibrotic therapy by overexpressing miR-29c with adeno-associated virus (AAV) mediated delivery in combination with micro-dystrophin to improve membrane stability. We injected scAAVrh.74.CMV. miR-29c alone, co-delivered with rAAVrh.74.MCK. micro-dystrophin, and rAAVrh.74.MCK. micro-dystrophin alone by intramuscular injection (IM) into the left gastrocnemius (GAS) muscle of 3 month old mdx/utrn+/- mice, a DMD mouse model. GAS muscle was analyzed 3 months post-injection to assess collagen accumulation by Sirius Red staining and subsequent quantification with ImageJ. Additional outcomes included miR-29c and collagen transcript levels, force measurements in the GAS muscle, fiber diameter measurements and western blot analysis for proteins involved in muscle regeneration (MyoD, Myogenin). Analogous to DMD tissue, we demonstrated a significant reduction in miR-29c levels in mdx/utrn+/− muscle correlated with increased fibrosis measured by Sirius red staining. Following 3 months of treatment with scAAV. miR-29c alone, there was a significant reduction in fibrosis (treated-23.5%±1.3 vs. untreated-27.8% ±0.6, p<0.01) in the GAS muscle. When co-delivered with micro-dystrophin we see further reduction in collagen (41%) by Sirius red staining along with significantly reduced mRNA levels of Col1A, Col3A, fibronectin and TGF-β levels. We observed an increase in specific and absolute force in the muscle treated with miR-29c alone compared to the untreated limb, which when combined with micro-dystrophin led to absolute and specific force that were not significantly different than wild-type (miR-29c treated-204.7±11.7 vs. untreated-151.6±14.5 vs. combined-244.2±6.6 vs. wild type-313.1±40.69 p<0.01). We also observed a significant increase in gastroc weight in those muscles that were co-treated. Demonstration of increased fibrosis and decreased miR-29c expression in the mdx/utrn+/− mice and dystrophin-deficient patients validates the mouse model as representative of the human disease. Initial results using AAV. miR-29c as an anti-fibrotic therapy suggest that there is beneficial effect with reduction in collagen levels, a key contributor in fibrosis. Moreover, when combined with micro-dystrophin to improve membrane stability, miR-29 upregulation normalized muscle force. These data provide rationale for overexpression of miR-29c to reduce fibrosis along with dystrophin replacement as a potential treatment for DMD.
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microRNA 29 overexpression delivered by adeno associated virus suppresses fibrosis in mdx utrn mice s61 003
Neurology, 2014Co-Authors: Eric Meadows, Janaiah Kota, Vinod Malik, Reed Clark, Zarife Sahenk, Scott Q Harper, Jerry R MendellAbstract:OBJECTIVE: To demonstrate the degree of muscle fibrosis in mdx:utrn +/- mice and in dystrophin-deficient patient-derived muscle biopsies, correlate changes with microRNA-29 (miR-29), and develop anti-fibrotic therapy using miR-29 delivered by adeno-associated virus (AAV). BACKGROUND: Duchenne muscular dystrophy (DMD) is caused by dystrophin deficiency resulting in muscle loss and progressive weakness, and fibrotic scarring. Muscle fibrosis impairs blood flow and excludes endomysial-derived constituents hampering muscle repair and regeneration. Fibrosis contributes to limb contractures and loss of ambulation. Irrespective of the success of gene restoration (molecular or pharmacologic), functional improvement mandates reduction of muscle fibrosis. mRNA-29 is a potential gene regulator and an ideal candidate for reducing muscle fibrosis. DESIGN/METHODS: Connective tissue was quantified from muscle biopsies of dystrophin-deficient patients, the mdx or mdx:utrn +/- mouse models. Endomysial fibrosis was quantified using Sirius Red-stained sections analyzed by NIH ImageJ Software. 59-nuclease assays were used to measure miR-29 expression correlated with levels of fibrosis. AAV was used to deliver miR-29 into mouse muscle via intramuscular injection. RESULTS: Endomysial connective tissue levels were uniformly increased in the muscles of mdx mice, and in muscle biopsies from DMD patients. Findings correlated with downregulation of miR-29a and miR-29c in the mouse disease model, and in patient biopsies. Initial findings from mdx:utrn +/- mouse muscles treated with AAV-miR29c suggest a favorable anti-fibrotic response that is varied among the treatment groups. CONCLUSIONS: Demonstration of increased fibrosis and decreased miR-29 expression in mdx:utrn +/- mice and dystrophin-deficient patients validates the mouse model as being representative of the human disease. Initial results using AAV-delivered miR29 as an anti-fibrotic therapy suggest that significant beneficial effects can be achieved with further optimization. These data provide a rationale for overexpression of miR-29 to reduce fibrosis in translational, preclinical studies in mdx:utrn +/- mice in preparation for clinical trials. Study Supported by: NIH Ruth L. Kirschstein National Research Service Award (F32) Disclosure: Dr. Meadows has nothing to disclose. Dr. Kota has nothing to disclose. Dr. Malik has nothing to disclose. Dr. Clark has nothing to disclose. Dr. Sahenk has received research support from Sarepta Therapeutics. Dr. Harper has nothing to disclose. Dr. Mendell has received research support from Sarepta Therapeutics Inc.
Wei Xiong - One of the best experts on this subject based on the ideXlab platform.
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microRNA 29 mediates tgfβ1 induced extracellular matrix synthesis by targeting wnt β catenin pathway in human orbital fibroblasts
International Journal of Clinical and Experimental Pathology, 2014Co-Authors: Boding Tong, Yujie Wu, Wei XiongAbstract:Purpose: Transforming growth factor β1 (TGFβ1) is very important in the synthesis and degradation of extracellular matrix (ECM) and also in the mediation of human orbital fibroblasts (OFs) proliferation. microRNA-29 (MiR-29) plays an important role in this process. In the present study, the effects of TGFβ1 on the expression of miR-29 and whether miR-29 is involved in pro-survival signaling pathways mediated by TGFβ1 were examined in human OFs. Methods: Detecting the influence of TGFβ1 on the expression of miR-29a/b/c by real-time PCR analysis. Using 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) to detecting the influence of miR-29 on the increased proliferation caused by TGF-β1 on the human orbital fibroblasts. Using soft agar assay to detecting the influence of miR-29 on the increased colony formation caused by TGF-β1 on the human orbital fibroblasts. Western blot was used to detect the specific mechanisin. Results: TGFβ1 treatment decreases the expression of miR-29 in OFs. In the cultured OFs, the value of optical density (OD) in the group treated with miR-29 is lower than that in the group treated without miR-29 (P < 0.05). In the cultured OFs, the ratio of colony formation in the group treated with miR-29 is lower than that in the group treated without miR-29 (P < 0.05). In OFs, miR-29 decreases the secretion of Wnt3a and activation of β-catenin whether the treatment of TGFβ1 was used or not. MiR-29 decreases expression of Collagen, type I, alpha 1 (COL1A1) through down-regulation of wnt/β-catenin pathway. Conclusions: In OFs TGFβ1 treatment decreases expression of miR-29 which can cause the inhibition of normal ability of TGFβ1. MiR-29 inhibits TGFβ1-induced proliferation of OFS cell and decreases colony formation of OFS cell after TGFβ1 treatment. MiR-29 Mediates TGFβ1-induced Extracellular matrix synthesis through activation of Wnt/β-catenin pathway in human OFs.
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microRNA-29 mediates TGFβ1-induced extracellular matrix synthesis by targeting wnt/β-catenin pathway in human orbital fibroblasts.
International Journal of Clinical and Experimental Pathology, 2014Co-Authors: Boding Tong, Yujie Wu, Wei XiongAbstract:Purpose: Transforming growth factor β1 (TGFβ1) is very important in the synthesis and degradation of extracellular matrix (ECM) and also in the mediation of human orbital fibroblasts (OFs) proliferation. microRNA-29 (MiR-29) plays an important role in this process. In the present study, the effects of TGFβ1 on the expression of miR-29 and whether miR-29 is involved in pro-survival signaling pathways mediated by TGFβ1 were examined in human OFs. Methods: Detecting the influence of TGFβ1 on the expression of miR-29a/b/c by real-time PCR analysis. Using 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) to detecting the influence of miR-29 on the increased proliferation caused by TGF-β1 on the human orbital fibroblasts. Using soft agar assay to detecting the influence of miR-29 on the increased colony formation caused by TGF-β1 on the human orbital fibroblasts. Western blot was used to detect the specific mechanisin. Results: TGFβ1 treatment decreases the expression of miR-29 in OFs. In the cultured OFs, the value of optical density (OD) in the group treated with miR-29 is lower than that in the group treated without miR-29 (P < 0.05). In the cultured OFs, the ratio of colony formation in the group treated with miR-29 is lower than that in the group treated without miR-29 (P < 0.05). In OFs, miR-29 decreases the secretion of Wnt3a and activation of β-catenin whether the treatment of TGFβ1 was used or not. MiR-29 decreases expression of Collagen, type I, alpha 1 (COL1A1) through down-regulation of wnt/β-catenin pathway. Conclusions: In OFs TGFβ1 treatment decreases expression of miR-29 which can cause the inhibition of normal ability of TGFβ1. MiR-29 inhibits TGFβ1-induced proliferation of OFS cell and decreases colony formation of OFS cell after TGFβ1 treatment. MiR-29 Mediates TGFβ1-induced Extracellular matrix synthesis through activation of Wnt/β-catenin pathway in human OFs.
Louise R Rodinoklapac - One of the best experts on this subject based on the ideXlab platform.
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microRNA 29 overexpression by adeno associated virus suppresses fibrosis and restores muscle function in combination with micro dystrophin
JCI insight, 2017Co-Authors: Jerry R Mendell, Kristin N Heller, Louise R RodinoklapacAbstract:: Duchenne muscular dystrophy (DMD) is caused by dystrophin deficiency resulting in progressive muscle weakness and fibrotic scarring. Muscle fibrosis impairs blood flow, hampering muscle repair and regeneration. Irrespective of the success of gene restoration, functional improvement is limited without reducing fibrosis. The levels of miR-29c, a known regulator of collagen, are reduced in DMD. Our goal is to develop translational, antifibrotic therapy by overexpressing miR-29c. We injected the gastrocnemius muscle with either self-complementary AAV.CMV.miR-29c or single-stranded AAV.MCK.micro-dystrophin alone or in combination in the mdx/utrn+/- mouse, a DMD mouse model. Treatment of 3-month-old mdx/utrn+/- mice with AAV.miR-29c showed a reduction in collagen and increased absolute and specific force compared with untreated animals, but neither parameter reached WT levels. Combinatorial gene delivery in 3-month-old mdx/utrn+/- mice further decreased fibrosis, and showed a reduction of transcript levels for Col1A, Col3A, fibronectin, and Tgfb1. In addition, absolute and specific force was normalized and equivalent to WT. However, protection against eccentric contraction fell short of WT levels at this time point. When this same mouse model was treated with miR-29c/micro-dystrophin combinatorial therapy at 1 month of age, there was complete normalization of specific and absolute force and protection against eccentric contraction-induced injury was comparable to WT. These findings highlight the potential for miR-29c as an important addition to the armamentarium for translational gene therapy, especially when used in combination with micro-dystrophin in DMD.
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379 microRNA 29 and micro dystrophin combinatorial therapy suppresses fibrosis and restores function to mdx utrn mice
Molecular Therapy, 2016Co-Authors: Kristin N Heller, Eric Meadows, Jerry R Mendell, Sarah Lewis, Louise R RodinoklapacAbstract:Duchenne muscular dystrophy (DMD) is caused by dystrophin deficiency resulting in muscle loss and progressive muscle weakness and fibrotic scarring. Muscle fibrosis impairs blood flow and excludes endomysial derived constituents hampering muscle repair and regeneration. Irrespective of the success of gene restoration (molecular or pharmacologic) functional improvement is limited without reduction of muscle fibrosis. miR-29c regulates collagen levels making it an ideal candidate for decreasing muscle fibrosis. miR-29c levels are reduced in DMD and our goal is to develop an anti-fibrotic therapy by overexpressing miR-29c with adeno-associated virus (AAV) mediated delivery in combination with micro-dystrophin to improve membrane stability. We injected scAAVrh.74.CMV. miR-29c alone, co-delivered with rAAVrh.74.MCK. micro-dystrophin, and rAAVrh.74.MCK. micro-dystrophin alone by intramuscular injection (IM) into the left gastrocnemius (GAS) muscle of 3 month old mdx/utrn+/- mice, a DMD mouse model. GAS muscle was analyzed 3 months post-injection to assess collagen accumulation by Sirius Red staining and subsequent quantification with ImageJ. Additional outcomes included miR-29c and collagen transcript levels, force measurements in the GAS muscle, fiber diameter measurements and western blot analysis for proteins involved in muscle regeneration (MyoD, Myogenin). Analogous to DMD tissue, we demonstrated a significant reduction in miR-29c levels in mdx/utrn+/− muscle correlated with increased fibrosis measured by Sirius red staining. Following 3 months of treatment with scAAV. miR-29c alone, there was a significant reduction in fibrosis (treated-23.5%±1.3 vs. untreated-27.8% ±0.6, p<0.01) in the GAS muscle. When co-delivered with micro-dystrophin we see further reduction in collagen (41%) by Sirius red staining along with significantly reduced mRNA levels of Col1A, Col3A, fibronectin and TGF-β levels. We observed an increase in specific and absolute force in the muscle treated with miR-29c alone compared to the untreated limb, which when combined with micro-dystrophin led to absolute and specific force that were not significantly different than wild-type (miR-29c treated-204.7±11.7 vs. untreated-151.6±14.5 vs. combined-244.2±6.6 vs. wild type-313.1±40.69 p<0.01). We also observed a significant increase in gastroc weight in those muscles that were co-treated. Demonstration of increased fibrosis and decreased miR-29c expression in the mdx/utrn+/− mice and dystrophin-deficient patients validates the mouse model as representative of the human disease. Initial results using AAV. miR-29c as an anti-fibrotic therapy suggest that there is beneficial effect with reduction in collagen levels, a key contributor in fibrosis. Moreover, when combined with micro-dystrophin to improve membrane stability, miR-29 upregulation normalized muscle force. These data provide rationale for overexpression of miR-29c to reduce fibrosis along with dystrophin replacement as a potential treatment for DMD.
Eric Meadows - One of the best experts on this subject based on the ideXlab platform.
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379. microRNA-29 and Micro-Dystrophin Combinatorial Therapy Suppresses Fibrosis and Restores Function to mdx/utrn+/− Mice
Molecular Therapy, 2016Co-Authors: Kristin N Heller, Eric Meadows, Jerry R Mendell, Sarah Lewis, Louise R. Rodino-klapacAbstract:Duchenne muscular dystrophy (DMD) is caused by dystrophin deficiency resulting in muscle loss and progressive muscle weakness and fibrotic scarring. Muscle fibrosis impairs blood flow and excludes endomysial derived constituents hampering muscle repair and regeneration. Irrespective of the success of gene restoration (molecular or pharmacologic) functional improvement is limited without reduction of muscle fibrosis. miR-29c regulates collagen levels making it an ideal candidate for decreasing muscle fibrosis. miR-29c levels are reduced in DMD and our goal is to develop an anti-fibrotic therapy by overexpressing miR-29c with adeno-associated virus (AAV) mediated delivery in combination with micro-dystrophin to improve membrane stability. We injected scAAVrh.74.CMV. miR-29c alone, co-delivered with rAAVrh.74.MCK. micro-dystrophin, and rAAVrh.74.MCK. micro-dystrophin alone by intramuscular injection (IM) into the left gastrocnemius (GAS) muscle of 3 month old mdx/utrn+/- mice, a DMD mouse model. GAS muscle was analyzed 3 months post-injection to assess collagen accumulation by Sirius Red staining and subsequent quantification with ImageJ. Additional outcomes included miR-29c and collagen transcript levels, force measurements in the GAS muscle, fiber diameter measurements and western blot analysis for proteins involved in muscle regeneration (MyoD, Myogenin). Analogous to DMD tissue, we demonstrated a significant reduction in miR-29c levels in mdx/utrn+/− muscle correlated with increased fibrosis measured by Sirius red staining. Following 3 months of treatment with scAAV. miR-29c alone, there was a significant reduction in fibrosis (treated-23.5%±1.3 vs. untreated-27.8% ±0.6, p
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379 microRNA 29 and micro dystrophin combinatorial therapy suppresses fibrosis and restores function to mdx utrn mice
Molecular Therapy, 2016Co-Authors: Kristin N Heller, Eric Meadows, Jerry R Mendell, Sarah Lewis, Louise R RodinoklapacAbstract:Duchenne muscular dystrophy (DMD) is caused by dystrophin deficiency resulting in muscle loss and progressive muscle weakness and fibrotic scarring. Muscle fibrosis impairs blood flow and excludes endomysial derived constituents hampering muscle repair and regeneration. Irrespective of the success of gene restoration (molecular or pharmacologic) functional improvement is limited without reduction of muscle fibrosis. miR-29c regulates collagen levels making it an ideal candidate for decreasing muscle fibrosis. miR-29c levels are reduced in DMD and our goal is to develop an anti-fibrotic therapy by overexpressing miR-29c with adeno-associated virus (AAV) mediated delivery in combination with micro-dystrophin to improve membrane stability. We injected scAAVrh.74.CMV. miR-29c alone, co-delivered with rAAVrh.74.MCK. micro-dystrophin, and rAAVrh.74.MCK. micro-dystrophin alone by intramuscular injection (IM) into the left gastrocnemius (GAS) muscle of 3 month old mdx/utrn+/- mice, a DMD mouse model. GAS muscle was analyzed 3 months post-injection to assess collagen accumulation by Sirius Red staining and subsequent quantification with ImageJ. Additional outcomes included miR-29c and collagen transcript levels, force measurements in the GAS muscle, fiber diameter measurements and western blot analysis for proteins involved in muscle regeneration (MyoD, Myogenin). Analogous to DMD tissue, we demonstrated a significant reduction in miR-29c levels in mdx/utrn+/− muscle correlated with increased fibrosis measured by Sirius red staining. Following 3 months of treatment with scAAV. miR-29c alone, there was a significant reduction in fibrosis (treated-23.5%±1.3 vs. untreated-27.8% ±0.6, p<0.01) in the GAS muscle. When co-delivered with micro-dystrophin we see further reduction in collagen (41%) by Sirius red staining along with significantly reduced mRNA levels of Col1A, Col3A, fibronectin and TGF-β levels. We observed an increase in specific and absolute force in the muscle treated with miR-29c alone compared to the untreated limb, which when combined with micro-dystrophin led to absolute and specific force that were not significantly different than wild-type (miR-29c treated-204.7±11.7 vs. untreated-151.6±14.5 vs. combined-244.2±6.6 vs. wild type-313.1±40.69 p<0.01). We also observed a significant increase in gastroc weight in those muscles that were co-treated. Demonstration of increased fibrosis and decreased miR-29c expression in the mdx/utrn+/− mice and dystrophin-deficient patients validates the mouse model as representative of the human disease. Initial results using AAV. miR-29c as an anti-fibrotic therapy suggest that there is beneficial effect with reduction in collagen levels, a key contributor in fibrosis. Moreover, when combined with micro-dystrophin to improve membrane stability, miR-29 upregulation normalized muscle force. These data provide rationale for overexpression of miR-29c to reduce fibrosis along with dystrophin replacement as a potential treatment for DMD.
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microRNA 29 overexpression delivered by adeno associated virus suppresses fibrosis in mdx utrn mice s61 003
Neurology, 2014Co-Authors: Eric Meadows, Janaiah Kota, Vinod Malik, Reed Clark, Zarife Sahenk, Scott Q Harper, Jerry R MendellAbstract:OBJECTIVE: To demonstrate the degree of muscle fibrosis in mdx:utrn +/- mice and in dystrophin-deficient patient-derived muscle biopsies, correlate changes with microRNA-29 (miR-29), and develop anti-fibrotic therapy using miR-29 delivered by adeno-associated virus (AAV). BACKGROUND: Duchenne muscular dystrophy (DMD) is caused by dystrophin deficiency resulting in muscle loss and progressive weakness, and fibrotic scarring. Muscle fibrosis impairs blood flow and excludes endomysial-derived constituents hampering muscle repair and regeneration. Fibrosis contributes to limb contractures and loss of ambulation. Irrespective of the success of gene restoration (molecular or pharmacologic), functional improvement mandates reduction of muscle fibrosis. mRNA-29 is a potential gene regulator and an ideal candidate for reducing muscle fibrosis. DESIGN/METHODS: Connective tissue was quantified from muscle biopsies of dystrophin-deficient patients, the mdx or mdx:utrn +/- mouse models. Endomysial fibrosis was quantified using Sirius Red-stained sections analyzed by NIH ImageJ Software. 59-nuclease assays were used to measure miR-29 expression correlated with levels of fibrosis. AAV was used to deliver miR-29 into mouse muscle via intramuscular injection. RESULTS: Endomysial connective tissue levels were uniformly increased in the muscles of mdx mice, and in muscle biopsies from DMD patients. Findings correlated with downregulation of miR-29a and miR-29c in the mouse disease model, and in patient biopsies. Initial findings from mdx:utrn +/- mouse muscles treated with AAV-miR29c suggest a favorable anti-fibrotic response that is varied among the treatment groups. CONCLUSIONS: Demonstration of increased fibrosis and decreased miR-29 expression in mdx:utrn +/- mice and dystrophin-deficient patients validates the mouse model as being representative of the human disease. Initial results using AAV-delivered miR29 as an anti-fibrotic therapy suggest that significant beneficial effects can be achieved with further optimization. These data provide a rationale for overexpression of miR-29 to reduce fibrosis in translational, preclinical studies in mdx:utrn +/- mice in preparation for clinical trials. Study Supported by: NIH Ruth L. Kirschstein National Research Service Award (F32) Disclosure: Dr. Meadows has nothing to disclose. Dr. Kota has nothing to disclose. Dr. Malik has nothing to disclose. Dr. Clark has nothing to disclose. Dr. Sahenk has received research support from Sarepta Therapeutics. Dr. Harper has nothing to disclose. Dr. Mendell has received research support from Sarepta Therapeutics Inc.
Boding Tong - One of the best experts on this subject based on the ideXlab platform.
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microRNA 29 mediates tgfβ1 induced extracellular matrix synthesis by targeting wnt β catenin pathway in human orbital fibroblasts
International Journal of Clinical and Experimental Pathology, 2014Co-Authors: Boding Tong, Yujie Wu, Wei XiongAbstract:Purpose: Transforming growth factor β1 (TGFβ1) is very important in the synthesis and degradation of extracellular matrix (ECM) and also in the mediation of human orbital fibroblasts (OFs) proliferation. microRNA-29 (MiR-29) plays an important role in this process. In the present study, the effects of TGFβ1 on the expression of miR-29 and whether miR-29 is involved in pro-survival signaling pathways mediated by TGFβ1 were examined in human OFs. Methods: Detecting the influence of TGFβ1 on the expression of miR-29a/b/c by real-time PCR analysis. Using 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) to detecting the influence of miR-29 on the increased proliferation caused by TGF-β1 on the human orbital fibroblasts. Using soft agar assay to detecting the influence of miR-29 on the increased colony formation caused by TGF-β1 on the human orbital fibroblasts. Western blot was used to detect the specific mechanisin. Results: TGFβ1 treatment decreases the expression of miR-29 in OFs. In the cultured OFs, the value of optical density (OD) in the group treated with miR-29 is lower than that in the group treated without miR-29 (P < 0.05). In the cultured OFs, the ratio of colony formation in the group treated with miR-29 is lower than that in the group treated without miR-29 (P < 0.05). In OFs, miR-29 decreases the secretion of Wnt3a and activation of β-catenin whether the treatment of TGFβ1 was used or not. MiR-29 decreases expression of Collagen, type I, alpha 1 (COL1A1) through down-regulation of wnt/β-catenin pathway. Conclusions: In OFs TGFβ1 treatment decreases expression of miR-29 which can cause the inhibition of normal ability of TGFβ1. MiR-29 inhibits TGFβ1-induced proliferation of OFS cell and decreases colony formation of OFS cell after TGFβ1 treatment. MiR-29 Mediates TGFβ1-induced Extracellular matrix synthesis through activation of Wnt/β-catenin pathway in human OFs.
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microRNA-29 mediates TGFβ1-induced extracellular matrix synthesis by targeting wnt/β-catenin pathway in human orbital fibroblasts.
International Journal of Clinical and Experimental Pathology, 2014Co-Authors: Boding Tong, Yujie Wu, Wei XiongAbstract:Purpose: Transforming growth factor β1 (TGFβ1) is very important in the synthesis and degradation of extracellular matrix (ECM) and also in the mediation of human orbital fibroblasts (OFs) proliferation. microRNA-29 (MiR-29) plays an important role in this process. In the present study, the effects of TGFβ1 on the expression of miR-29 and whether miR-29 is involved in pro-survival signaling pathways mediated by TGFβ1 were examined in human OFs. Methods: Detecting the influence of TGFβ1 on the expression of miR-29a/b/c by real-time PCR analysis. Using 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) to detecting the influence of miR-29 on the increased proliferation caused by TGF-β1 on the human orbital fibroblasts. Using soft agar assay to detecting the influence of miR-29 on the increased colony formation caused by TGF-β1 on the human orbital fibroblasts. Western blot was used to detect the specific mechanisin. Results: TGFβ1 treatment decreases the expression of miR-29 in OFs. In the cultured OFs, the value of optical density (OD) in the group treated with miR-29 is lower than that in the group treated without miR-29 (P < 0.05). In the cultured OFs, the ratio of colony formation in the group treated with miR-29 is lower than that in the group treated without miR-29 (P < 0.05). In OFs, miR-29 decreases the secretion of Wnt3a and activation of β-catenin whether the treatment of TGFβ1 was used or not. MiR-29 decreases expression of Collagen, type I, alpha 1 (COL1A1) through down-regulation of wnt/β-catenin pathway. Conclusions: In OFs TGFβ1 treatment decreases expression of miR-29 which can cause the inhibition of normal ability of TGFβ1. MiR-29 inhibits TGFβ1-induced proliferation of OFS cell and decreases colony formation of OFS cell after TGFβ1 treatment. MiR-29 Mediates TGFβ1-induced Extracellular matrix synthesis through activation of Wnt/β-catenin pathway in human OFs.