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Genetic silencing of Nrf2 enhances X-ROS in dysferlin-deficient muscle.

Kombairaju P, Kerr JP, Roche JA, Pratt SJ, Lovering RM, Sussan TE, Kim JH, Shi G, Biswal S, Ward CW - Front Physiol (2014)

Bottom Line: As nuclear factor erythroid 2-related factor 2 (Nrf2) plays an essential role in the transcriptional regulation of genes involved in redox homeostasis, we hypothesized that Nrf2 deficiency may contribute to enhanced X-ROS signaling by reducing redox buffering.To directly test the effect of diminished Nrf2 activity, Nrf2 was genetically silenced in the A/J model of dysferlinopathy-a model with a mild histopathologic and functional phenotype.Having identified that reduced Nrf2 activity is a negative disease modifier, we propose that strategies targeting Nrf2 activation may address the generalized reduction in redox homeostasis to halt or slow dystrophic progression.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Health Sciences, Bloomberg School of Public Health, Johns Hopkins University Baltimore, MD, USA.

ABSTRACT
Oxidative stress is a critical disease modifier in the muscular dystrophies. Recently, we discovered a pathway by which mechanical stretch activates NADPH Oxidase 2 (Nox2) dependent ROS generation (X-ROS). Our work in dystrophic skeletal muscle revealed that X-ROS is excessive in dystrophin-deficient (mdx) skeletal muscle and contributes to muscle injury susceptibility, a hallmark of the dystrophic process. We also observed widespread alterations in the expression of genes associated with the X-ROS pathway and redox homeostasis in muscles from both Duchenne muscular dystrophy patients and mdx mice. As nuclear factor erythroid 2-related factor 2 (Nrf2) plays an essential role in the transcriptional regulation of genes involved in redox homeostasis, we hypothesized that Nrf2 deficiency may contribute to enhanced X-ROS signaling by reducing redox buffering. To directly test the effect of diminished Nrf2 activity, Nrf2 was genetically silenced in the A/J model of dysferlinopathy-a model with a mild histopathologic and functional phenotype. Nrf2-deficient A/J mice exhibited significant muscle-specific functional deficits, histopathologic abnormalities, and dramatically enhanced X-ROS compared to control A/J and WT mice, both with functional Nrf2. Having identified that reduced Nrf2 activity is a negative disease modifier, we propose that strategies targeting Nrf2 activation may address the generalized reduction in redox homeostasis to halt or slow dystrophic progression.

No MeSH data available.


Related in: MedlinePlus

Histological evidence of enhanced pathology in Nrf2 silenced A/J muscle. (A) Cryosections were H&E stained to evaluate myofiber morphology and immune cell infiltrate (top) and Picrosirius red stained for assaying collagen content (bottom). (B) The Nrf2 silenced A/J muscle exhibited a decrease in total myofiber area, an increase in collagen content, the number of CNFs, and fatty deposits. *p < 0.05.
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Figure 4: Histological evidence of enhanced pathology in Nrf2 silenced A/J muscle. (A) Cryosections were H&E stained to evaluate myofiber morphology and immune cell infiltrate (top) and Picrosirius red stained for assaying collagen content (bottom). (B) The Nrf2 silenced A/J muscle exhibited a decrease in total myofiber area, an increase in collagen content, the number of CNFs, and fatty deposits. *p < 0.05.

Mentions: We have proposed that excessive X-ROS signaling is a proximal mechanism for the muscle damage and force deficits in dystrophic muscle. To assess the effect of Nrf2 silencing on the dysferlin-deficient A/J, we first evaluated nerve-evoked muscle function in vivo. Evaluation of quadriceps muscle function revealed a significant deficit in maximal isometric torque in the A/J-Nrf2−/− vs. the A/J (Figure 3A) despite no difference in bodyweight (27.2 ± 2.1 vs. 28.4 ± 1.9 g). To assess muscle specific force production, we analyzed electrically evoked contractions in single EDL muscles in vitro. An examination of the force (normalized to the muscle cross-sectional area) vs. stimulation frequency relationship revealed a significant deficit in force producing capacity in the A/J-Nrf2−/− (Figure 3B). Histological analysis of the quadriceps muscles provided insight for the deficits in muscle function following Nrf2 ablation, as A/J-Nrf2−/− mice exhibit enhanced degeneration compared to A/J mice (Figure 4). Furthermore, these muscles present with increased immune cell infiltrate (Figure 4A; top, right), fibrosis (Figure 4A; bottom, right), steatosis (fat accumulation), and a concomitant decrease in total myofiber area in cross-section (Figure 4B). Taken together with the increase in the occurrence of CNFs and evidence of increased oxidative stress (Figure 5), we conclude that the A/J-Nrf2−/− mouse presents with a significantly enhanced dystrophic phenotype compared to age-matched A/J mice.


Genetic silencing of Nrf2 enhances X-ROS in dysferlin-deficient muscle.

Kombairaju P, Kerr JP, Roche JA, Pratt SJ, Lovering RM, Sussan TE, Kim JH, Shi G, Biswal S, Ward CW - Front Physiol (2014)

Histological evidence of enhanced pathology in Nrf2 silenced A/J muscle. (A) Cryosections were H&E stained to evaluate myofiber morphology and immune cell infiltrate (top) and Picrosirius red stained for assaying collagen content (bottom). (B) The Nrf2 silenced A/J muscle exhibited a decrease in total myofiber area, an increase in collagen content, the number of CNFs, and fatty deposits. *p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3928547&req=5

Figure 4: Histological evidence of enhanced pathology in Nrf2 silenced A/J muscle. (A) Cryosections were H&E stained to evaluate myofiber morphology and immune cell infiltrate (top) and Picrosirius red stained for assaying collagen content (bottom). (B) The Nrf2 silenced A/J muscle exhibited a decrease in total myofiber area, an increase in collagen content, the number of CNFs, and fatty deposits. *p < 0.05.
Mentions: We have proposed that excessive X-ROS signaling is a proximal mechanism for the muscle damage and force deficits in dystrophic muscle. To assess the effect of Nrf2 silencing on the dysferlin-deficient A/J, we first evaluated nerve-evoked muscle function in vivo. Evaluation of quadriceps muscle function revealed a significant deficit in maximal isometric torque in the A/J-Nrf2−/− vs. the A/J (Figure 3A) despite no difference in bodyweight (27.2 ± 2.1 vs. 28.4 ± 1.9 g). To assess muscle specific force production, we analyzed electrically evoked contractions in single EDL muscles in vitro. An examination of the force (normalized to the muscle cross-sectional area) vs. stimulation frequency relationship revealed a significant deficit in force producing capacity in the A/J-Nrf2−/− (Figure 3B). Histological analysis of the quadriceps muscles provided insight for the deficits in muscle function following Nrf2 ablation, as A/J-Nrf2−/− mice exhibit enhanced degeneration compared to A/J mice (Figure 4). Furthermore, these muscles present with increased immune cell infiltrate (Figure 4A; top, right), fibrosis (Figure 4A; bottom, right), steatosis (fat accumulation), and a concomitant decrease in total myofiber area in cross-section (Figure 4B). Taken together with the increase in the occurrence of CNFs and evidence of increased oxidative stress (Figure 5), we conclude that the A/J-Nrf2−/− mouse presents with a significantly enhanced dystrophic phenotype compared to age-matched A/J mice.

Bottom Line: As nuclear factor erythroid 2-related factor 2 (Nrf2) plays an essential role in the transcriptional regulation of genes involved in redox homeostasis, we hypothesized that Nrf2 deficiency may contribute to enhanced X-ROS signaling by reducing redox buffering.To directly test the effect of diminished Nrf2 activity, Nrf2 was genetically silenced in the A/J model of dysferlinopathy-a model with a mild histopathologic and functional phenotype.Having identified that reduced Nrf2 activity is a negative disease modifier, we propose that strategies targeting Nrf2 activation may address the generalized reduction in redox homeostasis to halt or slow dystrophic progression.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Health Sciences, Bloomberg School of Public Health, Johns Hopkins University Baltimore, MD, USA.

ABSTRACT
Oxidative stress is a critical disease modifier in the muscular dystrophies. Recently, we discovered a pathway by which mechanical stretch activates NADPH Oxidase 2 (Nox2) dependent ROS generation (X-ROS). Our work in dystrophic skeletal muscle revealed that X-ROS is excessive in dystrophin-deficient (mdx) skeletal muscle and contributes to muscle injury susceptibility, a hallmark of the dystrophic process. We also observed widespread alterations in the expression of genes associated with the X-ROS pathway and redox homeostasis in muscles from both Duchenne muscular dystrophy patients and mdx mice. As nuclear factor erythroid 2-related factor 2 (Nrf2) plays an essential role in the transcriptional regulation of genes involved in redox homeostasis, we hypothesized that Nrf2 deficiency may contribute to enhanced X-ROS signaling by reducing redox buffering. To directly test the effect of diminished Nrf2 activity, Nrf2 was genetically silenced in the A/J model of dysferlinopathy-a model with a mild histopathologic and functional phenotype. Nrf2-deficient A/J mice exhibited significant muscle-specific functional deficits, histopathologic abnormalities, and dramatically enhanced X-ROS compared to control A/J and WT mice, both with functional Nrf2. Having identified that reduced Nrf2 activity is a negative disease modifier, we propose that strategies targeting Nrf2 activation may address the generalized reduction in redox homeostasis to halt or slow dystrophic progression.

No MeSH data available.


Related in: MedlinePlus