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Life or death by NFκB, Losartan promotes survival in dy2J/dy2J mouse of MDC1A.

Elbaz M, Yanay N, Laban S, Rabie M, Mitrani-Rosenbaum S, Nevo Y - Cell Death Dis (2015)

Bottom Line: Losartan treatment was associated with significantly increased serum tumor necrosis factor alpha (TNF-α) level, p65 nuclei accumulation, and decreased muscle IκB-β protein level, indicating NFκB activation.Losartan induced protein expression toward a pro-survival profile as BCL-2 expression levels were increased and Caspase-3 expression levels were decreased.Muscle apoptosis reduction was further confirmed using terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) assay.

View Article: PubMed Central - PubMed

Affiliation: Pediatric Neuromuscular Laboratory and Neuropediatric Unit, Hadassah - Hebrew University Medical Center, Jerusalem, Israel.

ABSTRACT
Inflammation and fibrosis are well-defined mechanisms involved in the pathogenesis of the incurable Laminin α2-deficient congenital muscular dystrophy (MDC1A), while apoptosis mechanism is barely discussed. Our previous study showed treatment with Losartan, an angiotensin II type I receptor antagonist, improved muscle strength and reduced fibrosis through transforming growth factor beta (TGF-β) and mitogen-activated protein kinases (MAPK) signaling inhibition in the dy(2J)/dy(2J) mouse model of MDC1A. Here we show for the first time that Losartan treatment up-regulates and shifts the nuclear factor kappa B (NFκB) signaling pathway to favor survival versus apoptosis/damage in this animal model. Losartan treatment was associated with significantly increased serum tumor necrosis factor alpha (TNF-α) level, p65 nuclei accumulation, and decreased muscle IκB-β protein level, indicating NFκB activation. Moreover, NFκB anti-apoptotic target genes TNF receptor-associated factor 1 (TRAF1), TNF receptor-associated factor 2 (TRAF2), cellular inhibitor of apoptosis (cIAP2), and Ferritin heavy chain (FTH1) were increased following Losartan treatment. Losartan induced protein expression toward a pro-survival profile as BCL-2 expression levels were increased and Caspase-3 expression levels were decreased. Muscle apoptosis reduction was further confirmed using terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) assay. Thus, along with TGF-β and MAPK signaling, NFκB serves as an important regulatory pathway which following Losartan treatment promotes survival in the dy(2J)/dy(2J) mouse model of MDC1A.

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Increased protein expression of anti-apoptotic BCL-2 and decreased protein expression of pro-apoptotic Caspase 3 following Losartan treatment. (a) Representative western blot gel and densitometry graph of BCL-2 expression in WT and dy2J/dy2J mice. A significantly higher BCL-2 protein expression level was noted in treated compared with untreated dy2J/dy2J mice (*P<0.01). (b) Representative western blot gel and densitometry graph of Caspase-3 expression in WT and dy2J/dy2J mice. A significantly lower Caspase-3 protein expression level was noted in treated compared with untreated dy2J/dy2J mice (*P<0.0001). Results of BCL-2 and Caspase-3 levels were obtained from densitometric analysis and expressed as ratio of BCL-2 and Caspase-3 to GAPDH and as change fold over control (WT group). These results represent three independent experiments. Each bar represents the mean±S.E.M. of 12 mice for BCL-2 (*P<0.01) and 12 mice for Caspase-3 (*/**P<0.0001)
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fig4: Increased protein expression of anti-apoptotic BCL-2 and decreased protein expression of pro-apoptotic Caspase 3 following Losartan treatment. (a) Representative western blot gel and densitometry graph of BCL-2 expression in WT and dy2J/dy2J mice. A significantly higher BCL-2 protein expression level was noted in treated compared with untreated dy2J/dy2J mice (*P<0.01). (b) Representative western blot gel and densitometry graph of Caspase-3 expression in WT and dy2J/dy2J mice. A significantly lower Caspase-3 protein expression level was noted in treated compared with untreated dy2J/dy2J mice (*P<0.0001). Results of BCL-2 and Caspase-3 levels were obtained from densitometric analysis and expressed as ratio of BCL-2 and Caspase-3 to GAPDH and as change fold over control (WT group). These results represent three independent experiments. Each bar represents the mean±S.E.M. of 12 mice for BCL-2 (*P<0.01) and 12 mice for Caspase-3 (*/**P<0.0001)

Mentions: Next using western blot analysis we examined the hind limb expression of B-cell lymphoma 2 (BCL-2), an anti-apoptotic protein. BCL-2 expression was significantly higher in Losartan treated compared with untreated dy2J/dy2J mice (treated dy2J/dy2J: 0.90±0.052-fold versus untreated dy2J/dy2J: 0.59±0.071-fold; *P<0.01). There was no significant difference in BCL-2 expression between treated and untreated WT mice (Figure 4a).


Life or death by NFκB, Losartan promotes survival in dy2J/dy2J mouse of MDC1A.

Elbaz M, Yanay N, Laban S, Rabie M, Mitrani-Rosenbaum S, Nevo Y - Cell Death Dis (2015)

Increased protein expression of anti-apoptotic BCL-2 and decreased protein expression of pro-apoptotic Caspase 3 following Losartan treatment. (a) Representative western blot gel and densitometry graph of BCL-2 expression in WT and dy2J/dy2J mice. A significantly higher BCL-2 protein expression level was noted in treated compared with untreated dy2J/dy2J mice (*P<0.01). (b) Representative western blot gel and densitometry graph of Caspase-3 expression in WT and dy2J/dy2J mice. A significantly lower Caspase-3 protein expression level was noted in treated compared with untreated dy2J/dy2J mice (*P<0.0001). Results of BCL-2 and Caspase-3 levels were obtained from densitometric analysis and expressed as ratio of BCL-2 and Caspase-3 to GAPDH and as change fold over control (WT group). These results represent three independent experiments. Each bar represents the mean±S.E.M. of 12 mice for BCL-2 (*P<0.01) and 12 mice for Caspase-3 (*/**P<0.0001)
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Related In: Results  -  Collection

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fig4: Increased protein expression of anti-apoptotic BCL-2 and decreased protein expression of pro-apoptotic Caspase 3 following Losartan treatment. (a) Representative western blot gel and densitometry graph of BCL-2 expression in WT and dy2J/dy2J mice. A significantly higher BCL-2 protein expression level was noted in treated compared with untreated dy2J/dy2J mice (*P<0.01). (b) Representative western blot gel and densitometry graph of Caspase-3 expression in WT and dy2J/dy2J mice. A significantly lower Caspase-3 protein expression level was noted in treated compared with untreated dy2J/dy2J mice (*P<0.0001). Results of BCL-2 and Caspase-3 levels were obtained from densitometric analysis and expressed as ratio of BCL-2 and Caspase-3 to GAPDH and as change fold over control (WT group). These results represent three independent experiments. Each bar represents the mean±S.E.M. of 12 mice for BCL-2 (*P<0.01) and 12 mice for Caspase-3 (*/**P<0.0001)
Mentions: Next using western blot analysis we examined the hind limb expression of B-cell lymphoma 2 (BCL-2), an anti-apoptotic protein. BCL-2 expression was significantly higher in Losartan treated compared with untreated dy2J/dy2J mice (treated dy2J/dy2J: 0.90±0.052-fold versus untreated dy2J/dy2J: 0.59±0.071-fold; *P<0.01). There was no significant difference in BCL-2 expression between treated and untreated WT mice (Figure 4a).

Bottom Line: Losartan treatment was associated with significantly increased serum tumor necrosis factor alpha (TNF-α) level, p65 nuclei accumulation, and decreased muscle IκB-β protein level, indicating NFκB activation.Losartan induced protein expression toward a pro-survival profile as BCL-2 expression levels were increased and Caspase-3 expression levels were decreased.Muscle apoptosis reduction was further confirmed using terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) assay.

View Article: PubMed Central - PubMed

Affiliation: Pediatric Neuromuscular Laboratory and Neuropediatric Unit, Hadassah - Hebrew University Medical Center, Jerusalem, Israel.

ABSTRACT
Inflammation and fibrosis are well-defined mechanisms involved in the pathogenesis of the incurable Laminin α2-deficient congenital muscular dystrophy (MDC1A), while apoptosis mechanism is barely discussed. Our previous study showed treatment with Losartan, an angiotensin II type I receptor antagonist, improved muscle strength and reduced fibrosis through transforming growth factor beta (TGF-β) and mitogen-activated protein kinases (MAPK) signaling inhibition in the dy(2J)/dy(2J) mouse model of MDC1A. Here we show for the first time that Losartan treatment up-regulates and shifts the nuclear factor kappa B (NFκB) signaling pathway to favor survival versus apoptosis/damage in this animal model. Losartan treatment was associated with significantly increased serum tumor necrosis factor alpha (TNF-α) level, p65 nuclei accumulation, and decreased muscle IκB-β protein level, indicating NFκB activation. Moreover, NFκB anti-apoptotic target genes TNF receptor-associated factor 1 (TRAF1), TNF receptor-associated factor 2 (TRAF2), cellular inhibitor of apoptosis (cIAP2), and Ferritin heavy chain (FTH1) were increased following Losartan treatment. Losartan induced protein expression toward a pro-survival profile as BCL-2 expression levels were increased and Caspase-3 expression levels were decreased. Muscle apoptosis reduction was further confirmed using terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) assay. Thus, along with TGF-β and MAPK signaling, NFκB serves as an important regulatory pathway which following Losartan treatment promotes survival in the dy(2J)/dy(2J) mouse model of MDC1A.

Show MeSH
Related in: MedlinePlus