AMP Deaminase 3 knockout mice and loss of mitochondrial proteins and enzyme activity during denervation atrophy

ECU Author/Contributor (non-ECU co-authors, if there are any, appear on document)

Nicolas Verhoeven (Creator)

Institution

East Carolina University (ECU )

Web Site: http://www.ecu.edu/lib/

Abstract: Muscle atrophy leads to decrements in muscle function , partly attributable to decreased mitochondrial content. One controller of mitochondrial content is AMP-activated protein kinase (AMPK) , which when bound by AMP activates PGC-1[alpha]leading to mitochondrial biogenesis and possibly greater mitochondrial content. AMP Deaminase (AMPD: AMP [right arrow]IMP + NH3) isoform 1 is the dominant isoform in skeletal muscle , yet AMPD3 is robustly upregulated during atrophy of skeletal muscle. This study will determine if loss of AMPD3 gene will reduce mitochondrial content loss typically seen with atrophy. Methods: Whole body AMPD3 knock out , heterozygous (HET) , and wildtype (WT) littermate mice (male & female , 10wk old , C57BL6 background) were unilaterally , surgically denervated by sectioning of the sciatic nerve. Contralateral limbs were sham treated. Two weeks later , body composition was measured by Echo MRI. Muscles were collected , weighed , and analyzed for citrate synthase enzyme activity , oxidative phosphorylation proteins , and COX IV protein (measures of mitochondrial content) and expression of AMPD proteins. ANOVA was used to detect significant differences (P[less than]0.05) . Results: AMPD3 genotype had no effect on total body weight , lean mass , or fat mass in either males or females. Likewise , non-denervated EDL and soleus muscle weights were not significantly different among genotypes. As expected , surgical denervation led to substantial muscle atrophy versus innervated muscle in female EDL (~22%) and soleus (~21%) , which were not different among genotypes. In males , similar atrophy occurred in EDL (~23%) and soleus (~22%) with no difference among genotypes. Citrate synthase activity in male innervated muscles was not significantly different among genotypes in EDL , but was significantly different in soleus (16.6±;1.8 WT , 22.3±;2.3 HET , 24.7±;1.4 KO)(p [less than]0.05 vs WT). Citrate synthase activity in female innervated muscles was not significantly different among genotypes in EDL or soleus. Predictably , denervation resulted in loss of CS activity in male EDL (~11%) and soleus (~14%) , as well as female EDL (~14%) and soleus (~14%) , but the loss was not significantly different between genotypes. Complexes I , III , and V proteins , in male EDL , significantly decreased with denervation (p[less than];0.05) , while complexes II and IV were not significantly decreased. All male soleus oxidative phosphorylation complexes decreased with denervation (p[less than];0.05). In female EDL , complexes I and III significantly decreased (p[less than];0.05) , while complex II (p[less than];0.051) and complex V (p[less than];0.09) trended toward significance , and complex IV increased with denervation (p[less than];0.05). Female soleus complexes all significantly decreased (p[less than];0.05). Male EDL and soleus COX IV protein significantly decreased with denervation , as did female EDL and soleus , yet there were no differences between genotypes (p[less than];0.05). AMPD3 protein was solely expressed in wild type denervated conditions and undetectable in knockout , in both muscles. Male and female EDL AMPD1 protein was significantly decreased (~90%) with denervation (p[less than];0.05). Male and female soleus AMPD1 protein was significantly decreased (42%) with denervation (p[less than];0.05). Female soleus AMPD1 was significantly increased versus wild type (p[less than];0.05). Conclusion: These data suggest AMPD3 does not affect mitochondrial content loss in denervation induced skeletal muscle atrophy. However , since AMPD3 is not present during development in our AMPD3 knockout mice , we cannot exclude compensation from other AMPD isoforms. Future research should investigate either inducible muscle specific AMPD3 knockouts or loss of both AMPD1 and AMPD3 , as well the control of AMPD activity.

Additional Information

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Thesis

Language: English

Date: 2018

Keywords

amp deaminase, denervation atrophy

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