Friedreich's ataxia (FRDA) is a neurodegenerative disease caused by reduced expression of the mitochondrial protein frataxin (FXN). Most FRDA patients are homozygous for large expansions of GAA ...repeats in intron 1 of FXN, while some are compound heterozygotes with an expanded GAA tract in one allele and a missense or nonsense mutation in the other. A missense mutation, changing a glycine to valine at position 130 (G130V), is prevalent among the clinical variants. We and others have demonstrated that levels of mature FXN protein in FRDA G130V samples are reduced below those detected in samples harboring homozygous repeat expansions. Little is known regarding expression and function of endogenous FXN-G130V protein due to lack of reagents and models that can distinguish the mutant FXN protein from the wild-type FXN produced from the GAA-expanded allele. We aimed to determine the effect of the G130V (murine G127V) mutation on Fxn expression and to define its multi-system impact in vivo. We used CRISPR/Cas9 to introduce the G127V missense mutation in the Fxn coding sequence and generated homozygous mice (FxnG127V/G127V). We also introduced the G127V mutation into a GAA repeat expansion FRDA mouse model (FxnGAA230/KO; KIKO) to generate a compound heterozygous strain (FxnG127V/GAA230). We performed neurobehavioral tests on cohorts of WT and Fxn mutant animals at three-month intervals for one year, and collected tissue samples to analyze molecular changes during that time. The endogenous Fxn G127V protein is detected at much lower levels in all tissues analyzed from FxnG127V/G127V mice compared to age and sex-matched WT mice without differences in Fxn transcript levels. FxnG127V/G127V mice are significantly smaller than WT counterparts, but perform similarly in most neurobehavioral tasks. RNA sequencing analysis revealed reduced expression of genes in oxidative phosphorylation and protein synthesis, underscoring the metabolic consequences in our mouse model expressing extremely low levels of Fxn. Results of these studies provide insight into the unique pathogenic mechanism of the FXN G130V mechanism and the tolerable limit of Fxn/FXN expression in vivo.
•Extremely low Fxn levels are detected in CNS and heart tissues of Fxn G127V mice.•Fxn G127V mice are small and develop a hunched posture.•Endurance and activity are reduced in adult Fxn G127V mice.•Motor coordination is not affected in adult Fxn G127V mice.
Nuclear factor erythroid 2 related factor 2 (Nrf2) signaling maintains the redox homeostasis and its activation is shown to suppress cardiac maladaptation. Earlier we reported that acute endurance ...exercise (2 days) evoked antioxidant cytoprotection in young WT animals but not in aged WT animals. However, the effect of repeated endurance exercise during biologic aging (WT) characterized by an inherent deterioration in Nrf2 signaling and pathological aging (pronounced oxidative susceptibility-Nrf2 absence) in the myocardium remains elusive. Thus, the purpose of our study was to determine the effect of chronic endurance exercise-induced cardiac adaptation in aged mice with and without Nrf2. Age-matched WT and Nrf2-null mice (Nrf2
) (>22 months) were subjected to 6 weeks chronic endurance exercise (25 meter/min, 12% grade). The myocardial redox status was assessed by expression of antioxidant defense genes and proteins along with immunochemical detection of DMPO-radical adduct, GSH-NEM, and total ubiquitination. Cardiac functions were assessed by echocardiography and electrocardiogram. At sedentary state, loss of Nrf2 resulted in significant downregulation of antioxidant gene expression (
, and
α) with decreased GSH-NEM immuno-fluorescence signals. While Nrf2
mice subjected to CEE showed an either similar or more pronounced reduction in the transcript levels of
, and
α in relation to WT littermates. In addition, the hearts of Nrf2
on CEE showed a substantial reduction in specific antioxidant proteins, G6PD and CAT along with decreased GSH, a pronounced increase in DMPO-adduct and the total ubiquitination levels. Further, CEE resulted in a significant upregulation of hypertrophy genes (
, and β
) (
< 0.05) in the Nrf2
hearts in relation to WT mice. Moreover, the aged Nrf2
mice exhibited a higher degree of cardiac remodeling in association with a significant decrease in fractional shortening, pronounced ST segment, and J wave elevation upon CEE compared to age-matched WT littermates. In conclusion, our findings indicate that while the aged WT and Nrf2 knockout animals both exhibit hypertrophy after CEE, the older Nrf2 knockouts showed ventricular remodeling coupled with profound cardiac functional abnormalities and diastolic dysfunction.
IntroductionMyocardial remodeling resulting in ventricular hypertrophy and dysfunction are either strongly correlated or causally linked to chronic redox imbalance. Nuclear erythroid-2 like factor-2 ...(Nrf2), a master transcriptional regulator plays a major role in cellular redox homeostasis. Here we investigated whether age-associated decline in Nrf2 is causal to stress induced cardiac remodeling and heart failure.HypothesisWe hypothesize that genetic ablation or age-dependent decline of Nrf2 impairs myocardial redox state and induces pathological ventricular hypertrophy upon resistance exercise (RE).Methods and resultsAge and sex matched WT and Nrf2-null mice (>22 months; n=6/gp.) were subjected to RE stress for 6 Weeks (25 meter/min, 12% grade) and cardiac functions were assessed by echocardiography and electrocardiogram. Biochemical, cellular and molecular indices related to redox and myocardial structural and functional remodeling were also analyzed.ResultsPre- and post- echocardiography analyses in response to RE indicated progressive cardiac hypertrophy in both WT and Nrf2-null mice on aging. Interestingly, the degree of cardiac remodeling was significantly higher in Nrf2-null when compared to WT mice. Dramatic increase in ejection fraction, cardiac output and R amplitude (V) suggesting that Nrf2 deficiency cause pathological remodeling. Transcriptional signatures of hypertrophy (Anf, Bnf and β-Mhc) were significantly (p<0.05) upregulated in Nrf2-null versus WT mice in response to RE. Under sedentary state, loss of Nrf2 was associated with significant downregulation of antioxidant genes (Nqo1, Ho1, Gclm, Cat and Gst-α). After RE, the transcript levels of Gclc, Nqo1, Gsr and Gst-α were profoundly diminished in Nrf2-null versus WT mice. Further, significant increase in DMPO-adduct coupled with the decreased myocardial glutathione levels illustrates oxidative stress in Nrf2-null mouse after RE. Further, increased accumulation of HNE-protein adducts and ubiquitinated proteins, known markers of proteasome inhibition were noted in Nrf2-null mice after RE.ConclusionsOur findings indicate that loss of Nrf2 could induce pathological ventricular remodeling and cardiac dysfunction over aging.
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