Leukocytes mediate disease pathogenesis in the Ndufs4(KO) mouse model of Leigh syndrome
Julia C. Stokes, Rebecca L. Bornstein, Katerina James, Kyung Yeon Park, Kira A. Spencer, Katie Vo, John C. Snell, Brittany M. Johnson, Philip G. Morgan, Margaret M. Sedensky, Nathan A. Baertsch, Simon C. Johnson
Julia C. Stokes, Rebecca L. Bornstein, Katerina James, Kyung Yeon Park, Kira A. Spencer, Katie Vo, John C. Snell, Brittany M. Johnson, Philip G. Morgan, Margaret M. Sedensky, Nathan A. Baertsch, Simon C. Johnson
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Research Article Inflammation Neuroscience

Leukocytes mediate disease pathogenesis in the Ndufs4(KO) mouse model of Leigh syndrome

Abstract

Symmetric, progressive, necrotizing lesions in the brainstem are a defining feature of Leigh syndrome (LS). A mechanistic understanding of the pathogenesis of these lesions has been elusive. Here, we report that leukocyte proliferation is causally involved in the pathogenesis of LS. Depleting leukocytes with a colony-stimulating factor 1 receptor inhibitor disrupted disease progression, including suppression of CNS lesion formation and a substantial extension of survival. Leukocyte depletion rescued diverse symptoms, including seizures, respiratory center function, hyperlactemia, and neurologic sequelae. These data reveal a mechanistic explanation for the beneficial effects of mTOR inhibition. More importantly, these findings dramatically alter our understanding of the pathogenesis of LS, demonstrating that immune involvement is causal in disease. This work has important implications for the mechanisms of mitochondrial disease and may lead to novel therapeutic strategies.

Authors

Julia C. Stokes, Rebecca L. Bornstein, Katerina James, Kyung Yeon Park, Kira A. Spencer, Katie Vo, John C. Snell, Brittany M. Johnson, Philip G. Morgan, Margaret M. Sedensky, Nathan A. Baertsch, Simon C. Johnson

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Figure 3

Leukocyte depletion prevents leukocyte/microglia accumulation and astrocytosis throughout the brain and rescues a range of systemic symptoms associated with LS in the Ndufs4(KO) mice.

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Leukocyte depletion prevents leukocyte/microglia accumulation and astroc...
(A–C) Iba1+ leukocytes/microglia (A) (see Discussion) and GFAP+ astrocytes (B) in cortex of control- and pexidartinib-treated control and Ndufs4(KO) (see Methods). Data points represent individual animals. n = 4 for untreated Ndufs4(KO), 3 for other groups. (C) Representative images of cortex. (D and E) Iba1+ leukocytes/microglia and GFAP+ astrocytes in brainstem regions outside overt lesions in control- and pexidartinib-treated control and Ndufs4(KO) mice (representative images in Figure 2). Data points represent individual animals. n = 4 for untreated Ndufs4(KO), 3 for other groups. *P < 0.05, **P < 0.005, ***P < 0.0005, and ****P < 0.0001, 2-way ANOVA with Tukey’s multiple-testing correction–adjusted P values for pairwise comparisons. (F) Rotarod-induced seizure frequency at P30 by treatment. *P < 0.016, Fisher’s exact test (Bonferroni-adjusted P value cutoff for significance = 0.0167). n indicated by bars. (G) Time to seizure in rotarod assay, P30. All data points shown. *P < 0.05, log-rank test. ns as in F. (H) Scatter plots of Ndufs4(KO) mouse weight as a function of age and treatment, with local regression (Lowess) curves to display population trends. (I) Cachexia onset (see Figure 1, Methods) in control- and pexidartinib-treated Ndufs4(KO) mice. Colors and ns as in G. *P < 0.0167, ****P < 0.00005, log-rank test (Bonferroni-corrected P value cutoff for significance = 0.0167). (J) Blood glucose by age in control- and pexidartinib-treated Ndufs4(KO) animals. Each point represents the median value for 1 animal during the period (data points are biological replicates/individual animals). ****P < 0.0001 by unpaired, unequal variances (Welch’s) t test (Bonferroni-corrected P value cutoff for significance = 0.0125). ns as indicated in bars. (K and L) Blood lactate in response to a glucose bolus (2 g/kg) in control and Ndufs4(KO) mice at predisease (P25) and early disease (P45). (K) Time course and (L) total AUC for blood lactate 0–90 minutes. *P < 0.008, 2-way ANOVA with Tukey’s multiple-testing correction–adjusted P values for pairwise comparisons. ns indicated in bars. (M–O) Blood lactate in response to glucose bolus (2 g/kg) in untreated control and Ndufs4(KO) mice and Ndufs4(KO) mice treated with pexidartinib (300 mg/kg/d in chow), IPI-549 (100 mg/kg/d in chow), or rapamycin (ABI-009 formulation, 8 mg/kg/d IP). Time course of blood lactate (M), baseline lactate (N), and total AUC for blood lactate (O) 0–90 minutes. *P < 0.0167, **P < 0.005, ****P < 0.0001 by unpaired, unequal variances (Welch’s) t test (Bonferroni-adjusted significance cutoff = 0.0167). ns indicated in bars (N and O). (P) Change in blood lactate in control and Ndufs4(KO) mice after 30-minute exposure to 0.4% isoflurane and impact of treatment with 300 mg/kg/d pexidartinib. n = 5, 7, and 3. **P < 0.005 by 1-way ANOVA with Tukey’s multiple-testing correction–adjusted P values for pairwise comparisons. (K and M) AUC, not individual time points, were compared. (Q) Minimum alveolar anesthetic concentration (MAC) of isoflurane associated with anesthesia in control- and pexidartinib-treated Ndufs4(KO) mice (see Methods). **P < 0.005 by unpaired, unequal variances (Welch’s) t test. n = 6/group. Data represent mean, error bars ± SEM, unless otherwise stated.