👤 Kari J Buck

Despite Alzheimer's disease (AD) disproportionately affecting women, the mechanisms remain elusive. In AD, microglia undergo 'metabolic reprogramming', which contributes to microglial dysfunction and AD pathology. However, how sex and age contribute to metabolic reprogramming in microglia is understudied. Here, we use metabolic imaging, transcriptomics, and metabolic assays to probe age- and sex-associated changes in brain and microglial metabolism. Glycolytic and oxidative metabolism in the whole brain was determined using Fluorescence Lifetime Imaging Microscopy (FLIM). Young female brains appeared less glycolytic than male brains, but with aging, the female brain became 'male-like.' Transcriptomic analysis revealed increased expression of disease-associated microglia (DAM) genes (e.g., ApoE, Trem2, LPL), and genes involved in glycolysis and oxidative metabolism in microglia from aged females compared to males. To determine whether estrogen can alter the expression of these genes, BV-2 microglia-like cell lines, which abundantly express DAM genes, were supplemented with 17β-estradiol (E2). E2 supplementation resulted in reduced expression of DAM genes, reduced lipid and cholesterol transport, and substrate-dependent changes in glycolysis and oxidative metabolism. Consistent with the notion that E2 may suppress DAM-associated factors, LPL activity was elevated in the brains of aged female mice. Similarly, DAM gene and protein expression was higher in monocyte-derived microglia-like (MDMi) cells derived from middle-aged females compared to age-matched males and was responsive to E2 supplementation. FLIM analysis of MDMi from young and middle-aged females revealed reduced oxidative metabolism and FAD+ with age. Overall, our findings show that altered metabolism defines age-associated changes in female microglia and suggest that estrogen may inhibit the expression and activity of DAM-associated factors, which may contribute to increased AD risk, especially in post-menopausal women. Show less

Despite Alzheimer's disease (AD) disproportionately affecting women, the mechanisms remain elusive. In AD, microglia undergo 'metabolic reprogramming', which contributes to microglial dysfunction and AD pathology. However, how sex and age contribute to metabolic reprogramming in microglia is understudied. Here, we use metabolic imaging, transcriptomics, and metabolic assays to probe age-and sex-associated changes in brain and microglial metabolism. Glycolytic and oxidative metabolism in the whole brain was determined using Fluorescence Lifetime Imaging Microscopy (FLIM). Young female brains appeared less glycolytic than male brains, but with aging, the female brain became 'male-like.' Transcriptomic analysis revealed increased expression of disease-associated microglia (DAM) genes (e.g., Show less

Genetic factors significantly affect alcohol consumption and vulnerability to withdrawal. Furthermore, some genetic models showing predisposition to severe withdrawal are also predisposed to low ethanol (EtOH) consumption and vice versa, even when tested independently in naïve animals. Beginning with a C57BL/6J × DBA/2J F2 intercross founder population, animals were simultaneously selectively bred for both high alcohol consumption and low acute withdrawal (SOT line), or vice versa (NOT line). Using randomly chosen fourth selected generation (S4) mice (N = 18-22/sex/line), RNA-Seq was employed to assess genome-wide gene expression in ventral striatum. The MegaMUGA array was used to detect genome-wide genotypic differences. Differential gene expression and the weighted gene co-expression network analysis were implemented as described elsewhere (Genes Brain Behav 16, 2017, 462). The new selection of the SOT and NOT lines was similar to that reported previously (Alcohol Clin Exp Res 38, 2014, 2915). One thousand eight hundred and sixteen transcripts were detected as differentially expressed between the lines. For genes more highly expressed in the SOT line, there was enrichment in genes associated with cell adhesion, synapse organization, and postsynaptic membrane. The genes with a cell adhesion annotation included 23 protocadherins, Mpdz and Dlg2. Genes with a postsynaptic membrane annotation included Gabrb3, Gphn, Grid1, Grin2b, Grin2c, and Grm3. The genes more highly expressed in the NOT line were enriched in a network module (red) with annotations associated with mitochondrial function. Several of these genes were module hub nodes, and these included Nedd8, Guk1, Elof1, Ndufa8, and Atp6v1f. Marked effects of selection on gene expression were detected. The NOT line was characterized by higher expression of hub nodes associated with mitochondrial function. Genes more highly expressed in the SOT aligned with previous findings, for example, Colville and colleagues (Genes Brain Behav 16, 2017, 462) that both high EtOH preference and consumption are associated with effects on cell adhesion and glutamate synaptic plasticity. Show less

Cardiac myosin-binding protein C (cMyBP-C) regulates actin-myosin interaction and thereby cardiac myocyte contraction and relaxation. This physiologic function is regulated by cMyBP-C phosphorylation. In our study, reduced site-specific cMyBP-C phosphorylation coincided with increased S-glutathiolation in ventricular tissue from patients with dilated or ischemic cardiomyopathy compared to nonfailing donors. We used redox proteomics, to identify constitutive and disease-specific S-glutathiolation sites in cMyBP-C in donor and patient samples, respectively. Among those, a cysteine cluster in the vicinity of the regulatory phosphorylation sites within the myosin S2 interaction domain C1-M-C2 was identified and showed enhanced S-glutathiolation in patients. In vitro S-glutathiolation of recombinant cMyBP-C C1-M-C2 occurred predominantly at Cys(249), which attenuated phosphorylation by protein kinases. Exposure to glutathione disulfide induced cMyBP-C S-glutathiolation, which functionally decelerated the kinetics of Ca(2+)-activated force development in ventricular myocytes from wild-type, but not those from Mybpc3-targeted knockout mice. These oxidation events abrogate protein kinase-mediated phosphorylation of cMyBP-C and therefore potentially contribute to the reduction of its phosphorylation and the contractile dysfunction observed in human heart failure.-Stathopoulou, K., Wittig, I., Heidler, J., Piasecki, A., Richter, F., Diering, S., van der Velden, J., Buck, F., Donzelli, S., Schröder, E., Wijnker, P. J. M., Voigt, N., Dobrev, D., Sadayappan, S., Eschenhagen, T., Carrier, L., Eaton, P., Cuello, F. S-glutathiolation impairs phosphoregulation and function of cardiac myosin-binding protein C in human heart failure. Show less