Short-Term Caloric Restriction and Subsequent Re-Feeding Compromise Liver Health and Associated Lipid Mediator Signaling in Aged Mice.

Aging is characterized by alterations in the inflammatory microenvironment, which is tightly regulated by a complex network of inflammatory mediators. Excessive calorie consumption contributes to age- and lifestyle-associated diseases like obesity, type 2 diabetes, cardiovascular disorders, and cancer, while limited nutrient availability may lead to systemic health-promoting adaptations. Geroprotective effects of short-term caloric restriction (CR) can beneficially regulate innate immune receptors and interferon signaling in the liver of aged mice, but how CR impacts the hepatic release of immunomodulatory mediators like cytokines and lipid mediators (LM) is elusive. Here, we investigated the impact of aging on the inflammatory microenvironment in the liver and its linkage to calorie consumption. The livers of female young and aged C57BL/6JRj mice, as well as of aged mice after caloric restriction (CR) up to 28 days, with and without subsequent re-feeding (2 days), were evaluated. Surprisingly, despite differences in the hepatic proteome of young and old mice, aging did not promote a pro-inflammatory environment in the liver, but it reduced lipoxygenase-mediated formation of LM from polyunsaturated fatty acids without affecting the expression of the involved lipoxygenases and related oxygenases. Moreover, CR failed to ameliorate the secretion of pro-inflammatory cytokines but shifted the LM production to the formation of monohydroxylated LM with inflammation-resolving features. Unexpectedly, re-feeding after CR even further decreased the inflammatory response as LM species were markedly downregulated. Our findings raise the question of how short-term CR is indeed beneficial as a nutritional intervention for healthy elderly subjects and further stress the necessity to address tissue-specific inflammatory states.

Side-by-side comparison of recombinant human glutathione peroxidases identifies overlapping substrate specificities for soluble hydroperoxides.

Five out of eight human glutathione peroxidases (GPXs) are selenoproteins, representing proteins that contain selenium as part of the amino acid selenocysteine. The GPXs are important for reducing hydroperoxides in a glutathione-consuming manner and thus regulate cellular redox homeostasis. GPX1, GPX2, and GPX4 represent the three main cytosolic GPXs, but they differ in their expression patterns with GPX1 and GPX4 being expressed ubiquitously, whereas GPX2 is mainly expressed in epithelial cells. GPX1 and GPX2 have been described to reduce soluble hydroperoxides, while GPX4 reduces complex lipid hydroperoxides, thus protecting cells from lipid peroxidation and ferroptosis. But most of these data are derived from cells that are devoid of one of the isoforms and thus, compensation or other cellular effects might affect the conclusions. So far, the use of isolated recombinant human selenoprotein glutathione peroxidases in pure enzyme assays has not been employed to study their substrate specificities side by side. Using recombinant GPX1, GPX2, and GPX4 produced in E. coli we here assessed their GPX activities by a NADPH-consuming glutathione reductase-coupled assay with 17 different peroxides (all at 50 µM) as substrates. GPX4 was clearly the only isoform able to reduce phosphatidylcholine hydroperoxide. In contrast, small soluble hydroperoxides such as H2O2, cumene hydroperoxide, and tert-butyl hydroperoxide were reduced by all three isoforms, but with approximately 10-fold higher efficiency for GPX1 in comparison to GPX2 and GPX4. Also, several fatty acid-derived hydroperoxides were reduced by all three isoforms and again GPX1 had the highest activity. Interestingly, the stereoisomerism of the fatty acid-derived hydroperoxides clearly affected the activity of the GPX enzymes. Overall, distinct substrate specificity is obvious for GPX4, but not so when comparing GPX1 and GPX2. Clearly GPX1 was the most potent isoform of the three GPXs in terms of turnover in reduction of soluble and fatty-acid derived hydroperoxides.