Prohibitin-1 Is a Dynamically Regulated Blood Protein With Cardioprotective Effects in Sepsis.

Background In sepsis, circulating cytokines and lipopolysaccharide elicit mitochondrial dysfunction and cardiomyopathy, a major cause of morbidity and mortality with this condition. Emerging research places the PHB1 (lipid raft protein prohibitin-1) at the nexus of inflammation, metabolism, and oxidative stress. PHB1 has also been reported in circulation, though its function in this compartment is completely unknown. Methods and Results Using a wide-ranging approach across multiple in vitro and in vivo models, we interrogated the functional role of intracellular and circulating PHB1 in the heart during sepsis, and elucidated some of the mechanisms involved. Upon endotoxin challenge or sepsis induction in rodent models, PHB1 translocates from mitochondria to nucleus in cardiomyocytes and is secreted into the circulation from the liver in a manner dependent on nuclear factor (erythroid-derived 2)-like 2, a key transcriptional regulator of the antioxidant response. Overexpression or treatment with recombinant human PHB1 enhances the antioxidant/anti-inflammatory response and protects HL-1 cardiomyocytes from mitochondrial dysfunction and toxicity from cytokine stress. Importantly, administration of recombinant human PHB1 blunted inflammation and restored cardiac contractility and ATP production in mice following lipopolysaccharide challenge. This cardioprotective, anti-inflammatory effect of recombinant human PHB1 was determined to be independent of nuclear factor (erythroid-derived 2)-like 2, but partially dependent on PI3K/AKT  signaling in the heart. Conclusions These findings reveal a previously unknown cardioprotective effect of PHB1 during sepsis, and illustrate a pro-survival, protective role for PHB1 in the circulation. Exploitation of circulating PHB1 as a biomarker and/or therapeutic could have widespread benefit in the clinical management of sepsis and other severe inflammatory disorders.

Estrogen receptor-α in female skeletal muscle is not required for regulation of muscle insulin sensitivity and mitochondrial regulation.

OBJECTIVE: Estrogen receptor-α (ERα) is a nuclear receptor family member thought to substantially contribute to the metabolic regulation of skeletal muscle. However, previous mouse models utilized to assess the necessity of ERα signaling in skeletal muscle were confounded by altered developmental programming and/or influenced by secondary effects, making it difficult to assign a causal role for ERα. The objective of this study was to determine the role of skeletal muscle ERα in regulating metabolism in the absence of confounding factors of development. METHODS: A novel mouse model was developed allowing for induced deletion of ERα in adult female skeletal muscle (ERαKOism). ERαshRNA was also used to knockdown ERα (ERαKD) in human myotubes cultured from primary human skeletal muscle cells isolated from muscle biopsies from healthy and obese insulin-resistant women. RESULTS: Twelve weeks of HFD exposure had no differential effects on body composition, VO2, VCO2, RER, energy expenditure, and activity counts across genotypes. Although ERαKOism mice exhibited greater glucose intolerance than wild-type (WT) mice after chronic HFD, ex vivo skeletal muscle glucose uptake was not impaired in the ERαKOism mice. Expression of pro-inflammatory genes was altered in the skeletal muscle of the ERαKOism, but the concentrations of these inflammatory markers in the systemic circulation were either lower or remained similar to the WT mice. Finally, skeletal muscle mitochondrial respiratory capacity, oxidative phosphorylation efficiency, and H2O2 emission potential was not affected in the ERαKOism mice. ERαKD in human skeletal muscle cells neither altered differentiation capacity nor caused severe deficits in mitochondrial respiratory capacity. CONCLUSIONS: Collectively, these results suggest that ERα function is superfluous in protecting against HFD-induced skeletal muscle metabolic derangements after postnatal development is complete.

Tissue-specific characterization of mitochondrial branched-chain keto acid oxidation using a multiplexed assay platform.

Alterations to branched-chain keto acid (BCKA) oxidation have been implicated in a wide variety of human diseases, ranging from diabetes to cancer. Although global shifts in BCKA metabolism-evident by gene transcription, metabolite profiling, and in vivo flux analyses have been documented across various pathological conditions, the underlying biochemical mechanism(s) within the mitochondrion remain largely unknown. In vitro experiments using isolated mitochondria represent a powerful biochemical tool for elucidating the role of the mitochondrion in driving disease. Such analyses have routinely been utilized across disciplines to shed valuable insight into mitochondrial-linked pathologies. That said, few studies have attempted to model in vitro BCKA oxidation in isolated organelles. The impetus for the present study stemmed from the knowledge that complete oxidation of each of the three BCKAs involves a reaction dependent upon bicarbonate and ATP, both of which are not typically included in respiration experiments. Based on this, it was hypothesized that the inclusion of exogenous bicarbonate and stimulation of respiration using physiological shifts in ATP-free energy, rather than excess ADP, would allow for maximal BCKA-supported respiratory flux in isolated mitochondria. This hypothesis was confirmed in mitochondria from several mouse tissues, including heart, liver and skeletal muscle. What follows is a thorough characterization and validation of a novel biochemical tool for investigating BCKA metabolism in isolated mitochondria.

Specialized Pro-Resolving Lipid Mediators Regulate Ozone-Induced Pulmonary and Systemic Inflammation.

Exposure to ozone (O3) induces lung injury, pulmonary inflammation, and alters lipid metabolism. During tissue inflammation, specialized pro-resolving lipid mediators (SPMs) facilitate the resolution of inflammation. SPMs regulate the pulmonary immune response during infection and allergic asthma; however, the role of SPMs in O3-induced pulmonary injury and inflammation is unknown. We hypothesize that O3 exposure induces pulmonary inflammation by reducing SPMs. To evaluate this, male C57Bl/6J mice were exposed to filtered air (FA) or 1 ppm O3 for 3 h and necropsied 24 h after exposure. Pulmonary injury/inflammation was determined by bronchoalveolar lavage (BAL) differentials, protein, and lung tissue cytokine expression. SPMs were quantified by liquid chromatography tandem mass spectrometry and SPM receptors leukotriene B4 receptor 1 (BLT-1), formyl peptide receptor 2 (ALX/FPR2), chemokine-like receptor 1 (ChemR23), and SPM-generating enzyme (5-LOX and 12/15-LOX) expression were measured by real time PCR. 24 h post-O3 exposure, BAL PMNs and protein content were significantly increased compared to FA controls. O3-induced lung inflammation was associated with significant decreases in pulmonary SPM precursors (14-HDHA, 17-HDHA), the SPM PDX, and in pulmonary ALX/FPR2, ChemR23, and 12/15-LOX expression. Exogenous administration of 14-HDHA, 17-HDHA, and PDX 1 h prior to O3 exposure rescued pulmonary SPM precursors/SPMs, decreased proinflammatory cytokine and chemokine expression, and decreased BAL macrophages and PMNs. Taken together, these data indicate that O3-mediated SPM reductions may drive O3-induced pulmonary inflammation.