Muscle mitochondrial partial pressure of oxygen (PmitoO2) is a key determinant of skeletal muscle oxygen consumption (V̇O2), metabolism, gene regulation, and adaptation in health and disease. Yet PmitoO2 remains difficult to measure directly in vivo in humans during exercise. Myoglobin-associated PO2 (PMbO2), measured non-invasively using proton magnetic resonance spectroscopy, may provide a measure of PmitoO2 during maximal exercise in humans because Mb is coupled to mitochondrial oxygen availability and PMbO2 falls in the low PO2 range expected for PmitoO2 in vivo. However, it remains unknown whether PMbO2 during maximal exercise varies systematically with muscle V̇O2max, as would be expected if PMbO2 reflects PmitoO2 in vivo. We analyzed data from five initially sedentary males before and after 8 weeks of single-leg knee-extensor exercise (KE) training. During maximal KE under 0.12, 0.21, and 1.00 fractions of inspired oxygen (FIO2), we measured PMbO2 using myoglobin desaturation measured by proton magnetic resonance spectroscopy and muscle V̇O2max from leg blood flow and the arterial-femoral venous O2 content difference. Across FIO2 conditions, muscle V̇O2max varied systematically with PMbO2 both before and after training, consistent with O2-dependent mitochondrial respiration. When interpreted within a canonical hyperbolic framework, the V̇O2-PMbO2 relationship yielded low apparent mitochondrial P50 values that were broadly consistent with values reported for human skeletal muscle isolated mitochondria studied in vitro. A complementary linear P50 analysis that did not assume a hyperbolic relationship supported the same interpretation. Together, these findings provide data-supported, proof-of-concept evidence that PMbO2 reflects in vivo PmitoO2 during maximal exercise in humans.
The pro-inflammatory cytokine tumor necrosis factor alpha (TNFα) mediates airway responses to acute inflammation. Previously we demonstrated that, in human airway smooth muscle (hASM) cells, TNFα increases reactive oxygen species (ROS) formation. TNFα also selectively activates the inositol-requiring enzyme 1α (pIRE1αS724 autophosphorylation) endoplasmic reticulum (ER) stress pathway involving splicing of X-box binding protein 1 (XBP1s) and transcriptionally activates cyclin-dependent kinases 1 and 5 (CDK1 and CDK5), promoting dynamin-related protein 1 (DRP1) phosphorylation at serine 616 (pDRP1S616) and mitochondrial fragmentation. In the present study, we hypothesized that in hASM, TNFα-induced ROS triggers pIRE1αS724/XBP1s ER stress pathway. To test this hypothesis, we examined the impact of the ROS scavenger Tempol on TNFα-induced pIRE1αS724/XBP1s ER stress pathway and downstream signaling mediating mitochondrial fragmentation. Bronchiolar tissue samples were obtained from 6 patients with no history of smoking or chronic pulmonary disease. The smooth muscle layer was dissected, and hASM cells were dissociated and randomly assigned to four treatment groups: 1) Vehicle, 2) Vehicle + TNFα (20 ng/mL, 6 h), 3) Tempol (500 μM) only, and 4) Tempol (500 μM) + TNFα (20 ng/mL, 6 h). ROS formation was determined by confocal imaging using MitoSOX™ Red. Mitochondria were labeled with MitoTracker Red and imaged using confocal microscopy. Using Western blot, we demonstrated that Tempol reduced cellular ROS formation and mitigated the TNFα-induced increase in pIRE1αS724, XBP1s, CDK1/5, pDRP1S616 protein levels and reduced mitochondrial fragmentation. These findings support our hypothesis and indicate a role of ROS in mediating TNFα-induced ER stress and mitochondrial fragmentation.
Circadian rhythms are endogenous ∼24-h cycles that regulate cardiovascular physiology. Although circadian regulation of the left ventricle (LV) is well established, whether the right ventricle (RV) exhibits intrinsic rhythmicity is not known. Here, we provide the first evidence that the healthy RV exhibits robust rhythms in function and molecular gene expression. Cardiovascular disease is the leading cause of mortality in type 1 diabetes (T1D). T1D disrupts circadian rhythms, yet how T1D alters chamber-specific circadian control remains unclear. We investigated RV and LV function and gene expression across the 24-h light-dark cycle in male streptozotocin-induced T1D and control mice. Echocardiographic assessment of diabetes-induced remodeling at ZT0-4 and ZT12-16 [zeitgeber time (ZT)] revealed time-dependent functional impairment, including reversal of the normal diurnal heart rate pattern. T1D impaired day-night RV and LV systolic function, with loss of day-night difference in LV ejection fraction and impaired active period RV stroke volume. Cosinor analysis of RV and LV gene expression demonstrated preserved 24-h rhythmicity of core clock genes Arntl1 and Per2 in both ventricles with T1D, whereas the clock output genes Nr1d1 and Dbp had dampened amplitude in the RV but amplified expression in the T1D LV. These findings demonstrate that T1D differentially disrupts circadian regulation of the RV and LV, with selective vulnerability of the clock-controlled output genes despite preservation of the core oscillator. Chamber-specific circadian remodeling may contribute to the elevated cardiovascular risk in T1D and has implications for the timing of diagnostic and therapeutic interventions.NEW & NOTEWORTHY This study provides the first evidence that the healthy right ventricle (RV), like the left ventricle (LV), possesses an intrinsic circadian clock with robust 24-h rhythmicity. Using a mouse model of type 1 diabetes, we show that diabetes does not uniformly suppress cardiac clocks but instead differentially rewires RV and LV rhythms, dampening clock-output gene amplitude in the RV while amplifying it in the LV. Together, we reveal chamber-specific circadian vulnerability with implications for diabetic cardiomyopathy and chronotherapy.
