Exercise training modifies skeletal muscle clock gene expression but not 24-hour rhythmicity in substrate metabolism of men with insulin resistance.

Twenty-four hour rhythmicity in whole-body substrate metabolism, skeletal muscle clock gene expression and mitochondrial respiration is compromised upon insulin resistance. With exercise training known to ameliorate insulin resistance, our objective was to test if exercise training can reinforce diurnal variation in whole-body and skeletal muscle metabolism in men with insulin resistance. In a single-arm longitudinal design, 10 overweight and obese men with insulin resistance performed 12 weeks of high-intensity interval training recurrently in the afternoon (between 14.00 and 18.00 h) and were tested pre- and post-exercise training, while staying in a metabolic research unit for 2 days under free-living conditions with regular meals. On the second days, indirect calorimetry was performed at 08.00, 13.00, 18.00, 23.00 and 04.00 h, muscle biopsies were taken from the vastus lateralis at 08.30, 13.30 and 23.30 h, and blood was drawn at least bi-hourly over 24 h. Participants did not lose body weight over 12 weeks, but improved body composition and exercise capacity. Exercise training resulted in reduced 24-h plasma glucose levels, but did not modify free fatty acid and triacylglycerol levels. Diurnal variation of muscle clock gene expression was modified by exercise training with period genes showing an interaction (time × exercise) effect and reduced mRNA levels at 13.00 h. Exercise training increased mitochondrial respiration without inducing diurnal variation. Twenty-four-hour substrate metabolism and energy expenditure remained unchanged. Future studies should investigate alternative exercise strategies or types of interventions (e.g. diet or drugs aiming at improving insulin sensitivity) for their capacity to reinforce diurnal variation in substrate metabolism and mitochondrial respiration. KEY POINTS: Insulin resistance is associated with blunted 24-h flexibility in whole-body substrate metabolism and skeletal muscle mitochondrial respiration, and disruptions in the skeletal muscle molecular circadian clock. We hypothesized that exercise training modifies 24-h rhythmicity in whole-body substrate metabolism and diurnal variation in skeletal muscle molecular clock and mitochondrial respiration in men with insulin resistance. We found that metabolic inflexibility over 24 h persisted after exercise training, whereas mitochondrial respiration increased independent of time of day. Gene expression of Per1-3 and Rorα in skeletal muscle changed particularly close to the time of day at which exercise training was performed. These results provide the rationale to further investigate the differential metabolic impact of differently timed exercise to treat metabolic defects of insulin resistance that manifest at a particular time of day.

Extracellular Granzyme K Modulates Angiogenesis by Regulating Soluble VEGFR1 Release From Endothelial Cells.

Angiogenesis is crucial for normal development and homeostasis, but also plays a role in many diseases including cardiovascular diseases, autoimmune diseases, and cancer. Granzymes are serine proteases stored in the granules of cytotoxic cells, and have predominantly been studied for their pro-apoptotic role upon delivery in target cells. A growing body of evidence is emerging that granzymes also display extracellular functions, which largely remain unknown. In the present study, we show that extracellular granzyme K (GrK) inhibits angiogenesis and triggers endothelial cells to release soluble VEGFR1 (sVEGFR1), a decoy receptor that inhibits angiogenesis by sequestering VEGF-A. GrK does not cleave off membrane-bound VEGFR1 from the cell surface, does not release potential sVEGFR1 storage pools from endothelial cells, and does not trigger sVEGFR1 release via protease activating receptor-1 (PAR-1) activation. GrK induces de novo sVEGFR1 mRNA and protein expression and subsequent release of sVEGFR1 from endothelial cells. GrK protein is detectable in human colorectal tumor tissue and its levels positively correlate with sVEGFR1 protein levels and negatively correlate with T4 intratumoral angiogenesis and tumor size. In conclusion, extracellular GrK can inhibit angiogenesis via secretion of sVEGFR1 from endothelial cells, thereby sequestering VEGF-A and impairing VEGFR signaling. Our observation that GrK positively correlates with sVEGFR1 and negatively correlates with angiogenesis in colorectal cancer, suggest that the GrK-sVEGFR1-angiogenesis axis may be a valid target for development of novel anti-angiogenic therapies in cancer.