GTP-cyclohydrolase deficiency induced peripheral and deep microcirculation dysfunction with age.

The present study assessed the impact of impaired tetrahydrobiopterin (BH4) production on vasoreactivity from conduit and small arteries along the vascular tree as seen during aging. For this purpose, the mutant hyperphenylalaninemic mouse (hph-1) was used. This model is reported to be deficient in GTP cyclohydrolase I, a rate limiting enzyme in BH4 biosynthesis. BH4 is a key regulator of vascular homeostasis by regulating the nitric oxide synthase 3 (NOS3) activity. In GTP-CH deficient mice, the aortic BH4 levels were decreased, by -77% in 12 week-middle-aged mice (young) and by -83% in 35-45 week-middle-aged mice (middle-aged). In young hph-1, the mesenteric artery ability to respond to flow was slightly reduced by 9%. Aging induced huge modification in many vascular functions. In middle-aged hph-1, we observed a decrease in aortic cGMP levels, biomarker of NO availability (-46%), in flow-mediated vasodilation of mesenteric artery (-31%), in coronary hyperemia response measured in isolated heart following transient ischemia (-27%) and in cutaneous microcirculation dilation in response to acetylcholine assessed in vivo by laser-doppler technic (-69%). In parallel, the endothelium-dependent relaxation in response to acetylcholine in conduit blood vessel, measured on isolated aorta rings, was unchanged in hph-1 mice whatever the age. Our findings demonstrate that in middle-aged GTP-CH depleted mice, the reduction of BH4 was characterized by an alteration of microcirculation dilatory properties observed in various parts of the vascular tree. Large conduit blood vessels vasoreactivity, ie aorta, was unaltered even in middle-aged mice emphasizing the main BH4-deletion impact on the microcirculation.

hsa-miR-548v controls the viscoelastic properties of human cardiomyocytes and improves their relaxation rates.

The impairment of left ventricular (LV) diastolic function with an inadequate increase in myocardial relaxation velocity directly results in lower LV compliance, increased LV filling pressures, and heart failure symptoms. The development of agents facilitating the relaxation of human cardiomyocytes requires a better understanding of the underlying regulatory mechanisms. We performed a high-content microscopy-based screening in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) using a library of 2,565 human miRNA mimics and measured relaxation kinetics via high-computing analyses of motion movies. We identified hsa-miR-548v, a primate-specific miRNA, as the miRNA producing the largest increase in relaxation velocities. This positive lusitropic effect was reproduced in engineered cardiac tissues generated with healthy and BRAF T599R mutant hiPSC-CMs and was independent of changes in calcium transients. Consistent with improvements in viscoelastic responses to mechanical stretch, RNA-Seq showed that hsa-miR-548v downregulated multiple targets, especially components of the mechanosensing machinery. The exogenous administration of hsa-miR-548v in hiPSC-CMs notably resulted in a significant reduction of ANKRD1/CARP1 expression and localization at the sarcomeric I-band. This study suggests that the sarcomere I-band is a critical control center regulating the ability of cardiomyocytes to relax and is a target for improving relaxation and diastolic dysfunction.

Aging increases circulating BH2 without modifying BH4 levels and impairs peripheral vascular function in healthy adults.

Little is known about the mechanisms of aging on vascular beds and its relationship with tetra and di-hydrobiopterin (BH4 and BH2) levels. This observational clinical study analyzed the impact of aging on plasma and platelet biopterins, cutaneous blood flow (CBF), and coronary flow reserve (CFR) in healthy adults. The study enrolled healthy adults in 3 age groups: 18-30, 50-59, and 60-70 years (n = 25/group). Biopterins were assessed by LC-MS/MS using newly defined pre-analytical conditions limiting BH4 oxidation and improving long-term stability. CBF was measured by Laser Speckle Contrast Imaging coupled with acetylcholine-iontophoresis and CFR by adenosine stress cardiac magnetic resonance. In healthy adults, aging (60-70 years vs 18-30 years) significantly increased platelet BH2 (+75%, P = 0.033) and BH2 + BH4 (+31%, P = 0.033), and to a lesser extent plasma BH2 (+29%, P = 0.009) without affecting BH4 and BH4/BH2. Simultaneously, CBF was decreased (-23%, P = 0.004) but not CFR, CBF being inversely correlated with platelet BH2 (r = -0.42, P = 0.001) and BH2 + BH4 (r = -0.41, P = 0.002). The proportion of adults with abnormal platelet BH2 increased with age (+28% in 60-70y). These abnormal BH2 levels were significantly associated with reduced CBF and CFR (-16%, P = 0.03 and -26%, P = 0.02). In conclusion, our study showed that age-related peripheral endothelial dysfunction was associated with an increase in circulating BH2 without decreasing BH4, the effect being more marked in platelets, the most relevant blood compartment to assess biopterin bioavailability. Peripheral but not coronary vascular function is progressively impaired with aging in healthy adults. All these findings support biopterins as therapeutic targets to improve vascular function.

Pore formation induced by an antimicrobial peptide: electrostatic effects.

We investigate the mode of action of Cateslytin, an antimicrobial peptide, on zwitterionic biomembranes by performing numerical simulations and electrophysiological measurements on membrane vesicles. Using this natural beta-sheet antimicrobial peptide secreted during stress as a model we show that a single peptide is able to form a stable membrane pore of 1 nm diameter of 0.25 nS conductance found both from calculation and electrical measurements. The resulting structure does not resemble the barrel-stave or carpet models earlier predicted, but is very close to that found in the simulation of alpha-helical peptides. Based on the simulation of a mutated peptide and the effects of small external electric fields, we conclude that electrostatic forces play a crucial role in the process of pore formation.