Age-related loss of muscle mass and force (sarcopenia) contributes to disability and increased mortality. Ryanodine receptor 1 (RyR1) is the skeletal muscle sarcoplasmic reticulum calcium release ...channel required for muscle contraction. RyR1 from aged (24 months) rodents was oxidized, cysteine-nitrosylated, and depleted of the channel-stabilizing subunit calstabin1, compared to RyR1 from younger (3–6 months) adults. This RyR1 channel complex remodeling resulted in “leaky” channels with increased open probability, leading to intracellular calcium leak in skeletal muscle. Similarly, 6-month-old mice harboring leaky RyR1-S2844D mutant channels exhibited skeletal muscle defects comparable to 24-month-old wild-type mice. Treating aged mice with S107 stabilized binding of calstabin1 to RyR1, reduced intracellular calcium leak, decreased reactive oxygen species (ROS), and enhanced tetanic Ca
2+ release, muscle-specific force, and exercise capacity. Taken together, these data indicate that leaky RyR1 contributes to age-related loss of muscle function.
► RyR1 in skeletal muscle is oxidized, nitrosylated, and depleted of calstabin1 with age ► Maladaptation of the RyR1 channel leads to SR Ca
2+ leak and muscle weakness ► Vicious cycle: Ca
2+ leak raises mitochondrial ROS, which oxidizes RyR1-enhancing leak ► The RyR1-stabilizing drug S107 fixes Ca
2+ leak and improves exercise capacity in aging
The type 2 ryanodine receptor/calcium release channel (RyR2), required for excitation-contraction coupling in the heart, is abundant in the brain. Chronic stress induces catecholamine biosynthesis ...and release, stimulating β-adrenergic receptors and activating cAMP signaling pathways in neurons. In a murine chronic restraint stress model, neuronal RyR2 were phosphorylated by protein kinase A (PKA), oxidized, and nitrosylated, resulting in depletion of the stabilizing subunit calstabin2 (FKBP12.6) from the channel complex and intracellular calcium leak. Stress-induced cognitive dysfunction, including deficits in learning and memory, and reduced long-term potentiation (LTP) at the hippocampal CA3-CA1 connection were rescued by oral administration of S107, a compound developed in our laboratory that stabilizes RyR2-calstabin2 interaction, or by genetic ablation of the RyR2 PKA phosphorylation site at serine 2808. Thus, neuronal RyR2 remodeling contributes to stress-induced cognitive dysfunction. Leaky RyR2 could be a therapeutic target for treatment of stress-induced cognitive dysfunction.
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► Leaky hippocampal RyR2 channels contribute to stress-induced cognitive dysfunction ► RyR2 PKA hyperphosphorylation and calstabin2 depletion cause intracellular Ca2+ leak ► Pharmacologic or genetic inhibition of Ca2+ leak prevent the cognitive dysfunction
Stabilizing a leaky calcium release channel with a small molecule alleviates the detrimental effects of long-term stress on learning and memory.
The regulation of intracellular calcium (Ca
) homeostasis is fundamental to maintain normal functions in many cell types. The ryanodine receptor (RyR), the largest intracellular calcium release ...channel located on the sarco/endoplasmic reticulum (SR/ER), plays a key role in the intracellular Ca
handling. Abnormal type 2 ryanodine receptor (RyR2) function, associated to mutations (ryanopathies) or pathological remodeling, has been reported, not only in cardiac diseases, but also in neuronal and pancreatic disorders. While animal models and in vitro studies provided valuable contributions to our knowledge on RyR2 dysfunctions, the human cell models derived from patients' cells offer new hope for improving our understanding of human clinical diseases and enrich the development of great medical advances. We here discuss the current knowledge on RyR2 dysfunctions associated with mutations and post-translational remodeling. We then reviewed the novel human cellular technologies allowing the correlation of patient's genome with their cellular environment and providing approaches for personalized RyR-targeted therapeutics.
Human-induced pluripotent stem cell (hiPSC)-derived cardiomyocytes raise the possibility of generating pluripotent stem cells from a wide range of human diseases. In the cardiology field, hiPSCs have ...been used to address the mechanistic bases of primary arrhythmias and in investigations of drug safety. These studies have been focused primarily on atrial and ventricular pathologies. Consequently, many hiPSC-based cardiac differentiation protocols have been developed to differentiate between atrial- or ventricular-like cardiomyocytes. Few protocols have successfully proposed ways to obtain hiPSC-derived cardiac pacemaker cells, despite the very limited availability of human tissues from the sinoatrial node. Providing an in vitro source of pacemaker-like cells would be of paramount importance in terms of furthering our understanding of the mechanisms underlying sinoatrial node pathophysiology and testing innovative clinical strategies against sinoatrial node dysfunction (i.e., biological pacemakers and genetic- and pharmacological- based therapy). Here, we summarize and detail the currently available protocols used to obtain patient-derived pacemaker-like cells.
External and internal, dark brown to red-brown lesions on branches of naturally infected plane trees in July and August 2022 in Cottbus, Germany Infected branches of different sizes (diameter 3–10 ...cm) were collected from four individual plane trees, with five samples being taken from each tree. To confirm the identification, genomic DNA was extracted from four isolates and the internal transcribed spacer (ITS) region of rDNA was amplified using the ITS1/ITS4 primers (White et al., 1990). Querciphoma minuta: (a) conidiomata on synthetic nutrient-poor agar; (b) conidiomata with setae; and (c-d) conidiomata with setae and conidia A pathogenicity test was performed using two isolates (22-784B, 22–1121) on twenty healthy plane tree seedlings (Platanus × hispanica), 120 to 150 cm in height.
A miniaturized paper-based lab-on-chip (LoC) was developed for the facile measurement of urinary Phe (phenylalanine) level on PKU (Phenylketonuria) treated patient. This system permits the monitoring ...of Phe in a dynamic range concentration of 20-3000 μM.
Numerous protocols of cardiac differentiation have been established by essentially focusing on specific growth factors on human pluripotent stem cell (hPSC) differentiation efficiency. However, the ...optimal environmental factors to obtain cardiac myocytes in network are still unclear. The mesoderm germ layer differentiation is known to be enhanced by low oxygen exposure. Here, we hypothesized that low oxygen exposure enhances the molecular and functional maturity of the cardiomyocytes. We aimed at comparing the molecular and functional consequences of low (5% O
or LOE) and high oxygen exposure (21% O
or HOE) on cardiac differentiation of hPSCs in 2D- and 3D-based protocols. hPSC-CMs were differentiated through both the 2D (monolayer) and 3D (embryoid body) protocols using several lines. Cardiac marker expression and cell morphology were assessed. The mitochondrial localization and metabolic properties were evaluated. The intracellular Ca
handling and contractile properties were also monitored. The 2D cardiac monolayer can only be differentiated in HOE. The 3D cardiac spheroids containing hPSC-CMs in LOE further exhibited cardiac markers, hypertrophy, steadier SR Ca
release properties revealing a better SR Ca
handling, and enhanced contractile force. Preserved distribution of mitochondria and similar oxygen consumption by the mitochondrial respiratory chain complexes were also observed. Our results brought evidences that LOE is moderately beneficial for the 3D cardiac spheroids with hPSC-CMs exhibiting further maturity. In contrast, the 2D cardiac monolayers strictly require HOE.
Type 2 Long QT Syndrome (LQT2) is a rare genetic heart rhythm disorder causing life-threatening arrhythmias. We derived induced pluripotent stem cell (iPSC) lines from two patients with LQT2, aged 18 ...and 6, both carrying a heterozygous missense mutation on the 3rd and 11th exons of KCNH2. The iPSC lines exhibited normal genomes, expressed pluripotent markers, and differentiated into trilineage embryonic layers. These patient-specific iPSC lines provide a valuable model to study the molecular and functional impact of the hERG channel gene mutation in LQT2 and to develop personalized therapeutic approaches for this syndrome.
Duchenne muscular dystrophy (DMD) is characterized by progressive muscle wasting following repeated muscle damage and inadequate regeneration. Impaired myogenesis and differentiation play a major ...role in DMD as well as intracellular calcium (Ca
) mishandling. Ca
release from the sarcoplasmic reticulum is mostly mediated by the type 1 ryanodine receptor (RYR1) that is required for skeletal muscle differentiation in animals. The study objective was to determine whether altered RYR1-mediated Ca
release contributes to myogenic differentiation impairment in DMD patients. The comparison of primary cultured myoblasts from six boys with DMD and five healthy controls highlighted delayed myoblast differentiation in DMD. Silencing
expression using specific si-RNA in a healthy control induced a similar delayed differentiation. In DMD myotubes, resting intracellular Ca
concentration was increased, but RYR1-mediated Ca
release was not changed compared with control myotubes. Incubation with the RYR-calstabin interaction stabilizer S107 decreased resting Ca
concentration in DMD myotubes to control values and improved calstabin1 binding to the RYR1 complex. S107 also improved myogenic differentiation in DMD. Furthermore, intracellular Ca
concentration was correlated with endomysial fibrosis, which is the only myopathologic parameter associated with poor motor outcome in patients with DMD. This suggested a potential relationship between RYR1 dysfunction and motor impairment. Our study highlights RYR1-mediated Ca
leakage in human DMD myotubes and its key role in myogenic differentiation impairment. RYR1 stabilization may be an interesting adjunctive therapeutic strategy in DMD.
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