Atomic Mechanisms of Timothy Syndrome-Associated Mutations in Calcium Channel Cav1.2

Timothy syndrome (TS) is a very rare multisystem disorder almost exclusively associated with mutations G402S and G406R in helix IS6 of Cav1.2. Recently, mutations R518C/H in helix IIS0 of the voltage sensing domain II (VSD-II) were described as a cause of cardiac-only TS. The three mutations are known to decelerate voltage-dependent inactivation (VDI). Here, we report a case of cardiac-only TS caused by mutation R518C. To explore possible impact of the three mutations on interdomain contacts, we modeled channel Cav1.2 using as templates Class Ia and Class II cryo-EM structures of presumably inactivated channel Cav1.1. In both models, R518 and several other residues in VSD-II donated H-bonds to the IS6-linked α1-interaction domain (AID). We further employed steered Monte Carlo energy minimizations to move helices S4–S5, S5, and S6 from the inactivated-state positions to those seen in the X-ray structures of the open and closed NavAb channel. In the open-state models, positions of AID and VSD-II were similar to those in Cav1.1. In the closed-state models, AID moved along the β subunit (Cavβ) toward the pore axis and shifted AID-bound VSD-II. In all the models R518 retained strong contacts with AID. Our calculations suggest that conformational changes in VSD-II upon its deactivation would shift AID along Cavβ toward the pore axis. The AID-linked IS6 would bend at flexible G402 and G406, facilitating the activation gate closure. Mutations R518C/H weakened the IIS0-AID contacts and would retard the AID shift. Mutations G406R and G402S stabilized the open state and would resist the pore closure. Several Cav1.2 mutations associated with long QT syndromes are consistent with this proposition. Our results provide a mechanistic rationale for the VDI deceleration caused by TS-associated mutations and suggest targets for further studies of calcium channelopathies

Generation of two iPSC lines (FAMRCi007-A and FAMRCi007-B) from patient with Emery-Dreifuss muscular dystrophy and heart rhythm abnormalities carrying genetic variant LMNA p.Arg249Gln.

Human iPSC lines were generated from peripheral blood mononuclear cells of patient carrying LMNA mutation associated with Emery–Dreifuss muscular dystrophy accompanied by atrioventricular block and paroxysmal atrial fibrillation. Reprogramming factors OCT4, KLF4, SOX2, CMYC were delivered using Sendai virus transduction. iPSCs were characterized in order to prove the pluripotency markers expression, normal karyotype, ability to differentiate into three embryonic germ layers. Generated iPSC lines would be useful model to investigate disease development associated with genetic variants in LMNA gene

Thr92Ala Polymorphism of Human Type 2 Deiodinase Gene (hD2) Affects the Development of Graves' Disease, Treatment Efficiency, and Rate of Remission

Clinical symptoms vary in thyrotoxicosis, and severity of these depends on many factors. Over the last years, impact of genetic factors upon the development and clinical significance of thyrotoxic symptoms became evident. It is known that a production of T3 in various tissues is limited by deiodinase 2 (D2). Recent studies revealed that certain single nucleotide polymorphisms (including threonine (Thr) to alanine (Ala) replacement in D2 gene codon 92, D2 Thr92Ala) affect T3 levels in tissues and in serum. Individuals with Ala92Ala genotype have lower D2 activity in tissues, compared with that in individuals with other genotypes. In our study, we have assessed an association of D2 Thr92Ala polymorphism with (1) frequency of disease development, (2) severity of clinical symptoms of thyrotoxicosis, and (3) rate of remissions, in Graves' disease patients

Thr92Ala Polymorphism of Human Type 2 Deiodinase Gene (hD2) Affects the Development of Graves' Disease, Treatment Efficiency, and Rate of Remission

Clinical symptoms vary in thyrotoxicosis, and severity of these depends on many factors. Over the last years, impact of genetic factors upon the development and clinical significance of thyrotoxic symptoms became evident. It is known that a production of T3 in various tissues is limited by deiodinase 2 (D2). Recent studies revealed that certain single nucleotide polymorphisms (including threonine (Thr) to alanine (Ala) replacement in D2 gene codon 92, D2 Thr92Ala) affect T3 levels in tissues and in serum. Individuals with Ala92Ala genotype have lower D2 activity in tissues, compared with that in individuals with other genotypes. In our study, we have assessed an association of D2 Thr92Ala polymorphism with (1) frequency of disease development, (2) severity of clinical symptoms of thyrotoxicosis, and (3) rate of remissions, in Graves' disease patients

Transgenerational and intergenerational effects of early childhood famine exposure in the cohort of offspring of Leningrad Siege survivors

Famine exposure during early life development can affect disease risk in late-life period, yet, transmission of phenotypic features from famine-exposed individuals to the next generations has not been well characterized. The purpose of our case–control study was to investigate the association of parental starvation in the perinatal period and the period of early childhood with the phenotypic features observed in two generations of descendants of Leningrad siege survivors. We examined 54 children and 30 grandchildren of 58 besieged Leningrad residents who suffered from starvation in early childhood and prenatal age during the Second World War. Controls from the population-based national epidemiological ESSE-RF study (n = 175) were matched on sex, age and body mass index (BMI). Phenotypes of controls and descendants (both generations, children and grandchildren separately) were compared, taking into account multiple testing. Comparison of two generations descendants with corresponding control groups revealed significantly higher creatinine and lower glomerular filtration rate (GFR), both in meta-analysis and in independent analyses. The mean values of GFR for all groups were within the normal range (GFR less than 60 mL/min/1.73 m2 was recorded in 2 controls and no one in DLSS). Additionally, independent of the creatinine level, differences in the eating pattern were detected: insufficient fish and excessive red meat consumption were significantly more frequent in the children of the Leningrad siege survivors compared with controls. Blood pressure, blood lipids and glucose did not differ between the groups. Parental famine exposure in early childhood may contribute to a decrease in kidney filtration capacity and altered eating pattern in the offspring of famine-exposed individuals.</p

Relationship Between Vitamin D Status and Vitamin D Receptor Gene Polymorphisms With Markers of Metabolic Syndrome Among Adults

Introduction: Recent studies have demonstrated that vitamin D deficiency contributes to the development of metabolic disorders, including obesity and type 2 diabetes mellitus (T2DM). Several vitamin D receptor (VDR) gene polymorphisms had been described to play a role in these conditions since vitamin D receptors were found in many tissues. The aim of this study was to assess the relationship between vitamin D status and VDR gene polymorphisms with metabolic syndrome (MS) parameters in Russian middle-aged women.Materials and Methods: A total of 697 women aged between 30 to 55 years were included in this cross-sectional study. Serum 25-hydroxyvitamin D (25(OH)D) level and four VDR gene polymorphisms rs1544410 (BsmI), rs7975232 (ApaI), rs731236 (TaqI), and rs2228570 (FokI) were measured. We applied the International Diabetes Federation (IDF) criteria to identify subjects with MS.Results: 9.3% of subjects had normal vitamin D level, while 90.7% were insufficient or deficient. Abdominal obesity (AO) was seen in 75.5%, impaired glucose tolerance (IGT) or T2DM was observed in 33.3%, reduced high-density lipoprotein cholesterol (HDL-C) level in 32.2% and hypertriglyceridemia in 23.4%. Serum 25(OH)D level in women with or without MS did not differ (48.6 ± 1.8 and 51.1 ± 1.5 nmol/l, p &gt; 0.05). Subjects with vitamin D deficiency showed an increased risk of AO [CI 95% 2.23; 1.15–4.30] and low HDL-C [CI95% 2.60; 1.04–6.49] compared to subjects with normal 25(OH)D level. IGT and T2DM risk was increased only when 25(OH)D concentration was less than 39.0 nmol/l [CI 95% 7.17; 2.99–17.7], but risk of MS did not differ in normal vitamin D status subjects and insufficient/deficient ones (p &gt; 0.05). T allele carriers (A) of rs7975232 had higher total cholesterol and low-density lipoprotein cholesterol levels compared with the GG (aa) genotypes. Similarly, GG (BB) genotype carriers of rs1544410 had higher triglyceride levels than subjects with A (b) allele carriers. However VDR gene polymorphisms did not seem to be associated with an increased risk of MS.Conclusions: Vitamin D deficiency, rs7975232, and rs1544410 VDR gene variants are associated with MS parameters in Russian middle-aged women

Inflammation and Mechanical Stress Stimulate Osteogenic Differentiation of Human Aortic Valve Interstitial Cells

Background: Aortic valve calcification is an active proliferative process, where interstitial cells of the valve transform into either myofibroblasts or osteoblast-like cells causing valve deformation, thickening of cusps and finally stenosis. This process may be triggered by several factors including inflammation, mechanical stress or interaction of cells with certain components of extracellular matrix. The matrix is different on the two sides of the valve leaflets. We hypothesize that inflammation and mechanical stress stimulate osteogenic differentiation of human aortic valve interstitial cells (VICs) and this may depend on the side of the leaflet.Methods: Interstitial cells isolated from healthy and calcified human aortic valves were cultured on collagen or elastin coated plates with flexible bottoms, simulating the matrix on the aortic and ventricular side of the valve leaflets, respectively. The cells were subjected to 10% stretch at 1 Hz (FlexCell bioreactor) or treated with 0.1 μg/ml lipopolysaccharide, or both during 24 h. Gene expression of myofibroblast- and osteoblast-specific genes was analyzed by qPCR. VICs cultured in presence of osteogenic medium together with lipopolysaccharide, 10% stretch or both for 14 days were stained for calcification using Alizarin Red.Results: Treatment with lipopolysaccharide increased expression of osteogenic gene bone morphogenetic protein 2 (BMP2) (5-fold increase from control; p = 0.02) and decreased expression of mRNA of myofibroblastic markers: α-smooth muscle actin (ACTA2) (50% reduction from control; p = 0.0006) and calponin (CNN1) (80% reduction from control; p = 0.0001) when cells from calcified valves were cultured on collagen, but not on elastin. Mechanical stretch of VICs cultured on collagen augmented the effect of lipopolysaccharide. Expression of periostin (POSTN) was inhibited in cells from calcified donors after treatment with lipopolysaccharide on collagen (70% reduction from control, p = 0.001), but not on elastin. Lipopolysaccharide and stretch both enhanced the pro-calcific effect of osteogenic medium, further increasing the effect when combined for cells cultured on collagen, but not on elastin.Conclusion: Inflammation and mechanical stress trigger expression of osteogenic genes in VICs in a side-specific manner, while inhibiting the myofibroblastic pathway. Stretch and lipopolysaccharide synergistically increase calcification