ProCMD: a database and 3D web resource for protein C mutants

Background: Activated Protein C (ProC) is an anticoagulant plasma serine protease which also plays an important role in controlling inflammation and cell proliferation. Several mutations of the gene are associated with phenotypic functional deficiency of protein C, and with the risk of developing venous thrombosis. Structure prediction and computational analysis of the mutants have proven to be a valuable aid in understanding the molecular aspects of clinical thrombophilia. Results: We have built a specialized relational database and a search tool for natural mutants of protein C. It contains 195 entries that include 182 missense and 13 stop mutations. A menu driven search engine allows the user to retrieve stored information for each variant, that include genetic as well as structural data and a multiple alignment highlighting the substituted position. Molecular models of variants can be visualized with interactive tools; PDB coordinates of the models are also available for further analysis. Furthermore, an automatic modelling interface allows the user to generate multiple alignments and 3D models of new variants. Conclusion: ProCMD is an up-to-date interactive mutant database that integrates phenotypical descriptions with functional and structural data obtained by computational approaches. It will be useful in the research and clinical fields to help elucidate the chain of events leading from a molecular defect to the related disease. It is available for academics at the URL http://www.itb.cnr.it/procmd/

The Organ Handling of Soluble Klotho in Humans

Chronic kidney disease (CKD) reduces both Klotho expression and its shedding into circulation, an effect that accelerates progression and cardiovascular complications. However, the mechanisms that regulate Klotho release by the human kidney are still unknown. Methods: We measured plasma Klotho across the kidney, splanchnic organs and lung in 22 patients (71 \ub1 2 years, estimated glomerular filtration rate [eGFR] 60 \ub1 5.4 mL/min 1.73 m2) during elective diagnostic cardiac catheterizations. Results: Although the Klotho average renal vein concentrations were remarkably higher (by ~9%) than arterial values, the kidney removed Klotho (or was at zero balance) in 7 subjects, indicating that the kidney contribution to systemic Klotho is not constant. Klotho fractional enrichment across the kidney was inversely related to plasma sodium (r = 0.43, p = 0.045) and acid uric acid levels (r = 0.38, p = 0.084) and directly, to renal oxygen extraction (r = 0.56, p = 0.006). In multivariate analysis, renal oxygen extraction was the only predictor of the enrichment of Klotho across the kidney, suggesting the dependence of renal Klotho release on tubular hypoxia or oxidative metabolism. Klotho balance was neutral across the lung. In patients with eGFR <60 mL/min, Klotho was also removed by splanchnic organs (single pass fractional extraction ~11%). Conclusions: The present study identifies kidney oxygen uptake as a predictor of Klotho release, and splanchnic organs as a site for Klotho removal. This study provides new understanding of kidney Klotho release and suggests that modulating kidney oxygen metabolism could increase Klotho delivery, as an option to slow disease progression and blunt organ damage

Morphology and Molecular Analysis of Ancient Grape Seeds

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BDNF Val66Met polymorphism and protein levels in Amniotic Fluid

Brain-Derived Neurotrophic Factor (BDNF) is a neurotrophin which plays survival- and growth-promoting activity in neuronal cells and it is involved in cellular plasticity mechanisms as it controls activity dependent synaptic transmission. A functional polymorphism (Val66Met) in the pro-region of BDNF, which affects the intracellular trafficking of proBDNF has been associated with memory and cognitive deficits as well as to an increased susceptibility for several psychiatric disorders especially those with a neurodevelopmental origin. To date, no study has evaluated the influence of the Val66Met polymorphism on BDNF levels in a peripheral system that may reflect fetal neurodevelopment. Therefore we investigated in amniotic fluids (AF) obtained from 139 healthy women during 15-17 week of pregnancy, BDNF protein levels in correlation with the Val66Met polymorphism

Efficacy of SARS-CoV-2 Vaccination in Dialysis Patients: Epidemiological Analysis and Evaluation of the Clinical Progress

This study investigated the impact of the fourth COVID-19 pandemic wave on dialysis patients of Romagna territory, assessing the associations of vaccination status with infection risk, clinical severity and mortality. From November 2021 to February 2022, an epidemiological search was conducted on 829 patients under dialysis treatment for at least one month. The data were then analyzed with reference to the general population of the same area. A temporal comparison was also carried out with the previous pandemic waves (from March 2020 to October 2021). The epidemiological evolution over time in the dialysis population and in Romagna citizens replicated the global trend, as the peak of the fourth wave corresponded to the time of maximum diffusion of omicron variant (B.1.1.529). Of 771 prevalent dialysis patients at the beginning of the study, 109 (14.1%) contracted SARS-CoV-2 infection during the 4-month observation period. Vaccine adherence in the dialysis population of the reference area was above 95%. Compared to fully or partially vaccinated subjects, the unvaccinated ones showed a significantly higher proportion of infections (12.5% vs. 27.0% p = 0.0341), a more frequent need for hospitalization (22.2% vs. 50.0%) and a 3.3-fold increased mortality risk. These findings confirm the effectiveness of COVID-19 vaccines in keeping infectious risk under control and ameliorating clinical outcomes in immunocompromised patients

Melorheostosis and Osteopoikilosis Clinical and Molecular Description of an Italian Case Series

Melorheostosis (MEL) is an uncommon, sclerosing disease, characterised by hyperostosis of long bones, resembling the flowing of candle wax. The disease is sporadic and the pathogenesis is still poorly understood. Occasionally, the same family can include individuals with MEL and Osteopoikilosis (OPK), a disease characterised by multiple round foci of increased bone density. LEMD3 gene mutations are related to OPK and Buschke–Ollendorff Syndrome, a genetic condition in which an association between MEL, OPK and skin lesions is observed. In rare cases, LEMD3 mutations and recently mosaic MAP2K1 gene mutations have been correlated to MEL suggesting that somatic mosaicism could be causative of the disease. In this study, we described the clinical, radiological and molecular findings of 19 individuals with MEL and 8 with OPK and compared the results to the medical literature. The molecular analyses of this case series corroborate the available data in the medical literature, indicating that LEMD3 germline mutations are not a major cause of isolated MEL and reporting five further cases of OPK caused by LEMD3 germline mutations

A novel de novo HCN1 loss-of-function mutation in genetic generalized epilepsy causing increased neuronal excitability

Abstract The causes of genetic epilepsies are unknown in the majority of patients. HCN ion channels have a widespread expression in neurons and increasing evidence demonstrates their functional involvement in human epilepsies. Among the four known isoforms, HCN1 is the most expressed in the neocortex and hippocampus and de novo HCN1 point mutations have been recently associated with early infantile epileptic encephalopathy. So far, HCN1 mutations have not been reported in patients with idiopathic epilepsy. Using a Next Generation Sequencing approach, we identified the de novo heterozygous p.Leu157Val (c.469C > G) novel mutation in HCN1 in an adult male patient affected by genetic generalized epilepsy (GGE), with normal cognitive development. Electrophysiological analysis in heterologous expression model (CHO cells) and in neurons revealed that L157V is a loss-of-function, dominant negative mutation causing reduced HCN1 contribution to net inward current and responsible for an increased neuronal firing rate and excitability, potentially predisposing to epilepsy. These data represent the first evidence that autosomal dominant missense mutations of HCN1 can also be involved in GGE, without the characteristics of epileptic encephalopathy reported previously. It will be important to include HCN1 screening in patients with GGE, in order to extend the knowledge of the genetic causes of idiopathic epilepsies, thus paving the way for the identification of innovative therapeutic strategies

Molecular Basis for the Dual Function of Eps8 on Actin Dynamics: Bundling and Capping

The unusual dual functions of the actin-binding protein EPS8 as an actin capping and actin bundling factor are mapped to distinct structural features of the protein and to distinct physiological activities in vivo

The Eps8/IRSp53/VASP Network Differentially Controls Actin Capping and Bundling in Filopodia Formation

There is a body of literature that describes the geometry and the physics of filopodia using either stochastic models or partial differential equations and elasticity and coarse-grained theory. Comparatively, there is a paucity of models focusing on the regulation of the network of proteins that control the formation of different actin structures. Using a combination of in-vivo and in-vitro experiments together with a system of ordinary differential equations, we focused on a small number of well-characterized, interacting molecules involved in actin-dependent filopodia formation: the actin remodeler Eps8, whose capping and bundling activities are a function of its ligands, Abi-1 and IRSp53, respectively; VASP and Capping Protein (CP), which exert antagonistic functions in controlling filament elongation. The model emphasizes the essential role of complexes that contain the membrane deforming protein IRSp53, in the process of filopodia initiation. This model accurately accounted for all observations, including a seemingly paradoxical result whereby genetic removal of Eps8 reduced filopodia in HeLa, but increased them in hippocampal neurons, and generated quantitative predictions, which were experimentally verified. The model further permitted us to explain how filopodia are generated in different cellular contexts, depending on the dynamic interaction established by Eps8, IRSp53 and VASP with actin filaments, thus revealing an unexpected plasticity of the signaling network that governs the multifunctional activities of its components in the formation of filopodia