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Molecular docking, pivotal in predicting small-molecule ligand binding modes, struggles with accurately identifying binding conformations and affinities. This is particularly true for neonicotinoids, insecticides whose impacts on ecosystems require precise molecular interaction modeling. This study scrutinizes the effectiveness of prominent docking software (Ledock, ADFR, Autodock Vina, CDOCKER) in simulating interactions of environmental chemicals, especially neonicotinoid-like molecules with nicotinic acetylcholine receptors (nAChRs) and acetylcholine binding proteins (AChBPs). We aimed to assess the accuracy and reliability of these tools in reproducing crystallographic data, focusing on semi-flexible and flexible docking approaches. Our analysis identified Ledock as the most accurate in semi-flexible docking, while Autodock Vina with Vinardo scoring function proved most reliable. However, no software consistently excelled in both accuracy and reliability. Additionally, our evaluation revealed that none of the tools could establish a clear correlation between docking scores and experimental dissociation constants (Kd) for neonicotinoid-like compounds. In contrast, a strong correlation was found with drug-like compounds, bringing to light a bias in considered software towards pharmaceuticals, thus limiting their applicability to environmental chemicals. The comparison between semi-flexible and flexible docking revealed that the increased computational complexity of the latter did not result in enhanced accuracy. In fact, the higher computational cost of flexible docking with its lack of enhanced predictive accuracy, rendered this approach useless for this class of compounds. Conclusively, our findings emphasize the need for continued development of docking methodologies, particularly for environmental chemicals. This study not only illuminates current software capabilities but also underscores the urgency for advancements in computational molecular docking as it is a relevant tool to environmental sciences.

Abstract An oxalate-degrading bacterium in the gut microbiota absorbs food-derived oxalate to use this as a carbon and energy source, thereby reducing the risk of kidney stone formation in host animals. The bacterial oxalate transporter OxlT selectively uptakes oxalate from the gut to bacterial cells with a strict discrimination from other nutrient carboxylates. Here, we present crystal structures of oxalate-bound and ligand-free OxlT in two distinct conformations, occluded and outward-facing states. The ligand-binding pocket contains basic residues that form salt bridges with oxalate while preventing the conformational switch to the occluded state without an acidic substrate. The occluded pocket can accommodate oxalate but not larger dicarboxylates, such as metabolic intermediates. The permeation pathways from the pocket are completely blocked by extensive interdomain interactions, which can be opened solely by a flip of a single side chain neighbouring the substrate. This study shows the structural basis underlying metabolic interactions enabling favourable symbiosis.

IntroductionSickle cell disease (SCD) is an inherited hemoglobinopathy disorder. The main consequence is synthesis of hemoglobin S leading to chronic hemolysis associated with morbidity. The aim of this study was to investigate Thrombin Generation Assay (TGA) to assess hypercoagulability in SCD and TGA parameters as biomarkers of vaso-occlusive crisis (VOC) risk and hospitalization within 1 year.Materials and methodsWe performed TGA in platelet poor plasma (PPP) with 1 pM of tissue factor and 4 μM of phospholipid-standardized concentration, in duplicate for patients and controls. We measured thrombomodulin (TM), soluble endothelial Protein C Receptor and Tissue Factor Pathway Inhibitor (TFPI).ResultsA total of 113 adult patients with SCD, 83 at steady state and 30 during VOC, and 25 healthy controls matched on age and gender were included. Among the 83 patients at steady state, (36 S/S-1 S/β0, 20 S/Sα3.7, and 19 S/C-7 S/β+) 28 developed a VOC within 1 year (median: 4 months [2.25–6]). We observed an increase of peak and velocity associated with a shortening of lagtime and time to peak (TTP) and no difference of endogenous thrombin potential (ETP) in patients compared to controls. TFPI (p < 0.001) and TM (p = 0.006) were significantly decreased. TGA confirmed hypercoagulability in all SCD genotypes and clinical status. The association of ETP > 1,207 nM.min and peak >228.5 nM presented a sensitivity of 73.5% and a specificity of 93.9% to predict VOC development within 1 year.ConclusionWe have demonstrated a hypercoagulable state in SCD associated with chronic hemolysis. These preliminary findings suggest that TGA parameters, as ETP and peak, could be used to predict VOC development within 1 year.

Sickle cell disease is a complex genetic disease involving cell adhesion between red blood cells, white blood cells, platelets and endothelial cells, inducing painful vaso-occlusive crisis (VOC). We assessed reticulocyte and erythrocyte parameters in a cohort of confirmed SCD patients, and investigated whether a combination of these routine laboratory biomarkers of haemolysis could be used to predict VOC development. Reticulocyte and erythrocyte parameters were evaluated using the Sysmex XN-9000 analyser. A total of 98 patients with SCD were included, 72 in steady state and 26 in VOC. Among the 72 patients in steady state, 22 developed a VOC in the following year (median: 3 months [2–6]). The following parameters were increased in SCD patients with VOC development compared to SCD patients without VOC development in the following year: reticulocyte count (94.6 109/L [67.8–128] vs. 48.4 109/L [24.9–87.5]), immature reticulocyte count (259 109/L [181–334] vs. 152 109/L [129–208]) reticulocyte/immature reticulocyte fraction (IRF) ratio (6.63 109/(L*%) [4.67–9.56] vs. 4.94 109/(L*%) [3.96–6.61]), and medium fluorescence reticulocytes (MFR) (19.9% [17.4–20.7] vs. 17.1% [15.95–19.75]). The association of a reticulocyte count of >189.4 109/L and an MFR of >19.75% showed a sensitivity of 81.8% and a specificity of 88% to predict VOC development in the following year. Based on our findings, a combination of routine laboratory biomarkers, as reticulocyte count, immature reticulocyte count and fluorescent reticulocyte fraction at steady state, could be used to predict VOC development in SCD.

Hereditary xerocytosis is a dominantly inherited red cell membrane disorder caused in most cases by gain-of-function mutations in PIEZO1, encoding a mechanosensitive ion channel that translates a mechanic stimulus into calcium influx. We found that PIEZO1 was expressed early in erythroid progenitor cells, and investigated whether it could be involved in erythropoiesis, besides having a role in the homeostasis of mature red cell hydration. In UT7 cells, chemical PIEZO1 activation using YODA1 repressed glycophorin A expression by 75%. This effect was PIEZO1-dependent since it was reverted using specific short hairpin-RNA knockdown. The effect of PIEZO1 activation was confirmed in human primary progenitor cells, maintaining cells at an immature stage for longer and modifying the transcriptional balance in favor of genes associated with early erythropoiesis, as shown by a high GATA2/GATA1 ratio and decreased α/β-globin expression. The cell proliferation rate was also reduced, with accumulation of cells in G0/G1 of the cell cycle. The PIEZO1-mediated effect on UT7 cells required calcium-dependent activation of the NFAT and ERK1/2 pathways. In primary erythroid cells, PIEZO1 activation synergized with erythropoietin to activate STAT5 and ERK, indicating that it may modulate signaling pathways downstream of erythropoietin receptor activation. Finally, we studied the in-vitro erythroid differentiation of primary cells obtained from 14 PIEZO1-mutated patients, from 11 families, carrying ten different mutations. We observed a delay in erythroid differentiation in all cases, ranging from mild (n=3) to marked (n=8). Overall, these data demonstrate a role for PIEZO1 during erythropoiesis, since activation of PIEZO1 - both chemically and through activating mutations - delays erythroid maturation, providing new insights into the pathophysiology of hereditary xerocytosis.

We describe the clinical, hematologic and genetic characteristics of a retrospective series of 126 subjects from 64 families with hereditary xerocytosis. Twelve patients from six families carried a KCNN4 mutation, five had the recurrent p.Arg352His mutation and one had a new deletion at the exon 7-intron 7 junction. Forty-nine families carried a PIEZO1 mutation, which was a known recurrent mutation in only one-third of the cases and private sequence variation in others; 12 new probably pathogenic missense mutations were identified. The two dominant features leading to diagnosis were hemolysis that persisted after splenectomy and hyperferritinemia, with an inconstant correlation with liver iron content assessed by magnetic resonance imaging. PIEZO1-hereditary xerocytosis was characterized by compensated hemolysis in most cases, perinatal edema of heterogeneous severity in more than 20% of families and a major risk of post-splenectomy thrombotic events, including a high frequency of portal thrombosis. In KCNN4-related disease, the main symptoms were more severe anemia, hemolysis and iron overload, with no clear sign of red cell dehydration; therefore, this disorder would be better described as a ‘Gardos channelopathy’. These data on the largest series to date indicate that PIEZO1-hereditary xerocytosis and Gardos channelopathy are not the same disease although they share hemolysis, a high rate of iron overload and inefficient splenectomy. They demonstrate the high variability in clinical expression as well as genetic bases of PIEZO1-hereditary xerocytosis. These results will help to improve the diagnosis of hereditary xerocytosis and to provide recommendations on the clinical management in terms of splenectomy, iron overload and pregnancy follow-up.

The risk and clinical significance of cardiac iron overload due to chronic transfusion varies with the underlying disease. Cardiac iron overload shortens the life expectancy of patients with thalassemia, whereas its effect is unclear in those with myelodysplastic syndromes (MDS). In patients with sickle cell anemia (SCA), iron does not seem to deposit quickly in the heart. Our primary objective was to assess through a multicentric study the prevalence of cardiac iron overload, defined as a cardiovascular magnetic resonance T2*8 ECs in the past year, and age older than 6 years. We included from 9 centers 20 patients with thalassemia, 41 with SCA, and 25 with MDS in 2012-2014. Erythrocytapharesis did not consistently prevent iron overload in patients with SCA. Cardiac iron overload was found in 3 (15%) patients with thalassemia, none with SCA, and 4 (16%) with MDS. The liver iron content (LIC) ranged from 10.4 to 15.2 mg/g dry weight, with no significant differences across groups (P = 0.29). Abnormal T2* was not significantly associated with any of the measures of transfusion or chelation. Ferritin levels showed a strong association with LIC. Non-transferrin-bound iron was high in the thalassemia and MDS groups but low in the SCA group (P

To assess the influence of initial diameter on the gender difference in flow-dependent dilatation (FDD) of the conduit artery, we measured radial artery internal diameter (echotracking), flow (Doppler) and total blood viscosity in 24 healthy (25 [+ or -] 0.8 yr) men and women during reactive hyperemia (RH) and during a gradual hand skin heating (SH). At baseline, mean diameter (men, 2.76 [+ or -] 0.09 vs. women, 2.32 [+ or -] 0.07 mm, P < 0.05), flow (men, 21 [+ or -] 4 vs. women, 10 [+ or -] 1 ml/min, P < 0.05), and blood viscosity (men, 4.13 [+ or -] 0.07 vs. women, 3.92 [+ or -] 0.13 cP, P < 0.05) were higher in men but mean shear stress (MSS) was not different between groups. During RH, the percent increase in diameter was lower in men (men, 9 [+ or -] 1 vs. women, 13 [+ or -] 1%, P < 0.05). This difference was suppressed after correction for baseline diameter. During SH, the increase in diameter with flow was higher in women (P < 0.01). However, the increase in MSS was higher in women because of their smaller diameter at each level of flow (P < 0.01) and there was no difference between groups for the increase in diameter at each level of MSS. These results demonstrate in a direct manner that initial diameter influences the magnitude of FDD of conduit arteries in humans by modifying the value of the arterial wall shear stress at each level of flow and support the interest of the heating method in presence of heterogeneous groups. endothelium; shear stress; skin heating; vasodilatation