Controversies in the pathophysiology of leg ulcers in sickle cell disease.

Patients with sickle cell disease (SCD) often experience painful vaso-occlusive crises and chronic haemolytic anaemia, as well as various acute and chronic complications, such as leg ulcers. Leg ulcers are characterized by their unpredictability, debilitating pain and prolonged healing process. The pathophysiology of SCD leg ulcers is not well defined. Known risk factors include male gender, poor social conditions, malnutrition and a lack of compression therapy when oedema occurs. Leg ulcers typically start with spontaneous pain, followed by induration, hyperpigmentation, blister formation and destruction of the epidermis. SCD is characterized by chronic haemolysis, increased oxidative stress and decreased nitric oxide bioavailability, which promote ischaemia and inflammation and consequently impair vascular function in the skin. This cutaneous vasculopathy, coupled with venostasis around the ankle, creates an ideal environment for local vaso-occlusive crises, which can result in the development of leg ulcers that resemble arterial ulcers. Following the development of the ulcer, healing is hindered as a result of factors commonly observed in venous ulceration, including venous insufficiency, oedema and impaired angiogenesis. All of these factors are modulated by genetic factors. However, our current understanding of these genetic factors remains limited and does not yet enable us to accurately predict ulceration susceptibility.

Two-Pole Nature of the Λ(1405) Resonance from Lattice QCD.

This Letter presents the first lattice QCD computation of the coupled channel πΣ-K[over ¯]N scattering amplitudes at energies near 1405 MeV. These amplitudes contain the resonance Λ(1405) with strangeness S=-1 and isospin, spin, and parity quantum numbers I(J^{P})=0(1/2^{-}). However, whether there is a single resonance or two nearby resonance poles in this region is controversial theoretically and experimentally. Using single-baryon and meson-baryon operators to extract the finite-volume stationary-state energies to obtain the scattering amplitudes at slightly unphysical quark masses corresponding to m_{π}≈200 MeV and m_{K}≈487 MeV, this study finds the amplitudes exhibit a virtual bound state below the πΣ threshold in addition to the established resonance pole just below the K[over ¯]N threshold. Several parametrizations of the two-channel K matrix are employed to fit the lattice QCD results, all of which support the two-pole picture suggested by SU(3) chiral symmetry and unitarity.

Metabolic Profile of Individuals with and without Type 2 Diabetes from Sub-Saharan Africa.

Epidemiological data predicts that sub-Saharan Africa will have the largest increase in type 2 diabetes (T2D) prevalence over the next two decades. Metabolomics studies have identified biomarkers that could improve T2D diagnosis and follow-up. However, no studies have characterized the metabolome of people from sub-Saharan Africa. Plasma samples from Senegalese individuals with T2D (n = 31) or without T2D (n = 34) were compared using measures of oxidative stress damage and plasma antioxidant enzyme activity and mass-spectrometry-based metabolomics analyses. Results showed that glucose, lactate, and tricarboxylic acid metabolites (fumarate, malate, and succinate) were increased in the T2D group, suggesting alterations in glycolysis and mitochondrial dysfunction. Several amino acids (leucine, isoleucine, valine, and tryptophan) and long-to-very-long-chain fatty acids were higher in the T2D group. Finally, elevated levels of ketone bodies and acylcarnitines were observed along with increased levels of oxidative stress damage and antioxidant activity. In conclusion, the T2D group exhibited modifications in metabolites previously shown to be associated with T2D risk in populations from other areas of the world. Future studies should seek to test whether these metabolites could be used as predictors for T2D-related complications in people from sub-Saharan Africa.

Red blood cell microvesicles activate the contact system, leading to factor IX activation via 2 independent pathways.

Storage lesion-induced, red cell-derived microvesicles (RBC-MVs) propagate coagulation by supporting the assembly of the prothrombinase complex. It has also been reported that RBC-MVs initiate coagulation via the intrinsic pathway. To elucidate the mechanism(s) of RBC-MV-induced coagulation activation, the ability of storage lesion-induced RBC-MVs to activate each zymogen of the intrinsic pathway was assessed in a buffer system. Simultaneously, the thrombin generation (TG) assay was used to assess their ability to initiate coagulation in plasma. RBC-MVs directly activated factor XII (FXII) or prekallikrein, but not FXI or FIX. RBC-MVs initiated TG in normal pooled plasma and in FXII- or FXI-deficient plasma, but not in FIX-deficient plasma, suggesting an alternate pathway that bypasses both FXII and FXI. Interestingly, RBC-MVs generated FIXa in a prekallikrein-dependent manner. Similarly, purified kallikrein activated FIX in buffer and initiated TG in normal pooled plasma, as well as FXII- or FXI-deficient plasma, but not FIX-deficient plasma. Dual inhibition of FXIIa by corn trypsin inhibitor and kallikrein by soybean trypsin inhibitor was necessary for abolishing RBC-MV-induced TG in normal pooled plasma, whereas kallikrein inhibition alone was sufficient to abolish TG in FXII- or FXI-deficient plasma. Heating RBC-MVs at 60°C for 15 minutes or pretreatment with trypsin abolished TG, suggesting the presence of MV-associated proteins that are essential for contact activation. In summary, RBC-MVs activate both FXII and prekallikrein, leading to FIX activation by 2 independent pathways: the classic FXIIa-FXI-FIX pathway and direct kallikrein activation of FIX. These data suggest novel mechanisms by which RBC transfusion mediates inflammatory and/or thrombotic outcomes.

Sex-related differences and effects of short and long trail running races on resting muscle-tendon mechanical properties.

The purpose of the study was to assess sex-related differences in resting mechanical properties and adaptations of skeletal muscles and tendons in response to trail running races of different distances using multi-site shear wave elastography assessments of the lower limb, force capacity and blood analyses. Sex differences in resting mechanical properties of knee extensor and plantar flexor muscles and tendons were characterized by shear wave velocity (SWV) measurements in healthy males (N = 42) and females (N = 25) trained in long-distance running. Effects of running distance on muscle and tendon properties were assessed in short (<60 km, N = 23) vs. long (>100 km, N = 26) distance races. Changes in isometric maximal voluntary contraction torque, serum C-reactive protein and creatine kinase activity were also quantified after running races. Higher SWV of relaxed triceps surae muscle was detected in females as compared to males before running races (+4.8%, p = 0.006), but the significant increases in triceps surae muscle group (+7.0%, p = 0.001) and patellar tendon SWV (+15.4%, p = 0.001) after short-distance races were independent of sex. A significant decrease in triceps surae muscle SWV was found after long-distance races in the whole experimental population (-3.1%, p = 0.049). Post-races increase in C-reactive protein and creatine kinase activity were significantly correlated to the relative decreases in triceps surae and quadriceps femoris skeletal muscle SWV (ρ = -0.56, p = 0.001 and ρ = -0.51, p = 0.001, respectively). Resting mechanical properties of muscles and tendons are affected by sex, and adaptations to trail races are related to running distance. Exercise-induced changes in resting skeletal muscle mechanical properties are associated with enhanced indirect markers of inflammation and muscle damage.

Oxidative stress, inflammation, blood rheology, and microcirculation in adults with sickle cell disease: Effects of hydroxyurea treatment and impact of sickle cell syndrome.

Inflammation and oxidative stress play a key role in the pathophysiology of sickle cell disease (SCD). However, the potential influence of different sickle genotypes, or hydroxyurea (HU) treatment, on these factors remains poorly documented. The present study compared several plasma markers of inflammation and oxidative stress, as well as microvascular function, between patients with sickle SC disease (HbSC, n = 19) and patients with sickle cell anemia (HbSS) under hydroxyurea (HU) treatment (n = 16), or not (n = 13). Hemorheological parameters and levels of inflammatory (IL-6, IL-8, IFN-γ, MCP-1, MIP-1ß, TNF-α) and oxidative stress (AOPP, MDA, MPO) markers were determined. Peripheral microcirculatory cutaneous blood flow and immediate microvascular response to local heat were evaluated using laser Doppler flowmetry. Oxidative stress and inflammation were lower in HbSC patients and HbSS patients under HU therapy compared to HbSS patients not treated with HU. Blood viscosity was higher in HbSC than in HbSS patients treated with or not with HU. Vasodilation response of the cutaneous microcirculation to heat stress was higher in HbSS patients receiving HU treatment. Our results clearly established that both sickle cell genotype and HU treatment modulate inflammation and oxidative stress.

Acute Cycling Exercise Induces Changes in Red Blood Cell Deformability and Membrane Lipid Remodeling.

Here we describe the effects of a controlled, 30 min, high-intensity cycling test on blood rheology and the metabolic profiles of red blood cells (RBCs) and plasma from well-trained males. RBCs demonstrated decreased deformability and trended toward increased generation of microparticles after the test. Meanwhile, metabolomics and lipidomics highlighted oxidative stress and activation of membrane lipid remodeling mechanisms in order to cope with altered properties of circulation resulting from physical exertion during the cycling test. Of note, intermediates from coenzyme A (CoA) synthesis for conjugation to fatty acyl chains, in parallel with reversible conversion of carnitine and acylcarnitines, emerged as metabolites that significantly correlate with RBC deformability and the generation of microparticles during exercise. Taken together, we propose that RBC membrane remodeling and repair plays an active role in the physiologic response to exercise by altering RBC properties.

Hemoglobin A1c and fructosamine correlate in a patient with sickle cell disease and diabetes on chronic transfusion therapy.

In patients with sickle cell disease (SCD) and diabetes mellitus (DM), hemoglobin A1c (HbA1c ) is unreliable and the American Diabetes Association recommends monitoring long-term glycemia by measuring serum glucose, but use of serum fructosamine (SF), a measurement independent of red cell lifespan, has been reported. SF as a screen for DM in SCD, however, is not standardized and its relationship to serum glucose has not been validated. Further, screening for DM was not adequately addressed in the 2014 National Heart, Lung, and Blood Institute (NHLBI) guidelines for SCD management. Blood transfusions, an important treatment for some patients with SCD, can also impact HbA1c . We present a case of a patient with SCD and cystic fibrosis-related diabetes on monthly chronic transfusions therapy (CTT) who had well-correlated "steady state" HbA1c and SF levels over time, suggesting for the first time these markers may actually be useful when following long-term glycemic control in patients with SCD on CTT programs.

Impact of a 10 km running trial on eryptosis, red blood cell rheology, and electrophysiology in endurance trained athletes: a pilot study.

PURPOSE: Blood rheology is a key determinant of blood flow and tissue perfusion. There are still large discrepancies regarding the effects of an acute running exercise on blood rheological properties and red blood cell (RBC) physiology. We investigated the effect of a 10 km running trial on markers of blood rheology and RBC physiology in endurance trained athletes. METHODS: Blood was sampled before and after the exercise to measure lactate and glucose, hematological and hemorheological parameters (blood viscosity, RBC deformability, and aggregation), eryptosis markers (phosphatidylserine and CD47 exposure, RBC reactive oxygen species), RBC-derived microparticles (RBC-MPs), and RBC electrophysiological activity. Weight was measured before and after exercise. Peripheral oxygen saturation and heart rate were monitored before and during the trial. RESULTS: Blood lactate and glucose levels increased after exercise and subjects significantly lost weight. All athletes experienced a significant fall in oxygen saturation. Mean corpuscular volume (MCV) was increased from 95.1 ± 3.2 to 96.0 ± 3.3 and mean corpuscular hemoglobin concentration (MCHC) decreased after exercise suggesting a slight RBC rehydration. Exercise increased RBC deformability from 0.344 ± 0.04 to 0.378 ± 0.07, decreased RBC aggregates strength and blood viscosity, while hematocrit (Hct) remained unaffected. While RBC electrophysiological recording suggested a modulation in RBC calcium content and/or chloride conductance, eryptosis markers and RBC-MPs were not modified by the exercise. CONCLUSION: A 10 km acute running exercise had no effect on RBC senescence and membrane blebbing. In contrast, this exercise increased RBC deformability, probably through rehydration process which resulted in a decrease in blood viscosity.

Impact of Trail Running Races on Blood Viscosity and Its Determinants: Effects of Distance.

Blood rheology is a key determinant of tissue perfusion at rest and during exercise. The present study investigated the effects of race distance on hematological, blood rheological, and red blood cell (RBC) senescence parameters. Eleven runners participated in the Martigny-Combes à Chamonix 40 km race (MCC, elevation gain: 2300 m) and 12 others in the Ultra-Trail du Mont Blanc (UTMB, 171 km, elevation gain: 10,000 m). Blood samples were collected before and after the races. After the UTMB, the percentage of RBC phosphatidylserine (PS) exposure was not affected while RBC CD235a levels decreased and RBC-derived microparticles increased. In contrast, after the MCC, RBC PS exposure increased, while RBC CD235a and RBC-derived microparticles levels were not affected. The free hemoglobin and hemolysis rate did not change during the races. RBC aggregation and blood viscosity at moderate shear rates increased after the MCC. RBC deformability, blood viscosity at a high shear rate, and hematocrit decreased after the UTMB but not after the MCC. Our results indicate that blood rheology behavior is different between a 40 km and a 171 km mountain race. The low blood viscosity after the ultra-marathon might facilitate blood flow to the muscles and optimize aerobic performance.

Altered blood rheology and impaired pressure-induced cutaneous vasodilation in a mouse model of combined type 2 diabetes and sickle cell trait.

OBJECTIVE: Type 2 diabetes (T2D)-related vascular dysfunction and hemorheological abnormalities could possibly be amplified by sickle cell trait (SCT). These alterations could potentially increase the risk of vascular complications in individuals with combined T2D and SCT. Therefore, this study used a mouse model to determine whether vascular function and blood rheology were more severely altered in combined T2D and SCT than in T2D or SCT alone. METHODS: Townes transgenic mice with or without SCT received a 12-week high fat high sucrose or standard diet to create models of combined T2D-SCT, T2D, SCT, and controls. Pressure-induced vasodilation (PIV) and sodium nitroprusside (SNP)-mediated vasodilation in-vivo, and hemorheological parameters were measured. RESULTS: No significant differences in blood viscosity, hematocrit, erythrocyte deformability, or PIV were observed between the control and T2D mice, or the control and SCT mice. However, blood viscosity, erythrocyte deformability, and PIV were significantly altered in the T2D-SCT mice compared to the control mice. There were no differences in SNP response between the groups. CONCLUSIONS: Although neither T2D nor SCT alone had significant effects on blood rheology parameters or vascular function, combined T2D-SCT mice had significantly altered blood rheology and significantly impaired vascular function.

Evaluation of the accuracy of multiparametric MRI for predicting prostate cancer pathology and tumour staging in the real world: an multicentre study.

OBJECTIVES: To assess the accuracy of multiparametric magnetic resonance imaging (mpMRI) for the detection of significant prostate cancer in men undergoing radical prostatectomy (RP) in an Australian multicentre setting, and to assess concordance between mpMRI and RP for local tumour staging and index lesion locations. PATIENTS AND METHODS: Men who underwent mpMRI within 12 months of RP between January 2013 and August 2016 at three Australian sites were included (Central Coast, NSW, St Vincents Hospital, Melbourne, Vic., and Bendigo Hospital, Vic.). The results of mpMRI were compared with the final RP specimen to analyse the performance of mpMRI for significant prostate cancer detection, index lesion localization, prediction of T3 disease and lymph node metastasis. A comparison between mpMRI cases performed using the technical and reporting specifications of Prostate Imaging Reporting and Data System (PI-RADS) version 1 and version 2 was also performed. Data analysis was performed using spss 24.0. RESULTS: A total of 235 cases were included for analysis. mpMRI PI-RADS score ≥3 had a 91% sensitivity and 95% positive predictive value (PPV) for significant prostate cancer at RP. The overall concordance between index lesion location on mpMRI and RP specimen was 75%. The sensitivity for predication of significant prostate cancer was higher in the PI-RADS version 2 cases compared with PI-RADS version 1 (87-99%; P = 0.005). Index lesion concordance was higher in the PI-RADS version 2 group (68% vs 91%; P = 0.002). mpMRI had a 38% sensitivity, 95% specificity, 90% PPV and 57% negative predictive value for extraprostatic disease. Sensitivity for prediction of T3 disease improved from 30% to 62% (P = 0.008) with PI-RADS version 2. CONCLUSIONS: In patients undergoing RP, an abnormal mpMRI is highly predictive (95% PPV) of significant prostate cancer, with an index lesion concordance of 75%. There has been a significant improvement in accuracy after the adoption of PI-RADS version 2 technical specifications and reporting criteria; however; further study is required to determine if this is attributable to improved experience with mpMRI or changes in the PI-RADS system.

Altered anabolic signalling and reduced stimulation of myofibrillar protein synthesis after feeding and resistance exercise in people with obesity.

KEY POINTS: Lifestyle modifications that include the regular performance of exercise are probably important for counteracting the negative consequences of obesity on postprandial myofibrillar protein synthetic responses to protein dense food ingestion. We show that the interactive effect of resistance exercise and feeding on the stimulation of myofibrillar protein synthesis rates is diminished with obesity compared to normal weight adults. The blunted myofibrillar protein synthetic response with resistance exercise in people with obesity may be underpinned by alterations in muscle anabolic signalling phosphorylation (p70S6K and 4E-BP1). The results obtained in the present study suggest that further exercise prescription manipulation may be necessary to optimize post-exercise myofibrillar protein synthesis rates in adults with obesity. ABSTRACT: We aimed to determine whether obesity alters muscle anabolic and inflammatory signalling phosphorylation and also muscle protein synthesis within the myofibrillar (MYO) and sarcoplasmic (SARC) protein fractions after resistance exercise. Nine normal weight (NW) (21 ± 1 years, body mass index 22 ± 1 kg m-2 ) and nine obese (OB) (22 ± 1 years, body mass index 36 ± 2 kg m-2 ) adults received l-[ring-13 C6 ]phenylalanine infusions with blood and muscle sampling at basal and fed-state of the exercise (EX) and non-exercise (CON) legs. Participants performed unilateral leg extensions and consumed pork (36 g of protein) immediately after exercise. Basal muscle Toll-like receptor 4 (TLR4) protein was similar between OB and NW groups (P > 0.05) but increased at 300 min after pork ingestion only in the OB group (P = 0.03). Resistance exercise reduced TLR4 protein in the OB group at 300 min (EX vs. CON leg in OB: P = 0.04). Pork ingestion increased p70S6K phosphorylation at 300 min in CON and EX of the OB and NW groups (P > 0.05), although the response was lower in the EX leg of OB vs. NW at 300 min (P = 0.05). Basal MYO was similar between the NW and OB groups (P > 0.05) and was stimulated by pork ingestion in the EX and CON legs in both groups (Δ from basal NW: CON 0.04 ± 0.01% h-1 ; EX 0.10 ± 0.02% h-1 ; OB: CON 0.06 ± 0.01% h-1 ; EX 0.06 ± 0.01% h-1 ; P < 0.05). MYO was more strongly stimulated in the EX vs. CON legs in NW (P = 0.02) but not OB (P = 0.26). SARC was feeding sensitive but not further potentiated by resistance exercise in both groups. Our results suggest that obesity may attenuate the effectiveness of resistance exercise to augment fed-state MYO.

Does physical activity increase or decrease the risk of sickle cell disease complications?

Sickle cell disease (SCD) is the most common inherited disease in the world. Red blood cell sickling, blood cell-endothelium adhesion, blood rheology abnormalities, intravascular haemolysis, and increased oxidative stress and inflammation contribute to the pathophysiology of SCD. Because acute intense exercise may alter these pathophysiological mechanisms, physical activity is usually contra-indicated in patients with SCD. However, recent studies in sickle-cell trait carriers and in a SCD mice model show that regular physical activity could decrease oxidative stress and inflammation, limit blood rheology alterations and increase nitric oxide metabolism. Therefore, supervised habitual physical activity may benefit patients with SCD. This article reviews the literature on the effects of acute and chronic exercise on the biological responses and clinical outcomes of patients with SCD.

Association between oxidative stress and vascular reactivity in children with sickle cell anaemia and sickle haemoglobin C disease.

Oxidative stress and haemolysis-associated nitric oxide (NO) depletion plays a crucial role in the development of vasculopathy in sickle cell anaemia (SS). However it remains unknown whether oxidative stress and haemolysis levels influence vascular function in patients with sickle haemoglobin C disease (SC). Microvascular response to heat (using Laser Doppler flowmetry on finger), oxidative stress biomarkers, NO metabolites, endothelin-1 and haematological parameters were compared between patients with SS and SC. Vascular function, oxidative and nitrosative markers were also measured in healthy (AA) children. SS and SC had increased plasma advanced oxidation protein products (AOPP), malondialdehyde, plasma antioxidant activities and NO end products, compared to AA. SC had lower catalase activity compared to AA and SS. Haemolytic rate, glutathione peroxidase and nitrotyrosine concentrations were significantly increased in children with SS compared to SC and AA. SS and SC had impaired microvascular reactivity compared to AA. In SS, the plateau phase of the response to local thermal heating was negatively associated with nitrotyrosine and AOPP. No association between vascular function parameters and oxidative stress markers was observed in SC. Mild haemolysis in SC, compared to SS, may limit oxidative and nitrosative stress and could explain the better preserved microvascular function in this group.

Alpha-thalassaemia promotes frequent vaso-occlusive crises in children with sickle cell anaemia through haemorheological changes.

BACKGROUND: Sickle cell anaemia (SCA) is a severe hereditary haemoglobinopathy characterised by haemorheological abnormalities, which play a role in the occurrence of several acute and chronic clinical complications. While ßS -haplotypes and alpha-thalassaemia modulate SCA clinical severity, their effects on blood rheology have been incompletely described. The aim of this study was to test the effects of these genetic modifiers on the haemorheological properties and clinical complication of children with SCA. PROCEDURE: Steady-state haemorheological profile, biological parameters, ßS -haplotypes, alpha-globin status, vaso-occlusive crisis (VOC) and acute chest syndrome frequencies were analysed in 128 children (aged 5 to 18 years) with SCA. RESULTS: Patients with alpha-thalassaemia showed increased red blood cell (RBC) deformability and aggregation compared to those without. Median VOC rate was higher in patients with homozygous alpha-thalassaemia compared to those with a normal alpha genotype. Conversely, the haemorheological profile and clinical complications were not influenced by the ßS -haplotypes in our study. CONCLUSION: Our results demonstrate that alpha-thalassaemia is associated with higher risk for VOC events in children with SCA, which may be due in part to its effects on RBC deformability and aggregation.

The maize brown midrib4 (bm4) gene encodes a functional folylpolyglutamate synthase.

Mutations in the brown midrib4 (bm4) gene affect the accumulation and composition of lignin in maize. Fine-mapping analysis of bm4 narrowed the candidate region to an approximately 105 kb interval on chromosome 9 containing six genes. Only one of these six genes, GRMZM2G393334, showed decreased expression in mutants. At least four of 10 Mu-induced bm4 mutant alleles contain a Mu insertion in the GRMZM2G393334 gene. Based on these results, we concluded that GRMZM2G393334 is the bm4 gene. GRMZM2G393334 encodes a putative folylpolyglutamate synthase (FPGS), which functions in one-carbon (C1) metabolism to polyglutamylate substrates of folate-dependent enzymes. Yeast complementation experiments demonstrated that expression of the maize bm4 gene in FPGS-deficient met7 yeast is able to rescue the yeast mutant phenotype, thus demonstrating that bm4 encodes a functional FPGS. Consistent with earlier studies, bm4 mutants exhibit a modest decrease in lignin concentration and an overall increase in the S:G lignin ratio relative to wild-type. Orthologs of bm4 include at least one paralogous gene in maize and various homologs in other grasses and dicots. Discovery of the gene underlying the bm4 maize phenotype illustrates a role for FPGS in lignin biosynthesis.

Inflammatory and oxidative stress phenotypes in transgenic sickle cell mice.

The Townes mouse model of homozygous sickle cell disease (SS) has emerged as the major experimental model for studying pathophysiological mechanisms of human sickle cell disease (SCD). We therefore investigated hematological and hemorheological parameters as well as organ-specific inflammatory and oxidative stress molecular profiles in these animals in steady state conditions. Evidences of SCD-related intravascular hemolysis, impaired red blood cell (RBC) deformability, leukocytosis and altered plasma nitric oxide byproducts (NOx) level were found in the SS mice. The SS mice have damaged, enlarged and dysfunctional spleen as attested by high AOPP levels, low SOD and GPx activities and low pro-inflammatory cytokines mRNA expression. SS mice exhibited cardiomegaly, high cardiac mRNA levels of proinflammatory markers and low cardiac GPx activity. While lungs did not display any noticeable defects, liver and kidney were particularly sensitive to oxidative stress and inflammation as suggested by high AOPP levels in both organs, elevated renal NF-κB and TNF-α, and increased hepatic VCAM-1 and IL-1ß. Our data indicate a tissue-specific phenotype regarding oxidative stress and inflammation in SS mice that may help to optimize the development of novel potential drug treatments.