Hemizygous nonsense variant in the moesin gene (MSN) leads to a new autoimmune phenotype of Immunodeficiency 50.

Here, we present the findings of an investigation involving two male siblings with juvenile total tooth loss, early-onset chronic leg ulcers, and autoimmune thyroiditis, as well as focal segmental glomerulosclerosis with associated pulmonary emphysema in one and diabetes mellitus in the other. The clinical picture and lupus anticoagulant, cryoglobulin, and cold agglutinin positivity suggested the diagnosis of antiphospholipid syndrome. Flow cytometry analysis showed immunophenotypes consistent with immune dysregulation: a low number of naive T cells, elevated CD4+ T cell counts, and decreased CD8+ T-cell counts were detected, and more than half of the T-helper population was activated. Considering the siblings' almost identical clinical phenotype, the genetic alteration was suspected in the background of the immunodeficiency. Whole exome sequencing identified a previously not described hemizygous nonsense variant (c.650G>A, p.W217X) within exon 6 of the moesin (MSN) gene localized on chromosome X, resulting in significantly decreased MSN mRNA expression compared to healthy controls. We present a putative new autoimmune phenotype of Immunodeficiency 50 (MIM300988) characterized by antiphospholipid syndrome, Hashimoto's thyroiditis, leg ulcers, and juvenile tooth loss, associated with W217X mutation of the MSN gene.

Different modes of barrel opening suggest a complex pathway of ligand binding in human gastrotropin.

Gastrotropin, the intracellular carrier of bile salts in the small intestine, binds two ligand molecules simultaneously in its internal cavity. The molecular rearrangements required for ligand entry are not yet fully clear. To improve our understanding of the binding process we combined molecular dynamics simulations with previously published structural and dynamic NMR parameters. The resulting ensembles reveal two distinct modes of barrel opening with one corresponding to the transition between the apo and holo states, whereas the other affecting different protein regions in both ligation states. Comparison of the calculated structures with NMR-derived parameters reporting on slow conformational exchange processes suggests that the protein undergoes partial unfolding along a path related to the second mode of the identified barrel opening motion.

Impact of tranexamic acid use on blood loss and transfusion rates following femoral varus derotational osteotomy in children with cerebral palsy.

PURPOSE: Previous studies have established the safety and efficacy of tranexamic acid (TXA) in reducing blood loss after total joint arthroplasty and spinal fusion surgery; however, literature regarding the effectiveness of intraoperative TXA in children with cerebral palsy (CP) is limited. The aim of this study was to investigate the safety and efficacy of intraoperative TXA in reducing blood loss and transfusion requirements for children with CP undergoing a proximal femoral varus derotational osteotomy (VDRO). METHODS: This is a retrospective review of 258 children with CP who underwent VDRO performed at the author's institution between 2004 and 2017. In all, 36 subjects underwent VDRO surgery with administration of intravenous TXA and 222 subjects underwent VDRO without administration of TXA. Outcome measures including blood loss, transfusion requirements and venous thromboembolic events were compared between groups using t-tests and chi-squared tests. RESULTS: No significant differences were seen in the rates of transfusion between groups for the entire hospitalization (TXA group: 11.1% versus No TXA group: 19.8%), intraoperatively (TXA: 2.8% versus No TXA: 9.0%) or postoperatively (TXA: 8.3% versus No TXA: 14.4%). Intraoperative estimated blood loss (TXA: 144.4 mL versus No TXA: 159.0 mL) and percentage blood loss (TXA: 8.9% versus No TXA: 9.2%) were similar between groups. No major thromboembolic complications events occurred in either group. CONCLUSION: The use of TXA was not associated with thromboembolic complications in this series of children with CP undergoing VDRO surgery. Though there was a trend toward lower rates of intraoperative and postoperative blood transfusion with TXA use in these patients, the differences were not significant, possibly due to low estimated blood loss in both groups and sample size. LEVEL OF EVIDENCE: III- retrospective comparative study.

Stomaching the pain of spinal fusion: gastrointestinal discomfort is as severe as back pain in 50% of adolescent idiopathic scoliosis patients following posterior spinal fusion.

PURPOSE: There have been no prospective studies investigating gastrointestinal (GI) symptoms of patients with adolescent idiopathic scoliosis (AIS) following posterior spinal fusion (PSF). The purpose of this study was to evaluate the incidence and severity of self-reported GI symptoms following PSF. METHODS: In all, 40 AIS patients undergoing PSF were prospectively enrolled between March 2015 and October 2016. Patients completed a survey on each postoperative, inpatient day regarding nausea, emesis, constipation, abdominal pain and back pain, rating their pain on a scale of 1 to 10. RESULTS: Abdominal pain (50%), emesis (63%), nausea (65%) and constipation (68%) were experienced by the majority of patients. Of those reporting back pain, the mean pain level during the postoperative period was 5.1 (0.2 to 9.6). Of those reporting abdominal pain, the mean pain level during the postoperative period was 5.5 (1.4 to 8.6), which was not different than the severity of their back-pain levels (mean = 6.0, p = 0.31). CONCLUSIONS: Gastrointestinal issues in AIS patients following PSF are common. Abdominal pain was as severe as the back pain for half of the patients. LEVEL OF EVIDENCE: II.

The effect of canal fill on paediatric femur fractures treated with titanium elastic nails.

PURPOSE: Traditional teaching for fixation of paediatric femur fractures recommends 80% nail diameter/medullary canal diameter ratio (ND/MCD) for successful maintenance of reduction. Prior studies have investigated this with stainless steel Enders nails. Our aim was to assess the impact of ND/MCD on maintenance of reduction and malunion rates in paediatric femur fractures treated with flexible intramedullary nails (FINs). METHODS: Retrospective data was collected on all paediatric patients treated with FINs for diaphyseal femur fractures at a single tertiary care institution over a ten-year period. Patients with co-morbidities affecting bone quality were excluded. Patients were subdivided into groups based on ND/MCD. RESULTS: A total of 66 patients met inclusion criteria. Mean ND/MCD was 76.3% (32.9% to 98.8%, SD 14.3). In all, 50% (n = 33/66) of patients had > 80% ND/MCD, and only 13.6% (n = 9/66) of patients had less than 60% ND/MCD. When controlling for fracture stability, ND/MCD had no correlation with mean shortening (p = 0.07) There was no correlation between ND/MCD and angulation in the sagittal (p = 0.96) or coronal plane (p = 0.20). Three patients fit malunion criteria. ND/MCD for these patients were 40%, 67% and 79%. CONCLUSION: There was no correlation between ND/MCD and shortening or malangulation. The majority of patients in this series with less than 80% fill with FIN healed within acceptable parameters. LEVEL OF EVIDENCE: III.

The effects of body mass index on treatment of paediatric femur fractures managed with flexible intramedullary nails.

PURPOSE: Traditionally, flexible intramedullary nails (FINs) are not to be used to fix femur fractures in patients > 50 kg (110 lbs). However, studies have not examined the efficacy of this technique in overweight and obese patients who may be under this 'weight cutoff'. The purpose of this study was to assess how patient body mass index (BMI) impacts the treatment of paediatric femur fractures managed with FINs. METHODS: Retrospective data was collected on all paediatric patients treated with FINs for diaphyseal femur fractures at a single tertiary care institution over a ten-year period. BMI was calculated and categorised according to the Centre for Disease Control BMI Calculator for Children and Teens. Patients with comorbidities affecting bone quality were excluded. RESULTS: A total of 54 patients met inclusion criteria. In all, 14 patients were underweight, 20 were within a normal weight range, and 20 were overweight/obese. There was no correlation between BMI and mean shortening (underweight: 7.1 mm, normal weight: 5.2 mm, overweight/obese: 7.2 mm; p = 0.55). There was no correlation between BMI and mean anterior/posterior angulation (underweight: 3.1°, normal weight: 3.8°, overweight/obese: 3.3°; p = 0.93). There was no correlation between BMI and varus/valgus angulation (underweight: -0.86°, normal weight: -0.5°, overweight/obese: -1.25°; p = 0.89). Three cases fit malunion criteria. One of these patients fell into the 'underweight' category and two patients fell into the 'normal weight' category. CONCLUSION: We found no association between BMI and malunion in FIN fixation of femoral diaphyseal fractures in children. All cases of malunion were seen in underweight or normal weight patients.

In Vivo Effects of Quercetin in Association with Moderate Exercise Training in Improving Streptozotocin-Induced Aortic Tissue Injuries.

BACKGROUND: Diabetes mellitus (DM) is a chronic endocrine-metabolic disorder associated with endothelial dysfunction. Hyperglycemia, dyslipidemia and abnormal nitric oxide-mediated vasodilatation are the major causal factors in the development of endothelial dysfunction in DM. The prevention of endothelial dysfunction may be a first target against the appearance of atherosclerosis and cardiovascular diseases. We have investigated the synergistic protective effects of quercetin administration and moderate exercise training on thoracic aorta injuries induced by diabetes. METHODS: Diabetic rats that performed exercise training were subjected to a swimming training program (1 h/day, 5 days/week, 4 weeks). The diabetic rats received quercetin (30 mg/kg body weight/day) for 4 weeks. At the end of the study, the thoracic aorta was isolated and divided into two parts; one part was immersed in 10% formalin for histopathological evaluations and the other was frozen for the assessment of oxidative stress markers (malondialdehyde, MDA and protein carbonyls groups, PC), the activity of antioxidant enzymes (superoxide dismutase, SOD and catalase, CAT), nitrite plus nitrate (NOx) production and inducible nitric oxide synthase (iNOS) protein expression. RESULTS: Diabetic rats showed significantly increased MDA and PC levels, NOx production and iNOS expression and a reduction of SOD and CAT activity in aortic tissues. A decrease in the levels of oxidative stress markers, NOx production and iNOS expression associated with elevated activity of antioxidant enzymes in the aortic tissue were observed in quercetin-treated diabetic trained rats. CONCLUSIONS: These findings suggest that quercetin administration in association with moderate exercise training reduces vascular complications and tissue injuries induced by diabetes in rat aorta by decreasing oxidative stress and restoring NO bioavailability.

Characterization of a novel acylaminoacyl peptidase with hexameric structure and endopeptidase activity.

We have overexpressed in E. coli, purified and investigated the kinetic, thermodynamic and biophysical properties of an acylaminoacyl peptidase (AAP), from the thermophile Pyrococcus horikoshii (PhAAP). It was shown that the electrostatic environment of the catalytic site of PhAAP substantially influenced the pH dependence of the specificity rate constant (k(cat)/K(m)). However, 0.3 M NaCl, which depressed the electrostatic effects, simplified the complex pH-rate profile. The rate of formation of the enzyme-substrate complex (k(1)) was obtained from a non-linear Arrhenius plot. The lack of substrate leaving group effects indicated that k(1) is the rate determining step in the catalysis. DSC and CD measurements demonstrated that PhAAP displayed a stable structure in the catalytically competent pH range. It was shown that PhAAP is not just an acylaminoacyl peptidase, but it also has an endopeptidase activity and so differs from the mammalian AAPs. Size exclusion chromatography with PhAAP revealed a hexameric structure, which is unique among the known members of the prolyl oligopeptidase family that includes AAPs and suggests that its cellular function may be different from that of the dimeric AAP also found in the same organism.

Structural and kinetic contributions of the oxyanion binding site to the catalytic activity of acylaminoacyl peptidase.

It is widely accepted that the catalytic activity of serine proteases depends primarily on the Asp-His-Ser catalytic triad and other residues within the vicinity of this motif. Some of these residues form the oxyanion binding site that stabilizes the tetrahedral intermediate by hydrogen bonding to the negatively charged oxyanion. In acylaminoacyl peptidase from the thermophile Aeropyrum pernix, the main chain NH group of Gly369 is one of the hydrogen bond donors forming the oxyanion binding site. The side chain of His367, a conserved residue in acylaminoacyl peptidases across all species, fastens the loop holding Gly369. Determination of the crystal structure of the H367A mutant revealed that this loop, including Gly369, moves away considerably, accounting for the observed three orders of magnitude decrease in the specificity rate constant. For the wild-type enzyme ln(k(cat)/K(m)) vs. 1/T deviates from linearity indicating greater rate enhancement with increasing temperature for the dissociation of the enzyme-substrate complex compared with its decomposition to product. In contrast, the H367A variant provided a linear Arrhenius plot, and its reaction was associated with unfavourable entropy of activation. These results show that a residue relatively distant from the active site can significantly affect the catalytic activity of acylaminoacyl peptidase without changing the overall structure of the enzyme.

[Concomitant septic and gouty olecranon bursitis]. / Bursite olécranienne d'origine simultanément goutteuse et septique.

A case of coexistent acute gout and septic olecranon bursitis is presented. Our hypothesis is that asymptomatic monosodium urate crystals, possibly present in the bursa could secondarily been triggered by the infection.

The acylaminoacyl peptidase from Aeropyrum pernix K1 thought to be an exopeptidase displays endopeptidase activity.

Mammalian acylaminoacyl peptidase, a member of the prolyl oligopeptidase family of serine peptidases, is an exopeptidase, which removes acylated amino acid residues from the N terminus of oligopeptides. We have investigated the kinetics and inhibitor binding of the orthologous acylaminoacyl peptidase from the thermophile Aeropyrum pernix K1 (ApAAP). Complex pH-rate profiles were found with charged substrates, indicating a strong electrostatic effect in the surroundings of the active site. Unexpectedly, we have found that oligopeptides can be hydrolysed beyond the N-terminal peptide bond, demonstrating that ApAAP exhibits endopeptidase activity. It was thought that the enzyme is specific for hydrophobic amino acids, in particular phenylalanine, in accord with the non-polar S1 subsite of ApAAP. However, cleavage after an Ala residue contradicted this notion and demonstrated that P1 residues of different nature may bind to the S1 subsite depending on the remaining peptide residues. The crystal structures of the complexes formed between the enzyme and product-like inhibitors identified the oxyanion-binding site unambiguously and demonstrated that the phenylalanine ring of the P1 peptide residue assumes a position different from that established in a previous study, using 4-nitrophenylphosphate. We have found that the substrate-binding site extends beyond the S2 subsite, being capable of binding peptides with a longer N terminus. The S2 subsite displays a non-polar character, which is unique among the enzymes of this family. The S3 site was identified as a hydrophobic region that does not form hydrogen bonds with the inhibitor P3 residue. The enzyme-inhibitor complexes revealed that, upon ligand-binding, the S1 subsite undergoes significant conformational changes, demonstrating the plasticity of the specificity site.

Crystallization and preliminary crystallographic analysis of porcine acylaminoacyl peptidase.

Acylaminoacyl peptidase (also known as acylamino-acid-releasing enzyme or acylpeptide hydrolase; EC 3.4.19.1) is an unusual member of the prolyl oligopeptidase family catalysing the hydrolysis of an N-acylated peptide to an acylamino acid and a peptide with a free N-terminus. Acylaminoacyl peptidase purified from porcine liver has been crystallized in mother liquor containing 0.1 M Tris-HCl pH 7.0, 10%(w/v) polyethylene glycol 8000, 50 mM MgCl2 and 1%(w/v) CHAPS using the hanging-drop vapour-diffusion technique. A full data set to 3.4 A resolution was collected at ESRF beamline ID14-4 and space group C222 was assigned, with unit-cell parameters a = 84.8, b = 421.1, c = 212.0 A and four molecules in the asymmetric unit.

His507 of acylaminoacyl peptidase stabilizes the active site conformation, not the catalytic intermediate.

Acylaminoacyl peptidase is a member of the prolyl oligopeptidase family. Amino acid sequence alignment suggests that the stabilization of the tetrahedral intermediate should be mediated by His507 rather than by a tyrosine residue found in the other family members of this serine peptidase group. The pH dependence of k(cat)/K(m) did not reveal any effect of His507. Substitution of an alanine for His507 gave the same bell-shaped pH rate profile with the same pK(a) values (7.0 and 8.7). However, the value of the rate constant was 85 times lower with the modified enzyme, which indicated that His507 is an important residue that is probably involved in the formation of the 3-dimensional structure.

Regulation of de novo sphingolipid biosynthesis and the toxic consequences of its disruption.

Complex sphingolipids are 'built' on highly bioactive backbones (sphingoid bases and ceramides) that can cause cell death when the amounts are elevated by turnover of complex sphingolipids, disruption of normal sphingolipid metabolism, or over-induction of sphingolipid biosynthesis de novo. Under normal conditions, it appears that the bioactive intermediates of this pathway (3-ketosphinganine, sphinganine and ceramides) are kept at relatively low levels. Both the intrinsic activity of serine palmitoyltransferase (SPT) and the availability of its substrates (especially palmitoyl-CoA) can have toxic consequences for cells by increasing the production of cytotoxic intermediates. Recent work has also revealed that diverse agonists and stresses (cytokines, UV light, glucocorticoids, heat shock and toxic compounds) modulate SPT activity by induction of SPTLC2 gene transcription and/or post-translational modification. Mutation of the SPTLC1 component of SPT has also been shown to cause hereditary sensory neuropathy type I, possibly via aberrant oversynthesis of sphingolipids. Another key step of the pathway is the acylation of sphinganine (and sphingosine in the recycling pathway) by ceramide synthase, and up-regulation of this enzyme (or its inhibition to cause accumulation of sphinganine) can also be toxic for cells. Since it appears that most, if not all, tissues synthesize sphingolipids de novo, it may not be surprising that disruption of this pathway has been implicated in a wide spectrum of disease.