Enzymatic detoxification of gluten by germinating wheat proteases: Implications for new treatment of celiac disease

Introduction. Currently the only treatment for celiac disease is a lifelong gluten-free diet. The diet is, however, often burdensome, and thus new treatment options are warranted. We isolated proteases from germinating wheat grain naturally meant for total digestion of wheat storage proteins and investigated whether these enzymes can diminish toxic effects of gluten in vitro and ex vivo.

Methods. Pepsin and trypsin digested (PT) gliadin was pretreated with proteases from germinating wheat, whereafter the degradation was analyzed by HPLC-MS (high-performance liquid chromatography and mass spectroscopy) and sodium dodecyl sulphate polyacrylamide gel electrophoresis. The toxicity of cleaved PT-gliadin products was assessed in Caco-2 epithelial cells, celiac patient-derived T cells, and in human small intestinal mucosal organ culture biopsies.

Results. Proteases from germinating wheat degraded gliadin into small peptide fragments, which, unlike unprocessed PT-gliadin, did not increase epithelial permeability, induce cytoskeletal rearrangement or changes in ZO-1 expression in Caco-2 cells. Pretreated gliadin did not stimulate T cell proliferation in vitro or enhance the production of autoantibodies to culture supernatants and the activation of CD25+ lymphocytes in the organ culture to the same extent as unprocessed PT-gliadin.

Discussion. Germinating wheat enzymes reduce the toxicity of wheat gliadin in vitro and ex vivo. Further studies are justified to develop an alternative therapy for celiac disease.     

Acute Kidney Injury Following Aortic Valve Replacement in Patients Without Chronic Kidney Disease

BackgroundThe data on acute kidney injury (AKI) in patients without chronic kidney disease (CKD) after transcatheter aortic valve replacement (TAVR) are limited. The study sought to compare the incidence of AKI and its impact on 5-year mortality after TAVR and surgical aortic valve replacement (SAVR) in patients without CKD.MethodsThis registry included data from 6463 consecutive patients who underwent TAVR or SAVR. CKD was defined as estimated glomerular filtration rate <60 mL/min/1.73 m². AKI was defined according to the Kidney Disease Improving Global Outcomes criteria. For sensitivity analysis, propensity-score matching between TAVR and SAVR was performed.ResultsThe study included 4555 consecutive patients (TAVR, n = 1215 and SAVR, n = 3340) without CKD. Propensity-score matching identified 542 pairs. Patients who underwent TAVR had a significantly lower incidence of AKI in comparison to those who underwent SAVR (unmatched 4.7% vs 16.4%, P < 0.001, multivariable analysis: odds ratio, 0.29, 95% confidence interval [CI], 0.20-0.41; matched 5.9% vs 19.0%, P < 0.001). Patients with AKI had significantly increased 5-year mortality compared with those without AKI (unmatched 36.0% vs 19.1%, log-rank P < 0.001; matched 36.3% vs 24.0%, log-rank P < 0.001). The adjusted hazard ratios for 5-year mortality were 1.58 (95% CI, 1.20-2.08) for AKI grade 1, 3.27 (95% CI, 2.09-5.06) for grade 2, and 4.82 (95% CI, 2.93-8.04) for grade 3.ConclusionsTAVR in patients without CKD was associated with a significantly less frequent incidence of AKI compared with SAVR. AKI significantly increased the risk of 5-year mortality after either TAVR or SAVR, and increasing severity of AKI was incrementally associated with 5-year mortality.     

Molecular characterization of HetRV8-ir1, a partitivirus of the invasive conifer pathogenic fungus Heterobasidion irregulare

The Heterobasidion annosum (Fr.) Bref. complex includes some of the most destructive conifer pathogenic fungi in the Boreal forest region. H. irregulare, formerly known as the North American pine type of H. annosum, was introduced from North America into Italy during the Second World War and occurs as an invasive pathogen in Pinus pinea stands together with the native European species H. annosum sensu stricto. We describe the complete nucleotide sequence of a new putative partitivirus from an Italian strain of H. irregulare. The bisegmented genome of HetRV8-ir1 encodes an RNA-dependent RNA polymerase of 704 aa and a capsid protein of 638 aa. The polymerase and capsid aa sequences are relatively similar (59-78 %) to those of Fusarium poae virus 1, Pleurotus ostreatus virus 1, and grapevine-associated partitivirus 1. HetRV8-ir1 is the first virus described from H. irregulare, and it is distantly related to previously known partitiviruses of Heterobasidion species.     

Transfer of intestinal microbiota from lean donors increases insulin sensitivity in individuals with metabolic syndrome

Alterations in intestinal microbiota are associated with obesity and insulin resistance. We studied the effects of infusing intestinal microbiota from lean donors to male recipients with metabolic syndrome on the recipients' microbiota composition and glucose metabolism. Subjects were assigned randomly to groups that were given small intestinal infusions of allogenic or autologous microbiota. Six weeks after infusion of microbiota from lean donors, insulin sensitivity of recipients increased (median rate of glucose disappearance changed from 26.2 to 45.3 μmol/kg/min; P < .05) along with levels of butyrate-producing intestinal microbiota. Intestinal microbiota might be developed as therapeutic agents to increase insulin sensitivity in humans; www.trialregister.nl; registered at the Dutch Trial Register (NTR1776).     

Oral and salivary parameters in patients with rheumatic diseases

We studied the presence of secondary Sjögren's syndrome (SS) and the composition of saliva, prevalence of oral pathogens, periodontitis, mouth mucosa, and teeth in patients with various rheumatic diseases and in healthy controls. The hypothesis was that different rheumatic diseases might cause differences in oral health characteristics because of the liability of secondary SS in the patients. The study involved 77 patients and 77 age-matched and sex-matched controls. Twenty patients were suffering from spondylarthropathy (SPA), 18 from ankylosing spondylitis (AS), 24 from rheumatoid arthritis (RA), and 15 from mixed connective tissue disease (MCTD). Clinical and radiographic oral health status was recorded and salivary flow rates were measured. Selected salivary proteins and immunoglobulins were analysed by routine methods. Minor salivary gland biopsy samples were taken from the patients for assessment of inflammatory focus scores. Differences between patients and controls and in between the different rheumatic diseases were analysed statistically. Secondary SS was diagnosed in 39% (30/77) of the patients. A severe periodontal condition (community periodontal index of treatment needs score 3 or 4) occurred in 58% (45/77) of the rheumatic patients compared with only 26% (20/77) of the controls (p<0.0001). The severity of focal sialadenitis (focus score) correlated significant with salivary IgA, IgG, and IgM concentrations. Salivary albumin, total protein, IgG, and IgM concentrations were higher in all patient groups than in the controls. The number of patients with low salivary flow rates was higher in all patient groups compared to controls. Oral yeast counts were significantly higher in the patients than in the controls (p<0.001). In a subgroup analysis, patients with SS had higher values for salivary IgA and IgM than patients without SS. Dental caries and oral lactobacilli were more frequent in patients with SS, but SS was not associated with periodontitis. No major differences were noted in other salivary biochemical parameters between these two groups. Patients with rheumatic diseases, irrespective of specific diagnosis, thus had various alterations in salivary flow and composition and oral health. The findings may reflect the autoimmune inflammation of the salivary glands frequently observed in these patients.     

Predictors of mortality following primary hip and knee replacement in the aged: A single-center analysis of 1,998 primary hip and knee replacements for primary osteoarthritis

Background and purpose High age is associated with increased postoperative mortality, but the factors that predict mortality in older hip and knee replacement recipients are not known.

Methods Preoperative clinical and operative data on 1,998 primary total hip and knee replacements performed for osteoarthritis in patients aged ≥ 75 years in a single institution were collected from a joint replacement database and compoared with mortality data. Average follow-up was 4.2 (2.2–7.6) years for the patients who survived. Factors associated with mortality were analyzed using Cox regression analysis, with adjustment for age, sex, operated joint, laterality, and anesthesiological risk score.

Results Mortality was 0.15% at 30 days, 0.35% at 90 days, 1.60% at 1 year, 7.6% at 3 years, and 16% at 5 years, and was similar following hip and knee replacement. Higher age, male sex, American Society of Anesthesiologists risk score of > 2, use of walking aids, preoperative walking restriction (inability to walk or ability to walk indoors only, compared to ability to walk > 1 km), poor clinical condition preoperatively (based on clinical hip and knee scores or clinical severity of osteoarthritis), preoperative anemia, severe renal insufficiency, and use of blood transfusions were associated with higher mortality. High body mass index had a protective effect in patients after hip replacement.

Interpretation Postoperative mortality is low in healthy old joint replacement recipients. Comorbidities and functional limitations preoperatively are associated with higher mortality and warrant careful consideration before proceeding with joint replacement surgery.     

Automated GMP production and long-term experience in radiosynthesis of CB1 tracer [18F]FMPEP-d2

Here, we describe the development of an in-house-built device for the fully automated multistep synthesis of the cannabinoid CB₁ receptor imaging tracer (3R,5R)-5-(3-([¹⁸F]fluoromethoxy-d₂)phenyl)-3-(((R)-1-phenylethyl)amino)-1-(4-(trifluoromethyl)phenyl)pyrrolidin-2-one ([¹⁸F]FMPEP-d₂), following good manufacturing practices. The device is interfaced to a HPLC and a sterile filtration unit in a clean room hot cell. The synthesis involves the nucleophilic ¹⁸F-fluorination of an alkylating agent and its GC purification, the subsequent ¹⁸F-fluoroalkylation of a precursor molecule, the semipreparative HPLC purification of the ¹⁸F-fluoroalkylated product, and its formulation for injection. We have optimized the duration and temperature of the ¹⁸F-fluoroalkylation reaction and addressed the radiochemical stability of the formulated product. During the past 5 years (2013–2018), we have performed a total of 149 syntheses for clinical use with a 90% success rate. The activity yield of the formulated product has been 1.0 ± 0.4 GBq starting from 11 ± 2 GBq and the molar activity 600 ± 300 GBq/μmol at the end of synthesis.