SETDB1 is required for intestinal epithelial differentiation and the prevention of intestinal inflammation.

OBJECTIVE: The intestinal epithelium is a rapidly renewing tissue which plays central roles in nutrient uptake, barrier function and the prevention of intestinal inflammation. Control of epithelial differentiation is essential to these processes and is dependent on cell type-specific activity of transcription factors which bind to accessible chromatin. Here, we studied the role of SET Domain Bifurcated Histone Lysine Methyltransferase 1, also known as ESET (SETDB1), a histone H3K9 methyltransferase, in intestinal epithelial homeostasis and IBD. DESIGN: We investigated mice with constitutive and inducible intestinal epithelial deletion of Setdb1, studied the expression of SETDB1 in patients with IBD and mouse models of IBD, and investigated the abundance of SETDB1 variants in healthy individuals and patients with IBD. RESULTS: Deletion of intestinal epithelial Setdb1 in mice was associated with defects in intestinal epithelial differentiation, barrier disruption, inflammation and mortality. Mechanistic studies showed that loss of SETDB1 leads to de-silencing of endogenous retroviruses, DNA damage and intestinal epithelial cell death. Predicted loss-of-function variants in human SETDB1 were considerably less frequently observed than expected, consistent with a critical role of SETDB1 in human biology. While the vast majority of patients with IBD showed unimpaired mucosal SETDB1 expression, comparison of IBD and non-IBD exomes revealed over-representation of individual rare missense variants in SETDB1 in IBD, some of which are predicted to be associated with loss of function and may contribute to the pathogenesis of intestinal inflammation. CONCLUSION: SETDB1 plays an essential role in intestinal epithelial homeostasis. Future work is required to investigate whether rare variants in SETDB1 contribute to the pathogenesis of IBD.

Occupational allergy to phytase: case series of eight production workers exposed to animal feed additives.

BACKGROUND: In agricultural meat production, adding enzymes such as phytase to animal feed is widespread, but there is little awareness of the allergenic potential and health risks of these fungal enzymes. PATIENTS AND METHODS: We report on eight patients working in a plant producing phytase granulates. All patients complained about work-related rhinitis occurring within six months of the onset of exposure to phytase dust. Asthmatic symptoms and contact urticaria also occurred. To detect sensitizations to phytase, skin prick-, patch-, and basophil activation test were carried out with the factory product. Levels of IgE and IgG against phytase were also measured. RESULTS: There was a positive reaction to phytase with skin prick testing in seven of the eight patients. IgE specific to phytase was detectable in four of the eight patients, and IgG specific to phytase was detectable in six of the eight patients. The basophil activation test was positive in four out of seven patients tested, but the patch test was negative in all patients tested. Transfer to a different workplace with no exposure to phytase completely eliminated the symptoms. CONCLUSIONS: Mold enzymes such as phytase are highly potent occupational allergens. Occupational safety measures must be strictly implemented in order to protect the health of workers.

Safety and tolerability during build-up phase of a rush venom immunotherapy.

BACKGROUND: Safety and tolerability of venom immunotherapy (VIT) in patients with concomitant disease and comedications, especially ß-blockers (BBs) and angiotensin-converting enzyme inhibitors (ACEIs), are under discussion. OBJECTIVE: To identify risk factors for the occurrence of systemic reactions (SRs) during the build-up phase of a 3-day rush VIT with focus on comorbidities and related comedications. METHODS: Data of 175 three-day rush VIT courses (Vespula venom, n = 161; honeybee venom, n = 14) were analyzed. Starting with 0.02 µg of venom, the maintenance dose of 100 µg was reached in 15-dose increments within 3 days. Local reactions (LRs) and SRs were documented during the build-up phase. Comorbidities and related comedications were registered. Univariate, bivariate, and multivariate data analysis was performed. RESULTS: During the 3-day rush VIT, an LR was seen in 74 (42.3%) of 175, a large LR (>10-cm diameter) in 82 (46.9%) of 175, and SRs in 19 (10.9%) of 175 VIT courses. Multivariate logistic regression revealed that female sex (P = .01), immunotherapy with honeybee venom (P = .003), and accompanying ACEI medication (P = .03) were independent predictors of the development of SRs during the build-up phase of VIT. CONCLUSION: This study revealed that female sex, honeybee VIT, and ACEI use independent predictors for SRs.

Comprehensive evaluation of eight commercial SARS-CoV-2 IgG assays.

Sensitivity and specificity of serological assays are key parameters for the accurate estimation of SARS-CoV-2 sero-prevalence. The aim of this study was to compare 8 readily available IgG antibody tests using a panel of well-defined serum samples of prepandemic and pandemic origin. A cross-reaction panel included samples of patients with recent infection with either of the endemic Coronaviruses 229E, NL63, HKU1, or OC43. Additionally, samples with high antibody levels against influenza virus, adenovirus, and during acute EBV infection were included. Previous infection with endemic coronaviruses caused a significant amount of cross-reactivity in two of the assays. In contrast, the confidence intervals for the assays of Abbott, DiaSorin, Euroimmun and Roche encompassed the value of 98% for samples with a previous endemic HCoV infection. For all assays, sensitivities were between 91.3% and 98.8%. Assay performance was independent of the usage of either nucleocapsid or spike proteins.

Mast cells enhance sterile inflammation in chronic nonbacterial osteomyelitis.

Chronic nonbacterial osteomyelitis (CNO) is an autoinflammatory bone disease, and patients with active or recurrent bone inflammation at multiple sites are diagnosed with chronic recurrent multifocal osteomyelitis (CRMO). The Chronic multifocal osteomyelitis (CMO) mouse model develops IL-1ß-driven sterile bone lesions reminiscent of severe CRMO. The goal of this study was to evaluate the potential involvement of mast cells in CMO/CRMO. Here, we show that mast cells accumulate in inflamed tissues from CMO mice and that mast cell protease Mcpt1 can be detected in the peripheral blood. A transgenic model of connective tissue mast cell depletion (Mcpt5-Cre:Rosa26-Stopfl/fl-DTa) was crossed with CMO mice and the resulting mice (referred to as CMO/MC-) showed a significant delay in disease onset compared with age-matched CMO mice. At 5-6 months of age, CMO/MC- mice had fewer bone lesions and immune infiltration in the popliteal lymph nodes that drain the affected tissues. In bone marrow-derived mast cell cultures from CMO mice, cytokine production in response to the alarmin IL-33 was elevated compared with wild-type cultures. To test the relevance of mast cells to human CRMO, we tested serum samples from a cohort of healthy controls and from CRMO patients at diagnosis. Interestingly, mast cell chymase was elevated in CRMO patients as well as in patients with oligoarticular juvenile arthritis. Tryptase-positive mast cells were also detected in bone lesions from CRMO patients and patients with bacterial osteomyelitis. Together, our results identify mast cells as cellular contributors to bone inflammation in CMO/CRMO and provide rationale for further study of mast cells as therapeutic targets.

Exhaled breath condensate nitrite--methodological problems of sample collection.

BACKGROUND: Exhaled breath condensate (EBC) analysis is a promising new method to monitor airway inflammation, however there are still multiple open methodological questions. The purpose of this study was therefore to investigate methodological influences on nitrite in EBC (surface contamination, flow dependency, storage time and inter- and within day variability). MATERIAL/METHODS: EBC samples of 10 healthy children, 10 healthy adults and 71 asthmatic children were collected and nitrite was analysed using the Griess reaction. Collector devices where either air-dried after disinfection or stored in distilled water till usage. Repeatability measurements were performed at three different times during one day and on five consecutive days. Flow dependency was analysed at 200 and 400 ml/s expiratory flow and storage stability was investigated at several time points within the first three hours after collection of the sample. In a preliminary study EBC nitrite was analysed in asthmatic children of different severity. RESULTS: Surface contamination appears to have a profound influence on nitrite levels and both within-day and inter-day variability is high. We found no flow dependency, and storage stability was satisfying although with considerable variety in several samples. There were no significant differences between the nitrite levels of the different asthmatic subgroups nor between the asthmatic children and the controls. CONCLUSIONS: These data indicate that EBC nitrite is a substance with a wide variety of influencing factors and different sources of origin. This has to be kept in mind when using exhaled nitrite as a biomarker for airway inflammation.

Statins potentiate the IFN-gamma-induced upregulation of group IIA phospholipase A2 in human aortic smooth muscle cells and HepG2 hepatoma cells.

The present study shows that the incubation of human aortic smooth muscle cells (HASMC) and HepG2 cells with atorvastatin and mevastatin as HMG-CoA reductase inhibitors potentiated the interferon-gamma (INF-gamma)-induced group IIA phospholipase A(2) (sPLA(2)-IIA) expression in a dose- and time-dependent manner. The effect of statins on sPLA(2)-IIA expression was reduced by mevalonate, farnesyl pyrophosphate and geranylgeranyl pyrophosphate. Inversely, inhibitors of the farnesyl transferase and geranylgeranyl transferase-I mimicked the effects of statins. Clostridium difficile toxin B (TcdB), Y-27632 and H-1152, functioning as inhibitors of Rho proteins and Rho-associated kinase, also augmented the sPLA(2)-IIA expression in combination with IFN-gamma. The same effects were observed when inhibitors of mitogen-activated/extracellular response protein kinase kinase (MEK), PD98059 or U0126 were used. Further, the Janus kinase-2 (Jak2)-specific inhibitor, AG-490 and inhibitors of nuclear factor-kappaB (NFkappaB) abrogated the sPLA(2)-IIA elevating effects of statins, TcdB and PD98059 in the presence of IFN-gamma. This cytokine alone increased the NFkappaB p65 and CAAT-enhancer-binding protein-beta (C/EBP-beta) activity in HASMC nuclear extract, but only C/EBP-beta was further augmented when the cells were incubated in addition to IFN-gamma with atorvastatin, H-1152, PD98059 or U0126. Moreover, after the incubation of cells with atorvastatin and IFN-gamma the stability of sPLA-(2)IIA mRNA significantly increased in comparison to those after incubation with IFN-gamma alone. In conclusion, the obtained data suggest that (i) the expression of sPLA(2)-IIA is negatively regulated by RhoA/Rho-associated kinase and MEK/ERK signaling pathways and (ii) statins, because of their ability to down-regulate these pathways, can potentiate the IFN-gamma-induced sPLA(2)-II expression at transcriptional and post-transcriptional levels.

Determination of the cholesterol-collagen ratio of arterial atherosclerotic plaques using near infrared spectroscopy as a possible measure of plaque stability.

Particular danger associated with an arteriosclerotic plaque consists in the possible rupture of its cap, dependent on the thickness of the cap covering the lipid core, its composition and different inflammatory changes. The purpose of this study was to compare the total cholesterol and collagen contents of arterial walls, both measured by near infrared spectroscopy (NIRS), and to test whether the ratios of cholesterol to collagen correlate with histochemical parameters possibly being indicators for plaque stability. NIR spectra of 118 sections from 36 human aortas were measured at 1000-2500 nm. Evaluation was performed by the partial least squares method (PLS), the chemical reference analysis by HPLC. Acceptable results were achieved for calibrations. With these calibrations 38 further aortic sections taken at autopsy were NIR-spectroscopically analysed and ordered in relation to histological findings of fatty deposits, cap thickness over the lipid core, and the ratio of fatty deposits to cap thickness. Correlations were found to exist between the spectroscopically determined total cholesterol concentrations and the histologically estimated fatty deposits (r=0.887), between the spectroscopically determined collagen concentrations and the cap thickness over the lipid core (r=0.441), and between the ratios total cholesterol to collagen and the ratios fatty deposits to cap thickness (r=0.575).

Serum levels of secreted group IIA phospholipase A(2) in benign prostatic hyperplasia and prostate cancer: a biomarker for inflammation or neoplasia?

Secreted group IIA phospholipase A(2) (sPLA(2)-IIA) is markedly up-regulated in human prostate cancer (PCa) specimens and in some PCa-derived cell lines, indicating an important role of this enzyme in tumourigenesis. In this study, we measured levels of sPLA(2)-IIA, C-reactive protein (CRP), and prostate-specific antigen (PSA) in serum samples obtained from patients with benign prostatic hyperplasia (BPH) and with PCa of different stages. We found that serum levels of sPLA(2)-IIA and CRP in BPH and PCa patients were significantly elevated compared to those of healthy individuals, but the concentrations of these inflammatory biomarkers did not differ between patients with BPH or PCa. Furthermore, serum levels of sPLA(2)-IIA correlated with concentrations of CRP, but not with PSA, Gleason grade or tumour stage. In conclusion, these findings suggest that cancer-related changes are not exclusive factors contributing to elevated serum sPLA(2)-IIA levels and emphasize the utility of sPLA(2)-IIA as a circulating marker of inflammation in patients with BPH and PCa.

Utility of measuring vitamin B12 and its active fraction, holotranscobalamin, in neurological vitamin B12 deficiency syndromes.

Vitamin B(12) (VitB(12), cobalamin) deficiency has been associated with various neuropsychiatric conditions, such as peripheral neuropathy, subacute combined degeneration, affective disorders, and cognitive impairment. Current assays analyze vitamin B(12), of which only a small percentage is metabolically active. Measurement of its active fraction, holotranscobalamin, might be of greater relevance, but data in populations with neuropsychiatric populations are lacking. In this study, in order to validate VitB(12) and holotranscobalamin (holoTC) serum levels for the detection of VitB(12) deficiency in neuropsychiatric conditions, we compared the validity of VitB(12) and holoTC in a patient cohort with neuropsychiatric conditions suspicious for VitB(12) deficiency. The cohort included all patients admitted to the Department of Neurology at our university between 2005 and 2009 with at least two parameters of the VitB(12) metabolism available (n = 1,279). We used elevated methylmalonic acid as the external validation criterion for VitB(12) deficiency and restricted our analyses to subjects with normal renal function. Among all normal renal function patients, 13.2% had VitB(12) deficiency. In receiver operating characteristic curve (ROC) analysis, correlation of VitB(12) and holoTC with vitamin B(12) deficiency was generally weak, and the areas under the curve (AUC) were not significantly different for holoTC compared to vitamin B(12) in all subjects (AUC: 0.66 [95%CI: 0.51-0.82]; p = 0.04 vs. 0.72 [0.65-0.78], p < 0.0001) and in subcohorts of patients with classical VitB(12) deficiency syndromes. The positive predictive values for holoTC and vitamin B(12) were low (14.7 vs. 21.0%) and both were associated with more false-positive than true-positive test results. holoTC does not show superior diagnostic accuracy compared to VitB(12) for the detection of VitB(12) deficiency in subjects with neuropsychiatric conditions. Neither test can be recommended to diagnose VitB(12) deficiency in subjects with neuropsychiatric disorders.

Exposure to chromium, cobalt and molybdenum from metal-on-metal total hip replacement and hip resurfacing arthroplasty.

BACKGROUND: All metal implants--and metal-on-metal bearings in particular--corrode and cause a release of metal ions. Because cobalt and chromium have been shown to be carcinogenic and mutagenic in human and animal models, systemic toxicity and cancer risk are considered to be possible disadvantages of the metal-on-metal articulation. This study was designed to investigate the serum concentration profiles of chromium, cobalt and molybdenum after implantation of a Birmingham hip resurfacing arthroplasty (BHR) and a cementless total hip replacement with a 28-mm Metasul articulation (MTHR), over the first 2 years after implantation. METHODS: We analyzed profiles of metal ion serum levels in 111 patients implanted with a BHR, in 74 patients implanted with an MTHR, and in 130 implant-free probands control subjects using atomic absorption spectrophotometry. RESULTS: Chromium and cobalt concentrations (in microg/L) of all BHR and MTHR patients differed significantly from those of control subjects (chromium: < 0.25; cobalt: 0.25). The median chromium and cobalt concentrations in BHR patients had increased to 5.1 and 4.3 microg/L 2 years after surgery. Concentrations in BHR patient exceeded those in the unilateral MTHR patients. Molybdenum serum concentrations hardly changed over time in either group and were not significantly different from the concentrations seen in the control subjects. INTERPRETATION: During the first 2 years after surgery, the Birmingham hip resurfacing arthroplasty leads to a significantly greater increase in serum chromium and cobalt levels than the 28-mm metal-on-metal MTHR. Observation of patients over a longer period will be necessary in order to evaluate any chronic adverse effects to the system due to elevated chromium and cobalt serum concentrations.