Notch signalling is an evolutionarily conserved pathway involved in cell-fate specification. The initiating event in this pathway is the binding of a Notch receptor to a DSL (Delta/Serrate/Lag-2) ...ligand on neighbouring cells triggering the proteolytic cleavage of Notch within its extracellular juxtamembrane region; a process known as proteolytic 'shedding' and catalysed by members of the ADAM (a disintegrin and metalloproteinase) family of enzymes. Jagged1 is a Notch-binding DSL ligand which is also shed by an ADAM-like activity raising the possibility of bi-directional cell-cell Notch signalling. In the present study we have unequivocally identified the sheddase responsible for shedding Jagged1 as ADAM17, the activity of which has previously been shown to be localized within specialized microdomains of the cell membrane known as 'lipid rafts'. However, we have shown that replacing the transmembrane and cytosolic regions of Jagged1 with a GPI (glycosylphosphatidylinositol) anchor, thereby targeting the protein to lipid rafts, did not enhance its shedding. Furthermore, the Jagged1 holoprotein, its ADAM-cleaved C-terminal fragment and ADAM17 were not enriched in raft preparations devoid of contaminating non-raft proteins. We have also demonstrated that wild-type Jagged1 and a truncated polypeptide-anchored variant lacking the cytosolic domain were subject to similar constitutive and phorbol ester-regulated shedding. Collectively these data demonstrate that Jagged1 is shed by ADAM17 in a lipid-raft-independent manner, and that the cytosolic domain of the former protein is not a pre-requisite for either constitutive or regulated shedding.
Background
There is an excess of mortality in patients with rheumatoid arthritis (RA) but no long‐term Australian cohort data.
Aims
To determine median life years lost, all‐cause standardised ...mortality ratio (SMR) and cause‐specific SMR, their predictors and secular change in Australian patients with RA.
Methods
Study population was all patients seen by a rheumatologist between 1990 and 1994. Record linkage with Australian National Death Index was performed to determine fact and cause of death up to 2004. All‐cause and cause‐specific SMR, and median life years lost were determined.
Results
There were 35 (31%) deaths in the early 1990s cohort (n = 113), SMR 1.31 (95% 0.93, 1.80). There were 216 (44%) deaths in the pre‐1990s established cohort (n = 495), SMR 1.73 (1.49, 1.95). Median life years lost in the early cohort was 6 years for males and 7 years for females compared with 8 and 10 years, respectively, in the established cohort. Patients with low disease activity score at baseline (DAS < 3.2), SMR was 0.8 (0.3, 2.2) and 1.5 (1.1, 2.2) for the early and established cohorts, and if DAS ≥3.2, SMR was 1.4 (1.02, 1.98) and 1.8 (1.5, 2.1) respectively. Primary cause of death was cardiovascular disease (SMR 1.43 (1.17, 1.74). Patients at most risk were those age 45–54 years. RA was listed as a comorbid condition on the death certificate in only 16% of patients.
Conclusions
Within a period of 14 years, median life expectancy of patients with RA with disease onset in the early 1990s is reduced by 6–7 years. However, our results also suggest a secular reduction in excess mortality.
Β2-Glycoprotein I (β2GPI) is an important anti-thrombotic protein and is the major auto-antigen in the antiphospholipid syndrome (APS). The clinical relevance of nitrosative stress in post ...translational modification of β2GPI was examined.The effects of nitrated (n)β2GPI on its anti-thrombotic properties and its plasma levels in primary and secondary APS were determined with appropriate clinical control groups.
β2-glycoprotein I was nitrated at tyrosines 218, 275 and 309. β2-glycoprotein I binds to lipid peroxidation modified products through Domains IV and V. Nitrated β2GPI loses this binding (p < 0.05) and had diminished activity in inhibiting platelet adhesion to vWF under high shear flow (p < 0.01). Levels of nβ2GPI were increased in patients with primary APS compared to patients with either secondary APS (p < 0.05), autoimmune disease without APS (p < 0.05) or non-autoimmune patients with arterial thrombosis (p < 0.01) and healthy individuals (p < 0.05).In conclusion tyrosine nitration of plasma β2GPI is demonstrated and has important implications with regards to the pathophysiology of platelet mediated thrombosis in APS. Elevated plasma levels of nβ2GPI in primary APS may be a risk factor for thrombosis warranting further investigation.
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•The major autoantigen, B2-GPI in antiphospholipid syndrome can be nitrated.•Nitration of B2-GPI inhibits binding to lipid peroxidation products.•Nitrated B2-GPI loses its anti-thrombotic activity.•Nitrated B2-GPI is increased in patients with primary APS compared to secondary APS and autoimmune patients without APS.
Abstract
Objectives
To examine the incidence, predictors and outcomes associated with severe gastrointestinal (GI) disease in a large inception SSc cohort.
Methods
SSc subjects with <2 years of ...disease duration were identified from two multicentre cohorts. Severe GI disease was defined as: malabsorption, hyperalimentation, pseudo-obstruction and/or ⩾10% weight loss in association with the use of antibiotics for bacterial overgrowth or oesophageal stricture. Kaplan-Meier, multivariate logistic regression and Cox proportional hazard analyses were performed to determine the cumulative incidence rate, independent clinical correlates and mortality rate associated with severe GI disease. A longitudinal mixed model was used to assess the impact of severe GI disease on the Short Form Health Survey.
Results
In this inception SSc cohort, the probability of developing severe GI disease was estimated at 9.1% at 2 years and 16.0% at 4 years. In multivariate analysis, severe GI disease was associated with inflammatory myositis (odds ratio 4.68, 95% CI 1.65, 13.24), telangiectasias (odds ratio 2.45, 95% CI 1.19, 5.04) and modified Rodnan skin score (odds ratio 1.03, 95% CI 1.01, 1.07). Severe GI disease was associated with a >2-fold increase in the risk of death (hazard ratio 2.27, 95% CI 1.27, 4.09) and worse health-related quality of life Short Form Health Survey physical (β = −2.37, P = 0.02) and mental (β = −2.86, P = 0.01) component summary scores.
Conclusion
Severe GI disease is common in early SSc and is associated with significant morbidity and increased mortality. More research is needed to understand, prevent and mitigate severe GI disease in SSc.
A disintegrin and metalloproteinases (ADAMs) and matrix metalloproteinases (MMPs) are zinc metalloproteinases (ZMPs) that catalyze the "ectodomain shedding" of a range of cell surface proteins ...including signaling and adhesion molecules. These "sheddases" are associated with the invasion and metastasis of a range of cancers. Increased serum and tumor tissue levels of copper are also observed in several cancers, although little is known about how the metal might promote disease progression at the molecular level. In the current study, we investigated whether copper might regulate the ectodomain shedding of two key cell surface proteins implicated in the invasion and metastasis of prostate cancer, the Notch ligand Jagged1 and the adhesion molecule E-cadherin, and whether the metal was able to influence the invasion of the prostate cancer epithelial cell line PC3. Physiological copper concentrations stimulated the ZMP-mediated proteolysis of Jagged1 and E-cadherin in cell culture models, whereas other divalent metals had no effect. Copper-mediated Jagged1 proteolysis was also observed following the pretreatment of cells with cycloheximide and in a cell-free membrane system, indicating a posttranslational mechanism of sheddase activation. Finally, the concentrations of copper that stimulated ZMP-mediated protein shedding also enhanced PC3 invasion; an effect that could be negated using a sheddase inhibitor or copper chelators. Collectively, these data implicate copper as an important factor in promoting prostate cancer cell invasion and indicate that the selective posttranslational activation of ZMP-mediated protein shedding might play a role in this process.
A search for heavy neutral lepton (N) production in K+→e+N decays using the data sample collected by the NA62 experiment at CERN in 2017–2018 is reported. Upper limits of the extended neutrino mixing ...matrix element |Ue4|2 are established at the level of 10−9 over most of the accessible heavy neutral lepton mass range 144–462 MeV/c2, with the assumption that the lifetime exceeds 50 ns. These limits improve significantly upon those of previous production and decay searches. The |Ue4|2 range favoured by Big Bang Nucleosynthesis is excluded up to a mass of about 340 MeV/c2.
The NA62 experiment at CERN reports searches for K+→μ+N and K+→μ+νX decays, where N and X are massive invisible particles, using the 2016–2018 data set. The N particle is assumed to be a heavy ...neutral lepton, and the results are expressed as upper limits of O(10−8) of the neutrino mixing parameter |Uμ4|2 for N masses in the range 200–384 MeV/c2 and lifetime exceeding 50 ns. The X particle is considered a scalar or vector hidden sector mediator decaying to an invisible final state, and upper limits of the decay branching fraction for X masses in the range 10–370 MeV/c2 are reported for the first time, ranging from O(10−5) to O(10−7). An improved upper limit of 1.0×10−6 is established at 90% CL on the K+→μ+ννν¯ branching fraction.
The structural stability of the
Haemophilus influenzae type b (Hib) capsular polysaccharide, polyribosylribitolphosphate (PRP) in an aluminum hydroxide adsorbed, polysaccharide–protein conjugate ...vaccine was monitored using modifications of an HPLC assay developed by Tsai et al. Tsai C-M, Gu X-X, Byrd RA. Quantification of polysaccharide in
Haemophilus influenzae type b conjugate and polysaccharide vaccines by high-performance anion-exchange chromatography with pulsed amperometric detection. Vaccine 1993;12:700–706.. As applied to products containing PRP conjugated to the outer membrane protein complex (OMPC) from
Neisseria meningitidis, this assay allows direct measurement of the total PRP content in very complex samples including commercial vaccine products. In addition, with the use of a high-speed centrifugation step, the assay can be used to directly quantify any PRP that is not conjugated to the OMPC carrier protein. These results provide evidence of what appears to be a catalytic reaction taking place between the phosphodiester bond of PRP and the aluminum hydroxide adjuvant that results in hydrolysis of the PRP polymer into smaller chain lengths and liberation of PRP oligomers from the conjugate particle. The reaction approaches an asymptotic limit after approximately two years at 2–8°C. Clinical studies which span this time period confirm that the modest decrease in conjugated PRP content over time does not impact the overall clinical effectiveness of PRP–OMPC-containing vaccines.
A search for heavy neutral lepton production in K+ decays using a data sample collected with a minimum bias trigger by the NA62 experiment at CERN in 2015 is reported. Upper limits at the 10−7 to ...10−6 level are established on the elements of the extended neutrino mixing matrix |Ue4|2 and |Uμ4|2 for heavy neutral lepton mass in the ranges 170–448 MeV/c2 and 250–373 MeV/c2, respectively. This improves on the previous limits from HNL production searches over the whole mass range considered for |Ue4|2, and above 300 MeV/c2 for |Uμ4|2.
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