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author:("gibbons, J")
1.  The role of plasma membrane STIM1 and Ca2 +entry in platelet aggregation. STIM1 binds to novel proteins in human platelets 
Cellular Signalling  2014;26(3):502-511.
Ca2 + elevation is essential to platelet activation. STIM1 senses Ca2 + in the endoplasmic reticulum and activates Orai channels allowing store-operated Ca2 + entry (SOCE). STIM1 has also been reported to be present in the plasma membrane (PM) with its N-terminal region exposed to the outside medium but its role is not fully understood. We have examined the effects of the antibody GOK/STIM1, which recognises the N-terminal region of STIM1, on SOCE, agonist-stimulated Ca2 + entry, surface exposure, in vitro thrombus formation and aggregation in human platelets. We also determined novel binding partners of STIM1 using proteomics. The dialysed GOK/STIM1 antibody failed to reduced thapsigargin- and agonist-mediated Ca2 + entry in Fura2-labelled cells. Using flow cytometry we detect a portion of STIM1 to be surface-exposed. The dialysed GOK/STIM1 antibody reduced thrombus formation by whole blood on collagen-coated capillaries under flow and platelet aggregation induced by collagen. In immunoprecipitation experiments followed by proteomic analysis, STIM1 was found to extract a number of proteins including myosin, DOCK10, thrombospondin-1 and actin. These studies suggest that PM STIM1 may facilitate platelet activation by collagen through novel interactions at the plasma membrane while the essential Ca2 +-sensing role of STIM1 is served by the protein in the ER.
Highlights
•STIM1 promotes collagen induced platelet aggregation and thrombus formation.•In human platelets SOCE activates but is not essential for platelet aggregation.•Plasma membrane STIM1 may facilitate platelet activation independent of SOCE.
doi:10.1016/j.cellsig.2013.11.025
PMCID: PMC4062937  PMID: 24308967
STIM1, stromal interaction molecule 1; SOCE, store operated Ca2 + entry; DAG, 1,2-diacyl-sn-glycerol; PM, plasma membrane; TG, thapsigargin; OAG, 1-Oleoyl-2-acetyl-sn-glycerol; TRPC, transient receptor potential canonical; Aggregation; Ca2 + entry; Collagen; STIM1; Thrombospondin-1
3.  Hospital admissions before the age of 2 years in Western Australia. 
Archives of Disease in Childhood  1994;70(3):205-210.
A linked data file of birth records and hospital admissions was used to investigate inpatient hospital morbidity before 2 years of age for all non-Aboriginal and Aboriginal children born in Western Australia in 1986. Of the non-Aboriginal children, 31.8% were admitted to hospital at least once before the age of 2 years, with an overall admission rate of 526/1000 live births; the corresponding figures for Aboriginal children were 68.7% and 2797. The mean number of days in hospital for each non-Aboriginal child admitted was 7.4, and 26.5 for Aboriginal children. Of the total cohort, 21% of non-Aboriginal and 20% of Aboriginal children were admitted only once, and 4% of non-Aboriginal and 36% of Aboriginal children were admitted at least three times; 23% of non-Aboriginal and 24% of Aboriginal children were admitted for only one major disease category, and 1% of non-Aboriginal and 16% of Aboriginal children were in at least four categories. The highest admission rates and highest percentages of the cohort admitted were for gastrointestinal and respiratory diseases and social admissions. These results illustrate the importance for both descriptive and analytical research of relating admissions to hospital for the total population to the individual child, and of using clinically relevant disease classifications.
PMCID: PMC1029743  PMID: 8135564
4.  Evaluation of ultra-short dialysis. 
British Medical Journal  1979;1(6164):680.
PMCID: PMC1598268  PMID: 435716
5.  Regular short haemodialysis in end-stage renal failure. 
British Medical Journal  1975;3(5986):758-759.
A study was made of thrice weekly haemodialysis of 3-3 1/2 hours' duration using a large surface area dialyser in patients with end-stage renal failure. Body water, potassium, and blood pressure control were satisfactory and comparable with the more widely used long dialysis schedules (6-9 hours thrice weekly). Patient rehabilitation was improved overall and the regimen enabled the dialysis unit to treat more patients despite a reduction in technical and nursing staff. The technique proved inadequate, however, in two patients with an intercurrent infection, and more intensive dialysis in recommended in such cases.
PMCID: PMC1674658  PMID: 1174885
7.  Platelet endothelial cell adhesion molecule-1 regulates collagen-stimulated platelet function by modulating the association of phosphatidylinositol 3-kinase with Grb-2-associated binding protein-1 and linker for activation of T cells 
Background: Platelet activation by collagen depends on signals transduced by the glycoprotein (GP)VI–Fc receptor (FcR)γ-chain collagen receptor complex, which involves recruitment of phosphatidylinositol 3-kinase (PI3K) to phosphorylated tyrosines in the linker for activation of T cells (LAT). An interaction between the p85 regulatory subunit of PI3K and the scaffolding molecule Grb-2-associated binding protein-1 (Gab1), which is regulated by binding of the Src homology 2 domain-containing protein tyrosine phosphatase-2 (SHP-2) to Gab1, has been shown in other cell types to sustain PI3K activity to elicit cellular responses. Platelet endothelial cell adhesion molecule-1 (PECAM-1) functions as a negative regulator of platelet reactivity and thrombosis, at least in part by inhibiting GPVI–FcRγ-chain signaling via recruitment of SHP-2 to phosphorylated immunoreceptor tyrosine-based inhibitory motifs in PECAM-1. Objective: To investigate the possibility that PECAM-1 regulates the formation of the Gab1–p85 signaling complexes, and the potential effect of such interactions on GPVI-mediated platelet activation in platelets. Methods: The ability of PECAM-1 signaling to modulate the LAT signalosome was investigated with immunoblotting assays on human platelets and knockout mouse platelets. Results: PECAM-1-associated SHP-2 in collagen-stimulated platelets binds to p85, which results in diminished levels of association with both Gab1 and LAT and reduced collagen-stimulated PI3K signaling. We therefore propose that PECAM-1-mediated inhibition of GPVI-dependent platelet responses result, at least in part, from recruitment of SHP-2–p85 complexes to tyrosine-phosphorylated PECAM-1, which diminishes the association of PI3K with activatory signaling molecules, such as Gab1 and LAT.
doi:10.1111/j.1538-7836.2010.04025.x
PMCID: PMC3298659  PMID: 20723025
GPVI; inhibitory; ITIM; PECAM-1; signaling
8.  Non-genomic effects of PPARγ ligands: inhibition of GPVI-stimulated platelet activation 
Background
Peroxisome proliferator-activated receptor-γ (PPARγ) is expressed in human platelets although in the absence of genomic regulation in these cells, its functions are unclear.
Objective
In the present study, we aimed to demonstrate the ability of PPARγ ligands to modulate collagen-stimulated platelet function and suppress activation of the glycoprotein VI (GPVI) signaling pathway.
Methods
Washed platelets were stimulated with PPARγ ligands in the presence and absence of PPARγ antagonist GW9662 and collagen-induced aggregation was measured using optical aggregometry. Calcium levels were measured by spectrofluorimetry in Fura-2AM-loaded platelets and tyrosine phosphorylation levels of receptor-proximal components of the GPVI signaling pathway were measured using immunoblot analysis. The role of PPARγ agonists in thrombus formation was assessed using an in vitro model of thrombus formation under arterial flow conditions.
Results
PPARγ ligands inhibited collagen-stimulated platelet aggregation that was accompanied by a reduction in intracellular calcium mobilization and P-selectin exposure. PPARγ ligands inhibited thrombus formation under arterial flow conditions. The incorporation of GW9662 reversed the inhibitory actions of PPARγ agonists, implicating PPARγ in the effects observed. Furthermore, PPARγ ligands were found to inhibit tyrosine phosphorylation levels of multiple components of the GPVI signaling pathway. PPARγ was found to associate with Syk and LAT after platelet activation. This association was prevented by PPARγ agonists, indicating a potential mechanism for PPARγ function in collagen-stimulated platelet activation. Conclusions: PPARγ agonists inhibit the activation of collagen-stimulation of platelet function through modulation of early GPVI signalling.
doi:10.1111/j.1538-7836.2009.03732.x
PMCID: PMC3298645  PMID: 20040043
glycoprotein VI; nuclear receptor; platelets; signaling
9.  The platelet-surface thiol isomerase enzyme ERp57 modulates platelet function 
Background: Thiol isomerases are a family of endoplasmic reticulum enzymes which orchestrate redox-based modifications of protein disulphide bonds. Previous studies have identified important roles for the thiol isomerases PDI and ERp5 in the regulation of normal platelet function. Aim: Recently, we demonstrated the presence of a further five thiol isomerases at the platelet surface. In this report we aim to report the role of one of these enzymes – ERp57 in the regulation of platelet function. Methods/Results: Using enzyme activity function blocking antibodies, we demonstrate a role for ERp57 in platelet aggregation, dense granule secretion, fibrinogen binding, calcium mobilisation and thrombus formation under arterial conditions. In addition to the effects of ERp57 on isolated platelets, we observe the presence of ERp57 in the developing thrombus in vivo. Furthermore the inhibition of ERp57 function was found to reduce laser-injury induced arterial thrombus formation in a murine model of thrombosis. Conclusions: These data suggest that ERp57 is important for normal platelet function and opens up the possibility that the regulation of platelet function by a range of cell surface thiol isomerases may represent a broad paradigm for the regulation of haemostasis and thrombosis.
doi:10.1111/j.1538-7836.2011.04593.x
PMCID: PMC3444690  PMID: 22168334
ERp5; ERp57; PDI; platelet activation; redox regulation; thiol isomerase
10.  The nucleotide sequence of the nitrogen-regulation gene ntrA of Klebsiella pneumoniae and comparison with conserved features in bacterial RNA polymerase sigma factors. 
Nucleic Acids Research  1985;13(21):7607-7620.
The nucleotide sequence of the Klebsiella pneumoniae ntrA gene has been determined. NtrA encodes a 53,926 Dalton acidic polypeptide; a calculated molecular weight which is significantly lower than that determined by SDS polyacrylamide gel analysis. NtrA is followed by another open-reading frame (orf) of at least 75 amino acids. In the spacer region between ntrA and orf there are no apparent transcription termination or promoter sequences and therefore orf may be co-transcribed with ntrA. Previous authors have proposed that NtrA could act as an RNA polymerase sigma factor but the NtrA amino acid sequence does not show a high level of homology to any known sigma factor. However analysis of sequences of five sigma factors from E. coli and B. subtilis has identified two conserved sequences at the C-terminal end of all these polypeptides. These sequences resemble those found in known site-specific DNA-binding domains and may be involved in recognition of conserved -35 and -10 promoter sequences. A similar pair of sequences is present at the C-terminus of NtrA and could play a role in recognition of ntr-activatable promoters.
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PMCID: PMC322074  PMID: 2999700

Results 1-10 (10)