Pharmacist and Homeless Outreach Engagement and Non-medical Independent prescribing Rx (PHOENIx): a study protocol for a pilot randomised controlled trial.

Introduction: The number of people experiencing homelessness (PEH) is increasing worldwide. Systematic reviews show high levels of multimorbidity and mortality. Integrated health and social care outreach interventions may improve outcomes.

Determination of ten monohydroxylated polycyclic aromatic hydrocarbons by liquid-liquid extraction and liquid chromatography/tandem mass spectrometry.

The aim of this study is to develop and validate an analytical method for the quantitation of ten urinary monohydroxylated polycyclic aromatic hydrocarbons (OH-PAHs) through high pressure liquid chromatography/tandem mass spectrometry (HPLC/MS/MS). After enzymatic deconjugation, urine samples were extracted by liquid-liquid extraction (LLE) and OH-PAHs were analyzed by HPLC/MS/MS operated in negative electrospray ionization (ESI) and multiple reaction monitoring (MRM) mode. LLE was conducted with the solvent mixture of pentane and toluene, which reduced the matrix interferences and enhanced the method sensitivity significantly.

Measurements of polybrominated diphenyl ethers and polychlorinated biphenyls in a single drop of blood.

A quantitative method that requires only a small volume (50μL) of blood has been developed for the determination of polybrominated diphenyl ethers (PBDEs) and polychlorinated biphenyls (PCBs). Target analytes in both plasma sample (DBSV) and dried blood spot (DBS) were analyzed by a gas chromatography/high resolution mass spectrometer (GC/HRMS). Measurements of standard reference materials by the developed method were in agreement with those certified values.

Fast and simultaneous determination of urinary 8-hydroxy-2'-deoxyguanosine and ten monohydroxylated polycyclic aromatic hydrocarbons by liquid chromatography/tandem mass spectrometry.

8-Hydroxy-2'-deoxyguanosine (8-OHdG), a biomarker of oxidative DNA damage, has been extensively studied to assess human exposure to carcinogenic compounds. Previous studies have associated levels of human urinary hydroxylated polycyclic aromatic hydrocarbons (OH-PAHs) with those of 8-OHdG. However, measurements of OH-PAHs and 8-OHdG in urine are often conducted with two different analytical methods, which is both costly and time-consuming.

9-Aminoacridine peptide derivatives as versatile reporter systems for use in fluorescence lifetime assays.

A novel long lifetime fluorescence reporter based on 9-aminoacridine was designed, the lifetime of which can be modulated in a defined manner when in proximity to a tryptophan residue enabling fluorescence lifetime based biochemical assays to be configured.

The antimicrobial antiproteinase elafin binds to lipopolysaccharide and modulates macrophage responses.

Lipopolysaccharides (LPS) of the outer membrane of Gram-negative bacteria represent a primary target for innate immune responses. We demonstrate here that the antimicrobial/anti-neutrophil elastase full-length elafin (FL-EL) is able to bind both smooth and rough forms of LPS. The N-terminus was shown to bind both forms of LPS more avidly.

Essential role for proteinase-activated receptor-2 in arthritis.

Using physiological, pharmacological, and gene disruption approaches, we demonstrate that proteinase-activated receptor-2 (PAR-2) plays a pivotal role in mediating chronic inflammation. Using an adjuvant monoarthritis model of chronic inflammation, joint swelling was substantially inhibited in PAR-2-deficient mice, being reduced by more than fourfold compared with wild-type mice, with virtually no histological evidence of joint damage. Mice heterozygous for PAR-2 gene disruption showed an intermediate phenotype.

Manganese superoxide dismutase gene therapy protects against irradiation-induced cystitis.

Urinary bladder cystitis occurs in patients receiving radiation therapy for pelvic tumors. Radiation-induced formation of superoxide radicals is believed to damage the urothelium, exposing the underlying bladder smooth muscle to urine, culminating in nerve irritation and muscle dysfunction. We tested whether overexpression of MnSOD could decrease superoxide levels and protect the bladder from radiation damage.

Bladder permeability barrier: recovery from selective injury of surface epithelial cells.

The mammalian bladder maintains high electrochemical gradients between urine and blood, permitting the kidney to modify body chemistries through urinary excretion. To perform this function, the urothelium maintains a tight permeability barrier. When this barrier is damaged, leakage of urine components into the underlying bladder layers results, with symptoms of cystitis.

Design of ET(B) receptor agonists: NMR spectroscopic and conformational studies of ET7-21[Leu7, Aib11, Cys(Acm)15].

In a previous report we have shown that the endothelin-B receptor-selective linear endothelin peptide, ET-1[Cys (Acm)1,15, Ala3, Leu7, Aib11], folds into an alpha-helical conformation in a methanol-d3/water co-solvent [Hewage et al. (1998) FEBS Lett., 425, 234-238].

Mapping the active site of endothelin-converting enzyme-1 through subsite specificity and mutagenesis studies: a comparison with neprilysin.

Endothelin-converting enzyme-1 (ECE-1) is a membrane-bound zinc metallopeptidase that is homologous to neprilysin in amino acid sequence. A major in vivo function of ECE-1 is the generation of endothelin-1, a potent vasoconstrictor, from big endothelin-1. ECE-1 is also potentially involved in the processing or degradation of other peptide hormones.

Innate immune system damage in human immunodeficiency virus type 1 infection. Implications for acquired immunity and vaccine design.

HIV infection affects the innate as well as the acquired immune systems. Critically, it changes the function of macrophages, which link the innate and acquired responses through their ability to present antigen to CD4(+) T lymphocytes. Patients with HIV infection have a reduced capacity to deal with subsequent pathogen exposure and many suffer from chronic pulmonary infections.

Inhibition of the ubiquitin-proteasome system in Alzheimer's disease.

Alzheimer's disease is the most common cause of dementia in the elderly. Although several genetic defects have been identified in patients with a family history of this disease, the majority of cases involve individuals with no known genetic predisposition. A mutant form of ubiquitin, termed Ub(+1), has been selectively observed in the brains of Alzheimer's patients, including those with nonfamilial Alzheimer's disease, but it has been unclear why Ub(+1) expression should be deleterious.

Chemically synthesized ubiquitin extension proteins detect distinct catalytic capacities of deubiquitinating enzymes.

We have used solid-phase chemistry to synthesize proteins equivalent to a human ubiquitin precursor (ubiquitin-52-amino-acid ribosomal protein fusion; UBICEP52) and representative of isopeptide-linked ubiquitin-protein conjugates [ubiquitin-(epsilonN)-lysine]; these proteins were precisely cleaved by a purified recombinant Drosophila deubiquitinating enzyme (DUB), UCH-D. Along with the previously synthesized ubiquitin-(alphaN)-valine, these synthetic proteins were used as substrates to assess the catalytic capacities of a number of diverse DUBs expressed in Escherichia coli: human HAUSP; mouse Unp; and yeast Ubps 1p, 2p, 3p, 6p, 11p, and 15p and Yuh1p. Distinct specificities of these enzymes were detected; notably, in addition to UCH-D, isopeptidase activity [ubiquitin-(epsilonN)-lysine cleavage] was only associated with Yuh1p, Unp, Ubp1p, and Ubp2p.

Selective isopeptide bond formation: coupling ubiquitin(67-76) with histone 2A(114-128) by use of the transfer active ester condensation technique.

A peptide, ubiquitin(67-76)-histone 2A(114-128) fragment (UBH2AF), was synthesized by selective formation of an isopeptide bond between the C-terminus of ubiquitin(67-76) and the epsilon-amino group of lysine-119 in histone 2A(114-128) which contained 4 lysine residues at positions 118, 119, 125 and 127, respectively. The transfer active ester condensation technique, together with the Tnm amine protecting group, were used successfully in the peptide segment coupling reaction. [structure: see text]

A linear endothelin-1 analogue: solution structure of ET-1[Aib1,3,11,15, Nle7] by nuclear magnetic resonance spectroscopy and molecular modelling.

Two-dimensional nuclear magnetic resonance techniques and a combination of distance geometry and molecular dynamics calculations were utilised to determine the three dimensional solution structure of an ET-1 analogue, ET-1[Aib1,3,11,15, Nle7], in a methanol-d3/water co-solvent. The modelled structure shows that the peptide folds into a consistent alpha-helical conformation between residues Ser4-His16 while the C-terminus prefers no fixed conformation. Our studies confirm that the disulphide links which are normally associated with the endothelin family of neuropeptides are not important for the formation of a helical conformation in solution.

Comparative studies of Nsc and Fmoc as N(alpha)-protecting groups for SPPS.

2-(4-Nitrophenyl)sulfonylethoxycarbonyl (Nsc) is an alternative base-labile N(alpha)-protecting group to 9-fluorenylmethoxycarbonyl (Fmoc) for amino acids. The UV spectrum of the Nsc group exhibits moderate absorption at 380 nm which is excellent for real-time monitoring of the deprotection process. It also decreases the rearrangement of X-Asp, which can be a serious problem in SPPS.

Solution structure of a novel ETB receptor selective agonist ET1-21 [Cys(Acm)1,15, Aib3,11, Leu7] by nuclear magnetic resonance spectroscopy and molecular modelling.

The solution structure of a biologically active modified linear endothelin-1 analogue, ET1-21[Cys(Acm)1,15, Aib3,11, Leu7], has been determined for the first time by two-dimensional nuclear magnetic resonance spectroscopy in a methanol-d3/water solvent mixture. Out of approximately one hundred linear peptide analogues tested by biological assay, this peptide, together with a dozen others, showed significant ETB selective agonist activity. Here we report the solution structure of an ETB selective agonist of a full-length, synthetic linear endothelin analogue.

Electrophoretic separation of ubiquitin and single amino acid residue ubiquitin extensions using a commercial modified acrylamide gel electrophoresis system: an assay to determine catalytic capacities of deubiquitinating enzymes.

We have chemically synthesised a number of ubiquitin extension proteins, with carboxyl-terminal single amino acid residue extensions, to use as substrates to assess the catalytic capacities of deubiquitinating enzymes (DUBs). Here we describe a modified acrylamide gel electrophoresis system which allows separation of peptide- or isopeptide-linked ubiquitin-lysine from ubiquitin (77 and 76 residue proteins respectively) in only 2 h. Western blotting, using antibodies against ubiquitin, allows both substrate (i.

A functional, discontinuous HIV-1 gp120 C3/C4 domain-derived, branched, synthetic peptide that binds to CD4 and inhibits MIP-1alpha chemokine binding.

This paper describes a branched synthetic peptide [3.7] that incorporates sequence discontinuous residues of HIV-1 gp120 constant regions. The approach was to bring together residues of gp120 known to interact with human cell membranes such that the peptide could fold to mimic the native molecule.

Development of ET(B) selective agonists: solution structure of a linear endothelin-1 analogue, ET-1 [Cys(Acm)(1,15), Ala3, Leu7, dAsp8, Aib11].

The solution structure of a synthetic ET(B) selective agonist, ET-1[Cys(Acm)(1,15), Ala3, Leu7, dAsp8, Aib11] has been solved by 1H NMR and molecular modelling studies. Such solution structures of linear modified peptides in aqueous methanol are being used in an ongoing program of research designed to assist in an understanding of the basic structural requirements for the biological activity of vasoconstrictors. The resulting structure of this peptide is characterised by an alpha-helical conformation between residues Leu6-His16 and by N- and C-termini which assume no defined conformation.

Synthetic peptides representing discontinuous CD4 binding epitopes of HIV-1 gp120 that induce T cell apoptosis and block cell death induced by gp120.

A vaccine against HIV-1 virus would block initial infection and must target conserved residues. Since initial infection depends on binding of the viral envelope protein gp120 to CD4 on the cell surface, the CD4 binding site of gp120 is a target for vaccine design. To identify the optimal biologically active site, we synthesized a series of 32-mer peptides, based on conserved residues in the C3 and C4 regions of gp120.

Solution conformation of an ET(B) selective agonist, ET-1[Cys(Acm)1,15,Ala3,Leu7,Aib11], in CD3OH/H2O by 1H NMR and molecular modelling.

To understand the basic structural requirements for the biological activity of endothelin peptides, the solution structure of an ETB selective agonist, ET-1[Cys-(Acm)1,15, Ala3,Leu7,Aib11, was investigated by 1H NMR spectroscopy and molecular modelling. The structure is characterised by an alpha-helical conformation between residues Ser5-His16 but is undefined at both the N and C termini. To date, neither the solution structures of linear modified peptides nor the effects of a methanol/water solvent system have been examined for endothelin or endothelin-like peptides.