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P Nash

Publications and source records attributed to P Nash.

At least 37 records · Page 2Linked to original sources
PubMed2000

Application of linear free energy relationships to the serpin-proteinase inhibition mechanism.

Linear free energy relationships can be used to link the changes in rate constant for a reaction to changes in the equilibrium caused by alterations in structure. While they have most often been used in the analysis of chemical reactions, they have also been employed to resolve questions in enzymology and protein folding. Here we analyze the reaction of a serpin with a panel of six serine proteinases, and observe that a linear free energy relationship exists between the true second-order rate constant for reaction, k(inh), and the inhibition constant, K(I), indicating that formation of the covalent serpin-enzyme complex may be reversible.

Animals↗
PubMed2000

Post-translational modification of the myxoma-virus anti-inflammatory serpin SERP-1 by a virally encoded sialyltransferase.

SERP-1 is a secreted serpin (serine-proteinase inhibitor) encoded by myxoma virus, a poxvirus pathogen of rabbits. SERP-1 is required for myxoma-virus virulence, and the purified protein has been shown to possess independent anti-inflammatory activity in animal models of restenosis and antigen-induced arthritis. As an inhibitor of serine proteinases, SERP-1 acts against tissue-type plasminogen activator, urokinase-type plasminogen activator, plasmin, thrombin and Factor Xa. In the present study, examination of SERP-1 glycosylation-site mutants showed that the N-linked glycosylation of Asn(172) was essential for SERP-1 secretion, whereas mutation of Asn(99) decreased secretion efficiency, indicating that N-linked glycosylation plays an essential role in the processing and trafficking of SERP-1. Furthermore, comparison of SERP-1 from wild-type myxoma virus and a virus containing a targeted disruption of the MST3N sialyltransferase locus demonstrated that SERP-1 is specifically modified by this myxoma-virus-encoded sialyltransferase, and is thus the first reported viral protein shown to by modified by a virally encoded glycosyltransferase. Sialylation of SERP-1 by the MST3N gene product creates a uniquely charged species of secreted SERP-1 that is distinct from SERP-1 produced from other eukaryotic expression systems, though this has no apparent effect upon the kinetics of in vitro proteinase inhibition. Rather, the role of viral sialylation of SERP-1 likely relates to masking antigenicity or targeting SERP-1 to specific sites of action in vivo.

Amino Acid Sequence↗
PubMed2000

Inhibition of transplant vasculopathy in a rat aortic allograft model after infusion of anti-inflammatory viral serpin.

BACKGROUND: Transplant vasculopathy remains a difficult therapeutic problem, resulting in the majority of late cardiac graft losses. This chronic vascular disease is thought to be triggered by alloantigen-dependent and alloantigen-independent inflammatory factors. Despite improved 1-year survival, the incidence of transplant vasculopathy has not improved with current immunosuppressive protocols. Highly effective strategies have evolved in the large DNA viruses that shield infecting viruses from host inflammatory responses. Serp-1 is a secreted myxoma virus anti-inflammatory serine proteinase inhibitor. Serp-1 inhibits plasminogen activators in a manner similar to plasminogen activator inhibitor (PAI-1), a vascular protein that plays a pivotal regulatory role in vascular wound healing. In this study, we tested the ability of purified Serp-1 protein to ameliorate posttransplant vasculopathy after rat aortic allograft surgery. METHODS AND RESULTS: Serp-1 protein or controls were infused into 98 rats immediately after segmental aortic allograft transplantation. After either late (28 days, 64 rats) or early (12 to 48 hours, 24 rats) follow-up, transplanted aortic segments were harvested for morphological and immunohistochemical analysis. Significant reductions in intimal plaque growth (P<0.002) and mononuclear cell invasion (P<0.033) were detected after Serp-1 infusion at nanogram doses. Serp-1 reduced early macrophage (P<0.0016) and nonspecific lymphocyte (P<0.0179) invasion into medial and adventitial layers and inhibited associated depletion of medial smooth muscle cells (P<0.0006). CONCLUSIONS: Infusion of a viral anti-inflammatory serpin, Serp-1, significantly reduces early inflammatory responses and later luminal occlusion in a rat aortic allograft model.

Animals↗
PubMed2000

Transplant vasculopathy: viral anti-inflammatory serpin regulation of atherogenesis.

BACKGROUND: Surgical and ischemic injury to the artery wall initiates vascular wound-healing responses that stimulate atherosclerotic plaque growth. The plasminogen activators have cellular chemotactic, adhesion, and proteolytic activity. Serp-1 is a secreted myxoma virus glycoprotein serpin that binds and inhibits plasminogen activators. We have examined the effects of Serp-1 on plaque growth and inflammatory cell invasion in animal models after balloon injury and after aortic allograft transplant. METHODS: We used histologic analysis to assess 4 animal models of angioplasty-mediated injury and 2 models of aortic allograft transplant for intimal hyperplasia and cellular invasion. We assessed plasminogen activator (uPA and tPA) and inhibitor (PAI-1) expression in rat iliofemoral arteries after balloon injury using Western blot, enzyme activity, and quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: Plaque growth after balloon injury decreased after Serp-1 treatment in all balloon-injury models tested. Transplant vasculopathy also significantly decreased in 2 rat models of aortic allograft transplant. Infusion of a Serp-1 active site mutant, that lacked plasminogen activator inhibiting activity, did not inhibit plaque growth. Quantitative RT-PCR detected increased transcription of PAI-1 mRNA. Increased PAI-1 protein and enzyme-inhibitory activity was also detected in Serp-1-treated arteries by activity assay and Western blot. CONCLUSIONS: Thrombolytic serpins are central regulatory agents in vascular wound-healing responses. Investigation of the inhibitory mechanisms of viral serpins may provide new insights into atherogenesis.

Angioplasty, Balloon↗
PubMed2000

Investigation of an acute chemical incident: exposure to fluorinated hydrocarbons.

OBJECTIVES: To assess whether attendance at the site after an incident in a sewer was associated with symptoms in emergency personnel and whether the prevalence of symptoms was associated with estimated levels of exposure to any chemical hazard. METHODS: Symptoms experienced by people attending an incident involving two dead sewer workers suggested the presence of a chemical hazard, before environmental sampling confirmed any toxic agent. Self reported symptoms, estimated exposures, and biomarkers of exposure for likely agents from all 254 people who attended the incident and a referent occupational group matching the 83 emergency personnel who went to the Accident and Emergency department (A and E) in the first 48 hours were recorded. The prevalence of symptoms and concentrations of creatine phosphokinase in serum of the 83 early patients at A and E were compared with their referent occupational group. In all workers who attended the incident, the trends in symptom prevalences and concentrations of creatine phosphokinase in serum were examined by distance from the site and predefined exposure category. RESULTS: Among all workers who attended the incident, symptoms of shortness of breath and sore throat were significantly associated with indirect estimates of exposure but not associated with concentrations of creatine phosphokinase. Freon was detected in two blood samples. The early patients at A and E reported more symptoms than their matched reference group and their median concentrations of creatine phosphokinase were higher. CONCLUSIONS: The association between symptoms and concentrations of creatine phosphokinase with attendance at the site indicated the presence of a continuing hazard at the site and led to extra precautions being taken. Comparison values from the referent occupational group prevented unnecessary medical follow up.

Adult↗
PubMed2000

Optical detection of triggered atherosclerotic plaque disruption by fluorescence emission analysis.

Fluorescence emission analysis (FEA) has proven to be very sensitive for the detection of elastin, collagen and lipids, which are recognized as the major sources of autofluorescence in vascular tissues. FEA has also been reported to detect venous thromboemboli. In this paper we have tested the hypothesis that FEA can reproducibly detect in vivo and in vitro triggered plaque disruption and thrombosis in a rabbit model. Fluorescence emission (FE) spectra, recorded in vivo, detected Russell's viper venom (RVV)-induced transformation of atherosclerotic plaque. FE intensity at 410-490 nm 4 weeks after angioplasty was significantly lower (P < 0.0033 by analysis of variance) in RVV-treated rabbits when compared to control animals with stable plaque. FE spectral profile analyses also demonstrated a significant change in curve shape as demonstrated by polynomial regression analysis (R2 from 0.980 to 0.997). We have also demonstrated an excellent correlation between changes in FE intensity and the structural characteristics detected at different stages of "unstable atherosclerotic plaque" development using multiple regression analysis (R2 = 0.989). Thus, FEA applied in vivo is a sensitive and highly informative diagnostic technique for detection of triggered atherosclerotic plaque disruption and related structural changes, associated with plaque transformation, in a rabbit model.

Angioplasty, Balloon↗
PubMed2000

Myxoma virus expresses a TNF receptor homolog with two distinct functions.

Myxoma virus, a member of the poxvirus family of DNA viruses, encodes many virulence factors to combat and evade the host immune responses. Among the virus-encoded immuno-modulators is M-T2, a tumor necrosis factor receptor (TNF-R) homologue. M-T2 is secreted as monomeric and dimeric species that bind and inhibit rabbit TNF in a species-specific manner. Deletion analysis indicates that the anti-TNF function is mediated by the first three of four cysteine rich domains (CRDs) of M-T2. In addition, the intracellular form of M-T2 has the ability to block virus-induced apoptosis in lymphocytes, and the first two CRDs appear to be sufficient for this function. Although the mechanisms for the anti-TNF and anti-apoptotic functions of M-T2 are not yet fully defined, we postulate that these dual activities of M-T2 are mediated through different functional motifs and abrogate distinct cellular responses to virus infection.

Amino Acid Sequence↗
PubMed1999

Detection of transplant vasculopathy in a rat aortic allograft model by fluorescence spectroscopic optical analysis.

BACKGROUND AND OBJECTIVE: Transplant vasculopathy is a leading cause of late cardiac graft loss. We have examined laser-induced fluorescence (LIF) spectroscopy as an optical diagnostic tool for detection of intimal plaque development and inflammatory cellular invasion in a rat model of aortic allograft transplant. STUDY DESIGN/MATERIALS AND METHODS: Infrarenal aortic segments were transplanted from Lewis to Sprague Dawley rats. A range of vasculopathy development was produced by treatment with a viral anti-inflammatory protein. LIF spectra were recorded from the intima of aortic implants at 28 days. Fluorescence intensity was analyzed for correlation with vasculopathy development. RESULTS: Significant differences in LIF intensity at 400-450 nm (P < or = 0.05 by ANOVA) were detected. LIF emission was correlated with plaque growth (R2 = 0.980), vessel narrowing (R2 = 0.964), and cellular invasion (R2 = 0.971) by regression analysis. CONCLUSION: LIF optical analysis provides a nontraumatic diagnostic approach for detection of atherosclerosis prior to cardiac transplant or during development of vasculopathy after transplant.

Animals↗
PubMed1999

Immunomodulation by viruses: the myxoma virus story.

Myxoma virus is a poxvirus pathogen of rabbits that has evolved to replicate successfully in the presence of an active immune response by an infected host. To accomplish this, the virus has developed a variety of strategies to avoid detection by or obstruct specific aspects of the antiviral response whose consolidated action is antagonistic to virus survival. We describe two distinct viral strategies carried out by viral proteins with which myxoma virus subverts the host immune response. The first strategy is the production of virus-encoded proteins known as viroceptors or virokines that mimic host receptors or cytokines. These seek to actively block extracellular immune signals required for effective virus clearance and produce a local environment in the infected tissue that is "virus friendly". The second strategy, carried out by intracellular viral proteins, seeks to retard the innate antiviral responses such as apoptosis, and hinder attempts by the infected cell to communicate with the cellular arm of the immune system. By studying these viral strategies of immune evasion, the myxoma system can provide insights into virus-host interactions and also provide new insights into the complex immune system.

Amino Acid Sequence↗
PubMed1999

Children's fractures: a population based study.

OBJECTIVE: To measure the incidence of childhood fractures in a defined population. SETTING: Accident and emergency (A&E) departments covering the Swansea and Neath Port Talbot areas of South Wales in 1996. METHODS: Linkage of data from A&E departments with population data to produce fracture incidence rates by anatomical site and cause in children aged 0-14 years. RESULTS: During 1996, 2463 new fractures occurred in 2399 residents yielding a fracture rate of 36.1/1000 children. Fractures were more common in boys than girls and increased with age in both groups. Sports and leisure activities accounted for 36% of fractures, assaults for 3.5%, and road traffic accidents 1.4%. Fractures of the radius/ulna were most frequent (36%). CONCLUSIONS: The fracture rate in South Wales children is twice the rate reported in previous studies. Further research is required to elucidate the reasons behind this high rate. Many fractures could be prevented by the use of safer surfaces in school playgrounds, and wrist protection in in-line skaters and possibly in soccer players.

Adolescent↗
PubMed1998

Inhibitory specificity of the anti-inflammatory myxoma virus serpin, SERP-1.

SERP-1 is a myxoma virus-encoded serpin, secreted from infected cells, that is required for virulence and has anti-inflammatory activity. We report that purified recombinant SERP-1 forms SDS-stable complexes with urokinase-type plasminogen activator (uPA), tissue-type plasminogen activator (tPA), plasmin, thrombin, and factor Xa. N-terminal sequencing confirmed Arg319-Asn320 as the site of reaction. Mutation of these residues to Ala-Ala abolished inhibitory activity but had no effect on the specific cleavage at Thr315-Leu316 seen with elastase and with cathepsin G. Kinetic analysis of the reactions with uPA, tPA, plasmin, thrombin, Xa, and C1s showed second-order rate constants to vary over 3 logs, from kinh = 3 x 10(5) M-1 s-1 with thrombin to approximately 600 M-1 s-1 with C1s, while steady-state inhibition constants ranged from KI = 10 pM with thrombin to approximately 100 nM with C1s. Stoichiometries of inhibition varied between SI = 1.4 +/- 0.1 for uPA to SI = 13 +/- 3 for thrombin. Analysis of the variations in inhibition kinetics shows that when serpins act at low concentrations, comparable with the target protease or with KI (as appears likely for SERP-1 in vivo), inhibitory specificity becomes less dominated by kinh and is increasingly dependent on partitioning within the branched reaction mechanism and on the lifetime of the inhibited complex.

Amino Acid Sequence↗
PubMed1998

Virus-encoded receptors for cytokines and chemokines.

A number of viruses, particularly members of the poxvirus, herpesvirus and retrovirus families, have adapted to the vertebrate immune responses by capturing and modifying cellular genes which regulate the host immune system. Included among these host-derived virus genes are modified versions of receptors for cytokines or chemokines. Most of these receptor homologs, also called viroceptors, are either secreted glycoproteins or are located at the infected cell surface. Although these viroceptors can act in different ways, collectively they function by modifying the cytokine network to the advantage of the virus rather than the host.

Animals↗
PubMed1996

Virus-encoded serine proteinase inhibitor SERP-1 inhibits atherosclerotic plaque development after balloon angioplasty.

BACKGROUND: Recurrent atherosclerotic plaque growth, restenosis, is a significant clinical problem after interventional procedures. Initiation of restenosis involves activation of inflammatory and thrombotic cascades, which are regulated by serine proteinase enzymes and inhibitors. We have investigated the use of a viral serine proteinase inhibitor, SERP-1, to reduce plaque development after primary balloon angioplasty. This is the first experimental report of the use of a viral anti-inflammatory protein for the prevention of atherosclerosis. METHODS AND RESULTS: Seventy-four cholesterol-fed rabbits were treated with either local or systemic infusions of SERP-1 protein (or control solutions) after balloon-mediated injury. Sites of SERP-1 infusion in rabbits had dramatically reduced plaque compared with control infusions at the 4-week follow-up. At low-dose infusions (30 to 300 pg), only the primary infusion site had a demonstrable decrease in plaque, whereas at higher-dose infusions (> 3000 pg), a generalized reduction in plaque development was detected. An associated decrease in mononuclear cell infiltration of the arterial wall was detected after SERP-1 infusion within the first 24 hours. Infusion of an active-site mutant of SERP-1 (P1-P1', ala-ala) lacking serine proteinase inhibitory activity failed to prevent plaque growth. CONCLUSIONS: Purified SERP-1, a virus-encoded secreted glycoprotein, reduces plaque growth after primary balloon-mediated injury. Plaque development is decreased by inhibition of serine proteinase activity and is associated with a focal reduction in macrophage infiltration immediately after injury. Investigation of serine proteinase inhibitors may provide new insight into the regulation of arterial responses to injury.

Angioplasty, Balloon↗
PubMed1996

Ultrasonographic appearance of necrotizing gangrene: aid in early diagnosis.

In 2 seriously ill patients with scrotal swelling of uncertain origin, scrotal and perineal ultrasonography demonstrated gas in the soft tissue before crepitus was detected on physical examination. Necrotizing infections of the scrotum and perineum have characteristic ultrasonographic features that can facilitate earlier diagnosis and treatment.

Aged↗