Biomedical subjects

Paul J McLaughlin

Publications and source records attributed to Paul J McLaughlin.

6 recordsLinked to original sources

PubMed2005

Methylation: lost in hydroxylation?

Methylation of histone tails is a key determinant in forming active and silent states of chromatin. Histone methylation was regarded as irreversible until the recent identification of a lysine-specific histone demethylase (LSD1), which acts specifically on mono- and dimethylated histone H3 lysine 4. Here, we propose that the fission yeast protein Epe1 is a putative histone demethylase that could act by oxidative demethylation. Epe1 modulates the stability of silent chromatin and contains a JmjC domain. The Epe1 protein can be modelled onto the structure of the 2-oxoglutarate-Fe(II)-dependent dioxygenase, factor inhibiting hypoxia inducible factor (FIH), which is a protein hydroxylase that also contains a JmjC domain. Thus, Epe1 and certain other chromatin-associated JmjC-domain proteins may be protein hydroxylases that catalyse a novel histone modification. Another intriguing possibility is that, by hydroxylating the methyl groups, Epe1 and certain other JmjC-domain proteins may be able to demethylate mono-, di- or trimethylated histones.

Amino Acid Sequence↗

PubMed2004

HLA-DR regulation and the influence of GM-CSF on transcription, surface expression and shedding.

Low surface HLA-DR expression is a feature in sepsis. However, the mechanisms that regulate HLA-DR expression have not been elucidated. The current study investigates regulation of HLA-DR gene transcription, post transcriptional events and shedding of surface HLA-DR, as well as the regulation of HLA-DR by GM-CSF and an immunomodulatory cytokine. Plasma and PBMC were collected from septic patients and healthy volunteers. An ELISA was developed to measure soluble HLA. PCR techniques were used to determine HLA-DR mRNA levels, and flow cytometry and fluorescent microscopy were used for measurement of surface expressed and intracellular HLA-DR. Septic patients fulfilling the criteria of the American College of Chest Physicians (ACCP) for sepsis were recruited for the study (n=70). HLA-DR was measured on three consecutive days, days seven and fourteen. Patients were excluded from the study if on immunosuppressive therapy. Results: Higher levels of shed HLA-DR were found in the plasma of septic patients compared to healthy controls. The level of HLA-DR mRNA was significantly lower in septic patients compared to healthy controls, however an increased intracellular HLA-DR expression was observed. When HL-60 cells were treated with GM-CSF, gene transcription, surface expression and shedding of HLA-DR were all up-regulated. These results indicate that the mechanisms involved in the regulation of HLA-DR in sepsis include shedding of HLA-DR from the cell surface and regulation of HLA-DR gene transcription. Post-translational processing of HLA-DR was also seen to be compromised. GM-CSF was shown to regulate HLA-DR at all these levels.

Journal Article↗

PubMed2003

Is low monocyte HLA-DR expression helpful to predict outcome in severe sepsis?

OBJECTIVE: Low monocyte human leukocyte antigen-DR (HLA-DR) expression has been reported to be an indicator of poor survival in critically ill septic patients. We assessed its usefulness as a prognostic indicator in order to identify possible interventions to normalise HLA-DR expression in those patients with lowered monocyte HLA-DR. DESIGN: HLA-DR expression was measured on separated monocytes of septic patients, using flow cytometry, and HLA-DR upregulation was measured by the same techniques after ex vivo stimulation with granulocyte macrophage colony stimulating factor (GM-CSF). APACHE II score, age, sex and outcome were determined for all patients. SETTING: A single-centre study at the Royal Liverpool University Hospital in a medico-surgical 13-bed intensive care unit. PATIENTS AND PARTICIPANTS: All septic patients ( n=70) fulfilling the criteria of the ACCP for the diagnosis of sepsis were recruited for the study with informed consent from day 1 of diagnosis of sepsis and monocyte HLA-DR expression measured on 3 consecutive days. Patients were excluded from the study if they were on immunosuppressive therapy. Normal healthy volunteers ( n=45) were included. RESULTS: Low monocyte surface expression and median fluorescence density HLA-DR expression was not associated with a high mortality. High APACHE II scores were not correlated with low HLA-DR expression. However, in those patients where HLA-DR expression was lowered, this could be restored ex vivo by GM-CSF. CONCLUSIONS: In the group of septic patients under study, HLA-DR was not a useful prognostic marker of outcome. We did not find a higher mortality in the group of patients who had low expression. These findings are contradictory to some previously reported findings, and the possible reasons are discussed.

Adult↗

PubMed2003

Gelsolin domains 4-6 in active, actin-free conformation identifies sites of regulatory calcium ions.

Structural analysis of gelsolin domains 4-6 demonstrates that the two highest-affinity calcium ions that activate the molecule are in domains 5 and 6, one in each. An additional calcium site in domain 4 depends on subsequent actin binding and is seen only in the complex. The uncomplexed structure is primed to bind actin. Since the disposition of the three domains is similar in different crystal environments, either free or in complex with actin, the conformation in calcium is intrinsic to active gelsolin itself. Thus the actin-free structure shows that the structure with an actin monomer is a good model for an actin filament cap. The last 13 residues of domain 6 have been proposed to be a calcium-activated latch that, in the inhibited form only, links two halves of gelsolin. Comparison with the active structure shows that loosening of the latch contributes but is not central to activation. Calcium binding in domain 6 invokes a cascade of swapped ion-pairs. A basic residue swaps acidic binding partners to stabilise a straightened form of a helix that is kinked in inhibited gelsolin. The other end of the helix is connected by a loop to an edge beta-strand. In active gelsolin, an acidic residue in this helix breaks with its loop partner to form a new intrahelical ion-pairing, resulting in the breakage of the continuous sheet between domains 4 and 6, which is central to the inhibited conformation. A structural alignment of domain sequences provides a rationale to understand why the two calcium sites found here have the highest affinity amongst the five different candidate sites found in other gelsolin structures.

Actins↗

PubMed2002

Low plasma granulocyte-macrophage colony stimulating factor is an indicator of poor prognosis in sepsis.

OBJECTIVE: Monocyte dysfunction has been shown to be associated with adverse consequences in septic patients. The cytokine growth factor granulocyte-macrophage colony stimulating factor (GM-CSF) may be required for optimal monocyte function in these patients. The current study investigates whether plasma GM-CSF levels were significantly different in septic patients and whether there was an association with prognosis. DESIGN: Plasma samples were collected from all septic patients from day 1 of the diagnosis of sepsis for 3 days. Healthy volunteer plasma served as control samples. A novel enzyme-linked immuno-adsorbent assay was developed with suitable sensitivity for detection of GM-CSF in patient and normal plasma. APACHE II score, age, sex and outcome were determined for all patients. SETTING: A single centre study at the Royal Liverpool University Hospital in a medico-surgical 13 bed intensive care unit. PATIENTS: All septic patients (n = 53) fulfilling the criteria of the APCC for the diagnosis of sepsis, were recruited for the study with informed consent from day 1 of the diagnosis of sepsis and plasma GM-CSF measured on three consecutive days. Patients were excluded from the study if on immunosuppressive therapy. Normal healthy volunteers (n = 33) were included in the study to serve as controls. RESULTS: Plasma GM-CSF levels were statistically significantly depressed in patients who died compared with those who survived, who had levels comparable with healthy controls. CONCLUSIONS: The results indicate that low plasma GM-CSF is associated with adverse consequences for septic patients. The measurement of GM-CSF in the plasma of septic patients merits further study for use as a prognostic marker and also to identify the type of immunotherapy the patient may benefit from.

APACHE↗

PubMed2002

Characterisation of heterogeneous solid surfaces by multiple probe, temperature-programmed inverse gas chromatography (IGC). A feasibility study.

A method is described for the characterisation of the surfaces of powdered solids by temperature programmed, multiple-probe, inverse gas chromatography. This modification of "traditional" inverse gas chromatography (IGC) allows rapid screening of solid surfaces to compare the adsorptive behaviour of pairs or sets of surfaces. This feasibility study examined the criteria for selection of a suitable probe set and approaches to the analysis and interpretation of the data generated. The technique was used to compare carbon black samples, chosen for their surface heterogeneity and the difficulty of surface characterisation using other techniques. The choice of a low surface area, methylated silica solid diluent and the elimination of its influence on the measurements is discussed. The probe set selected readily distinguished the carbon black samples and allowed tentative conclusions to be drawn regarding steric and electronic influences on probe-solid interactions.

Journal Article↗