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Biomedical subjects

P Harrison

Publications and source records attributed to P Harrison.

At least 55 records · Page 3Linked to original sources
PubMed2000

Mechanism of regulation of HGF/SF gene expression in fibroblasts by TGF-beta1.

The effect of transforming growth factor beta 1 (TGF-beta1) on levels of hepatocyte growth factor/scatter factor (HGF/SF) gene transcripts was investigated in the human lung embryonic fibroblast cell line, MRC-5. TGF-beta1 markedly reduced the expression of the 6. 0-kb and 3.0-kb HGF/SF mRNA, which encode full-length HGF/SF, but it had little effect on the expression of the alternatively spliced 1. 5-kb mRNA, which encodes NK2, a competitive HGF/SF antagonist. Using actinomycin D to block RNA synthesis, it was observed that TGF-beta1 had little effect on the stability of the 1.5-kb NK2 mRNA but increased the rate of degradation of the 6.0- and 3.0-kb HGF/SF mRNA transcripts by a mechanism that was dependent on new protein synthesis. TGF-beta1 minimally increased rather than reduced HGF/SF promoter activity in cells transiently transfected with chloramphenicol acetyltransferase (CAT) reporter genes driven by HGF/SF gene 5'-flanking sequences. In MRC-5 cells, TGF-beta1 modulates HGF/SF gene transcripts at the posttranscriptional level in order to favour expression of the 1.5-kb mRNA that encodes the truncated protein NK2.

3' Untranslated Regions↗
PubMed2000

Changes in gametogenesis and fecundity of acroporid corals that were exposed to elevated nitrogen and phosphorus during the ENCORE experiment.

Colonies of two scleractinian reef coral species, Acropora longicyathus and Acropora aspera were transplanted into patch reefs at One Tree Reef, Great Barrier Reef, Australia as part of the ENCORE experiment. These corals and colonies of A. aspera which were naturally present in the patch reefs were exposed to four treatments over two years: controls with normal seawater, elevated levels of nitrogen only, phosphorus only, or nitrogen plus phosphorus. These corals were sampled and used to determine whether gametogenic cycles and fecundity were affected by nutrient enrichment. Acropora longicyathus had a single annual gametogenic cycle. Corals exposed to elevated nitrogen produced significantly smaller and fewer eggs and contained less testes material than those which were not exposed to nitrogen. Exposure to elevated phosphorus only resulted in corals producing more but smaller eggs, and more testes material. Egg numbers of colonies from other treatments decreased as the gametogenic cycles continued, but those of the phosphorus colonies showed almost no reduction in egg numbers between the early and late stages of the gametogenic cycles. These results have important management implications for coral reefs as they demonstrate that small increases in concentrations of nitrogen and phosphorus can have severe effects on reproductive activity in these species of scleractinian corals.

Journal Article↗
PubMed2000

The interleukin-1 and interleukin-10 gene polymorphisms in primary sclerosing cholangitis: no associations with disease susceptibility/resistance.

BACKGROUND/AIMS: Although primary sclerosing cholangitis is believed to be an autoimmune disorder, no tissue-specific auto-antibodies have yet been identified, and the strongest support for an autoimmune aetiology comes from HLA-association studies. Three different HLA haplotypes are associated with susceptibility to the disease and one with protection from it. These HLA haplotypes, however, do not account for all of the disease risk and genes outside the HLA region may also have a role in disease pathogenesis. The aim of this study was to investigate, for the first time, polymorphic genes/sites within the interleukin-1 and interleukin-10 genes in a large well-characterised group of patients. METHODS: Ninety-six patients and 96 control subjects were studied. A single base-exchange polymorphism at position +3953 in the first exon of the IL-1B gene, a penta-allelic repeat sequence in the IL-1 receptor antagonist gene (IL-1RN) and three single base-exchange polymorphisms at positions -592, -819 and -1082 in the IL-10 gene promoter were determined by standard PCR-based techniques. RESULTS: Overall, there was no significant difference in the distribution of any of the IL-1B, IL-1RN or IL-10 alleles or genes sequences comparing patients and controls. In addition, there was no difference when the patients were stratified for the presence and absence of the HLA DRB1*0301 (DR3) allele or concurrent inflammatory bowel disease. CONCLUSION: Neither the IL-1B +3953, IL-1RN microsatellites polymorphisms on chromosome 2q13 nor the IL-10 -592, -819, -1082 promoter gene polymorphisms on chromosome 1q31-32 are associated with susceptibility or resistance to primary sclerosing cholangitis.

Adult↗
PubMed2000

Hepatic expression of hepatocyte growth factor gene mRNA in acute liver failure.

Hepatocyte growth factor plays a key role in liver regeneration but the role of liver in its synthesis in acute liver failure is unclear. We therefore measured hepatic expression of hepatocyte growth factor mRNA in this condition in comparison to H3 histone mRNA, a marker of cellular proliferation. Hepatocyte growth factor mRNA levels were quantified by specific RNase protection assay in nine patients with acute liver failure and found to be similar to those in six normal controls. Hepatocyte proliferation, as assessed by H3 histone mRNA expression, was not detected in normal liver but was present in six of nine patients with acute liver failure (P < 0.05) and was not correlated with expression of hepatocyte growth factor mRNA (rs = -0.28). Liver is unlikely to be the source of the high serum hepatocyte growth factor levels observed in acute liver failure.

Adolescent↗
PubMed2000

Immunoplatelet counting: a proposed new reference procedure.

Given the high degree of interoperator error and poor precision of manual platelet counting, it has recently been proposed that an immunoplatelet counting method could become the new reference procedure. Platelets are identified immunologically with a suitable monoclonal antibody, and the platelet count is derived from the ratio of fluorescent platelet events to collected red blood cell (RBC) events that are also counted by a reliable and calibrated standard impedance counter (RBC ratio). In this study, we have set up a rapid and simple method for immunoplatelet counting and simultaneously compared the RBC ratio with the bead ratio derived from two different preparations of commercial calibration beads (Trucount and FlowCount beads). Comparison of the level of imprecision of the RBC ratio with either the manual count or bead ratios revealed a superior coefficient of variation of < 5% even in samples with a platelet count < 20 x 10(9)/l. The RBC ratio correlated extremely well with the existing manual phase reference method (r2 = 0.93) and especially well with three different commercial impedance counters and a dual-angle optical counter (r2 = 0.98-0.99). However, at < 100 x 10(9)/l, the correlation of the RBC ratio with the dual-angle optical count (ADVIA 120) (r2 = 0.96) was superior to all impedance counters. This suggests that automated optical counting methods may be more accurate at determining platelet counts in thrombocytopenic samples. As the RBC ratio is rapid, cheap and relatively easy to perform, we propose that this method could replace the manual count as a new international reference method.

Calibration↗
PubMed2000

In vivo biotinylation studies: specificity of labelling of reticulated platelets by thiazole orange and mepacrine.

Animal in vivo biotinylation studies have demonstrated that thiazole orange (TO) labels the youngest cells in the circulation. TO has since been widely used for the measurement of reticulated platelets. As recent findings suggest that at high concentrations TO also labels platelet dense granules non-specifically, the value of previous work is unclear. Mepacrine also labels platelet dense granules and can detect storage pool defects. In this study, a mouse in vivo biotinylation model was used to determine the specificity of TO and mepacrine staining on platelets recently released into the circulation. The mean life span of biotin/TO (low), biotin/TO (high) and mepacrine/TO dual-positive platelets was 1.4 d (SD 0.5), 2.2 d (SD 0.2) and 2.3 d (SD 0.3) respectively (n = 6) compared with a life span for biotin-positive platelets of 4.9 d (SD 1.6). TO (low), TO (high) and mepacrine labelled 8.0% (SD 3.1), 43.9% (SD 8.3) and 40.0% (SD 9.9) of the total platelet population respectively (results of 30 samples from six mice), which decreased to 6.8% (SD 3. 9), 26.6% (SD 6.9) and 25.7% (SD 10.6) after thrombin degranulation. The shorter life span and lack of thrombin sensitivity of TO (low)-positive platelets, suggests that TO (low) measures reticulated platelets specifically. The comparative life spans and thrombin sensitivity of TO (high) and mepacrine-positive platelets suggest that TO (high) labels platelet dense granules. These data also suggest that dense granules are lost during platelet ageing.

Animals↗
PubMed2000

Von Willebrand factor--cleaving protease activity in congenital thrombotic thrombocytopenic purpura.

Thrombotic thrombocytopenic purpura (TTP) is characterized by microangiopathic haemolytic anaemia (MAHA), thrombocytopenia, fluctuating neurological impairment, renal dysfunction and fever. Both acquired and congenital forms are recognized. Recurrent episodes, which may be predictable (occurring every 21-28 d), are seen in congenital disease and may be treated by infusion with fresh-frozen plasma (FFP). Congenital TTP has recently been associated with deficiency of a novel von Willebrand factor (VWF)-cleaving protease. To investigate whether residual protease activity dictates clinical manifestations, we determined protease activity in three patients with congenital TTP of varying severity. Intrinsic VWF-cleaving protease activity of a range of plasma-derived products was also assessed as one patient had been successfully maintained for many years, initially using an intermediate-purity factor VIII concentrate (Kryobulin) and then cryoprecipitate. All three patients had a severe absolute deficiency of VWF-cleaving protease activity (< 3%) up to 5 months after clinical symptoms. Three relatives were also found to have a mild reduction in protease activity (25-50%). Nevertheless, the intrinsic VWF-cleaving protease activity of plasma-derived products correlated with their clinical efficacy: significant (100%) protease activity was found in FFP, cryosupernatant, solvent-detergent-treated plasma, cryoprecipitate and Kryobulin. Two clinically ineffective factor VIII products (Fahndi and Haemate P) possessed only low protease activity (6.25% and 12.5% respectively). Although this suggests that VWF-cleaving protease activity is central to the pathogenesis of congenital TTP, either small differences in protease activity below 3% or hitherto unknown factors have a profound influence on clinical phenotype. The possible use of factor VIII concentrates in the treatment of this condition also warrants further investigation.

ADAM Proteins↗
PubMed2000

New quantitative parameters on a recently introduced automated blood cell counter--the XE 2100.

The XE 2100 (Sysmex Corporation) is a cell counter that furthers the technology of fluorescent flow cytometry developed from the earlier range of Sysmex analysers. The new diagnostic features are a nucleated red cell count (NRBC), the ability to measure platelets by impedance as well as an 'optical' platelet count using a fluorescence dye and an immature granulocyte (IG) count. The NRBC count was highly correlated (r = 0.97) with the manual reference count. For counts below 100 x 109/l the 'optical' method and the immunocount gave good a correlation (r = 0.97) optical and impedance counts were also well correlated (r = 0.89). The use of the 'optical' platelet count significantly improves the reliability of low platelet counts. The IG count correlated with visual counts (r = 0.81) and allows the detection of immature cells at an earlier stage in the laboratory process. The introduction of fluorescent flow cytometric analysis allows extended quantification of additional cell populations and so potentially improves screening and monitoring of various pathological conditions.

Adult↗
PubMed1999

Novel cell lines for the analysis of preprotachykinin A gene expression identify a repressor domain 3' of the major transcriptional start site.

Until now, no clonal cells have been identified that support the expression of a marker gene expressed from the rat preprotachykinin A (rPPT) promoter. We have analysed recently available cell lines that are candidates for supporting reporter gene expression directed by the rPPT promoter. These are the neuronal-derived cell line NF2C and the pancreatic cell lines RINm5F and a derivative RIN-1027-B2. The NF2C line was derived from the brain homogenate of a transgenic animal in which a temperature-sensitive simian virus 40 large T antigen was expressed from a neurofilament promoter. All three lines are able to support expression of a reporter gene directed by a fragment of the 5' rPPT promoter. Analysis of reporter gene expression supported by various fragments of the rPPT promoter demonstrated that although -865 to +92 bp supported expression, the addition of fragments between +92 and +447 bp led to repression of expression. Subsequent analysis of reporter gene constructs microinjected into primary cultures of dorsal root ganglion neurons (DRG) confirmed the existence of this repressor domain. This repression could be relieved totally in both RIN cell lines and partly in NF2C cells by mutating residues between +373 and +396 bp. This indicates that these cell lines support PPT promoter activity similar to that observed in DRG and determines a novel repressor domain within the promoter.

Animals↗
PubMed1999

Platelet activation responses in vitro to human mast cell activation.

Mast cells are thought to play an important role in atherogenesis and plaque rupture, but their role in the subsequent platelet activation and thrombus formation is unclear. Tryptase positive cells (KU812T+) were established from the KU812 cell line as an in vitro model of human mast cells and used to study the effect of mast cell activation on human platelets. Overnight incubation of KU812T+ with IgE and subsequent challenge with anti-IgE caused the release of heparinoid substances which inhibited 1 microg/ml collagen-induced platelet aggregation. KU812T+ challenged with compound 48/80 produced a releasate that had no apparent heparinoid content but caused full platelet aggregation. These findings showed that, although activation of KU812T+ via FcepsilonR1 partially abrogated collagen-induced platelet aggregation, activation of the C5a receptor signalling pathway, by compound 48/80, caused the release of potent platelet-activating substances. This cell culture model offers a unique insight into the role of platelet-mast cell interactions in arterial thrombogenesis.

Cell Line↗
PubMed1999

Novel super-high affinity sheep monoclonal antibodies against CEA bind colon and lung adenocarcinoma.

We have developed technology to create monoclonal antibodies (MAbs) using lymphocytes from immunized sheep. The affinities of these sheep monoclonal antibodies (SMA) can be several orders of magnitude higher than mouse MAbs. This paper reports the development and validation of a modified enzyme-linked immunoadsorbent assay (ELISA) method to select high-affinity antibodies of the desired specificity at the first screen. Using this method, we have isolated high affinity SMA to carcinoembryonic antigen (CEA), a marker of colon cancer. Comparisons of our novel SMA with mouse MAbs using the new ELISA and BIAcore technology (BIAcore AB, Stevenage, Herts, UK) have confirmed that we have made super-high affinity antibodies to CEA. One of these has a t(1/2) for dissociation of 8 days, which could provide a longer therapeutic window than is available with murine monoclonals currently being used in the clinic. This antibody has a specific tissue staining profile; it thus appears to be an excellent candidate for use in the clinic.

Adenocarcinoma↗
PubMed1999

Performance of the platelet function analyser PFA-100 in testing abnormalities of primary haemostasis.

The PFA-100 device is a new instrument for the in-vitro testing of platelet function. Primary haemostasis is stimulated by recording the closure time taken for platelets to seal a 150 microm aperture in the centre of a membrane coated with collagen and either epinephrine or ADP. Patients with type 3 von Willebrand's disease (n = 4) all had infinitely prolonged closure times (> 200 s) with both types of cartridge. A patient with afibrinogenemia exhibited only slightly prolonged closure times of 111 and 166 s for the ADP and epinephrine membranes, respectively. Patients with Glanzmann's thrombasthenia (n = 6) and Bernard Soulier syndrome (n = 2) had grossly prolonged closure times (> 200 s) with both types of cartridges. These results confirmed that the PFA-100 system was highly dependent on normal von Willebrand factor, glycoprotein Ib and glycoprotein IIb/IIIa levels but not on plasma fibrinogen. Patients with storage pool disease (n = 6) and Hermansky Pudlak syndrome (n = 7) had prolonged closure times with the epinephrine cartridge. There was no evidence of enhanced platelet function in patients with antiphospholipid syndrome, in sickle-cell disease or thalassemia. However, ingestion of aspirin resulted in a near consistent and significant prolongation of the closure time for the epinephrine cartridge but not for the ADP cartridge in both normal subjects and patients. The test offers a reliable, reproducible, rapid and simple means of assessing high-shear platelet function in vitro.

Blood Platelets↗
PubMed1999

Gene transfer in the gastrointestinal tract.

The maximum in vivo transfer rate of plasmid pAMbeta1 in the gut was 0.03 transconjugant per recipient cell, and this rate could be simulated in vitro only by forced filter mating. Transfer was not detected in liquid culture matings. Our findings demonstrate that in vitro methods, such as forced filter mating and liquid mating, underestimate the in vivo rates of gene transfer.

Animals↗
PubMed1999

Consumption bomb.

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Conservation of Natural Resources↗
PubMed1998

Royal College debates whether MDs should promote moderate consumption of alcohol.

Emerging news about the potential beneficial effects of moderate alcohol consumption raises some interesting challenges for physicians, who often come face to face with problems created by alcohol. Physicians on the affirmative side won a debate on the pros and cons of moderate drinking that was held during this fall's Royal College meeting. Pam Harrison explains how they did it.

Alcohol Drinking↗