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Biomedical subjects

A MacLean

Publications and source records attributed to A MacLean.

At least 19 recordsLinked to original sources
PubMed2004

Acute hepatitis C virus seroconversion in a Scottish blood donor: HCV antigen is not comparable with HCV nucleic acid amplification technology screening.

BACKGROUND AND OBJECTIVES: This study was conducted to analyse the usefulness of hepatitis C virus (HCV) core antigen tests for the confirmation of HCV infection in a donor presenting as nucleic acid amplification technology (NAT) positive but negative for antibodies to HCV (anti-HCV). MATERIALS AND METHODS: Blood donations were screened, in parallel, for anti-HCV using the Abbott PRISM HCV Chemiluminescent immunoassay (ChLIA) and an 'in-house' HCV NAT (pools of up to 95 donations). An HCV NAT-positive antibody-negative donor was identified. Twelve follow-up samples were obtained and tested with various HCV antigen (including the recently marketed Trak-C second-generation assay) and HCV antibody assays. RESULTS: The single HCV NAT-positive, antibody-negative donation was identified from 1 117 681 donations screened in the 4-year period, July 1999 to June 2003. The index donation was positive by Ortho HCV core antigen enzyme immunoassay (EIA) and Ortho Trak-C (second-generation HCV core antigen EIA). An archive sample, taken 127 days prior to the index donation, was negative for all HCV markers. Subsequent samples demonstrated a loss of reactivity in the Ortho HCV core antigen EIA and reduced activity in the Ortho Trak-C until day 69. Immunoblot (Ortho RIBA-3) and HCV PRISM became positive on day 62, whilst Ortho HCV ELISA was not positive until day 132 or Biorad HCV ELISA until day 160. An alternative immunoblot (Innogenetics Innolia III) was positive from day 55. RNA levels fluctuated considerably during the follow-up period, being completely undetectable by routine screening methods at the time-point around seroconversion; subsequently, antibody was detected using all assays investigated. CONCLUSIONS: This HCV-converting blood donor provided a unique panel of samples for using to assess current (and future) HCV assay systems. The overall test results led to the conclusion that individual HCV antigen testing should not be considered as equivalent to HCV NAT minipool screening. Trak-C antigen testing may be considered as a suitable confirmatory assay for isolated HCV NAT reactivity.

Acute Disease↗
PubMed2003

A simple method for enriching populations of transfected CHO cells for cells of higher specific productivity.

To establish a simple and rapid method for the screening of stable recombinant Chinese hamster ovary (CHO) cell lines, we have developed a cell surface labeling technique using fluorescently tagged antibodies that bind to secreted target proteins at low temperature. Using fluorescence intensity as the sole criterion for selection of cells, we are able to enrich populations of highly productive cells using preparative flow cytometry sorting. Reiterative sorting based on selection of cells having the highest fluorescence intensity of cell surface labeled protein results in dramatic increases in specific cellular productivity. Using lymphotoxin-beta receptor IgG fusion protein as a model system, we have demonstrated a greater than 20-fold increase in specific productivity (0.49-11.5 pg cell(-1) day(-1)) (pcd) without the use of methotrexate (MTX)-mediated selection or amplification. In addition, the flow cytometry used to enrich for and clone high producer cell lines has reduced development time by more than 50% and the number of screening assays by more than 10-fold. When a transfected population of CHO cells expressing a humanized version of the murine monoclonal antibody (mAb) AQC2 directed against human alpha 1 beta 1 integrin was subjected to the same treatment, a 25-fold improvement in specific productivity (0.3-8.0 pcd) was observed. Furthermore, similar application of this technique to MTX-amplified clones resulted in up to 120-fold overall improvement in specific productivity (up to 42 pcd). Greater than 20 examples are also presented to demonstrate the robustness and performance of this technique.

Animals↗
PubMed2000

Toxicity evaluation of replication-competent herpes simplex virus (ICP 34.5 null mutant 1716) in patients with recurrent malignant glioma.

The herpes simplex virus (HSV) ICP34.5 null mutant 1716 replicates selectively in actively dividing cells and has been proposed as a potential treatment for cancer, particularly brain tumours. We present a clinical study to evaluate the safety of 1716 in patients with relapsed malignant glioma. Following intratumoural inoculation of doses up to 10(5) p.f.u., there was no induction of encephalitis, no adverse clinical symptoms, and no reactivation of latent HSV. Of nine patients treated, four are currently alive and well 14-24 months after 1716 administration. This study demonstrates the feasibility of using replication-competent HSV in human therapy.

Adult↗
PubMed1998

Mice lacking inducible nitric-oxide synthase are more susceptible to herpes simplex virus infection despite enhanced Th1 cell responses.

Mice deficient in the inducible nitric-oxide synthase (iNOS), constructed by gene-targeting, were significantly more susceptible to herpes simplex virus (HSV)-1 infection, displayed a delayed clearance of virus from the dorsal root ganglia (DRG) and exhibited an increase in the frequency of virus reactivation in DRG compared with similarly infected heterozygous mice. The infected iNOS-deficient mice developed enhanced Th1-type immune responses and their spleen cells produced higher concentrations of IL-12 than similarly infected heterozygous mice. This finding suggests that iNOS plays an important role in resistance against HSV-1 infection. Furthermore, nitric oxide (NO) may block the development of Th1 cells via inhibition of IL-12 synthesis and thereby play a role in immune regulation.

Animals↗
PubMed1997

The retrieval of thoracic organs: donor assessment and management.

The optimal management of the multi-organ donor is critical to the successful outcome of transplantation. It is a complex challenge demanding careful attention to detail, and requiring a shift in emphasis, since the pathophysiological processes have far reaching effects which many clinicians do not see on a day-to-day basis. The optimal management of haemodynamic and respiratory status is essential in order to maximise the yield of suitable thoracic donor organs, yet this process will also improve the condition of other organs at the time of procurement and thus enable prompt recovery of function following hepatic and renal transplantation. The process commences when a potential donor is identified, and is only complete after successful transplantation of all possible organs. In order to achieve this end, a dedicated, multi-disciplinary team is necessary, consisting not only of medical staff, but also support workers who organise logistics, and who play their own part in vital areas, such as transport of the donor team and organs. The co-ordinator's role is pivotal in bringing together, in harmony, teams from different centres. It is important to remember that the effort of every person involved in the management and procurement of donor organs is primarily directed towards maximisation of the donor pool, and that our main responsibility is to the recipients on our waiting lists.

Brain Death↗
PubMed1996

Herpes simplex 1716--an ICP 34.5 mutant--is severely replication restricted in human skin xenografts in vivo.

HSV-1716 is a replication-restricted, neuroattenuated ICP 34.5 gene mutant of herpes simplex virus type 1 (HSV-1). Because of the attenuated phenotype of ICP 34.5 mutants in rodent models of HSV disease, they have been promoted as potential vaccine strains and gene therapy vectors and have been used by us and others as therapeutic agents for the treatment of experimental malignant tumors. However, all data on the phenotype of HSV-1716 and other ICP 34.5 mutants are from animal model systems, while humans are the natural hosts of HSV-1. To achieve an initial characterization of the phenotype of 1716 in human tissue, we have studied its replication in mature human skin xenografts on SCID mice. We find that replication of 1716 is severely restricted in such human skin grafts relative to both parental wild-type HSV-1 strain 17+ and the HSV-1716 revertant virus 1716R, in which the 759-bp ICP 34.5 gene deletions have been repaired. Moreover, the replication of both 1716 and 17+ is significantly better in the human skin grafts than it is in mouse skin. The implications of these findings are discussed.

Animals↗
PubMed1996

Differential regulation of melatonin receptors in sheep, chicken and lizard brains by cholera and pertussis toxins and guanine nucleotides.

G-proteins define both the pharmacological characteristics and the signalling pathways of G-protein-coupled receptors. Melatonin receptors have been shown to belong to this class of receptors through their sensitivity to modulators of G-protein function. This study reveals that 2-125I-iodomelatonin (125I-MEL) binding to different target tissues is differentially affected by agents which disrupt the G-protein cycle. GTP gamma S, pertussis (PTX) and cholera (CTX) toxins each reduce 125I-MEL binding to ovine pars tuberalis (oPT) and lizard brain membranes, whereas chicken brain is affected only by GTP gamma S (guanosine 5'-O-(3-thiotriphosphate)) and CTX. In contrast, high affinity binding of 125I-MEL in the ovine hippocampus was not affected by any of these agents. This finding, together with the fact that neural binding sites of the sheep brain were found to have markedly lower molecular mass than those of the oPT on native gel electrophoresis (365 vs 525 kDa), suggests that the neural 125I-MEL binding sites in sheep may not be G-protein coupled. Pharmacologically, however, the binding sites in the hippocampus and oPT could not be distinguished using 11 analogues of melatonin. Therefore, these data support the notion not only of multiple forms of melatonin receptor/G-protein complex, but of high affinity binding sites for 125I-MEL which do not display sensitivity to guanine nucleotides.

Animals↗
PubMed1996

The distribution of immunocompetent cells in the genital tract of HIV-positive women.

OBJECTIVE: To study the distribution of immunocompetent cells in the female lower genital tract and to determine whether it is altered in HIV infection. METHODS: Colposcopically directed cervical biopsies were taken in the premenstrual phase of 10 HIV-positive women with sexually acquired infection, and matched HIV-negative controls. Peripheral blood was collected from the HIV-infected group for immune studies. Histological, immunohistochemical and double immunofluorescence techniques were applied to the biopsies to reveal immunocompetent cell distribution. Comparisons were made between the immune cell populations in the cervix in the HIV-positive and negative groups, and between the cervix and peripheral blood in the HIV-positive group. RESULTS: Cervical biopsies from HIV-positive women had significantly reduced Langerhans and plasma cell counts in the submucosa, but increased T lymphocyte counts compared with the HIV-negative group. There was an increase in CD8+ lymphocyte numbers in cervical biopsies of HIV-positive women, causing an inversion of the CD4/CD8 ratio. The majority of these CD8+ cells were 'memory' (CD45RO+) cells, with reduced proportions expressing perforin (cytoplasmic cytolytic granules) and TIA-1 (cytolytic granule-associated protein). Natural killer (CD57) cell markers were not detected. The levels of CD8+ cells expressing Bcl-2 (a gene product inhibiting cell apoptosis) in HIV-positive women with relatively high peripheral CD4 lymphocyte cell counts was higher than in HIV-negative women, but these levels fell with declining CD4 lymphocyte counts (< 300 x 10(6)/1). The reduction in CD4/CD8 ratio in the cervix occurred even with relatively high peripheral CD4 counts. CONCLUSIONS: HIV disease is associated with alterations in the proportions of immunocompetent cells in the cervix. There is an increase in CD8+ T-lymphocytes and evidence of reduced cytolytic capacity; thus, CD8+ cells may lack the ability to respond to viral infection. Reduction in Langerhans' and plasma cells may reflect loss of signals from CD4+ T cells. These findings may, in part, explain why HIV-infected women are susceptible to recurrent fungal and viral infections, even when peripheral immune function is intact. Further studies of immune cell function are urgently required to further elucidate these findings.

Adult↗
PubMed1996

Mature results of a randomized trial of two doses of cisplatin for the treatment of ovarian cancer. Scottish Gynecology Cancer Trials Group.

PURPOSE: In 1992, we reported the first results of a randomized study in ovarian cancer, comprising two doses of cisplatin and indicated a significant difference (P = .0008) in median survival. Four years later, we now describe the results of this trial. PATIENTS AND METHODS: After a median follow-up of 4 years and 9 months, 115 of 159 cases of advanced ovarian cancer, originally randomized to receive six cycles of cyclophosphamide 750 mg/m2 and either a high dose (HD) of 100 mg/m2 cisplatin or a low dose (LD) of 50 mg/m2 (LD) cisplatin, have now died. RESULTS: The overall survival for HD and LD patients is 32.4% and 26.6%, respectively, and the overall relative death rate is 0.68 (P = .043). This represents a reduction in overall benefit with longer follow-up compared with the first 2 years (relative death rate of 0.52). Toxicity, particularly neurotoxicity, is still evident in the fourth year (10/31 on HD compared with 1/24 on LD). CONCLUSION: Our recommended dose of cisplatin in combination schedule is therefore 75 mg/m2, representing the optimal balance between efficacy and toxicity.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1996

The ovine pars tuberalis secretes a factor(s) that regulates gene expression in both lactotropic and nonlactotropic pituitary cells.

The purpose of this study was to determine whether the cells of the ovine pars tuberalis (PT) secrete a factor(s) that can influence the activity of cells in the pars distalis (PD). By Northern blotting of total RNA isolated from PD cells that had been stimulated in the presence of cycloheximide (10 micrograms/ml), PT cell-conditioned medium was shown to induce a significant increase in the expression of the early response gene, c-fos, above both PD cell-conditioned and nonconditioned medium control levels (P < 0.05). Although forskolin (5 microM) induced a weak increase in c-fos expression in PD cells, the effect of PT medium conditioned in the presence of forskolin enhanced this expression more than additively (P < 0.05); furthermore, this effect was reversed by melatonin. These results are consistent with the release of a factor(s) from the PT, which for simplicity we have called tuberalin. This factor was released from PT cells in a time-dependent and cycloheximide-sensitive manner and was resistant to heating at 100 C for 10 min. Tuberalin activity could be size-fractionated using molecular size cut-off filters to produce activity in both the 1- to 10-kDa and more than 10-kDa size ranges. The activities in both of these fractions were sensitive to trypsin degradation and, therefore, appeared to be peptidergic. However, it was not clarified whether the biological activities were due to one or two components. Tuberalin also induced c-fos expression in other cell types, including GH3 and NIH3T3 cells. Dual labelling of PD cells by in situ hybridization using riboprobes for c-fos and PRL demonstrated that both the less than and more than 10-kDa fractions of tuberalin activated c-fos expression in some, but not all, lactotrophs in PD cell cultures, suggesting that a primary function of the PT is to regulate the activity of lactotrophs. This was supported further by enhanced secretion of PRL from PD cells in the presence of either PT-conditioned medium or PT cells in coculture. In addition, PT-conditioned medium was found to increase c-fos in a second cell type, which did not hybridize positively for PRL, indicating the existence of other endocrine interactions between the PT and PD.

3T3 Cells↗
PubMed1996

Regulation of the Mel 1a melatonin receptor mRNA and protein levels in the ovine pars tuberalis: evidence for a cyclic adenosine 3',5'-monophosphate-independent Mel 1a receptor coupling and an autoregulatory mechanism of expression.

The Mel 1a receptor gene has recently been cloned and sequenced, and its product has been found to represent a new class of the G protein-coupled receptor superfamily. A unique aspect of its endogenous hormonal ligand, melatonin, is the prolonged duration (16 h) required to initiate appropriate photoperiodic responses in the animal. This study provides the first data on the transcriptional and translational regulation of this receptor in primary cultures of its native tissue, the pars tuberalis of the ovine pituitary. Using RNAse protection assays we show Mel 1a mRNA expression is rapidly induced through changes in intracellular cAMP levels. Forskolin enhances Mel 1a mRNA expression, whereas melatonin reverses this effect. RNA stability is also a factor, as Mel 1a mRNA expression decreases rapidly in the presence of actinomycin D. Although the expression of receptor protein is sensitive to forskolin stimulation, the lag between induction of mRNA and protein suggests that other factors (e.g. RNA splicing) contribute to functional receptor expression. Furthermore we observed that both mRNA and receptor protein levels increase spontaneously during primary culture, and these rises could be blocked by melatonin. A spontaneous rise in Mel 1a mRNA can also occur in serum-depleted cultures. In the absence of corresponding changes in cAMP, these results suggest not only that the Mel 1a receptor can autoregulate its own expression by an undefined pathway, but also they provide the first evidence that melatonin can act via a cAMP-independent signal transduction pathway to repress transcription in this tissue.

Animals↗
PubMed1995

Effects of fosinopril on exercise tolerance and clinical deterioration in patients with chronic congestive heart failure not taking digitalis. Fosinopril Heart Failure Study Group.

A total of 241 men and women with mild to moderately severe chronic heart failure (New York Heart Association functional class II [90%] or III) and a mean (+/- SD) left ventricular ejection fraction of 25 +/- 7%, entered a 24-week, prospective, double-blind, placebo-controlled trial of 10 or 20 mg/day of fosinopril, a phosphinic acid angiotensin-converting enzyme inhibitor. Patients received concomitant diuretic therapy but not digitalis. Primary end points were mean change in maximal treadmill exercise time and occurrence of prospectively defined clinical events indicative of worsening heart failure (most to least severe): death, withdrawal for worsening heart failure, hospitalization for worsening heart failure, need for supplemental diuretic or emergency room visit for worsening heart failure, and no event. At study end point, treadmill exercise time had improved in the fosinopril versus the placebo group (+28.4 vs -13.5 seconds, p = 0.047). New York Heart Association functional class had improved at end point more frequently (24% vs 13%) and deteriorated less frequently (18% vs 32%) in the fosinopril group (p = 0.003). More patients treated with fosinopril (66% vs 50%) remained free of clinical events indicative of worsening heart failure, and fosinopril-treated patients had less severe clinical events (p = 0.004). Dyspnea, fatigue, and paroxysmal nocturnal dyspnea improved more often and worsened less often in this group (p < or = 0.002), and edema showed a trend toward improvement (p = 0.088). These clinical benefits did not require concomitant digitalis therapy. Fosinopril was associated with an acceptable safety profile.

Aged↗
PubMed1995

Fosinopril attenuates clinical deterioration and improves exercise tolerance in patients with heart failure. Fosinopril Efficacy/Safety Trial (FEST) Study Group.

This study was a 12-week, double-blind, placebo-controlled, multinational trial of fosinopril in 308 patients with mild to moderately severe heart failure (New York Heart Association [NYHA] functional class IIS 17%, IIM 48%, and III 35%; mean ejection fraction [+/-SD] 26.5% [+/-6.9%]; bicycle exercise duration 1 to 11 min). An initial dose of 10 mg once daily was titrated as tolerated to 40 mg once daily. Patients all received diuretic therapy; digoxin was optional. The primary endpoint was maximal bicycle exercise time; a secondary endpoint was occurrence of the following prospectively defined, ordered clinical events indicative of worsening heart failure: death, study discontinuation, hospitalization, emergency room visits, and need for supplemental diuretic. At study endpoint (last value obtained for each patient), bicycle exercise time increased more with fosinopril (38.1 s) than with placebo (23.5 s) (P = 0.101 by ANCOVA and 0.010 by prospectively defined dropout-adjusted endpoint analysis). More patients remained free of clinical events indicative of worsening heart failure when treated with fosinopril (89%) than with placebo (75%), and the worst events of fosinopril-treated patients tended to be less severe than those of placebo patients (P = 0.001). Analysis of the occurrence of individual clinical events showed that the need for supplemental diuretic was markedly reduced with fosinopril (8% vs 20%, of patients, P = 0.002), as were hospitalizations (3% vs 12% of patients, P = 0.002) and study discontinuations (2% vs 12% of patients, P < 0.001) for worsening heart failure; the two groups had similar incidences of death (3% of patients in the fosinopril group vs 2% in the placebo group, P = 0.723). In addition, symptoms of dyspnoea (P = 0.017), fatigue (P = 0.019), and NYHA functional class (P = 0.008) improved with fosinopril relative to placebo. In conclusion, fosinopril, at an initial dose of 10 mg once daily, subsequently titrated as tolerated to 40 mg once daily, increased exercise tolerance and reduced the frequency of clinical events indicative of worsening heart failure.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1995

Melatonin receptors couple through a cholera toxin-sensitive mechanism to inhibit cyclic AMP in the ovine pituitary.

The nature of melatonin receptor-G-protein coupling in ovine pars tuberalis (PT) cells of the pituitary was addressed using cholera (CTX) and pertussis (PTX) toxins. ADP-ribosylation of ovine PT membrane proteins using 32P-NAD in the presence of CTX radiolabelled several substrates including 44, 51, and 60 kD proteins. Each were clearly distinct from the 40 kD substrate radiolabelled in the presence of PTX. Acute incubation of PT membranes with either toxin reduced the number of high affinity binding sites for 125I-MEL, although the magnitude of the inhibition was much greater for CTX (56%) than for PTX (20%). A CTX-sensitive component also mediates the inhibition of forskolin-stimulated cyclic AMP accumulation as pre-treatment of PT cells with CTX (5 micrograms/ml) for 16 h blocked this response. Gs alpha is a major substrate for ADP-ribosylation by CTX, and 16 h pre-treatment of PT cells with CTX (5 micrograms/ml) caused a down-regulation of Gs alpha. Northern analysis showed only one major transcript of Gs alpha of about 2 kb, which would encompass all of the known splice variants of the Gs gene. Screening of a cDNA library from ovine PT for Gs-related genes and sequencing of clones, combined with RT-PCR of PT mRNA, revealed no novel products. On this basis it is concluded that the CTX substrate is unlikely to be a novel splice variant or related gene product of the Gs class of G-protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Diphosphate Ribose↗
PubMed1994

Evidence for multiple forms of melatonin receptor-G-protein complexes by solubilization and gel electrophoresis.

The daily production of melatonin from the pineal gland influences circadian and seasonal behaviour and physiology. To further understand how melatonin may function, it is important to characterize the receptor and signal transduction systems. Using the detergent digitonin, we were able to solubilize the receptor from the ovine pars tuberalis (PT) membrane. The receptor was isolated as a complex associated with its heterotrimeric G-protein. In the solubilized state, pre-bound 125I-2-iodomelatonin was stable at 4 degrees C, but was displaceable by GTP gamma S. The receptor-G-protein complex could be separated by molecular mass using native polyacrylamide gel electrophoresis. We demonstrate that the receptor-complex has a molecular mass of 525 kDa and differs from solubilized receptor-complexes isolated from either the lizard brain, chicken brain or the ovine hippocampus. Furthermore the receptor complex isolated from the hippocampus had the lowest molecular mass of these tissues (365 kDa) and was found not to be sensitive to GTP gamma S. This may indicate the existence of a distinct non-G-protein coupled form of the receptor.

Animals↗
PubMed1994

Intervening with high risk youth: a program model.

This paper describes a program model being implemented for older, high-risk adolescents who are beginning to exhibit significant substance use problems. The program provides an initial assessment and refers clients to appropriate services. Only a small percentage are being referred to inpatient treatment; most clients enter outpatient intervention groups, which are conducted by the program. These groups have three purposes: to allow staff to monitor and evaluate clients so that those who are in imminent danger can be placed in protective treatment settings; to help clients see themselves as candidates for recovery and to introduce them to treatment resources by accompanying them to AA and NA meetings; and to change behavior, a goal that seems more immediately achievable for youth with stable community ties who have more to lose if they get into further trouble. Besides describing the program model, this paper examines some of the difficulties of getting referrals from a variety of community agencies.

Adolescent↗