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Biomedical subjects

G A Rook

Publications and source records attributed to G A Rook.

At least 19 recordsLinked to original sources
PubMed2005

Smoking and tuberculosis: the epidemiological association and immunopathogenesis.

There is increasing evidence of a link between tuberculosis and smoking. This paper reviews the epidemiological evidence from the UK, China, India and the USA, summarizing some of the main papers which indicate an association. Where an association has been found there seems to be an increase in tuberculosis case rates of between two- and four-fold for those smoking in excess of 20 cigarettes a day, but it may be difficult to control for other factors, particularly alcohol consumption. The final part of the paper reviews possible mechanisms. A likely possibility is that nicotine turns off the production of TNF-alpha by the macrophages in the lungs, rendering the patient more susceptible to the development of progressive disease from latent Mycobacterium tuberculosis infection.

Adolescent↗
PubMed2001

Interleukin-4 and its alternatively spliced variant (IL-4delta2) in patients with atopic asthma.

The interleukin-4 (IL-4) splice variant (IL-4delta2) is known to antagonize many biological activities of IL-4, and this challenges our understanding of the role of IL-4 in asthma. Studies that have used nonspecific antibodies, probes, and/or primers to quantify IL-4 in clinical samples would not have distinguished the expression of IL-4 from IL-4delta2. This is the first study to examine patients with chronic asthma and atopy for IL-4delta2 mRNA in their peripheral blood mononuclear cells without antigen stimulation, using a quantitative nested reverse-transcription polymerase chain reaction (RT-PCR) protocol. The median IL-4 mRNA copy number in cells from the patients with asthma was 2.8 logs higher than in a comparator group of patients with tuberculosis (p = 0.0005) and 4.5 logs higher (p = 0.0004) than in healthy control subjects. In contrast, IL-4delta2 expression in cells from patients with asthma was similar to that seen in cells from patients with tuberculosis. Hence, the median ratio of IL-4 to IL-4delta2 was 500-fold higher in the patients with asthma when compared with either patients with tuberculosis or healthy control subjects. The relative expression of IL-4 and IL-4delta2 may be a reason for the functional diversity of Th2 cells in different clinical conditions, and a hitherto unexplored mechanism for the pulmonary pathology in patients with atopic asthma.

Adolescent↗
PubMed2001

Il-4 influences apoptosis of mycobacterium-reactive lymphocytes in the presence of TNF-alpha.

T cell apoptosis is associated with defective cell-mediated effector functions in several infectious diseases. In tuberculosis, there is evidence that T cell apoptosis may be cytokine mediated, but the mechanisms are not clearly understood. Type 2 cytokines have recently been associated with disease extent in human tuberculosis, but they have not previously been linked to apoptosis in mycobacterium-reactive T cells. This study presents evidence that PBLs from healthy donors respond to sonicated Mycobacterium tuberculosis Ags with increased IL-4 gene activation, CD30 expression, and apoptosis. The changes were significantly greater than those observed when cells were stimulated with Ags from nonpathogenic Mycobacterium vaccae. A hypothesis linking these observations was tested. CD30 expression and TNF-alpha-mediated lymphocyte apoptosis were both down-regulated by inhibiting IL-4 in this model. TNFR-associated factor 2 (TRAF2) expression was down-regulated in CD30(+) cells, and addition of anti-TNF-alpha Ab significantly reduced apoptosis in the CD30(+) but not the CD30(-) population. These observations support the hypothesis that increased IL-4 expression in M. tuberculosis-activated lymphocytes promotes CD30 expression, which sensitizes the lymphocytes to TNF-alpha-mediated apoptosis via TRAF2 depletion. This may be one mechanism by which IL-4 is associated with immunopathological consequences in human tuberculosis.

Antigens, Bacterial↗
PubMed2001

High levels of mRNA encoding IL-4 in unstimulated peripheral blood mononuclear cells from tuberculosis patients revealed by quantitative nested reverse transcriptase-polymerase chain reaction; correlations with serum IgE levels.

The dominant view has been that there is little or no activation of Type 2 cytokine production in human tuberculosis. A novel approach to quantitative nested reverse transcriptase-polymerase chain reaction has revealed that this conclusion was based on technical inadequacies of earlier studies, particularly the failure to discriminate between IL-4 and the IL-4 splice variant, IL4delta2. A new approach reveals that the largest cytokine change in tuberculosis is a 1-2 log increase in copy number for mRNAs encoding IL-4 and IL-13, accompanied by a small decrease in expression of mRNA encoding interferon-gamma. The increased IL-4 level correlates with disease severity and with serum levels of IgE and soluble CD30, and may be attributable to the recently observed increase in conversion of cortisone into cortisol in tuberculous lesions. The implications of these findings for pathogenesis, vaccine design and immunotherapy are discussed, as effective reagents will need to downregulate this inappropriate Th2 component.

Humans↗
PubMed2001

Advances in the immunopathogenesis of pulmonary tuberculosis.

Tuberculosis remains a global emergency because of our lack of understanding of the details of its pathogenesis. In the last 12 months there have been striking advances in the molecular genetics of the organism. Mutated strains of Mycobacterium tuberculosis have been used to study the genetic requirements for virulence and establishment of latency, and the biology of the interaction with host cells. Genes involved in lipid metabolism seem particularly important. The probable sites of latency within the host lungs have been identified by in situ polymerase chain reaction. The complex control by M. tuberculosis of apoptosis of T cells and macrophages has been somewhat clarified, and the data may suggest that M. tuberculosis causes death of a subset of T cells, while preserving some macrophages as hiding places with reduced microbicidal and antigen-presenting function. Similarly the demonstration of a very large relative increase in interleukin (IL)-4 and IL-13 expression, (together with IL-4delta2, the IL-4 splice variant), that correlates with lung damage, has been supported by data from flow cytometry and in situ hybridization, and indicates that a subversive T helper-2 (Th2) component in the response to M. tuberculosis may undermine the efficacy of immunity and contribute to immunopathology. Recently defined changes in metabolism of cortisol within the lesions may contribute to the development of the Th2 component. These findings underline the need to start testing vaccine candidates in models that mimic the situations in which bacille Calmette-Guerin fails, such as in the presence of latent infection, pre-existing Th2 responses to cross-reactive organisms, and stress.

Animals↗
PubMed2001

M. tuberculosis: immunology and vaccination.

Tuberculosis is increasing. Current treatment regimens require at least 6 months, because latent or stationary phase organisms are difficult to kill. Such regimens do not achieve full compliance, and "directly observed therapy short course" (DOTS) is having less impact than expected. This worrying situation is aggravated by coinfection with human immunodeficiency virus (HIV), and by the increase in drug-resistant strains. We need new insights that lead to more rapid therapies and immunotherapies, and more reliable vaccines. Recent insights have come from: understanding of the relationship between Mycobacterium tuberculosis and macrophages; the multiple T cell types that recognise mycobacterial peptides, lipids and glycolipids; the critical role of interferon-gamma (IFNgamma) and interleukin-12 (IL-12) in human mycobacterial infection revealed by genetically defective children; quantitation of the presence and importance of Th2 lymphocyte activation in human tuberculosis; the role of local conversion of inactive cortisone to active cortisol in the lesions; the recognition that some effective prophylactic vaccines also work as immumotherapeutics whereas others do not. In the longer term the recent sequencing of the M. tuberculosis genome will lead to further advances. In the short term, effective immunotherapy remains the most accessible breakthrough in the management of tuberculosis. The types of practical advance that will result from sequencing the genome are discussed speculatively, but cannot yet be predicted with certainty.

Animals↗
PubMed2000

Prevaccination with SRL172 (heat-killed Mycobacterium vaccae) inhibits experimental periodontal disease in Wistar rats.

Periodontal disease is a bacterial dental plaque-induced destructive inflammatory condition of the tooth-supporting tissues, which is thought to be mediated by T lymphocytes secreting T helper 2 (Th2) cytokines, resulting in recruitment of high numbers of antibody-producing B lymphocytes/plasma cells as well as polymorphonuclear leucocytes (PMN) secreting tissue-destructive components, such at matrix metalloproteinases and reactive oxygen metabolites into the gingival connective tissues. One treatment strategy may be to down-regulate the Th2 response to those dental plaque microorganisms which induce the destructive inflammatory response. In this study we have examined the effects of a potent down-regulator of Th2 responses on ligature-induced periodontal disease in an experimental rat model. A single s.c. injection into Wistar rats of 0.1 or 1 mg of SRL172, a preparation of heat-killed Mycobacterium vaccae (NCTC 11659), 13 days before application of the ligature, significantly reduced the subsequent destruction of the tooth-supporting tissues, as measured by loss of periodontal attachment fibres (P < 0.001) and bone (P < 0.002). This protective effect occurred not only on the experimental (ligatured) side but also on the control unligatured side. SRL172 has undergone extensive toxicological studies and safety assessments in humans, and it is suggested that it may provide a safe and novel therapeutic approach to periodontal disease.

Animals↗
PubMed2000

Interactions between hormone-mediated and vaccine-mediated immunotherapy for pulmonary tuberculosis in BALB/c mice.

Problems of logistics, compliance and drug resistance point to an urgent need for immunotherapeutic strategies capable of shortening the current 6-month chemotherapy regimens used to treat tuberculosis, or of supplementing ineffective therapy. In this study we sought to define the mechanism of action of two immunotherapies, both of which have previously been shown to prolong survival. Secondly, we wished to identify any clinically useful synergy between these therapies. In BALB/c mice infected via the trachea with Mycobacterium tuberculosis H37Rv there is an initial phase of partial resistance dominated by type 1 cytokines plus tumour necrosis factor-alpha (TNF-alpha) and interleukin-1 (IL-1), followed by a phase of progressive disease. This progressive phase is accompanied by increasing expression of IL-4, and diminished expression of IL-1 and TNF-alpha. Animals in this late progressive phase of the disease (day 60) were treated with two injections (day 60 and day 90) of 0.1 or 1.0 mg of heat-killed Mycobacterium vaccae, or with 3beta, 17beta-androstenediol (AED; 25 microg subcutaneously three times/week), or with both therapies. We show here using four techniques in parallel (morphometry, immunohistochemistry with automated cell counting, semiquantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assays of cytokines in lung extracts) that treatment with M. vaccae causes a switch back towards a type 1 cytokine profile, restoration of expression of IL-1alpha and TNF-alpha, and a switch from pneumonia to granuloma. This is very similar to the changes previously seen after treatment with AED. However, there was no evidence for synergy between M. vaccae and AED.

Androstenediol↗
PubMed2000

Type 2 cytokine gene activation and its relationship to extent of disease in patients with tuberculosis.

The extent of type 2 cytokine gene expression in patients with pulmonary tuberculosis (TB) was studied by use of quantitative nested reverse-transcription polymerase chain reaction on freshly isolated peripheral blood mononuclear cells. Interleukin (IL)-4 and IL-13 mRNA expression was significantly greater in patients-median mRNA copy numbers were 1.7 and 1.1 log10 higher, respectively-than in matched tuberculin-positive control subjects. Significant correlations with radiologic extent of disease and serum IgE levels supported the biologic significance of these results. Interferon-gamma mRNA copy numbers exceeded those of type 2 cytokines but were only marginally lower in patients than in control subjects. Gene expression of an IL-4 splice variant (IL-4delta2) was bimodally distributed in both patient and control groups. Patients with greater IL-4delta2 expression also expressed more IL-4 mRNA and had more extensive disease. Type 2 cytokines are associated with immunopathologic changes in TB patients but could be a cause or consequence of disease.

Adolescent↗
PubMed2000

Increased cortisol: cortisone ratio in acute pulmonary tuberculosis.

To evaluate a possible role for altered cortisol metabolism in mediating the immunoparesis associated with progressive tuberculosis (TB), we have studied the hypothalamic-pituitary-adrenal axis, and the activities of the 11beta-hydroxysteroid dehydrogenases (11-HSDs) that interconvert active cortisol and inactive cortisone. In active pulmonary tuberculosis (PTB), the ratio of cortisol/cortisone metabolites in 24-h urine showed a shift towards active cortisol (ratio, 1.19 +/- 0.1, n = 16 versus 0. 89 +/- 0.05 in cured pulmonary tuberculosis (CTB), n = 13, p < 0. 01; and 0.78 +/- 0.04 healthy volunteers (HV), n = 11, p < 0.005). Conversion of cortisone (administered as 25 mg orally) to cortisol in peripheral plasma was higher in PTB (peak 1,157 +/- 55 nM, n = 14 versus 862 +/- 50 nM in CTB, n = 10, p < 0.005, and 882 +/- 73 nM in HV, n = 10; p < 0.005). Cortisol/cortisone ratio was increased in bronchoalveolar lavage fluid in PTB (7.73 +/- 1.48, mean +/- SE, n = 13) compared with HV (4.05 +/- 0.38, n = 11, p < 0.05) but was not different in plasma (PTB, 3.25 +/- 0.68; HV, 4.01 +/- 0.92). Responses of plasma cortisol to dexamethasone, CRH stimulation, and multidose ACTH stimulation were not different. These data suggest that in pulmonary tuberculosis, central control of glucocorticoid production is normal but that peripheral metabolism, in particular in the lung, is deviated in favor of the active metabolite cortisol. This offers a possible mechanism to explain the immunoparesis observed in progressive pulmonary tuberculosis.

Adolescent↗
PubMed1999

A sensitive, non-radioactive quantitative method for measuring IL-4 and IL-4delta2 mRNA in unstimulated cells from multiple clinical samples, using nested RT-PCR.

Measurements of IL-4 mRNA directly from clinical samples are technically difficult as IL-4 is a low copy number cytokine. Moreover, most existing studies involving RT-PCR are confused by the use of primers which simultaneously amplify cDNA of IL-4 and its splice-variant (IL-4delta2). We describe a sensitive nested RT-PCR method to quantify mRNA expression of IL-4 and IL-4delta2 separately. It involves a simple method of generating cRNA standards without cloning. The use of external synthetic RNA standards, for which we validate that amplification kinetics are equivalent to the target, obviates the need for multiple sample dilutions. The assay is sensitive enough to measure IL-4 and IL-4delta2 mRNA expression in unstimulated PBMCs of normal subjects, and the reproducibility and throughput make this assay suitable for use in clinical studies with multiple samples.

Alternative Splicing↗
PubMed1999

Agalactosyl IgG is elevated in patients with active spondyloarthropathy.

Patients with rheumatoid arthritis or with Crohn's disease have deficient galactosylation of serum IgG [Gal(0)]. To study whether such deviation occurs in patients with spondyloarthropathy (SPA), we studied the percentage incidence of Gal(0) corrected for age [Gal(0)corr] in 47 SPA patients undergoing ileocolonoscopy and correlated the findings with clinical variables and prognosis of the patients. Gal(0)corr was elevated in 36% of the patients. Such patients had a higher number of inflamed joints (P < 0.02), higher ESR (P < 0.001) and CRP (P < 0.001). Elevated Gal(0)corr was also of prognostic significance: at 6-month follow-up those with elevated levels had a higher number of inflamed joints (P < 0.02) and ESR (P < 0.05). The presence of high Gal(0)corr did not associate with gut inflammation. In conclusion, a proportion of SPA patients has elevated levels of Gal(0), the amount of which correlates with severity of the disease and is a prognostic marker for chronicity of the disease.

Adult↗
PubMed1999

Possible improved survival of patients with stage IV AJCC melanoma receiving SRL 172 immunotherapy: correlation with induction of increased levels of intracellular interleukin-2 in peripheral blood lymphocytes.

PURPOSE: This phase I-II study was designed to assess safety and clinical efficacy of SRL 172 vaccine in patients with advanced stage IV (AJCC) malignant melanoma. Induction of intracellular cytokines (IL-2 and INF-gamma) in peripheral blood lymphocytes (PBLCs) from these patients was assayed and correlated to clinical outcome. PATIENTS AND METHODS: SRL 172 was administered intradermally to 24 patients with stage IV malignant melanoma, initially at 15-day intervals for three vaccinations and then at monthly intervals. Lymphocyte activation for cytokines in PBLCs was assayed prior to each vaccine administration using a FACS-based intracellular cytokine assay. Survival was compared to historical controls. RESULTS: The vaccination schedule resulted in sustained intracellular IL-2 induction in PBLCs in 9 of 23 patients (39%) who received at least three doses. Cytokine induction became apparent within the first three administrations of vaccine and was maximal at 8-12 weeks. Induction of intracellular IL-2 production (group 1) was associated with improved survival (P < 0.036). The median survival of the nine patients demonstrating IL-2 induction was 59 (95% confidence interval (95% CI): 47-71) weeks compared to 31 (95% CI: 18-44) weeks for the non-inducers. Induction of INF-gamma (group 2) was found in 10 patients and 6 patients had IL-2 and INF-gamma induction (group 3). There was no survival advantage for these patient groups. Although no objective responses were documented the group as a whole had a median survival of 44 (95% CI: 31-59) weeks which is better than that of historical controls. SRL 172 was safe and well tolerated. CONCLUSION: SRL 172 is effective in inducing intracellular IL-2 responses in PBLCs of a significant number of patients with stage IV (AJCC) melanoma. This is correlated with improved survival. The survival analysis is sufficiently encouraging to suggest that further prospective trials are justified. The methodology we present in this study may help in developing surrogate markers that will allow rational immunotherapeutic strategies to be designed for cancer patients.

Cancer Vaccines↗