Evaluation of cadmium, lead, nickel and zinc status in biological samples of smokers and nonsmokers hypertensive patients

The objective of this study was to evaluate the association between trace and toxic elements zinc (Zn), cadmium (Cd), nickel (Ni) and lead (Pb) in biological samples (scalp hair, blood and urine) of smoker and nonsmoker hypertensive patients (n=457), residents of Hyderabad, Pakistan. For the purpose of comparison, the biological samples of age-matched healthy controls were selected as referents. The concentrations of trace and toxic elements were measured by atomic absorption spectrophotometer prior to microwave-assisted acid digestion. The validity and accuracy of the methodology were checked using certified reference materials and by the conventional wet acid digestion method on the same certified reference materials and real samples. The recovery of all the studied elements was found to be in the range of 97.8–99.3% in certified reference materials. The results of this study showed that the mean values of Cd, Ni and Pb were significantly higher in scalp hair, blood and urine samples of both smoker and nonsmoker patients than in referents (P<0.001), whereas the concentration of Zn was lower in the scalp hair and blood, but higher in the urine samples of hypertensive patients. The deficiency of Zn and the high exposure of toxic metals as a result of tobacco smoking may be synergistic with risk factors associated with hypertension

Reporter enzyme fluorescence (REF) imaging and quantification of tuberculosis in live animals

The slow growth rate of Mycobacterium tuberculosis hinders research progress, since estimating the bacterial numbers present in all experiments normally relies on determination of colony forming units on agar plates. M. tuberculosis colonies can take as long as four to six weeks to become visible. Whole animal imaging is an emerging technology that has broad applications in all areas of biological sciences, including monitoring infections. Imaging allows bacterial numbers to be determined in real-time for each infected animal, individually, which allows inter-animal variability to be observed and controlled for. Reporter enzyme fluorescence (REF) utilizes custom substrates that allow production of a fluorescent product after cleavage by a bacterial enzyme. In our recently published studies, we demonstrate that the enzyme β-lactamase, a naturally occurring enzyme expressed by M. tuberculosis, can be used for REF. The resulting imaging system is the first that allows non-invasive detection of natural M. tuberculosis strains directly in pulmonary infected living animals. Use of REF for M. tuberculosis-infected mice allows detection of ∼104 CFU in the lungs, which is very sensitive. This system also displays promise for allowing rapid evaluation of differences in virulence of strains and efficacy of therapeutics and vaccines. This system could be developed into a diagnostic tool for tuberculosis through the use of REF to identify infected tissues or other diagnostic specimens

Assessment of in vitro immunity to Mycobacterium tuberculosis in a human peripheral blood infection model using a luciferase reporter construct of M. tuberculosis H37Rv

Protective immune responses to tuberculosis in man are primarily cell-mediated and require the interaction of specific T cells, cytokines and activated macrophages. In the present study, Mycobacterium tuberculosis H37Rv labelled with luciferase reporter enzyme was used to analyse the anti-mycobacterial immunity in man using an in vitro whole blood infection model. Peripheral blood samples obtained from M. bovis bacille Calmette–Guérin (BCG)-vaccinated tuberculin-positive healthy volunteers (n = 23) were cultured with M. tuberculosis H37Rv reporter strain. The growth of bacteria in the whole blood cultures was monitored after 48 and 96 h of infection. The results showed that the growth of M. tuberculosis was significantly inhibited after 96 h (P < 0·029) of culture. Among the cytokines studied, interleukin (IL)-10 and IL-12 were not detected at all, whereas low levels of interferon (IFN)-γ after 96 h (0·4 IU/ml) and tumour necrosis factor (TNF)-α after 48 (135 pg/ml) and 96 h (47 pg/ml) of culture were detected in the supernatants of whole blood infected with M. tuberculosis. The magnitude of bacterial growth correlated directly with the concentration of TNF-α detected after 48 h (r = 0·722) and 96 h (r = 0·747) of culture (P ≤ 0·0001 and P ≤ 0·0001, respectively). However, the addition of monoclonal antibodies specific to TNF-α and IFN-γ to the blood cultures did not alter mycobacterial growth indicating the role of other mechanisms/factors in restricting the growth of M. tuberculosis in whole blood cultures