Intra-Organ Variation in Age-Related Mutation Accumulation in the Mouse

Using a transgenic mouse model harboring chromosomally integrated lacZ mutational target genes, we previously demonstrated that mutations accumulate with age much more rapidly in the small intestine than in the brain. Here it is shown that in the small intestine point mutations preferentially accumulate in epithelial cells of the mucosa scraped off the underlying serosa. The mucosal cells are the differentiated villus cells that have undergone multiple cell divisions. A smaller age-related increase, also involving genome rearrangements, was observed in the serosa, which consists mainly of the remaining crypts and non-dividing smooth muscle cells. In the brain we observed an accumulation of only point mutations in no other areas than hypothalamus and hippocampus. To directly test for cell division as the determining factor in the generation of point mutations we compared mutation induction between mitotically active and quiescent embryonic fibroblasts from the same lacZ mice, treated with either UV (a point mutagen) or hydrogen peroxide (a clastogen). The results indicate that while point mutations are highly replication-dependent, genome rearrangements are as easily induced in non-dividing cells as in mitotically active ones. This strongly suggests that the point mutations found to have accumulated in the mucosal part of the small intestine are the consequence of replication errors. The same is likely true for point mutations accumulating in hippocampus and hypothalamus of the brain since neurogenesis in these two areas continues throughout life. The observed intra-organ variation in mutation susceptibility as well as the variation in replication dependency of different types of mutations indicates the need to not only extend observations made on whole organs to their sub-structures but also take the type of mutations and mitotic activity of the cells into consideration. This should help elucidating the impact of genome instability and its consequences on aging and disease

Identification of Inhibitors against Mycobacterium tuberculosis Thiamin Phosphate Synthase, an Important Target for the Development of Anti-TB Drugs

Tuberculosis (TB) continues to pose a serious challenge to human health afflicting a large number of people throughout the world. In spite of the availability of drugs for the treatment of TB, the non-compliance to 6–9 months long chemotherapeutic regimens often results in the emergence of multidrug resistant strains of Mycobacterium tuberculosis adding to the precariousness of the situation. This has necessitated the development of more effective drugs. Thiamin biosynthesis, an important metabolic pathway of M.tuberculosis, is shown to be essential for the intracellular growth of this pathogen and hence, it is believed that inhibition of this pathway would severely affect the growth of M.tuberculosis. In this study, a comparative homology model of M.tuberculosis thiamin phosphate synthase (MtTPS) was generated and employed for virtual screening of NCI diversity set II to select potential inhibitors. The best 39 compounds based on the docking results were evaluated for their potential to inhibit the MtTPS activity. Seven compounds inhibited MtTPS activity with IC50 values ranging from 20 – 100 µg/ml and two of these exhibited weak inhibition of M.tuberculosis growth with MIC99 values being 125 µg/ml and 162.5 µg/ml while one compound was identified as a very potent inhibitor of M.tuberculosis growth with an MIC99 value of 6 µg/ml. This study establishes MtTPS as a novel drug target against M.tuberculosis leading to the identification of new lead molecules for the development of antitubercular drugs. Further optimization of these lead compounds could result in more potent therapeutic molecules against Tuberculosis

Molecular identification of adenovirus causing respiratory tract infection in pediatric patients at the University of Malaya Medical Center

<p>Abstract</p> <p>Background</p> <p>There are at least 51 adenovirus serotypes (AdV) known to cause human infections. The prevalence of the different human AdV (HAdV) serotypes varies among different regions. Presently, there are no reports of the prevalent HAdV types found in Malaysia. The present study was undertaken to identify the HAdV types associated primarily with respiratory tract infections (RTI) of young children in Malaysia.</p> <p>Methods</p> <p>Archived HAdV isolates from pediatric patients with RTI seen at the University of Malaya Medical Center (UMMC), Kuala Lumpur, Malaysia from 1999 to 2005 were used. Virus isolates were inoculated into cell culture and DNA was extracted when cells showed significant cytopathic effects. AdV partial hexon gene was amplified and the sequences together with other known HAdV hexon gene sequences were used to build phylogenetic trees. Identification of HAdV types found among young children in Malaysia was inferred from the phylograms.</p> <p>Results</p> <p>At least 2,583 pediatric patients with RTI sought consultation and treatment at the UMMC from 1999 to 2005. Among these patients, 48 (< 2%) were positive for HAdV infections. Twenty-seven isolates were recovered and used for the present study. Nineteen of the 27 (~70%) isolates belonged to HAdV species C (HAdV-C) and six (~22%) were of HAdV species B (HAdV-B). Among the HAdV-C species, 14 (~74%) of them were identified as HAdV type 1 (HAdV-1) and HAdV type 2 (HAdV-2), and among the HAdV-B species, HAdV type 3 (HAdV-3) was the most common serotype identified. HAdV-C species also was isolated from throat and rectal swabs of children with hand, foot, and mouth disease (HFMD). Two isolates were identified as corresponding to HAdV-F species from a child with HFMD and a patient with intestinal obstruction.</p> <p>Conclusions</p> <p>HAdV-1 and HAdV-2 were the most common HAdV isolated from pediatric patients who sought treatment for RTI at the UMMC from 1999 to 2005. HAdV-B, mainly HAdV-3, was recovered from ~22% of the patients. These findings provide a benchmark for future studies on the prevalence and epidemiology of HAdV types in Malaysia and in the region.</p

Comparison of the elution properties of commercially available gentamicin and bone cement containing vancomycin with 'home-made' preparations

Introduction Vancomycin is commonly added to acrylic bone cement during revision arthroplasty surgery. Proprietary cement preparations containing vancomycin are available but significantly more expensive. We investigated whether the antibiotic elution and mechanical strength of ‘home-made’ vancomycin containing bone cement was comparable to commercial vancomycin-impregnated cement. Methods A total of 18 cement discs of constant size, containing either proprietary CopalG+V®; or ‘home-made’ CopalR+G® with vancomycin added by hand, were made. Each disc contained the same antibiotic quantities (0.5g gentamycin, 2g vancomycin) and was immersed in ammonium acetate buffer in a sealed container. Fluid from each container was sampled at eight time points over a two week period. The concentration of gentamicin and vancomycin in the fluid was analysed using high performance liquid chromatography mass spectrometry. The impact strength of each PMMA cement preparation was measured using a Charpy-type impact tester. Results Highest peak antibiotic concentrations were observed from the ‘home-made’ vancomycin containing cement, added as in the operating theatre. Overall antibiotic elution was, five-fold (vancomycin) and two-fold (gentamicin), greater from the ‘home-made’ mix compared to commercially mixed cement. However the ‘home-made’ cements showed greater variation in elution kinetics compared to the commercial mix. Use of a vacuum during mixing had no significant effect on antibiotic elution in any of the samples. Impact strength testing showed no significant differences between the groups. Discussion Our findings suggest the addition of 2g vancomycin powder to gentamicin-impregnated bone cement in theatre, significantly increases elution of both antibiotics, with no significant loss of strength, compared to commercially prepared cement. Conclusion We have found no significant advantages of expensive off-the-shelf vancomycin-impregnated bone cement and recommend the addition of vancomycin powder by hand when making cement beads and spacers