Biomedical subjects

S Phyu

Publications and source records attributed to S Phyu.

7 recordsLinked to original sources

PubMed2003

Rapid colorimetric method for testing susceptibility of Mycobacterium tuberculosis to isoniazid and rifampin in liquid cultures.

We have developed a rapid colorimetric method for testing the susceptibility of M. tuberculosis to isoniazid (INH) and rifampin (RIF) based on incorporation of nitrate in broth cultures containing growth supplements. The performance of this colorimetric nitrate reductase-based antibiotic susceptibility (CONRAS) test was compared with that of the radiometric BACTEC 460TB system in determining the susceptibilities of 74 M. tuberculosis strains to INH and RIF. By using the BACTEC 460TB system as the "gold standard," the sensitivity (i.e., the ability to detect true drug resistance) and specificity (i.e., the ability to detect true drug susceptibility) of the CONRAS test were 100 and 95% for INH and 94 and 100% for RIF, respectively. The repeatability of the CONRAS test was excellent (for INH, kappa = 1 and P < 0.001; for RIF, kappa = 0.88 and P < 0.001). For the majority of strains, results were obtained within 5 days. The CONRAS test is rapid, accurate, and inexpensive and is an adequate alternative, particularly for resource-poor countries.

Antitubercular Agents↗

PubMed2000

Diversity of lung and spleen immune responses in mice with slowly progressive primary tuberculosis.

The aim of the present study was to assess the compartmentalized immune response, in terms of cytokine secretion and cell activation, in lungs and spleens of mice with slowly progressive primary tuberculosis. Immunocyte populations from both organs were isolated and stimulated with concanavalin A, purified protein derivatives and MPT 59. Production of interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) was measured using an enzyme-linked immunosorbent assay, and cell activation was measured using a tetrazolium colorimetric assay. The IFN-gamma and IL-4 levels in the supernatants of Mycobacterium tuberculosis antigen (Ag)-stimulated lung immunocytes from infected mice were higher than the levels from uninfected mice. However, only IL-4 levels were raised in the supernatants of Ag-stimulated spleen immunocytes from infected mice. Spontaneous and Ag-stimulated immunocyte activation was lower only in the lungs of infected mice compared with uninfected mice. The level of lung immunocyte activation was inversely associated with the extent of gross pulmonary pathology. In conclusion, cytokine secretion and cell activation were different between lungs and spleens in slowly progressive primary murine tuberculosis. Cytokine diversity may explain the confinement of tuberculous lesions in the lungs and the absence of lesions in the spleens of mice with slowly progressive tuberculosis.

Animals↗

PubMed2000

In situ expression of cytokines and cellular phenotypes in the lungs of mice with slowly progressive primary tuberculosis.

The cellular phenotypes and the expression of cytokines were studied in the lungs of mice, using immunohistochemistry, during different phases of slowly progressive primary murine tuberculosis infection. During the first phase the small focal lesions in healthy mice contained predominantly interleukin-2 (IL-2)-expressing cells. A small number of tumour necrosis factor-alpha (TNF-alpha)-, monocyte chemoattractant protein-1 (MCP-1)- and IL-10-expressing cells were also present. IL-4-expressing cells were not detected. During the second phase the mice became unwell, but the bacterial counts and the size of focal lesions stabilized. IL-4-expressing cells appeared. The IL-10-, TNF-alpha- and MCP-1-expressing cells increased in number. On progression to phase three, the mice became seriously unwell and died rapidly. The inflammation spread to approximately 80% of the lung parenchyma. There was a marked increase in the number of IL-10-expressing cells. Expression of other cytokines was similar to that observed in the second phase. In the lesions, 3-6% of the macrophages (Mphi) containing mycobacterial antigens expressed high levels of IL-10 and TNF-alpha. The absolute numbers of CD3-, CD4- and CD11b-expressing cells in the lesions increased with the progression of infection. The numbers of CD8+ cells were reduced in the last phase of infection. The kinetics of T-lymphocyte subsets and the pattern of cytokine expression changed with the type and degree of tissue injury. The small number of Mphi with a heavy load of mycobacterial antigens may be the cause of this disturbance in cytokine balance, thus leading to progression of inflammation.

Animals↗

PubMed1999

Increased expression of Fas ligand on Mycobacterium tuberculosis infected macrophages: a potential novel mechanism of immune evasion by Mycobacterium tuberculosis?

We have studied the location and mechanism of apoptosis within the granulomas in the lungs at various stages of slowly progressive primary murine Mycobacterium tuberculosis infection. Parallel sections were analyzed for detection of mycobacterial antigens, Fas, and Fas ligand (FasL) by immunohistochemistry, and for apoptotic cells by terminal deoxynucleotidyl-transferase-mediated dUTP-digoxigenin nick end labeling (TUNEL) method. The frequency of apoptosis was high in the macrophage aggregates as compared to the lymphocyte aggregates and at the interface between them. Five to seven percent of the vacuolated macrophages in the granulomas expressed FasL intensely. These cells contained large amounts of mycobacterial antigens. These findings suggest that M. tuberculosis infection can induce increased expression of FasL in a population of infected macrophages. As a consequence the infected macrophages will be protected from the attack of cytotoxic T cells and activation of bactericidal mechanisms by Th1 type lymphocytes. This constitutes a novel evasion mechanism for M. tuberculosis possibly explaining the chronic course of infection.

Animals↗

PubMed1999

Changes in T-lymphocyte subsets in lungs and spleens of mice with slowly progressive primary mycobacterium tuberculosis infection: involvement of unconventional T-cell subsets.

Our previous study showed that the cell-activation responses and cytokine-secretion patterns were different in lungs and spleens of mice with slowly progressive primary Mycobacterium tuberculosis infection. The aim of the present study was to characterize the T-cell subsets in lungs and spleens of mice with a similar infection. The percentages of T-cell subsets were determined by flow cytometry and the absolute numbers were calculated. Spleens of infected mice showed a threefold expansion of CD4+ cells but no change in CD8+ cells, whereas lungs had a threefold increase of both subsets. A significant expansion of CD4-CD8-alphabeta+ [double negative (DN)alphabeta+] subsets was observed in the lungs of infected mice compared with uninfected mice. This was not the case in the spleens of infected mice. In infected mice the CD4-CD8- (DN) population preferentially expressed alphabeta-T-cell receptors (TCR) in the lungs but gammadelta-TCR in the spleens. The percentages of many T-cell subsets were significantly higher in the lungs than in the spleens of both uninfected and infected mice. However, the percentages of CD4+ and CD4-CD8+TCR- subsets in the lungs were significantly lower than in the spleens of infected mice. We also observed some previously unreported T-cell subsets: double positive-TCR- (DPTCR-), DPalphabeta+ and DPgammadelta+. So far their functions are unknown.

Animals↗

PubMed1999

A mouse model for slowly progressive primary tuberculosis.

The progression from primary Mycobacterium tuberculosis infection to disease is usually slow in humans. The aim of this study was to develop and characterize a mouse model for slowly progressive primary tuberculosis, using the intraperitoneal (i.p.) route of infection, and to compare it with our previously described model of latent M. tuberculosis infection. B6D2F1 hybrid mice inoculated with 1.5 x 106 colony-forming units (CFUs) of M. tuberculosis H37Rv were followed-up for 70 weeks. Lungs, livers and spleens were examined for bacillary growth, histopathological changes and mycobacterial antigens (MPT64, ManLAM and multiple antigens of M. tuberculosis), by immunohistochemical staining. The infection was found to pass through three distinctive phases. During phase 1, mice were healthy despite development of small granulomas and an increasing number of bacilli in the lungs. During phase 2, mice were unwell but mortality was low. The count of M. tuberculosis and the granuloma size stabilized. The granulomas contained an increasing population of large, vacuolated macrophages. During phase 3, mice became moribund and died rapidly, but the M. tuberculosis count remained relatively stable. The inflammatory infiltrates filled approximately 80% of the lung parenchyma and the lesions were not well demarcated. Rapidly progressing inflammation, rather than an increase in the M. tuberculosis count, seems to contribute more to mortality.

Animals↗

PubMed1998

A mouse model for latent tuberculosis.

The aim of the study was to establish a reproducible murine model for latent tuberculosis. We propose an operational definition of latent murine tuberculosis as a stable Mycobacterium-tuberculosis count in lungs and spleens without clinical signs or obvious histopathological changes in the lungs over a long period of time and without spontaneous reactivation of disease. B6D2F1Bom mice were inoculated with a wide range of Mycobacterium tuberculosis doses intraperitoneally or intravenously and followed for a long period to determine suitable conditions to produce latent infection. No anti-tuberculosis drug treatment was used. Microbiological and histopathological studies were carried out. Corticosterone challenge was used to reactivate the latent infection. Mice infected with 4 x 10(4) and 4 x 10(5) bacilli i.p. were followed up to 107 weeks without spontaneous reactivation. The present model is discussed in comparison with previous latent tuberculosis mouse models as well as the possible mechanisms of shift to stationary phase from multiplying bacilli.

Animals↗