Thesis (PhD (Biomedical Sciences. Molecular Biology and Human Genetics))--University of Stellenbosch, 2007. / Despite the ongoing global tuberculosis (TB) problem and extensive research into
protective immunity against this intracellular pathogen, mechanisms of protective
immunity against Mycobacterium tuberculosis (Mtb) in humans have not been fully
clarified. Numerous reports have addressed the potential immunological defect(s) in
infected individuals that have developed active TB in comparison to those who have
remained healthy in spite of infection. Markers of treatment response phenotypes are
still elusive. The aims of this study were to define lymphocyte subsets in the
peripheral blood of TB patients and controls, to determine intracellular interferon-γ
(IFN-γ) and interleukin-4 (IL-4) production and to find correlations of these data
with microbiologically-defined treatment response.
Methods
Whole blood tests were done on 30 HIV-negative, smear-positive pulmonary TB
patients and 18 healthy skin test positive volunteers resident in the same community.
Immunophenotyping was performed by flow cytometry, combined with routine
haematology, for the enumeration of peripheral blood immune cell subtypes. Whole
blood was also stimulated in vitro with anti-CD3 monoclonal antibody and
intracellular IFN-γ and IL-4 determined by flow cytometry. Lymphocyte
proliferation in response to heat-killed Mtb was determined by tritiated thymidine
incorporation. Routine microbiological monitoring by sputum smears and culture
was done throughout the patients’ 26 weeks of treatment.
Results
Compared to healthy controls, absolute numbers of peripheral blood lymphocytes
and lymphocyte subsets were significantly depressed in patients at diagnosis but
normalized during treatment with the exception of natural killer (NK) cells and
natural killer T (NKT) cells. A novel subset of the latter was found to correlate
significantly with treatment response. IFN-γ-producing T cells after a 4-hour T cell
receptor stimulation were significantly higher in patients at diagnosis and normalized
during treatment. Supplementary kinetic experiments showed that IFN-γ production
in patients at diagnosis seemed to be accelerated. Lymphocyte proliferation was
lower in patients at diagnosis and normalized during treatment. Neither IFN-γ
production nor lymphocyte proliferation correlated with treatment response. Low
intracellular IL-4 production was constitutive in patients and controls, was
insignificantly lower in patients at diagnosis than in controls and, in the slow
responder patient group, it was significantly lower than in the fast responder group.
High IL-4 expression was found in low numbers of T cells in patients and controls
and supplementary experiments showed co-expression of active caspase-3 in these
cells, which signified apoptosis.
Conclusions
Lymphocyte subset phenotypes associated with TB are largely abnormal only during
active infection and only a novel subset of NKT cells showed correlation with
treatment response. Intracellular IFN-γ production and lymphocyte proliferation is
increased and decreased, respectively, only during active infection and does not
correlate with treatment response. The T helper 1/T helper 2 (Th1/Th2) hypothesis
could not be confirmed in the context of tuberculosis but instead constitutive IL-4
production may play a role as a growth factor.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/1180 |
| Date | 03 1900 |
| Creators | Veenstra, Hannelore F. U. |
| Contributors | Walzl, Gerhard, Bouic, Patrick J. D., University of Stellenbosch. Faculty of Health Sciences. Dept. of Biomedical Sciences. Molecular Biology and Human Genetics. |
| Publisher | Stellenbosch : University of Stellenbosch |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | 1431899 bytes, application/pdf |
| Rights | University of Stellenbosch |
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