Changes in leucocyte and lymphocyte subsets during tuberculosis treatment; prominence of CD3dimCD56+ natural killer T cells in fast treatment responders.
H. Veenstra,* R. Baumann, *, N.M. Carroll, * P.T. Lukey, M. Kidd, N. Beyers, C.T. Bolliger, P. D. van Helden* and G. Walzl*.
Summary
The immune responses against pulmonary tuberculosis (TB) are still poorly defined. This study describes changes in leucocyte and lymphocyte subsets during treatment to find reliable immunological markers for the disease and treatment response. Flow cytometric peripheral blood immune phenotyping, routine haematology and sputum microbiology were performed on 21 HIV-negative adult TB patients with positive sputum cultures during therapy in comparison with 14 healthy purified protein derivative (PPD)-positive volunteers. Patients at diagnosis showed high absolute neutrophil and monocyte counts which fell during treatment but low lymphocyte subset counts which increased [except natural killer (NK) and NK T cells].
Introduction
The mechanisms of protective immunity against Mycobacterium tuberculosis (Mtb) infection and disease in humans have not been fully clarified. Many reports have addressed the potential immunological defect(s) by comparing immune phenotypes in actively diseased patients to those with latent infection. Most of these investigations have focused on T lymphocyte subsets, particularly CD4+ and γδ T cells, generally reporting depressed CD4+ cells in peripheral blood of tuberculosis (TB) patients [1-3], but results are discrepant for γδ T cells, where both elevated [4,5] and normal [6,7] numbers have been found. Only a few but inclusive reports of B lymphocyte and natural killer (NK) cell numbers in TB patients exit [1,3,8,9] and NK T cells have, to our knowledge, not been investigated in TB patients. Generally, contributors to TB susceptibility remain unclear and follow-up data during therapy are scanty.
The aim of our study was to investigate immune parameters during therapy and this report describes a systematic follow-up of leucocyte counts and lymphocyte subsets in TB patients for the entire 26-week treatment period. Furthermore, due to the fact that the identification of high-risk patients for slow response to chemotherapy would have important clinical implications, we analysed peripheral blood immunophenotypes as potential surrogate markers of early TB treatment response and applied a multivariate classification technique to identify fast and slow responders to treatment by immunophenotype at diagnosis.
Materials and methods
Settings
This study was conducted in an epidemiological field site in metropolitan Cape Town, where the incidence of new smear and/or culture-positive TB was on average 313/100 000 population/year (1993-980) [10].
Patients and controls
The study was approved by the Ethics Committee of the Faculty of Health Sciences at Stellenbosch University and written, informed consent was obtained from all participants.
Twenty-nine new smear-posivite pulmonary TB patients were screened fro this study. Inclusion criteria included: sputum culture-positive for Mtb, no multi-drug resistance, HIV-negative, taking at least 80% of prescribed doses during the intensive phase of treatment. Eight patients were excluded for the following reasons: non-compliance, multi-drug resistant TB, negative sputum culture, refusal of HIV testing or incomplete follow-up visits. Twenty-one patients with first-time TB were enrolled and studied throughout treatment. Blood samples were taken at diagnosis prior to initiation of treatment and at weeks 1, 5, 13 and 26 after start of treatment (the last blood sample being taken on the last day of chemotherapy).
Results
Differences between treatment response groups
When percentages and absolute numbers of each cell type at diagnosis in fast responders were compared to those at diagnosis of slow responders with a Mann-Whitney test, the percentages and absolute counts of CD3 dim/CD 56+ NK T cells at diagnosis were the only parameters that correlated significantly with the treatment response – they were significantly higher at diagnosis in fast responders (P=0.01).
Differences between early response phenotypes were most prominent at diagnosis and the variable at diagnosis that were used for the analysis were the absolute numbers of leucocyte, lymphocyte and T cell subsets.
Discussion
In this study we have shown significant changes in absolute numbers of neutrophils, monocytes and lymphocyte subsets during active TB. Our finding that these changes occur already during the first weeks of treatment is important, as it suggests strongly that TB patients tested at different time-points during their treatment should not be grouped together in the analysis of results.
In summary, peripheral blood white cell counts change rapidly during treatment and some counts of diagnosis hold promise as surrogate markers of treatment response. Further prospective studies with larger numbers of patients are now needed to evaluate the role of immunophenotyping in general and of CD3dim/CD56+ NK T cells specifically, including their functional characterization. The role of these cells in predicting differential outcomes at month 6 and the development of recurrence after cure need to be assessed.
Original article appeared in @2006 British Society for Immunology, Clinical and Experimental Immunology, 145: 252-260 |
