Immunophenotypic and DNA Genotypic Analysis of T-Cell and NK-Cell Subpopulations in Patients with B-Cell Chronic Lymphocytic Leukaemia (B-CLL)

Abstract

Absolute numbers and distributions of peripheral blood T-cells and NK cells were immunophenotypically determined in 21 patients with B-CLL and compared with those obtained from a series of 13 elderly normal controls with an age range of 60-87 years. For absolute CD3⁺, CD4⁺ and CD8⁺ T-cell, and CD16⁺ NK subpopulation numbers, there were no consistent differences between the normal and B-CLL groups although some individual patient variation was seen. Immunophenotypic analyses did however reveal that CD3+ T-cells in almost half (10/21) of the B-CLL patients were Ia⁺ (defined as <20% positive cells), compared to 0/13 of the elderly control group (p > 0.001), and that the proportions of CD4⁺ and CD8⁺ cells expressing membrane CD45RO were significantly increased compared to the control group. Subdivision of the B-CLL cases into those with low (>20%) and high (<20%) proportions of CD3+ T-cells co-expressing la further showed that CD45RO expression by CD4⁺ fractions was particularly prominent in the Ia⁺ subgroup, and that the relative increase of CD4⁺CD45RO⁺ cells was primarily a consequence of decreased absolute numbers of CD4⁺ CD45RA⁺ lymphocytes. This study also examined extracted DNA from enriched CD3+ T-cell fractions (obtained by immunomagnetic bead selection in 9 of the B-CLL cases) by PCR analysis with two primers for the T-cell gamma gene locus. With the VγC (consensus) primer, 8/9 cases were polyclonal and the remaining case was oligoclonal. For comparison, 7/9 CD3⁺ fractions were oligoclonal with the V-γ9 primer with the other two cases being polyclonal. No monoclonal CD3⁺ components were found. It is suggested that the observed increased la expression by CD3⁺ cells and the predominance of CD4⁺ cells expressing membrane CD45RO in patients with B-CLL may be of potential relevance to understanding the pathogenesis and patterns of disease progression.

Key Words:

• B-CLL
• T-cells
• immunophenotypes
• PCR analysis
• CD45R
• NK cells