Clinical Information

Normal immunity requires a balance between the activities of various lymphocyte subpopulations with different effector and regulatory functions.

Different immune cells can be characterized by unique surface membrane antigens described by a cluster of differentiation nomenclature (eg, CD3 is an antigen found on the surface of T lymphocytes). Abnormalities in the number and percent of T (CD3, CD4, CD8), B (CD19), and natural killer (CD16+CD56) lymphocytes have been described in a number of different diseases. In patients infected with HIV, the CD4 count is measured for AIDS diagnosis and initiation of antiviral therapy. The progressive loss of CD4 T lymphocytes in patients infected with HIV is associated with increased infections and complications.

The United States Public Health Service has recommended that all HIV-positive patients be tested every 3 to 6 months for the level of CD4 T lymphocytes.

The adaptive immune response includes both cell-mediated (mediated by T cells and natural killer [NK] cells) and humoral immunity (mediated by B cells). After antigen recognition and maturation in secondary lymphoid organs, some antigen-specific B cells terminally differentiate into antibody-secreting plasma cells or become memory B cells. Memory B cells are 3 subsets: marginal zone B cells (MZ or non-switched memory), class-switched memory B cells, and IgM-only memory B cells. Decreased B-cell numbers, B-cell function, or both result in immune deficiency states and increased susceptibility to infections. These decreases may be either primary (genetic) or secondary. Secondary causes include medications, malignancies, infections, and autoimmune disorders.

Common variable immunodeficiency (CVID), a disorder of B-cell function, is the most prevalent primary immunodeficiency with a prevalence of 1 to 25,000 to 1 to 50,000.(1) CVID has a bimodal presentation with a subset of patients presenting in early childhood and a second set presenting between 15 and 40 years or, occasionally, even later. Many different genetic defects have been associated with CVID, including variants in the ICOS, CD19, BAFF-R, and TACI genes. TACI variants account for 8% to 15% of CVID cases.

Common variable immunodeficiency is characterized by hypogammaglobulinemia, usually involving most or all of the immunoglobulin classes (IgG, IgA, IgM, and IgE), impaired functional antibody responses, and recurrent sinopulmonary infections.(1,2) B-cell numbers may be normal or decreased. A minority of patients with CVID (5%-10%) have very low B-cell counts (<1% of peripheral blood leukocytes), while another subset (5%-10%) exhibit noncaseating, sarcoid-like granulomas in different organs and also tend to develop a progressive T-cell deficiency.(1) Of all patients with CVID, 25% to 30% have increased numbers of CD8 T cells and a reduced CD4 to CD8 ratio (<1). Studies have shown the clinical relevance of classifying patients with CVID by assessing B-cell subsets since changes in different B-cell subsets are associated with specific clinical phenotypes or presentations.(3,4)

The B-cell phenotyping assay can be used in the diagnosis of hyper-IgM syndromes, which are characterized by increased or normal levels of IgM with low IgG and/or IgA.(5) Patients with hyper-IgM syndromes can have 1 of 5 known genetic defects- in the CD40L, CD40, AID (activation-induced cytidine deaminase), UNG (uracil DNA glycosylase), and NEMO (NF-kappa B essential modulator) genes.(5) Variants in CD40L and NEMO are inherited in an X-linked fashion, while variants in the other 3 genes are inherited in an autosomal recessive fashion. Patients with hyper-IgM syndromes have a defect in isotype class-switching, which leads to a decrease in class-switched memory B cells, with or without an increase in non-switched memory B cells and IgM-only memory B cells.

In addition to its utility in the diagnosis of the above-described primary immunodeficiencies, B-cell phenotyping may be used to assess reconstitution of B-cell subsets after hematopoietic stem cell or bone marrow transplant. This test is also used to monitor B-cell-depicting therapies, such as Rituxan (rituximab) and Zevalin (ibritumomab tiuxetan).

The etiology of CVID is heterogeneous. Variants of the gene that encodes TACI, TNFRSF13B (tumor necrosis factor receptor superfamily, member 13B), probably account for about 10% to 15% of all CVID cases.(6-8) Patients with variants in the TACI gene are particularly prone to developing autoimmune diseases, including cytopenias and lymphoproliferative disease. The other variants each have been reported in only a handful of patients. The etiopathogenesis is still undefined in 65% to 75% of patients with CVID.

The absolute counts of lymphocyte subsets are known to be influenced by a variety of biological factors, including hormones, the environment, and temperature. The studies on diurnal (circadian) variation in lymphocyte counts have demonstrated progressive increase in CD4 T-cell count throughout the day, while CD8 T cells and CD19+ B cells increase between 8:30 a.m. and noon, with no change between noon and afternoon. Natural killer (NK) cell counts, on the other hand, are constant throughout the day.(9) Circadian variations in circulating T-cell counts have been shown to be negatively correlated with plasma cortisol concentration.(10-12) In fact, cortisol and catecholamine concentrations control distribution and, therefore, numbers of naive versus effector CD4 and CD8 T cells.(10) It is generally accepted that lower CD4 T-cell counts are seen in the morning compared with the evening(13) and during summer compared to winter.(14) These data, therefore, indicate that timing and consistency in timing of blood collection is critical when serially monitoring patients for lymphocyte subsets.