INTRODUCTION — CD4 T cell laboratory testing through flow cytometry is considered an important part of HIV care since this parameter is used to stage disease and inform clinical management. With few exceptions, infectious and neoplastic complications of HIV disease are more common in patients with lower CD4 cell counts, so the results can guide clinicians as to how concerned they should be about a certain diagnosis. In addition, certain CD4 cell thresholds are used as benchmarks to either initiate or discontinue prophylaxis against opportunistic infections (OIs). These thresholds were previously used as an indication to begin antiretroviral therapy (ART); however, they now provide a sense of the urgency to start ART.
This topic will address flow cytometry and other techniques for measuring CD4 cells and the various factors that can lead to variations in CD4 cell counts, such as medications, infections, and other conditions. Issues related to the immunology and pathogenesis of HIV infection and the use of HIV RNA testing are discussed elsewhere. Guidance on when to start ART or OI prophylaxis is discussed elsewhere. (See "Techniques and interpretation of HIV-1 RNA quantitation" and "Selecting antiretroviral regimens for treatment-naïve persons with HIV-1: General approach" and "Overview of prevention of opportunistic infections in patients with HIV".)
CD4 and CD8 T cells — Human T lymphocytes can be functionally divided into cells that provide help for other immune cells and those that mediate cellular cytotoxicity. Helper T lymphocytes express cluster determinant 4 (abbreviated as "CD4") whereas cytotoxic T cells express cluster determinant 8 (abbreviated as "CD8"). The CD4 and CD8 molecules are members of the immunoglobulin superfamily and mediate adhesion to major histocompatibility complex class II and class I molecules, respectively. In addition, CD4 and CD8 amplify stimulatory signals through the T cell receptor [1-3].
A CD4:CD8 ratio is calculated by dividing the number of CD4+ T cells by the number of CD8+ T cells; this ratio is usually greater than 1 in immunocompetent individuals . However, in HIV infection, the CD4:CD8 ratio is usually less than 1. This reflects increasing numbers of CD8+ T cells and depletion of CD4+ T cells in chronic infection. This ratio usually increases with the initiation of antiretroviral therapy (ART) although the CD4:CD8 ratio normalizes in only a minority of patients .
How does HIV affect CD4 T cells? — HIV infection leads to severe depletion of CD4 T cells in the gut-associated lymphoid tissue with subsequent reduced levels of circulating CD4 lymphocytes in the peripheral blood . CD4 cells are reduced precipitously in acute HIV infection, but usually rebound in the blood over several weeks as HIV-specific CD8 T cells help to lower plasma viremia . In the untreated patient, CD4 T cells subsequently decline over several years. Population-based studies of the natural history of HIV infection among men who have sex with men (MSM) show that the mean CD4 count prior to seroconversion is about 1000 cells/microL; CD4 counts decline to a mean of 780 cells/microL at six months post-seroconversion and to 670 cells/microL at one year of follow-up . Subsequently, the CD4 cell count declines at an average yearly rate of approximately 50 cells/microL, but there is substantial variation among patients [8-10]. Significant depletion of CD4 T cells can lead to opportunistic infections and mortality in the untreated patient. In addition, in one report, low baseline CD4 counts were associated with increased rates of virologic failure . The stages and natural history of HIV infection are discussed elsewhere. (See "The natural history and clinical features of HIV infection in adults and adolescents".)
MECHANISMS OF CD4 T CELL DEPLETION IN HIV INFECTION — Although other viruses can infect CD4 T cells, HIV selectively targets and infects activated, expanding CD4 T cells .
During acute infection, viral replication occurs at an extremely rapid rate, often producing the highest level of circulating virus observed at any time during infection . Rapidly proliferating virus-specific CD4 T cells become activated and expand in the presence of high viremia, resulting in subsequent infection, functional impairment, and death of these CD4 T cells. Viremia declines with the recruitment of HIV-specific CD8 T cells (host immune response) and depletion of the pool of susceptible CD4 cells.
Within the first weeks of infection there is a massive loss of T cells in the gut-associated mucosa associated with disruption of mucosal integrity . This is followed by a progressive loss of CD4 cells in the peripheral blood, which is a hallmark of HIV infection. Disruption of the integrity of the gut mucosa is associated with translocation of microbial products from the gut, including lipopolysaccharide (LPS) . This "leaky gut" is associated with chronic immune activation, as evidenced by markers of T cell activation (ie, CD38 and HLA-DR) [15-17].
Treatment with antiretroviral therapy leads to viral suppression and immunologic improvement. The extent of the immune recovery is dependent in part on the degree of immune compromise prior to treatment. Incomplete immunologic recovery among patients with advanced disease may also be related to collagen formation in the gut and lymphoid tissues leading to disruption of normal architecture [18,19]. Other factors that influence immune recovery (eg, comorbid conditions, age) are reviewed below. (See 'The CD4 T cell response to antiretroviral therapy' below.)
TECHNIQUES FOR MEASURING CD4 CELL COUNTS — The CD4 cell count is usually determined by flow cytometry in which blood specimens must be processed within 18 hours of collection.
Alternative systems include: FACS Count System (Becton Dickinson); VCS Technology/Coulter Cyto-Spheres (Coulter Corp); Zymmune CD4/CD8 Cell Monitoring Kit (Zynaxis Inc); and TRAx CD4 Test Kit (T Cell Diagnostics). These systems perform well compared with standard flow cytometry and are cheaper and less technically demanding .
Absolute counts may differ when different assays are used, or even when done by the same test in different laboratories . As a result, sequential samples that are processed using different methods or by different laboratories should be interpreted with caution.
What is a normal CD4 cell count? — The normal adult CD4 count for most laboratories falls in a range of 800 to 1050 cells/microL; however, when considering laboratory variations of two standard deviations, the normal CD4 count range falls within 500 to 1400 cells/microL . This broad range in normal values reflects the fact that the CD4 cell count is the product of three variables: the white blood cell count, the percentage of lymphocytes, and the percentage of lymphocytes that bears the CD4 receptor. (See 'Variability of CD4 cell counts' below.)
Laboratory testing also reports CD4:CD8 ratios, which are usually greater than 1 in a normal host. (See 'CD4 and CD8 T cells' above.)
Absolute CD4 counts and percentages in HIV infection — Flow cytometry reports CD4 percentages; the absolute CD4 cell count is calculated by multiplying the percentage and the total white cell count (WBC). In general, these two numbers are concordant; corresponding values are listed below:
●An absolute CD4 count >500 cells/microL corresponds to a CD4 percentage of >29 percent
●An absolute CD4 count between 200 to 500 cells/microL corresponds to a CD4 percentage of 14 to 28 percent
●An absolute CD4 count <200 cells/microL corresponds to a CD4 percentage of <14 percent
Laboratory testing in people with HIV typically demonstrates an inversed CD4:CD8 ratio of <1. It is important for clinicians to be aware that significant changes in the total WBC will have an effect on the absolute CD4 cell count, while the CD4 percentage will remain the same. (See 'Variability of CD4 cell counts' below.)
People with HIV and a CD4 count <200 cells/microL are classified as having AIDS and are at risk for opportunistic infections. More detailed discussions on the clinical findings of patients with AIDS and the prevention of opportunistic infections are found elsewhere. (See "The natural history and clinical features of HIV infection in adults and adolescents", section on 'AIDS and advanced HIV infection' and "Overview of prevention of opportunistic infections in patients with HIV".)
What is a significant change in CD4 cell counts? — A significant change (eg, 2 standard deviations) between two tests is approximately a 30 percent change in the absolute count or an increase or decrease in CD4 percentage by 3 percentage points . An adequate response on treatment is defined as an increase in CD4 count in the range of 50 to 150 cells/microL per year. (See "Patient monitoring during HIV antiretroviral therapy".)
Definition of AIDS — A CD4 count of <200 cells/microL indicates the clinical stage of AIDS, which implies a high susceptibility to opportunistic infections, such as Pneumocystis pneumonia. Information on prophylaxis against opportunistic infections is discussed elsewhere. (See "Overview of prevention of opportunistic infections in patients with HIV".)
VARIABILITY OF CD4 CELL COUNTS
General principles — Although CD4 cell count monitoring can be a major parameter that influences clinical management, there is significant intraindividual variability; prior to the era of antiretroviral therapy (ART), a study of 1020 untreated patients found that the intraindividual coefficient of variation averaged by 25 percent .
As noted above, a number of factors other than HIV infection influence CD4 cell counts. Significant changes in the total WBC can lead to marked changes in the absolute CD4 cell count. Certain medications or infections associated with leukopenia may result in depression of the absolute CD4 cell count. In contrast, specific medications or infections which lead to leukocytosis can result in elevated CD4 cell counts. In such cases, the absolute CD4 cell count and percentage may be discordant. These types of changes do not indicate a true change in the patient’s immunologic status per se and should be interpreted with caution. If the CD4 cell count changes dramatically without significant changes in the CD4 percentage, the clinician needs to assess other factors that may be leading to artifactual changes in the absolute count. Flow cytometry results that would lead to a therapeutic intervention may need confirmation if the result is dramatically different from a prior result performed within three to six months.
The CD4 cell percentage is sometimes used to assess immune function instead of the absolute number since there is less variation with this parameter [23,24]. In the AIDS Clinical Trial Group laboratories, the within-subject coefficient of variation for the CD4 percentage was 18 percent compared with 25 percent for the CD4 cell count .
Even among healthy adults, there are seasonal or month-to-month changes in the CD4 cell count . There are also diurnal changes with the lowest levels of CD4 cell counts in the evening and peak values in the morning . These variations have only a minor impact on results of flow cytometry.
Factors that affect laboratory testing — The CD4 count can be influenced by factors other than HIV infection. The following factors do not appear to have an appreciable effect on the CD4 cell count: gender, HIV risk category, or psychologic or physical stress . However, several infectious and noninfectious causes can reduce or increase the CD4 cell count.
Infection — Modest decreases in the CD4 cell count have been noted in various acute infections (eg, cytomegalovirus, EB virus, hepatitis B, tuberculosis, some bacterial infections, and histoplasmosis). A study of patients coinfected with HIV and HTLV-I found that CD4 cell counts were 80 to 180 percent higher than in controls with HIV at comparable levels of immunosuppression . Some data suggest that hepatitis C virus (HCV) coinfection is associated with a less robust immune recovery after initiation of ART compared with patients who are HCV-seronegative . (See "Clinical manifestations and natural history of chronic hepatitis C virus infection", section on 'Natural history'.)
Medications — Depending on the effect of the medication on the total WBC count, absolute CD4 cell counts may rise or decline. These changes do not reflect true changes in the patient’s immunologic status.
●For example, declines in absolute CD4 cell counts have been observed in HIV/HCV coinfected patients who are undergoing interferon therapy; however, CD4 T cell percentages usually remain the same or increase throughout HCV therapy.
●In humans and laboratory animals, one dose of corticosteroids leads to a rapid and transient decline in the numbers of peripheral blood lymphocytes through a redistribution of circulating lymphocytes. This may lead to a profound drop in the absolute CD4 cell count . With chronic administration however, corticosteroids may lead to a leukocytosis; this may in turn lead to an elevated absolute CD4 cell count.
Alcohol use — Persons with alcohol use disorders may experience depression of CD4 cell counts, which improve after alcohol abstinence .
Chronic conditions — In people with HIV, having a splenectomy, advanced liver disease, and/or splenomegaly can affect the CD4 count. In such patients, the CD4 percentage may be a more reliable indicator of immunologic status than the absolute CD4 count. Splenectomy may lead to a deceptively high CD4 count. In contrast, advanced liver disease can be associated with a decrease in the CD4 count, with a lower absolute CD4 cell count relative to the CD4 percentage [31-33]; this discordance may be related to splenic sequestration and lymphopenia. A more detailed discussion of other chronic conditions that cause CD4 lymphopenia is found elsewhere. (See "Idiopathic CD4+ lymphocytopenia", section on 'Differential diagnosis'.)
Pregnancy — Pregnancy can also affect CD4 testing. A longitudinal analysis was performed of 51 HIV seronegative patients who had CD4 cell count testing during pregnancy and the postpartum period; 25 HIV-negative nonpregnant controls were also selected . The following results were found:
●The mean white blood cell count was elevated in pregnant subjects above that in the non-pregnant controls and this difference increased throughout the pregnancy.
●The mean absolute lymphocyte cell count, lymphocyte percentage, and absolute CD4+ cell count were significantly lower during pregnancy than during the 12-week post-delivery period. In contrast, the mean absolute CD8 count was not significantly different.
These data are important to bear in mind when evaluating the person with HIV who is pregnant. In this situation, the absolute CD4 count should be interpreted in the context of the CD4 percentage.
THE CD4 T CELL RESPONSE TO ANTIRETROVIRAL THERAPY — With antiretroviral therapy (ART) and effective viral suppression, the expected CD4 cell response is an increase of approximately 50 to 150 cells/microL at one year, followed by slower incremental increases of 50 to 100 cells/microL per year until a steady state level is reached [23,35,36]. The frequency of monitoring depends upon the patient's CD4 count and viral load, as well as the duration of ART. In some patients, monitoring CD4 cells while on therapy can be considered optional, provided there is viral suppression. A discussion of CD4 count monitoring in patients receiving ART is discussed in detail elsewhere. (See "Patient monitoring during HIV antiretroviral therapy", section on 'CD4 count'.)
Factors that correlate with reduced CD4 recovery include older age, male sex, the type of ART used, and the presence of certain coinfections (eg, hepatitis C virus) [28,37]. In addition, some studies have demonstrated that the baseline CD4 cell count affects the extent of immune recovery, suggesting that patients with advanced immune compromise may have limited immunologic reserve . When interleukin-2 was administered to people with HIV in conjunction with ART, there was an absolute increase in CD4 cell counts compared with ART alone, but no discernible clinical benefit .
If ART is discontinued, there is generally a rapid viral load rebound and sharp decline in CD4 count with a decrease of up to 100 to 150 cells/microL in three to four months [40-42]. However, if a patient develops virologic rebound due to resistance while taking ART, the CD4 may not decline for months . As a result, CD4 cell counts are not a reliable marker of virologic suppression or medication adherence, and in general, we do not make changes in ART based on CD4 changes. (See "Patient monitoring during HIV antiretroviral therapy", section on 'Virologic response' and "Evaluation of the treatment-experienced patient failing HIV therapy".)
CD4 T CELL TESTING IN RESOURCE-LIMITED SETTINGS — The World Health Organization (WHO) recommends CD4 cell count monitoring to assess when to start prophylaxis against opportunistic infections and when to initiate antiretroviral therapy (ART) . CD4 counts were previously advocated as a surrogate to monitor the response to ART; however, subsequent studies have shown that this, by itself, is an inadequate way to evaluate treatment response and that the viral load is also an invaluable parameter, when available [41,42,45,46].
The total lymphocyte count (TLC) is a crude surrogate for the CD4 cell count, but is advocated by the WHO in areas where flow cytometry may not be available.
Other alternatives to flow cytometry have been investigated in resource-poor areas:
●One study demonstrated that microcapillary flow cytometry correctly identified those individuals with a CD4 count <200 cells/microL (95 percent sensitivity and 100 percent specificity) . Although this technology is easier to use, requires little training of personnel, and is less expensive than flow cytometry, it still requires significant initial capital investment for the instrument.
●Manual immunobead-based methods have the advantage of only requiring routine light microscopy . However, the results with this technique may be less reproducible.
●The "CD4 Select" method, which utilizes only an automatic hematoanalyzer to assess both CD4 absolute counts and percentages, was found to have greater than 90 percent correlation with standard flow cytometry .
IDIOPATHIC CD4 LYMPHOCYTOPENIA — Idiopathic CD4 lymphocytopenia (ICL) is a rare syndrome characterized by low CD4 cell counts that are unexplained by HIV infection or other known medical conditions. Criteria for the definition of this disorder are :
●A CD4 count less than 300 cells/microL or a CD4 percentage less than 20 on two or more measurements
●Lack of laboratory evidence of HIV infection
●Absence of an alternative explanation for the CD4 lymphocytopenia
The characteristics of this syndrome are similar to HIV infection and include susceptibility to opportunistic infections and abnormalities of T cell homeostasis and chronic immune activation [30,51,52]. No etiologic infectious agent has been identified to date. This topic is discussed in detail elsewhere. (See "Idiopathic CD4+ lymphocytopenia".)
SOCIETY GUIDELINE LINKS — Links to society and government-sponsored guidelines from selected countries and regions around the world are provided separately. (See "Society guideline links: HIV screening and diagnostic testing".)
INFORMATION FOR PATIENTS — UpToDate offers two types of patient education materials, "The Basics" and "Beyond the Basics." The Basics patient education pieces are written in plain language, at the 5th to 6th grade reading level, and they answer the four or five key questions a patient might have about a given condition. These articles are best for patients who want a general overview and who prefer short, easy-to-read materials. Beyond the Basics patient education pieces are longer, more sophisticated, and more detailed. These articles are written at the 10th to 12th grade reading level and are best for patients who want in-depth information and are comfortable with some medical jargon.
Here are the patient education articles that are relevant to this topic. We encourage you to print or e-mail these topics to your patients. (You can also locate patient education articles on a variety of subjects by searching on "patient info" and the keyword(s) of interest.)
●Basics topics (see "Patient education: HIV/AIDS (The Basics)" and "Patient education: Tests to monitor HIV (The Basics)")
SUMMARY AND RECOMMENDATIONS
●Background – CD4 T cell laboratory testing through flow cytometry is considered an important part of HIV care since this parameter is used to stage disease and guide clinical management. (See 'Introduction' above.)
The normal CD4 count for most laboratories falls in a range of 800 to 1050 cells/microL. Flow cytometry reports CD4 percentages; the absolute CD4 cell count is calculated by multiplying the percentage and the total white blood cell count (WBC). In general, these two numbers are concordant. (See 'Absolute CD4 counts and percentages in HIV infection' above.)
●CD4 depletion in setting of HIV infection – During acute HIV infection, virus titers are extremely high. Rapidly proliferating virus-specific CD4 T cells become activated and expand in the presence of high viremia, resulting in subsequent infection, functional impairment, and death of CD4 T cells. In the untreated person with HIV, CD4 T cells subsequently decline over several years. (See 'Mechanisms of CD4 T cell depletion in HIV infection' above.)
A significant change (eg, two standard deviations) between two tests is approximately a 30 percent change in the absolute count or an increase or decrease in CD4 percentage by 3 percentage points. (See 'What is a significant change in CD4 cell counts?' above.)
Significant depletion of CD4 T-cells can lead to opportunistic infections and mortality in the untreated patient. A CD4 count of <200 cells/microL indicates the clinical stage of AIDS, which implies a high susceptibility to opportunistic infections, such as Pneumocystis pneumonia. (See 'Definition of AIDS' above.)
●Factors that can affect the CD4 count – A number of factors other than HIV infection influence CD4 cell counts. Significant changes in the total WBC can lead to marked changes in the absolute CD4 cell count. Certain medications or infections, which are associated with leukopenia, may result in depression of the absolute CD4 cell count. In contrast, specific medications or infections, which lead to leukocytosis, can result in elevated CD4 cell counts. These types of changes do not indicate a true change in the patient’s immunologic status per se and should be interpreted with caution. (See 'Variability of CD4 cell counts' above.)
●Response to antiretroviral therapy – With antiretroviral therapy and effective viral suppression, the expected CD4 cell response is an increase of approximately 50 to 150 cells/microL at one year, followed by slower incremental increases of 50 to 100 cells/microL per year until a steady state level is reached. The rate of increase in CD4 cells may be slower in certain patients (eg, those with severe immunocompromise at baseline). (See 'The CD4 T cell response to antiretroviral therapy' above.)
ACKNOWLEDGMENT — The UpToDate editorial staff acknowledges John G Bartlett, MD, who contributed to an earlier version of this topic review.
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