Primate Monocytes - CD14, CD16 - Ziegler-Heitbrock

Lymphocyte and monocyte flow cytometry immunophenotyping as a diagnostic tool in uncharacteristic inflammatory disorders.

Abstract

BACKGROUND: Patients with uncharacteristic inflammatory symptoms such as long-standing fatigue or pain, or a prolonged fever, constitute a diagnostic and therapeutic challenge. The aim of the present study was to determine if an extended immunophenotyping of lymphocytes and monocytes including activation markers can define disease-specific patterns, and thus provide valuable diagnostic information for these patients. METHODS: Whole blood from patients with gram-negative bacteraemia, neuroborreliosis, tuberculosis, acute mononucleosis, influenza or a mixed connective tissue disorders, as diagnosed by routine culture and serology techniques was analysed for lymphocyte and monocyte cell surface markers using a no-wash, no-lyse protocol for multi-colour flow cytometry method. The immunophenotyping included the activation markers HLA-DR and CD40. Plasma levels of soluble TNF alpha receptors were analysed by ELISA. RESULTS: An informative pattern was obtained by combining two of the analysed parameters: (i), the fractions of HLA-DR-expressing CD4+ T cells and CD8+ T cells, respectively, and (ii), the level of CD40 on CD14+ CD16- monocytes. Patients infected with gram-negative bacteria or EBV showed a marked increase in monocyte CD40, while this effect was less pronounced for tuberculosis, borrelia and influenza. The bacterial agents could be distinguished from the viral agents by the T cell result; CD4+ T cells reacting in bacterial infection, and the CD8+ T cells dominating for the viruses. Patients with mixed connective tissue disorders also showed increased activation, but with similar engagement of CD4+ and CD8+ T cells. Analysis of soluble TNF alpha receptors was less informative due to a large inter-individual variation. CONCLUSION: Immunophenotyping including the combination of the fractions of HLA-DR expressing T cell subpopulations with the level of CD40 on monocytes produces an informative pattern, differentiating between infections of bacterial and viral origin. Furthermore, a quantitative analysis of these parameters revealed the novel finding of characteristic patterns indicating a subacute bacterial infection, such as borreliosis or tuberculosis, or a mixed connective tissue disorder. The employed flow cytometric method is suitable for clinical diagnostic laboratories, and may help in the assessment of patients with uncharacteristic inflammatory symptoms. PMID: 20626864 [PubMed - indexed for MEDLINE]PMCID: PMC2912311Free PMC Article Images from this publication.See all images (4) Free text Figure 1 T lymphocytes: (A), ratio of CD4+/CD8+ T cells; (B), activation within the CD4+ and CD8+ T cell subsets. The result for T cell activation shows the percentage of HLA class II-expressing cells among all CD4+ T cells and CD8+ T cells, respectively. The indicated activation values are normalized with the result of the normal control individuals being expressed as 1.0, and the dashed lines indicate the ... Lymphocyte and monocyte flow cytometry immunophenotyping as a diagnostic tool in uncharacteristic inflammatory disorders BMC Infect Dis. 2010;10:205-205.Figure 2 Flow cytometry method for quantitation of T cell and monocyte activation status. Representative dot plots from analysis of patient samples are shown. CD4+ T cells and CD8+ T cells gated on CD3+ CD4+ (A,C,E) and CD3+ CD8+ (B,D,F) lymphocytes from (A-B), a healthy control; (C-D), tuberculosis; (E-F), influenza; the inserted percentages indicate the fraction of HLA-DR+ cells. Mo... Lymphocyte and monocyte flow cytometry immunophenotyping as a diagnostic tool in uncharacteristic inflammatory disorders BMC Infect Dis. 2010;10:205-205.Figure 3 NK cells, B cells and mucosal subtypes of B and T cells. The results show the fraction among all blood lymphocytes of (A), NK cells; (B), B cells; (C), CD5+ B1 subtype of B cells; (D), gamma/delta TCR+ subtype of T cells as determined by a CD3+ CD4- CD8- phenotype. The dashed lines indicate the level of the normal controls. ** p < 0.01. Lymphocyte and monocyte flow cytometry immunophenotyping as a diagnostic tool in uncharacteristic inflammatory disorders BMC Infect Dis. 2010;10:205-205.Figure 4 Monocytes: (A), fraction among all monocytes of the CD16+ subtype; and (B), activation status as determined by CD40 expression. The dashed lines indicate the level of the normal controls. *p < 0.02 or 0.05, ** p < 0.01. Lymphocyte and monocyte flow cytometry immunophenotyping as a diagnostic tool in uncharacteristic inflammatory disorders BMC Infect Dis. 2010;10:205-205.