Biomarkers in chronic fatigue syndrome: evaluation of natural killer cell function and dipeptidyl peptidase IV/CD26

Abstract

Background: Chronic Fatigue Syndrome (CFS) studies from our laboratory and others described decreased natural killer cell cytotoxicity (NKCC) and elevated proportion of lymphocytes expressing the activation marker, dipeptidyl peptidase IV (DPPIV) also known as CD26. However, neither these assays nor other laboratory tests are widely accepted for the diagnosis or prognosis of CFS. This study sought to determine if NKCC or DPPIV/CD26 have diagnostic accuracy for CFS.

Methods/results: Subjects included female and male CFS cases and healthy controls. NK cell function was measured with a bioassay, using K562 cells and (51)Cr release. Lymphocyte associated DPPIV/CD26 was assayed by qualitative and quantitative flow cytometry. Serum DPPIV/CD26 was measured by ELISA. Analysis by receiver operating characteristic (ROC) curve assessed biomarker potential. Cytotoxic function of NK cells for 176 CFS subjects was significantly lower than in the 230 controls. According to ROC analysis, NKCC was a good predictor of CFS status. There was no significant difference in NK cell counts between cases and controls. Percent CD2+ lymphocytes (T cells and NK cells) positive for DPPIV/C26 was elevated in CFS cases, but there was a decrease in the number of molecules (rMol) of DPPIV/C26 expressed on T cells and NK cells and a decrease in the soluble form of the enzyme in serum. Analyses by ROC curves indicated that all three measurements of DPPIV/CD26 demonstrated potential as biomarkers for CFS. None of the DPPIV/C26 assays were significantly correlated with NKCC.

Conclusions: By ROC analysis, NKCC and three methods of measuring DPPIV/C26 examined in this study had potential as biomarkers for CFS. Of these, NKCC, %CD2+CD26+ lymphocytes and rMol CD26/CD2+ lymphocyte, required flow cytometry, fresh blood and access to a high complexity laboratory. Soluble DPPIV/C26 in serum is done with a standard ELISA assay, or with other soluble factors in a multiplex type of ELISA. Dipeptidyl peptidase IV on lymphocytes or in serum was not predictive of NKCC suggesting that these should be considered as non-redundant biomarkers. Abnormalities in DPPIV/CD26 and in NK cell function have particular relevance to the possible role of infection in the initiation and/or the persistence of CFS.

Figure 1. ROC analyses were used to evaluate NKCC as a predictor of CFS.

The nonparametric ROC plot (blue curve) indicated the ability of NKCC to discriminate between CFS cases and healthy controls. Smaller values for NKCC were associated with CFS cases. The 45 degree line (green) indicates the theoretical plot of a test with no discrimination between CFS and controls.

Figure 2. ROC analyses were used to evaluate %CD26+CD2+ lymphocytes as a predictor of CFS.

The nonparametric ROC plot (purple curve) indicated the ability of %CD26+CD2+ lymphocytes to discriminate between CFS cases and healthy controls. Larger values for %CD26+CD2+ lymphocytes were associated with CFS cases. The 45 degree line (green) indicates the theoretical plot of a test with no discrimination between CFS and controls.

Figure 3. ROC analyses were used to evaluate serum dipeptidyl peptidase IV/CD26 as a predictor of CFS.

The nonparametric ROC plot (red curve) indicated the ability of serum dipeptidyl peptidase IV/CD26 to discriminate between CFS cases and healthy controls. Smaller values were associated with CFS cases. The 45 degree line (green) indicates the theoretical plot of a test with no discrimination between CFS and controls.

Figure 4. ROC analyses were used to evaluate relative number of molecules of dipeptidyl peptidase IV/CD26 on the surface of CD2+ lymphocytes as a predictor of CFS.

The nonparametric ROC plot (orange curve) indicated the ability of number of molecules of dipeptidyl peptidase IV/CD26 on the surface of CD2+ lymphocytes to discriminate between CFS cases and healthy controls. Smaller values were associated with CFS cases. The 45 degree line (green) indicates the theoretical plot of a test with no discrimination between CFS and controls. cell at saturating concentrations of antibody; rMol/cell) is shown.