Data from this and earlier studies gave credible support to diminished NKCC function in CFS. These effector cells of the innate immune system have an important role in antiviral, antibacterial, and antitumor immunity, but were deficient as measured by direct cytolysis of target cells, and as determined by measurement of intra cellular lytic proteins 
. In 60 to 80% of published samples, CFS presented with acute onset of illness, with systemic symptoms similar to influenza infection that did not subside 
. The sudden onset, the symptoms of myalgia, arthralgia, sore throat and tender lymphadenopathy prompted a theory of infection induced illness 
. Published reports both support and deny associated microbial infections, reactivation of latent herpes virus infections and/or retrovirus infections in CFS 
. Of interest is the finding by Glaser and colleagues that the adverse immunologic effects of persistent infections with Epstein Barr Virus (EBV) did not require viral DNA synthesis 
. Some published work suggested the possibility of elevated risk for cancer in patients with CFS 
, though to date there has been no long term natural history study to accurately assess this risk.
Previously, we showed that the proportion of lymphocytes (NK cells and T cells) expressing CD26 is elevated in CFS cases 
. In the present study, we found the density of DPPIV/CD26 on lymphocyte surfaces and the concentration of the enzyme in plasma is reduced in CFS subjects, compared to controls. We hypothesize that this reduction is due to chronic lymphocyte activation in CFS patients. The present study adds to the evidence of loss of innate immune function and chronic immune activation, resulting from the long term presence of antigenic stimulus, either self or foreign. Compared to healthy controls, chronic hepatitis C patients had significantly lower serum soluble CD26 levels 
. In another study, acute, self-limiting infection with live influenza vaccine and chronic infection with persistent antigen, such as with cytomegalovirus (CMV), EBV or human immunodeficiency virus (HIV), was compared using multi-parameter flow cytometry and tetramer technology. These analyses identified a unique pattern of high density DPPIV/CD26 expression among influenza-specific CD8 T cells, but not among CD8 T cells specific for CMV, EBV (three different epitopes) or HIV 
. These findings were interpreted as indicating that expression of CD26 (high) is characteristic of a memory cell, present in acute infection but not in chronic infection.
Dipeptidyl peptidase IV/CD26 cleaves N-terminal X-Pro dipeptides from peptides. The peptidase controls the in vivo
half-life of the proinflammatory chemokine stromal cell-derived factor-1 (SDF-1). Mice deficient in DPPIV/CD26 exhibited increased levels of circulating active SDF-1, associated with increased numbers of SDF-1 receptor (CXCR4)-positive cells infiltrating arthritic joints 
. In a clinical study, by the same researchers, plasma levels of DPPIV/CD26 from rheumatoid arthritis patients were significantly decreased when compared to those from osteoarthritis patients and inversely correlated with C-reactive protein levels. They postulated that decreased circulating soluble DPPIV/CD26 levels in arthritis may influence DPPIV/CD26-mediated regulation of the chemotactic SDF-1/CXCR4 axis. These patients have elevated number of T cells expressing DPPIV/CD26 and reduced DPPIV enzymatic activity and DPPIV/CD26 antigen in plasma compared to controls 
Dipeptidyl peptidase IV/CD26 causes the degradation of glucagon-like peptide 1 (GLP-1), an incretin hormone 
. Inhibitors of DPPIV/CD26 such as sitagliptin, which prevent the degradation of GLP-1 
, are now marketed for the treatment of type 2 diabetes mellitus (T2DM). Considering that DPPIV/CD26 has a key role in immune regulation as a T cell activation molecule and in immune-mediated disorders, it is noteworthy that the effects of inhibition of DPPIV/CD26 on the immune system have not been extensively investigated. There are reports that infections were increased after sitagliptin treatment 
. So far, only routine laboratory safety variables have been measured in published randomized controlled trials.
Administration of DPPIV/C26 inhibitors for the treatment of T2DM patients could influence immune function, including NKCC. A study of CD26 gene knockout mice concluded that DPPIV/C26 contributes to the regulation of development, maturation and migration of CD4 T, NK and NKT cells, cytokine secretion, T cell-dependent antibody production and immunoglobulin isotype switching of B cells 
. An initial diagnosis of CFS would not be made in the patient with obvious T2DM. However, the frequency of development of T2DM after diagnosis of CFS is not known–nor is the effects of a DPPIV/C26 inhibitor in the CFS patient.
Duration of illness typically exceeds 10 years. Persistence may involve complex interaction of immune, autonomic and neuroendocrine regulation and remains poorly understood. It is important to recall that the associated chronic inflammation can have important consequences on energy metabolism by promoting insulin resistance 
. This chronic inflammatory state would also support a concurrent low-grade Th1 response by inhibiting the protective effects of T regulatory cell subset via increased IL-6 expression. The decreased NKCC and the abnormal DPPIV/C26 manifestations in CFS would be compatible with the hypothesis that the immune system of affected individuals is biased towards a T- helper (Th) 2 type, or humoral immunity-oriented cytokine pattern.
The data obtained on NK cell function, immune activation and DPPIV/C26 on cell surfaces and in serum, are consistent with a viral etiology for CFS. The elevated proportion of activated CD4 and CD8 T cells and defective NKCC in CFS cases suggests that T cells are metabolically limited in performing their helper function. The abnormalities observed may have applications with other complex, chronic and poorly understood illnesses, including fibromyalgia, gulf war illness, rheumatologic disorders and multiple sclerosis—though the precise constellation of patterns observed with these biomarkers may differ in each. However, the specific panel that we have identified here are likely to be helpful as objective markers for diagnosing CFS, determining subgroups, following patients over time and as targets for therapeutic strategies. These indicators are parts of a complex and integrated system and their inter-dependency must be addressed 
. Accordingly, we are currently engaged in mapping the network structure of neuroendocrine-immune interaction in CFS
Obvious limitations of this study are that each patient sample represents a single point in time. To address this, we are conducting a large longitudinal study to follow 150 subjects over 18 months. Samples are collected during times of relative symptom remission and exacerbation. Completion of the study will allow the correlation of CFS related symptoms with lymphocyte function and activation. Because CFS is a condition that affects women in disproportionate numbers, over eighty percent of the cases in the present study were female. The larger study will have sufficient power to allow a sub study of biomarker patterns in men with CFS.
The predominance of evidence indicating that people with CFS have decreased function of NK cells and abnormal activation of T and NK cells was supported by this study. The purpose of the study was to determine usefulness of these measurements as biomarkers. By ROC analysis, NKCC and dipeptidyl peptidase/CD26 were identified as potential biomarkers for CFS through their demonstrated accuracy in discriminating CFS patients from healthy controls. Dipeptidyl peptidase/CD26 on lymphocytes or in serum was not correlated with NKCC, suggesting that these are non-redundant biomarkers. Current CFS treatments are directed at reducing symptom severity but no cure exists for this condition. The findings of this study give support to the concept that cause and/or the pathophysiology of CFS are related to infection. These findings may lead to therapeutic approaches. The specter of infectious disease further emphasizes the significance of this research to public health.