Journal of Internal Medicine 1997; 242: 465-478

"Elevated Apoptotic Cell Population in Patients With Chronic Fatigue Syndrome: The Pivotal Role of Protein Kinase RNA."

A. Vodjani, M. Ghoneum, P.C. Choppa, L. Magtoto, & C.W. Lapp. From the Immunosciences Laboratory Inc., Beverly Hills, and Charles Drew University School of Medicine and Science, Los Angeles, California and the Hunter-Hopkins Centre, Charlotte, North Carolina, USA.

ABSTRACT

Objectives. A prominent feature of chronic fatigue syndrome (CFS) is a disordered immune system. Recent evidence indicates that the induction of apoptosis might be mediated in a dysregulated immune system by the upregulation of growth inhibitory cytokines. Therefore, the purpose of this study was to evaluate the apoptotic cell population, interferon-alpha (IFN-a) and the IFN-induced protein kinase RNA (PKR) gene transcripts in peripheral blood lymphocytes (PBL) of CFS individuals, as compared to healthy controls.

Subjects and Methods. PBL were isolated from CFS (n=29) and healthy control individuals (n=15) and subjected to quanitative analysis of apoptotic cell population and cell cycle progression by flow cytometry. Quantitative competitive polymerase chain reaction (Q/C PCR) and Western blot analysis were used to assess the levels of PKR mRNA and protein in control and CFS individuals. In addition, circulating IFN-a was measured by ELISA assay.

Results. Increased apoptotic cell population was observed in CFS individuals, as compared to healthy controls (26.6+/- 12.9% and 9.9 +/- 4.2%, respectively). The increased apoptotic subpopulation in CFS individuals was accompanied by an abnormal cell arrest in the S phase and the G2/M boundary of the cell cycle as compared to the control group (8.6 +/- 1.2% to 22.8 +/- 2.4 and 3.6 +/- 0.82 to 24.3 +/- 3.4, respectively). In addition, CFS individuals exhibited enhanced PKR mRNA and protein levels (mean basal level 3538 +/- 1050 and 2.7 +/-0.26, respectively). In 50% of the CFS samples (n=29) treated with 2-aminopurine (2-AP) (a potent inhibitor of PKR) the apoptotic population was reduced by more than 50%.

Conclusions. PKR-mediated apoptosis in CFS individuals may contribute to the pathogenesis and the fatigue symptomology associated with CFS.

Keywords: apoptosis, chronic fatigue syndrome, PKR

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Other key points made in this paper:

1) Dysregulation of the IFN-induced antiviral antiproliferative pathways in CFS was reported from this report and other laboratories (i.e. Suhadolnik's findings in the changes of the 2-5a synthetase/RNaseL antiviral pathway in a controlled clinical trial) (pg.466).

2) Activation of RNaseL in either viral-infected or tumorigenic cells increases the rate of RNA degradation, resulting in the inhibition of viral replication and cellular proliferation, respectively (pg 466).

3) The other antiviral, antiproliferative IFN-induced gene product is PKR which, when upregulated, induces programmed cell death (apoptosis) and it has been shown to possess tumor suppressor properties (pgs. 466 and 470).

4) PKR protein levels increased approximately up to fivefold over basal levels and is manifested in abnormal metabolism in PWCs. Disregulated cellular metabolism due to inhibition of protein synthesis machinery might be the mechanism responsible for the fatigue observed in CFS individuals (Pgs. 471 and 475).

5) PKR upregulation is found in viral infection and cancer (Pg 471). The activation in CFS could result from disregulated immune system or chronic viral infection (Pg 475).

6) Apoptosis is a physiological mode of programmed cell death where the cell actively participates in its own death process. The morphological features of apoptosis are cell shrinkage, cytoplasmic condensation and the induction of double-stranded DNA fragmentation at linker region between nucleosomes (pg 466).

7) Growing evidence supports the theory that the induction of apoptosis through immune defense mechanisms is an important mechanism for elimination of cancer cells as well as virus infected cells (pgs. 466-467).

8) Inhibition of apoptosis may be one mechanism of tumor production; on the other hand, uncontrolled apoptosis can be detrimental to developing tissues and may facilitate the formation of degenerative diseases (pg. 466).

9) CFS patients exhibited enhanced levels of IFN-a in plasma and cell lysate indicating abnormal cytokine production (pg. 470).

10) PKR mRNA, protein and activity are upregulated predominantly in the PBL isolated from CFS individuals but not in those of the controls (pg 475).

11) The differences found between this study and other studies that didn't find elevated cytokines or apoptosis could be explained by differences in either symptomatic states of the population studied or differences between study sites or sample processing. Patient heterogeneity and and multiple patient subsets may also have contributed to the discrepancies (pg 476).

12) The fact that PKR levels, apotosis and cell cycle progression are disregulated, with the increased severity and duration of certain immunological markers, suggests that measurement of PKR levels together with apoptosis and cell cycle progression could be potentially useful for stratification of patients and possibly for monitoring therapy or the progression of CFS (pg 476).