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Index to Abstracts from AACFS Conference October 1998


Abstracts of Papers Presented at
The Bi-Annual Research Conference of the
American Association for
Chronic Fatigue Syndrome (AACFS)

October 10-11, 1998 -- Cambridge, Massachusetts

Poster Session 3: Immunology

Co-chairs: Sudhir Gupta, MD and Andrew Lloyd, MD
October 10, 1998, 4:00 pm-4:30 pm

1. A case series survey of silicone breast implant patients
Cichon, M., M.D.; Farrell, D.; Ganjo, S.; Sadler, G.
2. T-lymphocytes in CFS -- in vitro reaction to mutagens
I.Hauspie, I. Campine, P. De Becker, K. De Meirleir, M. Kirsch-Voldersl
3. Markers of immune activation are associated with psychological distress in patients with CFS
Lvnn Helder. Ph.D., Stacy Wagner, M.S., Robert Keller, M.D., Nancy Klimas, M.D. and Michael Antoni, Ph.D.
4. Immunocytology and in vitro reverse-transcriptase activity in CFS
M.I. Holmes, R. Easingwood, J. Cross, and J. Faed
5. Lymph node morphology and phenotype in Chronic Fatigue Syndrome
Nancy Klimas, Kevin Maher, Roberto Patarca, Jean Walling, Mack Smith, and Mary Ann Fletcher
6. Immunological response in Chronic Fatigue Syndrome (CFS) following a graded exercise test to exhaustion
J. Lamanca, S. Sisto, J. Ottenweller, A. Peckerman, S. Cook, T. Denny, W. Gause and B. H. Natelson
7. Could nickel allergy be involved in the pathogenesis of Chronic Fatigue Syndrome?
J.A. Marcusson , G. Lindh, B. Evengard
8. Serum neopterin and somatization in women with chemical intolerance, depressives, and normals
Roberto Patarca, Nancy G. Klimas, Iris R. Bell, and Elizabeth Hardin
9. Altered cytokine production in whole blood cultures of Chronic Fatigue Syndrome patients and a disturbed regulation by glucocorticoids
J.Visser, W. Graffelman, B Blauw, J. Haspels, IE.R. de Kloet, Lex Nagelkerten


A case series survey of silicone breast implant patients

Authors: Cichon M. M.D., Farrell, D., Ganjo, S., Sadler, G.

Objective: To survey the symptoms of a large group of breast implant patients displaying illness, and to determine if any clinical or serological features predominate.

Method: A private internal medicine practice.

Results: A referred sample of 415 patients with fatigue of long duration, followed by muscle/joint pain, cognition problems, polyneuropathy, and localized breast pain.

Conclusion: Silicone adjuvant breast disease may be a novel disorder, possibly auto-immune, producing atypical syndromes that do not fit within the classic diagnostic criteria of known conditions. Furthermore, the diversity and distinction of silicone adjuvant breast disease may require the medical community to accept it as a new entity, encompassing a neurological and connective tissue disorder.

Key Words: silicone, breast implants, atypical, children, women's health, polyneuropathy, scleroderma-like, lupus-like.

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T-lymphocytes in CFS -- in vitro reaction to mutagens

Authors:: I. Hauspie¹, I.Campine²*, P. De Becker², K. De Meirleir², M. Kirsch-Volders¹
¹Department of Human Genetics, Free University of Brussels, Brussels, Belgium
²Department of Human Physiology, Free University of Brussels, Brussels, Belgium
* I. Carnpine is supported by funds from the Foundation for Scientific Research, Belgium (F.W.O.).

Objective: Many studies in CFS patients suggest a virally-triggered onset, associated with an abnormal immune function. With recent findings in mind concerning the role of viral proteins taking in cell proliferation control, our aim was to test in vitro cell proliferation in CFS patients and healthy controls, both in normal conditions and after treatment with different mutagens.

Methods: Human lymphocytes from 8 patients and 14 controls were cultured in the presence and absence of different mutagens. Using the cytokinesis blocked micronucleus assay, the proliferation index was assessed and chromosomal abberations were detected. We also examined the effect of ethanol exposure on T-lymphocytes in vitro.

Results: The results show that the proliferation index in CFS patients (in absence of a mutagen) is slightly lower than in controls, but CFS patients show more spontaneous damage during cell division than controls. In the absence of S9, the cell division is reduced in the presence of ethanol in CFS patients. Ethanol also induces damage in dividing T-lymphocytes in CFS patients in the absence of S9. Finally, lymphocytes of CFS patients seem to be more resistant to in vitro treatment with MMS than controls.

Conclusion: These results add information to the existing knowledge of intracellular abnormalities in CFS. Our findings point toward abnormalities in intracellular protein metabolism, and increased sensitivity to alcohol. We also postulate, from the MMS experience, that lymphocyte cell membrane permeability is altered.

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Markers of immune activation are associated with psychological distress in patients with CFS

Authors: Lynn Helder. Ph.D., Stacy Wagner, M.S., Robert Keller, M.D., Nancy Klimas, M.D. and Michael Antoni, Ph.D.

Objective: Chronic immune activation, as suggested by abnormal T-cell phenotypes, and psychological distress have often been associated with Chronic Fatigue Syndrome (CFS), although conclusions remain controversial. Our purpose was to further explore the relationship between immune functioning and psychological distress in patients with CFS.

Methods: 25 subjects meeting the Centers for Disease Control criteria for CFS were examined prior to participation in a cognitive therapy intervention. Blood draws and psychological inventories were completed within two weeks prior to the group onset. Analysis of markers included activated T-cells (CD26+CD2+CD3+), T-helper cells (CD4+CD3+), T-suppressor/cytotoxic cells.(CD8+CD3+), and T4/T8 ratio (CD4/CD8). Subjects were also given the Profile of Mood States (POMS; with Anger, Anxiety, Depression, Confusion, Fatigue., and Vigor subscales), The Sickness Impact Profile (SIP; measuring psychosocial impairment), and the Attitudes Towards Self Scale (ATS; with Self-Criticism, Generalization, and High Standards factors).

Results: The POMS depression and anger subscales and the total POMS score were significantly correlated with the T4/T8 ratio (r = 44, p < .04, r = .50, p < .02; r = .44, p < .05, respectively). In addition, the POMS depression and anger subscales also showed a marginal inverse correlation with T-suppressor/cytotoxic cell percentage (r = .37, p < .09; r = -.40, p < .07, respectively). Similarly, the psychosocial impairment score of the SIP was also correlated with the T4/T8 ratio (r = .33, p < .09) and negatively associated with the percentage of T-suppressor/cytotoxic cells (r = -.36, p < .06). The Generalization factor of the ATS measuring the tendency to generalize negative events, was significantly correlated with activated T-cell numbers (r = .39, p < .04), T4-helper cell numbers (r =.37, p < .06), and T-helper cell percentages (r = .39, p < .05).

Conclusions: Our findings suggest that immune activation, as defined by elevated activated T-cells, T-helper cells, and T4/T8 ratios, is associated with measures of psychological distress and attitudes about oneself. The elevations in distress and impairment measures were also associated with a reduction in the percentage of T-suppressor/cytotoxic cells. There may be a subset of CFS patients exhibiting a depleted T-suppressor/cytotoxic cell population that tend to be more symptomatic and more emotionally distressed. Psychological distress may have a stronger relationship with immunologic functioning in those CFS patients who are the most ill, as suggested by increased activated T-cells, T-helper cells, and T4/T8 ratios, and a reduction in T-suppressor/cytotoxic percentages. Since the ATS was associated with immune activation, attitudes about oneself may mediate the association between psychological distress and decreased T-suppressor/cytotoxic cells. These findings are based on pilot data, and therefore, conclusions about whether or not the CFS subjects had significantly elevated T4/T8 ratios or abnormally low T-suppressor/cytotoxic percentages as compared to normal controls can not be made without further study.

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Immunocytology and in vitro reverse-transcriptase activity in CFS

AuthorsM.I. Holmes, R. Easingwood, J. Cross, and J. Faed

Objective: This paper describes two blind clinical trials of paired, age, sex and ethnically matched patients with CFS; 24 pairs of patients and controls in the first, and 18 in the second. In the first, the range of duration of symptoms was 1 to 3.5 (mean 1.5) years and in the second 1 to 5 (mean 1.7) years. Peripheral blood lymphocyte (PBL) cultures were assayed in triplicate for reverse-tran scriptase (RT) activity, and examined by EM for the presence of virus-like structures at days 0 and 12 and CD2, 3, 4, 8, 16, 20, and 31, and B1 phenotypes were counted at day 0 by FACScan.

Methods: A single dose of 1 µg ml-¹Concanavilin A (Con A) was given to all cultures at day 0. At days 4, 8 and 10 they were given 4.5 ng ml-¹ human recombinant IL-2. Cells were harvested at day 12 for EM studies, and ultracentrifuged supernatants and cells for RT assay using a poly rA:oligo dT template-primer and measuring RT activity by uptake of tritiated thymidine triphosphate.

Results: In Trial 1, RT activity up to 3 times background was observed in 9, and virus-like structures in 7 of 24 patients, and not in controls. Group means showed a significant CD4 cytopenia. In Trial 2, RT activity at levels of 2 to 4 times background were observed in 5 patients, and virus-like structures were observed in 4 of these, and not in controls. Group means showed significantly reduced CD4/CD8 ratios and an NK cytopenia. RT activity and EM virus-like structures were seen almost exclusively in the cohort of patients who identified the onset of their condition with a non-specific, acute febrile illness and whose duration of symptoms was 2 years or less.

Conclusion: These studies suggest the in vitro RT activity and the presence of virus-like structures in PBLs may correlate in CFS with patients who relate the onset of their condition to a non-specific, acute febrile illness.

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Lymph node morphology and phenotype in Chronic Fatigue Syndrome

Authors: Nancy Klimas, Kevin Maher, Roberto Patarca, Jean Walling, Mack Smith, and Mary Ann Fletcher. University of Miami School of Medicine and the Miami Veterans Affairs Medical Center, Miami, Florida.

Objective: Chronic Fatigue Syndrome is an illness which is associated with immune dysfunction, including abnormalities in the function and activation status of peripheral blood lymphocytes. There has been no study of the lymph node compartment in this illness.

Methods: Patients who had volunteered to undergo lymph node biopsy while participating in the pre-clinical phase and phase I studies of a novel immunomodulatory therapy were evaluated. At the time of this abstract 6 lymph nodes have been evaluated. An additional 9 will be studied prior to the October AACFS meeting.

Results: Of the 6 lymph nodes currently available for study, 1 is tonsillar, 2 posterior cervical, and three are axillary. Four demonstrated normal architecture and morphology. Two had a slight increase in the number of histiocytes. The tonsillar sample was, as expected, predominantly B-cells (56%). Excluding the tonsillar sample, the lymph nodes showed the following lymphocyte surface phenotype: The CD4 percent ranged from 61% to 83% (mean 70%), the CD8 percent from 8% to 19% (mean 11%) and B-cells (CD 19+) ranging from 13% to 30% (mean 20%). Few natural killer cells were present. CD56+CD3- lymphocytes ranged from 0.9% to 2.2%. The activation marker, CD26 was present on 63% to 77% of the CD2+ lymphocytes. Memory CD4 cells predominated with only 16% (range of 2% to 30%) expressing the naive subset marker, CD45RA. Not surprisingly, the majority of the CD4 cell population also expressed 62L (86%), which is considered to be a lymph node homing adhesion molecule. CD8 cells were predominantly CD38+ (95%) and DR negative.

Discussion: This initial study of the distribution of lymphocyte subsets in the lymph nodes from patients with CFS offers confirmation of our hypothesis regarding the immunopathogenesis of this syndrome. The data from the first 6 patients presented here indicates a preponderance of activated T-cells that is even higher than that reported in peripheral blood. The CD8 cells express the phenotype CD38+ and DR-, which is associated with cytotoxic T-cells. The ratio of memory to naive CD4 T-cells is skewed upward, as is the case in peripheral blood. These findings are compatible with a chronically activated immune status in this patient group.

(Supported in part by The CFIDS Association of America)

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Immunological response in Chronic Fatigue Syndrome (CFS) following a graded exercise test to exhaustion

Authors: J. LaManca, S. Sisto, J. Ottenweller, A. Peckerman, S. Cook, T. Denny, W. Gause and B.H. Natelson. NJ CFS Research Center, New Jersey Medical School, and VA NJ Health Care System, East Orange, NJ.

Objective: Data have indicated that CFS may involve abnormal immune system functioning. Exercise can be an useful tool in stimulating an immune response. This study was conducted to evaluate the immunological response of CFS patients to an exhaustive bout of exercise.

Methods: The two groups, which were matched for age, sex, and socioeconomic background, consisted of 19 CFS patients meeting the 1988 CDC criteria and 14 non-exercising (i.e. sedentary) controls respectively. All were females in the luteal phase of the menstrual cycle. Venipuncture was performed pre-, 4 minutes post-, 1 hour post-, and 24 hours post-exercise. The graded exercise test (GXT) consisted of walking on a treadmill with the work load increasing every 3 minutes to voluntary exhaustion. Expired gas was analyzed during the GXT. Peripheral blood cells were labeled with the following monoclonal antibody combinations: CD3/CD8, CD3/CD4, CD3/CDl9, and CD3/CD (16&56). The cytokines: IFN-gamma, IL-2, IL-4, IL-10, IL-12, and TNF-alpha were assayed utilizing a quantitative RT/PCR assay. White blood cells (WBC) and all subset concentrations were adjusted for plasma volume shifts.

Results: During the GXT no significant differences in VO2peak (28.6 ± 1.7 vs 30.8 ± 1.2 ml:kg-1min -1), HRpeak(175 ± 4 vs 183 ± 4 bpm) and RER (1.08 ± .02 vs 1.13 ± .02) were seen when comparing CFS to CON. The number of WBC, CD3+CD8+, CD3+CD4+, T-Cells, B-Cells, Natural Killer Cells, and IFN-gamma all were significantly changed across time (all comparisons, p < .0l). No significant group differences were seen for any of the immune variables. Also the changes across time were similar for both groups with the exception of IL-2, where a significant group-by-time interaction was seen (p < .03). Only one of the CFS patients had elevated blood lymphocyte levels 24 hours after the GXT.

Conclusion: The immune response of patients with CFS to a single bout of exhaustive exercise is not significantly different when compared to a group of sedentary healthy controls.

(This work was supported by NIH Center grant AI-32247)

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Could nickel allergy be involved in the pathogenesis of Chronic Fatigue Syndrome?

Authors: J.A. Marcusson¹ , G. Lindh², B. Evengård²
¹Department of Dermatology, Huddinge University Hospital, Karolinska Institute, Stockholm, Sweden
²Division of Infectious Diseases of the Department of Immunology, Microbiology, Pathology and Infectious Diseases, Huddinge University Hospital, Karolinska Institute, Stockholm, Sweden

Objective: Allergic reactions have been reported to occur more often in patients with CFS, and fatigue is also a common self-reported symptom among patients with allergy. The aim of this study was to study whether allergies to various metals might play a role in the pathogenesis of chronic fatigue syndrome.

Method: Fifty patients fulfilling the chronic fatigue syndrome (CFS) criteria according to CDC, and 73 sex and age-matched healthy controls without history of metal allergy or symptoms of fatigue, were percutaneously patch-tested with eight different metal salts. The following metal salts were used as test reagents: potassium dichromate, metallic mercury, cobalt chloride, nickel sulphate, gold sodium thiosulphate, palladium dichloride, tin and silver nitrate. Test reagents and test chambers were obtained from Chemotechnique Diagnostics AB, Malmö, Sweden. The test chambers were removed after 2 days and the skin reaction was graded after another 2 days as 0, 1+, 2+ and 3+ respectively. Only 2+ and 3+ reactions were accepted as true positive.

Result: Some type of contact allergy to tested metal salts was noted in 46% of the patients with CFS and in 36% of the healthy coritrols. Nickel sulphate allergy was significantly more common among the CFS patients, and this was even more noteworthy among the females. We found nickel allergy in 52% of the female CFS patients compared to 19% of the female control group (p < 0.05). No significant difference was found in skin reaction to the other metal salts tested in this study.

Conclusion: