Immunotherapy of Chronic Fatigue Syndrome: Therapeutic Interventions Aimed at Modulating the Th1/Th2 Cytokine Expression Balance
December 23, 2002
Source: Journal of Chronic Fatigue Syndrome, Vol. 8(1) 2001 © 2001
Roberto Patarca-Montero, MD, PhD Nancy G. Klimas, MD Mary Ann Fletcher, PhD
Roberto Patarca-Montero, Nancy G. Klimas, and Mary Ann Fletcher are Directors, E.M. Papper Laboratory of Clinical Immunology, Department of Medicine, University of Miami School of Medicine, 1600 NW 10th Street, Miami, FL 33136.
Address correspondence to: Roberto Patarca-Montero at the above address (E-mail: email@example.com).
ABSTRACT. Based on the postulates of viral and autoimmune etiologies of CFS, several interventions have been designed and tested by different research groups around the world, including the United States, Sweden, United Kingdom, Italy, and Japan. This review addresses those interventions aimed at altering the balance of certain cytokines, the mediators of immune responses. Patients with CFS who show evidence of activation of the immune system have poor immune cell function and a predominance of what is called a T -helper (Th)2-type cytokine response when their lymphocytes are activated. A Th2-type response, which is characterized by production of cytokines such as interleukin (IL)-4, -5, and -10, favors the function of B lymphocytes, the cellular factories of immunoglobulins.
A predominance of a Th2-type response is therefore consistent with pathologies, such as autoimmunity and atopy, which are based on inappropriate production of immunoglobulins. Many of the CFS therapies discussed decrease the Th2-type predominance seen at baseline in CFS patients, thereby allowing a greater predominance of a Th1-type response, which favors the function of macrophages and natural killer cells. The function of the latter cells, which have the natural ability of directly destroying invading microbes and cancer cells, is defective in untreated CFS patients. Typical Th1-type cytokines include IL-2 and interferon-gamma, and some of the therapies induce their production.
The interventions discussed in this review cover a wide spectrum of therapeutic tools ranging from lymph node cell immunotherapy, herbal products, and small molecules to vaccines. Despite the controversies on the etiology of CFS, immunotherapy research is useful and necessary. [Article copies available for a fee from The Haworth Document Delivery Service: 1-800-342-9678. E-mail address: Website: © 2001 by The Haworth Press, Inc. All rights reserved.]
KEYWORDS. Vaccine, lymph node, cytokines, viruses
Chronic fatigue syndrome (CFS) is characterized by debilitating fatigue that is not attributable to known clinical conditions, that has lasted for more than six months, that has reduced the activity level of a previously healthy person by more than 50 percent, and that has been accompanied by flu-like symptoms (e.g., pharyngitis, adenopathy, low-grade fever, myalgia, arthralgia, headache) and neuropsychological manifestations (e.g., difficulty concentrating, exercise intolerance, and sleep disturbances) (1-7).
Although syndromes are clusters of non-chance associations, and the components of a syndrome can be generally related to a common element, the cause of CFS still remains to be determined. CFS is frequently of a sudden onset. Possible precipitating factors include infections, psychiatric trauma, and exposure to toxins (7-10). Even among those who favor a microbial etiology for CFS, it is not yet clear whether CFS is a consequence of a chronic microbial infection or of an acute microbial infection which resolves but whose sequel in the form of autoimmunity or other manifestations is responsible for the pathology seen. Microbial reactivation may also play a causal or perpetuating role in disease manifestation.
A review of the literature on the immunology of CFS reveals that people who have Chronic Fatigue Syndrome (CFS) have two basic problems with immune function that have been documented by most research groups: 1. immune activation, as demonstrated by elevation of activated T lymphocytes, including cytotoxic T cells, as well as elevations of circulating cytokines; and 2. poor cellular function, with low natural killer cell cytotoxicity (NKCC), poor lymphocyte response to mitogens in culture, and frequent immunoglobulin deficiencies, most often IgG1 and IgG3. These findings have a waxing and waning temporal pattern which is consistent with episodic immune dysfunction (with predominance of so called T-helper type 2 and proinflammatory cytokines and low NKCC and lymphoproliferation) that can be associated as cause or effect of the physiological and psycho logical function derangement and/or activation of latent viruses or other pathogens.
The interplay of these factors can account for the perpetuation of disease with remission/exacerbation cycles. Therapeutic intervention aimed at induction of a more favorable cytokine expression pattern and immune status is discussed.
One of our models of CFS holds that the interaction of psychological factors (distress associated with either CFS-related symptoms or other stressful life events) and immunologic dysfunction (indicate din signs of chronic overactivation with cytokine abnormalities) contribute to: (a) CFS-related physical symptoms (e.g., fatigue, joint pain, cognitive difficulties, fever) and increases in illness burden; and (b) dysfunction in the immune system's ability to survey viruses including latent herpesviruses (indicated in impaired NKCC).
As discussed above, there is a decrease in the ratio of type 1/type 2 cytokines produced by lymphocytes in vitro following mitogen stimulation in CFS patients. This type of dysfunction should be expected to result in impaired immune surveillance associated with cytotoxic lymphocytes. For example, Cohen et al. (11) found an association between psychosocial stressors, immunomodulation, and the incidence and progression of rhinovirus infections in healthy normals. Here, the rates of respiratory infections and clinical colds increased in a dose-response fashion with increases in psychological stress across all five of the cold viruses studied.
If viruses related to upper respiratory tract infections (URIs) are not well controlled by immune surveillance mechanisms (e.g., NKCC) in CFS patients who are exposed to stressors, then patients may suffer more frequent and protracted URIs which are accompanied by prolonged elevations in proinflammatory cytokines. Stress-associated reactivation of latent herpesviruses may also play a role in modulating the production of cytokines that underlie CFS symptom exacerbations (12,13).
Alternatively, distress increases may more directly influence cytokine dysregulation by way of neuroendocrine changes which in turn intensify physical symptoms. Importantly, for all of the possible paths, further increases in distress as a "reaction" to mounting symptoms creates a vicious cycle. Such a recursive system may act as a positive feedback loop thereby accounting for the chronic nature of CFS and its refraction to interventions that focus solely on symptom reduction.
INFECTIOUS AGENTS AS POSSIBLE DIRECT CAUSES OF CFS
Many families of viruses summarized below have been studied in association with CFS, including herpesviruses, enteroviruses, retroviruses, lentiviruses, adenoviruses, Borna disease virus, parvoviruses, and arboviruses. There are also many historical accounts of diseases of presumed viral etiology that present similar to CFS, including George Reinhold Forster's description of the Tapanui flu and the documentation of Akureyri or Iceland disease (14,15). It has also been proposed that reactivation of certain viruses may play a role in the pathophysiology of CFS but may not be its primary cause.
Enteroviruses: Enteroviruses (Coxsackie virus A and B, echovirus, poliovirus) belong to a group of small RNA-viruses, picornavirus, which are widespread in nature. Enteroviruses cause a number of well known diseases and symptoms in humans, from subclinical infections and the common cold to poliomyelitis with paralysis. Serologic and molecular biology techniques have demonstrated that enteroviral genomes, in certain situations, persist after the primary infection, which is often silent. Persistent enteroviral infection or recurrent infections and/or virus-stimulated autoimmunity might contribute to the development of diseases with hitherto unexplained pathogenesis, such as post-polio syndrome, dilated cardiomyopathy, juvenile (type 1) diabetes and possibly some cases diagnosed as CFS (16-20). Several studies have failed to document persistent enteroviral infections in CFS (21-23).
Herpesviruses: Herpesviruses (Epstein-Barr virus, cytomegalovirus, human herpes virus types 6 and 7, herpes simplex virus types 1 and 2) have been associated with CFS. For instance, reactivation/replication of a latent herpesvirus (such as Epstein Barr virus) could modulate the immune system to induce CFS (24-26). In this respect, serologically proven acute infectious illness secondary to Epstein-Barr virus (EBV) is associated with a range of nonspecific somatic and psychological symptoms, particularly fatigue and malaise rather than anxiety and depression (27).
Although improvement in several symptoms occurs rapidly, fatigue commonly remains a prominent complaint at 4 weeks, and resolution of fatigue is associated with improvement in cell-mediated immunity. A prospective cohort study of 250 primary care patients also revealed a higher incidence and longer duration of an acute fatigue syndrome, and a higher prevalence of CFS, after glandular fever as compared to after an ordinary upper respiratory tract infection (28).
In another study, anti-EBV titers were higher among CFS patients and were associated with being more symptomatic (29). However, testing of 548 chronically fatigued, including patients with CFS, for antibodies to 13 viruses (herpes simplex virus 1 and 2, rubella, adenovirus, human herpesvirus 6, Epstein Barr virus, cytomegalovirus, and Coxsackie B virus, types 1-6) in patients found no consistent differences in any of the seroprevalences compared with controls (30).
Some studies suggest an association between human herpesvirus-6 (HHV 6) (Roseolovirus genus of the Betaherpesvirus subfamily) and CFS (31-34). One study found that a high proportion of CFS patients (50% by antibody testing and up to 80% by nested-PCR detection of viral DNA but not RNA) were infected with HHV 6 but with low viral load. The latter results do not support HHV 6 reactivation in CFS patients (33). Other studies have addressed a possible association between HHV 7 and CFS. Use of the supernatant fluid from HHV 7 infected cells as antigen in immunoassays yielded high and low HHV 7 antibody in sera from chronic fatigue patients and healthy donors as controls, respectively (35).
Stealth viruses: Cloned DNA obtained from the culture of an African green monkey simian cytomegalovirus-derived stealth virus contains multiple discrete regions of significant sequence homology to portions of known human cellular genes (36). The stealth virus has also been cultured from several CFS patient and a cytopathic stealth virus was also cultured from the cerebrospinal fluid of a nurse with CFS. The findings lend support to the possibility of replicative RNA forms of certain stealth viruses (37).
Review of the clinical histories and brain biopsy findings of 3 patients with severe stealth virus encephalopathy showed that the patients initially developed symptoms consistent with CFS (38). One patient has remained in a vegetative state for several years, while the other two patients have shown significant, although incomplete, recovery.
Histological and electron-microscopic studies revealed vacuolated cells with distorted nuclei and various cytoplasmic inclusions suggestive of incomplete viral expression. There was no significant inflammatory response. Viral cultures provided further evidence of stealth viral infections occurring in these patients (38). Partial sequencing of stealth virus segments isolated from a CFS patient revealed a fragmented genome and sequence microheterogeneity, observations that suggest that both the processivity and the fidelity of replication of the viral genome are defective (39). An unstable viral genome may provide a potential mechanism of recovery from stealth viral illness.
Retroviruses: Some studies (40,41) looked into a possible link between retroviruses and CFS but no conclusive evidence has been garnered.
Lentiviruses: Although structures consistent in size, shape and character with various stages of a Lentivirus replicative cycle were observed by electron microscopy in 12-day peripheral-blood lymphocyte cultures from 10 of 17 CFS patients and not in controls, attempts to identify a lymphoid phenotype containing these structures failed and the results of reverse-transcriptase assay of culture supernatant fluids were equivocal (42).
Parvovirus B19: The spectrum of disease caused by parvovirus B19 has been expanding in recent years because of improved and more sensitive methods of detection. There is evidence to suggest that chronic infection occurs in patients who are not detectably immunosuppressed. A young woman with recurrent fever and a syndrome indistinguishable from CFS was found to have persistent parvovirus B19 viremia, which was detectable by polymerase chain reaction despite the presence of IgM and IgG antibodies to parvovirus B19 (43). Testing of samples from this patient suggested that in some low viremic states, parvovirus B19 DNA is detectable by nested PCR in plasma but not in serum. The patient's fever resolved with the administration of intravenous immunoglobulin.
Ross River Virus: A prospective investigation revealed that serologically proven acute infectious illness due to Ross River virus is associated with a range of nonspecific somatic and psychological symptoms, particularly fatigue and malaise rather than anxiety and depression (44). Although improvement in several symptoms occurs rapidly, fatigue commonly remains a prominent complaint at 4 weeks. Resolution of fatigue is associated with improvement in cell-mediated immunity as measured by delayed-type hypersensitivity skin responses.
Borna disease virus: Borna disease virus (BDV) is a neurotropic, non-segmented, negative-sense single-strand RNA virus. Natural infection with this virus has been reported to occur in horses and sheep. Recent epidemiological data suggest that BDV may be closely associated with neuropsychiatric disease (depression and schizophrenia) in humans (45-52). In Japanese patients with CFS, the prevalence of BDV infection is up to 34%. Furthermore, anti-BDV antibodies and BDV RNA were detected in a family cluster with CFS. These results suggest that BDV or a related agent may contribute to or initiate CFS, although the single etiologic role of BDV is unlikely (45-52).
Borrelia: Despite antibiotic treatment, a sequel of Lyme disease may be a post-Lyme disease syndrome (PLS), which is characterized by persistent arthralgia, fatigue, and neurocognitive impairment (53-56). Although patients with CFS and PLS share many features, including symptoms of severe fatigue and cognitive impairment, patients with PLS show greater cognitive deficits than patients with CFS compared with healthy controls. This is particularly apparent among patients with PLS without premorbid psychiatric illness (57).
Chlamydiae: Some authors have proposed a link between Chlamydiae bacteria and CFS (58).
Mycoplasma: Multiplex polymerase chain reaction analysis to detect the presence of mycoplasma genus DNA sequences in 100 CFS patients revealed that 52% were infected with Mycoplasma genus as compared to 15% of healthy individuals. Mycoplasma fermentans, M. hominis and M. penetrans were detected in 32, 9 and 6% of the CFS patients but only in 8, 3 and 2% of the healthy control subjects, respectively (59). An analysis of studies based on the use of forensic polymerase chain reaction or polymerase chain reaction found that 52-63% of CFS/FMS patients (n-1000) had mycoplasmal infections, whereas 9-15% of controls (n-450) tested positive (59-63).
Rickettsiae: Several links have been proposed since 1991 between CFS and chronic Rickettsial infection (64-66), including: (a) CFS and Rickettsial infection present with a similar symptomatology; (b) CFS was reported in Incline, Nevada in 1984 (67) and developed into epidemic proportions. Rocky Mountain Spotted Fever originated form the same place in 1916 (68). Drury described the spirochete Borrelia duttoni, in 1702 as causing recurrent Malgach fever. In 1975, Borrelia burgdorferi was found in Connecticut, giving birth to a new name, Lyme disease; (c) a link has been established between CFS and Florence Nightingale (69) working surrounded by lice, fleas and ticks during the Crimean war. Soldiers were presenting with epidemic typhus, the common disease of wars, regularly reported since the time of Hannibal up to modern times; (d) lymphocyte studies conducted on sheep with tick-borne-diseases (70), CFS patients and patients with Q fever endocarditis (71) have shown similar results; (e) CFS was proposed to overlap with post Q fever syndrome (72).
Post-Q-fever fatigue syndrome (QFS) is characterized by inappropriate fatigue, myalgia and arthralgia, night sweats, changes in mood and sleep patterns following about 20% of laboratory-proven, acute primary Q-fever cases, a condition caused by Coxiella burnetti (72-74). The condition is associated with high levels of interleukin-6 and although improvement in several symptoms occurs rapidly, resolution of fatigue takes longer and it is associated-with improvement in cell-mediated immunity as measured by delayed-type hypersensitivity skin responses (75). The recovery rate associated with treatment of CFS patients with tetracyclines by several practitioners is 84-96% (58,76,77).
Yersinia: A study based on the detection of antibodies to various Yersinia outer membrane proteins (YOPs) in serum samples from 88 CFS patients and 77 healthy age- and gender-matched controls concluded that Yersinia enterocolitica is unlikely to play a major role in the etiology of CFS (78).
AUTOIMMUNITY AND CFS: INFECTIOUS AGENTS AS POSSIBLE INDIRECT CAUSES OF CFS
Another etiological hypothesis for CFS is that an acute microbial infection triggers an autoimmune response; i.e., when the body mounts an attack against the virus, it selects the production of immunoglobulins that can also recognize and attack the body itself. This happens because of what is termed "molecular mimicry" (79-93). The portions of the microbe that the antibodies recognize somehow resemble those of proteins that normally constitute the human body. The evidence for autoimmunity in CFS comes from several sources. One research team found that approximately 52% of sera from CFS patients react with nuclear envelope antigens (94). Some sera from CFS patients immunoprecipitated the nuclear envelope protein lamin 131 (95).
Another report documented a high frequency (83%) of autoantibodies to insoluble cellular antigens (vimentin and lamin B1) in CFS, a unique feature which might help to distinguish CFS from other rheumatic autoimmune diseases (96). The possible autoimmune etiology of CFS is further underscored by preliminary evidence for an association between CFS and the presence of HLA-DQ3 (97).
Several studies have documented the presence in CFS patients of rheumatoid factor (98-105), antinuclear antibodies (95,97-99,101-106), antithyroid antibodies (105,108,109), anti-smooth-muscle antibodies (108), antigliadin, cold agglutinins, cryoglobulins, and false serological positivity for syphilis (104,108). No circulating antimuscle and anti-CNS antibodies were found in 10 CFS patients (110), and one group found no significant differences in the number of positive tests for autoantibodies in CFS patients (111).
One team found that among children who chronically complain of nonspecific symptoms such as headache, fatigue, abdominal pain, and low grade fever, those who were ANA positive tended to have general fatigue and low grade fever, while gastrointestinal problems such as abdominal pain and diarrhea and orthostatic dysregulation symptoms were commonly seen in ANA negative patients (112). Children who were unable to go to school more than one day a week were seen significantly more among ANA positive patients than among negative patients. Based on these observations, the authors concluded that autoimmunity may play a role in childhood chronic nonspecific symptoms and proposed a new disease entity: the autoimmune fatigue syndrome in children.
The features shared between CFS and autoimmune diseases may complicate diagnosis. For instance, three cases of dermatomyositis had been erroneously diagnosed as CFS because of the presence of elevated titers of serum Epstein-Barr virus antibodies (113). In one study, one-third of CFS patients with sicca symptoms fulfilled the diagnostic criteria for Sjoegren's syndrome, but they were 'seronegative,' differing from the ordinary primary Sjoegren's syndrome (114).
An additional confounding feature is that patients with primary Sjoegren's syndrome report more fatigue than healthy controls on all the dimensions of the Multidimensional Fatigue Inventory and, when controlling for depression, significant differences remain on the dimensions of general fatigue, physical fatigue, and reduced activity (115). The negative correlation between levels of noradrenaline and general fatigue in patients with primary Sjoegren's syndrome may imply the involvement of the autonomic nervous system in the chronic fatigue reported in this syndrome (115,116).
Although fatigue in patients with systemic lupus erythematosus (SLE) does not correlate with disease activity, it is correlated with fibromyalgia, depression, and lower overall health status (117). Fatigue is also a major symptom in patients with ankylosing spondylitis and, unlike SLE, it is more likely to occur with active disease but it may occur as a lone symptom (118). Fatigue is also common in osteoarthritis and rheumatoid arthritis, associates with measures of distress, and is a predictor of work dysfunction and overall health status (119). Several studies have reported that rheumatoid arthritis-related fatigue is strongly associated with psychosocial variables, apart from disease activity per se (120,121).
Fatigue is associated to a large extent with pain, self-efficacy towards coping with disease, towards asking for help and problematic social support and female gender. One study found large individual differences in variation of pain and fatigue among rheumatoid arthritis patients (122). Stressors were associated with increased pain but not fatigue. Subjects with poor sleep had higher levels of pain and fatigue. Diurnal cycles of pain and fatigue were found, yet were observed for only some patients.
CYTOKINE IMBALANCES IN CFS AND RELATED DISORDERS
The increased autoimmune manifestations in CFS, along with the decreased natural killer cell cytotoxic and lymphoproliferative activities, would be compatible with the hypothesis that the immune systems of affected individuals are biased toward a T -helper (Th)2 type, or humoral immunity (antibody producing)-oriented cytokine expression pattern, over a Thl type, or cellular mediated (natural killer cell and macrophage activating) immunity-oriented one. Potent immunogens can have systemic long-lasting non-specific effects on the nature of the immune response to unrelated antigens.
In particular, vaccinations or infections can exert a systemic effect, and non-specifically increase or reduce the Th1/Th2 cytokine balance of the response to other unrelated antigens (123) and affect (positively or negatively) survival from unrelated diseases (124,125).
Based on the fact that Gulf War personnel were given multiple Th2-type response-inducing vaccinations, Rook, Stanford and Zumla in international published patent number WO-09826790 presented the hypothesis that Gulf War syndrome represents a special case of CFS, where the Th2-inducing stimuli can be identified. The authors pointed out that induction of a systemic Th2 switch is underscored by four features of the vaccination protocol used for the Gulf War troops:
1. Pertussis was used as an adjuvant in British troops in the Persian Gulf and its adjuvanticity is potently Th2-type response-inducing (126-128). This property of pertussis has recently led to discussion of the possibility that its use in children contributes to the contemporary increased prevalence of atopy (129,130).
2. Gulf War troops were given Th2-inducing immunogens against plague, anthrax, typhoid, tetanus and cholera. Such a cumulatively large antigen load would tend to drive the response towards a Th2-type response predominance (124,131,132). The measles vaccine, when used at the standard dose, reduced mortality by considerably more than can be accounted for by the incidence of measles in the unvaccinated population. It has been reported that diphtheria, tetanus, and pertussis vaccines (Th2-inducing) do not show this nonspecific protective effect (125). However, when a high titer measles vaccine was used the mortality increased, although protection from measles itself was maintained (124,125). There is evidence that this increase in mortality was accompanied by a switch towards a Th2-type response, and dose-related increases in the induction of a Th2-type component are well established for several other immunogens (131,132).
3. The vaccinations were given after deployment of the troops in the war zone, or just before they travelled there, at a time when stress levels would have been high. Immunization in the presence of raised levels of glucocorticoids (i.e., cortisol) drives the cytokine expression response by stimulated lymphocytes towards a Th2-type predominance (133-135). Several steroid hormones modulate T -cell responses. The nervous, endocrine and immune systems respond to internal and external challenges and communicate and regulate each other by means of shared or system-unique hormones, growth factors, neuro transmitters and neuromodulators.
For instance, similar alterations in central catecholamine neurotransmitter levels are associated with immune activity and stressor exposure, alterations that are more pronounced in aged as opposed to younger animals (136). For example, a decreased norepinephrine turnover in the hypothalami and brain stem of rats occurs at the peak of the immune response to sheep red blood cells (137,138), and increased serotonin metabolism is associated with depressed Arthus reaction and plaque-forming cell response in rats stressed either by overcrowding lasting two weeks or more or by repeated immobilization for four days (139,140).
The long-term effects of these acute changes are evidenced by chronic variable stress which facilitates tumor growth (141) and is associated with immune dysregulation in multiple sclerosis (142). The hypothalamic-pituitary-adrenal axis plays a pivotal role in stress-mediated changes, and stimulation of corticotropin-releasing factor in the central nervous system (143,144) has been shown to suppress rapidly a variety of immune responses, an effect which can be blocked by infusion into the brain of alpha-melanocyte-stimulating hormone, a tridecapeptide derived from pro-opiomelanocortin (145).
Besides external stimuli, intrinsic imbalances in neurotransmitter levels affect the immune system either directly by acting on immunocompetent cells or indirectly via induction of hormonal secretions. For instance, depression is associated with neurotransmitter imbalances and with decreased natural killer cell cytotoxic activity (146-149). Moreover, several studies have documented the existence of striking physiologic, neuroendocrine, metabolic, and pharmacologic differences between depressed and normal subjects and between depressed and severely ill subjects (150,151).
Dehydroepiandrosterone (DHEA) or unknown metabolites of DHEA, tend to promote a Th1-type response pattern. Thus, DHEA can restore immune functions in aged mice through correction of dysregulated cytokine release (152,153). DHEA has been tested for similar properties in aged humans (154) and found to enhance production of Th1-type cytokines, such as interleukin(IL)-2 and interferon(IFN)-gamma (155-158). DHEA also enhances IL-2 secretion from human peripheral blood T cells (159). These effects of DHEA are the reverse of the effects of glucocorticoids such as cortisol which enhance Th2-type response activity and synergize with Th2-type cytokines (160-163).
If proliferation of naive T lymphocytes is driven in the presence of a non-specific stimulus (134) or by an antigen (as follows vaccination), T lymphocytes with a Th2-type cytokine profile will develop. This has been rather clearly shown with spleen cells from laboratory rodents which have few memory cells under normal circumstances (135).
Overall, cortisol favors the development of a Th2 cytokine profile form naive cells (134). This point must not be confused with the fact that the cytokine secreting activity of established Th2-type cells is readily inhibited by cortisol. Thus, the use of cortisol analogs for conventional treatments of Th2-mediated diseases, such as eczema, asthma and hay fever may work via anti-inflammatory effects, and by reducing cytokine production by Th2-type cells (164), and yet at the same time the use of cortisol will encourage perpetuation of the underlying problem by driving newly recruited T cells towards a Th2-type response.
Psychological and physical stress activate the hypothalamic-pituitary-adrenal axis, and thereby lead to a variety of changes including increased production of cortisol. In this respect, excessive exercise and deprivation of food and sleep result in a falling ratio of DHEA to cortisol. The latter falling ratio correlates directly with a fall in delayed hypersensitivity (DTH) responsiveness (a Th1-type response marker), and there is a simultaneous rise in serum IgE levels. IgE is wholly dependent upon Th2-type cytokine production (133). This is to be expected in the light of known effects of DHEA and cortisol outlined above. A further example of the effect of stress on Th1- to Th2-type switching is the increase in antibody to Epstein-Barr virus in students reacting in a stressed manner to their exams. This virus is usually controlled by a Th1-type response and cytotoxic T cells. Loss of control results in virus replication and increased antibody production (165).
Similarly, peripheral blood leukocytes from medical students during exam periods showed lower mRNA for IFN-gamma, a Th1-type cytokine (166). Similar points can be demonstrated in a more controlled manner in animals. Stress secondary to crowding or restraint can increase mycobacterial growth in tuberculous mice (167,168). This is a model that is acutely sensitive to the presence of even a small Th2 component (169,170).
In tuberculosis, there is a systemic shift to a Th2-type response predominance (169,170), and an unusual pattern of metabolites of adrenal steroids is excreted in the urine (170). Treatment of the disease restores the Th1-type predominance and corrects the pattern of steroid metabolites, so that metabolites of cortisone increase relative to metabolites of cortisol (169,170). There is considerable evidence that depression can be associated with excessive cortisol-mediated effects in the brain (171,172), and stress can lead to depression. Thus, depression (as seen in CFS and Gulf War Syndrome) tends to associate with Th2-mediated disorders, such as asthma, eczema, and some endocrine changes are common to Th2 disorders and to depression (173,174).
Treatment of depression with the drug metyrapone causes the same change in steroid metabolites (e.g., increase in metabolites of cortisone relative to metabolites of cortisol) as those described above after treatment of tuberculosis (172,173).
4. The troops were also exposed to carbamate and organophosphorus insecticides, and these inhibit IL-2-driven phenomena essential for normal Th1-type function (175). The importance of this component is uncertain. However, it has been rumored that the insecticides were often obtained from local sources in the Gulf, so purity was not known, and even more toxic contaminants may have been present.
Thus, multiple vaccinations administered under these circumstances may have caused a long-lasting systemic cytokine imbalance. The same effect would occur sporadically in the general population, secondary to vaccinations or other Th2-type response-inducing environmental stimuli and infections, and could account for the widespread incidence of CFS. It should be stressed at this point that not all vaccines and infectious agents induce a preponderance of the Th2-type response.
For instance, measles infection reduced the incidence of atopy, and of allergic reactions to house dust mite (123). Similarly, Japanese children that are tuberculin skin-test positive are less likely to be atopic than are tuberculin negative children, and their ratio of circulating Th1/Th2 cytokines is higher. Moreover, after repeated injection of Bacillus Camelet-Guerin (BCG), those in whom tuberculin conversion occurs have an increased probability of losing their atopic symptoms.
THERAPIES AIMED AT CORRECTING Th1 /Th2 CYTOKINE IMBALANCES
Although vaccines, stressful stimuli and some pathogens have been shown to lead to long-term, nonspecific shifts in cytokine balance (124-126), the factors that could lead to a Th2 shift in CFS patients are unknown. Nevertheless, several therapeutic regimens which induce a systemic Th1 bias, some based on the use of certain vaccines, are being tested with preliminary success in subpopulations of CFS patients with documented baseline immune abnormalities.
Moreover, whether directly or indirectly, viruses may play a role in the etiology or the perpetuation of symptoms of CFS. Some authors have put forth the notion that reactivation of latent viruses, if not etiological, may serve as a perpetuation factor for CFS symptomatology and may account for the remission-exacerbation cycling nature of the disease. On the other hand, viral reactivation may be an epiphenomenon and not necessarily related to symptomatology. While mainly focused on therapeutic interventions, this section will address therapies based on the hypotheses discussed above and others on how local and/or systemic effects of acute or reactivated viral activity may underlie CFS.
The first half of the 20th century witnessed the first successful approach to control the spread of several viral infections: the development and worldwide use of vaccines. The concept of vaccination was originally developed by Jenner in 18th century England based on the observation that milkmaids exposed to cows with cowpox were protected from smallpox.
In this case, a subclinical infection with one virus was protective of an infection with a related one. The latter concept was also extended to the treatment of various infectious diseases by giving to the patient even unrelated but more innocuous infectious diseases. Although in many cases the treatment was worse than the disease, the therapeutic approach was somehow useful with particular combinations of infectious agents.
Outstanding triumphs from worldwide vaccination programs have been the eradication of smallpox and predictably soon of poliomyelitis (176). After smallpox was eliminated as an infectious disease in Great Britain in 1962, two outbreaks occurred, one in 1973 and one in 1978, when smallpox virus under study in laboratories infected susceptible individuals. In both incidents, deaths resulted (176). With the eradication of poliomyelitis throughout the world soon to be accomplished, steps are being taken to prevent polioviruses that remain in laboratories from escaping into the community and causing disease.
These examples stress the need for universal availability of vaccines to effectively eradicate the disease they cause. Unfortunately, we do not have vaccines against all viruses and even in the cases for which we do, the vaccine is not universally available. The dramatic success in immunizing children against childhood diseases stands in stark contrast to the much lower percentages of adults who are adequately immunized against common adult diseases. In the case of the flu vaccine, the influenza virus keeps changing and the vaccine has to be updated every year, and it is therefore not fully protective against all viral strains.
One alternative to vaccines has been the use of injections of immunoglobulins, the natural bullets that the body produces to kill foreign invaders. Not too long ago, physicians advocated the use of immunoglobulin injections as a way to boost the body's immune defenses and heighten resistance against microbes. The latter reasoning was perhaps again reflective of the old wisdom of using one infection to protect against another with the added refinement of using the natural mediators of the body's attack machinery against infections instead of the infectious agent itself. Some of the therapeutic interventions addressed below are based on the use of vaccines and the modulation of the body's defenses using certain microbes.
Lymph Node Cell-Based Immunotherapy
As presented in an accompanying article in this issue of the journal, the group of Drs. Nancy Klimas, Mary Ann Fletcher and Roberto Patarca at the University of Miami completed a safety and feasibility study using lymph node extraction, ex vivo cell culture, followed by autologous cell reinfusion as a treatment strategy to favor a Th2- to Th1-type cytokine expression shift in selected CFS patients (177).
Lymph nodes were obtained from patients who met the current case definition for CFS and the following inclusion criteria: a history of acute onset; a Karnofsky score less than 80; evidence of immune dysfunction in three or more of the following: greater than one standard deviation above controls for elevated soluble TNF receptor type I (sTNF-RI) levels in serum, elevated sTNF-RI production in phytohemmaglutinin (PHA)-stimulated blood culture or elevated IL-5 production in PHA-stimulated blood culture; lymphocyte activation (CD2 + CD26 + cells > 50%); or low NK cell cytotoxic activity (<20%). The lymph node cells were cultured for 10 to 12 days with anti-CD3 and IL-2. These cells were then reinfused into the donor who was monitored for safety and possible clinical benefit.
There were no adverse events noted in this Phase 1 clinical trial. Of 13 subjects, two had palpable lymph nodes that proved fibrotic with no viable cells. Of the remaining 11 subjects, all successfully underwent expansion and reinfusion. In some of the patients, there was an elevation in the expression of IL-2 receptor on CD4 T cells in the weeks following the reinfusion. There was a significant decrease in IL-5 production by PHA-stimulated blood cultures observed at 1 week which persisted for several weeks post-infusion. Levels of PHA-induced IFN-gamma production did not change.
There was a trend towards an increase in the ratio of IFN-gamma/IL-5 starting at week 1 and persisting at least 12 weeks post-infusion. Of the 11 subjects in the trial who had cells reinfused, 9 had significant cognitive improvement; other measures of severity of illness also trended towards improvement. The lack of adverse effects from this experimental approach to immunomodulation in CFS and the favorable clinical and immunologic results observed in the small number of patients studied suggest that further clinical trials are warranted.
The studies on CFS patients described above were preceded by studies of adoptive CD8+ T-cell immunotherapy of AIDS patients with Kaposi's sarcoma (178-183). The research group used a device developed to selectively capture CD8+ T cells for ex vivo culture, and instituted basic science and clinical evaluations of the consequences of the infusion, with rIL-2, of autologous, activated, and polyclonally-expanded CD8+ T cells in AIDS patients with Kaposi's sarcoma and oral hairy leukoplakia. Phase I and II trials showed safety and suggested efficacy in the treatment of the latter AIDS associated conditions. The intervention affected the patterns of cytokine expression of CD8+ T lymphocytes and favored a restoration of a strong type 1 cytokine expression response (183).
As described in international published patent number WO-09826790 by Rook, Stanford and Zumla, preparations of killed Mycobacterium vaccae are able to effect a nonspecific systemic Th1-type response bias, in particular by down-regulation of Th2-type activity without concomitant up-regulation of Th1-type activity. The latter feature is similar to the effect on the Th1/Th2 type cytokine proportions of the lymph node cell-based immunotherapy described above.
In experimental animals, a nonspecific systemic bias away from Th2-type activity on administration of M. vaccae can be seen as a reduction in the titer of an IL-4 (Th2)-dependent antibody response to ovalbumin (an allergen unrelated to M. vaccae itself), in mice pre-immunized so as to establish a Th2-type response. A single injection of M. vaccae is able to cause this effect, and further injections can enhance it. The effect is nonspecific because it does not require the presence of any component of ovalbumin in the injected preparation.
Briefly, Balb/c mice 6-8 weeks old were immunized with 50 µg ovalbumin emulsified in oil (incomplete Freund's adjuvant) on days 0 and 24. This is known to evoke a strong Th2-type pattern of response, accompanied by IgE production, and priming for release of two Th2-type cytokines, IL-4 and IL-5. Animals then received saline or 107 autoclaved M. vaccae on days 53 and 81 by subcutaneous injection. Injections of M. vaccae reduced the rise in IgE levels caused by immunization with ovalbumin. The reduction caused by treatment with M. vaccae was significant at all time points tested.
Similarly, spleen cells from the immunized animals failed to release IL-5 in vitro in response to ovalbumin if the donor animals had been treated with M. vaccae, while spleen cells from immunized animals treated with saline released large quantities of IL-5 in response to ovalbumin. The latter data shows that M. vaccae will reduce a Th2-type pattern of response, even when given after immunization with a potent allergen, and without epitopes of the Th2-inducing molecule. There is therefore a nonspecific systemic down-regulation of the Th2-type pattern of response, not dependent upon a direct adjuvant effect on the allergen itself.
In cancer patients, the effect of M. vaccae injection has been demonstrated by the appearance in the peripheral blood of lymphocytes that spontaneously secrete IL-2 (a characteristic Th1 cytokine) and decrease in T cells that secrete IL-4 (a characteristic Th2 cytokine) after stimulation with phorbol myristate acetate and calcium ionophore. The percentage of lymphocytes showing this activated TM-type phenotype increases progressively after each successive injection of M. vaccae, reaching a plateau in many individuals after 3-5 injections of 109 organisms (days 0, 15, 30, and then monthly).
The M. vaccae used for these therapies is grown on a solid medium including modified Sauton's medium solidified with 1.3% agar. The medium is inoculated with the microorganisms and incubated aerobically for 10 days at 32°C to enable growth of the microorganism to take place. The microorganisms are then harvested, weighed and suspended in diluent to give 100 mg of microorganisms/mL of diluent.
The suspension is then further diluted with buffered saline to give a suspension containing 10 mg wet weight (about 1010 cells) of microorganisms/mL of diluent and dispensed into 5 mL multidose vials. The vials containing the live microorganisms are then autoclaved (115-125° C) for 10 minutes at 69 kPa to kill the microorganisms. The therapeutic agent thus produced is stored at 4° C before use. A suspension of 0.1 mL containing 1 mg wet weight (about 109 cells) of M. vaccae, is shaken vigorously immediately before being administered by intradermal injection over the left deltoid muscle.
In the same patent publication, Rook, Stamford and Zumla describe their experience with CFS patients treated with M. vaccae. For instance, a CFS patient reported improvement after two injections of a M. vaccae preparation. A second one reported that, since she had been receiving a M.vaccae preparation at two month intervals, her CFS symptoms and food allergy had improved considerably and she believes to be very well as long as she continues with her regular injections.
In international published patent number WO-09829133 by Goteborg University Science Invest AB, Carl-Gergard Gottfries and Bjoern Regland describe the use of staphylococcal vaccine to favor a Th1-type predominance. In an accompanying article in this issue of the journal, these authors also addresses variables, such as smoking and nickel sensitivity that affect the outcome of this form of therapy. The treatment is preferably conducted as a series of administrations with increasing doses during a specific period.
Preferably, the vaccine is administered in 8-10 increasing doses during 4-12 weeks, preferably 8-10 weeks. The reason for the increasing doses is that during the first week or weeks, the patient will probably suffer from side effects, and it is therefore advantageous to start with a low dose. The side effects will diminish after some time. The first series of administrations is followed by repeated administrations given approximately once a week for 5-15 weeks, preferably for 10 weeks.
To prevent recurrence, the repeated administrations are then followed by a maintenance treatment with administrations approximately once a month, which preferably are continued for several years, such as 1-10 years, preferably approximately 5 years. The doses in the repeated administrations of the maintenance treatment are preferably constant and relatively high. Vitamin B12 and/or folacin is preferably administered simultaneously or in parallel with the staphylococcal preparation.
If the known staphylococcal vaccine Staphypan Berna from the Serum & Vaccine Institute, Bern, Switzerland is used, a typical treatment schedule may be as follows: 8-10 administrations are made during a period of 4-12 weeks, preferably 8-10 weeks, wherein the dose of the staphylococcal preparation is gradually increased from 0.1 to 1 mL of the pure vaccine. The increase depends on the response from the patient. It may be, for example, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 and 1.0 mL, respectively. If the patient shows a strong local reaction, it is possible to repeat a dose before increasing it. The dose of staphylococcal preparation in the repeated administration and in maintenance treatment is 1 mL.
As described in the published patent, after a pilot study comprising eight patients was made, a double-blind placebo-controlled study was performed, comprising a group of 24 women patients fulfilling both the criteria for fibromyalgia and for chronic fatigue syndrome. Seven of the 13 patients who received the staphylococcal preparation were assessed as being minimally improved, three as being much improved, and the remaining three were unchanged. In the placebo group, three patients were minimally improved, while the remaining eight were unchanged.
The improvement in the group with active treatment was statistically significant (p < 0.05) compared to the improvement in the placebo group. Following the controlled study, 24 patients chose to continue with the treatment and 20 of these have been treated between one and two years. Nineteen of these 20 patients were on the sick-list or received sickness pension prior to the start of treatment, and one patient was employed part-time. At a one-year follow-up after the completed study, nine of the 20 patients were in full- or part-time paid employment, while one patient was taking part in a work experience program and one was at the middle of a two-year training to become a nurse. The treatment strategy used in the above study is a series of administrations of staphylococcal preparations given approximately once a week during a period of some months, for example, three months and thereafter long-term treatment with monthly administrations. Further studies are being conducted by this Swedish group.
Sizofiran, an immunostimulant extracted from suehirotake mushroom (Schizophyllum commune) cultured fluid, is under development by Fidia Farmaceutici Italiani Deriviate Industriali a Affini for the potential treatment of cancer and hepatitis B. Sizofiran is licensed by Kaken Pharmaceutical Co., Ltd., Japan. Trials were underway for the treatment of gastric and lung tumor; phase III trials are underway for the treatment of hepatitis B; and the compound is in phase II trials for chronic fatigue syndrome. By August 1999, Kaken was preparing its NDA filing for hepatitis B.
Significant sizofiran-induced rises in IFN-gamma and IL-2 in culture medium of phytohemagglutinin (PHA) or concanavalin A-stimulated peripheral blood mononuclear cells have been observed. Further reports on sizofiran should become available in the near future.
In one study, an extract of the herb Panax ginseng was evaluated for its capacity to stimulate cellular immune function by peripheral blood mononuclear cells (PBMC) from normal individuals and patients with chronic fatigue syndrome. PBMC isolated on a Ficoll-Hypaque density gradient were tested in the presence or absence of varying concentrations of the extract for natural killer (NK) cell cytotoxic activity directed against K562 cell targets and for antibody-dependent cellular cytotoxicity (ADCC) directed against human herpesvirus 6-infected H9 cells (184).
Ginseng, at concentrations greater or equal to 10 micrograms/kg, significantly enhanced NK cell function in both groups. Similarly, the addition of the herb significantly increased ADCC of PBMC from the subject groups. Thus, an extract of Panax ginseng enhances cellular immune function of PBMC from normal individuals as well as from patients with depressed cellular immunity and CFS (184).
In line with the latter observations, Ginseng treatment was found in another study to lead to activation of neutrophils and modulation of the immunoglobulin G response to Pseudomonas aeruginosa, thereby enhancing the bacterial clearance and reducing the formation of immune complexes, effects which resulted in a milder lung pathology in chronic Pseudomonas aeruginosa lung infection in cystic fibrosis patients. The therapeutic effects of Ginseng may be related to activation of a Th1-type of cellular immunity and down-regulation of humoral immunity (185).
Although Ginseng is generally well tolerated, it has been implicated as a cause of decreased response to warfarin and may interfere with either digoxin pharmacodynamically or with digoxin monitoring (186-188). Nevertheless, some authors claim no relationship between cardiac glycosides and glycosides in Ginseng and attribute the effect on digoxin on contaminants (189,190). Additionally, ginseng may cause headache, tremulousness, and manic episodes in patients treated with phenelzine sulfate. Ginseng should also not be used with estrogens or corticosteroids because of possible additive effects. Ginseng may affect blood glucose levels and should not be used in patients with diabetes mellitus (187).
A group of Japanese herbal medicines called Hozai have been used to improve the physical condition of the elderly. One representative Hozai, Juzen-taiho-to, was shown to modulate antigen-specific T-cell responses toward more balanced Th1/Th2-type responses in old BALB/c mice, which have a preferential Th2-type cytokine response pattern (191). Such effects may help prevent the development of diseases associated with immunodysregulation, including chronic fatigue syndrome.
Immunotherapy of CFS holds promise and is an area of intense research that is being fed by a plethora of sources from herbal medicine, vaccines, and cell therapy to small molecules. Although the mechanisms whereby alterations in the immune system lead to changes in cognitive and functional status remain to be elucidated, the results of the different trials reinforce the need to integrate immunological, endocrinological and psychological approaches to the understanding of the different manifestations of CFS. Some of the clinical trials also emphasize the importance of appropriately categorizing patients for interventions and outcome assessments.
The experience garnered with CFS appears applicable to related conditions, such as fibromyalgia, Gulf War syndrome, sick building syndrome and multiple chemical sensitivity, among others. The etiology of CFS remains elusive but therapeutic intervention studies are opening the doors to control this ailment.
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