Transcriptomic signatures in whole blood of patients who acquire a chronic inflammatory response syndrome (CIRS) following an exposure to the marine toxin ciguatoxin.

BACKGROUND: Ciguatoxins (CTXs) are polyether marine neurotoxins found in multiple reef-fish species and are potent activators of voltage-gated sodium channels. It is estimated that up to 500,000 people annually experience acute ciguatera poisoning from consuming toxic fish and a small percentage of these victims will develop a chronic, multisymptom, multisystem illness, which can last years, termed a Chronic Inflammatory Response Syndrome (CIRS). Symptoms of ciguatera CIRS include fatigue, cognitive deficits, neurologic deficits, pain and sensitivity to light. There are few treatment options for ciguatera CIRS since little is known about its pathophysiology. METHODS: This study characterizes the transcriptional profile in whole blood of 11 patients with ciguatera-induced CIRS and 11 normal controls run in duplicate using Agilent one color whole genome microarrays. Differential expression was determined by using a combination of moderated t-test p-value and fold change (FC). Significant genes were subjected to gene ontology, principal component analysis and SVM classification. Seven significant genes found by microarray were validated by PCR. RESULTS: Using a low stringency (p < 0.05 and FC > 1.4) and a high stringency (p < 0.01 and FC > 1.5) filter, the resulting gene sets of 185 and 55, respectively, showed clear separation of cases and controls by PCA as well as 100% classification accuracy by SVM, indicating that the gene profiles can separate patients from controls. PCR results of 7 genes showed a 95% correlation to microarray data. Several genes identified by microarray are important in wound healing (CD9, CD36, vWF and Factor XIII), adaptive immunity (HLA-DQB1, DQB2, IL18R1 and IL5RA) and innate immunity (GZMK, TOLLIP, SIGIRR and VIPR2), overlapping several areas shown to be disrupted in a mouse model of acute exposure to ciguatoxin. Another area of interest was differential expression of long, non-coding sequences, or lncRNA. CONCLUSIONS: Disruptions of innate and adaptive immune mechanisms were recorded at both the genomic and proteomic level. A disruption in the HLA-T cell receptor axis could indicate HLA haplotype sensitivity for this chronic syndrome, as noted in many autoimmune conditions. Taken together, these indicators of illness provide additional insights into pathophysiology and potential therapies.

Structural brain abnormalities in patients with inflammatory illness acquired following exposure to water-damaged buildings: a volumetric MRI study using NeuroQuant®.

Executive cognitive and neurologic abnormalities are commonly seen in patients with a chronic inflammatory response syndrome (CIRS) acquired following exposure to the interior environment of water-damaged buildings (WDB), but a clear delineation of the physiologic or structural basis for these abnormalities has not been defined. Symptoms of affected patients routinely include headache, difficulty with recent memory, concentration, word finding, numbness, tingling, metallic taste and vertigo. Additionally, persistent proteomic abnormalities in inflammatory parameters that can alter permeability of the blood-brain barrier, such as C4a, TGFB1, MMP9 and VEGF, are notably present in cases of CIRS-WDB compared to controls, suggesting a consequent inflammatory injury to the central nervous system. Findings of gliotic areas in MRI scans in over 45% of CIRS-WDB cases compared to 5% of controls, as well as elevated lactate and depressed ratios of glutamate to glutamine, are regularly seen in MR spectroscopy of cases. This study used the volumetric software program NeuroQuant® (NQ) to determine specific brain structure volumes in consecutive patients (N=17) seen in a medical clinic specializing in inflammatory illness. Each of these patients presented for evaluation of an illness thought to be associated with exposure to WDB, and received an MRI that was evaluated by NQ. When compared to those of a medical control group (N=18), statistically significant differences in brain structure proportions were seen for patients in both hemispheres of two of the eleven brain regions analyzed; atrophy of the caudate nucleus and enlargement of the pallidum. In addition, the left amygdala and right forebrain were also enlarged. These volumetric abnormalities, in conjunction with concurrent abnormalities in inflammatory markers, suggest a model for structural brain injury in "mold illness" based on increased permeability of the blood-brain barrier due to chronic, systemic inflammation.

Defining the neurotoxin derived illness chronic ciguatera using markers of chronic systemic inflammatory disturbances: a case/control study.

BACKGROUND: Ciguatoxins are extremely potent neurotoxins, produced by tropical marine dinoflagellates, that persistently enter into our food web. Over 100,000 people annually experience acute ciguatera poisoning from consuming toxic fish. Roughly 5% of these victims will develop chronic ciguatera (CC), a widespread, multisymptom, multisystem, chronic illness that can last tens of years. CC is marked by disproportionate disability and non-specific refractory symptoms such as fatigue, cognitive deficits and pain, and is suggestive of other illnesses. Its unknown pathophysiology makes both diagnosis and treatment difficult. OBJECTIVES: We wanted to compare objective parameters of visual contrast sensitivity testing, measures of innate immune response and genetic markers in cases to controls to assess the potential for the presence of persistent inflammatory parameters that are demonstrated in other biotoxin associated illnesses at a single specialty clinic. METHODS: Using 59 CC cases and 59 controls we present in retrospective review, in all cases, abnormalities in immune responses paralleling the chronic systemic inflammatory response syndrome seen in several other chronic diseases. RESULTS: This study defines a preliminary case definition using medical history, total symptoms, visual contrast sensitivity, HLA DR genotype analysis, reduction of regulatory neuropeptides VIP and MSH, and multiple measures of inflammatory immune response, especially C4a and TGFß1, thereby providing a basis for identification and targeted therapy. CONCLUSIONS: CC provides a model for chronic human illness associated with initiation of inflammatory responses by biologically produced neurotoxins.

Complement split products C3a and C4a are early markers of acute lyme disease in tick bite patients in the United States.

BACKGROUND: Current laboratory markers do not readily detect acute Lyme disease. We assessed the utility of complement and its split products as markers of Lyme disease in patients shortly after a tick bite. METHODS: Thirty-one consecutive acute Lyme disease patients, 14 with and 17 without erythema migrans (EM) skin rash, seen by a physician within 96 h of a tick bite were matched with 24 consecutive tick bite patients without Lyme disease symptoms and 46 healthy control subjects. Complement and split products measured included factor B, Bb, C4, C3c, C3a(des Arg), C4a(des Arg), C1q- and C3d-containing immune complexes, and C2. RESULTS: C2, C4, C3 and factor B levels were within normal ranges in all groups. C3a and C4a levels were significantly higher in acute Lyme disease patients than in tick bite and healthy control groups (both p < 0.001). All acute Lyme disease patients, regardless of EM, had elevated levels of C3a or C4a. Few tick bite controls had elevated levels of C3a (2/20) or C4a (5/24) and only 1 of the healthy control subjects had elevated C3a (0/46) or C4a (1/32). CONCLUSIONS: These findings suggest that C3a and C4a may be useful markers of Lyme disease in patients seen shortly after tick bite, even in those without EM.

Sick building syndrome (SBS) and exposure to water-damaged buildings: time series study, clinical trial and mechanisms.

Occupants of water-damaged buildings (WDBs) with evidence of microbial amplification often describe a syndrome involving multiple organ systems, commonly referred to as "sick building syndrome" (SBS), following chronic exposure to the indoor air. Studies have demonstrated that the indoor air of WDBs often contains a complex mixture of fungi, mycotoxins, bacteria, endotoxins, antigens, lipopolysaccharides, and biologically produced volatile compounds. A case-series study with medical assessments at five time points was conducted to characterize the syndrome after a double-blinded, placebo-controlled clinical trial conducted among a group of study participants investigated the efficacy of cholestyramine (CSM) therapy. The general hypothesis of the time series study was that chronic exposure to the indoor air of WDBs is associated with SBS. Consecutive clinical patients were screened for diagnosis of SBS using criteria of exposure potential, symptoms involving at least five organ systems, and the absence of confounding factors. Twenty-eight cases signed voluntary consent forms for participation in the time-series study and provided samples of microbial contaminants from water-damaged areas in the buildings they occupied. Twenty-six participants with a group-mean duration of illness of 11 months completed examinations at all five study time points. Thirteen of those participants also agreed to complete a double-blinded, placebo-controlled clinical trial. Data from Time Point 1 indicated a group-mean of 23 out of 37 symptoms evaluated; and visual contrast sensitivity (VCS), an indicator of neurological function, was abnormally low in all participants. Measurements of matrix metalloproteinase 9 (MMP9), leptin, alpha melanocyte stimulating hormone (MSH), vascular endothelial growth factor (VEGF), immunoglobulin E (IgE), and pulmonary function were abnormal in 22, 13, 25, 14, 1, and 7 participants, respectively. Following 2 weeks of CSM therapy to enhance toxin elimination rates, measurements at Time Point 2 indicated group-means of 4 symptoms with 65% improvement in VCS at mid-spatial frequency-both statistically significant improvements relative to Time Point 1. Moderate improvements were seen in MMP9, leptin, and VEGF serum levels. The improvements in health status were maintained at Time Point 3 following a 2-week period during which CSM therapy was suspended and the participants avoid re-exposure to the WDBs. Participants reoccupied the respective WDBs for 3 days without CSM therapy, and all participants reported relapse at Time Point 4. The group-mean number of symptoms increased from 4 at Time Point 2 to 15 and VCS at mid-spatial frequency declined by 42%, both statistically significant differences relative to Time Point 2. Statistically significant differences in the group-mean levels of MMP9 and leptin relative to Time Point 2 were also observed. CSM therapy was reinstated for 2 weeks prior to assessments at Time Point 5. Measurements at Time Point 5 indicated group-means of 3 symptoms and a 69% increase in VCS, both results statistically different from those at Time Points 1 and 4. Optically corrected Snellen Distance Equivalent visual acuity scores did not vary significantly over the course of the study. Group-mean levels of MMP9 and leptin showed statistically significant improvement at Time Point 5 relative to Time Points 1 and 4, and the proportion of participants with abnormal VEGF levels was significantly lower at Time Point 5 than at Time Point 1. The number of participants at Time Point 5 with abnormal levels of MMP9, leptin, VEGF, and pulmonary function were 10, 10, 9, and 7, respectively. The level of IgE was not re-measured because of the low incidence of abnormality at Time Point 1, and MSH was not re-measured because previously published data indicated a long time course for MSH improvement. The results from the time series study supported the general study hypothesis that exposure to the indoor air of WDBs is associated with SBS. High levels of MMP9 indicated that exposure to the complex mixture of substances in the indoor air of the WDBs triggered a pro-inflammatory cytokine response. A model describing modes of action along a pathway leading to biotoxin-associated illness is presented to organize current knowledge into testable hypotheses. The model links an inflammatory response with tissue hypoxia, as indicated by abnormal levels of VEGF, and disruption of the proopiomelanocortin pathway in the hypothalamus, as evidenced by abnormalities in leptin and MSH levels. Results from the clinical trial on CSM efficacy indicated highly significant improvement in group-mean number of symptoms and VCS scores relative to baseline in the 7 participants randomly assigned to receive 2 weeks of CSM therapy, but no improvement in the 6 participants assigned placebo therapy during that time interval. However, those 6 participants also showed a highly significant improvement in group-mean number of symptoms and VCS scores relative to baseline following a subsequent 2-week period of CSM therapy. Because the only known benefit of CSM therapy is to enhance the elimination rates of substances that accumulate in bile by preventing re-absorption during enterohepatic re-circulation, results from the clinical trial also supported the general study hypothesis that SBS is associated with exposure to WDBs because the only relevant function of CSM is to bind and remove toxigenic compounds. Only research that focuses on the signs, symptoms, and biochemical markers of patients with persistent illness following acute and/or chronic exposure to WDBs can further the development of the model describing modes of action in the biotoxin-associated pathway and guide the development of innovative and efficacious therapeutic interventions.

Atovaquone plus cholestyramine in patients coinfected with Babesia microti and Borrelia burgdorferi refractory to other treatment.

Ten percent of US patients with Lyme disease are coinfected with Babesia microti. A double-blind, placebo-controlled, crossover trial enrolled 25 patients with confirmed Borrelia burgdorferi/B microti coinfection, abnormal visual contrast sensitivity (VCS), and persistent symptoms despite prior treatment with atovaquone and azithromycin. Patients were randomly assigned to atovaquone suspension or placebo plus cholestyramine for 3 weeks, were crossed over for 3 weeks, and then received open-label atovaquone and cholestyramine for 6 weeks. Symptoms and VCS scores were recorded at baseline and after weeks 3, 6, 9, and 12. Improvements in symptoms and VCS deficits were observed only after at least 9 weeks of treatment. At week 12, 5 patients were asymptomatic, and 16 had a notable reduction in the number of symptoms. The entire cohort demonstrated significant increases in VCS scores. Adverse effects were rare. Patients coinfected with B burgdorferi and B microti derive measurable clinical benefit from prolonged treatment with atovaquone and cholestyramine. Longer-term combination therapy may be indicated.

A time-series study of sick building syndrome: chronic, biotoxin-associated illness from exposure to water-damaged buildings.

The human health risk for chronic illnesses involving multiple body systems following inhalation exposure to the indoor environments of water-damaged buildings (WDBs) has remained poorly characterized and the subject of intense controversy. The current study assessed the hypothesis that exposure to the indoor environments of WDBs with visible microbial colonization was associated with illness. The study used a cross-sectional design with assessments at five time points, and the interventions of cholestyramine (CSM) therapy, exposure avoidance following therapy, and reexposure to the buildings after illness resolution. The methodological approach included oral administration of questionnaires, medical examinations, laboratory analyses, pulmonary function testing, and measurements of visual function. Of the 21 study volunteers, 19 completed assessment at each of the five time points. Data at Time Point 1 indicated multiple symptoms involving at least four organ systems in all study participants, a restrictive respiratory condition in four participants, and abnormally low visual contrast sensitivity (VCS) in 18 participants. Serum leptin levels were abnormally high and alpha melanocyte stimulating hormone (MSH) levels were abnormally low. Assessments at Time Point 2, following 2 weeks of CSM therapy, indicated a highly significant improvement in health status. Improvement was maintained at Time Point 3, which followed exposure avoidance without therapy. Reexposure to the WDBs resulted in illness reacquisition in all participants within 1 to 7 days. Following another round of CSM therapy, assessments at Time Point 5 indicated a highly significant improvement in health status. The group-mean number of symptoms decreased from 14.9+/-0.8 S.E.M. at Time Point 1 to 1.2+/-0.3 S.E.M., and the VCS deficit of approximately 50% at Time Point 1 was fully resolved. Leptin and MSH levels showed statistically significant improvement. The results indicated that CSM was an effective therapeutic agent, that VCS was a sensitive and specific indicator of neurologic function, and that illness involved systemic and hypothalamic processes. Although the results supported the general hypothesis that illness was associated with exposure to the WDBs, this conclusion was tempered by several study limitations. Exposure to specific agents was not demonstrated, study participants were not randomly selected, and double-blinding procedures were not used. Additional human and animal studies are needed to confirm this conclusion, investigate the role of complex mixtures of bacteria, fungi, mycotoxins, endotoxins, and antigens in illness causation, and characterize modes of action. Such data will improve the assessment of human health risk from chronic exposure to WDBs.