To examine change in health-related quality of life (HRQoL) in association with clinical outcomes of neuropsychiatric (NP) events in SLE.
An international study evaluated newly diagnosed SLE patients for NP events attributed to SLE and non-SLE causes. Outcome of events was determined by physician-completed 7-point scale and compared to patient-completed SF-36 questionnaires. Statistical analysis used linear mixed-effects regression models with patient specific random effects.
274 patients (92% female; 68% Caucasian), from a cohort of 1400, had ≥ 1 NP event where the interval between assessments was 12.3 ± 2 months. The overall difference in change between visits in mental component summary (MCS) scores of the SF-36 was significant (p<0.0001) following adjustments for gender, ethnicity, center and previous score. A consistent improvement in NP status (N=295) was associated with an increase in the mean(SD) adjusted MCS score of 3.66(0.89) in SF-36 scores. Between paired visits where NP status consistently deteriorated (N=30), the adjusted MCS score decreased by 4.00(1.96). For the physical component summary (PCS) scores the corresponding changes were +1.73(0.71) and −0.62(1.58) (p<0.05) respectively. Changes in SF-36 subscales were in the same direction (p<0.05; with the exception of role physical). Sensitivity analyses confirmed these findings. Adjustment for age, education, medications, SLE disease activity, organ damage, disease duration, attribution and characteristics of NP events did not substantially alter the results.
Changes in SF-36 summary and subscale scores, in particular those related to mental health, are strongly associated with the clinical outcome of NP events in SLE patients.
Systemic lupus erythematosus; Neuropsychiatric; Inception cohort; Health related quality of life; SF-36
Health administrative data are frequently used for diabetes surveillance. We aimed to determine the sensitivity and specificity of a commonly-used diabetes case definition (two physician claims or one hospital discharge abstract record within a two-year period) and their potential effect on prevalence estimation.
Following Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, we searched Medline (from 1950) and Embase (from 1980) databases for validation studies through August 2012 (keywords: “diabetes mellitus”; “administrative databases”; “validation studies”). Reviewers abstracted data with standardized forms and assessed quality using Quality Assessment of Diagnostic Accuracy Studies (QUADAS) criteria. A generalized linear model approach to random-effects bivariate regression meta-analysis was used to pool sensitivity and specificity estimates. We applied correction factors derived from pooled sensitivity and specificity estimates to prevalence estimates from national surveillance reports and projected prevalence estimates over 10 years (to 2018).
The search strategy identified 1423 abstracts among which 11 studies were deemed relevant and reviewed; 6 of these reported sensitivity and specificity allowing pooling in a meta-analysis. Compared to surveys or medical records, sensitivity was 82.3% (95%CI 75.8, 87.4) and specificity was 97.9% (95%CI 96.5, 98.8). The diabetes case definition underestimated prevalence when it was ≤10.6% and overestimated prevalence otherwise.
The diabetes case definition examined misses up to one fifth of diabetes cases and wrongly identifies diabetes in approximately 2% of the population. This may be sufficiently sensitive and specific for surveillance purposes, in particular monitoring prevalence trends. Applying correction factors to adjust prevalence estimates from this definition may be helpful to increase accuracy of estimates.
The Systemic Lupus Collaborating Clinics (SLICC) revised and validated the American College of Rheumatology (ACR) SLE classification criteria in order to improve clinical relevance, meet stringent methodology requirements and incorporate new knowledge in SLE immunology.
The classification criteria were derived from a set of 702 expert-rated patient scenarios. Recursive partitioning was used to derive an initial rule that was simplified and refined based on SLICC physician consensus. SLICC validated the classification criteria in a new validation sample of 690 SLE patients and controls.
Seventeen criteria were identified. The SLICC criteria for SLE classification requires: 1) Fulfillment of at least four criteria, with at least one clinical criterion AND one immunologic criterion OR 2) Lupus nephritis as the sole clinical criterion in the presence of ANA or anti-dsDNA antibodies. In the derivation set, the SLICC classification criteria resulted in fewer misclassifications than the current ACR classification criteria (49 versus 70, p=0.0082), had greater sensitivity (94% versus 86%, p<0.0001) and equal specificity (92% versus 93%, p=0.39). In the validation set, the SLICC Classification criteria resulted in fewer misclassifications (62 versus 74, p=0.24), had greater sensitivity (97% versus 83%, p<0.0001) but less specificity (84% versus 96%, p<0.0001).
The new SLICC classification criteria performed well on a large set of patient scenarios rated by experts. They require that at least one clinical criterion and one immunologic criterion be present for a classification of SLE. Biopsy confirmed nephritis compatible with lupus (in the presence of SLE autoantibodies) is sufficient for classification.
Our research objective was to estimate prostate cancer risk in systemic lupus (SLE), relative to the age-matched general population. A progressive literature review was performed to identify SLE cohort studies with cancer registry linkage for cancer ascertainment. Data were pooled from four studies of large SLE cohorts who met these criteria. The total number of prostate cancers observed was derived by pooling the incident cases across all studies. The total expected number of prostate, derived from applying appropriate general population cancer incidence data to the observed number of patient-years of follow-up for each study, was similarly determined. The parameter of interest was the standardized incidence ratio (SIR), the ratio of observed to expected malignancies.
The four studies together provided a pool of 6,068 male SLE patients observed for a total of 38,186 patient years (mean 6.3 years). Within these subjects, 80 prostate cancers observed. In each contributing study, the number of cancers expected far exceeded that observed. The pooled SIR estimate for prostate cancer risk in males with SLE, compared to the general population, was 0.72 (95% CI 0.57, 0.89).
These data suggest a decreased risk of prostate cancer in SLE; more definite conclusions require additional data. Since alterations in androgen pathways can potentially alter prostate risk, a lower risk of prostate cancer in SLE could possibly be due to low hypoadrenergic states which some believe may occur in men with SLE; underlying genetic factors could also be at play. Further study of these issues in large cohorts is needed.
Systemic lupus erythematosus; malignancy; prostate cancer
An increased lymphoma risk is well documented in systemic lupus (SLE). Less attention has been focused on women's cancers, even though SLE affects mostly females. Our objective was to estimate the risk of breast, ovarian, and endometrial cancers in SLE, relative to the general population.
Data were included from five recent studies of large SLE cohorts. The number of cancers observed was determined for each cancer type. The expected number of malignancies was ascertained from general population data. The parameter of interest was the standardised incidence ratio (SIR), the ratio of observed to expected malignancies.
The five studies included 47 325 SLE patients (42 171 females) observed for 282 553 patient years. There were 376 breast cancers, 66 endometrial cancers, and 44 ovarian cancers. The total number of cancers observed was less than that expected, with SIRs of 0.76 (95% CI: 0.69, 0.85) for breast cancer, 0.71 (95% CI: 0.55, 0.91) for endometrial cancer, and 0.66 (95% CI: 0.49, 0.90) for ovarian cancer.
Data strongly support a decreased risk of breast, ovarian, and endometrial cancers in SLE. This may be due to inherent differences in women in SLE (vs the general population) regarding endogenous oestrogen, other medications, and/or genetic make-up.
systemic lupus; SLE; malignancy
Objectives. We examined occupational and non-occupational exposures in relation to risk of SLE in a case–control study conducted through the Canadian Network for Improved Outcomes in SLE (CaNIOS).
Methods. SLE cases (n = 258) were recruited from 11 rheumatology centres across Canada. Controls (without SLE, n = 263) were randomly selected from phone number listings and matched to cases by age, sex and area of residence. Data were collected using a structured telephone interview.
Results. An association was seen with outdoor work in the 12 months preceding diagnosis [odds ratio (OR) 2.0; 95% CI 1.1, 3.8]; effect modification by sun reaction was suggested, with the strongest effect among people who reported reacting to midday sun with a blistering sunburn or a rash (OR 7.9; 95% CI 0.97, 64.7). Relatively strong but imprecise associations were seen with work as an artist working with paints, dyes or developing film (OR 3.9; 95% CI 1.3, 12.3) and work that included applying nail polish or nail applications (OR 10.2; 95% CI 1.3, 81.5). Patients were more likely than controls to report participation in pottery or ceramics work as a leisure activity, with an increased risk among individuals with a total frequency of at least 26 days (OR 2.1; 95% CI 1.1, 3.9). Analyses of potential respirable silica exposures suggested an exposure–response gradient (OR 1.0, 1.4. and 2.1 for zero, one and two or more sources of exposure, respectively; trend test P < 0.01).
Conclusions. This study supports the role of specific occupational and non-occupational exposures in the development of SLE.
Systemic lupus erythematosus; Risk factors; Silica; Ultraviolet radiation; Solvents; Occupation; Environment
To describe vascular events during an 8 year follow-up in a multicentre SLE inception cohort and their attribution to atherosclerosis.
Clinical data including co-morbidities are recorded yearly. Vascular events are recorded and attributed to atherosclerosis or not. All events met standard clinical criteria. Factors associated with atherosclerotic vascular events were analysed using descriptive statistics, t-tests and χ2. Stepwise multivariate logistic regression was used to assess the association of factors with vascular events attributed to atherosclerosis.
Since 2000, 1249 patients have been entered into the cohort. There have been 97 vascular events in 72 patients. These include: myocardial infarction (13), angina (15), congestive heart failure (24), peripheral vascular disease (8), transient ischemic attack (13), stroke (23), pacemaker insertion (1). Fifty of the events were attributed to active lupus, 31events in 22 patients were attributed to atherosclerosis, and 16 to other causes. Time from diagnosis to first atherosclerotic event was 2.0 ± 1.5 years. Compared to patients followed for 2 years without atherosclerosis events (615), at enrolment patients with AVE were more frequently Caucasian, male, older at diagnosis of SLE, obese, smokers, hypertensive and had a family history of coronary artery disease. On multivariate analysis only male gender and older age at diagnosis were associated factors.
In an inception cohort with SLE followed for up to 8 years there were 97 vascular events but only 31 were attributable to atherosclerosis. Patients with atherosclerotic events were more likely to be male and to be older at diagnosis of SLE.
To determine the frequency, accrual, attribution and outcome of neuropsychiatric (NP) events and impact on quality of life over 3 years in a large inception cohort of SLE patients.
The study was conducted by the Systemic Lupus International Collaborating Clinics. Patients were enrolled within 15 months of SLE diagnosis. NP events were identified using the ACR case definitions and decision rules were derived to determine the proportion of NP disease attributable to SLE. The outcome of NP events was recorded and patient perceived impact determined by the SF-36.
There were 1206 patients (89.6% female) with a mean (±SD) age of 34.5±13.2 years. The mean disease duration at enrollment was 5.4±4.2 months. Over a mean follow-up of 1.9±1.2 years 486/1206 (40.3%) patients had ≥1 NP events which were attributed to SLE in 13.0%–23.6% of patients using two a priori decision rules. The frequency of individual NP events varied from 47.1% (headache) to 0% (myasthenia gravis). The outcome was significantly better for those NP events attributed to SLE especially if they occurred within 1.5 years of the diagnosis of SLE. Patients with NP events, regardless of attribution, had significantly lower summary scores for both mental and physical health over the study.
NP events in SLE patients are variable in frequency, most commonly present early in the disease course and adversely impact patients’ quality of life over time. Events attributed to non-SLE causes are more common than those due to SLE, although the latter have a more favourable outcome.
Lupus; Neuropsychiatric; Prospective; Inception cohort
Glucocorticoid therapy is strongly associated with an elevated risk of serious infections in patients with rheumatoid arthritis (RA). The association between glucocorticoids and common non-serious infections (NSI) is not well studied.
A cohort of 16 207 patients with RA aged over 65 years was assembled using administrative data from Quebec. Glucocorticoid and disease-modifying antirheumatic drug (DMARD) therapy were identified from drug dispensing records. NSI cases were defined as first occurrence of a community physician billing code for infection or community-dispensed anti-infectives. A nested case–control analysis was performed considering drugs dispensed within 45 days of the index date, adjusting for age, sex, markers of disease severity, DMARD and comorbidity.
For 13 634 subjects, a NSI occurred during 28 695 person-years of follow-up, generating an incidence rate of 47.5/100 person-years. The crude rate of NSI in glucocorticoid-exposed and unexposed person time was 52.4 and 38.8/100 person-years, respectively. Glucocorticoid therapy was associated with an adjusted RR of 1.20 (95% CI 1.15 to 1.25). A dose response was seen, the adjusted RR increasing from 1.10 (<5 mg prednisolone/day) to 1.85 for doses greater than 20 mg/day. All glucocorticoid risk estimates (including <5 mg/day) were higher than that seen for methotrexate (adjusted RR 1.00; 0.95 to 1.04).
Glucocorticoid therapy is associated with an increased risk of NSI. The magnitude of risk increases with dose, and is higher than that seen with methotrexate, although residual confounding may exist. While the RR is low at 1.20, the absolute risk is high with one additional infection seen for every 13 patients treated with glucocorticoids for 1 year.
Systemic lupus erythematosus (SLE) is a chronic disease of unclear etiology, characterized by an overactive immune system and the production of antibodies that may target normal tissues of many organ systems, including the kidneys. It can arise at any age and occurs mainly in women.
Our aim was to evaluate the potential influence of particulate matter (PM) air pollution on clinical aspects of SLE.
We studied a clinic cohort of SLE patients living on the island of Montreal, followed annually with a structured clinical assessment. We assessed the association between ambient levels of fine PM [median aerodynamic diameter ≤ 2.5 μm (PM2.5)] measured at fixed-site monitoring stations and SLE disease activity measured with the SLE Disease Activity Index, version 2000 (SLEDAI-2K), which includes anti–double-stranded DNA (anti-dsDNA) serum-specific autoantibodies and renal tubule cellular casts in urine, which reflects serious renal inflammation. We used mixed effects regression models that we adjusted for daily ambient temperatures and ozone levels.
We assessed 237 patients (223 women) who together had 1,083 clinic visits from 2000 through 2007 (mean age at time of first visit, 41.2 years). PM2.5 levels were associated with anti-dsDNA and cellular casts. The crude and adjusted odds ratios (reflecting a 10-μg/m3 increase in PM2.5 averaged over the 48 hr prior to clinical assessment) were 1.26 [95% confidence interval (CI), 0.96–1.65] and 1.34 (95% CI, 1.02–1.77) for anti-dsDNA antibodies and 1.43 (95% CI, 1.05–1.95) and 1.28 (0.92–1.80) for cellular casts. The total SLEDAI-2K scores were not associated with PM2.5 levels.
We provide novel data that suggest that short-term variations in air pollution may influence disease activity in established autoimmune rheumatic disease in humans. Our results add weight to concerns that pollution may be an important trigger of inflammation and autoimmunity.
air pollution; antibodies; disease activity; PM2.5; SLE; SLEDAI-2K; systemic lupus erythematous
It is recommended that persons recently diagnosed with heart failure consult with a specialist in heart failure.
To determine whether patients who were diagnosed with new-onset chronic heart failure (CHF) by a noncardiologist consulted with a cardiologist, and identify the factors associated with delayed consultation.
Physician reimbursement administrative data were obtained for all adults with suspected new-onset CHF in the year 2000 in Quebec, defined operationally as a physician visit for CHF (based on the International Classification of Diseases, 9th Revision diagnostic codes), with no previous physician visit code for CHF in the preceding three years. Among those first diagnosed by a noncardiologist, Cox regression modelling was used to identify patient and physician characteristics associated with time to cardiology consultation.
Of the 13,523 persons coded as having incident CHF, 54.9% consulted a cardiologist within the next 2.5 to 3.5 years, and 67.4% were seen by an internist or cardiologist. Older patients, women, and those with lower comorbidity and socioeconomic status had significantly longer times to cardiology consultation.
The data suggest that many patients with suspected new-onset CHF do not receive prompt cardiology care, as stipulated by current recommendations. Equity of access for women and those with lower socioeconomic status appears to be problematic.
Congestive heart failure; Health policy; Referral to cardiologists
Hemiarthroplasty is often the treatment of choice after hip fracture, particularly in frail elderly patients. Such patients may benefit from home care after discharge. We assessed factors associated with the receipt of home care and evaluated the risk of death within three months after discharge.
We obtained administrative data for patients 65 years or older in the province of Quebec who were discharged alive from hospital after hemiarthroplasty during the period 1997–2004. We evaluated destination after discharge and mortality within three months after discharge.
Of 11 326 study patients, 5.6% were discharged home with home care, 29.9% home without home care, 2.0% to a rehabilitation centre, 24.2% to a nursing home and 38.3% to another hospital. Among patients who were discharged home, those who were older, had osteoarthritis, had an emergent admission and were admitted to a high-volume hospital were less likely to receive home care. Discharge with home care was most likely among patients admitted to teaching hospitals, those in hospital for more than seven days, those with atrial fibrillation and those with acute renal failure. Patients who received home care were at lower risk of death than those discharged home without care (hazard ratio 0.57, 95% confidence interval 0.39–0.85).
Less than 16% of the patients discharged home after hemiarthroplasty received home care. Those who received such care had a lower risk of death within three months after discharge.
We have previously developed and validated a self-administered questionnaire, modeled after the Systemic Lupus International Collaborating Clinics Damage Index (SDI), the Lupus Damage Index Questionnaire (LDIQ), which may allow the ascertainment of this construct in Systemic Lupus Erythematosus (SLE) patients followed in the community and thus expand observations made about damage. We have now translated, back-translated and adapted the LDIQ to Spanish, Portuguese and French and applied it to patients followed at academic and non-academic centers in North and South America, Portugal and Spain while their physicians scored the SDI. A total of 887 patients (659 Spanish-, 140 Portuguese- and 80 French-speaking) and 40 physicians participated. Overall patients scored higher than their physicians (total score and all domains) for all versions of the LDIQ. Infrequent manifestations had less than optional clinimetric properties but overall agreement was over 95% for the majority of items. The larger sample size may explain the higher correlations observed among the Spanish-speaking patients. Pending minor adjustments, the LDIQ may prove to be useful in community-based studies. The relationship between the LDIQ and other outcome parameters is currently being investigated in a different patient sample.
What have we learnt about cancer risk in systemic lupus erythematosus (SLE) over the past decade? One important lesson is that data do confirm a slight increased risk in SLE for all cancers combined, compared to the general population. However, it is clear that this is largely driven by an increased risk for hematological malignancies, particularly non-Hodgkin’s lymphoma (NHL), although Hodgkin’s lymphoma may be increased as well. In addition, there is evidence for a moderately increased risk of lung cancer, and possibly for rarer cancer types, such as hepatobiliary and vulvar/vaginal malignancies.
Unfortunately, the most clinically relevant question, the mechanism underlying the association between cancer and SLE, remains largely unanswered. Key issues remaining under study relate to the links between cancer risk, SLE disease activity, and medication exposures. Much of the recent data suggest that disease-related factors may be at least as important as medication exposures for certain cancers, such as NHL. The independent effects of drug exposures versus disease activity in mediating cancer risk in SLE remain unknown. Work is in progress to further elucidate these important issues.
Meanwhile, there is good evidence that cervical dysplasia is increased in women with SLE. This may be mediated by decreased clearance of the human papilloma virus, which some suggest is an innate characteristic of SLE patients. However, an increased risk of cervical dysplasia is also associated with immunosuppressive medication exposures, particularly cyclophosphamide. For these reasons, it is important that women with SLE follow established guidelines for cervical cancer screening.
Malignancy; cancer; systemic lupus erythematosus; SLE; lymphoma; NHL
To determine the population incidence of juvenile rheumatoid arthritis (JRA) in Quebec.
We obtained data from Quebec's physician claims database. Incident cases were defined as having a visit for JRA in 2000, no visit in the previous 3 years, a confirmed diagnosis by an arthritis specialist, or having ≥ 2 visits to any physician for JRA, ≥ 2 months apart but within 2 years.
Cumulative incidence of JRA was 17.8/100,000. Mean age at diagnosis was 9.8 ± 4.6 years, 68% were female and more persons were diagnosed in winter. Subjects had a median of 10 medical visits over the first year.
Our population based incidence estimate was similar to others. Children and adolescents with JRA are heavy users of medical care. Additional study of environmental or climate- related triggers may be warranted.
Objectives: To describe demographic factors, subtypes, and survival of patients with SLE who develop NHL.
Methods: A multi-site cohort of 9547 subjects with definite SLE was assembled. Subjects at each centre were linked to regional tumour registries to determine cancer cases occurring after SLE diagnosis. For the NHL cases ascertained, descriptive statistics were calculated, and NHL subtype frequency and median survival time of patients determined.
Results: 42 cases of NHL occurred in the patients with SLE during the 76 948 patient-years of observation. The median age of patients at NHL diagnosis was 57 years. Thirty six (86%) of the 42 patients developing NHL were women, reflecting the female predominance of the cohort. In the patients, aggressive histological subtypes appeared to predominate, with the most commonly identified NHL subtype being diffuse large B cell (11 out of 21 cases for which histological subtype was available). Twenty two of the patients had died a median of 1.2 years after lymphoma diagnosis.
Conclusions: These data suggest aggressive disease in patients with SLE who develop NHL. Continuing work should provide further insight into the patterns of presentation, prognosis, and aetiology of NHL in SLE.
Patients undergoing hip or knee replacement are at high risk of developing a postoperative venous thromboembolism even after discharge from hospital. We sought to identify hospital and patient characteristics associated with receiving thromboprophylaxis after discharge and to compare the risk of short-term mortality among those who did or did not receive thromboprophylaxis.
We conducted a retrospective cohort study using system-wide hospital discharge summary records, physician billing information, medication reimbursement claims and demographic records. We included patients aged 65 years and older who received a hip or knee replace ment and who were discharged home after surgery.
In total we included 10 744 patients. Of these, 7058 patients who received a hip replacement and 3686 who received a knee replacement. The mean age was 75.4 (standard deviation [SD] 6.8) years and 38% of patients were men. In total, 2059 (19%) patients received thomboprophylaxis at discharge. Patients discharged from university teaching hospitals were less likely than those discharged from community hospitals to received thromboprophylaxis after discharge (odds ratio [OR] 0.89, 95% confidence interval [CI] 0.80–1.00). Patients were less likely to receive thromboprophylaxis after discharge if they had a longer hospital stay (15–30 days v. 1–7 days, OR 0.69, 95% CI 0.59–0.81). Patients were more likely to receive thromboprophylaxis if they had hip (v. knee) replacement, osteoarthritis, heart failure, atrial fibrillation or hypertension, higher (v. lower) income or if they were treated at medium-volume hospitals (69–116 hip and knee replacements per year). In total, 223 patients (2%) died in the 3-month period after discharge. The risk of short-term mortality was lower among those who received thromboprophylaxis after discharge (hazard ratio [HR] 0.34, 95% CI 0.20–0.57).
Fewer than 1 in 5 elderly patients discharged home after a hip-or knee-replacement surgery received postdischarge thromboprophylaxis. Those prescribed these medications had a lower risk of short-term mortality. The benefits of and barriers to thromboprophylaxis therapy after discharge in this population requires further study.
To determine the proportion of family physicians who diagnose rheumatoid
arthritis (RA) correctly and to note how they report they would manage RA
Mailed survey (self-administered questionnaire) requesting comments on
Province of Quebec.
Computer-generated random sample of family physicians registered with the
Quebec College of Family Physicians.
MAIN OUTCOME MEASURES
The proportion of family physicians who recognized RA and their reported
Most respondents recognized the vignette presentation as a case of RA;
133/138 (96.4%) indicated RA as their provisional diagnosis, and all but 1
of the remaining respondents listed RA as a differential diagnosis. Of those
who considered RA as a provisional or possible diagnosis, 107 (77.5% of all
respondents) suggested referring the patient to a rheumatologist. Among the
physicians who suggested referral, none indicated they would initiate
disease-modifying antirheumatic drugs (DMARDs).
Almost all respondents considered RA as a provisional or differential
diagnosis. Although many suggested referring the patient to a
rheumatologist, almost a quarter did not. Initiating DMARDs before referring
patients to rheumatologists appears to be rare. Since DMARDs given during
the early stages of RA are known to decrease damage and dysfunction, ways to
increase their use and optimize care pathways for new-onset inflammatory
arthritis are urgently needed.
Twelve hundred blood samples were obtained in 1986 from pigs slaughtered in western Canada. The samples were assayed for ochratoxin as it is a potential contaminant in the food system. Blood ochratoxin concentration is a good indicator of tissue concentrations, particularly those in the liver and kidney. High performance liquid chromatography analysis of serum for ochratoxin demonstrated that 3.6 and 4.2% of the blood samples collected in February and March (n = 194) and May, June and July (n = 1006), respectively, had ochratoxin concentrations that exceeded 20 ng/mL. Overall the percent of samples that had concentrations of greater than 10, 20, 50, 100 and 200 ng/mL serum were 11.3, 4.1, 1.25, 0.42 and 0.08%, respectively. Samples that had ochratoxin concentrations that were greater than 20 ng/mL of serum were confirmed using two independent methods: liquid chromatography-mass spectrometry analysis and methyl ester derivatization followed by high performance liquid chromatography analysis.