The objective of the present study was to determine the relationship between 25(OH)D concentrations and inflammatory marker concentrations in healthy women. Although IL-6, IL-10, and CRP did not have a statistically significant relationship with 25(OH)D concentrations, linear regression models revealed a significant inverse relationship between serum 25(OH)D and serum TNF-α concentrations. This relationship remained significant after controlling for potential covariates such as body fat mass, menopausal status, age, or hormonal contraceptive use. Hinton et al
. found that hormonal contraceptive use was associated with greater TNF-α concentrations in young female athletes [32
]. Our data from healthy female non-athletes representing a much wider age range did not reveal such a relationship with TNF-α (P
= 0.2336); however, like Hinton et al
., there was a significant relationship between hormonal contraceptive and serum cortisol level (P
= 0.0030). Interestingly, in our study serum 25(OH)D remained a significant predictor of TNF-α even after controlling for contraceptive use and cortisol concentrations. The lack of significance between serum estradiol and any of the inflammatory markers (data not shown) supports previous research indicating that, in premenopausal women, menstrual phase may affect circulating cytokine concentrations, but the impact is generally not detectable [33
TNF-α is produced by numerous cell types, including macrophages, monocytes, T-cells, smooth muscle cells, adipocytes, and fibroblasts [34
] many of which also have VDR [14
]. Thus, it is difficult to discern the specific mechanisms by which elevations in systemic 25(OH)D attenuate circulating TNF-α concentrations. Nonetheless, our results agree with experimental data showing that vitamin D is capable of suppressing TNF-α production [36
]. Zhu et al
. recently showed that in the colonic tissue of mice with inflammatory bowel disease, 1,25(OH)2
D was capable of down-regulating several genes associated with TNFα, including proteins involved in the transcription of TNFα, one of its primary receptors, and TNF-α itself [39
Human studies of diseased populations have also shown beneficial effects of vitamin D status on TNF-α concentrations. Serum concentrations of TNF-α increased in unsupplemented congestive heart failure patients over a period of 9 months, whereas serum TNF-α concentrations in patients receiving daily supplementation of vitamin D (2000 IU) remained constant [12
]. Calcitriol (1,25(OH)2
) supplementation for 6 months in post-menopausal women with osteoporosis resulted in a significant reduction in serum TNF-α concentrations and an increase in bone mineral density [40
]. Additionally, six months of calcitriol supplementation in hemodialysis patients also caused significant decreases in serum TNF-α [41
]. Our study is the first to show a significant inverse relationship between serum 25(OH)D and TNF-α concentration in a healthy population.
TNF-α concentrations are increased in several disease states such multiple sclerosis (MS), inflammatory bowel disease (IBD), rheumatoid arthritis (RA), heart disease, and osteoporosis; and are often correlated with clinical impairment [42
]. Therefore, attenuating the concentrations of circulating TNF-α has the potential to positively impact the risk for or treatment of such conditions. Our data suggest that serum 25(OH)D status explains ~6% of the variation in TNF-α concentrations in healthy women, thus a mild relationship.
Even a slight drop in circulating TNF-α due to improved vitamin D status may have clinical significance. MS patients with < 2 active brain lesions visible on magnetic resonance imagery were shown to have serum TNF-α concentrations that were slightly but significantly (1.6 pg/mL) less than those with ≥ 2 active brain lesions [44
]. Patients with active ulcerative colitis were found to have 41% greater mean TNF-α concentrations than those with inactive disease (9.46 and 5.54 pg/mL, respectively); while, those with active Crohn's disease had TNF-α concentrations that were only 18% greater than patients with inactive Crohn's (14.0 and 11.5 pg/mL, respectively) [45
Increases in circulating TNF-α concentrations have been associated with heart disease progression. Koller-Strametz reported that TNF-α concentrations were 3.2 ± 0.2 pg/mL in patients with New York Heart Association (NYHA) function class II, 4.0 ± 0.3 pg/mL in NYHA function class III patients, and 5.3 ± 0.9 pg/mL in NYHA function class IV patients [46
Anti-TNF-α medications are efficacious in the management of IBD [47
]. Martinez-Borra et al
. found that patients with lower TNF-α concentrations (14 ± 25 pg/mL) prior to treatment with the anti-TNF drug, infliximab, responded to the treatment, whereas non-responders had significantly higher baseline serum concentrations (201 ± 362 pg/mL) [48
]. Therefore, it is possible vitamin D supplementation may be a viable adjunct to anti-TNF therapy.
Human studies involving diseased populations have shown positive relationships between 25(OH)D concentrations and IL-10 [12
]. Despite this evidence, in the present study, serum IL-10 was not significantly correlated with serum 25(OH)D, suggesting that in healthy adults, vitamin D status does not affect IL-10 secretion into systemic circulation.
Similarly, serum 25(OH)D and serum CRP were not correlated in the present study. As a non-specific inflammatory marker of general wellness, CRP increases with mild chronic infection, aging, and tissue damage [49
]. Research in diseased populations, such as diabetes [50
], arthritis [51
], prolonged chronic illness [53
], and clinical vitamin D deficiency (25(OH)D <27.5 nmol/L) [54
] have demonstrated negative associations between vitamin D status and CRP concentrations. Nevertheless, intervention studies of healthy post-menopausal women [55
] and patients with congestive heart failure [12
] failed to see changes in CRP concentrations after vitamin D supplementation.
Although the result of the linear regression analysis was not statistically significant, there appears to be a slight tendency towards an inverse relationship between 25(OH)D concentrations and serum IL-6 (P = 0.0909). Several in vitro
studies have shown that 1,25(OH)2
D and several of its analogs are capable of inhibiting the production of IL-6 in various cell types [38
]; while most published in vivo
studies have failed to show an effect of vitamin D status on circulating IL-6 concentrations in humans [12
]. One report, however, involving hemodialysis patients with elevated parathyroid hormone (PTH) demonstrated that both oral and intravenous 1,25(OH)2
D supplementation were capable of significantly decreasing serum IL-6 concentrations following 6 months of treatment [62
]. It has been well documented that PTH induces the production of IL-6 by osteoblasts [63
], thus, it is likely that the effects of vitamin D supplementation on serum IL-6 in this population were mediated primarily through the inverse relationship between 25(OH)D and PTH. In our study, there was no relationship between intact PTH and IL-6 concentrations (P = 0.8039). The significant relationship found between age and IL-6 (P = 0.0116) in this study was anticipated due to several reports showing that circulating IL-6 concentrations increase with advancing age [65
]. Further, IL-6 has been implicated in age-associated diseases (such as lymphoproliferative disorders, multiple myeloma, osteoporosis, and Alzheimer's disease) and frailty; and, it is postulated that certain clinically important late-life changes are due to an inappropriate presence of IL-6. Therefore, our results indicating a trend for a negative relationship between vitamin D status and IL-6 concentrations warrants further investigation. The lowering of circulating IL-6 through the improvement of vitamin D nutriture may have the potential to decrease disability and mortality in older populations in addition to helping maintain muscle strength and bone health.
The range of serum 25 (OH)D concentrations observed in our healthy female subjects are in accordance with the overwhelming number of reports documenting the prevalence of vitamin D deficiency and insufficiency in the general population [69
]. In recent years, mounting data has highlighted the need to re-examine vitamin D status and recommendations [76
]. Bischoff-Ferrari et al
. summarized results from randomized controlled trials, prospective and cross-sectional epidemiologic studies, strong mechanistic evidence, and dose-response relationships to determine an optimal serum 25(OH)D concentration [77
]. They showed that for all endpoints (bone mineral density, lower-extremity function, dental health, and risk of falls, fractures, and colorectal cancer), optimal 25(OH)D status began at 75 nmol/L. Our study demonstrates that like these other health outcomes, circulating TNF-α concentrations continue to be associated with serum 25(OH)D concentrations above this point, in a manner consistent with decreased disease risk/progression (i.e. lower TNF-α concentrations).
The primary limitation of this study was sample size. Women who were regularly exposed to UVB light and qualified to participate based on our inclusion and exclusion criteria were far more difficult to recruit than women with minimal UVB exposure. Additionally, women who tan regularly are inherently different from non-tanners. Frequent tanning bed use is associated with high risk behaviours, including frequent dieting, laxative use or vomiting to control weight, cigarette smoking, binge drinking, and recreational drug use. [78
]. In light of this, the present study was designed to control or account for these behaviors through subject inclusion/exclusion criteria and inclusion of pertinent questionnaire data in the multiple regression analysis.