Calcitriol modulates in vivo and in vitro cytokine production: A role for intracellular calcium

Abstract

Background. Several immunomodulatory properties of calcitriol are currently known, however, only little information is available regarding the in vivo and in vitro effects of calcitriol on cytokine production in chronic renal failure. Methods. To study the in vitro effect of calcitriol on lipopolysaccharide (LPS)-induced cytokine production, peripheral blood mononuclear cells (PBMC, 2.5 ml/ml) from 12 chronic dialytic (HD), 15 undialyzed chronic renal failure (CRF) patients and 10 normal subjects (N) were incubated at 37°for 12 hours with 100 ng of LPS (E. coli and P. maltofilia). Increasing doses of calcitriol from 10-10 to 10-9 M were added and cell associated TNF-α and IL-1β were determined by immunoreactive tests after three freeze-thaw cycles. The intradialytic TNF-α and IL-1β production were evaluated in vivo in 12 HD patients before and after three months of intravenous calcitriol treatment (6 μg/week). Intracellular calcium [Ca++ ](i) was determined on PBMC with a cytofluorimetric assay using FLUO-3 AM as the indicator. Results. In vitro, TNF-α increased from 3.6 ± 1.9 pg/cell to 1797 ± 337 in N, from 4.5 ± 1.7 to 1724 ± 232 in CRF and from 3.4 ± 2.3 to 1244 ± 553 in HD after the LPS stimulus. The production of TNF-α was inhibited by calcitriol in a dose-dependent manner [LPS + Vit.D3 100 ng, 2.9 ± 2.1 in N, 3.7 ± 1.9 in CRF and 3.4 ± 1.7 in HD; LPS + Vit.D3 50 ng, 263 ± 296 (N), 6.73 ± 11 (CRF), 38 ± 28 (HD); LPS + Vit.D3 25 ng = 873 ± 583 (N), 325 ± 483 (CRF), 588 ± 507 (HD); LPS + Vit.D3 12.5 ng, 954 ± 483 (N), 912 ± 510 (CRF), 875 ± 527 (HD)]. Comparable data were observed on IL-1β production. In vivo, the intradialytic TNF-α increase (from 8.5 ± 2.3 to 19 ± 5.6 pg/2.5 x 106 cell) during hemodialysis was markedly reduced after calcitriol therapy (from 6.6 ± 3.1 to 11 ± 4.7). [Ca++](i) decreased from 105 ± 25 to 72 ± 18 nM (P < 0.05) and a positive correlation between cytokine levels and [Ca++](i) was found (r = 0.79; P < 0.001). Conclusions. The in vitro increase of cell- associated cytokine after LPS challenge was inhibited by calcitriol in a dose-dependent manner. These data suggest a possible in vivo modulatory effect of calcitriol therapy on cytokine production in hemodialysis.