Oscillations in inositol 1,4,5-trisphosphate and diacyglycerol induced by vitamin D3 metabolites in confluent mouse osteoblasts

Abstract

For the last 5 years, attention has focused on the nongenomic effects of 1,25-(OH)2D3, but considerably less is known about the mechanisms of the nonnuclear actions of 24,25-(OH)2D3. The present study examines and compares the rapid (5-90 s) effects of 100 pM to 10 nM 24,25-(OH)2D3, 10 pM to 1 nM 1,25-(OH)2D3, and 1-100 nM 25-OHD3 on the formation of inositol phosphates and lipids in confluent mouse osteoblasts. 24,25-(OH)2D3 and 25-OHD3 effects were dose dependent; those of 1,25-(OH)2D3 were dose dependent in a bell-shaped manner. The two dihydroxylated metabolites induced a multiphasic response in inositol 1,4,5-trisphosphate (IP3) formation with three stimulation peaks; the IP3 response to 25-OHD3 was monophasic. The amplitude of the IP3 response to 24,25-(OH)2D3 was greater and its oscillation period was slower than that induced by 1,25-(OH)2D3. The diacylglycerol (DAG) responses to secosteroids showed two stimulation peaks that appeared at different times depending on the secosteroid used. Pretreatment with neomycin totally inhibited the first DAG response; neomycin had no effect on the second peak of DAG induced by 25-OHD3, whereas it partially blocked the second response of DAG to 24,25-(OH)2D3 and 1,25-(OH)2D3. These data show for the first time that 24,25-(OH)2D3 can modulate phospholipid metabolism in confluent mouse osteoblasts as early as 5-10 s. The first pathway used by all three secosteroids is that of the hydrolysis of phosphatidylinositol 4,5-bisphosphate via phospholipase C activation, leading to the formation of the two second messengers, IP3 and DAG, since neomycin totally blocked the response. Thus, the action of these secosteroids on the osteoblast membrane may also implicate several steps of the phosphatidylcholine cycle, according to the metabolite tested. Finally, these data point to a direct interaction of vitamin D metabolites with specific membrane recognition moieties.

Contributions of pro‐oxidant and anti‐oxidant conditions to the actions of 24,25‐dihydroxyvitamin D3 and 1,25‐dihydroxyvitamin D3 on phosphate uptake in intestinal cells

“…These observations are also in keeping with the earlier report that 24,25(OH) 2 D 3 inhibits 1,25(OH) 2 D 3 ‐induced increases in PKC activity [Nemere et al, 2000]. Other labs have also reported distinct effects of 24,25(OH) 2 D 3 on osteosarcoma cells [Yukihiro et al, 1994; Takeuchi and Guggino, 1996], mouse osteoblasts [Grosse et al, 1993], and rat chondrocytes (Pedrozo et al, 1999]. Our work has relevance at the physiological level: in an agricultural context, our findings have applicability to raising poultry in that the phosphate content of the manure can upset the ecological balance in waterways.…”