LYSOPHOSPHATIDYLCHOLINE STIMULATES INTERLEUKIN-6 RELEASE FROM RAT ANTERIOR-PITUITARY-CELLS IN-VITRO

Interleukin-6 (IL-6) is a B-cell differentiation-inducing cytokine tha t affects the secretion of several neuroendocrine hormones. Normal rat anterior pituitary (AP) cells synthesize and release IL-6, suggesting a paracrine role for the stimulation of AP hormone release by this cy tokine. We have previously reported that IL-1 beta enhances IL-6 relea se and phospholipase A(2) (PLA(2))-mediated hydrolysis of phosphatidyl choline (PC) in AP cells. Because lysophosphatidylcholine (LPC) may fu nction as a second messenger for IL-lp, we have investigated the effec ts of exogenous LPC on IL-6 release from AP cells in vitro. AP cells f rom male Long-Evans rats were dispersed and cultured for 5-6 days in 9 6-well(100,000 cells/well) culture plates. Cells were rinsed and incub ated in the absence or presence of 1.25-40 mu M LPC 18:0 (stearoyl) fo r 6 h, and IL-6 concentrations determined using the 7TD1 cell bioassay . LPC 18:0 significantly (P < 0.01) stimulated IL-6 release up to 10-f old in a concentration-related manner. In contrast, LPC 18:0 did not a ffect PRL release. LPC species substituted with progressively shorter saturated 1-acyl chains (16:0-10:0) were less effective for IL-6 induc tion. Examination of structurally related glycerophospholipid species revealed the specificity of the LPC stimulation of IL-6 release. Thus, 1.25-40 mu M lysophosphati-dylethanolamine (LPE; 18:0) and lysophosph atidic acid (LPA; 18:0) were without significant effect on AP IL-6 rel ease, demonstrating the specific functional requirement for the phosph orylcholine headgroup. Hydrolysis of the structurally related choline- linked phospholipid sphingomyelin (SM) has been implicated in IL-1 bet a action in certain cell types. Similarly, 1.25-20 mu M lysosphingomye lin (sphingosylphosphorylcholine; SPC) also significantly (P less than or equal to 0.01) stimulated IL-6 release from AF cells, although SPC exhibited discernibly lower potency and efficacy than LPC. An acyl an alog of platelet-activation factor (PAF), i.e. 18:0-2:0 PC tearoyl-2-a cetoyl-sn-glycero-3-phosphorylcholine), differs from LPC by an acetyl group in the sn-2 position; PAF was at least as effective as LPC for t he stimulation of IL-6 release from AP cells in vitro. Stimulation of IL-6 release by LPC 18:0 was completely suppressed by pharmacological inhibitors of protein kinase C such as H7 (20 mu M) and chelerythrine (5 mu M). In addition, H7 (20 mu M) abolished the stimulation of IL-6 release by IL-1 beta (0.16-100 ng/mL). These findings demonstrate that LPC, acyl PAF, or SPC (but not other lysophospholipids) stimulate IL- 6 release from AP cells in vitro. We conclude that LPC-mediated activa tion of protein kinase C is involved in the stimulatory actions of IL- 1 beta in AP cells.