Substance P Mediated Stimulation of Cytokine Levels in Cultured Murine Bone Marrow Stromal Cells

Substance P (SP) is a neuropeptide which has been reported to have immunomodulatory activity. Most studies on SP have been performed on cells of the peripheral immune system. More recently, SP has been reported to have stimulatory activity on human bone marrow cells in vitro, and this activity was dependent on the presence of an adherent layer of cells. The in vitro adherent layer represents the stromal cells of the marrow. In this study, we directly addressed the effect of SP on cultured bone marrow stromal cells. Since stromal cells play an important role in the regulation of hematopoiesis, interactions of neuropeptides such as SP with this cell population could lead to an alteration of stem cell development within the bone marrow. Previously we have shown that SP stimulates protein synthesis in this cell population with two waves of protein synthesis activation, after 2 hr and 48 hr of SP incubation. In this study, we asked whether levels of known stromal cell cytokines were altered in response to SP incubation. We assayed the levels of Interleukin-7 (IL-7) and Stem Cell Factor (SCF) associated with the stromal cell surface following 2 hr and 48 hr of SP incubation. Cells were stimulated with SP for 2 hr or 48 hr. Following SP incubation, cells were washed and the levels of cell associated cytokine was determined by ELISA. Following 2 hr of treatment, 0.1 nM of SP significantly increased (p = 0.05) the level of IL-7 as compared to untreated controls. After 48 hr of treatment, 1, 10, and 100 nM SP significantly increased the levels of IL-7 in this cell population. When SCF levels were assayed, SP at all concentrations tested was found to increase significantly the levels of SCF following 2 hr of incubation. Following 48 hr of incubation, 10 and 100 nM of SP significantly increased the levels of SCF. The ability of SP to affect cytokine levels varied with time. Following 2 hr of SP incubation, cytokine levels were enhanced at the lower end of the concentration range as compared to 48 hr of SP treatment. A 48 hr incubation with SP yielded the highest levels of cytokine at the higher end of the concentration range. Taken together, the results of these studies suggest that SP has an immunoregulatory effect on bone marrow stromal cells leading to alteration in the production and/or secretion of regulatory cytokines such as IL-7 and SCF.