Anti-apoptotic effects of IGF-1 and PDGF on human intervertebral disc cells in vitro

Study Design. Human cells from the anulus were grown in tissue culture in a n experimental design to study the anti-apoptotic effect of two selected cy tokines. Objectives. To determine whether two selected cytokines, insulin-like growt h factor-1 and platelet-derived growth factor, were effective in decreasing apoptosis in human cells from the anulus grown in culture for 10 days. Summary of the Background Data. Previous studies have shown that there is a small cell population in the aging human intervertebral disc, Earlier work from the authors' laboratory suggested that apoptosis (programmed cell dea th) may be a major contributing factor to the decrease in cell number. A wi de variety of inhibitors of apoptosis have now been identified; the present report presents findings on the actions of insulin-like growth factor-1 an d platelet-derived growth factor in retarding or preventing apoptosis. Methods. Using previously published culture methods ods, cells from the anu lus of 25 subjects (mean age, 41.7 years) were grown in monolayer culture f or 10 days and tested under the following conditions: 1) control growth in the presence of 20% fetal bovine serum; 2) positive control conditions prom oting the development of apoptosis in the absence of serum; or 3) in dose-r esponse regimes where insulin-like growth factor-1 or platelet-derived grow th factor were added in the presence of only 1% fetal bovine serum (necessa ry for basal cell maintenance). Specimens were derived from 18 lumbar, 9 ce rvical, and 1 thoracic sites; the average Thompson score was ill, Cells wer e grown on chambered slides and evaluated distinguish TdT in situ apoptosis detection identify apoptotic cells. An average of 300 cells were counted i n replicate cultures at each dose to deter mine the incidence of apoptosis; results were analyzed with standard statistical techniques. Cultured cells also were examined with transmission electron microscopy. Results, Serum withdrawal to a 1% level was used as a positive apoptosis co ntrol in vitro and resulted in a significantly greater percentage of apopto sis compared with the 20% serum negative control (1.02% +/- 0.34 (28) versu s 0.14% +/- 0.04 (27; mean +/- SEM (n)), P < 0.0001). Exposure to 50 ng/mL insulin-like growth factor-1 significantly reduced the percentage of apopto sis (vs. 1% serum) to 0.49 +/- 0.26 0.26 (P = 0.005); 500 ng/mL was also si gnificantly effective (% apoptosis = 0.09% +/- 0.04 (P = 0.0001). Platelet- derived growth factor at a dose of 100 ng/mL also significantly reduced apo ptosis (0.18 +/- 0.11, P = 0.0001). Conclusion. Data demonstrate a significant reduction in the percentage of a poptotic disc cells after exposure to 50-500 ng/mL insulin-like growth fact or-1 or exposure to 100 ng/mL platelet-derived growth factor. These finding s expand the understanding of the cell biology of the disc cell and show th at selected cytokines can retard or prevent programmed cell death in vitro. The administration of these cytokines may have future therapeutic potentia l treatment of disc degeneration.