The Roles of MAPKs in Rabbit Nucleus Pulposus Cell Apoptosis Induced by High Osmolality

Abstract

Introduction Earlier work has suggested that the p38 MAPK, JNK1/2, and ERK1/2 signal pathway existed in nucleus pulposus cells and the cell growth, differentiation, and apoptosis were regulated by them. Because osmotic fluctuations are inevitable in the physicochemical environment of intervertebral disc cells, high osmolality could activate p38 MAPK, JNK1/2, and ERK1/2 signal pathway. The effects of high osmolality on the catabolic program and proliferation of nucleus pulposus cells are still not clear. Materials and Methods Rabbit nucleus pulposus cells were cultured and divided into different group at random. The cells were pretreated with inhibitor for p38 MAPK, JNK1/2, and ERK1/2 signal pathway, respectively. In next step, the cells were cultured in different osmolality environment for different time at 37°C in 5% carbon dioxide incubator. After treatments, ratio of apoptosis was measured by flow cytometry, and Western blotting was performed to quantify the expression of the activated forms of p38 MAPK, JNK1/2, and ERK1/2. Furthermore, immunofluorescence analysis with confocal microscopy was performed to confirm the hyperosmolality effects on activation of p38 MAPK, JNK1/2, and ERK1/2 signal pathways in nucleus pulposus cells. Results Our results show that in 500 and 600 mOsm/kg medium, rabbit nucleus pulposus cell apoptosis increased, and a persistent phosphorylation of p38 MAPK, JNK1/2, and ERK1/2 proteins were observed. In the same condition, the apoptotic cells death remarkably decreased when the p38 MAPK and JNK1/2 signal pathways were blocked by their inhibitors SB203580 and SP600125, respectively. On the other side, the apoptotic cells death rate reraised greatly when the ERK1/2 signal pathways were blocked by its inhibitor PD98059. Conclusion High osmolality activated p38MAPK, JNK1/2, and ERK1/2 in rabbit nucleus pulposus cell, and the activated p38 MAPK and JNK1/2 induced cell apoptosis, on the contrary, the activated ERK1/2 made the cell survived. Disclosure of Interest None declared References Freemont AJ. The cellular pathobiology of the degenerate intervertebral disc and discogenic back pain. Rheumatology (Oxford) 2009;48(1):5–10 Urban JP, McMullin JF. Swelling pressure of the inervertebral disc: influence of proteoglycan and collagen contents. Biorheology 1985;22(2):145–157 Wilke HJ, Neef P, Caimi M, Hoogland T, Claes LE. New in vivo measurements of pressures in the intervertebral disc in daily life. Spine 1999;24(8):755–762 Takeno K, Kobayashi S, Negoro K, et al. Physical limitations to tissue engineering of intervertebral disc cells: effect of extracellular osmotic change on glycosaminoglycan production and cell metabolism. Laboratory investigation. J Neurosurg Spine 2007;7(6):637–644 Boyd LM, Richardson WJ, Chen J, Kraus VB, Tewari A, Setton LA. Osmolarity regulates gene expression in intervertebral disc cells determined by gene array and real-time quantitative RT-PCR. Ann Biomed Eng 2005;33(8):1071–1077 Wuertz K, Urban JP, Klasen J, et al. Influence of extracellular osmolarity and mec...

The protein tyrosine kinase inhibitor, Genistein, delays intervertebral disc degeneration in rats by inhibiting the p38 pathway-mediated inflammatory response

“…Some researchers believe that the p38 MAPK pathway increases the production of inflammatory cytokines and promotes the degradation of intervertebral protein polysaccharides, which leads to the degeneration of intervertebral discs [10]. Other researchers believe that the p38 MAPK pathway leads to disc degeneration by inducing excessive cell apoptosis [11]. Activation of the p38 MAPK pathway can also induce macrophages to secrete inflammatory cytokines (e.g., TNF-α, IL-1, and IL-6) [12].…”