age and fissure formation, resulting in structural disorganization. On the molecular level, the metabolism and biosynthetic functions of the cartilaginous endplate cells decrease as the matrix becomes degraded. The activity of matrix metalloproteinases is high in degenerative discs, and the balance between production of tissue inhibitors of metalloproteinase and MMPs appears to be altered. This is accompanied by the induction of collagenases that are known to be involved in disc degeneration. MMP-1 and MMP-13 are of particular importance, as they can cleave intact triple-helical collagen molecules. MMP-13 preferentially cleaves type II collagen. Anderson et al. confirmed that degenerative disc changes are associated with up-regulation of collagenases MMP-1 and MMP-13. In addition, human herniated 23796364 lumbar disc cultures spontaneously produce nitric oxide, a known mediator of proteoglycan synthesis. It has been reported that inflammatory cytokines are involved in the pathogenesis of IVD degeneration. Endothelin-1 has been recognized as one of the most potent vasoconstrictor agents. ET-1 was firstly discovered in aortic endothelial cells, and has since been found to be purchase TG 02 produced by many cell types. Interestingly, ET1 is not only a potent vasoconstrictor, but is also associated with inflammation in degenerative diseases mainly via endothelin receptor 1 ET-1 in Cartilaginous Endplate Degeneration type A. ET-1 causes excessive production of NO, which is generated following increases in inducible nitric oxide synthase levels. In addition, ET-1 promotes MMP-1 and MMP-13 synthesis and activation in osteoarthritis cartilage. As mentioned above, recent research has shown that ET-1 is an inflammatory cytokine involved in cartilage degenerative disease. It is not known if ET-1 is expressed by chondrocytes in human IVD endplates or if it mediates pathologic processes there. Therefore, the aim of this study was to determine if ET-1 is produced in human CEP and if activation or over-expression of ET-1 could alter the synthesis and retention of cartilage matrix molecules, MMPs, or otherwise play an important role in IVD tissue degeneration. Materials and Methods Ethics Statement The Institutional Ethics Committee Board of 22286128 Zhongshan Hospital, Fudan University approved the study protocol and the use of human tissues. All study participants gave written informed consent before enrolment. with phosphate buffered saline, a small piece of each endplate was prepared for hematoxylin and eosin and immunohistochemical staining. A second sample was reserved for western blot assay. The remainder of each sample was minced into pieces,1 mm3 with sterile ophthalmic scissors, and digested with 0.15% collagenase type II in Dulbecco’s Minimum Essential Medium containing 5% fetal calf serum for 12 h at 37uC with shaker agitation. The cell suspension was passed through a 70 mm filter to remove aggregates and was then centrifuged for 10 min at 2000 rpm. The supernatant was discarded; the cells were washed three times with PBS and centrifuged again to obtain a pellet. Finally, the cells were cultured in 8 cm2 Petri dishes in DMEM with 10% FCS and 1% penicillin/streptomycin. Cultures were incubated at 37uC and 95% relative humidity in a 5% CO2 atmosphere. Cells for all experiments were used at the third passage of culture after isolation. Histochemical Staining The morphology of cultured cells was evaluated in H&E. To confirm the deposition of sulfated glycosaminoglycan, cell pr