8:30 PH and anti-inflammatory agents modulate nucleus pulposus cytokine secretion

Abstract

Purpose of study: Increased levels of interleukin (IL)-6 and IL-8 have been found in intervertebral disc (IVD) tissue from patients undergoing fusion for discogenic low back pain. The stimuli that induce these mediators in degenerate discs remain unknown. Impaired diffusion of nutrients and wastes to and from the nucleus pulposus (NP) is believed to be an important factor in the degenerative process. The oxygen tension and pH in the NP of degenerating discs are significantly decreased. The aims of this study were to 1) demonstrate the ability of porcine NP to respond to a proinflammatory stimulus (lipopolysaccharide [LPS]) in vitro, 2) investigate the effects of pH, pO2 and glucose concentration on NP proinflammatory mediator secretion and 3) determine if methylprednisolone (MP) or indomethacin (IND) can block NP proinflammatory mediator secretion.Methods used: IVDs were harvested from 6-month-old pigs and dissected under sterile conditions in the laboratory. Two hundred milligram samples of NP were cultured under optimal conditions (control), in a 1% O2 environment, at pH 6 and in culture medium without glucose for 72 hours. Blocking experiments were performed by culturing LPS-stimulated samples with either methylprednisolone or indomethacin for 24 hours. IL-6 and IL-8 levels were estimated by ELISA.of findings: See Tables 1 and 2Table 1ControlLPS (5 ug/ml)HypoxiaGlucose- freepH 6IL-6 pg/ml6.2 ± 1.3371 ± 1013.3 ± 42.3 ± 327.5 ± 1*IL-8 pg/ml13.9 ± 1.42068 ± 745†12.4 ± 2.13.7 ± 3.6559 ± 8‡Data=mean ± SD. Statistical analysis was by Students t test. A significant result between control and stimulated groups is indicated by:*p=.024.†p=.0007.‡p=.012.Table 2ControlLPS (5 ug/ml)LPS MPLPS INDIL-6 pg/ml7.5 ± 6.359 ± 662 ± 2.4*0.5 ± 1†IL-8 pg/ml9.1 ± 9.33,212 ± 99810 ± 11.2‡21.5 ± 14.3‡LPS MP = LPS-stimulated tissue incubated with 2 mg/ml MP; LPS IND = LPS-stimulated tissue incubated with 500 uM IND. Data = mean ± SD. Statistical analysis was by Students t test. A significant result between control and stimulated groups is indicated by:*p=.044.†p=.04.‡p=.00004..Relationship between findings and existing knowledge: In recent years evidence has accumulated that biochemical events play an integral role in IVD degeneration and discogenic low back pain. Kang et al. have reported increased levels of nitric oxide, IL-6 and PGE2 in herniated IVDs. Burke et al. have linked IVD production of elevated levels of IL-6 and IL-8 with discogenic low back pain. The stimuli responsible for proinflammatory mediator production in degenerating disc tissue remain unknown. This study has demonstrated that decreased pH stimulates NP proinflammatory mediator secretion at values that have been recorded in vivo. This study confirms the findings of Takahashi et al. that corticosteroids reduce disc proinflammatory mediator production. No previous reports have examined the effects of nonsteroidal anti-inflammatory drugs on disc mediator secretion.Overall significance of findings: Our results confirm that NP is a biochemically active tissue capable of producing proinflammatory mediators in response to environmental stresses. Our findings suggest that decreased intradiscal pH may be an important factor inducing IL-6 and IL-8 secretion in painful degenerative disc disease. This study demonstrates that both methylprednisolone and indomethacin can reduce proinflammatory cytokine production in NP tissue. Intradiscal administration of these agents may represent a novel therapeutic approach to the management of discogenic pain.Disclosures: Device or drug: methylprednisolone. Status: approved. Device or drug: indomethacin. Status: approved. Conflict of interest: No conflicts.