P12. Comparisons of patterns of upregulation of inflammatory cytokines in herniated nucleus pulposus, disc and nerve root lavagates and in the serum of patients with acute sciatica secondary to lumbar disc herniation undergoing surgery

BACKGROUND CONTEXT The pathogenesis of acute sciatica secondary to lumbosacral spine nucleus pulposus extrusion (AS/NPE) is mediated through an acute inflammatory response. Thus, AS/NPE patients are of particular interest in the study of the complex interaction of cytokines that regulate the severity of inflammation. Marked upregulation of TNFα and some interleukins (IL) has been demonstrated in the serum of AS/LPE patients. TNFα and IL-1β also activate extracellular matrix metalloproteinases and therefore also play an important role in enhancing the degradation/resorption of acute herniated disc tissue. IL-8 is known to be a major initiator of the acute inflammatory response by specifically activating neutrophils and RANTES (Regulation on Activation Normal T Cell Expressed and Secreted) is an inflammatory cytokine that has been correlated to intensity of disc-related pain. To date, research studies correlating patient clinical parameters to elevated cytokine levels have relied upon measurements in serum or percutaneously-obtained disc lavagate and/or nerve root lavagate. Whether or not these methods of measuring cytokine levels, particularly in serum, provide an accurate representation of the true levels and patterns of the up-regulated cytokine response in extruded disc in vivo, has yet to be determined. PURPOSE To compare the patterns of up-regulation of key inflammatory cytokines in extruded disc tissue with that identified in the serum, disc lavagate (DL) and nerve root lavagate (NRL) in patients with AS/NPE undergoing surgery. STUDY DESIGN/SETTING ELISA cytokine immunoassay. PATIENT SAMPLE Inclusion: AS/HNP of less than 16 weeks duration undergoing laminectomy discectomy surgery; spinal nerve root motor score of >3/5; McCulloch score of 5/5 Exclusion: Red flag condition; prior lumbar surgery, other active inflammatory diseases. OUTCOME MEASURES TNFα, IL-1β, 2, 6, 8, 9, 10 and RANTES patterns of upregulation and concentrations (pg/ml) in herniated nucleus pulposus (HNP), DL, NRL and in serum. METHODS At the time of surgery, 5cc saline lavagates of the spinal nerve root (NRL) and at the site of the annular disruption (DL), the extruded disc tissue removed surgically and a venous blood sample, were obtained for study. Blood samples were centrifuged in a standardized manner and the serum collected. The serum and HNP, DL, NRT samples (placed in physiological buffer with protease inhibitors) were stored at -80°C. The concentrations of TNFα, IL-2, 6, 8, 10 and RANTES in all serum and tissue samples were determined using ELISA immunoassay. Research ethics approval was obtained. RESULTS Twenty-four patients had laminectomy discectomy surgery carried out by eight different spine surgeons. In extruded disc tissue, 22/24 (92%) of the patients studied,cytokine concentrations showed a consistent pattern of upregulation with highest concentrations of RANTES (>500 pg/ml), mid-range concentrations of TNFα, IL-1β,IL-6 and IL-8 (100-200 pg/ml) and lower concentrations of IL-2, 9, 10 (< 50 pg/ml). Disc lavagate cytokines levels showed a consistently similar pattern of cytokine elevation. Serum analyses showed consistently high RANTES, TNFα, and IL concentrations in 20/24 patients. In 20/24 patients (83%), serum TNFα, IL-1β and IL-6 and 8 concentrations were clearly elevated, but to a lesser degree (40- 100 pg/ml). IL-2, 9, 10 were detected at trace levels. The nerve root lavage cytokine concentrations and patterns of up-regulation were the least consistent and in 8/24 patients (33%), significant variations in NRL cytokine concentrations were seen. CONCLUSIONS In patients with acute sciatica secondary to lumbar disc herniation, serum and intraoperative disc lavagate cytokine content patterns of upregulation were similar to those identified in extruded disc tissue.The cytokine concentrations in nerve root lavagate samples showed significant variation and correlations with clinical parameters in patients with AS/NPE should be interpreted with caution. Funded by a research grant from the National Institutes of Health, Bethesda USA. FDA DEVICE/DRUG STATUS This abstract does not discuss or include any applicable devices or drugs. The pathogenesis of acute sciatica secondary to lumbosacral spine nucleus pulposus extrusion (AS/NPE) is mediated through an acute inflammatory response. Thus, AS/NPE patients are of particular interest in the study of the complex interaction of cytokines that regulate the severity of inflammation. Marked upregulation of TNFα and some interleukins (IL) has been demonstrated in the serum of AS/LPE patients. TNFα and IL-1β also activate extracellular matrix metalloproteinases and therefore also play an important role in enhancing the degradation/resorption of acute herniated disc tissue. IL-8 is known to be a major initiator of the acute inflammatory response by specifically activating neutrophils and RANTES (Regulation on Activation Normal T Cell Expressed and Secreted) is an inflammatory cytokine that has been correlated to intensity of disc-related pain. To date, research studies correlating patient clinical parameters to elevated cytokine levels have relied upon measurements in serum or percutaneously-obtained disc lavagate and/or nerve root lavagate. Whether or not these methods of measuring cytokine levels, particularly in serum, provide an accurate representation of the true levels and patterns of the up-regulated cytokine response in extruded disc in vivo, has yet to be determined. To compare the patterns of up-regulation of key inflammatory cytokines in extruded disc tissue with that identified in the serum, disc lavagate (DL) and nerve root lavagate (NRL) in patients with AS/NPE undergoing surgery. ELISA cytokine immunoassay. Inclusion: AS/HNP of less than 16 weeks duration undergoing laminectomy discectomy surgery; spinal nerve root motor score of >3/5; McCulloch score of 5/5 Exclusion: Red flag condition; prior lumbar surgery, other active inflammatory diseases. TNFα, IL-1β, 2, 6, 8, 9, 10 and RANTES patterns of upregulation and concentrations (pg/ml) in herniated nucleus pulposus (HNP), DL, NRL and in serum. At the time of surgery, 5cc saline lavagates of the spinal nerve root (NRL) and at the site of the annular disruption (DL), the extruded disc tissue removed surgically and a venous blood sample, were obtained for study. Blood samples were centrifuged in a standardized manner and the serum collected. The serum and HNP, DL, NRT samples (placed in physiological buffer with protease inhibitors) were stored at -80°C. The concentrations of TNFα, IL-2, 6, 8, 10 and RANTES in all serum and tissue samples were determined using ELISA immunoassay. Research ethics approval was obtained. Twenty-four patients had laminectomy discectomy surgery carried out by eight different spine surgeons. In extruded disc tissue, 22/24 (92%) of the patients studied,cytokine concentrations showed a consistent pattern of upregulation with highest concentrations of RANTES (>500 pg/ml), mid-range concentrations of TNFα, IL-1β,IL-6 and IL-8 (100-200 pg/ml) and lower concentrations of IL-2, 9, 10 (< 50 pg/ml). Disc lavagate cytokines levels showed a consistently similar pattern of cytokine elevation. Serum analyses showed consistently high RANTES, TNFα, and IL concentrations in 20/24 patients. In 20/24 patients (83%), serum TNFα, IL-1β and IL-6 and 8 concentrations were clearly elevated, but to a lesser degree (40- 100 pg/ml). IL-2, 9, 10 were detected at trace levels. The nerve root lavage cytokine concentrations and patterns of up-regulation were the least consistent and in 8/24 patients (33%), significant variations in NRL cytokine concentrations were seen. In patients with acute sciatica secondary to lumbar disc herniation, serum and intraoperative disc lavagate cytokine content patterns of upregulation were similar to those identified in extruded disc tissue.The cytokine concentrations in nerve root lavagate samples showed significant variation and correlations with clinical parameters in patients with AS/NPE should be interpreted with caution. Funded by a research grant from the National Institutes of Health, Bethesda USA.