Effect of a nonprotein bioactive agent on the reduction of cyclooxygenase-2 and tumor necrosis factor–α in human intervertebral disc cells in vitro

Laboratory investigation

Tatsuhiro Yoshida M.D., Jin Soo Park M.D., Ph.D., Kimiaki Yokosuka M.D., Ph.D., Kotaro Jimbo M.D., Ph.D., Kei Yamada M.D., Ph.D., Kimiaki Sato M.D., Ph.D., and Kensei Nagata M.D., Ph.D.
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  • Department of Orthopaedic Surgery, Kurume University School of Medicine, Kurume, Fukuoka, Japan
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Object

Neurotropin is a nonprotein extract from the inflamed skin of rabbits inoculated with vaccinia virus. In the present study the authors sought to clarify the focal antiinflammatory effects of Neurotropin in intervertebral disc cells, and these effects were compared with those induced by the selective cyclooxygenase (COX)–2 inhibitor 6-methoxy-2-naphthylacetic acid (nabumetone).

Methods

Six human intervertebral disc specimens were harvested during spinal surgery for lumbar disc herniation. Cells were stimulated with 500 pg/ml of interleukin (IL)–1β in the presence of various concentrations of Neurotropin (0, 10−5, 10−4, and 10−3 Neurotropin Units/ml) or 50 μg/ml of nabumetone for 3 hours. The mRNA was extracted for polymerase chain reaction (PCR), and real-time PCR was used to quantify the mRNA levels of COX- 2, tumor necrosis factor (TNF)–α, and phospholipase A2. Cyclooxygenase-2, TNFα, and prostaglandin E2 (PGE2) protein concentrations were each determined by enzyme-linked immunosorbent assay.

Results

Neurotropin was found to significantly suppress the expression of COX-2 and TNFα at mRNA levels as well as the concentration of COX-2 at protein levels in a dose-dependent manner. Nabumetone was found to significantly increase COX-2 at mRNA levels but directly suppress the concentration of PGE2 in culture medium.

Conclusions

Results in this study suggest that Neurotropin has an analgesic effect through the suppression of COX-2 and TNFα in a focal area, and nabumetone shows this same effect through the suppression of PGE2 production. Thus, Neurotropin could decrease pain by blocking the central pain pathway or increasing focal antiinflammatory effects.

Abbreviations used in this paper:

COX-2 = cyclooxygenase 2; ELISA = enzyme-linked immunosorbent assay; FBS = fetal bovine serum; IDH = intervertebral disc herniation; IL = interleukin; MMP = matrix metalloproteinase; NSAID = nonsteroidal antiinflammatory drug; NU = Neurotropin Unit; PGE2 = prostaglandin E2; PLA2 = phospholipase A2; RT-PCR = real-time polymerase chain reaction; SART = specific alternation of rhythm in environmental temperature; TNFα = tumor necrosis factor–α.

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