| Objective: Human nucleus pulposus cells from lumbar disc herniation were isolated and cultured in vitro,and cultured with different concentrations of Bugan Jianyao Decoction containing serum to observe IL-1β、 IL-6、TNF-α、 NF-κB p65 m RNA、NF-κBp65 protein,to explore whether Bugan Jianyao Decoction formula regulates NF-κ B signaling pathway can protect LDH nucleus pulposus cells.Methods: The medicated serum of Bugan Jianyao Decoction and the serum of normal rats were prepared.The human lumbar intervertebral disc was taken,and the nucleus pulposus cells were obtained by sequential digestion method.The nucleus pulposus cells were identified as nucleus pulposus cells by type II collagen immunofluorescence staining,and cultured in a constant temperature and humidity carbon dioxide incubator.After the first generation nucleus pulposus cells were in logarithmic proliferation,group intervention was carried out.The experiment was divided into eight groups: blank group,si RNA blocking group and NF-κ Bp65 si RNA blocking group,5% normal serum group,10% normal serum group,20% normal serum group,5% medicated serum group,10% medicated serum group and 20% medicated serum group.6 parallel holes are set in each group,with 200μl.The culture time was set at 24 h and 48 h respectively,and IL-1β、 IL-6、TNF-α was detected by ELISA,The NF-κ Bp65 m RNA was detected by RT q PCR and NF-κ Bp65 protein were detected by Western blot.Results: In terms of time,48 h compared with 24 h,the blank group,si RNA blocking group comparison,IL-1β、 IL-6、TNF-α、 NF-κ B p65 m RNA 、 NF-κ B p65 protein(P > 0.05);NF-κ B p65 si RNA blocking group,normal serum group and drug containing serum group of Bugan Jianyao Decoction formula for 48 hours IL-1β、 IL-6、TNF-α、NF-κ B p65 m RNA、NF-κ B p65 protein decreased(P < 0.05);In the group,there was no significant change in the content of each index between the blank group and the si RNA blocking group at 24 h and 48h(P > 0.05).Compared with the blank group and the si RNA blocking group,the IL-1 of the other groups at 24 h and 48 h IL-1β、 IL-6、TNF-α、 NF-κ B p65 m RNA、NF-κ B p65 protein was decreased There was no significant change(P < 0.05).The expression of 20%medicated serum group decreased significantly compared with other medicated serum groups.Conclusion: Bugan Jianyao Decoction can regulate NF-κ B p65 signaling pathway inhibits IL-1 β 、 IL-6 、 TNF-α 、 NF-κ B p65 m RNA、NF-κ B p65 and the inflammatory response of nucleus pulposus cells may be one of the mechanisms of Bugan Jianyao formula in the treatment of lumbar disc herniation. |
