Exploration on the Effect of Bushen Huoxue Prescription on Necroptosis in Human Nucleus Pulposus Cells Based on RIPK1/RIPK3/MLKL Pathway
10.19879/j.cnki.1005-5304.202409495
- VernacularTitle:基于RIPK1/RIPK3/MLKL通路探讨补肾活血方对髓核细胞坏死性凋亡的影响
- Author:
Wei PENG
1
;
Liguo ZHU
;
Xunlu YIN
;
Kexin YANG
;
Jie LUO
;
Minshan FENG
;
Jie YU
;
Linghui LI
;
Jiawen ZHAN
;
Tao HAN
;
Long LIANG
;
Mingyi LUO
;
Dian ZHANG
Author Information
1. 北京中医药大学,北京 100029
- Publication Type:Journal Article
- Keywords:
Bushen Huoxue Prescription;
nucleus pulposus cells;
necroptosis;
intervertebral disc degeneration;
RIPK1/RIPK3/MLKL pathway
- From:
Chinese Journal of Information on Traditional Chinese Medicine
2025;32(3):69-75
- CountryChina
- Language:Chinese
-
Abstract:
Objective To observe the effects of Bushen Huoxue Prescription on pressure-induced necroptosis in human nucleus pulposus cells and the expressions of RIPK1/RIPK3/MLKL pathway;To explore its potential mechanism in delaying intervertebral disc degeneration.Methods Human primary nucleus pulposus cells were cultured in vitro,and a model of nucleus pulposus cell degeneration was established using continuous load pressure method.After modeling,the nucleus pulposus cells were divided into model group,Bushen Huoxue Prescription group and inhibitor group,blank serum,Bushen Huoxue Prescription containing serum and necroptotic apoptosis inhibitor(Nec-1)intervention were administered,respectively.Normal group nucleus pulposus cells were cultured routinely.AO/EB fluorescence dual staining method was used for detecting cell apoptosis,flow cytometry was used to detect the necroptosis rate of nucleus pulposus cells,Western blot was used to detect the protein expressions of p-receptor interacting protein kinase(RIPK)1,p-RIPK3 and p-mixed lineage kinase domain like protein(MLKL),RT-qPCR was used to detect the mRNA expressions of RIPK1,RIPK3 and MLKL.Results Compared with the normal group,the model group showed more red fluorescence under AO/EB staining of nucleus pulposus cells,which were round and condensed,the necroptosis rate of nucleus pulposus cells increased(P<0.05),the protein expressions of p-RIPK1,p-RIPK3 and p-MLKL increased(P<0.05,P<0.01),while there was no significant difference in RIPK1 mRNA expression(P>0.05),and RIPK3 and MLKL mRNA expression increased(P<0.01).Compared with the model group,Bushen Huoxue Prescription group and the inhibitor group had less red condensed chromatin in the nucleus pulposus cells,Bushen Huoxue Prescription group had a lower rate of necroptosis(P<0.05),while the inhibitor group showed a decreasing trend in necroptosis rate(P>0.05),the protein expressions of p-RIPK1,p-RIPK3 and p-MLKL decreased in Bushen Huoxue Prescription group and the inhibitor group(P<0.05,P<0.01),while there was no significant difference in RIPK1 mRNA expression(P>0.05),and RIPK3 and MLKL mRNA expressions decreased(P<0.01).Conclusion Bushen Huoxue Prescription can alleviate pressure-induced damage to nucleus pulposus cells and inhibit necroptosis,thereby slowing the progression of intervertebral disc degeneration.Its mechanism may be related to the RIPK1/RIPK3/MLKL pathway mediated necroptosis.
