Dynamic Mechanical Loading Mediates Human Nucleus Pulposus Cell Proliferation And Matrix Metabolism Via Activation Of NF-?B Pathway

Background:Low back pain(LBP)is a common symptom in orthopedic clinics.Static mechanical loading may be an inducer of disc degeneration.However,static mechanical stress cannot better explain the effects of mechanical loading on people's daily life(walking,lifting,moving,etc).Meanwhile,a large number of studies have found that nuclear factor-kappa B(NF-kB)can promote inflammation,induce apoptosis,activate tissue aging,and accelerate matrix degradation in human body.These effects can cause the onset and exacerbation of LBP.Objective:1.To explore the effects of dynamic mechanical loading on the proliferation of human nucleus pulposus cells.2.To explore the effects of dynamic mechanical loading on matrix metabolism.3.To explore the relationship between dynamic mechanical loading pattern and IDD via NF-kB pathway.Methods:A dynamic mechanical loading model of human nucleus pulposus cells(NP)in vitro was established.According to different strength,time and pattern in dynamic mechanical loading,three groups were established:control group(no dynamic mechanical loading),load group 0.1 MPa(20 h)+0.8 MPa(4 h),and overload group 0.1 Mpa(4 h)+0.8 MPa(20 h).CCK-8 cell proliferation assay,flow cytometry,WesternenBlot,PCR,RT-PCR,ELISA,and cellular immunohistochemical staining were used to evaluate cell viability,apoptosis rate,the relative expression of apoptosis factors Bax/Bcl-2,Matrix degrading-enzyme familys(MMPs+ADAMTS),collagen-II fiber synthesis and degradation,aggrecan as well as the NF-kB system(p-IKba,IKba,p-p65,p65).Results:1.The human nucleus pulposus cell viability was declined in the overloading group while it showed a high level in cells culured in the loading pressure group.In the overloading group,the level of Bax protein expression was upregulated,while that of Bcl-2 protein was downregulated.In addition,proper Dynamic loading does not alter the level of Bax and Bcl-2 proteins expression.The ratio of apoptosis in the excessive pressure group was higher than that in the control group and the pressure group.2.The levels of matrix degrading-enzymes familys(MMPs+ADAMTS)expression were elevated in the overloading group,while the levels of collagen-II fibers and aggrecans expression were declined.The level of matrix-degrading enzymes(MMPs+ADAMTS)was declined in the loading group,while that of collagen-II fibers and aggrecan was elevated.3.The level of NF-kB pathway activators expresion in the overload group was ascensused,meanwhile that of collagen-II degeneration observed by biochemistry was declined when the NF-kB cell pathway was blocked in overloading group on the fifth day,indicating that the NF-kB cell pathway induced collagen-II degeneration via an excessive dynamic mechanical loading.Conclusion:Excessive dynamic mechanical loading group(long-hours high stress compression+short-hours stress remittence)activated NF-kB pathway,leading proliferation declination and apoptosis of human nucleus pulposus cells as well as elevation in level of matrix-degrading enzymes(ADAMTS-4,ADAMTS-5,MMP-3,MMP-13)expression and degradation of collagen fibers and aggrecan in the extracellular matrix,promoting IDD;conversely,reasonable dynamic mechanical loading(short-hours high stress compression+long-hours stress remittence)would be beneficial to human nucleus pulposus cells proliferation and the physiological metabolic activities of extracellular matrix.Therefore,the lifestyle as reasonable work hours with relaxation have certain protective fuction to human intervertebral disc.The development of targeted drugs that block NF-kB on the disc may disturb IDD.