Periodic mechanical stress induces the extracellular matrix expression and migration of rat nucleus pulposus cells by upregulating the expression of intergrin α1 and phosphorylation of downstream phospholipase Cγ1

Abstract

Intervertebral disk degeneration (IDD) is a major cause of low back pain and an important socioeconomic burden. Degradation of the extracellular matrix (ECM) of nucleus pulposus (NP) cells in the interverterbal disk is important for IDD. Stress of a suitable frequency and amplitude promotes the synthesis of the ECM of NP cells, however, the associated mechanisms remain to be fully elucidated The present study aimed to investigate the effect of integrin α1 on the migration and ECM synthesis of NP cells under soft periodic mechanical stress. Rat NP cells were isolated and plated onto slides, and were then treated with or without the use of a periodic mechanical stress system. The expression levels of integrin α1, α5 and αv, ECM collagen 2A1 (Col2A1) and aggrecan, and the phosphorylation of phospholipase C-γ1 (PLCγ1) were measured using reverse transcription-quantitative polymerase chain reaction and western blot analyses. Cell migration was assayed using a scratch experiment. The results showed that exposure to periodic mechanical stress significantly induced the mRNA expression levels of Col2A1 and aggrecan, cell migration, mRNA expression of integrin α1 and phosphorylation of PLC-γ1 of the NP, compared with the control (P<0.05). Inhibition of the PLCγ1 protein by U73122 significantly decreased the ECM expression under periodic mechanical stress (P<0.05). Small interfering RNA-mediated integrin α1 gene knockdown suppressed the mRNA expression levels of Col2A1 and aggrecan, and suppressed the migration and phosphorylation of PLCγ1 of the NP cells under periodic mechanical stress, compared with the control (P<0.05). In conclusion, periodic mechanical stress induced ECM expression and the migration of NP cells via upregulating the expression of integrin α1 and the phosphorylation of downstream PLCγ1. These findings provide novel information to aid the understanding of the pathogenesis and development of IDD.

Identification of Core Genes and Screening of Potential Targets in Intervertebral Disc Degeneration Using Integrated Bioinformatics Analysis

“…Among the five KEGG pathways, the chemokine pathway was identified as being strongly associated with the development of IDD. Previous studies have confirmed that multiple molecules involved in these pathways were associated with pathological changes in that intervertebral disc tissue ( Gao et al, 2016 ; Wang J. et al, 2019 ; Liu et al, 2020 ). Our data mining results further confirm that the chemokine pathway plays a crucial role in the development of IDD.…”

Flow Cytometry Has a Significant Impact on the Cellular Metabolome

“…Sorting decreased the abundance of the DG family members DG(44:12) (depleted in Ab+Sort), DG(36:5) (FC −1.52), DG(36:2) (FC −1.64), DG(38:5) (FC −1.59), DG(40:4) (FC −1.40), DG(42:8) (FC −1.60), DG(36:8) (FC −0.95), DG­(P-32:1) (depleted in Ab+Sort), DG(34:2) (FC −1.58), DG(36:4) (depleted in Ab+Sort), DG(38:7) (FC −1.39), DG(38:4) (FC −1.54), DG(40:7) (FC −1.48), DG(42:10) (FC −1.32), DG(40:3) (FC −1.18), and increased species DG(32:3) (appeared in Ab+Sort, when absent in Ctrl), DG(34:5) (FC 1.09 in Sort), altogether possibly reflecting PLC and DG lipase activity. PLC activity was previously found to depend upon periodic mechanical stress, electrostatic potential, and elastic stress of lipid membranes. − The observed increase in DG consumption in FACS sorted macrophages might be associated with PLC and DG lipase activities, which are crucial in the biosynthesis of 2-arachidonoylglycerol (2-AG), , the AA precursor in the endocannabinoid signaling pathway . The outcome is an inflammatory-like status featuring increased energy consumption and cell damage, potentially compromising cell homeostasis.…”

“…in Sort), altogether possibly reflecting PLC and DG lipase activity. PLC activity was previously found to depend upon periodic mechanical stress, electrostatic potential, and elastic stress of lipid membranes [39][40][41] . The observed increase in DG consumption in FACS sorted macrophages might be associated with PLC and DG lipase activities, which are crucial in the biosynthesis of 2-arachidonoylglycerol (2-AG) 42,43 , the AA precursor in the endocannabinoid signalling pathway 43 .…”

Bioengineering the ameloblastoma tumour to study its effect on bone nodule formation

“…Introduction of an AM-3 tumour mass downregulated 30 genes and upregulated 1 gene in the osteoblasts. ALPL and bone development COL2A1 58 , were under-expressed by 5.5-fold (p = 0.003) and 4.2-fold (p = 0.01) respectively. A differentiation factor GDF10 was under-expressed by 17.8-fold (p = 0.0007), which indicated strong inhibition of osteoblast differentiation by AM-3 cells.…”