Melatonin reverses tumor necrosis factor-alpha-induced metabolic disturbance of human nucleus pulposus cells via MTNR1B/Gαi2/YAP signaling

Abstract

Background: Intervertebral disc degeneration (IDD), the main cause of low back pain, is closely related to the inflammatory microenvironment in the nucleus pulposus (NP). Tumor necrosis factor-α (TNF-α) plays an important role in inflammation-related metabolic disturbance of NP cells. Melatonin has been proven to regulate the metabolism of NP cells, but whether it can protect NP cells from TNF-α-induced damage is still unclear. Therefore, this study aims to investigate the role and specific mechanism of melatonin on regulating the metabolism of NP cells in the inflammatory microenvironment. Methods: Western blotting, RT-qPCR and immunohistochemistry were used to detect the expression of melatonin membrane receptors (MTNR1A/B) and TNF-α in human NP tissues. In vitro, human primary NP cells were treated with or without vehicle, TNF-α and melatonin. And the metabolic markers were also detected by western blotting and RT-qPCR. The activity of NF-κB signaling and Hippo/YAP signaling were assessed by western blotting and immunofluorescence. Membrane receptors inhibitors, pathway inhibitors, lentiviral infection, plasmids transfection and immunoprecipitation were used to explore the specific mechanism of melatonin. In vivo, the rat IDD model was constructed and melatonin was injected intraperitoneally to evaluate its therapeutical effect on IDD. Results:The upregulation of TNF-α and downregulation of melatonin membrane receptors (MTNR1A/B) were observed in degenerative NP tissues. Then we demonstrated that melatonin could alleviate the development of IDD in a rat model and reverse TNF-α-impaired metabolism of NP cells in vitro. Further investigation revealed that the protective effects of melatonin on NP cells mainly rely on MTNR1B, which subsequently activates Gαi2 protein. The activation of Gαi2 could upregulate the yes-associated protein (YAP) level, resulting in anabolic enhancement of NP cells. In addition, melatonin-mediated YAP upregulation increased the expression of IκBα and suppressed the TNF-αinduced activation of the NF-κB pathway, thereby inhibiting the catabolism of NP cells. Conclusions:Our results revealed that melatonin can reverse TNF-α-impaired metabolism of NP cells via the MTNR1B/Gαi2/YAP axis and suggested that melatonin can be used as a potential therapeutic drug in the treatment of IDD.

Irisin Ameliorates Intervertebral Disc Degeneration by Activating LATS/YAP/CTGF Signaling

“…Ample studies have shown that the Hippo signaling pathway is related closely to mechanical stress and is involved in bone and cartilage degenerative diseases [ 42 , 43 ]. According to the available evidence, the YAP protein has been reported to promote the anabolic metabolism of ECM in NPCs [ 31 , 32 ]. CTGF, as the target molecule of the Hippo signaling pathway, has been reported to be involved in the ECM metabolism of nucleus pulposus cells [ 31 , 32 ].…”

“…According to the available evidence, the YAP protein has been reported to promote the anabolic metabolism of ECM in NPCs [ 31 , 32 ]. CTGF, as the target molecule of the Hippo signaling pathway, has been reported to be involved in the ECM metabolism of nucleus pulposus cells [ 31 , 32 ]. Nevertheless, the mechanisms governing LATS/YAP/CTGF in the regulation of NPCs have remained elusive.…”

“…SYSU-IACUC-2022-B0403). As previously described [ 32 ], briefly, a total of eighteen 12-week-old rats were housed in a vivarium with a 12-hour light/dark cycle. After being anesthetized, the rats were placed on a board, and 21 G needle punctures were performed in two adjacent caudal intervertebral discs at a depth of approximately 5 mm.…”

“…As reported, the key effector, the yes-associated protein (YAP), was downregulated, while the phosphorylated YAP protein was upregulated in degenerative NP tissues [ 30 ]. Recently, it has been reported that YAP mediates the unbalanced ECM metabolism in NPCs and is found to be regulative in the progression of IDD [ 31 , 32 ]. As the target gene of YAP, the connective tissue growth factor (CTGF), also known as CCN2, is a secreted peptide consisting of 349 amino acids, composed of N-terminal signal peptide, intermediate protein-binding domain, and C-terminal-binding domain [ 33 ].…”

Therapeutic effect of chinese herbal medicine gu-ben-hua-shi (AESS) formula on atopic dermatitis through regulation of yes-associated protein

“…The crosstalk between Hippo-YAP and NF-κB signaling pathways also seems complex. Qiu et al (2022) showed that melatonin-mediated YAP upregulation attenuates TNF-α–induced NF-κB pathway activation by enhancing the expression of IκBα in nucleus pulposus cells. YAP also inhibits the activation of NF-κB signaling prompted by TNF-α in murine preosteoblastic cells ( Yang et al, 2020 ) and cementoblast cells ( Zhang et al, 2020 ).…”