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Platelet-rich plasma attenuates intervertebral disc degeneration via delivering miR-141-3p-containing exosomes.

ABSTRACT: Oxidative stress mediated apoptotic and pyroptotic cell death contributes to intervertebral disc (IVD) degeneration, and platelet-rich plasma (PRP) exerts protective effects to attenuate IVD degeneration. Hence, the present study aimed to validate this issue and uncover the potential underlying mechanisms. The mice and cellular models for IVD degeneration were established by using puncture method and H₂O₂ exposure, respectively, and we evidenced that NLRP3-mediated cell pyroptosis, apoptosis and inflammatory responses occurred during IVD degeneration progression in vitro and in vivo. Then, the PRP-derived exosomes (PRP-exo) were isolated and purified, and we noticed that both PRP-exo and ROS scavenger (NAC) reversed the detrimental effects of H₂O₂ treatment on the nucleus pulposus (NP) cells. Further results supported that PRP-exo exerted its protective effects on H₂O₂ treated NP cells by modulating the Keap1-Nrf2 pathway. Mechanistically, PRP-exo downregulated Keap1, resulting in the release of Nrf2 from the Keap1-Nrf2 complex, which further translocated from cytoplasm to nucleus to achieve its anti-oxidant biological functions, and H₂O₂ treated NP cells with Nrf2-deficiency did not respond to PRP-exo treatment. In addition, miR-141-3p was enriched in PRP-exo, and miR-141-3p targeted the 3' untranslated region (3'UTR) of Keap1 mRNA for its degradation, leading to Nrf2 translocation. Furthermore, overexpression of miR-141-3p ameliorated the cytotoxic effects of H₂O₂ on NP cells, which were abrogated by upregulating Keap1 and silencing Nrf2. Taken together, we concluded that PRP secreted exosomal miR-141-3p to activate the Keap1-Nrf2 pathway, which helped to slow down IVD degeneration.

SUBMITTER: Xu J

PROVIDER: S-EPMC8354670 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Background contextNonphysiological mechanical loading and inflammation are both critically involved in intervertebral disc (IVD) degeneration, which is characterized by an increase in cytokines and matrix metalloproteases (MMPs) in the nucleus pulposus (NP). This process is known to be mediated by the NF-κB pathway.Clinical significanceCurrent clinical treatments for IVD degeneration focus on the alleviation of symptoms rather than targeting the underlying mechanism. Injection of an NF-κB inhibitor may attenuate the progression of IVD degeneration.PurposeTo investigate the ability of the NF-κB inhibitor, NEMO binding domain peptide (NBD), to alter IVD degeneration processes by reducing IL-1β- and mechanically-induced cytokine and MMP levels in human nucleus pulposus cells in vitro, and by attenuating IVD degeneration in an in vivo rat model for disc degeneration.Study designExperimental in vitro and animal model.Patient sampleDiscarded specimens of lumbar disc from 21 patients, and 12 Sprague Dawley rats.Outcome measuresGene and protein expression, cell viability, µMRI and histology.MethodsIL-1β-prestimulated human nucleus pulposus cells embedded into fibrin constructs were loaded in the Flexcell FX-5000 compression system at 5 kPa and 1 Hz for 48 hours in the presence and absence of NBD. Unloaded hNPC/fibrin constructs served as controls. Cell viability in loaded and unloaded constructs was quantified, and gene and protein expression levels determined. For in vivo testing, a rat needle disc puncture model was employed. Experimental groups included injured discs with and without NBD injection and uninjured controls. Levels of disc degeneration were determined via µMRI, qPCR and histology. Funding sources include $48,874 NASS Young Investigator Research Grant and $119,174 NIH 5K01AR071512-02. There were no applicable financial relationships or conflicts of interest.ResultsMechanical compression of hNPC/fibrin constructs resulted in upregulation of MMP-3 and IL-8. Supplementation of media with 10 μM NBD during loading increased cell viability, and decreased MMP-3 gene and protein levels. IVD injury in rat resulted in an increase in MMP-3, IL-1β and IL-6 gene expression. Injections of 250 µg of NBD during disc injury resulted in decreased IL-6 gene expression. µMRI analysis demonstrated a reduction of disc hydration in response to disc needle injury, which was attenuated in NBD-treated IVDs. Histological evaluation showed NP and AF lesion in injured discs, which was attenuated by NBD injection.ConclusionsThe results of this study show NBD peptide's capacity to reduce IL-1β- and loading-induced MMP-3 levels in hNPC/fibrin constructs while increasing the cells' viability, and to attenuate IVD degeneration in rat, involving downregulation of IL-6. Therefore, NBD may be a potential therapeutic agent to treat IVD degeneration.

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Platelet-rich plasma attenuates intervertebral disc degeneration via delivering miR-141-3p-containing exosomes.

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