A thermosensitive, reactive oxygen species-responsive, MR409-encapsulated hydrogel ameliorates disc degeneration in rats by inhibiting the secretory autophagy pathway

Abstract

Lumbar disc degeneration is a common cause of chronic low back pain and an important contributor to various degenerative lumbar spinal disorders. However, currently there is currently no effective therapeutic strategy for treating disc degeneration. The pro-inflammatory cytokine interleukin-1β (IL-1β) mediates disc degeneration by inducing apoptotic death of nucleus pulposus (NP) cells and degradation of the NP extracellular matrix. Here, we confirmed that extracellular secretion of IL-1β via secretory autophagy contributes to disc degeneration, and demonstrate that a thermosensitive reactive oxygen species (ROS)-responsive hydrogel loaded with a synthetic growth hormone-releasing hormone analog (MR409) can protect against needle puncture-induced disc degeneration in rats. Methods: The expression levels of proteins related to secretory autophagy such as tripartite motif-containing 16 (TRIM16) and microtubule-associated protein light chain 3B (LC3B) were examined in human and rat disc tissues by histology and immunofluorescence. The effects of TRIM16 expression level on IL-1β secretion were examined in THP-1 cells transfected with TRIM16 plasmid or siRNA using ELISA, immunofluorescence, and immunoblotting. The in vitro effects of MR409 on IL-1β were examined in THP-1 cells and primary rat NP cells using ELISA, immunofluorescence, immunoblotting, and qRT-PCR. Further, MR409 was subcutaneously administered to aged mice to test its efficacy against disc degeneration using immunofluorescence, X-ray, micro-CT, and histology. To achieve controllable MR409 release for intradiscal use, MR409 was encapsulated in an injectable ROS-responsive thermosensitive hydrogel. Viscosity, rheological properties, release profile, and biocompatibility were evaluated. Thereafter, therapeutic efficacy was assessed in a needle puncture-induced rat model of disc degeneration at 8 and 12 weeks post-operation using X-ray, magnetic resonance (MR) imaging, histological analysis, and immunofluorescence. Results: Secretory autophagy-related proteins TRIM16 and LC3B were robustly upregulated in degenerated discs of both human and rat. Moreover, while upregulation of TRIM16 facilitated, and knockdown of TRIM16 suppressed, secretory autophagy-mediated IL-1β secretion from THP-1 cells under oxidative stress, MR409 inhibited ROS-induced secretory autophagy and IL-1β secretion by THP-1 cells as well as IL-1β-induced pro-inflammatory and pro-catabolic effects in rat NP cells. Daily subcutaneous injection of MR409 inhibited secretory autophagy and ameliorated age-related disc degeneration in mice. The newly developed ROS-responsive MR409-encapsulated hydrogel provided a reliable delivery system for controlled MR409 release, and intradiscal application effectively suppressed secretory autophagy and needle puncture-induced disc degeneration in rats. Conclusion: Secretory autophagy and associated IL-1β secretion contribute to the pathogenesis of disc degener...

Aggressive strategies for regenerating intervertebral discs: stimulus-responsive composite hydrogels from single to multiscale delivery systems

“…173 ROS-responsive capsule hydrogels loaded with MR409 effectively inhibited secretory autophagy and acupuncture-induced exacerbation of IVDD in rats. 174 Notably, glucose oxidase and ferrous glycinate triggered cascade enzyme polymerization reactions leading to fast curing of chondroitin sulfate methacrylate (CSMA) and acrylamide hydrogels. 175 This facilitates rapid healing of bone defects and provides a tunable strength bionic strategy for IVD tissue engineering of bioimplant materials and 3D printed scaffolds.…”

Interaction between N6-methyladenosine and autophagy in the regulation of bone and tissue degeneration

“…Prior studies have revealed the marked impairment of autophagy in degenerative tissues ( Zhong et al, 2022 ). Enhanced autophagy flux ameliorates oxidative stress, alleviates the progression of degenerative alterations, and enhances the cellular regenerative activity ( Yao et al, 2018 ; Zheng et al, 2021 ). Increasing number of studies have indicated that m6A-mediated autophagy was involved in the regulation of IVDD and other degenerative diseases ( Yao et al, 2018 ).…”

Colloidally Stabilized DSPE-PEG-Glucose/Calcium Phosphate Hybrid Nanocomposites for Enhanced Photodynamic Cancer Therapy via Complementary Mitochondrial Ca2+ Overload and Autophagy Inhibition

“…23−25 It is demonstrated that autophagy would capture and degrade dysfunctional organelles to sustain metabolism and homeostasis. 24,26,27 Nevertheless, there is a complex relationship between autophagy and tumor treatment. use of block copolymer/CP hybrid composites has been a promising approach to enhance the stability of CP particles.…”

“…Moreover, there remains an obstacle underlying that cancer could develop protective mechanisms to skip cell apoptosis in PDT. − It is demonstrated that autophagy would capture and degrade dysfunctional organelles to sustain metabolism and homeostasis. ,, Nevertheless, there is a complex relationship between autophagy and tumor treatment. Collective studies have documented that autophagy has currently exhibited an essential role for cancer cell survival, hindering the success of the current treatment paradigms. − Autophagy inhibition has been accepted as an adjuvant therapy in the context of other therapies, , which has paved the way for the potential combined approach for tumor treatment.…”