Myristic acid beneficially modulates intervertebral disc degeneration by preventing endplate osteochondral remodeling and vertebral osteoporosis in naturally aged mice

Background: The origin of intervertebral disc degeneration (IDD) is highly complex, where both cartilage endplate remodeling and vertebral osteoporosis are of utmost importance. Myristic acid (MA), a saturated fatty acid derived from nutmeg, a traditional Chinese herb, has been shown to boost memory. Additionally, its isomers have been verified to have antiosteoporotic characteristics. However, the precise mechanism by which MA functions in relation to IDD remains unclear.

Methods: In vivo, a natural aging animal model was used. The drug—administration method of MA was intraperitoneal injection to mice aged 22 months at a dose of 2 mg/kg·d for 2 months. Micro-CT observed vertebral bone mass and endplate changes, followed by Hematoxylin‒eosin (H&E), Masson, and Safranin-O staining of tissues. TRAP staining counted osteoclasts; immunohistochemistry detected the expressions of Aggrecan and Collagen II. Bioinformatics explored MA’s anti-IDD mechanism. In vitro, MA-treated senescent endplate chondrocytes (induced by TBHP) were analyzed by Real-Time PCR (qPCR) and immunofluorescence (IF) for senescence and matrix synthesis markers. TRAP and F-actin detected MA’s effect on RAW264.7 osteoclast differentiation (induced by RANKL); qPCR examined the expressions of osteoclast genes.

Results: Using the natural aging model, we found that MA tended to improve vertebral osteoporosis and endplate osteochondral remodeling, decreased the TRAP activity of the endplate, and alleviated IDD in naturally aging mice. Bioinformatics analysis suggested that the relationships among IDD, osteoporosis, and endplate degeneration were mainly linked to cellular senescence. In vitro, MA postponed the senescence of TBHP-induced endplate chondrocytes by increasing the expression of aggrecan and decreasing the expressions of MMP-3, MMP-9, and the senescence markers p16 and p21. Additionally, MA notably inhibited osteoclast activity, as evidenced by a decrease in the number of osteoclasts and a significant suppression of F-actin formation. At the molecular level, MA efficiently reduced the expressions of osteoclast marker genes like ACP-5, CTSK, and DC-STAMP.

Conclusion: The findings of this research suggest that MA is capable of inhibiting endplate osteochondral remodeling and vertebral osteoporosis, diminishing osteoclastogenesis to preserve bone mass, and consequently delaying IDD in naturally aging mice. Hence, MA holds the potential to serve as an alternative therapeutic approach for IDD.

Methods
This research dataset focuses on the physiological mechanism of naturally aging mice, and the content is as follows: 1. Micro-CT data: It includes all the relevant data of naturally aging mice in the article, such as the microscopic structure of the cartilaginous endplate bone, the fine structural parameters of the vertebral bone, as well as the staining results of multiple methods like immunohistochemistry (IHC), hematoxylin-eosin (HE) staining, safranin O-fast green staining, and tartrate-resistant acid phosphatase (TRAP) staining. These data can reflect the morphological characteristics of the skeletal tissues and the cellular features of the mice. 2. Cell experiment data: Firstly, it includes the data of PCR, immunofluorescence (IF), Cell Counting Kit-8 (CCK8) assay, and flow cytometry identification of cartilaginous endplate cells, which conduct multi-dimensional analysis from aspects such as gene expression, protein expression, cell viability, and cell population characteristics. Secondly, it involves the data of F-actin staining, PCR, and TRAP staining related to the osteoclast differentiation of RAW264.7 cells, which helps to understand the mechanism of osteoclast differentiation. 3. Bioinformatics data: It covers the data related to molecular docking, Gene Ontology (GO) functional enrichment analysis, and Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway. It explores the biological significance behind the data from the molecular level and biological pathways, providing support for revealing the molecular mechanism of aging.