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miR-29b-3p Affects the Hypertrophy of Ligamentum Flavum in Lumbar Spinal Stenosis and its Mechanism

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Abstract

To explore the effect of miR-29b-3p on fibrosis and hypertrophy of ligamentum flavum (LF) in lumbar spinal stenosis (LSS) and its underlying mechanism. Patients with LSS and lumbar disc herniation (LDH) (control) undergoing posterior lumbar laminectomy were included in this study. Human LF samples were obtained for LF cell isolation, RNA, and protein extraction. Histomorphological analysis of LF was performed using hematoxylin-eosin (HE) staining. After isolation, culture, and transfection of primary LF cells, different transfection groups were constructed: NC-mimic, miR-29b-3p-mimic, NC-inhibitor, and miR-29b-3p-inhibitor. Quantitative real time polymerase chain reaction (qRT-PCR) was performed to detect the expression of miR-29b-3p in LF and LF cells. Western blot analysis detected the protein expressions of P16 and CyclinD1. ELISA detected the protein expressions of TGF-β1, Smad2, Smad3, TLR4, Type I collagen, and Type III collagen. Finally, LF cell viability was detected using the Cell Counting Kit-8 (CCK8) assay. The thickness of LF was significantly thicker in the LSS group compared to the LDH group (p < 0.05), accompanied by a higher calcification degree, more fibroblasts, and a larger area of collagen fiber proliferation. miR-29b-3p expression was significantly lower in LSS-derived LF tissues and cells than in LDH-derived tissues and cells (both p < 0.05). Compared to the NC-mimic group, the miR-29b-3p-mimic group exhibited significantly higher miR-29b-3p expression, decreased protein expressions of Type I collagen, Type III collagen, TGF-β1, Smad2, Smad3, TLR4, P16, and CyclinD1, and inhibited LF cell proliferation (all p < 0.05). As expected, the miR-29b-3p-inhibitor group displayed contrasting expression patterns (all p < 0.05). Compared to the phosphate buffer saline (PBS) group, the Trimethylamine-N-Oxide (TMAO) group showed significantly increased expressions of TGF-β1, Smad2, Smad3, TLR4, Type I collagen, Type III collagen, P16, and CyclinD1, as well as enhanced LF cell proliferation (all p < 0.05). However, there was no significant difference between the TMAO group and the Ang II group (all p > 0.05). Upregulation of miR-29b-3p expression may play a role in improving LF fibrosis and hypertrophy in LSS by inhibiting P16 expression and suppressing the activation of the TGF-β/Smad signaling pathway. This finding offers new insights into future gene modification therapy for this patient population.

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No datasets were generated or analysed during the current study.

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Acknowledgements

We would like to acknowledge the hard and dedicated work of all the staff that implemented the intervention and evaluation components of the study.

Funding

This work was supported by the Dehong Prefecture science and technology plan project (ZC202318), Tianjin Health Commission science and technology youth project (TJWJ2022QN040) and Tianjin Key Medical Discipline (Specialty) Construction Project (TJYXZDXK-064B).

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Author notes

  1. Hongjie Zhang, Zhixiong Hong and Zehua Jiang contributed equally.

Authors and Affiliations

  1. Department of Orthopedics, Dehong People’s Hospital, Kunming Medical University Affiliated Dehong Hospital, Dehong, No.13 Yonghan Road, Mangshi District, 678400, China

    Hongjie Zhang, Zhixiong Hong & Jiashao Hu

  2. College of Integrative Chinese and Western Medicine, Tianjin University of Traditional Chinese Medicine, No.10, Poyang Lake Road, Jinghai District, Tianjin, 301617, China

    Hongjie Zhang

  3. Department of Spine Surgery, Tianjin Union Medical Center, No.190 Jieyuan Road, Hongqiao District, Tianjin, 300122, China

    Zehua Jiang, Wei Hu & Rusen Zhu

Authors
  1. Hongjie Zhang
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  2. Zhixiong Hong
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  3. Zehua Jiang
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  4. Wei Hu
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  5. Jiashao Hu
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  6. Rusen Zhu
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Contributions

ZHJ, HZX and JZH conceived the idea and conceptualized the study. ZHJ, HW and ZRS collected the data. JZH and HW analyzed the data. ZHJ and ZRS drafted the manuscript, then HJS and HZX reviewed the manuscript. All authors read and approved the final draft.

Corresponding author

Correspondence to Rusen Zhu.

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Ethics Approval and Consent to Participate

This study was conducted with approval from the Ethics Committee of Tianjin People’s Hospital (2016-YXLL-000490). This study was conducted in accordance with the declaration of Helsinki. Written informed consent was obtained from all participants.

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Not applicable.

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The authors declare no competing interests.

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Zhang, H., Hong, Z., Jiang, Z. et al. miR-29b-3p Affects the Hypertrophy of Ligamentum Flavum in Lumbar Spinal Stenosis and its Mechanism. Biochem Genet (2024). https://doi.org/10.1007/s10528-024-10811-8

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  • DOI: https://doi.org/10.1007/s10528-024-10811-8

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