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Toxicity Effects of Methylene Blue on Rat Intervertebral Disc Annulus Fibrosus Cells

Liang Zhang1, Yang Liu2, Zenan Huang1

  • 1Clinical College of Yangzhou University, China.

Pain Physician
|March 30, 2019
PubMed
Abstract

Insights

Methylene blue (MB) can inhibit annulus fibrosus cell proliferation and induce apoptosis in a concentration-dependent manner. These findings suggest careful consideration of MB concentration in clinical applications for discogenic low back pain.

Area of Science:

  • Biomedical Science
  • Cell Biology
  • Orthopedics

Background:

  • Methylene blue (MB) is increasingly used in treatments for discogenic low back pain (LBP) and percutaneous transforaminal endoscopic discectomy (PTED).
  • While MB is known to cause DNA damage and apoptosis in various cells, its specific effects on intervertebral disc (IVD) annulus fibrosus (AF) cells remain unclear.

Purpose of the Study:

  • To investigate the in vitro effects of varying concentrations of methylene blue (MB) on rat AF cells.
  • To assess the cytotoxic impact and cellular responses of AF cells exposed to MB.

Main Methods:

  • AF cells were cultured with different MB concentrations (0, 2, 20, 200 μg/mL).
  • Cell viability was assessed using CCK-8 assays.
  • Apoptosis was evaluated via microscopy and flow cytometry.
  • Gene and protein expression of apoptosis markers (caspase-3, Bcl-2, Bax) and functional genes (collagen type I, TGF-β1, bFGF, TIMP-1) were analyzed using RT-PCR and Western blotting.

Main Results:

  • MB reduced AF cell viability and proliferation in a concentration- and time-dependent manner.
  • MB induced significant AF cell apoptosis, evidenced by morphological changes, flow cytometry, and increased caspase-3 expression.
  • MB upregulated Bax and caspase-3, downregulated Bcl-2, and decreased collagen type I, TGF-β1, bFGF, and TIMP-1 expression in a concentration-dependent manner.

Conclusions:

  • High concentrations of MB inhibit AF cell proliferation and paracrine function while inducing apoptosis.
  • These findings highlight the necessity of using low MB concentrations in clinical practice.
  • The use of MB for discogenic LBP should be limited to research protocols until further validation.

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