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Adenosine blocks aminopterin-induced suppression of osteoclast differentiation

Junpei Teramachi1, Akiko Kukita, Pengfei Qu

  • 1Department of Molecular Cell Biology and Oral Anatomy, Faculty of Dental Sciences, Kyushu University, 3-1-1 Maidashi, Fukuoka, 812-8582, Japan.

Journal of Bone and Mineral Metabolism
|November 1, 2012
PubMed

Insights

Aminopterin in hybridoma selection medium (HAT) inhibits osteoclastogenesis. Adding adenosine to cultures overcomes this suppression, enabling effective screening of antibodies targeting osteoclast formation.

Area of Science:

  • Immunology
  • Cell Biology
  • Biochemistry

Background:

  • Osteoclastogenesis, the formation of multinucleated osteoclasts, is crucial for bone remodeling.
  • Identifying molecules that regulate osteoclast fusion is key to understanding bone diseases.
  • Monoclonal antibodies are valuable tools for studying cell surface molecules involved in osteoclastogenesis.

Purpose of the Study:

  • To identify cell surface molecules regulating osteoclastogenesis, particularly during cell fusion.
  • To develop a reliable method for screening hybridoma supernatants for osteoclastogenesis-inhibiting monoclonal antibodies.

Main Methods:

  • Whole bone marrow cultures were used to study osteoclastogenesis.
  • The effects of hypoxanthine-aminopterin-thymidine (HAT) selection medium and its components on osteoclast formation were investigated.
  • The role of nucleosides, specifically adenosine, in abrogating the suppressive effects of aminopterin and methotrexate (MTX) was examined.

Main Results:

  • Hybridoma supernatants containing HAT medium significantly suppressed osteoclastogenesis.
  • Aminopterin and methotrexate were identified as the HAT components responsible for inhibiting osteoclastogenesis.
  • The addition of all nucleosides, particularly adenosine, to the culture medium abrogated the suppressive effects of aminopterin and MTX.

Conclusions:

  • Aminopterin and MTX inhibit osteoclastogenesis by interfering with essential cellular pathways.
  • Adenosine can rescue osteoclastogenesis from the suppressive effects of aminopterin and MTX.
  • Direct bio-assay of hybridoma supernatants is feasible for screening osteoclastogenesis-regulating antibodies when using adenosine-supplemented culture media.

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