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Pathophysiological Role of Endothelin in Ectopic Ossification of Human Spinal Ligaments Induced by Mechanical Stress

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Abstract

Ossification of the posterior longitudinal ligament (OPLL) of the spine is characterized by progressive ectopic bone formation in the spinal ligament. To identify the genes related to ossification affected by mechanical stress during OPLL, analyses using cDNA microarray were carried out using cultured human spinal ligament cells that had been subjected to uniaxial cyclic stretching. Samples were obtained from a total of 14 patients: seven cervical or thoracic OPLL patients and seven control patients. Spinal ligament cells derived from tissues of OPLL (OPLL cells) and control (non-OPLL cells) patients were subjected to uniaxial sinusoidal cyclic stretching (0.5 Hz, 20% stretch) for various time periods (0–9 hours). cDNA microarrays revealed that ranges of distribution of both up- and downregulated genes evoked by cyclic stretching were significantly wider in OPLL cells than in non-OPLL cells. Increases in the mRNA expression of endothelin-1 (ET-1) as well as various marker genes related to ossification were also observed. mRNA expression of ET-1 and alkaline phosphatase was increased by mechanical stress in a time-dependent manner, while addition of ET-1 to static cultures of OPLL cells increased mRNA expression of alkaline phosphatase in a dose-dependent manner. During 9 hours of cyclic stretching, ET-1 release increased to about sixfold the amount observed in nonstretched cells. In non-OPLL cells, neither cyclic stretching nor ET-1 induced any increase in alkaline phosphatase expression. These results suggest that mechanical stress promotes the progression of ossification in OPLL cells through autocrine and/or paracrine mechanisms of ET-1.

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Acknowledgements

We thank Drs. Masahiko Tanno and Hirotaka Ohishi of the Department of Orthopedic Surgery, Hirosaki University School of Medicine, for their technical assistance and valuable discussions. We also acknowledge Professors Hiroto Kimura and Hideki Mizunuma and Associate Professor Shinji Nishikawa (Hirosaki University School of Medicine) for their valuable suggestions. This work was supported in part by a grant-in-aid from the Ministry of Education, Science, Sports, and Culture of Japan and from the Ministry of Health, Labor, and Welfare. Also, we thank the Karohji-Memorial Aid of Medical Study and Hirosaki University Educational Improvement Promotional Aid for financial support.

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Authors and Affiliations

  1. Department of Orthopedic Surgery, Hirosaki University School of Medicine, 5 Zaifu-cho, Hirosaki, 036-8562, Japan

    T. Iwasawa, A. Okada & S. Toh

  2. Department of Pharmacology, Hirosaki University School of Medicine, 5 Zaifu-cho, Hirosaki, 036-8562, Japan

    K. Iwasaki, T. Sawada, S. Motomura & K.-I. Furukawa

  3. Department of Orthopedic Surgery, Hirosaki Memorial Hospital, 59-1 Nishida Sakaizeki, Hirosaki, 036-8076, Japan

    K. Ueyama

  4. Department of Orthopedic Surgery, Aomori Central Hospital, 2-1-1 Higashitsukurimichi, Aomori, 036-8553, Japan

    S. Harata

  5. Division of Genetic Diagnosis, Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo, 108-8639, Japan

    I. Inoue

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  1. T. Iwasawa
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  2. K. Iwasaki
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  3. T. Sawada
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  4. A. Okada
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  5. K. Ueyama
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  7. S. Harata
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  8. I. Inoue
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  9. S. Toh
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  10. K.-I. Furukawa
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Correspondence to K.-I. Furukawa.

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Iwasawa, T., Iwasaki, K., Sawada, T. et al. Pathophysiological Role of Endothelin in Ectopic Ossification of Human Spinal Ligaments Induced by Mechanical Stress. Calcif Tissue Int 79, 422–430 (2006). https://doi.org/10.1007/s00223-006-0147-7

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  • DOI: https://doi.org/10.1007/s00223-006-0147-7

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