2009   Volume No 17 – pages 15-22

Title: Low-intensity pulsed ultrasound stimulates cell proliferation, proteoglycan synthesis and expression of growth factor-related genes in human nucleus pulposus cell line

Author: Y Kobayashi, D Sakai, T Iwashina, S Iwabuchi, J Mochida

Address: Department of Orthopaedic Surgery, Surgical Science, and Research Center for Regenerative Medicine, Tokai University School of Medicine, 143 Shimokasuya, Isehara, Kanagawa, 259-1193, Japan

E-mail: daisakai at is.icc.u-tokai.ac.jp

Key Words: LIPUS, ultrasound, intervertebral disc, growth factor.

Publication date: June 30th 2009

Abstract: Low-intensity pulsed ultrasound (LIPUS) stimulation has been shown to effect differentiation and activation of human chondrocytes. A study involving stimulation of rabbit disc cells with LIPUS revealed upregulation of cell proliferation and proteoglycan (PG) synthesis. However, the effect of LIPUS on human nucleus pulposus cells has not been investigated. In the present study, therefore, we investigated whether LIPUS stimulation of a human nucleus pulposus cell line (HNPSV-1) exerted a positive effect on cellular activity. HNPSV-1 cells were encapsulated in 1.2% sodium alginate solution at 1x105 cells/ml and cultured at 10 beads/well in 6-well plates. The cells were stimulated for 20 min each day using a LIPUS generator, and the effects of LIPUS were evaluated by measuring DNA and PG synthesis. Furthermore, mRNA expression was analyzed by cDNA microarray using total RNA extracted from the cultured cells. Our study revealed no significant difference in cell proliferation between the control and the ultrasound treated groups. However, PG production was significantly upregulated in HNPSV cells stimulated at intensities of 15, 30, 60, and 120 mW/cm2 compared with the control. The results of cDNA array showed that LIPUS significantly stimulated the gene expression of growth factors and their receptors (BMP2, FGF7, TGFbetaR1 EGFRF1, VEGF). These findings suggest that LIPUS stimulation upregulates PG production in human nucleus pulposus cells by the enhancement of several matrix-related genes including growth factor-related genes. Safe and non-invasive stimulation using LIPUS may be a useful treatment for delaying the progression of disc degeneration.

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