2002 CTOS Annual Meeting Oral Presentations — Biology

ANTISENSE INHIBITION OF HYALURONAN SYNTHASE-2 IN HUMAN OSTEOSARCOMA CELL LINE, MG-63, INHIBITS HYALURONAN RETENTION AND TUMORIGENICITY OF THE CELLS
[Abstract ID: 35]

Category: Biology

Presentation: Oral

Authors: Yoshihiro Nishida¹, Warren Knudson², Cheryl B Knudson², Izuru Tabata¹, Naoki Ishiguro¹

Author Institutions: ¹Department of Orthopaedic Surgery Nagoya University School of Medicine, Japan; ²Department of Biochemistry Rush Medical College, IL, United States

Presenter: Yoshihiro Nishida
ynishida@med.nagoya-u.ac.jp

Correspondent: Yoshihiro Nishida
ynishida@med.nagoya-u.ac.jp
Nagoya Japan 466-8550
Ph: +81-52-741-2111
Fax: +81-52-744-2260

Objectives: Osteosarcoma is the common primary malignant tumor of bone and is represented by a heterogenous group of lesions with diverse histopathology and clinical behavior. Hyaluronan (HA) has been thought to play significant roles in tumor progression. The aim of this study was to determine the differential pattern of expression of three hyaluronan synthase genes (HAS), that are responsible for the synthesis of HA in osteosarcomas. An additional goal was to analyze the effects of the inhibition of HAS genes by means of antisense oligonucleotides on the behavior of osteosarcoma cells.

Methods: Total RNA was isolated from osteosarcoma cell lines, MG-63, which were cultured in monolayer, and subjected to competitive, quantitative RT-PCR to determine the mRNA copy numbers of HAS genes. Antisense oligonucleotides complimentary to the predominant HAS gene was transfected into osteosarcoma cells using a lipofection facilitator. The inhibition of HAS and HA expression was determined competitive RT-PCR and HA staining, respectively. Cell proliferation and invasion were analyzed by MTT assay and matrigel assay.

Results: HAS-2 mRNA expression was 90-hold higher than HAS-3. Antisense oligos treatment for HAS-2 resulted in a 60% inhibition of HAS-2 mRNA levels and a remarkable decrease of HA accumulation around the cells. The suppression of HAS-2 also lead to the suppression of cell proliferation and invasion.

Conclusions: HA inhibition via the down-regulation of HAS genes could control the behavior of osteosarcoma cells. Thus, the use of small antisense oligonucleotides to affect the selective inhibition of genes may provide a useful tool for the control of osteosarcomas.