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Modulation of chondrocyte proliferation by ascorbic acid and BMP-2
R Venezian1, B J Shenker, S Datar
1Department of Biochemistry, School of Dental Medicine, University of Pennsylvania, Philadelphia 19104-6003, USA.
Journal of Cellular Physiology
|February 14, 1998
Summary
Ascorbic acid significantly boosts chondrocyte proliferation in serum-free cultures, increasing thymidine incorporation. However, this proliferation boost is linked to increased apoptosis, especially during chondrocyte hypertrophy.
Area of Science:
- Cell Biology
- Biochemistry
- Developmental Biology
Background:
- Chondrocytes can survive serum-free conditions with specific supplements like insulin, thyroxine, and cysteine.
- Serum-free culture of chondrocytes shows limited thymidine incorporation compared to fetal bovine serum (FBS).
- Skin fibroblast proliferation is significantly reduced (<5%) in serum-free conditions.
Purpose of the Study:
- To investigate the effect of ascorbic acid on chondrocyte proliferation in serum-free medium.
- To analyze the impact of ascorbic acid on cell cycle progression and apoptosis in chondrocytes.
- To explore the relationship between chondrocyte hypertrophy, proliferation, and apoptosis.
Main Methods:
- Culturing sternal chondrocytes from chick embryos under serum-free conditions.
- Stimulating proliferation using varying concentrations of ascorbic acid.
- Assessing cell proliferation via thymidine incorporation and Bromo-deoxyuridine (BrdU) labeling.
- Quantifying apoptosis using nuclear morphology, TUNEL assay, and FACS analysis.
- Inducing hypertrophy in prehypertrophic chondrocytes using rhBMP-2 and ascorbic acid.
Main Results:
- Ascorbic acid (30-50 microM) stimulated chondrocyte proliferation 4-5x, exceeding levels seen with 10% FBS.
- Proliferation stimulation occurred within 3-5 hours of ascorbate exposure, peaking at 12-15 hours.
- Approximately 25% of chondrocytes entered S phase 16-20 hours post-ascorbate treatment.
- Ascorbate treatment increased chondrocyte apoptosis from 5% to 15%.
- Hypertrophy induction with rhBMP-2 and ascorbate in prehypertrophic chondrocytes showed high apoptosis levels, similar to ascorbate alone.
- Caudal chondrocytes treated with BMP and ascorbate did not hypertrophy and had lower apoptosis rates.
Conclusions:
- Ascorbic acid is a potent stimulator of chondrocyte proliferation in serum-free culture.
- Increased proliferation stimulated by ascorbate is accompanied by a significant rise in apoptosis.
- The transition of chondrocytes to hypertrophy is associated with reduced proliferation and elevated apoptosis.