Implications in Osteoporosis: Caffeine Impairs, Dexamethasone Increases Osteogenesis in MC3T3 Osteoblast Cells

Overview

Objectives/Goals To study the effect of caffeine and dexamethasone on osteoblast cell line MC3T3-E1, and determine the toxicity level of caffeine in these cells in vitro, so as to better understand the pathogenesis of osteoporosis. Hypotheses: 1)Caffeine will interfere with MC3T3-E1 osteogenesis in a dose-responsive manner as evidenced by decreased amounts of mineralized nodules on Von Kossa staining. 2)Osteoblasts grown in culture media supplemented with dexamethasone will show greater mineralization as compared to cultures grown without dexamethasone, but this will also be negatively influenced by caffeine. 3)Caffeine will affect osteoblast growth in a dose-responsive manner as seen by the toxicity study. Methods/Materials Cell line MC3T3-E1 was maintained in DMEM with 10%FBS and gentamicin at 37 deg C., humidified and 5%CO2. For Investigations #1 and #2, culture solutions with or without the inducers glycerol phosphate and ascorbic acid were compared to cultures treated with various concentrations of caffeine, and a subset of cells enhanced with dexamethasone. At 19 days, these cells were stained by Von Kossa technique to determine formation of nodules. For investigation # 3, caffeine (0 mM control; 0.1mM; 0.3mM; 1mM; 3mM; and 10mM) was added to nonconfluent cells. Cells were fixed at days 3, 4, and 5. Cell counts were done to determine caffeine toxicity and effect on growth. Results Investigations #1 and #2 show osteoblasts grown with caffeine form fewer nodules, regardless of induction (10mM Caffeinated=238, vs. Non-caffeinated Control=937). Cells grown with dexamethasone show much greater matrix nodule formation, but this, too, is diminished by caffeine(10mM Caffeinated=726, vs. Non-caffeinated Control=10,436). A novel finding was that dexamethasone was seen to act as an independent inducer of osteogenesis. Investigation #3 shows that caffeine is toxic to osteoblasts and negatively affects growth,primarily occurring at levels of 1.0 mM and above. Conclusions/Discussion The results support my original hypotheses. Caffeine interferes with osteogenesis in a dose-responsive manner shown by decreased amounts of mineralized nodules. Cells grown in culture media supplemented with dexamethasone show greater mineralization as compared to cultures grown without dexamethasone, but were also negatively influenced by caffeine. Caffeine did affect osteoblast growth in a dose-responsive manner as seen by the toxicity study.

Awards (1)

• First Place

Competition history

• CSEF 2009 Pharmacology (Senior Division Only) · Entry S1809

Resources

• CSEF project summary ↗

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