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A Collagen/DBP Sponge System Designed for in Vitro Analysis of Chondroinduction

Published online by Cambridge University Press:  15 February 2011

Shuichi Mizuno ,
Chris Lycette ,
Charlene Quinto  and
Julie Glowacki
Shuichi Mizuno
Affiliation:
Orthopedic Research, Brigham and Women's Hospital, Harvard Medical School, 75 Francis St., Boston, MA 02115
Chris Lycette
Affiliation:
Orthopedic Research, Brigham and Women's Hospital, Harvard Medical School, 75 Francis St., Boston, MA 02115
Charlene Quinto
Affiliation:
Orthopedic Research, Brigham and Women's Hospital, Harvard Medical School, 75 Francis St., Boston, MA 02115
Julie Glowacki
Affiliation:
Orthopedic Research, Brigham and Women's Hospital, Harvard Medical School, 75 Francis St., Boston, MA 02115
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Abstract

In response to subcutaneous implants of demineralized bone powder (DBP), cells are attracted to the DBP, are converted to chondroblasts, and produce a cartilage matrix that is resorbed and replaced by bone. In order to define the cellular mechanisms of this induction, we developed a collagen sponge model for simulating the in vivo environment and for promoting the ingrowth and viability of cells cultured in them in vitro. Reconstituted pepsin–digested type I collagen from bovine hide was neutralized. Rat DBP (75–250 εm) was added into the collagen mixture (20 mg/ml). In order to simulate the connective tissue environment, modified chondroitin sulfate, heparan sulfate, or hyaluronic acid was added into the mixture. Aliquots (0.2 ml) were placed in 3/8 inch diameter molds and freeze-dried. Human dermal fibroblasts were cultured from minced fresh tissue and inoculated at 1.5 × 105 cells/sponge. Fifteen hours later, some sponges were transferred to medium which contained growth factors (PDGF or TGF-β). At intervals, samples were examined histologically. The inoculated cells attached to the collagen fibers and migrated into the sponge. Eventually the sponges contracted and acquired an oval shape. Cells on or near DBP were ovoid or stellate in shape. Cell morphology was modulated by glycosaminoglycan composition of the sponge. Increasing doses of PDGF or TGF-β promoted cellularity within the sponges. In conclusion, this system simulates the in vivo environment but allows accessibility for analysis. This three-dimensional matrix culture system will enable investigation of mechanisms of chondroinduction by morphogenic material.

Type
Research Article
Information
MRS Online Proceedings Library (OPL) , Volume 252: Symposium T – Tissue-Inducing Biomaterials , 1991 , 133
Copyright
Copyright © Materials Research Society 1992

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References

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