New chondrocyte genes discovered by representational difference analysis of chondroinduced human fibroblasts.
This report includes a review of the potential for gene expression analyses to provide new information for solving problems in skeletal repair and regeneration. It focuses on two approaches: high-throughput gene array methods and representational difference analysis (RDA). The principles underlying these methods are presented with experimental tutorials and some applications. Second, this report includes a review of results from applying both approaches to an in vitro model of postnatal chondroinduction by demineralized bone powder (DBP). Human dermal fibroblasts (hDFs) cultured with DBP acquire a chondroblast phenotype and express cartilage-specific matrix proteins after 7 days. We used cDNA macroarrays and RDA to identify the genes that were altered prior to expression of the chondroblast phenotype, i.e., after only 3 days' culture with DBP. Using a strategy of data management and reduction based upon biological functions, we reported several functional families of genes (cytoskeletal elements, protein synthesis/trafficking, and matrix molecules and their modifiers) that are upregulated during chondroinduction of hDFs. Together with histological and biochemical evidence of the chondroblast phenotype, the gene expression patterns indicate that there are specific stages of induced chondrocyte differentiation in this experimental system. Third, this report includes a new study, in which DBP-regulated genes were used as a data base to derive new information on the cell biology of chondrocytes. The objective was to determine whether a set of genes expressed during induction of chondrocyte differentiation is also expressed by mature articular chondrocytes. Our search of the literature for 59 of the DBP-regulated genes disclosed that expression of 20 of them (33%) had been documented in mature cartilage or chondrocytes. Of the 39 genes not previously documented in cartilage, 11 were tested by RT-PCR and all were found to be expressed in freshly isolated adult human chondrocytes. This review and these new data show how the strategy of high-throughput methods and functional data reduction can expand our knowledge of chondrocyte cell biology.
Yates
Karen E
KE
Department of Orthopedic Surgery, Brigham and Women's Hospital, Boston, Mass., USA. kyates@partners.org
Forbes
Rachel L
RL
Glowacki
Julie
J
D000818:Animals; D018775:Bone Demineralization Technique; D001842:Bone and Bones / Q000737:chemistry; D015815:Cell Adhesion Molecules / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics*; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000235:genetics; D016326:Extracellular Matrix Proteins / Q000235:genetics; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects; D020869:Gene Expression Profiling; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics; D011208:Powders / Q000494:pharmacology; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012196:Review Literature as Topic; D015398:Signal Transduction / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics; D015854:Up-Regulation / Q000235:genetics
D015815:Cell Adhesion Molecules; D018797:Cell Cycle Proteins; D003598:Cytoskeletal Proteins; D016326:Extracellular Matrix Proteins; D036341:Intercellular Signaling Peptides and Proteins; D011208:Powders; D014157:Transcription Factors