DISCORDANT EXPRESSION OF OSTEOBLAST MARKERS IN MC3T3-E1 CELLS THAT SYNTHESIZE A HIGH TURNOVER MATRIX

To examine the autocrine effects that an organizing extracellular matr ix has on osteoblast precursors, we created MC3T3-E1 cell lines that s tably expressed pro-alpha 1(I) collagen chains with a truncated triple helical domain. Cells that had incorporated the pro-alpha 1(I) expres sion plasmid (pMG155) expressed shortened pro-alpha 1(I) transcripts a t high levels and efficiently secreted the expression gene products in to culture media. Those cells lost over 30% of newly deposited collage nous matrix compared with virtually no loss in control cultures, and m edia from the abnormal cells had qualitative differences in matrix met alloprotinase production. Electron micrographs strongly suggested that type I collagen molecules containing the truncated pro-alpha 1(I) cha ins dramatically interfered with collagen fibrillogenesis in newly for ming osteoblast matrix. Abnormal collagen fibrillogenesis was also ass ociated with altered characteristics of cellular differentiation in th at abnormal cells displayed a delayed and attenuated increase in alkal ine phosphatase activity. Surprisingly, synthesis of osteocalcin was m ore than 5-fold higher than control cultures. These findings demonstra te that osteoblasts require a normally structured collagenous matrix f or up-regulation of alkaline phosphatase activity. However, in the pre sence of rapid turnover of osteoblast matrix, osteocalcin gene express ion may be up-regulated in response to local signals by an unknown mec hanism.