MARROW STROMAL CELL COMMITMENT TO MINERALIZATION UNDER THE EFFECT OF A PROLYL HYDROXYLASE INHIBITOR

Mitochondrial response to the effect of a hydroxylase (PH) inhibitor w as tested in marrow stromal cells during stimulation of osteoprogenito r cell (OPC) differentiation. The rationale for this was to explore pa thways of regulatory interactions between procollagen synthesis and mi tochondrial respiration that could be linked to the commitment of OPCs to mineralization. Stimulated OPCs exposed to the PH inhibitor cis-hy droxyproline (cis-HP), compared to the noninhibiting isomer trans-HP, for 11 days showed a dose-dependent decrease in cell proliferation, th e surviving cells showed increased alkaline phosphatase activity. Tran s-HP did not influence the cis-HP effect on ALP and on proliferation a rrest. Short time exposures, 2-3 days, to cis-HP at different periods suggested that Days 0-3 and 3-5 were critical for the commitment to Da y 21 mineralization of OPCs. On Days 0-3 cells were most sensitive to cis-HP, since on Day 11, 8 days after removal of cis-HP, they were too scarce to be counted by the staining method. However, the presence of 5.0 mM cis-HP in the cultures during Days 3-5 has induced on Day 21 c lose to 24-fold more mineralization/cell than controls, compared to th e trans-HP effect, which was only close to 3-fold. The presence of cis -HP in the cultures on Days 0-3 has augmented the mitochondrial Day 3 retention of rhodamine 123 (Rho) in the stromal cells, relative to con trols, compared to the presence of trans-HP. However, the presence of cis-HP during Days 3-5 or 3-6 resulted in lower Day 5 Rho retention, r elative to controls, which was not significantly different from the re tention that resulted from trans-HP. Since Rho retention is related to the final result of aerobic respiration level, these results are inte rpreted as a cis-HP inhibitory effect on procollagen peptidyl-proline hydroxylation, which may in turn release oxygen surpluses, to be avail able for mitochondrial consumption. The fall in Rho retention response s to cis-HP between Days 0-3 and 3-5 is suggesting either abrupt decre ase in proline hydroxylation or poor mitochondrial sensitivity to oxyg en in Day 3-5 cultures. (C) 1994 Wiley-Liss, Inc.