Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Biochemical and molecular studies of osteoclasts generally require cells in a reasonable degree of purity. The chicken has been extremely useful in this regard, as abundant avian osteoclasts can be generated in vitro entirely from pure populations of marrow macrophage precursors. Propagation of murine osteoclasts is, in contrast, far less efficient, demanding the presence of stromal cells. The aims of this study were to develop a method by which murine osteoclasts generated in culture, can be effectively enriched while maintaining viability and, to explore the mechanisms by which stromal cells promote murine osteoclast generation and survival. We find that 10(6) fractionated murine marrow cells enriched, for marrow-residing colony-forming units (CFU-cs), yield 3000-4000 tartrate-resistant acid phosphatase (TRAP)-expressing multinucleated giant cells when cultured for 12 days with ST-2 stromal cells. These cells are osteoclasts as evidenced by their ability to "pit" bone slices, resorb radiolabeled bone particles, and generate cyclic AMP in response to calcitonin. Treatment of these generated osteoclast cultures with bacterial collagenase for 2 hours at 37 degrees selectively removes virtually all ST-2 cells, yielding a > 60% pure population of TRAP and calcitonin receptor-expressing cells, 90% of which are viable. These cells continue to respond to calcitonin and survive for 24 hours in the absence of ST-2 cells. We also found that murine osteoclast generation depends upon contact of osteoclast precursors with viable ST-2 cells. Furthermore, the stromal cells secrete macrophage colony-stimulating factor (CSF-1), and the anti-CSF-1 antibody 5A1 inhibits murine osteoclastogenesis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Enrichment of generated murine osteoclasts. 753 41

Numerous resorptive stimuli have been shown to enhance osteoclast differentiation, increasing osteoclast numbers and accelerating bone resorption. Currently, there is much less understanding of regulation of mature osteoclast activity. Indeed, there is presently only minimal evidence of changes in gene expression as a mechanism for altering bone resorption. We investigate here, in the mature osteoclast, regulation of 2 genes-carbonic anhydrase II (CAII) and calcitonin receptor (CTR) in response to acidosis, which is known to increase bone resorption. We studied the effect of acid pH on CAII and CTR mRNA expression in mature osteoclasts raised in coculture of ST-2 and primary marrow cells. On day 6 of culture, stromal cells were removed with collagenase, the remaining osteoclasts were incubated overnight, and then exposed to varying pH. RT-PCR was performed on total RNA using primers for CAII, CTR, or glyceraldehyde dehydrogenase phosphate (GAP). Expression of CTR mRNA was increased 2.14 +/- 0.41 and 2.56 +/- 0.45 (P < 0.05)-fold by a 4-hour exposure to pH 6.75 and 6.5, respectively. CAII mRNA was similarly increased 2.18 +/- 0.42 and 2.63 +/- 0.48 (P < 0.05)-fold by pH 6.75 and 6.5, respectively. Increased expression of CAII and CTR mRNA was seen by 2 hours and maximally by 4 hours. Increased expression of CTR and CAII mRNA was not explained by increases in osteoclast numbers: pH 7.4-100 +/- 3.7, 6.75-133 +/- 8.3, 6.5-124 +/- 7.8. These results demonstrate upregulation of two osteoclast genes in response to acidosis, illustrating the ability of the mature osteoclast to respond to resorptive signals with increased functional gene expression.
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PMID:Acid pH increases carbonic anhydrase II and calcitonin receptor mRNA expression in mature osteoclasts. 1092 Feb 24