CD34 human hematopoietic progenitor cell line, MUTZ-3, differentiates into functional osteoclasts.
OBJECTIVE
CD14(+) monocyte cell lines can differentiate into an osteoclast (OC)-like lineage. However, the identification of human cell lines with stem cell characteristics, capable of differentiating into OCs, would provide a tool for the study of the molecular mechanisms regulating their commitment, differentiation, and function. Since the human acute myeloid leukemia cell line MUTZ-3 contains both CD34(+) stem cell and CD14(+) cell populations, we investigated the capacity of the stem/progenitor CD34(+) population to differentiate into functional OCs.


METHODS
Sorted MUTZ-3-CD34(+) and MUTZ-3-CD14(+) cells were cultured in presence of M-CSF, RANK-L, and TNF-alpha to generate OCs. Differentiation was evaluated by TRAP staining and RT-PCR, which assessed the expression of c-fms, RANK, MMP-9, CATK, TRAP, and CTR in -CD34(+)OC and -CD14(+)OC cells. Resorption pit formation was also evaluated. CD34, CD14, M-CSF-R, RANK, and CTR expression was assessed by FACS analysis.


RESULTS
MUTZ-3-CD34(+) differentiated into OCs, displaying the full range of differentiation markers; MMP-9, CATK, TRAP, and RANK mRNA were detected from day 3 of culture, whereas CTR from day 12. Stimulated MUTZ-3-CD34(+) generated functional osteoclasts that formed extensive resorption lacunae on both mineralized surface and bone slices. Surprisingly, in both sorted populations we identified a population M-CSF-R(+)/RANK(+) that at the same time co-expressed CD14 and CD34.


CONCLUSIONS
These findings demonstrate that MUTZ-3 cells constitute an invaluable model to study the expression pattern in different developmental stages of commitment and differentiation. Importantly, the data indicate that the CD14(+)CD34(+)M-CSF-R(+)RANK(+) population represents an intermediate stage of differentiation from CD34 precursors and monocytes to osteoclast.
Ciraci
Elisa
E
Laboratory of Stem Cells, IRCCS-Pediatric Hospital of Bambino Gesù, Rome, Italy.
Barisani
Donatella
D
Parafioriti
Antonina
A
Formisano
Giuseppe
G
Arancia
Giuseppe
G
Bottazzo
GianFranco
G
Berardi
Anna C
AC
D018952:Antigens, CD34; D000943:Antigens, Differentiation / Q000096:biosynthesis; D002460:Cell Line; D016207:Cytokines / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000502:physiology* / Q000648:ultrastructure; D006801:Humans; D018950:Lipopolysaccharide Receptors; D008855:Microscopy, Electron, Scanning; D009000:Monocytes / Q000502:physiology* / Q000648:ultrastructure; D010010:Osteoclasts / Q000502:physiology* / Q000648:ultrastructure; D053246:Receptor Activator of Nuclear Factor-kappa B; D016186:Receptor, Macrophage Colony-Stimulating Factor; D013997:Time Factors
D018952:Antigens, CD34; D000943:Antigens, Differentiation; D016207:Cytokines; D018950:Lipopolysaccharide Receptors; D053246:Receptor Activator of Nuclear Factor-kappa B; C505686:TNFRSF11A protein, human; D016186:Receptor, Macrophage Colony-Stimulating Factor