Lead induces chondrogenesis and alters transforming growth factor-beta and bone morphogenetic protein signaling in mesenchymal cell populations.
BACKGROUND
It has been established that skeletal growth is stunted in lead-exposed children. Because chondrogenesis is a seminal step during skeletal development, elucidating the impact of Pb on this process is the first step toward understanding the mechanism of Pb toxicity in the skeleton.


OBJECTIVE
The aim of this study was to test the hypothesis that Pb alters chondrogenic commitment of mesenchymal cells and to assess the effects of Pb on various signaling pathways.


METHODS
We assessed the influence of Pb on chondrogenesis in murine limb bud mesenchymal cells (MSCs) using nodule formation assays and gene analyses. The effects of Pb on transforming growth factor-beta (TGF-beta) and bone morphogenetic protein (BMP) signaling was studied using luciferase-based reporters and Western analyses, and luciferase-based assays were used to study cyclic adenosine monophosphate response element binding protein (CREB), beta-catenin, AP-1, and nuclear factor-kappa B (NF-kappaB) signaling. We also used an ectopic bone formation assay to determine how Pb affects chondrogenesis in vivo.


RESULTS
Pb-exposed MSCs showed enhanced basal and TGF-beta/BMP induction of chondrogenesis, evidenced by enhanced nodule formation and up-regulation of Sox-9, type 2 collagen, and aggrecan, all key markers of chondrogenesis. We observed enhanced chondrogenesis during ectopic bone formation in mice preexposed to Pb via drinking water. In MSCs, Pb enhanced TGF-beta but inhibited BMP-2 signaling, as measured by luciferase reporter assays and Western analyses of Smad phosphorylation. Although Pb had no effect on basal CREB or Wnt/beta-catenin pathway activity, it induced NFkappaB signaling and inhibited AP-1 signaling.


CONCLUSIONS
The in vitro and in vivo induction of chondrogenesis by Pb likely involves modulation and integration of multiple signaling pathways including TGF-beta, BMP, AP-1, and NFkappaB.
Zuscik
Michael J
MJ
Center for Musculoskeletal Research, University of Rochester School of Medicine and Dentistry, Rochester, New York, USA.
Ma
Lin
L
Buckley
Taylor
T
Puzas
J Edward
JE
Drissi
Hicham
H
Schwarz
Edward M
EM
O'Keefe
Regis J
RJ
D000818:Animals; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000187:drug effects*; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D005260:Female; D006609:High Mobility Group Proteins / Q000235:genetics; D007854:Lead / Q000633:toxicity*; D059630:Mesenchymal Stem Cells / Q000187:drug effects* / Q000502:physiology; D051379:Mice; D008815:Mice, Inbred Strains; D016328:NF-kappa B / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D055758:SOX9 Transcription Factor; D051785:Smad Proteins / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics; D016212:Transforming Growth Factor beta / Q000378:metabolism*