1: J Cell Biochem. 2005 May 1;95(1):53-63. 

N-cadherin mediated distribution of beta-catenin alters MAP kinase and BMP-2
signaling on chondrogenesis-related gene expression.

Modarresi R, Lafond T, Roman-Blas JA, Danielson KG, Tuan RS, Seghatoleslami MR.

Department of Orthopaedic Surgery Research, Thomas Jefferson University,
Philadelphia, Philadelphia 19107, USA.

We have examined the effect of calcium-dependent adhesion, mediated by
N-cadherin, on cell signaling during chondrogenesis of multipotential embryonic
mouse C3H10T1/2 cells. The activity of chondrogenic genes, type II collagen,
aggrecan, and Sox9 were examined in monolayer (non-chondrogenic), and micromass
(chondrogenic) cultures of parental C3H10T1/2 cells and altered C3H10T1/2 cell
lines that express a dominant negative form of N-cadherin (delta390-T1/2) or
overexpress normal N-cadherin (MNCD2-T1/2). Our findings show that
missexpression or inhibition of N-cadherin in C3H10T1/2 cells results in
temporal and spatial changes in expression of the chondrogenic genes Sox9,
aggrecan, and collagen type II. We have also analyzed activity of the serum
response factor (SRF), a nuclear target of MAP kinase signaling implicated in
chondrogenesis. In semi-confluent monolayer cultures (minimum cell-cell contact)
of C3H10T1/2, MNCD2-T1/2, or delta390-T1/2 cells, there was no significant
change in the pattern of MAP kinase or bone morphogenetic protein-2 (BMP-2)
regulation of SRF. However, in micromass cultures, the effect of MAP kinase and
BMP-2 on SRF activity was proportional to the nuclear localization of
beta-catenin, a Wnt stabilized cytoplasmic factor that can associate with
lymphoid enhancer-binding factor (LEF) to serve as a transcription factor. Our
findings suggest that the extent of adherens junction formation mediated by
N-cadherin can modulate the potential Wnt-induced nuclear activity of
beta-catenin. 2005 Wiley-Liss, Inc

PMID: 15723280 [PubMed - indexed for MEDLINE]


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2: J Cell Biochem. 2003 Apr 15;88(6):1129-44. 

Progression of chondrogenesis in C3H10T1/2 cells is associated with prolonged
and tight regulation of ERK1/2.

Seghatoleslami MR, Roman-Blas JA, Rainville AM, Modaressi R, Danielson KG, Tuan
RS.

Division of Rheumatology, Department of Medicine, Thomas Jefferson University,
Philadelphia, Pennsylvania 19107, USA. reza.seghatoleslami@mexil.tju.edu

Close contact of mesenchymal cells in vivo and also in super dense micromass
cultures in vitro results in cellular condensation and alteration of existing
cellular signaling required for initiation and progression of chondrogenesis. To
investigate chondrogenesis related changes in the activity of ubiquitous cell
signaling mediated by mitogen-activated protein kinases (MAP kinase), we have
compared the effect of cell seeding of pluripotent C3H10T1/2 mesenchymal cells
as monolayers (non-chondrogenic culture) or high density micromass cultures
(chondrogenic) on the regulation and phosphorylation state of extracellular
signal-regulated kinase 1 and 2 (ERK1/2) and also on regulation of ERK1/2
nuclear targets, namely, activation protein-1 (AP-1) and serum response factor
(SRF). Increasing cell density resulted in reduced DNA binding as well as
activity of AP-1. SRF activity, on the other hand, was up-regulated in confluent
monolayer cultures but like AP-1 was inhibited in micromass cultures. Low levels
of PD 98059 (5 microM), a specific inhibitor of ERK1/2, resulted in delayed
induction of AP-1 and SRF activity whereas higher concentrations of this
inhibitor (10-50 microM) conferred an opposite effect. Increasing concentrations
of the PD 98059 inhibitor in long term monolayer or micromass cultures (2.5 day)
resulted in differential regulation of c-Fos and c-Jun protein levels as well as
total expression and phosphorylation levels of ERK1/2. PD 98059 treatment of
C3H10T1/2 micromass cultures also resulted in up-regulation of type IIB collagen
and Sox9 gene expression. While high expression of aggrecan and type IIB
collagen genes were dependent on BMP-2 signaling, ERK inhibition of BMP-2
treated micromass cultures resulted in reduced activity of both genes. Our
findings show that the activity of ERK1/2 in chondrogenic cultures of C3H10T1/2
cells is tightly controlled and can cross interact with other signaling
activities mediated by BMP-2 to positively regulate chondrogensis.

PMID: 12647296 [PubMed - indexed for MEDLINE]


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