Enhancement of Connexin 43 Expression Increases Proliferation and Differentiation of an Osteoblast-like Cell Line

Gramsch, B.; Gabriel, Heinz-Dieter; Wiemann, Martin; Grümmer, Ruth; Winterhager, Elke; Bingmann, D.; Schirrmacher, K.

doi:10.1006/excr.2000.5145

Cited by 103 publications

(68 citation statements)

References 42 publications

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“…Our assessment of functional GJIC in HOS cells done by DEPArrayª microfluidic approach is in agreement with a recent new high throughput screening proposed by Lee et al (2015) between the two cell types during this crucial phase (Pollmann et al, 2005, Saito-Katsuragi et al 2007). In line with several studies (Gramsch et al 2001, Matemba et al 2006, we found an increase in Cx43 expression in the differentiated osteoblasts. This may explain why we observed GJIC between hOS cells and differentiated osteoblasts at either 7 or 21 days, whereas no cell-cell communication was observed between osteoblast precursors (MSC) and tumour cells.…”

Section: Osteosarcoma Cells Transduced Withsupporting

confidence: 93%

Analysis of gap junctional intercellular communications using a dielectrophoresis-based microchip

Gabriel

Charrier

Royer

et al. 2017

European Journal of Cell Biology

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International audiencePlease cite this article in press as: Tellez-Gabriel, M., et al., Analysis of gap junctional intercellular communications using a dielectrophoresis-based microchip. Gap junctions are transmembrane structures that directly connect the cytoplasm of adjacent cells, making intercellular communications possible. It has been shown that the behaviour of several tumours – such as bone tumours – is related to gap junction intercellular communications (GJIC). Several methodologies are available for studying GJIC, based on measuring different parameters that are useful for multiple applications, such as the study of carcinogenesis for example. These methods nevertheless have several limitations. The present manuscript describes the setting up of a dielectrophoresis (DEP)-based lab-on-a-chip platform for the real-time study of Gap Junctional Intercellular Communication between osteosarcoma cells and the main cells accessible to their microenvironment. We conclude that using the DEParray technology for the GJIC assessment has several advantages comparing to current techniques. This methodology is less harmful for cells integrity; cells can be recovered after interaction to make further molecular analysis; it is possible to study GJIC in real time; we can promote cell interactions using up to five different populations. The setting up of this new methodology overcomes several difficulties to perform experiments for solving questions about GJIC process that we are not able to do with current technics

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Section: Osteosarcoma Cells Transduced Withsupporting

confidence: 93%

Analysis of gap junctional intercellular communications using a dielectrophoresis-based microchip

Gabriel

Charrier

Royer

et al. 2017

European Journal of Cell Biology

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Section: Discussionmentioning

confidence: 99%

“…Likewise, cells from Cx43 ϩ/Ϫ mice have deficient calcium mobilization in response to endothelin-1. Additionally, several in vitro studies have upregulated (Steinberg et al, 1994;Gramsch et al, 2001) or down-regulated Cx43 abundance or function (Van der Molen et al, 1996;Lecanda et al, 1998;Li et al, 1999Li et al, , 2006Upham et al, 2003;Plotkin et al, 2008) with profound effects on osteoblast function.Our ultimate goals are to gain insight into how osteoblasts utilize intercellular communication in order to regulate function and to identify biologically relevant molecules being passed through Cx43 gap junction channels in osteoblasts. Accordingly, our approach is to work our way from a defined transcriptional response affected by gap junctions back through the signaling cascade-systematically examining a path from the nucleus to membrane.…”

mentioning

confidence: 99%

Connexin43 Potentiates Osteoblast Responsiveness to Fibroblast Growth Factor 2 via a Protein Kinase C-Delta/Runx2–dependent Mechanism

Lima

Niger

Hebert

et al. 2009

MBoC

100

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In this study, we examine the role of the gap junction protein, connexin43 (Cx43), in the transcriptional response of osteocalcin to fibroblast growth factor 2 (FGF2) in MC3T3 osteoblasts. By luciferase reporter assays, we identify that the osteocalcin transcriptional response to FGF2 is markedly increased by overexpression of Cx43, an effect that is mediated by Runx2 via its OSE2 cognate element, but not by a previously identified connexin-responsive Sp1/Sp3-binding element. Furthermore, disruption of Cx43 function with Cx43 siRNAs or overexpression of connexin45 markedly attenuates the response to FGF2. Inhibition of protein kinase C delta (PKC␦) with rottlerin or siRNA-mediated knockdown abrogates the osteocalcin response to FGF2. Additionally, we show that upon treatment with FGF2, PKC␦ translocates to the nucleus, PKC␦ and Runx2 are phosphorylated and these events are enhanced by Cx43 overexpression, suggesting that the degree of activation is enhanced by increased Cx43 levels. Indeed, chromatin immunoprecipitations of the osteocalcin proximal promoter with antibodies against Runx2 demonstrate that the recruitment of Runx2 to the osteocalcin promoter in response to FGF2 treatment is dramatically enhanced by Cx43 overexpression. Thus, Cx43 plays a critical role in regulating the ability of osteoblasts to respond to FGF2 by impacting PKC␦ and Runx2 function. INTRODUCTIONBone formation and remodeling is a tightly organized and dynamic process that requires the coordinated action of osteoblasts, osteocytes, and osteoclasts to maintain bone homeostasis. It is hypothesized that osteoblasts and osteocytes coordinate their activities, at least in part, through direct cell-to-cell communication through gap junctions. Gap junctions are composed of connexins, a family of transmembrane proteins that constitute the intercellular gap junction channels (Beyer et al., 1990). Six connexins assemble to make up the gap junction hemichannel on the plasma membrane of one cell, which docks with a hemichannel on an adjacent cell to form an aqueous gap junction channel. The resultant gap junctions provide direct conduits for the passage of ions and other low molecular weight molecules, including second messengers, among cells.The gap junction protein connexin43 (Cx43) is abundantly expressed in both osteoblasts and osteocytes, where it has been hypothesized to transmit hormonal signals, mechanical load, and growth factor cues among cells in order to coordinate the synthesis of new bone (reviewed in Stains and Civitelli, 2005a;Jiang et al., 2007). Genetic ablation of gja1, the gene encoding Cx43, in mice leads to a severe delay in the ossification of both intramembranous and endochondral derived skeletal elements during embryonic development (Lecanda et al., 2000). The bones of these animals are remarkably brittle, and the osteoblasts isolated from the Cx43 null animals are dysfunctional, with reduced osteogenic and mineralizing capacity (Lecanda et al., 2000). These Cx43-deficient mice die at birth because of a defect in cardiac f...

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“…Two genes encode for proteins involved in adhesion and cell-cell contact, periostin (already mentioned above) and connexin 43. Periostin, previously called osteoblast-specific-factor-2 (Osf-2), plays a role in the recruitment and attachment of osteoblast precursors in the periosteum (Horiuchi et al 1999), and enhancement of connexin 43 expression has been shown to increase both proliferation and differentiation of osteoblasts (Gramsch et al 2001). Three genes encode for transcription factorrelated proteins, the homeobox transcription factor Prx2, the AP-1 family member fra-1, and Smad6.…”

Section: Validation Of Bone Class Membersmentioning

confidence: 99%

Finding Genes in the C2C12 Osteogenic Pathway by k-Nearest-Neighbor Classification of Expression Data

Theilhaber¹,

Connolly²,

Roman-Roman³

et al. 2002

Genome Res.

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A supervised classification scheme for analyzing microarray expression data, based on the k-nearest-neighbor method coupled to noise-reduction filters, has been used to find genes involved in the osteogenic pathway of the mouse C2C12 cell line studied here as a model for in vivo osteogenesis. The scheme uses as input a training set embodying expert biological knowledge, and provides internal estimates of its own misclassification errors, which furthermore enables systematic optimization of the classifier parameters. On the basis of the C2C12-generated expression data set with 34,130 expression profiles across 2 time courses, each comprised of 6 points, and a training set containing known members of the osteogenic, myoblastic, and adipocytic pathways, 176 new genes in addition to 28 originally in the training set are selected as relevant to osteogenesis. For this selection, the estimated sensitivity is 42% and the posterior false-positive rate (fraction of candidates that are spurious) is 12%. The corresponding sensitivity and false-positive rate for detection of myoblastic genes are 9% and 31%, respectively, and only 4% and ∼100%, respectively, for adipocytic genes, in accordance with an experimental design that predominantly stimulated the osteogenic pathway. Validation of this selection is provided by examining expression of the genes in an independent biological assay involving mouse calvaria (skull bone) primary cell cultures, in which a large fraction of the 176 genes are seen to be strongly regulated, as well as by case-by-case analysis of the genes on the basis of expert domain knowledge. The methodology should be generalizable to any situation in which enough a priori biological knowledge exists to define a training set.

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Enhancement of Connexin 43 Expression Increases Proliferation and Differentiation of an Osteoblast-like Cell Line

Cited by 103 publications

References 42 publications

Analysis of gap junctional intercellular communications using a dielectrophoresis-based microchip

Analysis of gap junctional intercellular communications using a dielectrophoresis-based microchip

Connexin43 Potentiates Osteoblast Responsiveness to Fibroblast Growth Factor 2 via a Protein Kinase C-Delta/Runx2–dependent Mechanism

Finding Genes in the C2C12 Osteogenic Pathway by k-Nearest-Neighbor Classification of Expression Data

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