Development of mice with osteoblast-specific connexin43 gene deletion.
Genetic deficiency of Cx43 in vivo causes skeletal developmental defects, osteoblast dysfunction and perinatal lethality. To determine the role of Cx43 in the adult skeleton, we developed two models of osteoblast-specific Cx43 gene deletion using Cre mediated replacement of a "floxed" Cx43 allele with a LacZ reporter gene. Cre recombinase expression in osteoblasts was driven by either the osteocalcin OG2 promoter or the 2.3 kb fragment of the Colalpha1(I) promoter. Homozygous Cx43(fl/fl) mice, in which the Cx43 coding region is flanked by two loxP sites, were crossed with Cre expressing mice in a heterozygous Cx43-null background [Cx43(+/-); Colalpha1(I)-Cre or Cx43(+/-); OG2-Cre]. Cx43 gene ablation was demonstrated in tissues by selective X-gal staining of cells lining the endosteal surface, and in cultured osteoblastic cells from calvaria using different approaches. Although no LacZ expression was observed in proliferating calvaria cells, before osteoblast differentiation begins, post-proliferative cells isolated from conditional knockout mice [Cx43(fl/-); Colalpha1(I)-Cre or Cx43(fl/-); OG2-Cre] developed strong LacZ expression as they differentiated, in parallel to a progressive disappearance of Cx43 mRNA and protein abundance relative to controls. Selective Cre mediated Cx43 gene inactivation in bone forming cells will be useful to determine the role of Cx43 in adult skeletal homeostasis and overcome the perinatal lethality of the conventional null model.
Castro
Charles H M
CH
Division of Bone and Mineral Diseases, Washington University School of Medicine, St. Louis, MO 63110, USA.
Stains
Joseph P
JP
Sheikh
Sharmin
S
Szejnfeld
Vera Lucia
VL
Willecke
Klaus
K
Theis
Martin
M
Civitelli
Roberto
R
D000818:Animals; D018031:Connexin 43 / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D017930:Genes, Reporter; D051379:Mice; D018345:Mice, Knockout; D010006:Osteoblasts / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011995:Recombination, Genetic
D018031:Connexin 43