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. 2014 Jan 15;9(1):e85161.
doi: 10.1371/journal.pone.0085161. eCollection 2014.

Osx-Cre targets multiple cell types besides osteoblast lineage in postnatal mice

Jianquan Chen  1 , Yu Shi  1 , Jenna Regan  1 , Kannan Karuppaiah  2 , David M Ornitz  2 , Fanxin Long  3
Affiliations

Osx-Cre targets multiple cell types besides osteoblast lineage in postnatal mice

Jianquan Chen et al. PLoS One. .

Abstract

Osterix (Osx or Sp7) is a zinc-finger-family transcriptional factor essential for osteoblast differentiation in mammals. The Osx-Cre mouse line (also known as Osx1-GFP::Cre) expresses GFP::Cre fusion protein from a BAC transgene containing the Osx regulatory sequence. The mouse strain was initially characterized during embryogenesis, and found to target mainly osteoblast-lineage cells. Because the strain has been increasingly used in postnatal studies, it is important to evaluate its targeting specificity in mice after birth. By crossing the Osx-Cre mouse with the R26-mT/mG reporter line and analyzing the progenies at two months of age, we find that Osx-Cre targets not only osteoblasts, osteocytes and hypertrophic chondrocytes as expected, but also stromal cells, adipocytes and perivascular cells in the bone marrow. The targeting of adipocytes and perivascular cells appears to be specific to those residing within the bone marrow, as the same cell types elsewhere are not targeted. Beyond the skeleton, Osx-Cre also targets the olfactory glomerular cells, and a subset of the gastric and intestinal epithelium. Thus, potential contributions from the non-osteoblast-lineage cells should be considered when Osx-Cre is used to study gene functions in postnatal mice.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Osx-Cre targets osteoblast lineage cells, hypertrophic chondrocytes and bone marrow stromal cells.
(A–B) Confocal images for direct fluorescence from EGFP (A, B) or EGFP/tdTomato (A′, B′) on longitudinal tibial sections from two-month-old R26-mT/mG (A, A′) or Osx-Cre; R26-mT/mG mice (B, B′); 1: chondro-osseous junction of primary ossification center; 2: secondary ossification center. (C–F, C′–F′) Higher magnification images for EGFP (C–F) or EGFP/tdTomato: trabecular bone (C, C′), cortical bone (D, D′), growth plate (E, E′) and bone marrow (F, F′). Red: membrane-targeted tdTomato; Green: membrane-targeted EGFP; Blue: DAPI. Arrow: osteocyte.
Figure 2
Figure 2. Osx-Cre marks adipocytes in bone marrow.
(A–B) Double immunostaining for EGFP (A) and perilipin (B) on longitudinal sections of tibias from two-month-old Osx-Cre; R26-mT/mG mice. (C) Co-localization of EGFP and perilipin. Arrows denote co-expression of GFP and perilipin. Green: EGFP; magenta: perilipin; blue: DAPI.
Figure 3
Figure 3. Osx-Cre does not mark non-bone marrow adipocytes.
(A–B) Images for direct fluorescence from tdTomato (A) or EGFP (B) in whole-mount gonadal fat depots from two-month-old Osx-Cre; R26-mT/mG mice. (C–F) Direct fluorescence for tdTomato (C) and immunofluorescence for perilipin (D) and EGFP (E) on sections of gonadal fat depots from two-month-old Osx-Cre;R26-mT/mG mice. (G–K) Imaging of longitudinal sections of an intramuscular fat depot associated with a tibia from two-month-old Osx-Cre;R26-mT/mG mice. G: perilipin immunofluorescence; H: EGFP immunofluorescence; I: direct fluorescence for tdTomato; J: merged view of G–I; K: DAPI staining. Arrow: GFP-positive periosteum.
Figure 4
Figure 4. Osx-Cre marks perivascular smooth muscle cells in bone marrow.
(A–C) Confocal images of EGFP (A), tdTomato (B), and αSMA (C) on longitudinal sections of tibias from two-month-old Osx-Cre; R26-mT/mG mice. (D–F) Merged images. Arrow: co-expression of EGFP and αSMA. EGFP and αSMA: immunofluorescence; tdTomato: direct fluorescence. DAPI stains DNA blue.
Figure 5
Figure 5. Osx-Cre targets olfactory glomerular cells.
(A–D) Direct fluorescence of whole-mount rostral brain from a two-month-old Osx-Cre; R26-mT/mG mouse. ob: olfactory bulb. (E–H) Direct fluorescence of tdTomato (E, F) and EGFP (G, H) on sagittal sections through the olfactory bulb of Osx-Cre (E, G) or Osx-Cre; R26-mT/mG (F, H) mice at two months of age. GL: glomerular layer.
Figure 6
Figure 6. Osx-Cre targets gastric and intestinal epithelia.
(A–D) Direct fluorescence of tdTomato (A, C) and EGFP (B, D) in whole-mount small intestine of two-month-old R26-mT/mG (A, B) or Osx-Cre; R26-mT/mG mice (C, D). (E–G) Direct fluorescence on cross sections of small intestine from a two-month-old Osx-Cre; R26-mT/mG mouse. Red-boxed area in F shown at a higher magnification in G. (H–J) Direct fluorescence on stomach sections from a two-month-old Osx-Cre; R26-mT/mG mouse. Red-boxed area in I shown at a higher magnification in J. White arrow denotes parietal cells.
See this image and copyright information in PMC

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