Use of Type I Collagen Green Fluorescent Protein Transgenes to Identify Subpopulations of Cells at Different Stages of the Osteoblast Lineage

Kalajzić, Ivo; Kalajzić, Žana; Kaliterna, M.; Gronowicz, Gloria; Clark, Stephen H.; Lichtler, Alexander C.; Rowe, David W.

doi:10.1359/jbmr.2002.17.1.15

Cited by 359 publications

(477 citation statements)

References 51 publications

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“…Animals were kindly provided by Dr. David Rowe and Dr. Alexander Lichtler from University of Connecticut Health Center, Farmington, CT. These transgenic lines have been characterized previously, and promoter constructs have been shown to support endogenous type I collagen like tissue expression pattern (3,17). In addition, our data indicate that changes in the reporter gene closely resemble changes in mature collagen synthesis estimated by Western blot analysis of the growth media.…”

Section: Methodssupporting

confidence: 61%

Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α₅β₁-integrin

Dodig¹,

Ogunwale²,

Dasarathy³

et al. 2007

American Journal of Physiology-Gastrointestinal and Liver Physi

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Hepatic stellate cells (HSC) differ in their phenotype depending on the initiation and progression of their activation. Our hypothesis was that different mechanisms govern type I collagen synthesis depending on stage of HSC activation. We investigated the role of ␣5␤1-integrin as a regulator of type I collagen gene COL1A1 expression in primary and passaged HSC cultures using transgenic mouse containing type I collagen gene COL1A1 promoter linked to the chloramphenicol acetyltransferase (CAT) reporter gene. The ␣5␤1 protein levels increased during the activation and were highest in day 6 primary cultures but decreased in passaged HSC. CAT activity, reflecting COL1A1 expression, was upregulated by ␣5␤1-integrin. Inhibition of ␣5␤1-integrin by echistatin and blocking antibody resulted in reduced transgene activity only in early primary cultures (compared with the control, 53.3 Ϯ 12% echistatin and 58.8 Ϯ 7% blocking antibody, respectively, P Ͻ 0.05). Treatment of passaged HSC with either echistatin or blocking antibody had no effect. Fibronectin, an ␣5␤1-integrin ligand, increased transgene activity in primary (210 Ϯ 33%, P Ͻ 0.05) but not in passaged HSC cultures (119 Ϯ 8%). This ␣5␤1-integrin effect appears to be at least in part mediated by CCAAT enhancer binding protein-␤ (C/EBP␤), because fibronectin increased and ␣5-gene silencing by small interfering RNA decreased C/EBP␤ levels. In addition, C/EBP␤ knockout mice showed reduced type I collagen synthesis compared with wild-type littermates. Therefore ␣5␤1-integrin is an important regulator of type I collagen production in early primary HSC cultures but appears to have no direct role once the HSC are fully activated.

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Section: Methodssupporting

confidence: 61%

Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α₅β₁-integrin

Dodig¹,

Ogunwale²,

Dasarathy³

et al. 2007

American Journal of Physiology-Gastrointestinal and Liver Physi

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“…To explain the cellular basis for the reduced mineralization in Psen1 +/− mice, we tracked cells of the osteoblast lineage using reporter mice that expressed GFP during early and late osteoblast progression. We crossed Psen1 +/− mice with Col3.6-GFPtpz or Col2.3-EGFP mice (24), respectively, and lineage tracked GFP + cells in adult mice in vivo, as well as ex vivo in BMSC cultures at 15 and 21 d. Previous studies with Col3.6-GFPtpz mice have shown that the 3.6 kb Col1a1 promoter is extensively expressed, in addition to skin, in osteoblasts lining the periosteal and endosteal trabecular surfaces, with weak expression in periosteal spindle-shaped preosteoblasts (24). In contrast, the 2.3-kb Col1a1 promoter marks mature osteoblasts and osteocytes in Col2.3EGFP mice (24).…”

Section: Resultsmentioning

confidence: 99%

“…We crossed Psen1 +/− mice with Col3.6-GFPtpz or Col2.3-EGFP mice (24), respectively, and lineage tracked GFP + cells in adult mice in vivo, as well as ex vivo in BMSC cultures at 15 and 21 d. Previous studies with Col3.6-GFPtpz mice have shown that the 3.6 kb Col1a1 promoter is extensively expressed, in addition to skin, in osteoblasts lining the periosteal and endosteal trabecular surfaces, with weak expression in periosteal spindle-shaped preosteoblasts (24). In contrast, the 2.3-kb Col1a1 promoter marks mature osteoblasts and osteocytes in Col2.3EGFP mice (24). In differentiating bone marrow stromal cell cultures, GFP expression is seen as early as days 7 and 14, respectively, for Col3.6-GFPtpz and Col2.3EGFP mice (24) (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Of note is that the expression of the 3.2-kb Col1a1 promoter is restricted to mature osteoblasts and osteocytes similarly to the 2.3-kb Col1a1 promoter, but is distinct from that of the Col3.6 promoter. Notably, the 3.2-kb promoter is not active in spindle-shaped periosteal preosteoblasts, where the 3.6-kb promoter is predominantly expressed (24). As such, our premise is that expression of Nicd using the inducible 3.2-kb promoter will closely mimic any effects driven by the 2.3-kb promoter (26,29).…”

Section: Controlled Notch Overexpression In Osteoblast Lineage Cellsmentioning

confidence: 97%

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Anabolic actions of Notch on mature bone

Liu

Ping

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Notch controls skeletogenesis, but its role in the remodeling of adult bone remains conflicting. In mature mice, the skeleton can become osteopenic or osteosclerotic depending on the time point at which Notch is activated or inactivated. Using adult EGFP reporter mice, we find that Notch expression is localized to osteocytes embedded within bone matrix. Conditional activation of Notch signaling in osteocytes triggers profound bone formation, mainly due to increased mineralization, which rescues both age-associated and ovariectomy-induced bone loss and promotes bone healing following osteotomy. In parallel, mice rendered haploinsufficient in γ-secretase presenilin-1 (Psen1), which inhibits downstream Notch activation, display almost-absent terminal osteoblast differentiation. Consistent with this finding, pharmacologic or genetic disruption of Notch or its ligand Jagged1 inhibits mineralization. We suggest that stimulation of Notch signaling in osteocytes initiates a profound, therapeutically relevant, anabolic response.

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“…The 2.3 kb type I collagen promoter is strongly expressed in endosteal osteoblasts and osteocytes, but not in the periosteum, 10,11 whereas the 3.6 kb type I collagen promoter has been shown to be active in the periosteum. 9,12 A full AR knockout has been shown to exhibit increased bone turnover and reduced trabecular and cortical bone volume in male mice. [13][14][15][16] Further elimination of estrogen receptor a (ERa) caused an additional reduction in cortical bone mass.…”

Section: Introductionmentioning

confidence: 99%

Inactivation of the androgen receptor in bone-forming cells leads to trabecular bone loss in adult female mice

et al. 2013

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Removal of the androgen receptor (AR) from bone-forming cells has been shown to reduce trabecular bone volume in male mice. In female mice, the role of AR in the regulation of bone homeostasis has been poorly understood. We generated a mouse strain in which the AR is completely inactivated only in mineralizing osteoblasts and osteocytes by breeding mice carrying osteocalcin promoter-regulated Cre-recombinase with mice possessing loxP recombination sites flanking exon 2 of the AR gene (AR DOB/DOB mice). In female AR DOB/DOB mice, the trabecular bone volume was reduced owing to a smaller number of trabeculae at 6 months of age compared with the control AR fl/fl animals. In male AR DOB/DOB mice, an increase in trabecular bone separation could already be detected at 3.5 months of age, and at 6 months, the trabecular bone volume was significantly reduced compared with that of male AR fl/fl mice. No AR-dependent changes were observed in the cortical bone of either sex. On the basis of micro-computed tomography and histomorphometry, we conclude that in male mice, the AR is involved in the regulation of osteoclast number by osteoblasts, whereas in female mice, the lack of the AR in the bone-forming cells leads to a decreased number of trabeculae upon aging.

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Use of Type I Collagen Green Fluorescent Protein Transgenes to Identify Subpopulations of Cells at Different Stages of the Osteoblast Lineage

Cited by 359 publications

References 51 publications

Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α₅β₁-integrin

Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α₅β₁-integrin

Anabolic actions of Notch on mature bone

Inactivation of the androgen receptor in bone-forming cells leads to trabecular bone loss in adult female mice

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Use of Type I Collagen Green Fluorescent Protein Transgenes to Identify Subpopulations of Cells at Different Stages of the Osteoblast Lineage

Cited by 359 publications

References 51 publications

Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α5β1-integrin

Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α5β1-integrin

Anabolic actions of Notch on mature bone

Inactivation of the androgen receptor in bone-forming cells leads to trabecular bone loss in adult female mice

Contact Info

Product

Resources

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Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α₅β₁-integrin

Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α₅β₁-integrin