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Insulin exerts direct, IGF-1 independent actions in growth plate chondrocytes.

Zhang F, He Q, Tsang WP, Garvey WT, Chan WY, Wan C - Bone Res (2014)

Bottom Line: Subsequently, IGF-1 induced phosphorylation of Akt and ERK was enhanced, while this action was eliminated when the cells were treated with IGF-1R inhibitor Picropodophyllin.Deletion of the IR impaired chondrogenic differentiation, and the effect could not be restored by treatment of insulin, but partially rescued by IGF-1 treatment.These results suggest that deletion of the IR in chondrocytes sensitizes IGF-1R signaling and action, IR and IGF-1R coordinate to regulate the proliferation, differentiation and hypertrophy of growth plate chondrocytes.

View Article: PubMed Central - PubMed

Affiliation: Ministry of Education Key Laboratory for Regenerative Medicine, School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong , Hong Kong SAR, China ; School of Biomedical Sciences Core Laboratory, Institute of Stem Cell, Genomics and Translational Research, Shenzhen Research Institute, The Chinese University of Hong Kong , Shenzhen, China.

ABSTRACT
Insufficient insulin production or action in diabetic states is associated with growth retardation and impaired bone healing, while the underling mechanisms are unknown. In this study, we sought to define the role of insulin signaling in the growth plate. Insulin treatment of embryonic metatarsal bones from wild-type mice increased chondrocyte proliferation. Mice lacking insulin receptor (IR) selectively in chondrocytes (CartIR (-/-)) had no discernable differences in total femoral length compared to control littermates. However, CartIR (-/-) mice exhibited an increase in chondrocyte numbers in the growth plate than that of the controls. Chondrocytes lacking IR had elevated insulin-like growth factor (IGF)-1R mRNA and protein levels. Subsequently, IGF-1 induced phosphorylation of Akt and ERK was enhanced, while this action was eliminated when the cells were treated with IGF-1R inhibitor Picropodophyllin. Deletion of the IR impaired chondrogenic differentiation, and the effect could not be restored by treatment of insulin, but partially rescued by IGF-1 treatment. Intriguingly, the size of hypertrophic chondrocytes was smaller in CartIR (-/-) mice when compared with that of the control littermates, which was associated with upregulation of tuberous sclerosis complex 2 (TSC2). These results suggest that deletion of the IR in chondrocytes sensitizes IGF-1R signaling and action, IR and IGF-1R coordinate to regulate the proliferation, differentiation and hypertrophy of growth plate chondrocytes.

No MeSH data available.


Related in: MedlinePlus

Insulin promotes growth of fetal metatarsal bones in vitro. (a) Metatarsalbones from E17.5 wild-type mouse embryos were cultured for 6 days in the absence or presence of insulin or IGF-1. (b) Quantitation of the actual length of metatarsal bones from (a) as shown. *P<0.05, n=6. (c) Representative histological images of metatarsal bones treated with insulin or IGF-1, with none treatment as control. PZ, proliferating zone; HZ, hypertrophic zone; MZ, mineralizing zone. (d) Quantitation of the proliferating zone of the cartilage rudiments from (c). *P<0.05, n=6. (e) Representative histological sections of metatarsal bones treated with insulin or IGF-1 after staining with antibody against PCNA as described in the section on ‘Materials and methods’. Sections were counterstained with hematoxylin. Scale bar=50 µm. (f) Quantitation for the percentage of PCNA+ cells from (e). *P<0.05, n=6. IGF-1 treatment shows more dramatic effects on the total length in the cultures as positive control. CON, none treatmentcontrol; IGF-1, insulin-like growth factor-1; INS, insulin.
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fig1: Insulin promotes growth of fetal metatarsal bones in vitro. (a) Metatarsalbones from E17.5 wild-type mouse embryos were cultured for 6 days in the absence or presence of insulin or IGF-1. (b) Quantitation of the actual length of metatarsal bones from (a) as shown. *P<0.05, n=6. (c) Representative histological images of metatarsal bones treated with insulin or IGF-1, with none treatment as control. PZ, proliferating zone; HZ, hypertrophic zone; MZ, mineralizing zone. (d) Quantitation of the proliferating zone of the cartilage rudiments from (c). *P<0.05, n=6. (e) Representative histological sections of metatarsal bones treated with insulin or IGF-1 after staining with antibody against PCNA as described in the section on ‘Materials and methods’. Sections were counterstained with hematoxylin. Scale bar=50 µm. (f) Quantitation for the percentage of PCNA+ cells from (e). *P<0.05, n=6. IGF-1 treatment shows more dramatic effects on the total length in the cultures as positive control. CON, none treatmentcontrol; IGF-1, insulin-like growth factor-1; INS, insulin.

Mentions: To begin to examine the effects of insulin on chondrocytes, we determined the effects of insulin and IGF-1 on the growth of fetal metatarsal rudiments in vitro. Metatarsal bones from E17.5 wild-type mouse embryos were cultured for 6 days in the absence or presence of 10 nmol⋅L−1 insulin or 13 nmol⋅L−1 IGF-1. Both ligands significantly increased the total length (Figure 1a and 1b) primarily by increasing the lengths and numbers of cells in the proliferating zone (Figure 1c and 1d). Immunostaining for PCNA also showed that treatment with insulin and IGF-1 increased PCNA positive cell numbers in the cultured metatarsal bone sections (Figure 1e and 1f). These data demonstrate that insulin exerts mitogenic effects in growth plate chondrocytes that are qualitatively similar to those of IGF-1.


Insulin exerts direct, IGF-1 independent actions in growth plate chondrocytes.

Zhang F, He Q, Tsang WP, Garvey WT, Chan WY, Wan C - Bone Res (2014)

Insulin promotes growth of fetal metatarsal bones in vitro. (a) Metatarsalbones from E17.5 wild-type mouse embryos were cultured for 6 days in the absence or presence of insulin or IGF-1. (b) Quantitation of the actual length of metatarsal bones from (a) as shown. *P<0.05, n=6. (c) Representative histological images of metatarsal bones treated with insulin or IGF-1, with none treatment as control. PZ, proliferating zone; HZ, hypertrophic zone; MZ, mineralizing zone. (d) Quantitation of the proliferating zone of the cartilage rudiments from (c). *P<0.05, n=6. (e) Representative histological sections of metatarsal bones treated with insulin or IGF-1 after staining with antibody against PCNA as described in the section on ‘Materials and methods’. Sections were counterstained with hematoxylin. Scale bar=50 µm. (f) Quantitation for the percentage of PCNA+ cells from (e). *P<0.05, n=6. IGF-1 treatment shows more dramatic effects on the total length in the cultures as positive control. CON, none treatmentcontrol; IGF-1, insulin-like growth factor-1; INS, insulin.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4472128&req=5

fig1: Insulin promotes growth of fetal metatarsal bones in vitro. (a) Metatarsalbones from E17.5 wild-type mouse embryos were cultured for 6 days in the absence or presence of insulin or IGF-1. (b) Quantitation of the actual length of metatarsal bones from (a) as shown. *P<0.05, n=6. (c) Representative histological images of metatarsal bones treated with insulin or IGF-1, with none treatment as control. PZ, proliferating zone; HZ, hypertrophic zone; MZ, mineralizing zone. (d) Quantitation of the proliferating zone of the cartilage rudiments from (c). *P<0.05, n=6. (e) Representative histological sections of metatarsal bones treated with insulin or IGF-1 after staining with antibody against PCNA as described in the section on ‘Materials and methods’. Sections were counterstained with hematoxylin. Scale bar=50 µm. (f) Quantitation for the percentage of PCNA+ cells from (e). *P<0.05, n=6. IGF-1 treatment shows more dramatic effects on the total length in the cultures as positive control. CON, none treatmentcontrol; IGF-1, insulin-like growth factor-1; INS, insulin.
Mentions: To begin to examine the effects of insulin on chondrocytes, we determined the effects of insulin and IGF-1 on the growth of fetal metatarsal rudiments in vitro. Metatarsal bones from E17.5 wild-type mouse embryos were cultured for 6 days in the absence or presence of 10 nmol⋅L−1 insulin or 13 nmol⋅L−1 IGF-1. Both ligands significantly increased the total length (Figure 1a and 1b) primarily by increasing the lengths and numbers of cells in the proliferating zone (Figure 1c and 1d). Immunostaining for PCNA also showed that treatment with insulin and IGF-1 increased PCNA positive cell numbers in the cultured metatarsal bone sections (Figure 1e and 1f). These data demonstrate that insulin exerts mitogenic effects in growth plate chondrocytes that are qualitatively similar to those of IGF-1.

Bottom Line: Subsequently, IGF-1 induced phosphorylation of Akt and ERK was enhanced, while this action was eliminated when the cells were treated with IGF-1R inhibitor Picropodophyllin.Deletion of the IR impaired chondrogenic differentiation, and the effect could not be restored by treatment of insulin, but partially rescued by IGF-1 treatment.These results suggest that deletion of the IR in chondrocytes sensitizes IGF-1R signaling and action, IR and IGF-1R coordinate to regulate the proliferation, differentiation and hypertrophy of growth plate chondrocytes.

View Article: PubMed Central - PubMed

Affiliation: Ministry of Education Key Laboratory for Regenerative Medicine, School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong , Hong Kong SAR, China ; School of Biomedical Sciences Core Laboratory, Institute of Stem Cell, Genomics and Translational Research, Shenzhen Research Institute, The Chinese University of Hong Kong , Shenzhen, China.

ABSTRACT
Insufficient insulin production or action in diabetic states is associated with growth retardation and impaired bone healing, while the underling mechanisms are unknown. In this study, we sought to define the role of insulin signaling in the growth plate. Insulin treatment of embryonic metatarsal bones from wild-type mice increased chondrocyte proliferation. Mice lacking insulin receptor (IR) selectively in chondrocytes (CartIR (-/-)) had no discernable differences in total femoral length compared to control littermates. However, CartIR (-/-) mice exhibited an increase in chondrocyte numbers in the growth plate than that of the controls. Chondrocytes lacking IR had elevated insulin-like growth factor (IGF)-1R mRNA and protein levels. Subsequently, IGF-1 induced phosphorylation of Akt and ERK was enhanced, while this action was eliminated when the cells were treated with IGF-1R inhibitor Picropodophyllin. Deletion of the IR impaired chondrogenic differentiation, and the effect could not be restored by treatment of insulin, but partially rescued by IGF-1 treatment. Intriguingly, the size of hypertrophic chondrocytes was smaller in CartIR (-/-) mice when compared with that of the control littermates, which was associated with upregulation of tuberous sclerosis complex 2 (TSC2). These results suggest that deletion of the IR in chondrocytes sensitizes IGF-1R signaling and action, IR and IGF-1R coordinate to regulate the proliferation, differentiation and hypertrophy of growth plate chondrocytes.

No MeSH data available.


Related in: MedlinePlus