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Bone growth during rapamycin therapy in young rats.

Sanchez CP, He YZ - BMC Pediatr (2009)

Bottom Line: Although body and tibial length remained short after 4 weeks of rapamycin, changes in the expression of chondrocyte proliferation, chondrocyte differentiation and chondro/osteoclastic resorption which were significant after 2 weeks of rapamycin improved at the end of 4 weeks.When given to young rats, 2 weeks of rapamycin significantly decreased endochondral bone growth.No catch-up growth was demonstrated at the end of 4 weeks, although markers of chondrocyte proliferation and differentiation improved.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pediatrics, University of Wisconsin School of Medicine & Public Health, Madison, Wisconsin, USA. cpsanchez@pediatrics.wisc.edu

ABSTRACT

Background: Rapamycin is an effective immunosuppressant widely used to maintain the renal allograft in pediatric patients. Linear growth may be adversely affected in young children since rapamycin has potent anti-proliferative and anti-angiogenic properties.

Methods: Weanling three week old rats were given rapamycin at 2.5 mg/kg daily by gavage for 2 or 4 weeks and compared to a Control group given equivalent amount of saline. Morphometric measurements and biochemical determinations for serum calcium, phosphate, iPTH, urea nitrogen, creatinine and insulin-growth factor I (IGF-I) were obtained. Histomorphometric analysis of the growth plate cartilage, in-situ hybridization experiments and immunohistochemical studies for various proteins were performed to evaluate for chondrocyte proliferation, chondrocyte differentiation and chondro/osteoclastic resorption.

Results: At the end of the 2 weeks, body and tibia length measurements were shorter after rapamycin therapy associated with an enlargement of the hypertrophic zone in the growth plate cartilage. There was a decrease in chondrocyte proliferation assessed by histone-4 and mammalian target of rapamycin (mTOR) expression. A reduction in parathyroid hormone/parathyroid hormone related peptide (PTH/PTHrP) and an increase in Indian hedgehog (Ihh) expression may explain in part, the increase number of hypertrophic chondrocytes. The number of TRAP positive multinucleated chondro/osteoclasts declined in the chondro-osseous junction with a decrease in the receptor activator of nuclear factor kappa beta ligand (RANKL) and vascular endothelial growth factor (VEGF) expression. Although body and tibial length remained short after 4 weeks of rapamycin, changes in the expression of chondrocyte proliferation, chondrocyte differentiation and chondro/osteoclastic resorption which were significant after 2 weeks of rapamycin improved at the end of 4 weeks.

Conclusion: When given to young rats, 2 weeks of rapamycin significantly decreased endochondral bone growth. No catch-up growth was demonstrated at the end of 4 weeks, although markers of chondrocyte proliferation and differentiation improved. Clinical studies need to be done to evaluate these changes in growing children.

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The panels on the left show representative photomicrographs of IGF-I protein expression (4A), 50× (denoted by the brown color and arrows); ap < 0.04 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.03 Rapamycin (2 weeks) vs Rapamycin (4 weeks); cp < 0.03 Control (2 weeks) vs Control (4 weeks). IGFBP3 protein expression (4B, denoted by arrows and brown color), 50×; ap < 0.002 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.03 Rapamycin (2 weeks) vs Rapamycin (4 weeks). Type × collagen (col10a) protein expression localized only to the hypertrophic chondrocytes (4C, denoted by purple color and brackets), 20×; ap < 0.005 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.02 Rapamycin (4 weeks) vs Control (4 weeks). The panels on the right show the quantification of the protein expression for IGF-I (upper panel); IGFBP3 (middle panel); and type × collagen (lower panel) after 2 weeks and 4 weeks in Rapamycin and Control groups expressed as number of labeled cells to the total number of cells in the appropriate zone (Labeling Index).
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Figure 4: The panels on the left show representative photomicrographs of IGF-I protein expression (4A), 50× (denoted by the brown color and arrows); ap < 0.04 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.03 Rapamycin (2 weeks) vs Rapamycin (4 weeks); cp < 0.03 Control (2 weeks) vs Control (4 weeks). IGFBP3 protein expression (4B, denoted by arrows and brown color), 50×; ap < 0.002 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.03 Rapamycin (2 weeks) vs Rapamycin (4 weeks). Type × collagen (col10a) protein expression localized only to the hypertrophic chondrocytes (4C, denoted by purple color and brackets), 20×; ap < 0.005 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.02 Rapamycin (4 weeks) vs Control (4 weeks). The panels on the right show the quantification of the protein expression for IGF-I (upper panel); IGFBP3 (middle panel); and type × collagen (lower panel) after 2 weeks and 4 weeks in Rapamycin and Control groups expressed as number of labeled cells to the total number of cells in the appropriate zone (Labeling Index).

Mentions: The protein expression of IGF-I which was restricted to the hypertrophic chondrocytes decreased after 2 weeks of rapamycin compared to Control (Figure 4A). In agreement with other published studies, IGF-I staining was 20 percent lower in the 2 weeks Control animals compared to 4 weeks Control (Figure 4A). IGF-II and not IGF-I has been demonstrated to be more abundant in younger animals and that IGF-I may be associated with chondrocyte hypertrophy and mineralization [17]. The expression of IGF-II was not assessed in the current study.


Bone growth during rapamycin therapy in young rats.

Sanchez CP, He YZ - BMC Pediatr (2009)

The panels on the left show representative photomicrographs of IGF-I protein expression (4A), 50× (denoted by the brown color and arrows); ap < 0.04 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.03 Rapamycin (2 weeks) vs Rapamycin (4 weeks); cp < 0.03 Control (2 weeks) vs Control (4 weeks). IGFBP3 protein expression (4B, denoted by arrows and brown color), 50×; ap < 0.002 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.03 Rapamycin (2 weeks) vs Rapamycin (4 weeks). Type × collagen (col10a) protein expression localized only to the hypertrophic chondrocytes (4C, denoted by purple color and brackets), 20×; ap < 0.005 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.02 Rapamycin (4 weeks) vs Control (4 weeks). The panels on the right show the quantification of the protein expression for IGF-I (upper panel); IGFBP3 (middle panel); and type × collagen (lower panel) after 2 weeks and 4 weeks in Rapamycin and Control groups expressed as number of labeled cells to the total number of cells in the appropriate zone (Labeling Index).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2629763&req=5

Figure 4: The panels on the left show representative photomicrographs of IGF-I protein expression (4A), 50× (denoted by the brown color and arrows); ap < 0.04 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.03 Rapamycin (2 weeks) vs Rapamycin (4 weeks); cp < 0.03 Control (2 weeks) vs Control (4 weeks). IGFBP3 protein expression (4B, denoted by arrows and brown color), 50×; ap < 0.002 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.03 Rapamycin (2 weeks) vs Rapamycin (4 weeks). Type × collagen (col10a) protein expression localized only to the hypertrophic chondrocytes (4C, denoted by purple color and brackets), 20×; ap < 0.005 Rapamycin (2 weeks) vs Control (2 weeks); bp < 0.02 Rapamycin (4 weeks) vs Control (4 weeks). The panels on the right show the quantification of the protein expression for IGF-I (upper panel); IGFBP3 (middle panel); and type × collagen (lower panel) after 2 weeks and 4 weeks in Rapamycin and Control groups expressed as number of labeled cells to the total number of cells in the appropriate zone (Labeling Index).
Mentions: The protein expression of IGF-I which was restricted to the hypertrophic chondrocytes decreased after 2 weeks of rapamycin compared to Control (Figure 4A). In agreement with other published studies, IGF-I staining was 20 percent lower in the 2 weeks Control animals compared to 4 weeks Control (Figure 4A). IGF-II and not IGF-I has been demonstrated to be more abundant in younger animals and that IGF-I may be associated with chondrocyte hypertrophy and mineralization [17]. The expression of IGF-II was not assessed in the current study.

Bottom Line: Although body and tibial length remained short after 4 weeks of rapamycin, changes in the expression of chondrocyte proliferation, chondrocyte differentiation and chondro/osteoclastic resorption which were significant after 2 weeks of rapamycin improved at the end of 4 weeks.When given to young rats, 2 weeks of rapamycin significantly decreased endochondral bone growth.No catch-up growth was demonstrated at the end of 4 weeks, although markers of chondrocyte proliferation and differentiation improved.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pediatrics, University of Wisconsin School of Medicine & Public Health, Madison, Wisconsin, USA. cpsanchez@pediatrics.wisc.edu

ABSTRACT

Background: Rapamycin is an effective immunosuppressant widely used to maintain the renal allograft in pediatric patients. Linear growth may be adversely affected in young children since rapamycin has potent anti-proliferative and anti-angiogenic properties.

Methods: Weanling three week old rats were given rapamycin at 2.5 mg/kg daily by gavage for 2 or 4 weeks and compared to a Control group given equivalent amount of saline. Morphometric measurements and biochemical determinations for serum calcium, phosphate, iPTH, urea nitrogen, creatinine and insulin-growth factor I (IGF-I) were obtained. Histomorphometric analysis of the growth plate cartilage, in-situ hybridization experiments and immunohistochemical studies for various proteins were performed to evaluate for chondrocyte proliferation, chondrocyte differentiation and chondro/osteoclastic resorption.

Results: At the end of the 2 weeks, body and tibia length measurements were shorter after rapamycin therapy associated with an enlargement of the hypertrophic zone in the growth plate cartilage. There was a decrease in chondrocyte proliferation assessed by histone-4 and mammalian target of rapamycin (mTOR) expression. A reduction in parathyroid hormone/parathyroid hormone related peptide (PTH/PTHrP) and an increase in Indian hedgehog (Ihh) expression may explain in part, the increase number of hypertrophic chondrocytes. The number of TRAP positive multinucleated chondro/osteoclasts declined in the chondro-osseous junction with a decrease in the receptor activator of nuclear factor kappa beta ligand (RANKL) and vascular endothelial growth factor (VEGF) expression. Although body and tibial length remained short after 4 weeks of rapamycin, changes in the expression of chondrocyte proliferation, chondrocyte differentiation and chondro/osteoclastic resorption which were significant after 2 weeks of rapamycin improved at the end of 4 weeks.

Conclusion: When given to young rats, 2 weeks of rapamycin significantly decreased endochondral bone growth. No catch-up growth was demonstrated at the end of 4 weeks, although markers of chondrocyte proliferation and differentiation improved. Clinical studies need to be done to evaluate these changes in growing children.

Show MeSH
Related in: MedlinePlus