Limits...
1,25(OH)2D3 alters growth plate maturation and bone architecture in young rats with normal renal function.

Idelevich A, Kerschnitzki M, Shahar R, Monsonego-Ornan E - PLoS ONE (2011)

Bottom Line: Concomitant with altered chondrocyte maturation, 1,25(OH)(2)D(3) increased chondrocyte proliferation and apoptosis in terminal hypertrophic cells.In vitro treatment of the chondrocytic cell line ATDC5 with 1,25(OH)(2)D(3) lowered differentiation and increased proliferation dose and time-dependently.Micro-CT analysis of femurs from 1-week 1,25(OH)(2)D(3)-treated group revealed reduced cortical thickness, elevated cortical porosity, and higher trabecular number and thickness. 1-month administration resulted in a similar cortical phenotype but without effect on trabecular bone.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biochemistry, Food Science and Nutrition, Hebrew University of Jerusalem, Rehovot, Israel.

ABSTRACT
Whereas detrimental effects of vitamin D deficiency are known over century, the effects of vitamin D receptor activation by 1,25(OH)(2)D(3), the principal hormonal form of vitamin D, on the growing bone and its growth plate are less clear. Currently, 1,25(OH)(2)D(3) is used in pediatric patients with chronic kidney disease and mineral and bone disorder (CKD-MBD) and is strongly associated with growth retardation. Here, we investigate the effect of 1,25(OH)(2)D(3) treatment on bone development in normal young rats, unrelated to renal insufficiency. Young rats received daily i.p. injections of 1 µg/kg 1,25(OH)(2)D(3) for one week, or intermittent 3 µg/kg 1,25(OH)(2)D(3) for one month. Histological analysis revealed narrower tibial growth plates, predominantly in the hypertrophic zone of 1,25(OH)(2)D(3)-treated animals in both experimental protocols. This phenotype was supported by narrower distribution of aggrecan, collagens II and X mRNA, shown by in situ hybridization. Concomitant with altered chondrocyte maturation, 1,25(OH)(2)D(3) increased chondrocyte proliferation and apoptosis in terminal hypertrophic cells. In vitro treatment of the chondrocytic cell line ATDC5 with 1,25(OH)(2)D(3) lowered differentiation and increased proliferation dose and time-dependently. Micro-CT analysis of femurs from 1-week 1,25(OH)(2)D(3)-treated group revealed reduced cortical thickness, elevated cortical porosity, and higher trabecular number and thickness. 1-month administration resulted in a similar cortical phenotype but without effect on trabecular bone. Evaluation of fluorochrome binding with confocal microscopy revealed inhibiting effects of 1,25(OH)(2)D(3) on intracortical bone formation. This study shows negative effects of 1,25(OH)(2)D(3) on growth plate and bone which may contribute to the exacerbation of MBD in the CKD pediatric patients.

Show MeSH

Related in: MedlinePlus

1-week continuous administration of 1,25(OH)2D3 induces chondrocyte proliferation and compresses chondrocytes maturation zone in young rats.4 weeks old male Spague Dawley rats received daily i.p. injections of 1 µg/kg 1,25(OH)2D3 (n = 6, VD) or vehicle (n = 6, Ctr) for a period of 7 days. (A) Safranin O staining of proximal tibial growth plates. Arrows indicate reduced hypertrophic zone width in 1,25(OH)2D3-treated compared to the control group. (B) Width measurements of total growth plate, proliferative zone, and hypertrophic zone. (C) Percent of PCNA positive proliferating cells out of total proliferating cell count. (D) Immunhistochemistry of PCNA positive proliferating cells. (E) In-situ hybridization (ISH) of collagen type II mRNA. (F) ISH of aggrecan mRNA. (G) ISH of collagen type X mRNA. (H) TUNEL detection of apoptotic cells, indicated by white arrows. Cell nuclei were counterstained with DAPI. (I) TRAP staining for chondroclasts, indicated by arrows. In all studies, results are shown as means (n = 6) ± SD. * denotes p<0.05 comparing 1,25(OH)2D3-treated to control groups. Open squares in the images on the left represent areas magnified in the images on the right. Abbreviations: PZ-proliferative zone, HZ-hypertrophic zone.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3113808&req=5

pone-0020772-g001: 1-week continuous administration of 1,25(OH)2D3 induces chondrocyte proliferation and compresses chondrocytes maturation zone in young rats.4 weeks old male Spague Dawley rats received daily i.p. injections of 1 µg/kg 1,25(OH)2D3 (n = 6, VD) or vehicle (n = 6, Ctr) for a period of 7 days. (A) Safranin O staining of proximal tibial growth plates. Arrows indicate reduced hypertrophic zone width in 1,25(OH)2D3-treated compared to the control group. (B) Width measurements of total growth plate, proliferative zone, and hypertrophic zone. (C) Percent of PCNA positive proliferating cells out of total proliferating cell count. (D) Immunhistochemistry of PCNA positive proliferating cells. (E) In-situ hybridization (ISH) of collagen type II mRNA. (F) ISH of aggrecan mRNA. (G) ISH of collagen type X mRNA. (H) TUNEL detection of apoptotic cells, indicated by white arrows. Cell nuclei were counterstained with DAPI. (I) TRAP staining for chondroclasts, indicated by arrows. In all studies, results are shown as means (n = 6) ± SD. * denotes p<0.05 comparing 1,25(OH)2D3-treated to control groups. Open squares in the images on the left represent areas magnified in the images on the right. Abbreviations: PZ-proliferative zone, HZ-hypertrophic zone.

Mentions: Previously, VDR activation was shown to affect growth plate development [18], [19], and induce vascular calcification [30], in rats with chronic renal damage. To assess whether pharmacological administration of 1,25(OH)2D3 influences bone formation in young rats with normal renal function we established two regiments where 1,25(OH)2D3, was administered to young rats for a period of one week or one month. 1-week and 1-month treatment with 1,25(OH)2D3 induced vascular calcification, as evident by intense Alizarin Red staining of the aortae, compared to control (Fig S1A,B). Staining of tibiae with Safranin O (stain for cartilage) revealed marked reduction in the total growth plate width in both experiments (Fig. 1A, 2A). Measurement of individual zones showed that this effect is attributed predominantly to the narrowing of the hypertrophic zone of the growth plate, since the proliferative zone width remained unchanged (Fig. 1B, 2B). Hypertrophic zone width was reduced by 47% after 1-week 1,25(OH)2D3 treatment and 32% after 1-month treatment compared to the relevant controls. While continuous 1-week 1,25(OH)2D3 administration did not significantly alter the morphological appearance of chondrocytes, intermittent 1-month treatment resulted in disordered chondrocyte morphology, noticeably disrupting the columnar organization of the growth plate (Fig. 2A,E). Growth plate phenotypes were not manifested in significant tibial length differences (and final weight) between experimental groups, suggesting presence of compensating mechanisms which regulate longitudinal growth in this model.


1,25(OH)2D3 alters growth plate maturation and bone architecture in young rats with normal renal function.

Idelevich A, Kerschnitzki M, Shahar R, Monsonego-Ornan E - PLoS ONE (2011)

1-week continuous administration of 1,25(OH)2D3 induces chondrocyte proliferation and compresses chondrocytes maturation zone in young rats.4 weeks old male Spague Dawley rats received daily i.p. injections of 1 µg/kg 1,25(OH)2D3 (n = 6, VD) or vehicle (n = 6, Ctr) for a period of 7 days. (A) Safranin O staining of proximal tibial growth plates. Arrows indicate reduced hypertrophic zone width in 1,25(OH)2D3-treated compared to the control group. (B) Width measurements of total growth plate, proliferative zone, and hypertrophic zone. (C) Percent of PCNA positive proliferating cells out of total proliferating cell count. (D) Immunhistochemistry of PCNA positive proliferating cells. (E) In-situ hybridization (ISH) of collagen type II mRNA. (F) ISH of aggrecan mRNA. (G) ISH of collagen type X mRNA. (H) TUNEL detection of apoptotic cells, indicated by white arrows. Cell nuclei were counterstained with DAPI. (I) TRAP staining for chondroclasts, indicated by arrows. In all studies, results are shown as means (n = 6) ± SD. * denotes p<0.05 comparing 1,25(OH)2D3-treated to control groups. Open squares in the images on the left represent areas magnified in the images on the right. Abbreviations: PZ-proliferative zone, HZ-hypertrophic zone.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3113808&req=5

pone-0020772-g001: 1-week continuous administration of 1,25(OH)2D3 induces chondrocyte proliferation and compresses chondrocytes maturation zone in young rats.4 weeks old male Spague Dawley rats received daily i.p. injections of 1 µg/kg 1,25(OH)2D3 (n = 6, VD) or vehicle (n = 6, Ctr) for a period of 7 days. (A) Safranin O staining of proximal tibial growth plates. Arrows indicate reduced hypertrophic zone width in 1,25(OH)2D3-treated compared to the control group. (B) Width measurements of total growth plate, proliferative zone, and hypertrophic zone. (C) Percent of PCNA positive proliferating cells out of total proliferating cell count. (D) Immunhistochemistry of PCNA positive proliferating cells. (E) In-situ hybridization (ISH) of collagen type II mRNA. (F) ISH of aggrecan mRNA. (G) ISH of collagen type X mRNA. (H) TUNEL detection of apoptotic cells, indicated by white arrows. Cell nuclei were counterstained with DAPI. (I) TRAP staining for chondroclasts, indicated by arrows. In all studies, results are shown as means (n = 6) ± SD. * denotes p<0.05 comparing 1,25(OH)2D3-treated to control groups. Open squares in the images on the left represent areas magnified in the images on the right. Abbreviations: PZ-proliferative zone, HZ-hypertrophic zone.
Mentions: Previously, VDR activation was shown to affect growth plate development [18], [19], and induce vascular calcification [30], in rats with chronic renal damage. To assess whether pharmacological administration of 1,25(OH)2D3 influences bone formation in young rats with normal renal function we established two regiments where 1,25(OH)2D3, was administered to young rats for a period of one week or one month. 1-week and 1-month treatment with 1,25(OH)2D3 induced vascular calcification, as evident by intense Alizarin Red staining of the aortae, compared to control (Fig S1A,B). Staining of tibiae with Safranin O (stain for cartilage) revealed marked reduction in the total growth plate width in both experiments (Fig. 1A, 2A). Measurement of individual zones showed that this effect is attributed predominantly to the narrowing of the hypertrophic zone of the growth plate, since the proliferative zone width remained unchanged (Fig. 1B, 2B). Hypertrophic zone width was reduced by 47% after 1-week 1,25(OH)2D3 treatment and 32% after 1-month treatment compared to the relevant controls. While continuous 1-week 1,25(OH)2D3 administration did not significantly alter the morphological appearance of chondrocytes, intermittent 1-month treatment resulted in disordered chondrocyte morphology, noticeably disrupting the columnar organization of the growth plate (Fig. 2A,E). Growth plate phenotypes were not manifested in significant tibial length differences (and final weight) between experimental groups, suggesting presence of compensating mechanisms which regulate longitudinal growth in this model.

Bottom Line: Concomitant with altered chondrocyte maturation, 1,25(OH)(2)D(3) increased chondrocyte proliferation and apoptosis in terminal hypertrophic cells.In vitro treatment of the chondrocytic cell line ATDC5 with 1,25(OH)(2)D(3) lowered differentiation and increased proliferation dose and time-dependently.Micro-CT analysis of femurs from 1-week 1,25(OH)(2)D(3)-treated group revealed reduced cortical thickness, elevated cortical porosity, and higher trabecular number and thickness. 1-month administration resulted in a similar cortical phenotype but without effect on trabecular bone.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biochemistry, Food Science and Nutrition, Hebrew University of Jerusalem, Rehovot, Israel.

ABSTRACT
Whereas detrimental effects of vitamin D deficiency are known over century, the effects of vitamin D receptor activation by 1,25(OH)(2)D(3), the principal hormonal form of vitamin D, on the growing bone and its growth plate are less clear. Currently, 1,25(OH)(2)D(3) is used in pediatric patients with chronic kidney disease and mineral and bone disorder (CKD-MBD) and is strongly associated with growth retardation. Here, we investigate the effect of 1,25(OH)(2)D(3) treatment on bone development in normal young rats, unrelated to renal insufficiency. Young rats received daily i.p. injections of 1 µg/kg 1,25(OH)(2)D(3) for one week, or intermittent 3 µg/kg 1,25(OH)(2)D(3) for one month. Histological analysis revealed narrower tibial growth plates, predominantly in the hypertrophic zone of 1,25(OH)(2)D(3)-treated animals in both experimental protocols. This phenotype was supported by narrower distribution of aggrecan, collagens II and X mRNA, shown by in situ hybridization. Concomitant with altered chondrocyte maturation, 1,25(OH)(2)D(3) increased chondrocyte proliferation and apoptosis in terminal hypertrophic cells. In vitro treatment of the chondrocytic cell line ATDC5 with 1,25(OH)(2)D(3) lowered differentiation and increased proliferation dose and time-dependently. Micro-CT analysis of femurs from 1-week 1,25(OH)(2)D(3)-treated group revealed reduced cortical thickness, elevated cortical porosity, and higher trabecular number and thickness. 1-month administration resulted in a similar cortical phenotype but without effect on trabecular bone. Evaluation of fluorochrome binding with confocal microscopy revealed inhibiting effects of 1,25(OH)(2)D(3) on intracortical bone formation. This study shows negative effects of 1,25(OH)(2)D(3) on growth plate and bone which may contribute to the exacerbation of MBD in the CKD pediatric patients.

Show MeSH
Related in: MedlinePlus