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Pomegranate Peel Extract Prevents Bone Loss in a Preclinical Model of Osteoporosis and Stimulates Osteoblastic Differentiation in Vitro.

Spilmont M, Léotoing L, Davicco MJ, Lebecque P, Miot-Noirault E, Pilet P, Rios L, Wittrant Y, Coxam V - Nutrients (2015)

Bottom Line: The nutritional benefits of pomegranate have attracted great scientific interest.Among chronic diseases, osteoporosis, which is associated with bone remodelling impairment leading to progressive bone loss, could eventually benefit from antioxidant compounds because of the involvement of oxidative stress in the pathogenesis of osteopenia.In addition, PGPE appeared to substantially stimulate osteoblastic MC3T3-E1 alkaline phosphatase (ALP) activity at day 7, mineralization at day 21 and the transcription level of osteogenic markers.

View Article: PubMed Central - PubMed

Affiliation: Unité de Nutrition Humaine, CRNH Auvergne, UMR 1019, INRA, F-63000 Clermont-Ferrand, France. mel.spilmont@gmail.com.

ABSTRACT
The nutritional benefits of pomegranate have attracted great scientific interest. The pomegranate, including the pomegranate peel, has been used worldwide for many years as a fruit with medicinal activity, mostly antioxidant properties. Among chronic diseases, osteoporosis, which is associated with bone remodelling impairment leading to progressive bone loss, could eventually benefit from antioxidant compounds because of the involvement of oxidative stress in the pathogenesis of osteopenia. In this study, with in vivo and ex vivo experiments, we investigated whether the consumption of pomegranate peel extract (PGPE) could limit the process of osteopenia. We demonstrated that in ovariectomized (OVX) C57BL/6J mice, PGPE consumption was able to significantly prevent the decrease in bone mineral density (-31.9%; p < 0.001 vs. OVX mice) and bone microarchitecture impairment. Moreover, the exposure of RAW264.7 cells to serum harvested from mice that had been given a PGPE-enriched diet elicited reduced osteoclast differentiation and bone resorption, as shown by the inhibition of the major osteoclast markers. In addition, PGPE appeared to substantially stimulate osteoblastic MC3T3-E1 alkaline phosphatase (ALP) activity at day 7, mineralization at day 21 and the transcription level of osteogenic markers. PGPE may be effective in preventing the bone loss associated with ovariectomy in mice, and offers a promising alternative for the nutritional management of this disease.

No MeSH data available.


Related in: MedlinePlus

Osteoblast viability, differentiation and transcription in culture, after exposure to serum harvested from mice raised either under control conditions or treated with pomegranate peel extract. * p < 0.05 PGPE versus Control condition. (A) Cell proliferation test performed on MC3T3-E1 cells cultured in minimal medium (2% Fetal Bovine Serum, FBS), supplemented with serum from mice given either physiological serum (1.25% FBS + 0.75 Control mice serum) or pomegranate peel extract, PGPE (1.25% FBS + 0.75 PGPE mice serum), after 2 and 7 days of treatment. Results are expressed as mean ± standard error of the mean (SEM) (n = 6). (B) Alkaline phosphatase, ALP activity of MC3T3-E1 cells cultured in optimized medium with 10% FBS or with 7.5% FBS + 2.5% serum from mice fed either physiological serum (Control) or pomegranate peel extract (PGPE), after 0, 2, 7 and 14 days of treatment. Results are expressed as mean ± SEM (n = 8). (C,D) Mineralized nodules stained with alizarin red S on Day 21 in MC3T3-E1 cells cultured in optimized (C+) medium with 10% FBS or with 7.5% FBS + 2.5% serum from mice exposed to either physiological serum (Control) or pomegranate peel extract (PGPE). (C) Representative image of alizarin red staining for each condition. (D) Relative quantification of alizarin red staining with picture analyser software ImageJ. Results are presented as mean ± standard deviation, SD (n = 6). (E) Transcriptomic analysis of the main osteoblast markers in MC3T3-E1 cells cultured in optimized medium after 7 days of treatment with serum from mice exposed to either physiological serum (Control) or pomegranate peel extract (PGPE). Transcriptomic analysis of MC3T3-E1 mRNA levels determined by Taqman Low-Density Arrays are presented as fold change compared to the Control condition (fold change = 1) as mean ± SD (n = 6). Genes: ALP: alkaline phosphatase; βCat: β-catenin;Coll1a1: collagen 1a1; Coll2a1: collagen 2a1; DDR2: discoidin domain receptor 2; BSP: bone sialoprotein; Lrp5; OCN: osteocalcin; OPG: osteoprotegerin; osteri RunX2 and Wnt.
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nutrients-07-05465-f002: Osteoblast viability, differentiation and transcription in culture, after exposure to serum harvested from mice raised either under control conditions or treated with pomegranate peel extract. * p < 0.05 PGPE versus Control condition. (A) Cell proliferation test performed on MC3T3-E1 cells cultured in minimal medium (2% Fetal Bovine Serum, FBS), supplemented with serum from mice given either physiological serum (1.25% FBS + 0.75 Control mice serum) or pomegranate peel extract, PGPE (1.25% FBS + 0.75 PGPE mice serum), after 2 and 7 days of treatment. Results are expressed as mean ± standard error of the mean (SEM) (n = 6). (B) Alkaline phosphatase, ALP activity of MC3T3-E1 cells cultured in optimized medium with 10% FBS or with 7.5% FBS + 2.5% serum from mice fed either physiological serum (Control) or pomegranate peel extract (PGPE), after 0, 2, 7 and 14 days of treatment. Results are expressed as mean ± SEM (n = 8). (C,D) Mineralized nodules stained with alizarin red S on Day 21 in MC3T3-E1 cells cultured in optimized (C+) medium with 10% FBS or with 7.5% FBS + 2.5% serum from mice exposed to either physiological serum (Control) or pomegranate peel extract (PGPE). (C) Representative image of alizarin red staining for each condition. (D) Relative quantification of alizarin red staining with picture analyser software ImageJ. Results are presented as mean ± standard deviation, SD (n = 6). (E) Transcriptomic analysis of the main osteoblast markers in MC3T3-E1 cells cultured in optimized medium after 7 days of treatment with serum from mice exposed to either physiological serum (Control) or pomegranate peel extract (PGPE). Transcriptomic analysis of MC3T3-E1 mRNA levels determined by Taqman Low-Density Arrays are presented as fold change compared to the Control condition (fold change = 1) as mean ± SD (n = 6). Genes: ALP: alkaline phosphatase; βCat: β-catenin;Coll1a1: collagen 1a1; Coll2a1: collagen 2a1; DDR2: discoidin domain receptor 2; BSP: bone sialoprotein; Lrp5; OCN: osteocalcin; OPG: osteoprotegerin; osteri RunX2 and Wnt.

Mentions: To insure that mouse sera (harvested from mice fed either a standard diet (Control) or the control diet enriched with PGPE) were not deleterious for MC3T3-E1 cells, a cell viability test was performed. Incubation with serum from PGPE animals did not modify the proliferation of the MC3T3-E1 cells, on days 2 and 7, in comparison with cells cultured with serum from CT mice (Figure 2A).


Pomegranate Peel Extract Prevents Bone Loss in a Preclinical Model of Osteoporosis and Stimulates Osteoblastic Differentiation in Vitro.

Spilmont M, Léotoing L, Davicco MJ, Lebecque P, Miot-Noirault E, Pilet P, Rios L, Wittrant Y, Coxam V - Nutrients (2015)

Osteoblast viability, differentiation and transcription in culture, after exposure to serum harvested from mice raised either under control conditions or treated with pomegranate peel extract. * p < 0.05 PGPE versus Control condition. (A) Cell proliferation test performed on MC3T3-E1 cells cultured in minimal medium (2% Fetal Bovine Serum, FBS), supplemented with serum from mice given either physiological serum (1.25% FBS + 0.75 Control mice serum) or pomegranate peel extract, PGPE (1.25% FBS + 0.75 PGPE mice serum), after 2 and 7 days of treatment. Results are expressed as mean ± standard error of the mean (SEM) (n = 6). (B) Alkaline phosphatase, ALP activity of MC3T3-E1 cells cultured in optimized medium with 10% FBS or with 7.5% FBS + 2.5% serum from mice fed either physiological serum (Control) or pomegranate peel extract (PGPE), after 0, 2, 7 and 14 days of treatment. Results are expressed as mean ± SEM (n = 8). (C,D) Mineralized nodules stained with alizarin red S on Day 21 in MC3T3-E1 cells cultured in optimized (C+) medium with 10% FBS or with 7.5% FBS + 2.5% serum from mice exposed to either physiological serum (Control) or pomegranate peel extract (PGPE). (C) Representative image of alizarin red staining for each condition. (D) Relative quantification of alizarin red staining with picture analyser software ImageJ. Results are presented as mean ± standard deviation, SD (n = 6). (E) Transcriptomic analysis of the main osteoblast markers in MC3T3-E1 cells cultured in optimized medium after 7 days of treatment with serum from mice exposed to either physiological serum (Control) or pomegranate peel extract (PGPE). Transcriptomic analysis of MC3T3-E1 mRNA levels determined by Taqman Low-Density Arrays are presented as fold change compared to the Control condition (fold change = 1) as mean ± SD (n = 6). Genes: ALP: alkaline phosphatase; βCat: β-catenin;Coll1a1: collagen 1a1; Coll2a1: collagen 2a1; DDR2: discoidin domain receptor 2; BSP: bone sialoprotein; Lrp5; OCN: osteocalcin; OPG: osteoprotegerin; osteri RunX2 and Wnt.
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Related In: Results  -  Collection

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nutrients-07-05465-f002: Osteoblast viability, differentiation and transcription in culture, after exposure to serum harvested from mice raised either under control conditions or treated with pomegranate peel extract. * p < 0.05 PGPE versus Control condition. (A) Cell proliferation test performed on MC3T3-E1 cells cultured in minimal medium (2% Fetal Bovine Serum, FBS), supplemented with serum from mice given either physiological serum (1.25% FBS + 0.75 Control mice serum) or pomegranate peel extract, PGPE (1.25% FBS + 0.75 PGPE mice serum), after 2 and 7 days of treatment. Results are expressed as mean ± standard error of the mean (SEM) (n = 6). (B) Alkaline phosphatase, ALP activity of MC3T3-E1 cells cultured in optimized medium with 10% FBS or with 7.5% FBS + 2.5% serum from mice fed either physiological serum (Control) or pomegranate peel extract (PGPE), after 0, 2, 7 and 14 days of treatment. Results are expressed as mean ± SEM (n = 8). (C,D) Mineralized nodules stained with alizarin red S on Day 21 in MC3T3-E1 cells cultured in optimized (C+) medium with 10% FBS or with 7.5% FBS + 2.5% serum from mice exposed to either physiological serum (Control) or pomegranate peel extract (PGPE). (C) Representative image of alizarin red staining for each condition. (D) Relative quantification of alizarin red staining with picture analyser software ImageJ. Results are presented as mean ± standard deviation, SD (n = 6). (E) Transcriptomic analysis of the main osteoblast markers in MC3T3-E1 cells cultured in optimized medium after 7 days of treatment with serum from mice exposed to either physiological serum (Control) or pomegranate peel extract (PGPE). Transcriptomic analysis of MC3T3-E1 mRNA levels determined by Taqman Low-Density Arrays are presented as fold change compared to the Control condition (fold change = 1) as mean ± SD (n = 6). Genes: ALP: alkaline phosphatase; βCat: β-catenin;Coll1a1: collagen 1a1; Coll2a1: collagen 2a1; DDR2: discoidin domain receptor 2; BSP: bone sialoprotein; Lrp5; OCN: osteocalcin; OPG: osteoprotegerin; osteri RunX2 and Wnt.
Mentions: To insure that mouse sera (harvested from mice fed either a standard diet (Control) or the control diet enriched with PGPE) were not deleterious for MC3T3-E1 cells, a cell viability test was performed. Incubation with serum from PGPE animals did not modify the proliferation of the MC3T3-E1 cells, on days 2 and 7, in comparison with cells cultured with serum from CT mice (Figure 2A).

Bottom Line: The nutritional benefits of pomegranate have attracted great scientific interest.Among chronic diseases, osteoporosis, which is associated with bone remodelling impairment leading to progressive bone loss, could eventually benefit from antioxidant compounds because of the involvement of oxidative stress in the pathogenesis of osteopenia.In addition, PGPE appeared to substantially stimulate osteoblastic MC3T3-E1 alkaline phosphatase (ALP) activity at day 7, mineralization at day 21 and the transcription level of osteogenic markers.

View Article: PubMed Central - PubMed

Affiliation: Unité de Nutrition Humaine, CRNH Auvergne, UMR 1019, INRA, F-63000 Clermont-Ferrand, France. mel.spilmont@gmail.com.

ABSTRACT
The nutritional benefits of pomegranate have attracted great scientific interest. The pomegranate, including the pomegranate peel, has been used worldwide for many years as a fruit with medicinal activity, mostly antioxidant properties. Among chronic diseases, osteoporosis, which is associated with bone remodelling impairment leading to progressive bone loss, could eventually benefit from antioxidant compounds because of the involvement of oxidative stress in the pathogenesis of osteopenia. In this study, with in vivo and ex vivo experiments, we investigated whether the consumption of pomegranate peel extract (PGPE) could limit the process of osteopenia. We demonstrated that in ovariectomized (OVX) C57BL/6J mice, PGPE consumption was able to significantly prevent the decrease in bone mineral density (-31.9%; p < 0.001 vs. OVX mice) and bone microarchitecture impairment. Moreover, the exposure of RAW264.7 cells to serum harvested from mice that had been given a PGPE-enriched diet elicited reduced osteoclast differentiation and bone resorption, as shown by the inhibition of the major osteoclast markers. In addition, PGPE appeared to substantially stimulate osteoblastic MC3T3-E1 alkaline phosphatase (ALP) activity at day 7, mineralization at day 21 and the transcription level of osteogenic markers. PGPE may be effective in preventing the bone loss associated with ovariectomy in mice, and offers a promising alternative for the nutritional management of this disease.

No MeSH data available.


Related in: MedlinePlus