Synergistic effects of combined therapy of curcumin and Fructus Ligustri Lucidi for treatment of osteoporosis: cellular and molecular evidence of enhanced bone formation
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摘要
Objective: The present study explored the effects of the combined herbal therapy consisting of curcumin(CUR) and Fructus Ligustri Lucidi(FLL) on aspects of bone regeneration.Methods: Prior to analyzing the ability of this novel combined herbal therapy to promote aspects of bone regeneration, its cytotoxicity was determined using MC3 T3-E1 cells(pre-osteoblast model). Cell proliferation was evaluated using phase-contrast microscopy and cell differentiation was estimated using alkaline phosphatase activity. The effect of the combined herbal therapy(CUR + FLL) was also assessed in terms of mineralization in the extracellular matrix(ECM) of cultured cells. Further, to explore the molecular mechanisms of bone formation, time-dependent expression of bone-regulating protein biomarkers was also evaluated.Results: Combined herbal therapy(CUR + FLL) significantly upregulated the viability, proliferation and differentiation of MC3 T3-E1 cells compared to the monotherapy of CUR or FLL. The magnitude of ECM mineralization(calcium deposition) was also higher in MC3 T3-E1 cells treated with combined therapy.The time-dependent expression of bone-forming protein biomarkers revealed that the tendency of expression of these bone-regulating proteins was remarkably higher in cells treated with combined therapy.Conclusion: The co-administration of CUR and FLL had superior promotion of elements of bone regeneration in cultured cells, thus could be a promising alternative herbal therapy for the management of bone erosive disorders such as osteoporosis.
Objective: The present study explored the effects of the combined herbal therapy consisting of curcumin(CUR) and Fructus Ligustri Lucidi(FLL) on aspects of bone regeneration.Methods: Prior to analyzing the ability of this novel combined herbal therapy to promote aspects of bone regeneration, its cytotoxicity was determined using MC3 T3-E1 cells(pre-osteoblast model). Cell proliferation was evaluated using phase-contrast microscopy and cell differentiation was estimated using alkaline phosphatase activity. The effect of the combined herbal therapy(CUR + FLL) was also assessed in terms of mineralization in the extracellular matrix(ECM) of cultured cells. Further, to explore the molecular mechanisms of bone formation, time-dependent expression of bone-regulating protein biomarkers was also evaluated.Results: Combined herbal therapy(CUR + FLL) significantly upregulated the viability, proliferation and differentiation of MC3 T3-E1 cells compared to the monotherapy of CUR or FLL. The magnitude of ECM mineralization(calcium deposition) was also higher in MC3 T3-E1 cells treated with combined therapy.The time-dependent expression of bone-forming protein biomarkers revealed that the tendency of expression of these bone-regulating proteins was remarkably higher in cells treated with combined therapy.Conclusion: The co-administration of CUR and FLL had superior promotion of elements of bone regeneration in cultured cells, thus could be a promising alternative herbal therapy for the management of bone erosive disorders such as osteoporosis.
Objective: The present study explored the effects of the combined herbal therapy consisting of curcumin(CUR) and Fructus Ligustri Lucidi(FLL) on aspects of bone regeneration.Methods: Prior to analyzing the ability of this novel combined herbal therapy to promote aspects of bone regeneration, its cytotoxicity was determined using MC3 T3-E1 cells(pre-osteoblast model). Cell proliferation was evaluated using phase-contrast microscopy and cell differentiation was estimated using alkaline phosphatase activity. The effect of the combined herbal therapy(CUR + FLL) was also assessed in terms of mineralization in the extracellular matrix(ECM) of cultured cells. Further, to explore the molecular mechanisms of bone formation, time-dependent expression of bone-regulating protein biomarkers was also evaluated.Results: Combined herbal therapy(CUR + FLL) significantly upregulated the viability, proliferation and differentiation of MC3 T3-E1 cells compared to the monotherapy of CUR or FLL. The magnitude of ECM mineralization(calcium deposition) was also higher in MC3 T3-E1 cells treated with combined therapy.The time-dependent expression of bone-forming protein biomarkers revealed that the tendency of expression of these bone-regulating proteins was remarkably higher in cells treated with combined therapy.Conclusion: The co-administration of CUR and FLL had superior promotion of elements of bone regeneration in cultured cells, thus could be a promising alternative herbal therapy for the management of bone erosive disorders such as osteoporosis.
引文
[1]Lee YS,Choi EM.Costunolide stimulates the function of osteoblastic MC3T3-E1cells.Int Immunopharmacol 2011;11(6):712-8.
[2]Miao Q,Li JG,Miao S,Hu N,Zhang J,Zhang S,et al.The bone-protective effect of genistein in the animal model of bilateral ovariectomy:roles of phytoestrogens and PTH/PTHR1 against post-menopausal osteoporosis.Int J Mol Sci 2012;13(1):56-70.
[3]Kaunitz AM,McClung MR,Feldman RG,Wysocki S.Postmenopausal osteoporosis:fracture risk and prevention.J Fam Pract 2009;58(Suppl 11):S1-6.
[4]Leboime A,Confavreux CB,Mehsen N,Paccou J,David C,Roux C.Osteoporosis and mortality.Joint Bone Spine 2010;77(Suppl 2):S107-12.
[5]Thu HE,Mohamed IN,Hussain Z,Jayusman PA,Shuid AN.Eurycoma longifolia as a potential adoptogen of male sexual health:a systematic review on clinical studies.Chin J Nat Med 2017;15(1):71-80.
[6]Thu HE,Mohamed IN,Hussain Z,Shuid AN.Eurycoma longifolia as a potential alternative to testosterone for the treatment of osteoporosis:exploring timemannered proliferative,differentiative and morphogenic modulation in osteoblasts.J Ethnopharmacol 2017;195:143-58.
[7]Thu HE,Mohamed IN,Hussain Z,Shuid AN.Eurycoma longifolia,a Malaysian medicinal herb,significantly upregulate proliferation and differentiation in pre-osteoblasts(MC3T3-E1):an in vitro model.Int J Pharm Pharm Sci 2016;8(11):199-204.
[8]Hussain Z,Thu HE,Shuid AN,Kesharwani P,Khan S,Hussain F.Phytotherapeutic potential of natural herbal medicines for the treatment of mild-to-severe atopic dermatitis:a review of human clinical studies.Biomed Pharmacother 2017;93:596-608.
[9]Hussain F,Rana Z,Shafique H,Malik A,Hussain Z.Phytopharmacological potential of different species of Morus alba and their bioactive phytochemicals:a review.Asian Pac J Trop Biomed 2017;7(10):950-6.
[10]Ahn MS,Kim HJ,Han MN.Won JS.A study on the antioxidative and antimicrobial activities of the chopi(Zanthoxylum Pipperitum DC.)solvent extract.J Korean Soc Food Cult 2004;19:170-6.
[11]Effendy NM,Mohamed N,Muhammad N,Mohamad IN,Shuid AN.Eurycoma longifolia:medicinal plant in the prevention and treatment of male osteoporosis due to androgen deficiency.Evid Based Complement Altern Med 2012;2012:1-9.
[12]Menon VP,Sudheer AR.Antioxidant and anti-inflammatory properties of curcumin.Adv Exp Med Biol 2007;595:105-25.
[13]Hussain Z,Thu HE,Ng SF,Khan S,Katas H.Nanoencapsulation,an efficient and promising approach to maximize wound healing efficacy of curcumin:a review of new trends and state-of-the-art.Colloids Surf B Biointerfaces2017;150:223-41.
[14]Hussain Z,Thu HE,Amjad MW,Hussain F,Ahmed TA,Khan S.Exploring recent developments to improve antioxidant,anti-inflammatory and antimicrobial efficacy of curcumin:a review of new trends and future perspectives.Mater Sci Eng C Mater Biol Appl 2017;77:1316-26.
[15]Moran JM,Roncero-Martin R,Rodriguez-Velasco FJ,Calderon-Garcia JF,ReySanchez P,Vera V,et al.Effects of curcumin on the proliferation and mineralization of human osteoblast-like cells:implications of nitric oxide.Int J Mol Sci 2012;13(12):16104-18.
[16]Zhang Y,Lai WP,Leung PC,Wu CF,Yao XS,Wong MS.Effects of Fructus Ligustri Lucidi extract on bone turnover and calcium balance in ovariectomized rats.Biol Pharm Bull 2006;29(2):291-6.
[17]Ko CH,Siu WS,Lau CP,Lau CB,Fung KP,Leung PC.Osteoprotective effects of Fructus Ligustri Lucidi aqueous extract in aged ovariectomized rats.Chin Med2010;5:39.
[18]Zhang Y,Leung PC,Che CT,Chow HK,Wu CF,Wong MS.Improvement of bone properties and enhancement of mineralization by ethanol extract of Fructus Ligustri Lucidi.Br J Nutr 2008;99(3):494-502.
[19]Gao L,Li C,Wang Z,Liu X,You Y,Wei H,et al.Ligustri Lucidi Fructus as a traditional Chinese medicine:a review of its phytochemistry and pharmacology.Nat Prod Res 2015;29:493-510.
[20]Jin H.Present situation of chemical compositions of Fructus Ligustri Lucidi.Ji Lin Hua Gong Xue Yuan Xue Bao 2008;25(2):35-7[Chinese with abstract in English].
[21]Liu J,Si S,Qin Y,Zhang B,Song S,Guo Y.The effect of different molecular weight collagen peptides on MC3T3-E1 cells differentiation.Biomed Mater Eng Suppl 2015;26:S2041-7.
[22]Weinreb M,Shinar D,Rodan GA.Different pattern of alkaline phosphatase,osteopontin,and osteocalcin expression in developing rat bone visualized by in situ hybridization.J Bone Miner Res 1990;5:831-42.
[23]Orimo H.The mechanism of mineralization and the role of alkaline phosphatase in health and disease.J Nippon Med Sch 2010;77(1):4-12.
[24]Bonewald LF,Harris SE,Rosser J,Dallas MR,Dallas SL,Camacho NP,et al.von Kossa staining alone is not sufficient to confirm that mineralization in vitro represents bone formation.Calcif Tissue Int 2003;72:537-47.
[25]Tsutsumi K,Saito N,Kawazoe Y,Ooi HK,Shiba T.Morphogenetic study on the maturation of osteoblastic cell as induced by inorganic polyphosphate.PLoSONE 2014;9(2):e86834.
[26]Liu J,Zhang B,Song S,Ma M,Si S,Wang Y,et al.Bovine collagen peptides compounds promote the proliferation and differentiation of MC3T3-E1 preosteoblasts.PLoS ONE 2014;9(6):e99920.
[27]Yamaguchi A,Komori T,Suda T.Regulation of osteoblast differentiation mediated by bone morphogenetic proteins,hedgehogs,and Cbfa1.Endocr Rev2000;21:393-411.
[28]Luppen CA,Leclerc N,Noh T,Barski A,Khokhar A,Boskey AL,et al.Brief bone morphogenetic protein 2 treatment of glucocorticoid-inhibited MC3T3-E1osteoblasts rescues commitment-associated cell cycle and mineralization without alteration of Runx2.J Biol Chem 2003;278(45):44995-5003.
[29]Raida M,Heymann AC,Gunther C,Niederwieser D.Role of bone morphogenetic protein 2 in the crosstalk between endothelial progenitor cells and mesenchymal stem cells.Int J Mol Med 2006;18:735-9.
[30]Ohba S,Nakajima K,Komiyama Y,Kugimiya F,Igawa K,Itaka K,et al.A novel osteogenic helioxanthin-derivative acts in a BMP-dependent manner.Biochem Biophys Res Commun 2007;357(4):854-60.
[31]Zhao J,Ohba S,Shinkai M,Chung UI,Nagamune T.Icariin induces osteogenic differentiation in vitro in a BMP-and Runx2-dependent manner.Biochem Biophys Res Commun 2008;369(2):444-8.
[32]Suzawa M,Takeuchi Y,Fukumoto S,Kato S,Ueno N,Miyazono K,et al.Extracellular matrix associated bone morphogenetic proteins are essential for differentiation of murine osteoblastic cells in vitro.Endocrinology1999;140:2125-33.
[33]Thu HE,Mohamed IN,Hussain Z,Shuid AN.Exploring molecular mechanism of bone-forming capacity of Eurycoma longifolia:evidence of enhanced expression of bone-related biomarkers.J Ayurveda Integr Med 2017.in press.
[34]Komori T.Regulation of bone development and extracellular matrix protein genes by RUNX2.Cell Tissue Res 2010;339(1):189-95.
[35]Sakamoto W,Isomura H,Fujie K,Deyama Y,Kato A,Nishihira J,et al.Homocysteine attenuates the expression of osteocalcin but enhances osteopontin in MC3T3-E1 preosteoblastic cells.Biochim Biophys Acta2005;1740(1):12-6.
[2]Miao Q,Li JG,Miao S,Hu N,Zhang J,Zhang S,et al.The bone-protective effect of genistein in the animal model of bilateral ovariectomy:roles of phytoestrogens and PTH/PTHR1 against post-menopausal osteoporosis.Int J Mol Sci 2012;13(1):56-70.
[3]Kaunitz AM,McClung MR,Feldman RG,Wysocki S.Postmenopausal osteoporosis:fracture risk and prevention.J Fam Pract 2009;58(Suppl 11):S1-6.
[4]Leboime A,Confavreux CB,Mehsen N,Paccou J,David C,Roux C.Osteoporosis and mortality.Joint Bone Spine 2010;77(Suppl 2):S107-12.
[5]Thu HE,Mohamed IN,Hussain Z,Jayusman PA,Shuid AN.Eurycoma longifolia as a potential adoptogen of male sexual health:a systematic review on clinical studies.Chin J Nat Med 2017;15(1):71-80.
[6]Thu HE,Mohamed IN,Hussain Z,Shuid AN.Eurycoma longifolia as a potential alternative to testosterone for the treatment of osteoporosis:exploring timemannered proliferative,differentiative and morphogenic modulation in osteoblasts.J Ethnopharmacol 2017;195:143-58.
[7]Thu HE,Mohamed IN,Hussain Z,Shuid AN.Eurycoma longifolia,a Malaysian medicinal herb,significantly upregulate proliferation and differentiation in pre-osteoblasts(MC3T3-E1):an in vitro model.Int J Pharm Pharm Sci 2016;8(11):199-204.
[8]Hussain Z,Thu HE,Shuid AN,Kesharwani P,Khan S,Hussain F.Phytotherapeutic potential of natural herbal medicines for the treatment of mild-to-severe atopic dermatitis:a review of human clinical studies.Biomed Pharmacother 2017;93:596-608.
[9]Hussain F,Rana Z,Shafique H,Malik A,Hussain Z.Phytopharmacological potential of different species of Morus alba and their bioactive phytochemicals:a review.Asian Pac J Trop Biomed 2017;7(10):950-6.
[10]Ahn MS,Kim HJ,Han MN.Won JS.A study on the antioxidative and antimicrobial activities of the chopi(Zanthoxylum Pipperitum DC.)solvent extract.J Korean Soc Food Cult 2004;19:170-6.
[11]Effendy NM,Mohamed N,Muhammad N,Mohamad IN,Shuid AN.Eurycoma longifolia:medicinal plant in the prevention and treatment of male osteoporosis due to androgen deficiency.Evid Based Complement Altern Med 2012;2012:1-9.
[12]Menon VP,Sudheer AR.Antioxidant and anti-inflammatory properties of curcumin.Adv Exp Med Biol 2007;595:105-25.
[13]Hussain Z,Thu HE,Ng SF,Khan S,Katas H.Nanoencapsulation,an efficient and promising approach to maximize wound healing efficacy of curcumin:a review of new trends and state-of-the-art.Colloids Surf B Biointerfaces2017;150:223-41.
[14]Hussain Z,Thu HE,Amjad MW,Hussain F,Ahmed TA,Khan S.Exploring recent developments to improve antioxidant,anti-inflammatory and antimicrobial efficacy of curcumin:a review of new trends and future perspectives.Mater Sci Eng C Mater Biol Appl 2017;77:1316-26.
[15]Moran JM,Roncero-Martin R,Rodriguez-Velasco FJ,Calderon-Garcia JF,ReySanchez P,Vera V,et al.Effects of curcumin on the proliferation and mineralization of human osteoblast-like cells:implications of nitric oxide.Int J Mol Sci 2012;13(12):16104-18.
[16]Zhang Y,Lai WP,Leung PC,Wu CF,Yao XS,Wong MS.Effects of Fructus Ligustri Lucidi extract on bone turnover and calcium balance in ovariectomized rats.Biol Pharm Bull 2006;29(2):291-6.
[17]Ko CH,Siu WS,Lau CP,Lau CB,Fung KP,Leung PC.Osteoprotective effects of Fructus Ligustri Lucidi aqueous extract in aged ovariectomized rats.Chin Med2010;5:39.
[18]Zhang Y,Leung PC,Che CT,Chow HK,Wu CF,Wong MS.Improvement of bone properties and enhancement of mineralization by ethanol extract of Fructus Ligustri Lucidi.Br J Nutr 2008;99(3):494-502.
[19]Gao L,Li C,Wang Z,Liu X,You Y,Wei H,et al.Ligustri Lucidi Fructus as a traditional Chinese medicine:a review of its phytochemistry and pharmacology.Nat Prod Res 2015;29:493-510.
[20]Jin H.Present situation of chemical compositions of Fructus Ligustri Lucidi.Ji Lin Hua Gong Xue Yuan Xue Bao 2008;25(2):35-7[Chinese with abstract in English].
[21]Liu J,Si S,Qin Y,Zhang B,Song S,Guo Y.The effect of different molecular weight collagen peptides on MC3T3-E1 cells differentiation.Biomed Mater Eng Suppl 2015;26:S2041-7.
[22]Weinreb M,Shinar D,Rodan GA.Different pattern of alkaline phosphatase,osteopontin,and osteocalcin expression in developing rat bone visualized by in situ hybridization.J Bone Miner Res 1990;5:831-42.
[23]Orimo H.The mechanism of mineralization and the role of alkaline phosphatase in health and disease.J Nippon Med Sch 2010;77(1):4-12.
[24]Bonewald LF,Harris SE,Rosser J,Dallas MR,Dallas SL,Camacho NP,et al.von Kossa staining alone is not sufficient to confirm that mineralization in vitro represents bone formation.Calcif Tissue Int 2003;72:537-47.
[25]Tsutsumi K,Saito N,Kawazoe Y,Ooi HK,Shiba T.Morphogenetic study on the maturation of osteoblastic cell as induced by inorganic polyphosphate.PLoSONE 2014;9(2):e86834.
[26]Liu J,Zhang B,Song S,Ma M,Si S,Wang Y,et al.Bovine collagen peptides compounds promote the proliferation and differentiation of MC3T3-E1 preosteoblasts.PLoS ONE 2014;9(6):e99920.
[27]Yamaguchi A,Komori T,Suda T.Regulation of osteoblast differentiation mediated by bone morphogenetic proteins,hedgehogs,and Cbfa1.Endocr Rev2000;21:393-411.
[28]Luppen CA,Leclerc N,Noh T,Barski A,Khokhar A,Boskey AL,et al.Brief bone morphogenetic protein 2 treatment of glucocorticoid-inhibited MC3T3-E1osteoblasts rescues commitment-associated cell cycle and mineralization without alteration of Runx2.J Biol Chem 2003;278(45):44995-5003.
[29]Raida M,Heymann AC,Gunther C,Niederwieser D.Role of bone morphogenetic protein 2 in the crosstalk between endothelial progenitor cells and mesenchymal stem cells.Int J Mol Med 2006;18:735-9.
[30]Ohba S,Nakajima K,Komiyama Y,Kugimiya F,Igawa K,Itaka K,et al.A novel osteogenic helioxanthin-derivative acts in a BMP-dependent manner.Biochem Biophys Res Commun 2007;357(4):854-60.
[31]Zhao J,Ohba S,Shinkai M,Chung UI,Nagamune T.Icariin induces osteogenic differentiation in vitro in a BMP-and Runx2-dependent manner.Biochem Biophys Res Commun 2008;369(2):444-8.
[32]Suzawa M,Takeuchi Y,Fukumoto S,Kato S,Ueno N,Miyazono K,et al.Extracellular matrix associated bone morphogenetic proteins are essential for differentiation of murine osteoblastic cells in vitro.Endocrinology1999;140:2125-33.
[33]Thu HE,Mohamed IN,Hussain Z,Shuid AN.Exploring molecular mechanism of bone-forming capacity of Eurycoma longifolia:evidence of enhanced expression of bone-related biomarkers.J Ayurveda Integr Med 2017.in press.
[34]Komori T.Regulation of bone development and extracellular matrix protein genes by RUNX2.Cell Tissue Res 2010;339(1):189-95.
[35]Sakamoto W,Isomura H,Fujie K,Deyama Y,Kato A,Nishihira J,et al.Homocysteine attenuates the expression of osteocalcin but enhances osteopontin in MC3T3-E1 preosteoblastic cells.Biochim Biophys Acta2005;1740(1):12-6.