Category Archives: Flavonoids

Naringenin Derivative has Bone Anabolic Effects

Abstract

A naturally occurring naringenin derivative exerts potent bone anabolic effects by mimicking oestrogen action on osteoblasts.

Naringenin and its derivatives have been assessed in bone health for their oestrogen-‘like’ effects but low bioavailability impedes clinical potential. This study was aimed at finding a potent form of naringenin with osteogenic action.
Osteoblast cultures were harvested from mouse calvaria to study differentiation by naringenin, isosakuranetin, poncirin, phloretin and naringenin-6-C-glucoside (NCG). Balb/cByJ ovariectomized (OVx) mice without or with osteopenia were given naringenin, NCG, 17β-oestradiol (E2) or parathyroid hormone (PTH). Efficacy was evaluated by bone microarchitecture using microcomputed tomography and determination of new bone formation by fluorescent labelling of bone. Plasma levels of NCG and naringenin were determined by HPLC.
NCG stimulated osteoblast differentiation more potently than naringenin, while isosakuranetin, poncirin or phloretin had no effect. NCG had better oral bioavailability than naringenin. NCG increased the mRNA levels of oestrogen receptors (ERs) and bone morphogenetic protein (an ER responsive gene) in vivo, more than naringenin. In OVx mice, NCG treatment in a preventive protocol increased bone formation rate (BFR) and improved trabecular microarchitecture more than naringenin or E2. In osteopenic mice, NCG but not naringenin, in a therapeutic protocol, increased BFR and improved trabecular microarchitecture, comparable with effects of PTH treatment. Stimulatory effects of NCG on osteoblasts were abolished by an ER antagonist. NCG transactivated ERβ but not ERα. NCG exhibited no uterine oestrogenicity unlike naringenin.
NCG is a potent derivative of naringenin that has bone anabolic action through the activation of osteoblast ERs and exhibited substantial oral bioavailability.

Swarnkar G, Sharan K, Siddiqui JA, Mishra JS…
Br. J. Pharmacol. Mar 2012
PMID: 21864313 | Free Full Text

Apigenin Inhibits Osteoclasts in Mouse Cells

Abstract

Attenuation of osteoclastogenesis and osteoclast function by apigenin.

The physiological effects of the flavone, apigenin on bone cells were studied. We first show that apigenin inhibits tumor necrosis factor alpha (TNFalpha)- and interferon gamma (IFNgamma)-induced secretion of several osteoclastogenic cytokines from MC3T3-E1 mouse calvarial osteoblast cell line. Ligands of the TNF receptor family constitute the most potent osteoclastic cytokines. In MC3T3-E1 cells, apigenin dose-dependently (from 5 to 20 microM) inhibits TNFalpha-induced production of the osteoclastogenic cytokines, IL-6 (interleukin-6), RANTES (regulated upon activation, normal T cell-expressed and -secreted), monocyte chemoattractant protein-1 (MCP-1) and MCP-3. In addition, apigenin inhibits IFNgamma-stimulated secretion of monokines, CXCL-9, and -10 in MC3T3-E1 cells. Next, we show that apigenin strongly inhibits differentiation of 3T3-L1 preadipocytes to adipocytes with attendant inhibition of adipocyte differentiation-induced IL-6, MCP-1, and leptin production. Inhibition of adipogenic differentiation by apigenin could be due to induction of osteogensis as it robustly upregulates mRNA levels of bone morphogenetic protein-6 (BMP-6). Finally, the presence of apigenin inhibited osteoclast differentiation from the RAW 264.7 cell line by reducing receptor activator of nuclear factor kappa ligand (RANKL)-induced expression of tartrate-resistant acid phosphatase (TRAP), RANK, and calcitonin receptor but not CCR1, resulting in the inhibition of multinucleated osteoclast formation. Similarly, apigenin inhibited expression of the osteoclast differentiation markers TRAP, RANK, and c-Fms in osteoclast precursor cells obtained from mouse bone marrow following treatment with RANKL and macrophage colony stimulating factor (MCSF). Furthermore, apigenin induced apoptosis of mature osteoclasts obtained from rabbit long bone and inhibited bone resorption. In all instances, a structurally related compound, flavone had no significant effect. These data suggest that apigenin has multiple effects on all three bone cells that could prevent bone loss in vivo.

Bandyopadhyay S, Lion JM, Mentaverri R, Ricupero DA…
Biochem. Pharmacol. Jul 2006
PMID: 16750176

Phloretin Inhibits Osteoclasts In Vitro

Abstract

Novel antiosteoclastogenic activity of phloretin antagonizing RANKL-induced osteoclast differentiation of murine macrophages.

Bone-remodeling imbalance resulting in more bone resorption than bone formation is known to cause skeletal diseases such as osteoporosis. Phloretin, a natural dihydrochalcone compound largely present in apple peels, possesses antiphotoaging, and antiinflammatory activity.
Phloretin inhibited receptor activator of NF-κB ligand (RANKL)-induced formation of multinucleated osteoclasts and diminished bone resorption area produced during the osteoclast differentiation process. It was also found that ≥ 10 μM phloretin reduced RANKL-enhanced tartrate-resistance acid phosphatase activity and matrix metalloproteinase-9 secretion in a dose-dependent manner. The phloretin treatment retarded RANKL-induced expression of carbonic anhydrase II, vacuolar-type H(+) -ATPase D2 and β3 integrin, all involved in the bone resorption. Furthermore, submicromolar phloretin diminished the expression and secretion of cathepsin K elevated by RANKL, being concurrent with inhibition of TRAF6 induction and NF-κB activation. RANKL-induced activation of nuclear factor of activated T cells c1 (NFATc1) and microphthalmia-associated transcription factor was also suppressed by phloretin.
These results demonstrate that the inhibition of osteoclast differentiation and bone resorption by phloretin entail a disturbance of TRAF6-NFATc1-NF-κB pathway triggered by RANKL. Therefore, phloretin may be a potential therapeutic agent targeting osteoclast differentiation and bone resorption in skeletal diseases such as osteoporosis.

Kim JL, Kang MK, Gong JH, Park SH…
Mol Nutr Food Res Aug 2012
PMID: 22700286

Glabridin Inhibits Osteoclasts In Vitro

Abstract

The inhibitory effect and the molecular mechanism of glabridin on RANKL-induced osteoclastogenesis in RAW264.7 cells.

Osteoblastic bone formation and osteoclastic bone resorption are in balance to maintain a constant, homeostatically controlled amount of bone. Excessive bone resorption by osteoclasts is involved in the pathogenesis of bone-related disorders. In the present study, we evaluated the inhibitory effects of glabridin, a flavonoid purified from licorice root, on the receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation and its molecular mechanisms in murine osteoclast progenitor RAW264.7 cells. Glabridin significantly inhibited RANKL-induced tartrate-resistant acid phosphatase (TRAP) activity, the formation of multinucleated osteoclasts and resorption-pit formation. In mechanistic studies of the anti-osteoclastogenic potential of glabridin, we found that glabridin inhibited RANKL-induced expression of c-Fos and subsequent expression of NFATc1, which is a master regulator of osteoclastogenesis. Interestingly, glabridin inhibited the RANKL-induced expression of signaling molecules (TRAF6, GAB2, ERK2, JNK1 and MKK7) and osteoclast survival-related signaling pathways such as c-Src, PI3K and Akt2. Glabridin also inhibited the bone resorptive activity of mature osteoclasts by inhibiting osteoclast-associated genes (cathepsin K, MMP-9, CAII, TCIRG1, OSTM1 and CLCN7). Taken together, our data suggest that glabridin holds great promise for use in preventing osteoclastogenesis by inhibiting RANKL-induced activation of signaling molecules and subsequent transcription factors in osteoclast precursors and these findings may be useful for evaluating treatment options in bone-destructive diseases.

Kim HS, Suh KS, Sul D, Kim BJ…
Int. J. Mol. Med. Feb 2012
PMID: 22038020

Review: Green Tea May Decrease Fractures by Improving Bone Density and Osteoblasts and Suppressing Osteoclasts

Abstract

Green tea and bone metabolism.

Osteoporosis is a major health problem in both elderly women and men. Epidemiological evidence has shown an association between tea consumption and the prevention of age-related bone loss in elderly women and men. Ingestion of green tea and green tea bioactive compounds may be beneficial in mitigating bone loss of this population and decreasing their risk of osteoporotic fractures. This review describes the effect of green tea or its bioactive components on bone health, with an emphasis on (i) the prevalence and etiology of osteoporosis; (ii) the role of oxidative stress and antioxidants in osteoporosis; (iii) green tea composition and bioavailability; (iv) the effects of green tea and its active components on osteogenesis, osteoblastogenesis, and osteoclastogenesis from human epidemiological, animal, as well as cell culture studies; (v) possible mechanisms explaining the osteoprotective effects of green tea bioactive compounds; (vi) other bioactive components in tea that benefit bone health; and (vii) a summary and future direction of green tea and bone health research and the translational aspects. In general, tea and its bioactive components might decrease the risk of fracture by improving bone mineral density and supporting osteoblastic activities while suppressing osteoclastic activities.

Shen CL, Yeh JK, Cao JJ, Wang JS
Nutr Res Jul 2009
PMID: 19700031

Flavonoids Associated with Increased Bone Density in Women

Abstract

Associations between dietary flavonoid intakes and bone health in a Scottish population.

Flavonoids are bioactive polyphenols found particularly in fruit and vegetables, but little is known about their role in bone health in humans. The aim of this observational study was to investigate whether dietary flavonoid intake was associated with bone mineral density (BMD) and bone resorption in a large group of perimenopausal Scottish women. Over 3000 women completed a food frequency questionnaire as part of an osteoporosis screening study. The diets were analyzed for flavonoid intake using a food composition database. BMD was measured at the femoral neck (FN) and lumbar spine (LS) by dual-energy X-ray absorptiometry (DXA). Free pyridinoline (PYD) and deoxypyridinoline (DPD) were measured by high-performance liquid chromatography (HPLC) in second early morning fasted urine samples. The mean flavonoid intake of the diet was 307 ±199 mg/d. The catechin family contributed the most to flavonoid intakes (55%), and the flavones the least (<1%). Associations were found between energy-adjusted total flavonoid intakes and BMD at the FN and LS (FN r = 0.054, LS r = 0.036, p ≤ .05). Annual percent change in BMD was associated with intakes of procyanidins and catechins (p ≤ .05), and flavanones were negatively associated with bone-resorption markers (PYD r = -0.049, DPD r = -0.057, p ≤ .001). These associations were still seen after adjusting for confounders. It is concluded that dietary flavonoid intakes are associated with BMD, supporting the evidence from animal and cellular studies.

Hardcastle AC, Aucott L, Reid DM, Macdonald HM
J. Bone Miner. Res. May 2011
PMID: 21541996

Phloridzin Prevents Bone Loss in Ovariectomized Rats

Abstract

Prevention of bone loss by phloridzin, an apple polyphenol, in ovariectomized rats under inflammation conditions.

Aging and sex hormones related changes lead to inflammatory and oxidant conditions, which are involved in the pathogenesis of osteoporosis. Recent studies have suggested that polyphenols may exert a protective effect in such conditions. We assessed the effect of phloridzin (Phlo), a flavonoid exclusively found in apple, on bone metabolism in ovariectomized (OVX) or sham-operated (SH) rats with and without inflammation. Six-month-old Wistar rats were allocated to two equal groups that received either a control diet or a diet supplemented with 0.25% Phlo for 80 days. Three weeks before necropsy, inflammation was induced by subcutaneous injection of talc in 10 animals of each group. At necropsy, ovariectomy decreased both total (T-BMD) and metaphyseal (M-BMD) femoral bone mineral density (P < 0.01). Inflammation conditions, checked by an increase in the spleen weight and alpha1-acid glycoprotein concentration in OVX rats, exacerbated the decrease in T-BMD (g/cm2) (as well as M-BMD) observed in castrated animals (P < 0.05). Daily Phlo intake prevented ovariectomy-induced bone loss in conditions of inflammation as shown by T-BMD and M-BMD (P < 0.05). At the diaphyseal site, BMD was improved by Phlo in OVX rats with or without inflammation (P < 0.05). These results could be explained by changes in bone remodeling as the increased urinary deoxypyridinoline excretion in OVX and OVXinf animals was prevented by the polyphenol-rich diet (P < 0.001), while plasma osteocalcin concentration was similar in all experimental groups. In conclusion, Phlo consumption may provide protection against ovariectomy-induced osteopenia under inflammation conditions by improving inflammation markers and bone resorption.

Puel C, Quintin A, Mathey J, Obled C…
Calcif. Tissue Int. Nov 2005
PMID: 16307390

p-Hydroxycinnamic Acid Stimulates Osteoblastogenesis Mouse Cells

Abstract

Bioactive flavonoid p-hydroxycinnamic acid stimulates osteoblastogenesis and suppresses adipogenesis in bone marrow culture.

The bioactive flavonoid p-hydroxycinnamic acid (HCA), which is an intermediate-metabolic substance in plants and fruits, is synthesized from tyrosine. The biological effect of HCA is poorly understood. Among cinnamic acid and its related compounds, HCA has a specific-anabolic effect on bone, being found to stimulate osteoblastogenesis and to inhibit osteoclastogenesis through the suppression of NF-κB signaling, thereby preventing bone loss. Bone marrow mesenchymal stem cells give rise to ostoblasts and adipocytes. HCA might therefore have effects on osteoblastogenesis and adipogenesis in bone marrow culture. This study demonstrates (1) that HCA has stimulatory effects on osteoblastogenesis and mineralization and suppressive effects on adipogenesis in mouse bone marrow culture and (2) that HCA depresses adipogenesis in mouse 3T3-L1 preadipocytes in vitro. Such effects of HCA might be involved in the differentiation of mesenchymal stem cells.

Yamaguchi M, Baile CA, Zhu S, Shoji M
Cell Tissue Res. Dec 2013
PMID: 24026435 | Free Full Text

p-Hydroxycinnamic Acid Stimulates Mineralization in Mouse Osteoblastic Cells

Abstract

Phytocomponent p-hydroxycinnamic acid stimulates mineralization in osteoblastic MC3T3-E1 cells.

Phytocomponent p-hydroxycinnamic acid (HCA) has been shown to have stimulatory effects on bone calcification and inhibitory effects on bone resorption in rat femoral tissues in vitro. Whether HCA has a stimulatory effect on mineralization in osteoblastic cells is unknown. This study was undertaken to determine the effect of HCA on mineralization in osteoblastic MC3T3-E1 cells in vitro.Cells were cultured for 72 h in a minimum essential medium (alpha-MEM) containing 10% fetal bovine serum (FBS), and the cells with subconfluency were changed to a medium containing either vehicle or HCA (10(-7)-10(-5) M) without FBS. Culture with HCA (10(-7)-10(-5) M) did not have a significant effect on cell proliferation and cell death. Deoxyribonucleic acid (DNA) content in osteoblastic cells was significantly increased after culture with HCA (10(-6) or 10(-5) M) for 48 or 72 h. Alkaline phosphatase activity in osteoblastic cells was significantly increased after culture with HCA (10(-7)-10(-5) M) for 24, 48, or 72 h. The results with Alizarin red staining for calcium showed that mineralization was significantly stimulated after culture with HCA (10(-8)-10(-5) M) for 7, 14, or 21 days. This study demonstrates that HCA has stimulatory effects on mineralization in osteoblastic MC3T3-E1 cells.

Yamaguchi M, Lai YL, Uchiyama S, Nakagawa T
Int. J. Mol. Med. Sep 2008
PMID: 18698486

p-Hydroxycinnamic Acid Prevents Bone Loss in Diabetic Rats

Abstract

Oral administration of phytocomponent p-hydroxycinnamic acid has a preventive effect on bone loss in streptozotocin-induced diabetic rats.

The phytocomponent p-hydroxycinnamic acid (HCA) has been shown to have a stimulatory effect on bone formation and an inhibitory effect on bone resorption in rat femoral tissues in vitro. The preventive effect of HCA on bone loss induced in streptozotocin (STZ)-diabetic rats was investigated in vivo. Rats received a single subcutaneous administration of STZ (6.0 mg/100 g body weight), and then the animals were orally administered HCA (0.25, 0.5, or 1.0 mg/100 g body weight) once daily for 14 days. STZ administration caused a significant decrease in body weight and a significant increase in serum glucose, triglyceride, and calcium levels, indicating a diabetic state. These alterations were significantly prevented by administration of HCA (0.25, 0.5, or 1.0 mg/100 g). Calcium content in the femoral-diaphyseal and -metaphyseal tissues was significantly decreased in STZ-diabetic rats. This decrease was significantly prevented after administration of HCA (0.25, 0.5, or 1.0 mg/100 g). Alkaline phosphatase activity in the diaphyseal and metaphyseal tissues was significantly decreased in STZ-diabetic rats. The decrease in diaphyseal alkaline phosphatase activity in STZ-diabetic rats was significantly prevented after administration of HCA (0.5 and 1.0 mg/l00 g). The diaphyseal DNA content was also significantly decreased in STZ-diabetic rats. Administration of HCA (0.25, 0.5, or 1.0 mg/100 g) caused a significant increase in DNA content in the diaphyseal and metaphyseal tissues in STZ-diabetic rats. This study demonstrates that the intake of HCA has preventive effects on bone loss in STZ-diabetic rats, and that the intake has partially restorative effects on serum biochemical findings in the diabetic state.

Yamaguchi M, Uchiyama S, Lai YL
Int. J. Mol. Med. May 2007
PMID: 17390086