EGCG Suppresses Osteoclasts and Arthritis in Mice

Abstract

(-)-Epigallocatechin-3-gallate suppresses osteoclast differentiation and ameliorates experimental arthritis in mice.

To verify the effects of (-)-epigallocatechin-3-gallate (EGCG) on osteoclast differentiation and on experimental arthritis in mice.
Human osteoclasts were differentiated from peripheral blood monocytes. The effects of EGCG were examined by tartrate-resistant acid phosphatase (TRAP) staining, bone resorption assay, Western blotting, and quantitative real-time polymerase chain reaction.

Arthritis was induced in mice by injecting a cocktail of monoclonal antibodies against collagen. EGCG (20 microg/gm body weight) was administered intraperitoneally every day from day 0 through the end of the experiments (day 15). The effects of EGCG were determined by assessments of joint swelling, histologic changes, and TRAP staining on day 15.
EGCG reduced the generation of TRAP-positive multinucleated cells, bone resorption activity, and osteoclast-specific gene expression without affecting cell viability. EGCG down-regulated expression of nuclear factor of activated T cells c1 (NF-ATc1), but not of NF-kappaB, c-Fos, and c-Jun, suggesting that down-regulation of NF-ATc1 is one of the molecular bases of EGCG action. Additionally, EGCG treatment ameliorated clinical symptoms and reduced histologic scores in arthritic mice (P < 0.05). The in vivo effect of EGCG on osteoclast differentiation was not clear in this model, probably because EGCG suppressed the inflammation itself.
EGCG suppressed osteoclast differentiation and ameliorated experimental arthritis in mice over the short term. It remains to be established whether EGCG is useful for the prevention and treatment of osteoporosis and rheumatoid arthritis.

Morinobu A, Biao W, Tanaka S, Horiuchi M…
Arthritis Rheum. Jul 2008
PMID: 18576345 | Free Full Text

EGCG Inhibits Osteoclasts in Mice

Abstract

Epigallocatechin-3-gallate inhibits osteoclastogenesis by down-regulating c-Fos expression and suppressing the nuclear factor-kappaB signal.

Epigallocatechin-3-gallate (EGCG), the major anti-inflammatory compound in green tea, has been shown to suppress osteoclast differentiation. However, the precise molecular mechanisms underlying the inhibitory action of EGCG in osteoclastogenesis and the effect of EGCG on inflammation-mediated bone destruction remain unclear. In this study, we found that EGCG inhibited osteoclast formation induced by osteoclastogenic factors in bone marrow cell-osteoblast cocultures but did not affect the ratio of receptor activator of nuclear factor kappaB (NF-kappaB) ligand (RANKL) to osteoprotegerin induced by osteoclastogenic factors in osteoblasts. We also found that EGCG inhibited osteoclast formation from bone marrow macrophages (BMMs) induced by macrophage colony-stimulating factor plus RANKL in a dose-dependent manner without cytotoxicity. Pretreatment with EGCG significantly inhibited RANKL-induced the gene expression of c-Fos and nuclear factor of activated T-cells (NFATc1), essential transcription factors for osteoclast development. EGCG suppressed RANKL-induced activation of c-Jun N-terminal protein kinase (JNK) pathway, among the three well known mitogen-activated protein kinases and also inhibited RANKL-induced phosphorylation of the NF-kappaB p65 subunit at Ser276 and NF-kappaB transcriptional activity without affecting the degradation of IkappaBalpha and NF-kappaB DNA-binding in BMMs. The inhibitory effect of EGCG on osteoclast formation was somewhat reversed by retroviral c-Fos overexpression, suggesting that c-Fos is a downstream target for antiosteoclastogenic action of EGCG. In addition, EGCG treatment reduced interleukin-1-induced osteoclast formation and bone destruction in mouse calvarial bone in vivo. Taken together, our data suggest that EGCG has an antiosteoclastogenic effect by inhibiting RANKL-induced the activation of JNK/c-Jun and NF-kappaB pathways, thereby suppressing the gene expression of c-Fos and NFATc1 in osteoclast precursors.

Lee JH, Jin H, Shim HE, Kim HN…
Mol. Pharmacol. Jan 2010
PMID: 19828731 | Free Full Text

EGCG Improves Bone in Ovariectomized Rats

Abstract

(-)-Epigallocatechin-3-gallate improves bone microarchitecture in ovariectomized rats.

Previously, we reported that (-)-epigallocatechin-3-gallate (EGCG), a green tea polyphenol, increased the osteogenic differentiation of murine bone marrow mesenchymal stem cells by increasing the messenger RNA expression of osteogenesis-related genes, alkaline phosphatase activity, and, eventually, mineralization. The present study further investigated the effects of EGCG on bone microstructure change and possible mechanisms in ovariectomy (OVX)-induced osteopenic rats.
Rats subjected to OVX were administered EGCG systemically for 12 weeks. Proximal tibial bone mineral densities before and after treatment were compared between groups. Changes in the microarchitecture of both the proximal tibia and the third lumbar spine were compared between EGCG-treated and nontreated groups using micro-CT (μCT). Bone histology and immunohistochemistry in the proximal tibia were evaluated.
Results showed that EGCG 3.4 mg/kg/day (estimated peak serum concentration, 10 μmol/L) hampered the decrease in bone mineral density (from 7.97% to 3.96%) and improved the parameters of μCT measurements, including bone volume (from 18% to 27%), trabecular thickness (from 0.17 to 0.22 mm), trabecular number (from 1.13 to 1.37 mm(-1)), and trabecular separation (from 0.91 to 0.69 mm), compared with nontreated ovariectomized rats. Similar improvements in bone volume (from 30% to 49%) and trabecular thickness (from 0.14 to 0.26 mm) were also found in the third lumbar spine. Bone volume in the tibial cortex also increased after EGCG treatment (from 9% to 28%). A higher trabecular number and greater trabecular volume were also seen in histology, further confirming the results of μCT. The immunolocalized bone morphogenetic protein 2 brown-stained area increased from 31% in the OVX group to 53% in the OVX + 10 EGCG group (P < 0.01). Serial biochemistry data revealed no significant systemic toxic effect of EGCG.
Intraperitoneal treatment with EGCG 3.4 mg/kg/day for 3 months can mitigate bone loss and improve bone microarchitecture in ovariectomized rats, and increased expression of bone morphogenetic protein 2 may contribute to this effect.

Chen CH, Kang L, Lin RW, Fu YC…
Menopause Jun 2013
PMID: 23511703

Coffee Delays Bone Repair in Rats

Abstract

Effects of cigarette smoke inhalation and coffee consumption on bone formation and osseous integration of hydroxyapatite implant.

The present study aims to assess the effects of cigarette smoke inhalation and/or coffee consumption on bone formation and osseous integration of a dense hydroxyapatite (DHA) implant in rats. For this study, 20 male rats were divided into four groups (n = 5): CT (control) group, CE (coffee) group, CI (cigarette) group and CC (coffee + cigarette) group. During 16 weeks, animals in the CI group were exposed to cigarette smoke inhalation equivalent to 6 cigarettes per day; specimens in the CE group drank coffee as liquid diet; and rats in the CC group were submitted to both substances. In the 6th week a 5 mm slit in the parietal bone and a 4 mm slit in the tibia were performed on the left side: the former was left open while the latter received a DHA implant. As soon as surgeries were finished, the animals returned to their original protocols and after 10 weeks of exposure they were euthanised (ethically sacrificed) and the mentioned bones collected for histological processing. Data showed that exposure to cigarette smoke inhalation and coffee consumption did not interfere in weight gain and that solid and liquid diet consumption was satisfactory. Rats in the CC group showed a decrease in bone neoformation around the tibial DHA implant (31.8 ± 2.8) as well as in bone formation in the parietal slit (28.6 ± 2.2). On their own, cigarette smoke inhalation or coffee consumption also led to diminished bone neoformation around the implant and delayed the bone repair process in relation to the CT group. However, reduction in the bone repair process was accentuated with exposure to both cigarette smoke inhalation and coffee consumption in this study.

Andrade AR, Sant’Ana DC, Mendes JA, Moreira M…
Braz J Biol Feb 2013
PMID: 23644799 | Free Full Text

Tea Protects Bone in Older Women

Abstract

Tea drinking is associated with benefits on bone density in older women.

Impaired hip structure assessed by dual-energy X-ray absorptiometry (DXA) areal bone mineral density (aBMD) is an independent predictor for osteoporotic hip fracture. Some studies suggest that tea intake may protect against bone loss.
Using both cross-sectional and longitudinal study designs, we examined the relation of tea consumption with hip structure. Randomly selected women (n = 1500) aged 70-85 y participated in a 5-y prospective trial to evaluate whether oral calcium supplements prevent osteoporotic fractures. aBMD at the hip was measured at years 1 and 5 with DXA. A cross-sectional analysis of 1027 of these women at 5 y assessed the relation of usual tea intake, measured by using a questionnaire, with aBMD. A prospective analysis of 164 women assessed the relation of tea intake at baseline, measured by using a 24-h dietary recall, with change in aBMD from years 1 to 5.
In the cross-sectional analysis, total hip aBMD was 2.8% greater in tea drinkers (x: 806; 95% CI: 797, 815 mg/cm(2)) than in non-tea drinkers (784; 764, 803 mg/cm(2)) (P < 0.05). In the prospective analysis over 4 y, tea drinkers lost an average of 1.6% of their total hip aBMD (-32; -45, -19 mg/cm(2)), but non-tea drinkers lost 4.0% (-13; -20, -5 mg/cm(2)) (P < 0.05). Adjustment for covariates did not influence the interpretation of results.
Tea drinking is associated with preservation of hip structure in elderly women. This finding provides further evidence of the beneficial effects of tea consumption on the skeleton.

Devine A, Hodgson JM, Dick IM, Prince RL
Am. J. Clin. Nutr. Oct 2007
PMID: 17921409 | Free Full Text

Cola Associated with Low Bone Density in Older Women

Abstract

Colas, but not other carbonated beverages, are associated with low bone mineral density in older women: The Framingham Osteoporosis Study.

Soft drink consumption may have adverse effects on bone mineral density (BMD), but studies have shown mixed results. In addition to displacing healthier beverages, colas contain caffeine and phosphoric acid (H3PO4), which may adversely affect bone.
We hypothesized that consumption of cola is associated with lower BMD. BMD was measured at the spine and 3 hip sites in 1413 women and 1125 men in the Framingham Osteoporosis Study by using dual-energy X-ray absorptiometry. Dietary intake was assessed by food-frequency questionnaire. We regressed each BMD measure on the frequency of soft drink consumption for men and women after adjustment for body mass index, height, age, energy intake, physical activity score, smoking, alcohol use, total calcium intake, total vitamin D intake, caffeine from noncola sources, season of measurement, and, for women, menopausal status and estrogen use.
Cola intake was associated with significantly lower (P < 0.001-0.05) BMD at each hip site, but not the spine, in women but not in men. The mean BMD of those with daily cola intake was 3.7% lower at the femoral neck and 5.4% lower at Ward’s area than of those who consumed <1 serving cola/mo. Similar results were seen for diet cola and, although weaker, for decaffeinated cola. No significant relations between noncola carbonated beverage consumption and BMD were observed. Total phosphorus intake was not significantly higher in daily cola consumers than in nonconsumers; however, the calcium-to-phosphorus ratios were lower.
Intake of cola, but not of other carbonated soft drinks, is associated with low BMD in women. Additional research is needed to confirm these findings.

Tucker KL, Morita K, Qiao N, Hannan MT…
Am. J. Clin. Nutr. Oct 2006
PMID: 17023723 | Free Full Text

Mediterranean Diet or Nuts May Benefit Bones

Abstract

Mediterranean diet and bone mineral density in two age groups of women.

We hypothesized that adherence to the Mediterranean diet measured as a Mediterranean diet score (MDS) has a beneficial effect on bone mineral density (BMD). For the purposes of this study, a sample of healthy women from Southern Spain was chosen. Subjects were grouped into two major groups: a first group consisted of women of reproductive age (premenopausal, pre-M) and a second group consisted of postmenopausal women (pos-M). The consumption of vegetables and fruit was found to be significantly related to BMD in both groups of subjects studied. In the pre-M group, the lipid ratio was positively associated with BMD and in pos-M women nuts intake was also associated with BMD. After implementing the analysis of covariance analysis, significant linear trends between the MDS and BMD were observed in all subjects studied. Our results indicate that a varied diet based on Mediterranean diet patterns may be beneficial in the prevention of osteoporosis.

Rivas A, Romero A, Mariscal-Arcas M, Monteagudo C…
Int J Food Sci Nutr Mar 2013
PMID: 22946650

Mediterranean Diet + Nuts Does Not Significantly Increase Resorption

Abstract

Mediterranean diet and high dietary acid load associated with mixed nuts: effect on bone metabolism in elderly subjects.

Objectives: To analyze the effect of differing diet on the acid load content on bone metabolism.
Design: Multicentric, randomized, single-blind, parallel-group clinical trial.
Setting: Outpatient clinics.
Two hundred thirty-eight elderly men and women aged 60 to 80 at high risk for cardiovascular disease were randomly assigned to three interventional groups: a recommended low-fat diet (control diet group), a Mediterranean diet supplemented with virgin olive oil, or a Mediterranean diet supplemented with mixed nuts.
Main outcomes were 12-month changes from baseline in bone formation and resorption markers and bone mass measured according to quantitative ultrasound scanning.
The baseline data on the anthropometric, bone densitometry, and biochemical variables did not differ between the three groups. Dietary potential renal acid load (PRAL) and daily net endogenous acid production (NEAP) at baseline did not differ between groups. After intervention, subjects allocated to the Mediterranean diet with mixed nuts had a significant increase of PRAL and NEAP. In comparison, subjects in the Mediterranean diet with nuts group had higher parathyroid hormone (PTH) levels (2.63, 95% confidence interval (CI)=-1.01-6.35, P=.02) and a nonsignificantly higher (0.31, 95% CI=-0.13-0.74, P=.14) urine free deoxypyridoxine:creatinine ratio, a marker of bone resorption, than the control group and the Mediterranean diet with virgin olive oil group.
A Mediterranean dietary pattern associated with a high dietary acid load derived from consumption of mixed nuts does not seem to have a much greater effect on bone metabolism biomarkers, with the exception of PTH levels, than a Mediterranean diet without mixed nuts or a control diet in elderly subjects.

Bulló M, Amigó-Correig P, Márquez-Sandoval F, Babio N…
J Am Geriatr Soc Oct 2009
PMID: 19807791

High Protein + Low Calcium Increases Fracture – 2008

Abstract

Proteins, dietary acid load, and calcium and risk of postmenopausal fractures in the E3N French women prospective study.

Excess dietary proteins and “acid ash” diets have been suspected to increase the risk of osteoporosis, but experimental and epidemiological evidence is mixed. We aimed to determine whether the association between protein intake and the overall acid-base equilibrium of the diet (as renal net acid excretion [RNAE] estimate) and fracture risk vary according to calcium intake. During an average of 8.37 +/- 1.73 yr of follow-up, 2408 women reported a fracture (excluding high-impact trauma) among 36,217 postmenopausal women from the E3N prospective study. We used Cox regression models to study the interaction between calcium and, respectively, proteins and RNAE, from the 1993 dietary questionnaire for fracture risk determination, adjusting for potential confounders. There was no overall association between fracture risk and total protein or RNAE. However, in the lowest quartile of calcium (<400 mg/1000 kcal), high protein intake was associated with a significant increased fracture risk (RR = 1.51 for highest versus lowest quartile; 95% CI, 1.17-1.94). An increasing fracture risk with increasing animal protein intake was also observed (trend, p < 0.0001). A similar pattern of interaction for fracture risk was observed between RNAE and calcium. In this Western population of postmenopausal women with normal to high protein intake and fairly high calcium intake, there was no overall association between total protein or RNAE and fracture risk. However, there was some evidence that high protein-high acid ash diets were associated with an increased risk of fracture when calcium intake was low (<400 mg/1000 kcal).

Dargent-Molina P, Sabia S, Touvier M, Kesse E…
J. Bone Miner. Res. Dec 2008
PMID: 18665794

Fish or Fish Oil No Association With Fracture – 2010

Abstract

Fish consumption, bone mineral density, and risk of hip fracture among older adults: the cardiovascular health study.

Marine n-3 polyunsaturated fatty acids (PUFAs) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) may be beneficial for bone health, but few studies have investigated the association with fish consumption. Our aim was to study associations of fish and EPA + DHA consumption with bone mineral density (BMD) and hip fracture risk and determine whether high linoleic acid (LA) intake, the major dietary n-6 PUFA, modifies the associations. The study population consisted of 5045 participants aged 65 years and older from the Cardiovascular Health Study. Data on BMD were available for 1305 participants. Food-frequency questionnaire was used to assess dietary intake, and hip fracture incidence was assessed prospectively by review of hospitalization records. After multivariable adjustment, femoral neck BMD was 0.01 g/cm(2) lower in the highest versus lowest tuna/other-fish intake category (p = .05 for trend). EPA + DHA intake (higher versus lower median of 0.32 g/day) was associated with lower femoral neck BMD (0.66 versus 0.71 g/cm(2), p < .001) among those with LA intake greater than the median 12.1 g/day (p = .03 for interaction). No significant associations were found with total-hip BMD. During mean follow-up of 11.1 years, 505 hip fractures occurred. Fish or EPA + DHA consumption was not significantly associated with fracture incidence [hazard ratio (HR) for extreme categories: HR = 1.23, 95% confidence interval (CI) 0.83-1.84 for tuna/other fish; HR = 1.16, 95% CI 0.91-1.49 for fried fish; and HR = 0.98, 95% CI 0.71-1.36 for EPA + DHA]. High LA intake did not modify these associations. In this large prospective cohort of older adults, fish consumption was associated with very small differences in BMD and had no association with hip fracture risk.

Virtanen JK, Mozaffarian D, Cauley JA, Mukamal KJ…
J. Bone Miner. Res. Sep 2010
PMID: 20572022 | Free Full Text