Elsevier

Bone

Volume 83, February 2016, Pages 127-140
Bone

Original Full Length Article
Lycopene treatment against loss of bone mass, microarchitecture and strength in relation to regulatory mechanisms in a postmenopausal osteoporosis model

https://doi.org/10.1016/j.bone.2015.10.017Get rights and content

Highlights

  • We investigated the effect of lycopene treatment on bone health in ovariectomized (OVX) Wistar rats.

  • Lycopene treatment suppressed bone turnover to restore bone strength and microarchitecture.

  • Lycopene treatment down-regulated osteoclast differentiation and upregulated osteoblast function.

Abstract

Lycopene supplementation decreases oxidative stress and exhibits beneficial effects on bone health, but the mechanisms through which it alters bone metabolism in vivo remain unclear. The present study aims to evaluate the effects of lycopene treatment on postmenopausal osteoporosis. Six-month-old female Wistar rats (n = 264) were sham-operated (SHAM) or ovariectomized (OVX). The SHAM group received oral vehicle only and the OVX rats were randomized into five groups receiving oral daily lycopene treatment (mg/kg body weight per day): 0 OVX (control), 15 OVX, 30 OVX, and 45 OVX, and one group receiving alendronate (ALN) (2 μg/kg body weight per day), for 12 weeks. Bone densitometry measurements, bone turnover markers, biomechanical testing, and histomorphometric analysis were conducted. Micro computed tomography was also used to evaluate changes in microarchitecture. Lycopene treatment suppressed the OVX-induced increase in bone turnover, as indicated by changes in biomarkers of bone metabolism: serum osteocalcin (s-OC), serum N-terminal propeptide of type 1 collagen (s-PINP), serum crosslinked carboxyterminal telopeptides (s-CTX-1), and urinary deoxypyridinoline (u-DPD). Significant improvement in OVX-induced loss of bone mass, bone strength, and microarchitectural deterioration was observed in lycopene-treated OVX animals. These effects were observed mainly at sites rich in trabecular bone, with less effect in cortical bone. Lycopene treatment down-regulated osteoclast differentiation concurrent with up-regulating osteoblast together with glutathione peroxidase (GPx) catalase (CAT) and superoxide dismutase (SOD) activities. These findings demonstrate that lycopene treatment in OVX rats primarily suppressed bone turnover to restore bone strength and microarchitecture.

Introduction

Postmenopausal osteoporosis is a serious public health concern associated with significant morbidity, mortality, deterioration of quality of life, and high health care costs [1]. Current FDA-approved therapies include antiresorptive (e.g., bisphosphonate and denosumab) and anabolic (e.g., teriparatide) agents [2]. While denosumab is relatively new and unstudied, bisphosphonate and teriparatide therapies have contributed to a documented decrease in fracture risk among treated patients [3]. However, the potentially undesirable side effects associated with these pharmacological therapies, including atypical fractures, osteonecrosis of the jaw, gastro-oesophageal adverse events, and dizziness, along with poor compliance and cost concerns for teriparatide, continue to challenge their overall efficacy [4], [5]. Thus, research efforts are directed towards discovering more effective, lower-cost therapeutic strategies, including natural alternatives with minimal side effects and fewer compliance challenges.

There is growing evidence that oxidative stress, induced by reactive oxygen species (ROS) that increase with aging or with the onset of an inflammatory state, can adversely affect bone homeostasis [6]. Recent studies have suggested that postmenopausal bone loss may be caused by ROS, which induce a more oxidized bone microenvironment [7], and an excess of ROS may inhibit osteoblast differentiation and proliferation [8]. ROS generated in the extra- or intra-osteoclasts act as signals to enhance osteoclastic differentiation, resulting in more bone resorption [9], [10]. Thus, an imbalance in ROS levels can accelerate bone resorption, resulting in bone fragility and fracture: accordingly, eliminating excessive ROS is an effective approach for maintaining bone integrity [11].

Estrogen deficiency, an independent risk factor for bone fragility [12], has been linked to an increase in oxidative stress. Ovariectomy in rats induces oxidative stress and attenuates levels of antioxidants [13]. In addition, bone marrow of OVX rats exhibits increased ROS levels and decreased activity of antioxidative enzymes (e.g., glutathione reductase) [14]. In a mouse model, higher levels of the antioxidant glutathione prevent bone loss during estrogen deficiency, whereas depletion of antioxidants increases bone loss [15]. Moreover, in OVX rats, plasma lipid peroxidation levels increase as compared with sham-operated controls [16], [17]. In postmenopausal women, Bednarek-Tupikowksa et al. demonstrated higher serum lipid peroxide levels and lower antioxidative potency (indicated by decreased glutathione levels and glutathione peroxidase (GPx) activity), as compared with premenopausal women [18]. Taken together, these studies suggest that estrogen deficiency-induced bone loss is accompanied by higher local and systemic oxidative stress.

Dietary supplementation or treatment with antioxidants is an effective approach to counteract and ameliorate excessive ROS production. Lycopene, a carotenoid found in red fruits and vegetables, especially tomatoes and tomato products, is one of the most potent antioxidants, with a scavenging capacity for singlet oxygen molecules 100 times higher than that of other carotenoids [19], thus, it has been associated with a decreased risk of chronic diseases (for review see [20]). Ben-Dor et al. [21], proposed that the effect of lycopene may be attributed to the induction of anti-oxidant and phase II enzymes. The transcriptional up-regulation of the genes encoding such enzymes is mediated by cis-acting DNA sequences located within their promoter regions known as anti-oxidant response elements (AREs). The major ARE transcription factor is Nrf2 (nuclear factor E2-related factor 2). It plays key roles in the detoxification processes and modulation of anti-oxidant cellular defense system promoting the up-regulation of stress-induced cyto-protective enzymes [e.g., superoxide dismutase (SOD), GPx, heme oxygenase 1 (HO-1)] [22]. The Nrf2 is normally sequestered in the cytoplasm by Kelch-like ECH-associated protein-1 (Keap1). Oxidative stress promotes the dissociations of Nrf2 from Keap1, translocates to the nucleus and complexes with other factors and binds to AREs to regulate the expression of target genes [23], [24]. In addition, Nrf2 also elicits anti-inflammatory effects. Lycopene and its metabolites were found to mediate the activation of Nrf2-ARE signaling and subsequent induction of gene expression [25]. Moreover, lycopene might target other signaling pathways for its cellular anti-oxidant actions (for review see [26]).

Recently, Mackinnon et al. showed that a lycopene-restricted diet significantly decreased circulating lycopene and decreased the antioxidant enzymes SOD and catalase (CAT) in healthy postmenopausal women [27]. In addition, the same group showed in a pilot study using a small group of healthy postmenopausal women that lycopene supplementation significantly decreased oxidative stress markers and a bone resorption marker [28]. In OVX rats, lycopene supplementation prevents bone loss and restores bone strength [29], [30]. However, how lycopene exerts its bone-protective effects remains uncertain.

To provide more insight into the possible mechanisms of the bone-sparing effects of lycopene, we investigated the effect of lycopene treatment on bone loss in an OVX rat model. We comprehensively assessed bone health with measurements of bone turnover markers (BTMs), bone mass, bone dynamics, bone microarchitecture, and bone strength parameters. In conjunction, we evaluated alterations in regulators of oxidative stress and of osteoblast and osteoclast differentiation and activity at the tissue level. Additionally, we used alendronate (ALN) treatment as a positive control to compare the bone metabolic response to lycopene treatment with that of an established antiresorptive therapy.

Section snippets

Animals and experimental design

A total of 264 Wistar female rats, ~ 6 months old, were supplied by the Animal House at King Fahd Medical Research Center (KFMRC), KAU. Rats were housed in individual cages and maintained at 22 °C with a 12-h light/dark cycle. During the study period, rats were given standard rodent chow diet ad libitum (commercial rat cubes containing approximately 18% protein, 3% fat, 77% carbohydrate, and 2% of an inorganic-salt mixture with a vitamin supplement by weight, supplied by Grain Silos and Flour

Body weight, uterine weight, body composition, and serum lycopene levels

Moderate body weight gain was evident in all OVX groups with no significant differences observed in final body weights among the OVX groups (Table 1). Lycopene treatment did not significantly influence the final body weight, body weight gain, or food intake (data not shown). Body composition analysis by DXA, however, showed that while fat mass increased about 22% in the OVX control group, groups with higher dosages of lycopene had 8.9% and 16.4% decreases in fat mass (P < 0.003), lean body mass

Discussion

We investigated the efficacy of lycopene for treatment of postmenopausal physiological changes. We used an ovariectomized rat model to mimic the loss of estrogen associated with menopause. As expected, we found that the mean uterine weight in OVX rats was significantly lower than in the SHAM group (P < 0.001). Lycopene supplementation increased uterine weight in a dose-dependent way (P < 0.01), but uterine weight was still lower in all lycopene-treated OVX groups than the SHAM group (P < 0.001).

Conclusions

In conclusion, lycopene treatment for 12 weeks demonstrated bone-protective effects similar to ALN, improving the biomechanical properties of bone and inhibiting bone resorption in OVX rats. Such effects appear to be primarily due to decreased bone turnover, as indicated by changes in BTMs and other microarchitecture parameters. These findings extend the previously reported effects of lycopene both in experimental animals [29], [30], [59] and postmenopausal women [28], [64]. Moreover, these

Acknowledgements

This study was supported partly by grants from the Ministry of Higher Education (now Education) to the Center of Excellence for Osteoporosis Research (CEOR) at King Abdulaziz University (Grants # CEOR/001-08 and CEOR/004-08) Jeddah, Saudi Arabia and partly by the National Plan for Science, Technology and Innovation (MAARIFAH) – King Abdulaziz City for Science and Technology (KACST), The Kingdom of Saudi Arabia – award no. 11-BIO1552-03.

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    Conflict of interest: The authors have no conflicts of interest.

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