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1.  The Effects of Tualang Honey on Bone Metabolism of Postmenopausal Women 
Osteoporosis which is characterized by low bone mass and microarchitectural deterioration with a consequent increase in bone fragility can be associated with various stimuli such as oxidative stress and inflammation. Postmenopausal women are more prone to osteoporosis due to reduction in estrogen which may further lead to elevation of oxidative stress and lipid accumulation which will promote osteoblasts apoptosis. Proinflammatory cytokines are elevated following estrogen deficiency. These cytokines are important determinants of osteoclasts differentiation and its bone resorption activity. The main treatment for postmenopausal osteoporosis is estrogen replacement therapy (ERT). Despite its effectiveness, ERT, however, can cause many adverse effects. Therefore, alternative treatment that is rich in antioxidant and can exert an anti-inflammatory effect can be given to replace the conventional ERT. Tualang honey is one of the best options available as it contains antioxidant as well as exerting anti-inflammatory effect which can act as a free radical scavenger, reducing the oxidative stress level as well as inhibiting proinflammatory cytokine. This will result in survival of osteoblasts, reduced osteoclastogenic activity, and consequently, reduce bone loss. Hence, Tualang honey can be used as an alternative treatment of postmenopausal osteoporosis with minimal side effects.
doi:10.1155/2012/938574
PMCID: PMC3437962  PMID: 22973408
2.  Bone Micro-CT Assessments in an Orchidectomised Rat Model Supplemented with Eurycoma longifolia 
Recent studies suggested that Eurycoma longifolia, a herbal plant, may have the potential to treat osteoporosis in elderly male. This study aimed to determine the effects of Eurycoma longifolia supplementation on the trabecular bone microarchitecture of orchidectomised rats (androgen-deficient osteoporosis model). Forty-eight-aged (10–12 months old) Sprague Dawley rats were divided into six groups of sham-operated (SHAM), orchidectomised control (ORX), orchidectomised + 7 mg/rat testosterone enanthate (TEN) and orchidectomised + Eurycoma longifolia 30 mg/kg (EL30), orchidectomised + Eurycoma longifolia 60 mg/kg (EL60), orchidectomised + Eurycoma longifolia 90 mg/kg (EL90). Rats were euthanized following six weeks of treatment. The left femora were used to measure the trabecular bone microarchitecture using micro-CT. Orchidectomy significantly decreased connectivity density, trabecular bone volume, and trabecular number compared to the SHAM group. Testosterone replacement reversed all the orchidectomy-induced changes in the micro-CT parameters. EL at 30 and 60 mg/kg rat worsened the trabecular bone connectivity density and trabecular separation parameters of orchidectomised rats. EL at 90 mg/kg rat preserved the bone volume. High dose of EL (90 mg/kg) may have potential in preserving the bone microarchitecture of orchidectomised rats, but lower doses may further worsen the osteoporotic changes.
doi:10.1155/2012/501858
PMCID: PMC3431134  PMID: 22952556
3.  Combined Effects of Eurycoma longifolia and Testosterone on Androgen-Deficient Osteoporosis in a Male Rat Model 
Androgen-deficient osteoporosis in men is treated with testosterone therapy, which is associated with side effects. Eurycoma longifolia (EL) is known to possess androgenic properties and has been reported to protect bone from androgen-deficient osteoporosis in experimental animal models. The present study aimed to determine the effectiveness of combination therapy of EL and testosterone (T) in treating androgen-deficient osteoporosis. Forty male Sprague-Dawley rats were divided into: sham-operated (SHAM), orchidectomized-control (ORX), orchidectomized with testosterone (ORX + T), orchidectomized with EL (ORX + EL), and orchidectomized with combined T and EL therapy (ORX + T + EL). EL was administered via oral gavages daily at the dose of 15 mg/kg. T was injected intramuscularly at 8 mg/kg and 4 mg/kg for the ORX + T and ORX + T + EL groups, respectively. Following 6 weeks of treatment, the osteocalcin levels of ORX + T and ORX + T + EL groups were significantly lower than the SHAM group (P < 0.05). The posttreatment CTX levels of ORX + T and ORX + T + EL groups were significantly lower than their pretreatment levels (P < 0.05). Biomechanically, the strain parameter of the ORX + T + EL group was significantly higher than the ORX group (P < 0.05). Thus, the combination therapy of EL and low-dose T has potential for treatment of androgen-deficient osteoporosis. The lower T dose is beneficial in reducing the sideeffects of testosterone therapy.
doi:10.1155/2012/872406
PMCID: PMC3424595  PMID: 22924057
4.  Eurycoma longifolia: Medicinal Plant in the Prevention and Treatment of Male Osteoporosis due to Androgen Deficiency 
Osteoporosis in elderly men is now becoming an alarming health issue due to its relation with a higher mortality rate compared to osteoporosis in women. Androgen deficiency (hypogonadism) is one of the major factors of male osteoporosis and it can be treated with testosterone replacement therapy (TRT). However, one medicinal plant, Eurycoma longifolia Jack (EL), can be used as an alternative treatment to prevent and treat male osteoporosis without causing the side effects associated with TRT. EL exerts proandrogenic effects that enhance testosterone level, as well as stimulate osteoblast proliferation and osteoclast apoptosis. This will maintain bone remodelling activity and reduce bone loss. Phytochemical components of EL may also prevent osteoporosis via its antioxidative property. Hence, EL has the potential as a complementary treatment for male osteoporosis.
doi:10.1155/2012/125761
PMCID: PMC3403331  PMID: 22844328
5.  Tocotrienol supplementation in postmenopausal osteoporosis: evidence from a laboratory study 
Clinics  2013;68(10):1338-1343.
OBJECTIVE:
Accelerated bone loss that occurs in postmenopausal women has been linked to oxidative stress and increased free radicals. We propose the use of antioxidants to prevent and reverse postmenopausal osteoporosis. This study aimed to examine the effects of tocotrienol, a vitamin E analog, on bone loss due to estrogen deficiency. Our previous study showed that tocotrienol increased the trabecular bone volume and trabecular number in ovariectomized rats. In the current study, we investigated the effects of tocotrienol supplementation on various biochemical parameters in a postmenopausal osteoporosis rat model.
MATERIALS AND METHODS:
A total of 32 female Wistar rats were randomly divided into four groups. The baseline group was sacrificed at the start of the study, and another group was sham operated. The remaining rats were ovariectomized and either given olive oil as a vehicle or treated with tocotrienol at a dose of 60 mg/kg body weight. After four weeks of treatment, blood was withdrawn for the measurement of interleukin-1 (IL1) and interleukin-6 (IL6) (bone resorbing cytokines), serum osteocalcin (a bone formation marker) and pyridinoline (a bone resorption marker).
RESULTS:
Tocotrienol supplementation in ovariectomized rats significantly reduced the levels of osteocalcin, IL1 and IL6. However, it did not alter the serum pyridinoline level.
CONCLUSION:
Tocotrienol prevented osteoporotic bone loss by reducing the high bone turnover rate associated with estrogen deficiency. Therefore, tocotrienol has the potential to be used as an anti-osteoporotic agent in postmenopausal women.
doi:10.6061/clinics/2013(10)08
PMCID: PMC3798611  PMID: 24212841
Estrogen Deficiency; Ovariectomy; Tocotrienol
6.  Labisia pumila regulates bone-related genes expressions in postmenopausal osteoporosis model 
Background
Labisia Pumila var. alata (LPva) has shown potential as an alternative to estrogen replacement therapy (ERT) in prevention of estrogen-deficient osteoporosis. In earlier studies using postmenopausal model, LPva was able to reverse the ovariectomy-induced changes in biochemical markers, bone calcium, bone histomorphometric parameters and biomechanical strength. The mechanism behind these protective effects is unclear but LPva may have regulated factors that regulate bone remodeling. The aim of this study is to determine the bone-protective mechanism of LPva by measuring the expressions of several factors involved in bone formative and resorptive activities namely Osteoprotegerin (OPG), Receptor Activator of Nuclear Factor kappa-B Ligand (RANKL), Macrophage-Colony Stimulating Factor (MCSF) and Bone Morphogenetic Protein-2 (BMP-2).
Methods
Thirty-two female Wistar rats were randomly divided into four groups: Sham-operated (Sham), ovariectomized control (OVXC), ovariectomized with Labisia pumila var. alata (LPva) and ovariectomized with ERT (Premarin®) (ERT). The LPva and ERT were administered via daily oral gavages at doses of 17.5 mg/kg and 64.5 μg/kg, respectively. Following two months of treatment, the rats were euthanized and the gene expressions of BMP-2, OPG, RANKL and MCSF in the femoral bones were measured using a branch - DNA technique.
Results
The RANKL gene expression was increased while the OPG and BMP-2 gene expressions were reduced in the OVXC group compared to the SHAM group. There were no significant changes in the MCSF gene expressions among the groups. Treatment with either LPva or ERT was able to prevent these ovariectomy-induced changes in the gene expressions in ovariectomized rats with similar efficacy.
Conclusion
LPva may protect bone against estrogen deficiency-induced changes by regulating the RANKL, OPG and BMP-2 gene expressions.
doi:10.1186/1472-6882-13-217
PMCID: PMC3847139  PMID: 24007208
Labisia pumila; Postmenopausal osteoporosis; Estrogen; OPG; BMP-2; RANKL; MCSF
8.  Palm Tocotrienol Supplementation Enhanced Bone Formation in Oestrogen-Deficient Rats 
Postmenopausal osteoporosis is the commonest cause of osteoporosis. It is associated with increased free radical activity induced by the oestrogen-deficient state. Therefore, supplementation with palm-oil-derived tocotrienols, a potent antioxidant, should be able to prevent this bone loss. Our earlier studies have shown that tocotrienol was able to prevent and even reverse osteoporosis due to various factors, including oestrogen deficiency. In this study we compared the effects of supplementation with palm tocotrienol mixture or calcium on bone biomarkers and bone formation rate in ovariectomised (oestrogen-deficient) female rats. Our results showed that palm tocotrienols significantly increased bone formation in oestrogen-deficient rats, seen by increased double-labeled surface (dLS/Bs), reduced single-labeled surface (sLS/BS), increased mineralizing surface (MS/BS), increased mineral apposition rate (MAR), and an overall increase in bone formation rate (BFR/BS). These effects were not seen in the group supplemented with calcium. However, no significant changes were seen in the serum levels of the bone biomarkers, osteocalcin, and cross-linked C-telopeptide of type I collagen, CTX. In conclusion, palm tocotrienol is more effective than calcium in preventing oestrogen-deficient bone loss. Further studies are needed to determine the potential of tocotrienol as an antiosteoporotic agent.
doi:10.1155/2012/532862
PMCID: PMC3485551  PMID: 23150728
9.  Vitamin E and Bone Structural Changes: An Evidence-Based Review 
Purpose. This paper explores the effects of vitamin E on bone structural changes. Methods. A systematic review of the literature was conducted to identify relevant studies about vitamin E and osteoporosis/bone structural changes. A comprehensive search in Medline and CINAHL for relevant studies published between the years 1946 and 2012 was conducted. The main inclusion criteria were published in English, studies had to report the association or effect of vitamin E and osteoporosis-related bone changes, and the osteoporosis-related bone changes should be related to lifestyle variables, aging, or experimentally-induced conditions. Results. The literature search identified 561 potentially relevant articles, whereby 11 studies met the inclusion criteria. There were three human epidemiological studies and eight animal experimental studies included in this paper. Four animal studies reported positive bone structural changes with vitamin E supplementation. The rest of the studies had negative changes or no effect. Studies with positive changes reported better effects with tocotrienol vitamin E isomer supplementation. Conclusions. This evidence-based review underscores the potential of vitamin E being used for osteoporosis. The effect of one of the vitamin E isomers, tocotrienols, on bone structural changes warrants further exploration. Controlled human observational studies should be conducted to provide stronger evidence.
doi:10.1155/2012/250584
PMCID: PMC3483709  PMID: 23118786
10.  Two Different Isomers of Vitamin E Prevent Bone Loss in Postmenopausal Osteoporosis Rat Model 
Postmenopausal osteoporotic bone loss occurs mainly due to cessation of ovarian function, a condition associated with increased free radicals. Vitamin E, a lipid-soluble vitamin, is a potent antioxidant which can scavenge free radicals in the body. In this study, we investigated the effects of alpha-tocopherol and pure tocotrienol on bone microarchitecture and cellular parameters in ovariectomized rats. Three-month-old female Wistar rats were randomly divided into ovariectomized control, sham-operated, and ovariectomized rats treated with either alpha-tocopherol or tocotrienol. Their femurs were taken at the end of the four-week study period for bone histomorphometric analysis. Ovariectomy causes bone loss in the control group as shown by reduction in both trabecular volume (BV/TV) and trabecular number (Tb.N) and an increase in trabecular separation (Tb.S). The increase in osteoclast surface (Oc.S) and osteoblast surface (Ob.S) in ovariectomy indicates an increase in bone turnover rate. Treatment with either alpha-tocopherol or tocotrienol prevents the reduction in BV/TV and Tb.N as well as the increase in Tb.S, while reducing the Oc.S and increasing the Ob.S. In conclusion, the two forms of vitamin E were able to prevent bone loss due to ovariectomy. Both tocotrienol and alpha-tocopherol exert similar effects in preserving bone microarchitecture in estrogen-deficient rat model.
doi:10.1155/2012/161527
PMCID: PMC3484319  PMID: 23118785
11.  Curcumin Protects against Ovariectomy-Induced Bone Changes in Rat Model 
Osteoporosis is a metabolic disease affecting both men and women especially in postmenopausal women. Curcumin possesses many medicinal properties. In this study, thirty two female Sprague-Dawley rats were used to determine the potential effect of curcumin in prevention of bone loss following ovariectomy. The animals were divided into Sham group, ovariectomised control, ovariectomised treated with curcumin 110 mg/kg and ovariectomised treated with Premarin 100 μg/kg. The treatments were given via daily oral gavages for 60 days. The structural parameters such as bone volume, trabecular number, trabecular thickness and trabecular separation were found to be deteriorated in ovariectomised rats compared to Sham group. Moreover, the reduced osteoblast count, the increased osteoclast count and increased eroded surface were found in ovariectomised groups. Treatment with curcumin was able to reverse all these ovariectomy-induced deteriorations. Curcumin treatment was as effective as Premarin in most parameters except the bone volume and eroded surface, which were better than Premarin. The high dose of curcumin treatment was not only able to reduce the osteoclast number but also increase the osteoblast count. Therefore, the potential effect of curcumin can be applied as an alternative to oestrogen for prevention of postmenopausal osteoporosis.
doi:10.1155/2012/174916
PMCID: PMC3463175  PMID: 23049604
12.  Eurycoma longifolia upregulates osteoprotegerin gene expression in androgen- deficient osteoporosis rat model 
Background
Eurycoma longifolia (EL) has been shown recently to protect against bone calcium loss in orchidectomised rats, the model for androgen-deficient osteoporosis. The mechanism behind this is unclear but it may be related to its ability to elevate testosterone levels or it may directly affect bone remodeling. The aim of this study is to determine the mechanism involved by investigating the effects of EL extract on serum testosterone levels, bone biomarkers, biomechanical strength and gene expression of Receptor Activator of Nuclear Factor kappa-B ligand (RANKL), Osteoprotegerin (OPG) and Macrophage-Colony Stimulating Factor (MCSF) in orchidectomised rats.
Methods
Thirty-two male Sprague–Dawley rats were divided into: Sham-operated group (SHAM); orchidectomised-control group (ORX); orchidectomised and given 15 mg/kg EL extract (ORX + EL) and orchidectomised and given 8 mg/kg testosterone (ORX + T). The rats were treated for 6 weeks. The serum levels of testosterone, osteocalcin and C-terminal telopeptide of type I collagen (CTX) were measured using the ELISA technique. The femoral bones were subjected to biomechanical testing. The tibial bone gene expressions of RANKL, OPG and MCSF were measured using the branch DNA technique.
Results
The post-treatment level of testosterone was found to be significantly reduced by orchiectomy (p < 0.05). Both ORX + EL and ORX + T groups have significantly higher post-treatment testosterone levels compared to their pre-treatment levels (p < 0.05). The bone resorption marker (CTx) was elevated after orchiectomy but was suppressed after treatment in the ORX + EL and ORX + T groups (p < 0.05). There was no significant finding for the femoral biomechanical parameters. The tibial OPG gene expression in the ORX group was significantly lower compared to the SHAM and ORX + EL groups (p < 0.05).
Conclusion
Supplementation with EL extract elevated the testosterone levels, reduced the bone resorption marker and upregulated OPG gene expression of the orchidectomised rats. These actions may be responsible for the protective effects of EL extract against bone resorption due to androgen deficiency.
doi:10.1186/1472-6882-12-152
PMCID: PMC3493384  PMID: 22967165
Eurycoma longifolia; Osteoporosis; Orchiectomy; OPG; RANKL
13.  Labisia pumila Prevents Complications of Osteoporosis by Increasing Bone Strength in a Rat Model of Postmenopausal Osteoporosis 
Estrogen replacement therapy (ERT) is the main treatment postmenopausal osteoporosis. However, ERT causes serious side effects, such as cancers and thromboembolic problems. Labisia pumila var. alata (LPva) is a herb with potential as an alternative to ERT to prevent complications of osteoporosis, especially fragility fractures. This study was conducted to determine the effects of LPva on the biomechanical strength of femora exposed to osteoporosis due to estrogen deficiency, using the postmenopausal rat model. Thirty-two female rats were randomly divided into four groups: Sham-operated (Sham), ovariectomized control (OVXC), ovariectomized with Labisia pumila var. alata (LP), and ovariectomized with ERT (Premarin) (ERT). The LPva and ERT were administered via oral gavage daily at doses of 17.5 mg/kg and 64.5 μg/kg, respectively. Following two months of treatment, the rats were euthanized, and their right femora were prepared for bone biomechanical testing. The results showed that ovariectomy compromised the femoral strength, while LPva supplementation to the ovariectomized rats improved the femoral strength. Therefore, LPva may be as effective as ERT in preventing fractures due to estrogen-deficient osteoporosis.
doi:10.1155/2012/948080
PMCID: PMC3443997  PMID: 22991574
14.  Nigella sativa: A Potential Antiosteoporotic Agent 
Nigella sativa seeds (NS) has been used traditionally for various illnesses. The most abundant and active component of NS is thymoquinone (TQ). Animal studies have shown that NS and TQ may be used for the treatment of diabetes-induced osteoporosis and for the promotion of fracture healing. The mechanism involved is unclear, but it was postulated that the antioxidative, and anti-inflammatory activities may play some roles in the treatment of osteoporosis as this bone disease has been linked to oxidative stress and inflammation. This paper highlights studies on the antiosteoporotic effects of NS and TQ, the mechanisms behind these effects and their safety profiles. NS and TQ were shown to inhibit inflammatory cytokines such as interleukin-1 and 6 and the transcription factor, nuclear factor κB. NS and TQ were found to be safe at the current dosage for supplementation in human with precautions in children and pregnant women. Both NS and TQ have shown potential as antiosteoporotic agent but more animal and clinical studies are required to further assess their antiosteoporotic efficacies.
doi:10.1155/2012/696230
PMCID: PMC3438907  PMID: 22973403
15.  Role of Medicinal Plants and Natural Products on Osteoporotic Fracture Healing 
Popularly known as “the silent disease” since early symptoms are usually absent, osteoporosis causes progressive bone loss, which renders the bones susceptible to fractures. Bone fracture healing is a complex process consisting of four overlapping phases—hematoma formation, inflammation, repair, and remodeling. The traditional use of natural products in bone fractures means that phytochemicals can be developed as potential therapy for reducing fracture healing period. Located closely near the equator, Malaysia has one of the world's largest rainforests, which are homes to exotic herbs and medicinal plants. Eurycoma longifolia (Tongkat Ali), Labisia pumila (Kacip Fatimah), and Piper sarmentosum (Kaduk) are some examples of the popular ethnic herbs, which have been used in the Malay traditional medicine. This paper focuses on the use of natural products for treating fracture as a result of osteoporosis and expediting its healing.
doi:10.1155/2012/714512
PMCID: PMC3438813  PMID: 22973405
16.  The effects of alpha-tocopherol supplementation on fracture healing in a postmenopausal osteoporotic rat model 
Clinics  2012;67(9):1077-1085.
OBJECTIVE:
Osteoporosis increases the risk of bone fractures and may impair fracture healing. The aim of this study was to investigate whether alpha-tocopherol can improve the late-phase fracture healing of osteoporotic bones in ovariectomized rats.
METHOD:
In total, 24 female Sprague-Dawley rats were divided into three groups. The first group was sham-operated, and the other two groups were ovariectomized. After two months, the right femora of the rats were fractured under anesthesia and internally repaired with K-wires. The sham-operated and ovariectomized control rat groups were administered olive oil (a vehicle), whereas 60 mg/kg of alpha-tocopherol was administered via oral gavage to the alpha-tocopherol group for six days per week over the course of 8 weeks. The rats were sacrificed, and the femora were dissected out. Computed tomography scans and X-rays were performed to assess fracture healing and callus staging, followed by the assessment of callus strengths through the biomechanical testing of the bones.
RESULTS:
Significantly higher callus volume and callus staging were observed in the ovariectomized control group compared with the sham-operated and alpha-tocopherol groups. The ovariectomized control group also had significantly lower fracture healing scores than the sham-operated group. There were no differences between the alpha-tocopherol and sham-operated groups with respect to the above parameters. The healed femora of the ovariectomized control group demonstrated significantly lower load and strain parameters than the healed femora of the sham-operated group. Alpha-tocopherol supplementation was not able to restore these biomechanical properties.
CONCLUSION:
Alpha-tocopherol supplementation appeared to promote bone fracture healing in osteoporotic rats but failed to restore the strength of the fractured bone.
doi:10.6061/clinics/2012(09)16
PMCID: PMC3438250  PMID: 23018307
Bone; Fracture; Osteoporosis; Vitamin E; Alpha-tocopherol
17.  Effects of Eurycoma longifolia on Testosterone Level and Bone Structure in an Aged Orchidectomised Rat Model 
Testosterone replacement is the choice of treatment in androgen-deficient osteoporosis. However, long-term use of testosterone is potentially carcinogenic. Eurycoma longifolia (EL) has been reported to enhance testosterone level and prevent bone calcium loss but there is a paucity of research regarding its effect on the bone structural parameters. This study was conducted to explore the bone structural changes following EL treatment in normal and androgen-deficient osteoporosis rat model. Thirty-six male Sprague-Dawley rats aged 12 months were divided into normal control, normal rat supplemented with EL, sham-operated, orchidectomised-control, orchidectomised with testosterone replacement, and orchidectomised with EL supplementation groups. Testosterone serum was measured both before and after the completion of the treatment. After 6 weeks of the treatment, the femora were processed for bone histomorphometry. Testosterone replacement was able to raise the testosterone level and restore the bone volume of orchidectomised rats. EL supplementation failed to emulate both these testosterone actions. The inability of EL to do so may be related to the absence of testes in the androgen deficient osteoporosis model for EL to stimulate testosterone production.
doi:10.1155/2012/818072
PMCID: PMC3433727  PMID: 22966245
18.  Effects of Low-Dose versus High-Dose γ-Tocotrienol on the Bone Cells Exposed to the Hydrogen Peroxide-Induced Oxidative Stress and Apoptosis 
Oxidative stress and apoptosis can disrupt the bone formation activity of osteoblasts which can lead to osteoporosis. This study was conducted to investigate the effects of γ-tocotrienol on lipid peroxidation, antioxidant enzymes activities, and apoptosis of osteoblast exposed to hydrogen peroxide (H2O2). Osteoblasts were treated with 1, 10, and 100 μM of γ-tocotrienol for 24 hours before being exposed to 490 μM (IC50) H2O2 for 2 hours. Results showed that γ-tocotrienol prevented the malondialdehyde (MDA) elevation induced by H2O2 in a dose-dependent manner. As for the antioxidant enzymes assays, all doses of γ-tocotrienol were able to prevent the reduction in SOD and CAT activities, but only the dose of 1 μM of GTT was able to prevent the reduction in GPx. As for the apoptosis assays, γ-tocotrienol was able to reduce apoptosis at the dose of 1 and 10 μM. However, the dose of 100 μM of γ-tocotrienol induced an even higher apoptosis than H2O2. In conclusion, low doses of γ-tocotrienol offered protection for osteoblasts against H2O2 toxicity, but itself caused toxicity at the high doses.
doi:10.1155/2012/680834
PMCID: PMC3432387  PMID: 22956976
19.  The Effects of Virgin Coconut Oil on Bone Oxidative Status in Ovariectomised Rat 
Virgin coconut oil (VCO) was found to have antioxidant property due to its high polyphenol content. The aim of this study was to investigate the effect of the virgin coconut oil on lipid peroxidation in the bone of an osteoporotic rat model. Normal female Sprague-Dawley rats aged 3 months old were randomly divided into 4 groups, with 8 rats in each group: baseline, sham, ovariectomised (OVX) control group, and OVX given 8% VCO in the diet for six weeks. The oxidative status of the bone was assessed by measuring the index of lipid peroxidation, which is malondialdehyde (MDA) concentration, as well as the endogenous antioxidant enzymes glutathione peroxidase (GPX) and superoxide dismutase (SOD) in the tibia at the end of the study. The results showed that there was a significant decrease in MDA levels in the OVX-VCO group compared to control group. Ovariectomised rats treated with VCO also had significantly higher GPX concentration. The SOD level seemed to be increased in the OVX-VCO group compared to OVX-control group. In conclusion, VCO prevented lipid peroxidation and increased the antioxidant enzymes in the osteoporotic rat model.
doi:10.1155/2012/525079
PMCID: PMC3426286  PMID: 22927879
20.  The Effects of Cosmos caudatus on Structural Bone Histomorphometry in Ovariectomized Rats 
Osteoporosis is considered a serious debilitating disease. Cosmos caudatus (ulam raja), a plant containing antioxidant compounds and minerals, may be used to treat and prevent osteoporosis. This study determines the effectiveness of C. caudatus as bone protective agent in postmenopausal osteoporosis rat model. Thirty-two female rats, aged 3 months old, were divided into 4 groups. Group one was sham operated (sham) while group two was ovariectomized. These two groups were given ionized water by forced feeding. Groups three and four were ovariectomized and given calcium 1% ad libitum and force-fed with C. caudatus at the dose of 500 mg/kg, respectively. Treatments were given six days per week for a period of eight weeks. Body weight was monitored every week and structural bone histomorphometry analyses of the femur bones were performed. Ovariectomy decreased trabecular bone volume (BV/TV), decreased trabecular number (Tb.N), and increased trabecular separation (Tb.Sp). Both calcium 1% and 500 mg/kg C. caudatus reversed the above structural bone histomorphometric parameters to normal level. C. caudatus shows better effect compared to calcium 1% on trabecular number (Tb.N) and trabecular separation (Tb.Sp). Therefore, Cosmos caudatus 500 mg/kg has the potential to act as the therapeutic agent to restore bone damage in postmenopausal women.
doi:10.1155/2012/817814
PMCID: PMC3424602  PMID: 22924056
21.  Tocotrienol Supplementation Improves Late-Phase Fracture Healing Compared to Alpha-Tocopherol in a Rat Model of Postmenopausal Osteoporosis: A Biomechanical Evaluation 
This study investigated the effects of α-tocopherol and palm oil tocotrienol supplementations on bone fracture healing in postmenopausal osteoporosis rats. 32 female Sprague-Dawley rats were divided into four groups. The first group was sham operated (SO), while the others were ovariectomised. After 2 months, the right femora were fractured under anesthesia and fixed with K-wire. The SO and ovariectomised-control rats (OVXC) were given olive oil (vehicle), while both the alpha-tocopherol (ATF) and tocotrienol-enriched fraction (TEF) groups were given alpha-tocopherol and tocotrienol-enriched fraction, respectively, at the dose of 60 mg/kg via oral gavages 6 days per week for 8 weeks. The rats were then euthanized and the femora dissected out for bone biomechanical testing to assess their strength. The callous of the TEF group had significantly higher stress parameter than the SO and OVXC groups. Only the SO group showed significantly higher strain parameter compared to the other treatment groups. The load parameter of the OVXC and ATF groups was significantly lower than the SO group. There was no significant difference in the Young's modulus between the groups. In conclusion, tocotrienol is better than α-tocopherol in improving the biomechanical properties of the fracture callous in postmenopausal osteoporosis rat model.
doi:10.1155/2012/372878
PMCID: PMC3398681  PMID: 22829855
22.  Protective effects of Tualang honey on bone structure in experimental postmenopausal rats 
Clinics  2012;67(7):779-784.
OBJECTIVE:
The objective of this study was to evaluate the effects of Tualang honey on trabecular structure and compare these effects with those of calcium supplementation in ovariectomized rats.
METHODS:
Forty female, Sprague-Dawley rats were randomly divided into five groups (n = 8): four controls and one test arm. The control arm comprised a baseline control, sham-operated control, ovariectomized control, and ovariectomized calcium-treated rats (receiving 1% calcium in drinking water ad libitum). The test arm was composed of ovariectomized, Tualang honey-treated rats (received 0.2 g/kg body weight of Tualang honey). Both the sham-operated control and ovariectomized control groups received vehicle treatment (deionized water), and the baseline control group was sacrificed without treatment.
RESULTS:
All rats were orally gavaged daily for six weeks after day one post-surgery. The bone structural analysis of rats in the test arm group showed a significant increase in the bone volume per tissue volume (BV/TV), trabecular thickness (Tb.Th) and trabecular number (Tb.N) and a significant decrease in inter-trabecular space (Tb.Sp) compared with the ovariectomized control group. The trabecular thickness (Tb.Th) in the test arm group was significantly higher compared with the ovariectomized-calcium treated group, and the inter-trabecular space (Tb.Sp) in the test arm group was significantly narrower compared with the ovariectomized-calcium treated group.
CONCLUSION:
In conclusion, ovariectomized rats that received Tualang honey showed more improvements in trabecular bone structure than the rats that received calcium.
doi:10.6061/clinics/2012(07)13
PMCID: PMC3400169  PMID: 22892923
Tualang honey; Ovariectomy; Osteoporosis; Trabecular; Structural
23.  Expression of TGF-β1 in the blood during fracture repair in an estrogen-deficient rat model 
Clinics  2011;66(12):2113-2119.
OBJECTIVES:
Previous studies have reported that osteoporosis due to estrogen deficiency influences fracture healing. Transforming growth factor (TGF-β) has been found to be involved in fracture healing via the regulation of the differentiation and activation of osteoblasts and osteoclasts. The current study aimed to determine the effects of estrogen on the expression of TGF-β1 during fracture healing in ovariectomized rats.
METHODS:
Thirty female Sprague-Dawley rats weighing 200–250 g were assigned to: (i) a sham-operated group that was given a normal saline; (ii) an ovariectomized control group that was given a normal saline; or (iii) an ovariectomized + estrogen (100 µg/kg/day) group that was treated with conjugated equine estrogen. The right femur of all rats was fractured, and a Kirschner wire was inserted six weeks post-ovariectomy. Treatment with estrogen was given for another six weeks post-fracture. At the end of the study, blood samples were taken, and the right femur was harvested and subjected to biomechanical strength testing.
RESULTS:
The percentage change in the plasma TGF-β1 level before treatment was significantly lower in the ovariectomized control and estrogen groups when compared with the sham group (p<0.001). After six weeks of treatment, the percentage change in the plasma TGF-β1 level in the estrogen group was significantly higher compared with the level in the ovariectomized control group (p = 0.001). The mean ultimate force was significantly increased in the ovariectomized rats treated with estrogen when compared with the ovariectomized control group (p = 0.02).
CONCLUSION:
These data suggest that treatment with conjugated equine estrogen enhanced the strength of the healed bone in estrogen-deficient rats by most likely inducing the expression of TGF-β1.
doi:10.1590/S1807-59322011001200018
PMCID: PMC3226608  PMID: 22189738
Estrogen; Biomechanics; Fracture; Osteoporosis; Ovariectomy; Repair; TGF-β1
24.  Histological Changes in the Fracture Callus Following the Administration of Water Extract of Piper Sarmentosum (Daun Kadok) in Estrogen-Deficient Rats 
Background: The fracture healing is impaired in osteoporosis. Piper sarmentosum is a plant, which contains potent antioxidant, naringenin that may enhance fracture healing. The present histological study aimed to determine the effects of water extract of Piper sarmentosum on the late phase of fracture healing in estrogen-deficient rats.
Methods: Twenty four female Sprague-Dawley rats (200-250 gm) were obtained. Six rats underwent sham operation and the rest were ovariectomized. Six weeks post-ovariectomy all the rats were fractured at the mid-diaphysis of the right femur and a K-wire was inserted for internal fixation. The sham group was given vehicle (normal saline) and the ovariectomized group was randomly subdivided into three groups: (i) ovariectomized-control group supplemented with vehicle; (ii) ovariectomized+estrogen replacement therapy group treated with estrogen (100 µg/kg/day) and (iii) ovariectomized+Piper sarmentosum group treated with Piper sarmentosum water extract (125 mg/kg). Following six weeks of treatment, the rats were sacrificed and the right femora were harvested for histological assessment of fracture callus.
Results: The ovariectomized-control group showed a significant delay in fracture healing compared to the sham, ovariectomized-estrogen replacement therapy and ovariectomized-Piper sarmentosum groups. The median callus score for the ovariectomized-Piper sarmentosum group was 4.50 (range, 4-5), which was significantly higher than the median callus score 3.50 (range, 3-4) for the ovariectomized-control group (P=0.019). However, there was no significant (P>0.05) difference in the callus score among the sham, ovariectomized-estrogen replacement therapy and ovariectomized-Piper sarmentosum groups groups.
Conclusion: Treatment with water extract of Piper sarmentosum proved beneficial in the fracture healing in estrogen-deficient rats.
PMCID: PMC3470271  PMID: 23115413
Antioxidant; callus; fracture healing; histology; osteoporosis; ovariectomy
25.  Piper sarmentosum enhances fracture healing in ovariectomized osteoporotic rats: a radiological study 
Clinics  2011;66(5):865-872.
INTRODUCTION:
Osteoporotic fractures are common during osteoporotic states. Piper sarmentosum extract is known to possess antioxidant and anti-inflammatory properties.
OBJECTIVES:
To observe the radiological changes in fracture calluses following administration of a Piper sarmentosum extract during an estrogen-deficient state.
METHODS:
A total of 24 female Sprague-Dawley rats (200-250 g) were randomly divided into 4 groups: (i) the sham-operated group; (ii) the ovariectomized-control group; (iii) the ovariectomized + estrogen-replacement therapy (ovariectomized-control + estrogen replacement therapy) group, which was supplemented with estrogen (100 µg/kg/day); and (iv) the ovariectomized + Piper sarmentosum (ovariectomized + Piper sarmentosum) group, which was supplemented with a water-based Piper sarmentosum extract (125 mg/kg). Six weeks after an ovariectomy, the right femora were fractured at the mid-diaphysis, and a K-wire was inserted. Each group of rats received their respective treatment for 6 weeks. Following sacrifice, the right femora were subjected to radiological assessment.
RESULTS:
The mean axial callus volume was significantly higher in the ovariectomized-control group (68.2±11.74 mm3) than in the sham-operated, estrogen-replacement-therapy and Piper sarmentosum groups (20.4±4.05, 22.4±4.14 and 17.5±3.68 mm3, respectively). The median callus scores for the sham-operated, estrogen-replacement-therapy and Piper sarmentosum groups had median (range, minimum - maximum value) as 1.0 (0 - 2), 1.0 (1 - 2) and 1.0 (1 - 2), respectively, which were significantly lower than the ovariectomized-control group score of 2.0 (2 - 3). The median fracture scores for the sham-operated, estrogen-replacement-therapy and Piper sarmentosum groups were 3.0 (3 - 4), 3.0 (2 - 3) and 3.0 (2 - 3), respectively, which were significantly higher than the ovariectomized-control group score of 2.0 (1 - 2) (p<0.05).
CONCLUSION:
The Piper sarmentosum extract improved fracture healing, as assessed by the reduced callus volumes and reduced callus scores. This extract is beneficial for fractures in osteoporotic states.
doi:10.1590/S1807-59322011000500025
PMCID: PMC3109388  PMID: 21789393
Piper sarmentosum; Antioxidant; Fracture: Healing; Osteoporosis; Ovariectomy

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