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. 2014 Aug 5:9:1247-59.
doi: 10.2147/CIA.S67016. eCollection 2014.

Effects of annatto-derived tocotrienol supplementation on osteoporosis induced by testosterone deficiency in rats

Kok-Yong Chin  1 , Soelaiman Ima-Nirwana  1
Affiliations

Effects of annatto-derived tocotrienol supplementation on osteoporosis induced by testosterone deficiency in rats

Kok-Yong Chin et al. Clin Interv Aging. .

Abstract

Background: Previous animal models have demonstrated that tocotrienol is a potential treatment for postmenopausal osteoporosis. This study evaluated the antiosteoporotic effects of annatto-derived tocotrienol (AnTT) using a testosterone-deficient osteoporotic rat model.

Methods: Forty rats were divided randomly into baseline, sham, orchidectomized, AnTT, and testosterone groups. The baseline group was euthanized without undergoing any surgical treatment or intervention. The remaining groups underwent orchidectomy, with the exception of the sham group. AnTT 60 mg/kg/day was given orally to the AnTT group, while the testosterone group received testosterone enanthate 7 mg/kg per week intramuscularly for 8 weeks. Structural changes in trabecular bone at the proximal tibia were examined using microcomputed tomography. Structural and dynamic changes at the distal femur were examined using histomorphometric methods. Serum osteocalcin and C-terminal of type 1 collagen crosslinks were measured. Bone-related gene expression in the distal femur was examined.

Results: There were significant degenerative changes in structural indices in the orchidectomized group (P<0.05), but no significant changes in dynamic indices, bone remodeling markers, or gene expression (P>0.05) when compared with the sham group. The AnTT group showed significant improvement in structural indices at the femur (P<0.05) and significantly increased expression of bone formation genes (P<0.05). Testosterone was more effective than AnTT in preventing degeneration of bone structural indices in the femur and tibia (P<0.05).

Conclusion: AnTT supplementation improves bone health in testosterone-deficient rats by enhancing bone formation. Its potential should be evaluated further by varying the dosage and treatment duration.

Keywords: bone remodeling; osteoporosis; testosterone; tocotrienol.

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Figures

Figure 1
Figure 1
Percentage changes in body weight, serum terminal-C type 1 collagen crosslinks, and osteocalcin levels in rats. The bar charts show the percentage change in body weight (A), serum terminal-C type 1 collagen crosslinks (B), and osteocalcin levels (C) before and after treatment. The data are shown as the mean with the standard error of the mean. Notes: bSignificant difference versus the sham group; csignificant difference versus the orchidectomized group; dsignificant difference versus the AnTT group. The statistical significance value is set at P<0.05. Abbreviations: AnTT, annatto tocotrienol-supplemented group; BL, baseline group; CTX-1, terminal-C type 1 collagen crosslinks; ORX, orchidectomized group; SH, sham-operated group; TE, testosterone enanthate-supplemented group.
Figure 2
Figure 2
Bone structural indices in proximal tibia of rats evaluated using μCT. The bar charts (A–F) show bone structural indices at the proximal tibia evaluated using μCT. The data are shown as the mean and standard error of the mean. Notes: aSignificant difference versus the baseline group; bsignificant difference versus the sham group; csignificant difference versus the orchidectomized group. The statistical significance value is set at P<0.05. Abbreviations: AnTT, annatto tocotrienol-supplemented group; BL, baseline group; ORX, orchidectomized group; SH, sham-operated group; TE, testosterone enanthate-supplemented group; SMI, Structural model index; Conn.D, connectivity density; BV/TV, bone volume over total volume; Tb. N., trabecular number; Tb. Th., trabecular thickness; Tb. Sp., trabecular separation.
Figure 3
Figure 3
Bone structural and dynamic indices at the distal femur in rats evaluated using the bone histomorphometry technique. The bar charts show the bone structural (A–D) and dynamic (E–I) indices at the distal femur evaluated using the bone histomorphometry technique. The data are shown as the mean and standard error of the mean. Notes: aSignificant difference versus the baseline group; bsignificant difference versus the sham group; csignificant difference versus the orchidectomized group; dsignificant difference versus the AnTT group. The statistical significance value is set at P<0.05. Abbreviations: AnTT, annatto tocotrienol-supplemented group; BL, baseline group; ORX, orchidectomized group; SH, sham-operated group; TE, testosterone enanthate-supplemented group; BV/TV, bone volume over total volume; Tb.N., trabecular number; Tb. Th., trabecular thickness; Tb. Sp., trabecular separation; dLS, double-labeled surface; sLS, single-labeled surface; MAR, mineral apposition rate; MS, mineralizing surface; BFR, bone formation rate.
Figure 3
Figure 3
Bone structural and dynamic indices at the distal femur in rats evaluated using the bone histomorphometry technique. The bar charts show the bone structural (A–D) and dynamic (E–I) indices at the distal femur evaluated using the bone histomorphometry technique. The data are shown as the mean and standard error of the mean. Notes: aSignificant difference versus the baseline group; bsignificant difference versus the sham group; csignificant difference versus the orchidectomized group; dsignificant difference versus the AnTT group. The statistical significance value is set at P<0.05. Abbreviations: AnTT, annatto tocotrienol-supplemented group; BL, baseline group; ORX, orchidectomized group; SH, sham-operated group; TE, testosterone enanthate-supplemented group; BV/TV, bone volume over total volume; Tb.N., trabecular number; Tb. Th., trabecular thickness; Tb. Sp., trabecular separation; dLS, double-labeled surface; sLS, single-labeled surface; MAR, mineral apposition rate; MS, mineralizing surface; BFR, bone formation rate.
Figure 4
Figure 4
Relative expression of genes related to osteoblast differentiation/bone formation in rats. The bar charts (A–I) show the relative expression of genes related to osteoblast/bone formation. The data are shown as the mean with standard error of the mean. Notes: bSignificant difference versus the sham group; csignificant difference versus the orchidectomized group; dsignificant difference versus the AnTT group. The statistical significance value is set at P<0.05. Abbreviations: AnTT, annatto tocotrienol-supplemented group; BL, baseline group; ORX, orchidectomized group; SH, sham-operated group; TE, testosterone enanthate-supplemented group; MFI, mean fluorescence intensity.
Figure 4
Figure 4
Relative expression of genes related to osteoblast differentiation/bone formation in rats. The bar charts (A–I) show the relative expression of genes related to osteoblast/bone formation. The data are shown as the mean with standard error of the mean. Notes: bSignificant difference versus the sham group; csignificant difference versus the orchidectomized group; dsignificant difference versus the AnTT group. The statistical significance value is set at P<0.05. Abbreviations: AnTT, annatto tocotrienol-supplemented group; BL, baseline group; ORX, orchidectomized group; SH, sham-operated group; TE, testosterone enanthate-supplemented group; MFI, mean fluorescence intensity.
Figure 5
Figure 5
Relative expression of genes related to osteoclastogenesis/osteoclast function/bone resorption and PPARG in rats. The bar charts (A–H) show the relative expression of genes related to osteoclastogenesis/osteoclast function/bone resorption and PPARG. The data are shown as the mean and the standard error of the mean. Notes: bSignificant differences versus the sham group; csignificant differences versus the orchidectomized group; dsignificant difference versus the AnTT group. The statistical significance value is set at P<0.05. Abbreviations: AnTT, annatto tocotrienol-supplemented group; BL, baseline group; ORX, orchidectomized group; SH, sham-operated group; TE, testosterone enanthate-supplemented group; MFI, mean fluorescence intensity.
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