Lycopene Promotes Osteogenesis and Reduces Adipogenesis through Regulating FoxO1/PPARγ Signaling in Ovariectomized Rats and Bone Marrow Mesenchymal Stem Cells

Abstract

Recent interest in preventing the development of osteoporosis has focused on the regulation of redox homeostasis. However, the action of lycopene (LYC), a strong natural antioxidant compound, on osteoporotic bone loss remains largely unknown. Here, we show that oral administration of LYC to OVX rats for 12 weeks reduced body weight gain, improved lipid metabolism, and preserved bone quality. In addition, LYC treatment inhibited ROS overgeneration in serum and bone marrow in OVX rats, and in BMSCs upon H₂O₂ stimulation, leading to inhibiting adipogenesis and promoting osteogenesis during bone remodeling. At the molecular level, LYC improved bone quality via an increase in the expressions of FoxO1 and Runx2 and a decrease in the expressions of PPARγ and C/EBPα in OVX rats and BMSCs. Collectively, these findings suggest that LYC attenuates osteoporotic bone loss through promoting osteogenesis and inhibiting adipogenesis via regulation of the FoxO1/PPARγ pathway driven by oxidative stress, presenting a novel strategy for osteoporosis management.

Keywords: BMSCs; FoxO1/PPARγ signaling; OVX rats; adipogenesis; lycopene; osteogenesis.

Figure 1

The flow chart for animal experiments.

Figure 2

Lycopene preserves bone micro-architecture, strength, and material properties in OVX rats. Representative images of H&E (A) and μCT scanning (B) in the femurs of the different groups of rats. The BMD (C), BV/TV (D), BS/TV (1/mm) (E), Tb.N (1/mm) (F), Tb.Th (mm) (G), Tb.Sp (mm) (H), Conn.D (1/mm³) (I), and DA (J) in the femoral metaphysis were analyzed by the Analyzer Software. Elastic Modulus/d (K), Bending strength/d (L), Ultimate Load/d (M), and carbonate to phosphate (N) were determined by a three-point bending assay and Fourier-Transform Infrared Spectroscopy (FTIR), respectively. Data are presented as mean ± SD. The black arrow in panel (A) denotes lipid droplet. SHAM denotes the sham operation group, OVX denotes the ovariectomized model group, EV denotes the estradiol treatment group, LYCH denotes the high-dose lycopene treatment group, and LYCL denotes the low-dose lycopene treatment group. n = 5, ^# vs. the SHAM group, * vs. the OVX group. p < 0.05 was considered statistically significant.

Figure 3

Lycopene inhibits oxidative stress in OVX rats and in BMSCs. Serum and bone marrow levels of oxidative stress markers were determined by biochemical assays, including serum total antioxidant capacity (Serum T-AOC, (A)), serum superoxide dismutase (Serum SOD, (B)), malondialdehyde (Serum MDA, (C)), bone marrow total antioxidant capacity (BM-T-AOC, (D)) and bone marrow superoxide dismutase (BM-SOD, (E)). The primary BMSCs were characterized by flow cytometry (F). Effects of lycopene on BMSCs proliferation with/without H₂O₂ exposure after 24, 48, and 72 h (G–I) were determined by a CCK-8 assay. The intracellular levels of ROS were determined by the DCFH-DA (J). SHAM denotes the sham operation group, OVX denotes the ovariectomized model group, EV denotes the estradiol treatment group, LYCH denotes the high-dose lycopene treatment group, LYCL denotes the low-dose lycopene treatment group. CON denotes the blank control, H₂O₂ denotes the stimulation with H₂O₂ for 1 h, LYC2 denotes the 2 μM of lycopene treatment, LYC4 denotes 4 μM of lycopene treatment. n = 5, ^# vs. the SHAM or CON group, * vs. the OVX or H₂O₂ group. p < 0.05 was considered statistically significant.

Figure 4

Lycopene improves lipid metabolism in OVX rats and in BMSCs. Body weight (A), serum triacylglycerols (Serum TAGs, (B)), serum total cholesterol (Serum TC, (C)), serum low-density lipoprotein (Serum LDL, (D)), serum high-density lipoprotein (Serum HDL, (E)), bone marrow TAGs (BM-TAGs, (F)) and bone marrow TC (BM-TC, (G)). The Oil Red O staining and its analysis in BMSCs (I,H). SHAM denotes the sham operation group, OVX denotes the ovariectomized model group, EV denotes the estradiol treatment group, LYCH denotes the high-dose lycopene treatment group, LYCL denotes the low-dose lycopene treatment group. CON denotes the blank control, H₂O₂ denotes the stimulation with H₂O₂ for 1 h, LYC2 denotes 2 μM of lycopene treatment, LYC4 denotes 4 μM of lycopene treatment. n = 5 (rats), n = 3 (cells), ^# vs. the SHAM or CON group, * vs. the OVX or H₂O₂ group. p < 0.05 was considered statistically significant.

Figure 5

Lycopene promotes osteogenesis in OVX rats and in BMSCs. Serum levels of PINP (A) and CTX-I (B). (C,D) The representative images of Safranin O/fast green staining and their analyses show the GAG levels in the femurs. The representative images of alizarin red S staining and their analyses show the osteogenesis and calcium nodules in the different groups of rats (E,F) and BMSCs (G,H). Blue star denotes calcium nodules (E). SHAM denotes the sham operation group, OVX denotes the ovariectomized model group, EV denotes the estradiol treatment group, LYCH denotes the high-dose lycopene treatment group, LYCL denotes the low-dose lycopene treatment group. CON denotes the blank control, H₂O₂ denotes the stimulation with H₂O₂ for 1 h, LYC2 denotes 2 μM of lycopene treatment, LYC4 denotes 4 μM of lycopene treatment. n = 5 (rats), n = 3 (cells), ^# vs. the SHAM or CON group, * vs. the OVX or H₂O₂ group. p < 0.05 was considered statistically significant.

Figure 6

Lycopene increases FoxO1, Runx2, and OCN expressions, and inhibits PPARγ and C/EBPα expressions in the femurs and tibias of OVX rats and in BMSCs. The expressions of FoxO1, PPARγ, Runx2, OCN, and C/EBPα in the femurs and tibias were determined by immunohistochemical staining (A,B,D,E,G,H,J,K) and/or western blot (C,F,I,L). The expressions of FoxO1, Runx2, and PPARγ in BMSCs were determined by western blot (M–O). SHAM denotes the sham operation group, OVX denotes the ovariectomized model group, LYCL denotes the low-dose lycopene treatment group. CON denotes the blank control, H₂O₂ denotes the stimulation with H₂O₂ for 1 h, LYC2 denotes 2 μM of lycopene treatment, AS1842856 denotes FoxO1 inhibitor, AS1842856+LYC2 denotes the co-treatment of FoxO1 inhibitor and 2 μM of lycopene. n = 5 (rats), n = 3 (cells), ^# vs. the SHAM or CON group, * vs. the OVX or H₂O₂ group. p < 0.05 was considered statistically significant.