Curcumin improves osteoporosis in type 2 diabetic mice by inhibiting RANBP3L expression

doi:10.12122/j.issn.1673-4254.2026.02.10

Abstract

Abstract:

Objective To investigate the therapeutic mechanism of curcumin for osteoporosis induced by type 2 diabetes mellitus (T2DM). Methods In cultured MC3T3-E1 osteoblasts exposed to high glucose (HG), the effects of curcumin treatment on osteogenic differentiation and mineralization were examined by assessing alkaline phosphatase (ALP) activity and using alizarin red S (ARS) staining. RNA-seq sequencing was used to analyze the transcriptional characteristics of MC3T3-E1 osteoblasts in HG culture after curcumin treatment. In a mouse model of T2DM established by high-fat diet feeding and streptococcal injection, the effects of solvent vehicle, vehicle+RANBP3L knockdown, and RANBP3L knockdown+curcumin treatment on blood lipids, bone microstructure and calcium deposition was evaluated using ELISA, HE staining, and Alizarin red S staining. Osteogenic differentiation ability of mouse femoral tissues were assessed by detecting ALP, RANBP3L, OCN and RANKL expressions using immunohistochemical staining, RT-qPCR and Western blotting, and the changes in the TNF‑α/ NF‑κB pathway were detected with Western blotting. Results Curcumin obviously promoted proliferation and osteogenic differentiation of MC3T3-E1 cells in HG culture and down-regulated cellular expression of RANBP3L protein. In the diabetic mouse models, RANBP3L knockdown significantly improved bone microstructure and blood lipid balance, increased expression levels of osteogenic markers, decreased blood glucose level, and caused inhibition of the TNF‑α/NF‑κB signaling pathway. The combined treatment with curcumin further reduced RANBP3L expression levels in the bone tissue and significantly enhanced the therapeutic effect. Conclusion Curcumin promotes osteogenesis, alleviates glucose and lipid metabolism disorders, and improves osteoporosis in type 2 diabetic mice possibly by inhibiting TNF‑α/NF‑κB signaling pathway via suppressing RANBP3L expression.

Key words: curcumin, RANBP3L, type 2 diabetes osteoporosis, mRNA sequencing

Xiaosong ZOU, Xing ZHANG, Ping LI, Ruixue TIAN, Xiaomiao LU. Curcumin improves osteoporosis in type 2 diabetic mice by inhibiting RANBP3L expression[J]. Journal of Southern Medical University, 2026, 46(2): 325-334.

Figures/Tables 7

Tab.1 Primer sequences for RT-qPCR

Gene	Primer sequences (5'to3')	Size (bp)
GAPDH-m	F:AGGTCGGTGTGAACGGATTTG	95
GAPDH-m	R:GGGGTCGTTGATGGCAACA	95
RANBP3L	F:GAGAAAGCGTGTAAGGTCTTCA	82
RANBP3L	R:CGTTTGAGACAAAGTTGCCAC	82
OCN	F:AGTGTGAGCTTAACCCTGCT	156
OCN	R:GAGGATCAAGTCCCGGAGAG	156
RANKL	F:CGCTCTGTTCCTGTACTTTCG	114
RANKL	R:GAGTCCTGCAAATCTGCGTT	114

Fig.1 Curcumin promotes proliferation and osteogenic differentiation of MC3T3-E1 cells in high-glucose (HG) culture. A： CCK-8 assay for assessing cell viability following HG culture and treatment with different concentrations of curcumin. B： ALP staining of the cells after curcumin treatment for 7 days (Scale bar=50 μm). C： Alizarin red staining of the cells after curcumin treatment for 14 days. D： Relative area of ALP staining (Scale bar=50 μm). E： Relative area of Alizarin red staining. n=3. *P<0.05 vs HG group, **P<0.01, ***P<0.001 vs Control; ##P<0.01, ###P<0.001 vs HG.

Fig.2 Curcumin reduces RANBP3L expression in MC3T3-E1 cells cultured in high glucose. A: Volcano plot showing the distribution of differentially expressed mRNAs. B: KEGG pathway analysis of the pivotal signaling pathways within the differentially expressed genes. C: mRNA expression of RANBP3L detected by qRT-PCR. D, E: Protein expression of RANBP3L detected by Western blotting. n=3. *P<0.05, ***P<0.001 vs Control; ##P<0.01, ###P<0.001 vs HG.

Fig.3 Blood glucose and lipid profiles of the mice in each group. A: Schematic diagram of the experimental process. B: Fasting blood glucose of the mice after STZ injection. C-F: TC, TG, LDL-C and HDL-C at the 17rd week. n=5. **P<0.01, ***P<0.001 vs Ctrl+Veh; ##P<0.01, ###P<0.001 vs T2DM+Veh; ‡P<0.05, ‡‡P<0.01 vs T2DM+KD+Veh.

Fig.4 Diabetic mice show reduced bone formation. A： HE staining of the bone tissues (Scale bar=200 μm). B： Alizarin Red Staining of the bone tissues (Scale bar=200 μm). C, D： Immunohistochemistry for ALP and RANBP3L （Scale bar=100 μm）. E-G： mRNA expression of RANBP3L, OCN and RANKL detected by qRT-PCR. H-J： Protein expressions of OCN and RANKL detected by Western blotting. n=5. ***P<0.001 vs Ctrl+Veh.

Fig.5 Curcumin down-regulates RANBP3L expression and promotes bone formation in T2DM mice. A： HE staining of the bone tissues (Scale bar=200 μm). B： Alizarin Red Staining of the bone tissues (Scale bar=200 μm). C-F： Immunohistochemistry for ALP and RANBP3L and quantitative histograms (Scale bar =100 μm). G, I and J： mRNA expressions of RANBP3L, OCN and RANKL detected by qRT-PCR. H, K and L： Protein expressions of OCN and RANKL detected by Western blotting. n=5. **P<0.01, ***P<0.001 vs Ctrl+Veh; #P<0.05, ##P<0.01, ###P<0.001 vs T2DM+Veh; ‡P<0.05, ‡‡P<0.01, ‡‡‡P<0.001 vs T2DM+KD+Veh.

Fig.6 Effects of RANBP3L knockdown and curcumin treatment on the TNF‑α/NF‑κB signaling pathway. A-E: Protein expressions of TNF-α, p-p65, p-IKKβ and p-IκB-α detected by Western blotting. n=5. ***P<0.001 vs Ctrl+Veh; #P<0.05, ##P<0.01 vs T2DM+Veh; ‡P<0.05, ‡‡‡P<0.001 vs T2DM+KD+Veh.

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