Curcumin inhibits the proliferation and migration of osteosarcoma by regulating the expression of super-enhancer-associated genes.

Abstract

Objectives: Super-enhancer-associated genes may be closely related to the progression of osteosarcoma, curcumin exhibits a certain inhibitory effect on tumors such as osteosarcoma. This study aims to investigate the effects of curcumin on osteosarcoma in vitro and in vivo, and to determine whether curcumin can inhibit the progression of osteosarcoma by suppressing the expression of super-enhancer-associated genes LIM and senescent cell antigen-like-containing domain 1 (LIMS1), secreted protein acidic and rich in cysteine (SPARC), and sterile alpha motif domain containing 4A (SAMD4A).

Methods: Human osteosarcoma cell lines (MG63 cells or U2OS cells) were treated with 5 to 50 μmol/L curcumin for 24, 48, and 72 hours, followed by the methyl thiazolyl tetrazolium (MTT) assay to detect cell viability. Cells were incubated with dimethyl sulfoxide (DMSO) or curcumin (2.5, 5.0 μmol/L) for 7 days, and a colony formation assay was used to measure in vitro cell proliferation. After treatment with DMSO or curcumin (10, 15 μmol/L), a scratch healing assay and a transwell migration assay were performed to evaluate cell migration ability. Real-time reverse transcription polymerase chain reaction (real-time RT-PCR) and Western blotting were used to detect mRNA and protein expression levels of LIMS1, SPARC, and SAMD4A in the cells. An osteosarcoma-bearing nude mouse model was established, and curcumin was administered via gavage for 14 days to assess the impact of curcumin on tumor volume and weight in vivo. Real-time RT-PCR was used to measure mRNA expression levels of LIMS1, SPARC, and SAMD4A in the cancer and adjacent tissues from 12 osteosarcoma patients.

Results: After treating cells with different concentrations of curcumin for 24, 48, and 72 hours, cell viability were all significantly decreased. Compared with the DMSO group, the colony formation rates in the 2.5 μmol/L and 5.0 μmol/L curcumin groups significantly declined (both P<0.01). The scratch healing assay showed that, compared with the DMSO group, the migration rates of cells in the 10 μmol/L and 15 μmol/L curcumin groups were significantly reduced. The exception was the 10 μmol/L curcumin group at 24 h, where the migration rate of U2OS cells did not show a statistically significant difference (P>0.05), while all other differences were statistically significant (P<0.01 or P<0.001). The transwell migration assay results showed that the number of migrating cells in the 10 μmol/L and 15 μmol/L curcumin groups was significantly lower than that in the DMSO group (both P<0.001). In the in vivo tumor-bearing mouse experiment, the curcumin group showed a reduction in tumor mass (P<0.01) and a significant reduction in tumor volume (P<0.001) compared with the control group. Compared with the DMSO group, the mRNA expression levels of LIMS1, SPARC, and SAMD4A in the 10 μmol/L and 15 μmol/L curcumin groups were significantly down-regulated (all P<0.05). Additionally, the protein expression level of LIMS1 in U2OS cells in the 10 μmol/L curcumin group was significantly lower than that in the DMSO group (P<0.05). Compared with adjacent tissues, the mRNA expression level of SPARC in osteosarcoma tissues was significantly increased (P<0.001), while the mRNA expression levels of LIMS1 and SAMD4A did not show statistically significant differences (both P>0.05).

Conclusions: Curcumin inhibits the proliferation and migration of osteosarcoma both in vitro and in vivo, which may be associated with the inactivation of super-enhancer-associated gene LIMS1.