Expression of miR-664-3p in Osteosarcoma and Its Effects on the Proliferation and Apoptosis of Osteosarcoma Cells

Abstract

Background: To explore the expression level of miR-664-3p in osteosarcoma and its effects on the proliferation and apoptosis of osteosarcoma cells.

Methods: Specimens of osteosarcoma tissues were collected from 41 cases undergoing surgical treatment in the Orthopedics Department of Wuhan Puai Hospital, Wuhan, China from January 2015 to February 2018. Another 40 cases of normal bone tissue were collected. The expression of miR-664-3p were detected using quantificational real-time polymerase chain reaction. miR-664-3p mimics, miR-664-3p inhibitor and miR-664-3p negative control (NC) were used to transfect U2-OS, which were named as mimics group, inhibitor group and NC group, respectively. MTT assay was adopted to detect the effects of microRNA-664-3p on the proliferation of U2-OS after 24, 48, 72, 96 and 120 hours of transfection. Flow cytometry was applied to measure the apoptosis rate of U2-OS after miR-664-3p transfection. Finally, Western Blot was employed to detect the expression of proteolipid protein 2 (PLP2).

Results: The total apoptosis rate of cells in the inhibitor group was obviously higher than those in the mimics group and the NC group (P<0.001). The relative expression level of PLP2 in the inhibitor group was significantly lower than those in the mimics group and the NC group (P<0.001).

Conclusion: MiR-664-3p may be involved in the occurrence and development of osteosarcoma, and can regulate the proliferation and apoptosis of U2-OS cells, and the expression of PLP2. Besides, miR-664-3p may become a novel molecular biological indicator for the diagnosis, targeted treatment and prognosis assessment of osteosarcoma.

Keywords: Apoptosis; Osteosarcoma; Proliferation; U2-OS; miR-664-3p.

Fig. 1:

Prediction of miR-664-3p related target genes

Fig. 2:

Expressions of miR-664-3p in osteosarcoma tissues and normal bone tissues. The expression level of miR-664-3p in osteosarcoma tissues was significantly higher than that in normal bone tissues (t=19.680, P<0.001). Note: ^****P<0.001

Fig. 3:

Expression of miR-664-3p in osteosarcoma cells. the relative expression levels of miR-664-3p in SOSP-9607 and U2-OS were significantly higher than that in hFOB1.19. The relative expression level of miR-664-3p in U2-OS was significantly higher than that in SOSP-9607 (P<0.001) Note: ^****P<0.001

Fig. 4:

Expression of miR-664-3p in U2-OS cells after transfection The results of qRT-PCR showed that the relative expression of miR-664-3p in U2-OS cells in the mimics group was significantly higher than that in the inhibitor group and the NG group (p<0.001), and the relative expression of miR-664-3p in U2-OS cells in the NG group was significantly higher than that in the inhibitor group (P<0.001). Note: ^****P<0.001

Fig. 5:

Effects of miR-664-3p expression on the proliferation of osteosarcoma cell line U2-OS. The results of MTT assay manifest that the OD value detected in the mimics group is obviously higher than that in NC group from 48 h (P<0.001). The OD value detected in the inhibitor group is significantly lower than those in the mimics group and the NC group from 48 h (P<0.001). Note: ^****P<0.001 vs. mimics group

Fig. 6:

Effects of the expression of miR-664-3p on the apoptosis of osteosarcoma cell line U2-OS. The results of flow cytometry indicate that the total apoptosis rate in the inhibitor group is significantly higher than those in the mimics group and the NC group The total apoptosis rate in the mimics group is significantly lower than that in the NC group (P<0.001). Note: ^****P<0.001

Fig. 7:

Effects of the expression of miR-664-3p on the expression of PLP2 in osteosarcoma cell line U2-OS. Western Blot results reveal that the relative expression level of PLP2 in the inhibitor group is obviously lower than those in the mimics group and the NC group. The relative expression level of PLP2 in the mimics group is remarkably higher than those in the NC group (P<0.001). Note: ^****P<0.001