ABSTRACT
Objective To investigate the effect of knock⁃down 3⁃phosphoglycerin dehydrogenase (PHGDH) targeting energy metabolism on malignant biological behavior and osteogenic differentiation of human osteosarcoma 143B cells. Methods Real⁃time PCR and Western blotting were used to detect the expression of PHGDH in osteoblasts hFOB1. 19 and osteosarcoma cells TE85, MG63 and 143B with different malignant degrees. The short hairpin RNA (shRNA)⁃PHGDH recombinant plasmid was transfected into 143 B cells by liposome transfection method. The expression of PHGDH was detected by Real⁃time PCR and Western blotting. Crystal violet staining, cell counting and CCK⁃8 assay were used to detect cell proliferation; wound healing assay was used to detect cell parallel migration ability, and Transwell assay was used to detect cell vertical migration and invasion ability. Annexin V⁃FITC/ PI double staining and DAPI staining were used to detect apoptosis; Alkaline phosphatase(ALP) staining and alizarin red S staining were used to detect osteogenic differentiation. Western blotting was used to detect the expression of Runt related transcription factor 2 (Runx2) and osteocalcin (OC) . The expression of genes related to energy metabolism, glucose transporter⁃1 (GLUT1), 6⁃ phosphofructokinase⁃1(PFK1), pyruvate kinae subtype M2 (PKM2), lactate dehydrogenase A (LDHA) was detected by Real⁃time PCR. Lactic acid secretion was detected by lactic acid detection kit. Adenosine triphosphate(ATP) production was detected by ATP detection kit. Results The expression of PHGDH in 143B cells was significantly higher than that in hFOB1. 19, MG63 and TE85 cells (P < 0. 01) . After the transfection of shRNA⁃PHGDH recombinant plasmid, the expression of PHGDH in 143 B cells decreased (P<0. 01), proliferation ability decreased (P<0. 01), cell migration and invasion ability decreased (P < 0. 01), apoptosis rate increased (P < 0. 01), ALP staining positive rate increased (P < 0. 01), alizarin red staining positive rate increased (P < 0. 05), Runx2 (P < 0. 05) and OC expression increased (P < 0. 01), expression of genes related to energy metabolism (GLUT1, PFK1, PKM2, LDHA) decreased (P < 0. 01), lactic acid decreased (P < 0. 01), ATP increased (P < 0. 05) . Conclusion Knocking down of PHGDH can inhibit the proliferation, migration and invasion of human osteosarcoma 143B cells through energy metabolism, promote their apoptosis and promote their osteogenic differentiation.