Expression of STOML2 promotes proliferation and glycolysis of multiple myeloma cells via upregulating PAI-1

Abstract

Background: This study aimed to investigate the effects of STOML2 and the relationship between STOML2 and PAI-1 in the development of multiple myeloma (MM).

Methods: Cell proliferation was tested using CCK-8 assay and cell colony formation assay. Glucose consumption, lactate production and ATP level were measured using commercial kits. The mRNA and protein expression were assessed using quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting, respectively.

Results: Both mRNA and protein expression of STOML2 were upregulated in MM patients compared to healthy volunteers. CCK-8 and colony formation assays demonstrated that STOML2 silencing inhibited cell proliferation in MM cells. Knockdown of STOML2 reduced glucose consumption, lactate production and ATP/ADP ratios. STOML2 silencing by shSTOML2 led to reduced PAI-1 expression. Overexpression of PAI-1 reversed the inhibitory effects of shSTOML2 on MM cell growth.

Conclusion: Results from this study demonstrated that STOML2 silencing inhibits cell proliferation and glycolysis through downregulation of PAI-1 expression, suggesting a new therapeutic target for MM.

Keywords: Cell proliferation; Glycolysis; Multiple myeloma; PAI-1; STOML2.

Fig. 1

STOML2 is overexpressed in MM sera and cell lines. A The expression of STOML2 mRNA is upregulated in MM patients; B the expression of STOML2 protein is upregulated in MM patients; C the expression of STOML2 mRNA is upregulated in MM cell lines; D the expression of STOML2 protein is upregulated in MM cell lines; E statistical analysis shows that the expression of STOML2 protein is upregulated in MM cell lines. *p < 0.05 versus (vs) nPCs; **p < 0.01 vs Normal or nPCs; ***p < 0.005 vs Normal or nPCs. MM: multiple myeloma

Fig. 2

Knockdown of STOML2 inhibits MM cell growth. A Protein expression of STOML2 is reduced by shSTOML2-1# and shSTOML2-2# in MM cell lines; B MM cell viability is reduced by shSTOML2-1# and shSTOML2-2#; C the number of cell colonies is reduced by shSTOML2-1# and shSTOML2-2# in MM cell lines; D the activity of Caspase-3 is increased by shSTOML2-1# and shSTOML2-2# in MM cell lines; E the protein expression of Bcl2 and PCNA is downregulated, and the protein expression of Bax and Clv-PARP is upregulated by shSTOML2-1# and shSTOML2-2# in MM cell lines. *p < 0.05 vs shNC; **p < 0.01 vs shNC. shSTOML2: short hairpin RNAs of STOML2; shNC: scrambled RNA of shSTOML2

Fig. 3

Knockdown of STOML2 represses abnormal glycolysis in MM. A The protein expression of GLUT1, HK2 and LDHA is downregulated by shSTOML2-1# and shSTOML2-2# in MM cell lines; B statistical analysis shows that the protein expression of GLUT1, HK2 and LDHA is downregulated by shSTOML2-1# and shSTOML2-2# in MM cell lines; C the relative glucose consumption, lactate production and ATP/ADP ratio are reduced by shSTOML2-1# and shSTOML2-2# in MM cell lines. *p < 0.05 vs shNC; **p < 0.01 vs shNC

Fig. 4

Expression of PAI-1 is regulated by STOML2 in MM. A The mRNA expression of PAI-1 is downregulated y shSTOML2-1# and shSTOML2-2# in MM cell lines; B the protein expression of PAI-1 is downregulated y shSTOML2-1# and shSTOML2-2# in MM cell lines. *p < 0.05 vs shNC; **p < 0.01 vs shNC

Fig. 5

PAI-1 reverses STOML2 silencing-mediated tumor cell growth inhibition in MM. A PAI-1 reverses shSTOML2-mediated cell proliferation inhibition; B statistical analysis shows that PAI-1 reverses shSTOML2-mediated cell proliferation inhibition; C PAI-1 reverses shSTOML2-mediated increase in Caspase-3 activity; D PAI-1 reverses shSTOML2-mediated decrease in GLUT1, HK2 and LDHA; E–G Statistical analyses show that PAI-1 reverses shSTOML2-mediated downregulation of GLUT1 E, HK2 F, and LDHA G. **p < 0.01 vs shNC; ^@p < 0.05 vs shSTOML2-2# + NC; ^@@p < 0.01 vs shSTOML2-2# + NC; ^#p < 0.05 vs PAI-1 + shNC; ^##p < 0.05 vs PAI-1 + shNC. PAI-1: pcDNA3.1 plasmid containing full coding sequence of PAI-1; NC: scrambled sequence of PAI-1