Fig. 1

LPA enhances MM cell resistance to PIs through LPAR2-mediated MEK1/2-ERK1/2 signal pathways and enhanced OXPHOS in mitochondria. a Levels of LPA in serum of normal healthy controls and MM patients. b Levels of LPA in serum of 5TGM1 or Vk*MYC MM-tumor free (Ctr) and tumor bearing (TB) mice. Ctr, control, MM-tumor free mice; TB: MM-tumor-bearing mice. c The primary MM cells isolated from BM of MM patients (n = 20) were divided into LPA^low and LPA^high groups based on their serum levels of LPA and treated with BTZ or CFZ for one hour, then the apoptosis was measured after 24-h incubation. d Human primary MM cells isolated from BM of MM patients were treated with BTZ or CFZ for one hour, after wash and 24-h incubation with or without 4 μg/mL LPA, the apoptosis of the cells was determined. e Relative mRNA expression of LPA receptors in CD138⁺ cells of MM patients and plasma cells of normal healthy controls from GSE5900 array data. Values were normalized with normal healthy controls. f Overall survival of MM patients with high (LPAR2^High) or low (LPAR2^Low) LPAR2 expression based on published Oncomine data (GSE9782). g Surface expression of different LPA receptors on human primary MM cells isolated from BM of MM patients (n = 10). h Ctr-KO and LPAR2-KO ARP1 or MM.1S cells were pulsed with BTZ or CFZ for 1-h and then incubated with or without 4 μg/mL LPA for 24 h. The apoptosis of the cells were determined. Ctr-KO, MM cells transfected with lentivirus containing empty vector; LPAR2-KO, MM cells transfected with lentivirus containing LPAR2 sgRNA. i, j Ctr-KO and LPAR2-KO ARP1 or MM.1S cells were treated without (PBS) or with 4 μg/mL LPA and phosphorylation level of MEK1/2 and ERK1/2 was determined by western blot. Mock, MM cells without treatment; Ctr-KO, MM cells transfected with lentivirus containing empty vector; LPAR2-KO, MM cells transfected lentivirus containing LPAR2 sgRNA. k Ctr-KO and LPAR2-KO ARP1 or MM.1S MM cells were pulsed with BTZ or CFZ for one hour, followed by wash and culture with kinase inhibitors PD184352 (PD, 5 μM) or SCH772984 (SCH, 20 μM) for 24 h in the present/absent 4 μg/mL LPA, then the apoptotic rates were determined. l IPA analysis of canonical signaling pathway in MM cells treated without (PBS) or with 4 μg/mL LPA. The circle surface area is proportional to -log (P value) and the color intensity of circles indicates the Z score. m GSEA result of GO_OXIDATIVE_PHOSPHORYLATION gene signatures. NES, normalized enrichment score; FDR, false discovery rate. n, o OCRs of Ctr-KO and LPAR2-KO ARP1 cells treated with or without LPA (n) and summarized result of the basal respiration, ATP-linked respiration, maximal respiration, and spare capacity for Ctr-KO and LPAR2-KO ARP1 and MM.1S cells treated with or without LPA (o). p ARP1 and MM.1S cells were pulsed with BTZ or CFZ for 1 h and the cells were washed and cultured with or without LPA for another 24 h. Representative summarized results of the basal respiration, maximal respiration, and spare capacity for ARP1 and MM.1S cells. q–s Relative production of NAD⁺ (q), ATP (r), and the relative proteasome activity (s) of Ctr-KO and LPAR2-KO ARP1, U266, and MM.1R cells treated with vehicle (PBS) or LPA (4 μg/mL) for 24 h. Results are shown as means ± S.E.M.. The survival rate was analyzed by log-rank (Mantel–Cox) test. *P < 0.05; **P < 0.01; ***P < 0.001; n.s., not significant