Fig 2: MiR-106b-5p targeting to HPGD promoted cell cycle progression and suppressed cell apoptosis of ESCC cells. A Cell cycle was detected in KYSE450 and KYSE510 cells transfected with NC, OE-HPGD, miR-106b-5p mimic, and OE-HPGD+miR-106b-5p mimic by flow cytometry assay. B Cell apoptosis rate was detected in KYSE450 and KYSE510 cells transfected with NC, OE-HPGD, miR-106b-5p mimic, and OE-HPGD+miR-106b-5p mimic by flow cytometry assay. C Cell apoptosis was determined in KYSE450 and KYSE510 cells transfected with NC, OE-HPGD, miR-106b-5p mimic, and OE-HPGD+miR-106b-5p mimic by caspase-3/7 activity assay kit. D The protein expression of Bax and Bcl-2 were determined in KYSE450 and KYSE510 cells transfected with NC, OE-HPGD, miR-106b-5p mimic, and OE-HPGD+miR-106b-5p mimic by western blot analysis. Data are presented as means± SD of at least three independent tests per experiment. *, P < 0.05; **, P < 0.001 compared to CON group. CON, blank control; NC, empty vectors+mimic-NC; OE-HPGD, overexpression-HPGD; OE-HPGD+miR-106b-5p mimic, overexpression-HPGD+ miR-106b-5p mimic

Fig 3: HPGD and miR-106b-5p were selected to be further investigate in ESCC by microarray analysis. A 85 DEGs were overlapped from GSE38129 and GSE17351 by Venny 2.1.0. GSE38129 and GSE17351 were the mRNA expression profiles. B The positive regulation of cell death containing 9 DEGs was screen out as the key progress by Metascape analysis. C The expression of HPGO and MAL in ESCA was significant reduced. D The expression of HPGO and MAL was detected in the ESCC tissue samples (n = 45) and normal tissues (n = 45) by qRT-PCR. E miR-106b-5p and miR-31-5p were overlapped from GSE114110, TargetScan, and starBase. GSE114110 was the miRNA expression microarray. TargetScan and starBase were used to predict the miRNAs targeting HPGD. F The expression of miR-106b-5p was significant down-regulated in ESCA by starBase analysis

Fig 4: HPGD was a direct target ofmiR-106b-5p. A Bioinformatics analysis showed the predicted binding sequence of HPGD 3’-UTR. B Dual luciferase assay was performed in cells co-transfected with plasmids HPGD-Wt or HPGD-Mut and miR-NC or miR-106b-5p mimic in KYSE450 and KYSE510 cells. **, P < 0.001 compared to co-transfection of HPGD-Wt and miR-NC group. C RNA pull down assay was used to detect the association between HPGD and miR-106b-5p in KYSE450 and KYSE510 cells. **, P < 0.001 compared to Bio-NC group. D RT-qPCR detection of expression of HPGD in Het-1A, KYSE450 and KYSE510 cells. **, P < 0.001 compared to Het-1A cells. E Western blot detection of protein expression of HPGD in Het-1A, KYSE450 and KYSE510 cells. Data are presented as means ± SD of at least three independent tests per experiment. *, P < 0.05; **, P < 0.001 compared to Het-1A cells. Wt, wild-type; Mut, mutant; NC, negative control

Fig 5: The expression levels of Hpgd in hPAECs treated with hypoxia for 0, 6, 12, 24 and 48 h. (A) Western Blot results showed that the expression of Hpgd decreased in hypoxia-treated hPAECs. (B) qRT-PCR showed that the longer the time of hypoxic treatment of hPAECs, the lower the expression of Hpgd. (*P<0.05, **P<0.01)