Fig 2: Sch A suppresses DON-induced heme oxygenase-1 (HO-1) mRNA and protein expression. HT-29 cells were treated with 1 µM of DON for 24 hours in the presence and absence of Sch A pre-treatment (2.5, 5 and 10 µM) for 24 hours. (A) HO-1 mRNA expression was measured by qPCR, with GAPDH as the internal control; (B) HO-1 protein expression was measured by Western blotting, with beta actin as the internal control. (C) Representative photos of western blotting of HO-1 and beta actin. Results were shown as mean of ± SEM (n = 6), which are at least six separate experiments. Different letters indicate significant differences at p < 0.05.

Fig 3: Effects of water extracts of djulis (WECF) on PM-induced Nrf2 and HO-1 protein expression in A549 cells. (A) Effects of WECF on PM-induced expression of Nrf2 in A549 cells. The cells were treated with WECF and exposed to 400 µg/mL PM for 12 h. # (p < 0.05) compared with the control group and * (p < 0.05) compared with 400 µg/mL PM-induced cells alone. (B) Effects of WECF on PM-induced HO-1 activity in A549 cells. The cells were treated with WECF and exposed to 400 µg/mL PM for 15 h. # (p < 0.05) compared with the control group and * (p < 0.05) compared with 400 µg/mL PM-induced cells alone. (C) Effects of p38 inhibitor (SB203580, SB), JNK inhibitor (SP600125, SP), and ERK inhibitor (U0126, U) on WECF-induced Nrf2 protein expression in PM-treated A549 cells. Control, cultured with medium alone for 12 h; PM, incubated with PM 400 µg/mL for 12 h; P + W, incubated with WECF and PM for 12 h (WECF was added 30 min before PM); SB + W + P, SP + W + P, and U + W + P, treated as described for P + W except that the p38 MAPK inhibitor SB203580, the JNK inhibitor SP600125, or the ERK inhibitor U0126 were added to the medium before WECF. # (p < 0.05) compared with the control group and * (p < 0.05) compared with WECF+ 400 µg/mL PM-induced cells. (D) Effects of p38 inhibitor (SB203580, SB), JNK inhibitor (SP600125, SP), and ERK inhibitor (U0126, U) on WECF-induced HO-1 protein expression in PM-treated A549 cells. Control, cultured with medium alone for 15 h; PM, incubated with PM 400 µg/mL for 15 h; P + W, incubated with WECF and PM for 15 h (WECF was added 30 min before PM); SB + W + P, SP + W + P, and U + W + P, treated as described for P + W except that the p38 MAPK inhibitor SB203580, the JNK inhibitor SP600125, or the ERK inhibitor U0126 were added to the medium before WECF. # (p < 0.05) compared with the control group and * (p < 0.05) compared with WECF+ 400 µg/mL PM-induced cells. Values in each sample with different lowercase letters are significantly different (p < 0.05). Data are presented as means ± SD (n = 3).

Fig 4: Expression of histone modification-related proteins in SNUC5 and SNUC5/5-FURA. Expression patterns of HDAC1, HAT1, and H3K9Ac in SNUC5 and SNUC5/5-FUR cells determined by Western blot analysis. *Significantly different from SNUC5 cells (p<0.05). B. Expression patterns of MLL, H3K4Me3, G9a, and H3K9Me2 in both cells determined by Western blot analysis. TBP antibody was used as loading control for nuclear fractions. *Significantly different from SNUC5 cells (p<0.05). The cells were transfected with non-targeting siRNA (siControl) or siMLL RNA or siHAT1 RNA for 24 h. C. Expression patterns of MLL, HAT1, Nrf2, and HO-1 in SNUC5/5-FUR determined by Western blot analysis. *Significantly different from untreated siControl-transfected cells (p<0.05).

Fig 5: HMOX1 product biliverdin suppresses SARS-CoV-2 replication. A A schematic diagram showing the heme oxygenase pathway. Vero-E6 cells were infected with recombinant GFP-SARS-CoV-2 at an MOI of 0.01. After incubation with virus for 1 h, the cells were treated with the indicated doses of Fe2+ (B) hemoglobin (HB) D or biliverdin (F) for 24 h. C, E, G The amount of viral RNA in the cell culture medium was quantified by real-time PCR assay. H Fluorescence microscopy images of GFP-SARS-CoV-2 in biliverdin-treated ACE2-HeLa cells. Blue: DAPI (nuclear staining). I The dose response curve of biliverdin against SARS-CoV-2. HeLa-ACE2 cells were infected with recombinant GFP-SARS-CoV-2. After incubation with virus for 1 h, the cells were treated with biliverdin at different concentrations for an additional 24 h. The viral NP expression in the SARS-CoV-2-infected cells was quantified by real-time PCR and normalized to the expression of the 36B4 gene. The results are expressed as the mean ± SD (error bar) of 3 independent experiments; asterisks represent statistical significance based on two-tailed unpaired Student’s t test (*P < 0.05, **P < 0.01)