Fig. 6: Endogenous formaldehyde reacts with GSH altering the GSH:GSSG ratio. | Nature Communications

Fig. 6: Endogenous formaldehyde reacts with GSH altering the GSH:GSSG ratio.

From: Endogenous formaldehyde scavenges cellular glutathione resulting in redox disruption and cytotoxicity

Fig. 6

a Scheme showing the spontaneous reaction between formaldehyde (FA) and glutathione (GSH) yielding S-hydroxymethylglutathione (HSMGSH). b HSMGSH content relative to GSH in Wild type (WT) and ∆ADH5 HCT116 cells (n = 9, two-tailed Mann–Whitney test, ****P < 0.0001). The box and whiskers plots: the line in the box corresponds to the median; the edges are the 25th and 75th percentiles, and the whiskers extend to the most extreme values. c Net HSMGSH content in WT and ∆ADH5 cells calculated as normalized peak areas relative to the number of viable cells (n = 9, two-tailed Mann–Whitney test, ****P < 0.0001). Box and whiskers plotted as (b). d GSH content in WT and ∆ADH5 HCT116 cells exposed to 150 μmol L−1 FA over 48 h (n = 9 for untreated (UT) WT and ∆ADH5 samples, n = 5 for WT and ∆ADH5 FA-treated samples, two-tailed Mann–Whitney test, ***P = 0.001, ****P < 0.0001). Box and whiskers plotted as (b). e GSH disulfide (GSSG) content in WT and ∆ADH5 HCT116 cells exposed to 150 μmol L−1 FA over 48 h (n = 9, WT and ∆ADH5 UT, n = 5 WT and ∆ADH5 FA-treated samples, two-tailed Mann–Whitney test, **P = 0.002 (WT vs WT + FA), **P = 0.0055 (∆ADH5 vs ∆ADH5 + FA), n.s.: not significant). Box and whiskers plotted as (b). f Total GSH (GSSG plus GSH) content in WT and ∆ADH5 HCT116 cells exposed to 150 μmol L−1 FA over 48 h (n = 9 for untreated WT and ∆ADH5 samples, n = 5 for WT and ∆ADH5 FA-treated samples, two-tailed Mann–Whitney test, ***P = 0.001 (WT vs WT + FA), ***P = 0.001 (∆ADH5 vs ∆ADH5 + FA), ****P < 0.0001 (WT vs ∆ADH5)). Box and whiskers plotted as (b). g. GSH:GSSG ratio in WT and ∆ADH5 HCT116 cells (n = 9 for untreated WT and ∆ADH5 samples, n = 5 for WT and ∆ADH5 FA-treated samples, two-tailed Mann–Whitney test, ****P < 0.0001 (WT vs ∆ADH5), ***P = 0.001 (∆ADH5 vs ∆ADH5 + FA). Box and whiskers plotted as (b). h Flow cytometry representative plots from WT and ∆ADH5 HCT116 cells harbouring the cytosolic Grx1-roGFP2 reporter. i Quantification of oxidized Grx1-roGFP2 [OxD (Grx1-roGFP2)] (n = 4, mean ± SEM, one-way ANOVA, Tukey’s test). Left: Cytoplasmic Grx1-roGFP2 (∆ADH5 0 vs ∆ADH5 + 150 µmol L−1 FA: *P = 0.0186). Right: Mitochondrial Grx1-roGFP2 (WT vs WT + 150 µmol L−1 FA: *P = 0.023, ∆ADH5 0 vs ∆ADH5 + 150 µmol L−1 FA: **P = 0.002).

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