Research ArticleHEALTH AND MEDICINE

The essential role of the transporter ABCG2 in the pathophysiology of erythropoietic protoporphyria

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Science Advances  18 Sep 2019:
Vol. 5, no. 9, eaaw6127
DOI: 10.1126/sciadv.aaw6127
  • Fig. 1 Essential role of ABCG2 in EPP-associated phototoxicity.

    (A) Genotyping results of WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice. Fech-mut/Abcg2-null mice are deficient in both Fech and Abcg2. (B) Gross appearance of mice after light exposure. The back skin of mice was shaved and exposed to UV light (395 to 410 nm) for 30 min each day for 5 days. (C to F) Histologic sections of mouse skin after exposure to UV light, hematoxylin and eosin (H&E) staining. Scale bars, 40 μm. (G) PPIX levels in RBCs. (H) PPIX levels in serum. (I) PPIX levels in the skin of mice after exposure to UV light. PPIX was analyzed by UPLC-QTOFMS. All data are expressed as means ± SEM (n = 4 per group). *P < 0.05, ***P < 0.001, one-way ANOVA. (J) A schematic showing that deficiency of ABCG2 decreases PPIX distribution to the skin and abrogates PPIX-mediated phototoxicity in EPP. (Photo credit: Pengcheng Wang, University of Pittsburgh.)

  • Fig. 2 EPP-associated hepatotoxicity is dependent on ABCG2.

    WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice were kept under the same environment and euthanized at a similar age. (A to C) Serum activities of alanine transaminase (ALT), aspartate transaminase (AST), and alkaline phosphatase (ALP). (D) Serum total bilirubin. (E) PPIX in the liver, analyzed by UPLC-QTOFMS. The data are expressed as means ± SEM (n = 4 per group). ***P < 0.001, one-way ANOVA. (F to I) Representative liver sections with H&E staining. Arrows indicate bile plugs. Scale bars, 10 μm. PV, portal vein; BD, bile duct.

  • Fig. 3 PPIX accumulation and hepatotoxicity in WT and Abcg2-null mice treated with DDC.

    (A to C) Serum activities of ALT, AST, and ALP. (D) PPIX in the liver, analyzed by UPLC-QTOFMS. All data are expressed as means ± SEM (n = 3 to 4 per group). **P < 0.01, ***P < 0.001, two-way ANOVA. (E and F) Histologic analysis of the liver with H&E staining. Arrows point to bile plugs. Scale bars, 10 μm.

  • Fig. 4 PPIX accumulation and hepatotoxicity in hPXR and hPXR/Abcg2-null mice treated with RIF and INH.

    (A) Genotyping results of hPXR and hPXR/Abcg2-null mice. (B and C) Serum activities of ALT and ALP. (D) PPIX in the liver, analyzed by UPLC-QTOFMS. All data are expressed as means ± SEM (n = 3 to 4 per group). *P < 0.05, **P < 0.01, two-way ANOVA. (E and F) Histologic analysis of liver with H&E staining. Arrows point to bile plugs.

  • Fig. 5 Deficiency of ABCG2 modulates PPIX distribution, metabolism, and excretion.

    (A to F) Metabolomic analyses in WT and Abcg2-null mice treated with deuterium-labeled aminolevulinic acid (D2-ALA), a precursor of PPIX. Liver and bile samples were collected at 1 hour after D2-ALA treatment. (A) Score plots of liver samples generated by principal components analysis. Each point represents a mouse sample. (B and C) S-plots of liver and bile metabolome generated by orthogonal partial least-squares discriminant analysis. Each point represents a metabolite. All metabolites were analyzed by UPLC-QTOFMS. (D) D16-PPIX in the bile. (E and F) D16-protoporphyrin-1-O-acyl-glucouronide (D16-PPIX-glu) in the liver and bile. All the data are expressed as means ± SEM. The data in WT were set as 100% or 1. *P < 0.05, **P < 0.01, two-tailed Student’s t test. (G) The structures of PPIX and its conjugated metabolites with glucuronic acid, xylose, and glucose. (H) The percentages of PPIX and its conjugated metabolites in the bile of Fech-mut and Fech-mut/Abcg2-null mice. (I) A schematic showing deficiency of ABCG2 abolishes EPP-associated liver injury by modulating PPIX distribution, metabolism, and excretion.

  • Fig. 6 A summary of the roles of ABCG2 in the pathophysiology of EPP.

    ABCG2 is expressed in RBCs and hepatocytes. (A) A schematic showing ABCG2 drives phototoxicity and hepatotoxicity in EPP by (1) increasing PPIX distribution to the skin and increasing photosensitivity; (2) increasing PPIX delivery to the hepatobiliary system and causing bile duct blockage and cholestatic liver injury; and (3) ABCG2-dependent bile duct blockage further increases PPIX accumulation in the body, which, in turn, potentiates both phototoxicity and hepatotoxicity. (B) A schematic showing deficiency of ABCG2 abolishes phototoxicity and hepatotoxicity in EPP by (1) decreasing PPIX distribution to the skin and decreasing photosensitivity; (2) decreasing PPIX delivery to the hepatobiliary system and relieving PPIX-mediated bile duct blockage; (3) the retained PPIX in hepatocytes can be further metabolized to conjugated products to facilitate their excretion; and (4) prevention of PPIX-mediated bile duct blockage decreases PPIX accumulation in the body and attenuates both phototoxicity and hepatotoxicity.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/5/9/eaaw6127/DC1

    Supplementary Materials and Methods

    Fig. S1. Histology, oxidative stress, and inflammation in the skin of WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice after the exposure to UV light (395 to 410 nm).

    Fig. S2. Gross appearance of Fech-mut and Fech-mut/Abcg2-null mice before and after exposure to red light (630 nm).

    Fig. S3. Fluorescence of liver samples from WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice.

    Fig. S4. Evaluation of liver fibrosis in WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice.

    Fig. S5. ABCG2 deficiency abolishes GSF-induced PPIX accumulation and hepatotoxicity.

    Fig. S6. Effects of ABCG2 deficiency on the expression of efflux transporters in the liver.

    Fig. S7. Identification of PPIX-glu.

    Fig. S8. Hematological analysis of blood samples from WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice.

    Fig. S9. The ratios of spleen to body weight and PPIX levels in the spleen of WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice.

    Table S1. Primers for quantitative PCR analysis.

  • Supplementary Materials

    This PDF file includes:

    • Supplementary Materials and Methods
    • Fig. S1. Histology, oxidative stress, and inflammation in the skin of WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice after the exposure to UV light (395 to 410 nm).
    • Fig. S2. Gross appearance of Fech-mut and Fech-mut/Abcg2-null mice before and after exposure to red light (630 nm).
    • Fig. S3. Fluorescence of liver samples from WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice.
    • Fig. S4. Evaluation of liver fibrosis in WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice.
    • Fig. S5. ABCG2 deficiency abolishes GSF-induced PPIX accumulation and hepatotoxicity.
    • Fig. S6. Effects of ABCG2 deficiency on the expression of efflux transporters in the liver.
    • Fig. S7. Identification of PPIX-glu.
    • Fig. S8. Hematological analysis of blood samples from WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice.
    • Fig. S9. The ratios of spleen to body weight and PPIX levels in the spleen of WT, Abcg2-null, Fech-mut, and Fech-mut/Abcg2-null mice.
    • Table S1. Primers for quantitative PCR analysis.

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