Akkermansia muciniphila ameliorates doxorubicin-induced cardiotoxicity by regulating PPARα-dependent mitochondrial biogenesis

Figures

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  Fig. 1. Akkermansia muciniphila (A. muciniphila) was depleted in C57BL/6 mice subjected to doxorubicin (DOX) and in breast cancer patients treated with anthracycline-based chemotherapy. a Principal component analysis (PCA) score plot for the feces metabolome of breast cancer patients with or without anthracycline (AC, n = 14) or other chemotherapy therapy (nAC, n = 16). Each point represented a sample. b At the genus level, a heatmap showed the relative abundance of bacteria in breast cancer patients from AC (n = 14) and nAC groups (n = 16). c Relative abundance of A. muciniphila in the AC and nAC groups. d Receiver operating characteristic analysis with the area under curve (AUC) is used to determine the efficiency of A. muciniphila in distinguishing anthracycline utilization (n = 30). e, f PCA score plot for the feces metabolome of control mice (e) and DOX-treated mice (f) at the indicated time (n = 4). g At the genus level, relative abundance of bacteria (percentage of total bacteria) in mice treated with or without DOX at the indicated time (week0: before DOX administration; week1–4: on the first day following DOX administration during the specified week). Different colors represent…

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  Fig. 2. Akkermansia muciniphila (A. muciniphila) colonization alleviates doxorubicin (DOX)-induced cardiomyopathy in antibiotic cocktail (Abx)-treated mice. a Changes in qPCR-based absolute abundances of fecal A. muciniphila in mice from indicated group. b Anatomic M-mode echocardiography and corresponding electrocardiographic images of hearts from Abx-treated mice, DOX plus Abx-treated mice, and DOX plus A. muciniphila and Abx-treated mice (n = 6). Changes of echocardiographic c left ventricular ejection fraction (LVEF) and d left ventricular fraction shortening (LVFS) in the indicated groups (n = 6). e, f Changes of serum lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) levels in the three groups (n = 6). g Representative H&E staining images of the heart tissue section from mice in the indicated groups. Bar = 100 μm (n = 6). h Wheat germ agglutinin (WGA) staining of myocardial tissue sections to evaluate cardiac atrophy. Bar = 50 μm. i Quantitation of myocardial cell size of mice in the indicated groups (n = 6). j Representative images of myocardial tissue sections stained with Masson’s trichrome (from top to bottom, the images are a panoramic view of the heart, a transverse…

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  Fig. 3. Supplement with Akkermansia muciniphila (A. muciniphila) improves cardiac dysfunction by regulating mitochondrial bioenergetics. a KEGG enrichment analysis of RNA-sequencing data from DOX and control hearts. b Heatmap showing differentially expressed mitochondrial respiratory electron transport chain (ETC, Complex I–V) genes in the mouse hearts from indicated groups (n = 3). c The activity of mitochondrial respiratory chain complexes in the hearts of mice from various groups was assessed using commercial kits (n = 6). d Representative immunoblotting images and quantification of mitochondrial OXPHOS proteins in mouse heart lysates (antibodies against Ndufb8, Sdhb, Uqcrc2, Mtco1, and Atp5a1 used as representatives for mitochondrial complex I, II, III, IV, and V, and GAPDH used as an internal control, n = 3 for each group). e Real-time monitoring of the oxygen consumption rate (OCR) in neonatal mouse ventricular cardiomyocytes (NMVMs) with or without DOX/A. muciniphila treatment. Data are presented as the mean ± SD (n = 3). f Representative transmission electron microscopy images of mice hearts from the control, DOX and DOX + A. muciniphila group. Scale bar = 2 μm. Quantification of…

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  Fig. 4. Akkermansia muciniphila (A. muciniphila) administration regulates mitochondrial bioenergetics by activating PPARα/PGC1α signaling pathway. a Pearson correlation analysis between the 11 mitochondrial OXPHOS-related genes and A. muciniphila levels, as well as cardiac injury markers in mice (n = 3). b, c Representative immunoblotting images and quantification of PPARα and PGC1α in the heart tissues from the control, DOX, and DOX + A. muciniphila groups (n = 3). d RT-qPCR was used to measure the mRNA levels of PPARA and PGC1α, from the heart tissues in the indicated groups (n = 3). e Immunofluorescence staining of heart tissues from the indicated groups with cardiac PPARα (green) DAPI (blue), as well as semi-quantitative analysis of PPAR fluorescence. f Schematic representation of treatment with DOX, A. muciniphila, or PPARα inhibitor NXT629 (30 mg/kg). g Anatomic M-mode echocardiography and corresponding electrocardiographic images of hearts from DOX, DOX + A. muciniphila, and DOX + A. muciniphila + NXT629 (n = 6). Changes of echocardiographic h left ventricular ejection fraction (LVEF) and i left ventricular fraction shortening (LVFS) in the indicated groups (n = 6). j…

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  Fig. 5. Fecal microbiome analysis of Akkermansia muciniphila (A. muciniphila)-treated mice under doxorubicin (DOX) stimulation. a Boxplots of a-diversity calculated by Chao, Shannon_2, Simpson, and observed reads (n = 4). b PCoA plot based on unweighted Unifrac distance was used to test the variations of microbial communities among groups (n = 4). c Relative abundance of differential genera in the three groups were illustrated (n = 4 per group). d Statistical differences among the control, DOX, and DOX + A. muciniphila groups were identified using the linear discriminant analysis (LDA) effect size (LEfSe) method. Cladogram illustrating the output of the LEfSe algorithm. Significantly distinct taxonomic nodes are colored, and the branch areas are shaded according to the effect size of the taxa (n = 4). e Ternary plots reveal OTUs relative abundance (dot size) at the genus level among the three sample groups. Generalist taxa are represented as circles in the middle of the triangle, sample-specific bacterial taxa are represented as circles in the summit or along the edges of the triangle (n = 4). f Representative hematoxylin and eosin (H&E) staining and periodic acid Schiff (PAS) staining…

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  Fig. 6. Akkermansia muciniphila (A. muciniphila) modulates the microbial tryptophan metabolites profile of doxorubicin (DOX)-treated mice. Principal component analysis of a fecal metabolites and b serum metabolites among the control, DOX, and DOX + AKK groups (n = 6). c Bar chart of significantly different KEGG metabolic pathways (based on serum metabolites) among the control, DOX, and DOX + AKK groups (n = 6). d Serum concentrations of metabolites in tryptophan metabolism (ArAT aromatic amino acid aminotransferase, IPyA indole-3-pyruvate, fldH phenyllactate dehydrogenase, ILA indole-3-lactic acid, fldBC phenyllactate dehydratase, acdA acyl-Coa dehydrogenase, IPA indole-3-propionic acid, TMO tryptophan 2-monooxygenase, TrD tryptophan decarboxylase, IAM indole-3-acetamide, IAA indole acetic acid, IAId indole-3-aldehyde) among three groups of mice (n = 6). e Spearman correlation analysis between the IPA levels and cardiac injury markers (LVEF left ventricular ejection fraction, LVFS left ventricular fraction shortening, LDH lactate dehydrogenase, CK-MB creatine kinase-MB) in mice from the three groups (n = 18). f Pairwise comparisons of tryptophan metabolites, with a color gradient…

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  Fig. 7. Indole-3-propionic acid (IPA) supplementation alleviates doxorubicin (DOX)-induced cardiac dysfunction, atrophy, and fibrosis in C57BL/6 mice. a Schematic representation of treatment with doxorubicin (DOX) and low IPA (20 mg/kg) or high IPA (40 mg/kg) (n = 6 per group). b Anatomic M-mode echocardiography and corresponding electrocardiographic images of hearts from the indicated groups (n = 6). Changes of echocardiographic c left ventricular ejection fraction (LVEF) and d left ventricular fraction shortening (LVFS) in the indicated groups (n = 6). e, f Changes of serum lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) levels in the four groups (n = 6). g Upper panel: representative H&E staining images of the heart tissue section from mice in the indicated groups. Bar = 100 μm. Bottom panel: wheat germ agglutinin (WGA) staining of myocardial tissue sections to evaluate cardiac atrophy. Bar = 50 μm. Right panel: quantitation of myocardial cell size of DIC mice in the indicated groups (n = 6). h Representative ×200 images of myocardial tissue sections stained with Masson’s trichrome (from top to bottom, the images are a panoramic view of the heart, a transverse section, and…

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  Fig. 8. Indole-3-propionic acid (IPA) administration improves doxorubicin (DOX)-induced mitochondrial dysfunction. a Representative transmission electron microscopy images of mice hearts from the control, DOX, and DOX + IPA group. Scale bar = 2 μm. Quantification of mitochondria-related parameters from (b). Mitochondria/μm2 refers to the average number of mitochondria (n = 6). c % mitochondrial area refers to the ratio of mitochondrial area to image area (n = 6). d The activity of mitochondrial respiratory chain complexes in the hearts of mice from various groups was assessed using commercial kits (n = 6). e Representative immunoblotting images and quantification of mitochondrial OXPHOS proteins in mouse heart lysates (antibodies against Ndufb8, Sdhb, Uqcrc2, Mtco1, and Atp5a1 used as representatives for mitochondrial complex I, II, III, IV, and V, and GAPDH used as an internal control, n = 3 for each group). f Representative immunoblotting images and quantification of DRP1, MFN2 and OPA1 in the heart tissues from the control, DOX, and DOX + IPA groups (n = 3). g–k Real-time monitoring of the oxygen consumption rate (OCR) in neonatal mouse ventricular cardiomyocytes (NMVMs) with or…

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  Fig. 9. The schematic diagram of Akkermansia muciniphila (AKK) ameliorating doxorubicin (DOX)-induced cardiotoxicity. AKK colonization increased serum tryptophan metabolite indole-3-propionic acid (IPA), which improved DOX-induced mitochondrial dysfunction by activating cardiac aryl hydrocarbon receptor (AhR)-dependent PPARα/PGC1α signaling pathway, which could attenuate DOX-induced cardiotoxicity.