Sex Differences in Circulating MicroRNA Profiles in Heart Failure with Preserved and Reduced Ejection Fraction

Figures

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  Figure 1. Study cohort and analytic pipeline. (A) Study flow diagram showing participant selection and sex distribution across the control, HFpEF, and HFrEF groups. (B) Analytic workflow for whole-blood miRNA sequencing. After sequencing and generation of raw miRNA counts, miRNAs were filtered using the main filter (≥10 raw counts in ≥ 50% of samples within at least one of the six sex-by-phenotype subgroups). Primary analyses comprised sex-stratified differential-expression analyses, between-sex comparison of log2 fold changes, and principal component analysis (PCA) with permutational multivariate analysis of variance (PERMANOVA).

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  Figure 2. Sex-stratified volcano plots of circulating miRNA expression changes in HFpEF and HFrEF. (A, B) Volcano plots showing age- and BMI-adjusted differential-expression results for sex-stratified comparisons of HFpEF vs. Control (A) and HFrEF vs. Control (B). The x-axis indicates log2 fold change for the HF phenotype relative to Control, and the y-axis indicates −log10(FDR). Points passing FDR < 0.05 are highlighted. In males, 50 miRNAs were significantly altered in HFpEF (31 up-regulated and 19 down-regulated), and 59 miRNAs were significantly altered in HFrEF (34 up-regulated and 25 down-regulated). No miRNAs met the FDR threshold in females for either comparison.

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  Figure 3. Between-sex comparison of HF-associated miRNA changes. (A) HFpEF vs. Control. (B) HFrEF vs. Control. In each panel, the left scatter plot compares age- and BMI-adjusted log2 fold changes relative to Control in males (x-axis) and females (y-axis) for the 229 filtered miRNAs, with each point representing one miRNA. The corresponding Spearman correlation coefficients were 0.559 for HFpEF vs. Control and 0.513 for HFrEF vs. Control (both P < 0.001). The right box plot compares absolute age- and BMI-adjusted log2 fold changes between males and females across the same set of analysed miRNAs. Boxes indicate the inter-quartile range (IQR), with the median shown as the centre line; whiskers indicate the range within 1.5 × IQR. Paired Wilcoxon signed-rank tests were used.

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  Figure 4. Principal component analysis of circulating miRNA profiles. (A, B) Principal component analysis (PCA) plots based on variance-stabilized expression values from the 229 filtered miRNAs for HFpEF vs. Control (A) and HFrEF vs. Control (B). Males and females are shown separately. Control samples are shown as open circles and HF samples as filled circles. Ellipses indicate 95% normal-data regions for each group.