Combined cardioprotective effects of molecular hydrogen combined with Vericiguat via the NO-sGC-cGMP signaling pathway in heart failure rats
Abstract
Heart failure is a life-threatening end-stage cardiovascular condition associated with high rates of hospital readmission and mortality. Conventional therapeutic interventions cannot fully reverse the complex pathological remodeling associated with heart failure. Vericiguat, a guideline-recommended soluble guanylate cyclase (sGC) stimulator, enhances cGMP signaling, whereas molecular hydrogen has antioxidant and anti-apoptotic properties that may confer cardioprotection. In this study, we established an isoproterenol (ISO)-induced heart failure model in rats and an ISO-induced H9C2 cardiomyoblast injury model to evaluate the combined effects of molecular hydrogen and vericiguat and to investigate the associated molecular pathways. Echocardiography, histopathological staining, transmission electron microscopy, ELISA, western blotting, and flow cytometry were used to assess cardiac function, myocardial fibrosis, mitochondrial ultrastructure, apoptosis, and signaling-related molecular changes. Combined treatment improved left ventricular systolic and diastolic function, reduced myocardial collagen deposition and mitochondrial structural damage, and decreased apoptosis. These effects were accompanied by changes in TGF-β/Smad signaling, mitophagy- and endoplasmic reticulum stress-related markers, and increased NO-sGC-cGMP signaling. These findings demonstrate the cardioprotective effects of combined molecular hydrogen and vericiguat treatment in preclinical models and provide a methodological basis for further investigation of this therapeutic approach.
Dataset DOI: 10.5061/dryad.h9w0vt500
Description of the data and file structure
Archive name: 73109.zip
This compressed archive contains all raw experimental data generated from the present study, which investigated the protective effects of hydrogen combined with vericiguat against isoproterenol-induced myocardial injury and heart failure in rats and H9C2 cardiomyocytes. The entire dataset is organized into six categorized directories, including echocardiography, ELISA, histological morphology, TEM, in vitro cell assays, and western blot data.
Directory and File Description
1. Echocardiography
This directory contains raw cardiac function parameters and representative echocardiographic images of experimental rats.
- echo_raw_output.xlsx: Raw numerical data of all echocardiographic indices for each animal, including cardiac structure and function parameters.
- Echocardiography.tif: Representative original echocardiogram images of rats from each experimental group.
2. ELISA
This directory contains raw optical density data of inflammatory and myocardial injury biomarkers detected by ELISA. Individual Excel files are not listed one by one.
File naming convention: ELISA_GroupAbbreviation_SampleNumber.xlsx
Group abbreviations include CON (control), ISO (model), ISO+H₂, ISO+Veri, ISO+H₂+Veri. Each file corresponds to one independent biological sample and contains original OD values obtained from microplate detection.
3. Morphological results
This directory contains myocardial histopathological staining images and quantitative scoring results, divided into HE and Masson subdirectories.
3.1 HE
- HE.tif: Representative original HE-stained myocardial tissue images for histological observation of inflammatory infiltration and myofibril damage.
- Myocardial histopathological injury score.xlsx: Semi-quantitative histological injury scores based on myocardial structural damage, inflammatory infiltration, and edema.
3.2 MASSON
- Masson.tif: Representative original Masson trichrome staining images for myocardial fibrosis observation.
- Fibrosis_quantification.xlsx: Quantitative data of myocardial fibrosis area percentage in each group.
4. TEM
This directory contains transmission electron microscopy original images and scoring data of cardiomyocyte mitochondrial ultrastructure.
- TEM_mitochondria.tif: Original TEM images showing mitochondrial morphology, cristae structure, and ultrastructural changes in cardiomyocytes.
- Mitochondrial_injury_score.xlsx: Semi-quantitative scoring results of mitochondrial ultrastructural damage.
5. vitro_cell_assays
This folder contains experimental raw and processed data from H9C2 cardiomyocyte in‑vitro experiments.
cell_cycle/directory: Flow‑cytometry cell‑cycle distribution data. Contains raw flow output and processed percentage values for G0‑G1, S and G2‑M cell‑cycle phases.cell cycle distribution.xlsx: Processed percentage of cells in each cell‑cycle phase across Control, ISO, ISO+Veri, ISO+H₂, ISO+Veri+H₂ groups, n=3 biological replicates.cell cycle distribution.tif: Raw exported flow‑cytometry dot‑plot image for cell‑cycle analysis.
AnnexinV_PI_flow_cytometry/directory: Apoptosis detection dataset using Annexin V‑FITC / PI double‑staining flow cytometry.The total apoptosis rate.xlsx: Processed total apoptosis percentage for each experimental group, n=3 biological replicates. Includes replicate values for Control, ISO, ISO+Veri, ISO+H₂, ISO+Veri+H₂.The total apoptosis rate.tif: Raw flow‑cytometry dot‑plot image for Annexin V‑PI apoptosis detection.
F_actin_stained/directory: Phalloidin‑DAPI immunofluorescence dataset for F‑actin cytoskeleton observation.relative fluorescence intensity.xlsx: Quantified relative fluorescence intensity of F‑actin, n=3 biological replicates across all five in‑vitro treatment groups.relative fluorescence intensity.tif: Representative confocal immunofluorescence raw image of F‑actin‑stained H9C2 cardiomyocytes.
6. WB
This directory contains Western‑blot band data for protein expression analysis. It comprises 16 sub‑directories, each named according to the specific protein analysed (e.g., BAX, BCL‑2, Cleaved‑caspase3, COL‑I, COL‑III, GUCY1B1, GUCY1B3, LC3, Nrf2, PARKIN/PINK1, PRKG1, sGC, Smad2, TGF‑β, BNP, and others).
Each protein‑named sub‑directory contains two files:
- One .tif image file of the corresponding Western‑blot membrane/band (named after the protein);
- One .xlsx Excel file with the quantified band‑intensity data (also named after the protein).
Variable Dictionary for All Tabular Data
All column variables, abbreviations, units and interpretation for Excel datasets are defined below:
- Group: Experimental grouping; CON (normal control), ISO (ISO-induced heart failure model), ISO+H₂ (hydrogen treatment), ISO+Veri (vericiguat treatment), ISO+H₂+Veri (combination treatment)
- Sample_ID: Unique identification number of each independent biological sample
- EF%: Ejection fraction, cardiac systolic function index; unit: %
- FS%: Fractional shortening; unit: %
- LVIDd/s: Left ventricular internal dimension at diastole/systole; unit: mm
- OD value: Optical density; unitless absorbance value for ELISA detection
- Injury score: Myocardial histopathological injury score; 0–4 semi-quantitative scoring scale (0 = normal, 4 = severe injury)
- Fibrosis area %: Percentage of fibrotic area in myocardial tissue; unit: %
- Apoptosis rate: Total cardiomyocyte apoptosis rate; unit: %
- G0/G1, S, G2/M%: Cell cycle phase proportion; unit: %
- Relative protein expression: Normalized relative grey value of target protein bands; unitless