EP_052. Hibiscus sabdariffa modulates inflammatory and matrix genes in senescent cardiac fibroblasts

Mei Siu Lau^1*, Wan Mohamad Adham Afiq Wan Zainuzzaman¹, Siti Khadijah Mustapha Kamal¹, Wan Nurul Nadia Wan Seman¹, Nik Aina Syazana Nik Zainuddin¹, Chin Long Poo¹, Mohd Rahimi Ashraf Abd Rahman¹, Nur Liana Md Nasir¹, Hussin Muhammad¹, Norizah Awang¹

1. Unit Toxicology & Pharmacology Unit, Herbal Medicine Research Centre, Institute for Medical Research, National Institutes of Health, 40170 Shah Alam, Selangor, Malaysia

*Corresponding author: Mei Siu Lau. This email address is being protected from spambots. You need JavaScript enabled to view it.

ABSTRACT

INTRODUCTION: Oxidative stress-induced cellular senescence is a major contributor to cardiac ageing and fibrosis, mediated by the upregulation of pro-inflammatory cytokines and extracellular matrix (ECM) remodelling genes. Hibiscus sabdariffa (Roselle), known for its antioxidant and cardioprotective properties, may modulate these responses. 

OBJECTIVE(S): To evaluate the effects of H. sabdariffa aqueous extract on senescence-associated secretory phenotype (SASP) and ECM gene expression in hydrogen peroxide (H₂O₂)-induced senescent human ventricular cardiac fibroblasts (NHCF-V).

MATERIALS & METHODS: Cellular senescence was induced in NHCF-V cells using H₂O₂ (50–500 µM) over 24, 48, and 72 hours. Based on senescence-associated β-galactosidase (SA-β-Gal) quantification, 100 µM H₂O₂ for 48 hours was selected as the optimal senescence-inducing condition, allowing sustained SASP gene expression with minimal cytotoxicity. Cells were pre-treated with H. sabdariffa extract (3.9–250 µg/mL) for four hours before H₂O₂ exposure. For gene expression analysis, non-toxic concentrations (15.625, 31.25 and 62.5 µg/mL) were selected. Total RNA was extracted, reverse-transcribed, and analysed by reverse transcription quantitative PCR to quantify SASP markers (IL-1B, IL-6, IL-18, TNF-α) and ECM genes (COL1A1 and COL3A1). GAPDH served as the reference gene.

RESULTS: H₂O₂ significantly upregulated IL-1B (~2.9-fold vs untreated cells, p = 0.001). Pre-treatment with H. sabdariffa at 62.5 µg/mL slightly reduced IL-1B (~4% reduction vs H₂O₂ group, p = 0.037). Lower concentrations showed similar but non-significant trends. TNF-α,  IL-6, IL-18, COL1A1, and COL3A1 expressions did not differ significantly. GAPDH Ct values remained stable.

CONCLUSION: H. sabdariffa pre-treatment significantly reduced IL-1B expression in senescent NHCF-V cells. While other genes showed no significant changes, variable responses in TNF-α, COL1A1, and COL3A1 warrant further investigation in time-course and higher-powered studies.

KEYWORDS: Hibiscus sabdariffa, Cardiac Fibroblast, Senescence, Senescence-Associated Secretory Phenotype, Reverse Transcription Quantitative PCR