Rosmarinic acid enhances apoptosis and cell cycle arrest induced by Carnosic acid on Human Lung Fibroblasts

Abstract

Background: Pulmonary Fibrosis (PF) is a chronic progressive disease characterized by aberrant fibroblasts proliferation and collagen accumulation in the interstitial tissue. Considering the partial efficacy and side effects of current treatments, several therapeutic trials have been carried out with plant extracts rich in bioactive molecules. Objective: The aim of this study was to investigate the effects of Carnosic acid (CA) and Rosmarinic acid (RA), two compounds extracted from Rosemary, on Human Lung Fibroblasts (HLF) cultured in vitro. Methods: HLF were purchased from Lonza group and cultured in FGM-2 culture medium (Lonza, Verviers, Belgium) at 37 °C and 5% CO2. The colorimetric determination of the number of living cells was carried out using the Cell Counting Kit-8 (CCK-8, Sigma-Aldrich, Leuven, Belgium). The effect of RA and CA on cell cycle progression and apoptosis were detected using the MuseTM Cell Analyser (MerckKGaA, Darmstadt, Germany). Akt, MAPK and p21 expressions were evaluated by western blot. Results: CA induced HLF apoptosis and cell cycle arrest in G0 /G1 while RA had no cytotoxic effect. Addition of RA enhanced pro-apoptotic effect of CA, with complete disappearance of Akt, p38-MAPK and p21 expression 24 hours after starting CA/RA treatment. Inhibitors of Akt, p38-MAPK and pan-caspase doubled the rate of HLF living cells in response to CA/RA treatment, proving the implication of Akt /p38-MAPK and caspases activation in HLF apoptosis induced by these biomolecules. Conclusion: This study demonstrated a significant synergistic pro-apoptotic effect of CA/RA on HLF in vitro and may be completed by an in vivo study.