Induction of p53 mediated mitochondrial apoptosis and cell cycle arrest in human breast cancer cells by plant mediated synthesis of silver nanoparticles from Bergenia ligulata (Whole plant).

The biological synthesis of nanoparticles is a growing research trend because it has numerous pharmaceutical and biomedical applications. The present study describes the preparation, characterization and anti-cancer evaluation of silver nanoparticles synthesized using an aqueous extract of Bergenia ligulata whole plant as a reducing agent. The physiochemical properties of the Bergenia ligulata silver nanoparticles (BgAgNPs) were measured by ultraviolet-visible spectrophotometry, Fourier transform infrared spectrophotmetry (FTIR), X-ray powder diffraction (XRD) and Scanning electron microscopy (SEM) analysis for identifying functional groups, crystallinity, structural and morphological features, respectively. Further, BgAgNps, along with the Bergenia ligulata aqueous extract (BgAE), were investigated for their effects on cell proliferation and apoptosis through MTT, colony-forming assay, wound-healing assay and flow cytometry-based approaches. The cytotoxic effects were more pronounced in cells treated with BgAgNps in comparison to BgAE. These effects were evidenced by the decreasing cell viability, migration capacity and loss of characteristic morphological features. In addition, BgAgNps unveiled significant induction of apoptosis in human breast cancer (MCF-7) cells, possibly through oxidative stress-mediated reactive oxygen species (ROS) generation and loss of mitochondrial membrane potential (MMP). Moreover, molecular mechanism-based studies revealed that BgAgNps robustly augmented p53 levels and pro-apoptotic downstream targets of p53 like Bax and cleaved caspase 3 in MCF-7 cells. Of note, BgAgNps had little or no cytotoxic effect on p53-deficient cancer cells (Mda-mb-231 and SW-620). These findings confirm that the BgAgNPs exhibited superior anti-cancer potential and could be exploited as a promising, cost-effective, and environmentally benign strategy in treating this disease in the future.

Par-4 mediated Smad4 induction in PDAC cells restores canonical TGF-ß/ Smad4 axis driving the cells towards lethal EMT.

Deregulation of TGF-ß signaling is intricately engrossed in the pathophysiology of pancreatic adenocarcinomas (PDACs). The role of TGF-ß all through pancreatic cancer initiation and progression is multifarious and somewhat paradoxical. TGF-ß plays a tumor suppressive role in early-stage pancreatic cancer by promoting apoptosis and inhibiting epithelial cell cycle progression, but incites tumor promotion in late-stage by modulating genomic instability, neo-angiogenesis, immune evasion, cell motility, and metastasis. Here, we provide evidences that Par-4 acts as one of the vital mediators to regulate TGF-ß/Smad4 pathway, wherein, Par-4 induction/over-expression induced EMT which was later culminated in to apoptosis in presence of TGF-ß via positive regulation of Smad4. Intriguingly, Par-4-/- cells were devoid of significant Smad4 induction compared to Par-4+/+ cells in presence of TGF-ß and ectopic Par-4 steadily augmented Smad4 expression by restoring TGF-ß/Smad4 axis in Panc-1 cells. Further, our FACS and western blotting results unveiled that Par-4 dragged the PDAC cells to G1 arrest in presence of TGF-ß byelevating p21 and p27 levels while attenuating Cyclin E and A levels and augmenting caspase 3 cleavage triggering lethal EMT. Through restoration of Smad4, we further establish that in BxPC3 cell line (Smad4-/-), Smad4 is essential for Par-4 to indulge TGF-ß dependent lethal EMT program. The mechanistic relevance of Par-4 mediated Smad4 activation was additionally validated by co-immunoprecipitation wherein disruption of NM23H1-STRAP interaction by Par-4 rescues TGF-ß/Smad4 pathway in PDAC and mediates the tumor suppressive role of TGF-ß, therefore serving as a vital cog to restore the apoptotic functions of TGF-ß pathway.