Anticancer and immunomodulatory effect of rhaponticin on Benzo(a)Pyrene-induced lung carcinogenesis and induction of apoptosis in A549 cells

In worldwide, one of the most important cancer-related death is lung cancer. Also has the highest mortality rate between various cancer types. The count of lung cancer occurrence is increasing with an increased frequency by smoking. Proficient chemoprevention approaches are needed to prevent the occurrence of lung cancer. Therefore, the aim of this exploration is to determine the therapeutic impact on the immune modulatory effect of rhaponticin on lung tumorigenesis in vivo and in vitro cytotoxicity effect in A549 cells of human lung cancer. Lung cancer tumorigenesis in mice was challenged with benzo(a)pyrene (BaP) with 50 mg/kg bodyweight (b.wt) as oral administration for 6 weeks (two times/week). Rhaponticin were given orally 30 mg/kg b.wt (two times/week) in BaP induced mice from 12 weeks to 18 weeks. After treatment completes, the body weight was measured and then blood, lung tissue was collected for various parameters detection. The results evidenced that BaP induced mice decreased the bodyweight, increased lung weight, increased tumor markers (AHH, CEA and LDH), and increased the proinflammatory cytokines. The enzyme catalase, superoxide dismutase activity was decreased and increased lipid peroxidation in immune comprising cells compared with the control cells. Moreover, rhaponticin treatment improves in chemical assays and also the histopathological alteration of lung tissues. The present findings provide evidence about the therapeutic potentials of rhaponticin against BaP triggered lung tumorigenesis.     

Rhaponticin suppresses osteosarcoma through the inhibition of PI3K-Akt-mTOR pathway

Osteosarcoma is the frequent pediatric bone cancer where pediatric osteosarcoma incidences are more than 10% within the population. Most of the patients with osteosarcoma fall within the age of 15–30 years. Therefore, in this research, we examined the anticancer effect of Rhaponticin against the human osteosarcoma (MG-63) cells. The cytotoxicity of Rhaponticin on the MC3T3-E1 and MG-63 cells was examined through the MTT assay. The intracellular ROS accumulation, cell nuclear morphological alterations, apoptotic cell death and nuclear damages, and MMP status of Rhaponticin administered MG-63 cells were inspected by fluorescent staining techniques. The cell migration was assessed through scratch assay. The mRNA expressions of PI3K-Akt-mTOR signaling proteins were studied by RT-PCR analysis. Rhaponticin showed potent cytotoxicity, substantially inhibited the MG-63 cell growth, and displayed morphological alterations. However, rhaponticin did not affect the MC3T3-E1 cell viability. Rhaponticin administered MG-63 cells demonstrated augmented intracellular ROS accretion, weakened MMP, increased nuclear damages, and increased apoptosis. Rhaponticin effectively down-regulated the PI3K-Akt-mTOR signaling cascade in the MG-63 cells. These outcomes proved that the Rhaponticin can be a hopeful chemotherapeutic agent in the future to treat human osteosarcoma.     

Arginase inhibition by rhaponticin increases L-arginine concentration that contributes to Ca2+-dependent eNOS activation

Although arginase primarily participates in the last reaction of the urea cycle, we have previously demonstrated that arginase II is an important cytosolic calcium regulator through spermine production in a p32-dependent manner. Here, we demonstrated that rhaponticin (RPT) is a novel medicinal-plant arginase inhibitor and investigated its mechanism of action on Ca²⁺-dependent endothelial nitric oxide synthase (eNOS) activation. RPT was uncompetitively inhibited for both arginases I and II prepared from mouse liver and kidney. It also inhibited arginase activity in both aorta and human umbilical vein endothelial cells (HUVECs). Using both microscope and FACS analyses, RPT treatments induced increases in cytosolic Ca²⁺ levels using Fluo-4 AM as a calcium indicator. Increased cytosolic Ca²⁺ elicited the phosphorylations of both CaMKII and eNOS Ser1177 in a time-dependent manner. RPT incubations also increased intracellular L-arginine (L-Arg) levels and activated the CaMKII/AMPK/Akt/eNOS signaling cascade in HUVECs. Treatment of L-Arg and ABH, arginase inhibitor, increased intracellular Ca²⁺ concentrations and activated CaMKII-dependent eNOS activation in ECs of WT mice, but, the effects were not observed in ECs of inositol triphosphate receptor type 1 knockout (IP3R1^−/−) mice. In the aortic endothelium of WT mice, RPT also augmented nitric oxide (NO) production and attenuated reactive oxygen species (ROS) generation. In a vascular tension assay using RPT-treated aortic tissue, cumulative vasorelaxant responses to acetylcholine (Ach) were enhanced, and phenylephrine (PE)-dependent vasoconstrictive responses were retarded, although sodium nitroprusside and KCl responses were not different. In this study, we present a novel mechanism for RPT, as an arginase inhibitor, to increase cytosolic Ca²⁺ concentration in a L-Arg-dependent manner and enhance endothelial function through eNOS activation.