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PI3K/AKT/mTOR signaling promotes cell survival, proliferation and progression in cancer cells. Targeting this pathway may lead to the development of novel therapeutic approaches for human cancers. Here, we examined the effects of (-)-epigallocatechin-3-gallate (EGCG) on the PI3K/AKT/mTOR pathway in pancreatic cancer cells, and assessed its therapeutic potential. In this study, the proliferation and apoptosis of PANC-1 cells were examined by MTT assay and flow cytometry, respectively. The expression of genes and proteins involved in the PI3K/AKT/mTOR pathway were measured by RT-PCR and western blotting, respectively. Our results revealed that EGCG dramatically inhibited the proliferation of PANC-1 cells and induced apoptosis simultaneously. Furthermore, it upregulated PTEN mRNA and protein expression levels, as well as downregulating the expression of phospho-AKT and phospho-mTOR. In conclusion, these results suggest that EGCG can suppress proliferation and induce apoptosis of PANC-1 cells in a time- and dose-dependent manner; moreover, EGCG also can upregulate PTEN expression and downregulate the expression of pAKT and p-mTOR to modulate the PI3K/AKT/mTOR signaling pathway.

Several studies demonstrate that estradiol can prevent arterial calcification. However, little is known regarding the effect of puerarin, a phytoestrogen extracted from Radix Puerariae, on arterial calcification. The aim of the present study was to determine whether puerarin reduced osteoblastic differentiation of calcifying vascular smooth muscle cells (CVSMCs). The CVSMCs were isolated from mice aorta and treated with different concentrations of puerarin. The alkaline phosphatase (ALP) activity, osteocalcin secretion and Runx2 expression were determined. To examine whether estrogen receptors (ERs) PI3K and Akt play a role in this effect, ICI182789, phosphoinositide 3-kinase (PI3K) inhibitor, LY294002, or the Akt inhibitor, 1L-6-hydroxymethyl-chiro-inositol 2-(R)-2-O-methyl-3-O-octadecylcarbonate (HIMO) was used. Our results showed puerarin could inhibit ALP activity, osteocalcin secretion and Runx2 expression in CVSMCs. Puerarin could induce the activation of Akt. Furthermore, pretreatment of ICI182780, LY294002, HIMO could abolish the effect of puerarin on ALP activity in CVSMCs. Our experiment demonstrated that puerain could attenuate the osteoblastic differentiation of VSMCs through the ER/PI3K-Akt signal pathway.

Dried roots of Gastrodia elata have traditionally been used in Korean medicine for the treatment of neurological disorders such as scotodinia, paralysis, and epilepsy. In our study, we attempted to investigate the neuroprotective effects of methanol extract from G. elata (MEGE) against glutamate-mediated oxidative stress and to explore underlying neuroprotective mechanisms. Analyses for cell viability, lactate dehydrogenase (LDH), flow cytometry, Western blot, and reactive oxygen species (ROS) were performed in HT22 hippocampal cells. Pretreatment with MEGE resulted in a potent neuroprotective effect against oxidative glutamate toxicity and these effects were exerted mainly by the abrogation of glutamate-induced apoptotic death. Treatment with glutamate resulted in a significant expression of both phosphorylated p38 and dephosphorylated phosphatidylinositol-3-kinase (PI3K). However, pretreatment with MEGE resulted in the inhibition of these expressions. In the inhibitor studies, treatment with PI3K inhibitor LY294002 resulted in the abrogation of the neuroprotective effect of MEGE. In addition, pretreatment with MEGE also resulted in the suppression of the glutamate-induced production of ROS. Treatment with MEGE and anti-oxidant N-acetyl-L-cysteine (NAC) resulted in the enhanced phosphorylation of both PI3K and cAMP responsive element binding protein (CREB), and, in particular, treatment with MEGE resulted in significantly enhanced expression of mature brain-derived neurotrophic factor (BDNF). These results suggest that the extract from G. elata mainly exerted neuroprotective effects through the up-regulation of the PI3K signaling pathway in association with BDNF and may be a useful therapeutic agent for treatment of oxidative neuronal death.

Prunella vulgaris (PV) is a traditional Chinese medicine that has been used clinically for centuries in Asian countries to treat herpetic keratitis. In previous studies, PV was shown to suppress TPA-induced activation of MMP-9 and inhibit cell invasion and migration in hepatoma cell lines. However, the detailed molecular mechanism underlying these effects is still unclear. In this study, we investigated the mechanisms underlying PV-mediated inhibition of 12-O-Tetradecanoylphorbol-13-acetate (TPA)-induced cell invasion and inhibition of secreted and cytosolic MMP-9 production in human hepatoma cells (Huh-7 and HA22T). PV suppressed VEGF and MMP-9 transcription by inhibiting activator protein (AP)-1 and nuclear factor-$\kappa$B (NF-$\kappa$B) activity. PV suppressed TPA-induced AP-1 activity by inhibiting phosphorylation of the extracellular signal-related kinase (ERK), downregulating p38 signaling pathways, and suppressing TPA-induced inhibition of NF-$\kappa$B nuclear translocation through I$\kappa$B. PV suppressed TPA-induced activation of ERK/phosphatidylinositol-3-kinase/Akt upstream of NF-$\kappa$B and AP-1. These data suggest that PV modifies the metastatic microenvironment of hepatocellular carcinoma (HCC) by inhibiting multiple signal transduction pathways. PV thus may have the therapeutic potential to inhibit the migration and invasion of HCC and act as potential agent for systemic therapies.

Baizhu, the dried rhizome of Atractylodes Macrocephala Koidz (Compositae), is one of the most important traditional Chinese herbal medicines. Baizhu is generally used to treat digestive disorders and diabetes in Asian countries. This study investigates the activity of two sesquiterpenes isolated from Baizhu, atractylenolide I (AT-I) and atractylenolide II (AT-II), for their effects on glucose uptake in mouse skeletal muscle C2C12 cells, and the corresponding mechanism. These compounds show a significant stimulatory effect on glucose uptake in C2C12 myotubes. Both AT-I and AT-II significantly increased GLUT4 but not GLUT1 protein levels, and promoted GLUT4 translocation to the plasma membrane. The increased glucose uptake induced by these compounds is associated with activation of AMP-activated protein kinase (AMPK) and PI3K/Akt pathways in these cells. Further studies have indicated that AT-I and AT-II ameliorate TNF-$\alpha$-induced insulin resistance in C2C12 myotubes. In summary, our findings highlight the insulin mimetic activity of Baizhu in myotubes, and provide insights into the action mechanism underlying these effects. Our findings may also prove highly relevant to the development of novel therapeutic applications for these compounds.

Aloe-emodin (AE) is derived from Aloe vera and rhubarb (Rheum palmatum) and exhibits anticancer activities via multiple regulatory mechanisms in various cancers. AE can also enhance the anticancer efficacy of cisplatin, doxorubicin, docetaxel, and 5-fluorouracil; however, its effects remain poorly characterized. MCF-7, MDA-MB-231, MDA-MB-468, BT-474, and HCC-1954 breast cancer cell lines were treated with the indicated conditions of AE, and cell viability assays were performed. The expression levels of signaling proteins were determined by western blot analysis, intracellular reactive oxygen species (ROS), cell cycle distributions, and rates of apoptosis as estimated by flow cytometry. In comparison with other cells, MCF-7 cells were more sensitive to AE treatment; AE enhanced the cytotoxicity of 9$\mu$g/ml tamoxifen by reducing EGFR, ER$\alpha$, Ras, ERK, c-Myc, and mTOR protein expression and blocking PI3K and mTOR activation. Finally, although co-treatment of AE with tamoxifen increased intracellular ROS, there were no effects on cell cycle progression. Besides facilitating tamoxifen-induced cell death, AE also enhanced the antiproliferative activity of tamoxifen by blocking Ras/ERK and PI3K/mTOR pathways in breast cancer cells, thus demonstrating the chemosensitizing potential of AE.

Diabetic nephropathy (DN) is one of the most serious complications of diabetes and the most common cause of death. The autophagy of podocytes plays an important role in the pathogenesis of DN. Here, through screening the constituent compounds of practical and useful Chinese herbal formulas, we identified that isoorientin (ISO) strongly promoted the autophagy of podocytes and could effectively protect podocytes from high glucose (HG)-induced injury. ISO significantly improved autophagic clearance of damaged mitochondria under HG conditions. Through a proteomics-based approach, we identified that ISO could reverse the excessive phosphorylation of TSC2 S939 under HG conditions and stimulate autophagy through inhibition of the PI3K-AKT-TSC2-mTOR pathway. Furthermore, ISO was predicted to bind to the SH2 domain of PI3Kp85$\beta$, which is crucial for the recruitment and activation of PI3K. The protective effect of ISO and its effects on autophagy and particularly on mitophagy were further proved using a DN mice model. To summarize, our study identified the protective effects of ISO against DN and demonstrated that ISO was a strong activator of autophagy, which could provide a basis for drug development.

Breast cancer (BC) is a critical health issue that affects countless women, and it is the second leading reason of death worldwide. The phosphatidylinositol 3 kinases (PI3Ks) constitute a group of lipid kinases that play a role in tumorigenesis, development, migration, infiltration, programmed cell death, glycogen synthesis, DNA correction and viability by the PI3K/Akt cascade. The PI3K pathway has been linked to a variety of malignancies and increases the activation rate of cancer. Here, focus was given to the study of PI3K pathway involved in BC and emphasis was given on a particular nSH₂ domain that resides in the regulatory subunit of PI3K to find a potent inhibitor. A detailed pathway and interaction study was performed from KEGG pathway database and from the cystoscope. A total list of 60 compounds, comprises phytochemicals, and herbal compounds were screened based on structural similarity and eight FDA-approved drugs were considered. The docking analysis was carried over through the AutoDock software and Ligplot analysis was performed to investigate the interaction between the nSH₂ domain and the potent inhibitors. To ensure the complex stability, 20 ns of simulation run was also performed on the best complexes using GROMACS. From this study, it can be concluded that Evodia fruit has the maximum stability in the catalytic region among all the listed inhibitors against the target proteins and can act as a potent inhibitor among the others.