Emodin-induced autophagy against cell apoptosis through the PI3K/AKT/mTOR pathway in human hepatocytes.

BACKGROUND: Emodin, a major component of Polygonum multiflorum (PM), has been reported to exert both protective and toxic effects in several cell types. However, the effects and underlying mechanisms of action of emodin in hepatic cells are still obscure. METHODS: The present study used the normal human liver cell line L02 to investigate the effects and mechanisms of emodin in hepatic cells. After treatment with emodin, L02 cells were examined for viability, apoptosis and autophagy with the Cell Counting Kit-8 (CCK-8), annexin V/PerCP staining and GFP-LC3 plasmid transfection. The expression of proteins including cleaved caspase-3, LC3B-I/II, p-PI3K, PI3K, p-AKT, AKT, p-mTOR, mTOR and actin was examined by using Western blot. RESULTS: Emodin significantly inhibited the viability of and induced apoptosis in L02 cells in a dose- and time-dependent manner. In addition, emodin increased the number of GFP-LC3 puncta in L02 cells and upregulated the expression of LC3B-II compared to those in control cells. Furthermore, emodin significantly decreased the expression of p-PI3K, p-AKT and p-mTOR in a dose-dependent manner compared to that in control cells without altering the expression of PI3K, AKT and mTOR. Notably, cotreatment with emodin and 3-methyladenine (3-MA) or rapamycin significantly increased and decreased the apoptosis rate of L02 cells, respectively, compared to that of cells treated with emodin alone. CONCLUSION:  In conclusion, emodin exhibited cytotoxicity in the L02 human hepatic cell line by promoting apoptosis, and it also induced autophagy through the suppression of the PI3K/AKT/mTOR signalling pathway. The autophagy could play a protective role following emodin treatment.

Lycorine Induces Mitochondria-Dependent Apoptosis in Hepatoblastoma HepG2 Cells Through ROCK1 Activation.

Lycorine, a naturally occurring compound extracted from the Amaryllidaceae plant family, has been reported to exhibit antitumor activity in various cancer cell types. In the present study, we investigated the molecular mechanisms underlying lycorine-induced apoptosis in hepatoblastoma HepG2 cells. We found that lycorine induced mitochondria-dependent apoptosis in HepG2 cells accompanied by mitochondrial permeability transition pore (mPTP) opening, mitochondrial membrane potential (MMP) loss, adenosine triphosphate (ATP) depletion, Ca2+ and cytochrome c (Cyto C) release, as well as caspase activation. Furthermore, we found Rho associated coiled-coil containing protein kinase 1 (ROCK1) cleavage/activation played a critical role in lycorine-induced mitochondrial apoptosis. In addition, the ROCK inhibitor Y-27632 was employed, and we found that co-treatment with Y-27632 attenuated lycorine-induced mitochondrial injury and cell apoptosis. Meanwhile, an in vivo study revealed that lycorine inhibited tumor growth and induced apoptosis in a HepG2 xenograft mouse model in association with ROCK1 activation. Taken together, all these findings suggested that lycorine induced mitochondria-dependent apoptosis through ROCK1 activation in HepG2 cells, and this may be a theoretical basis for lycorine's anticancer effects.

Mitochondrial fission and mitophagy depend on cofilin-mediated actin depolymerization activity at the mitochondrial fission site.

Mitochondria fission and mitophagy are fundamentally crucial to cellular physiology and play important roles in cancer progression. Developing a comprehensive understanding of the molecular mechanism underlying mitochondrial fission and mitophagy will provide novel strategies for cancer prevention and treatment. Actin has been shown to participate in mitochondrial fission and mitophagy regulation. Cofilin is best known as an actin-depolymerizing factor. However, the molecular mechanism by which cofilin regulates mitochondrial fission and mitophagy remains largely unknown. Here we report that knockdown of cofilin attenuates and overexpression of cofilin potentiates mitochondrial fission as well as PINK1/PARK2-dependent mitophagy induced by staurosporine (STS), etoposide (ETO), and carbonyl cyanide 3-chlorophenylhydrazone (CCCP). Cofilin-mediated-PINK1 (PTEN-induced putative kinase 1) accumulation mainly depends on its regulation of mitochondrial proteases, including peptidase mitochondrial processing beta (MPPß), presenilin-associated rhomboid-like protease (PARL), and ATPase family gene 3-like 2 (AFG3L2), via mitochondrial membrane potential activity. We also found that the interaction and colocalization of G-actin/F-actin with cofilin at mitochondrial fission sites undergo constriction after CCCP treatment. Pretreatment with the actin polymerization inhibitor latrunculin B (LatB) increased and actin-depolymerization inhibitor jasplakinolide (Jas) decreased mitochondrial translocation of actin induced by STS, ETO, and CCCP. Both LatB and Jas abrogated CCCP-mediated mitochondrial fission and mitophagy. Our data suggest that G-actin is the actin form that is translocated to mitochondria, and the actin-depolymerization activity regulated by cofilin at the mitochondrial fission site is crucial for inducing mitochondrial fission and mitophagy.

Polyphyllin I induces mitophagic and apoptotic cell death in human breast cancer cells by increasing mitochondrial PINK1 levels.

The molecular mechanisms underlying the anti-breast cancer effects of polyphyllin I, a natural compound extracted from Paris polyphylla rhizomes, are not fully understood. In the present study, we found that polyphyllin I induces mitochondrial translocation of DRP1 by dephosphorylating DRP1 at Ser637, leading to mitochondrial fission, cytochrome c release from mitochondria into the cytosol and, ultimately apoptosis. Polyphyllin I also increased the stabilization of full-length PINK1 at the mitochondrial surface, leading to the recruitment of PARK2, P62, ubiquitin, and LC3B-II to mitochondria and culminating in mitophagy. PINK1 knockdown markedly suppressed polyphyllin I-induced mitophagy and enhanced polyphyllin I-induced, DRP1-dependent mitochondrial fission and apoptosis. Furthermore, suppression of DRP1 by mdivi-1 or shRNA inhibited PINK1 knockdown/polyphyllin I-induced mitochondrial fragmentation and apoptosis, suggesting that PINK1 depletion leads to excessive fission and, subsequently, mitochondrial fragmentation. An in vivo study confirmed that polyphyllin I greatly inhibited tumor growth and induced apoptosis in MDA-MB-231 xenografts, and these effects were enhanced by PINK1 knockdown. These data describe the mechanism by which PINK1 contributes to polyphyllin I-induced mitophagy and apoptosis and suggest that polyphyllin I may be an effective drug for breast cancer treatment.

Xinqin exhibits the anti-allergic effect through the JAK2/STAT5 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Xinqin, a polyherbal medicine, is an important traditional Chinese herbal formula used in traditional oriental medicine for treatment of allergic rhinitis (AR). The formula is based on the Chinese Pharmacopoeia AIM OF THE STUDY: Previously, Xinqin exhibited potent anti-allergic effect in a guinea pig model of AR. In this study, we explored the molecular mechanism of the anti-allergic effect mediated by Xinqin. MATERIALS AND METHODS: AR was induced in guinea pigs (Hartley) with toluene-2, 4-diisocyanate (TDI) in vivo and in HMC-1 mast cells with A23187/phorbol 12-myristate-13-acetate (PMA) in vitro. The releases of allergic inflammatory mediators such as histamine, leukotriene (LT) D4, immunoglobulin (Ig) E, TNF-α, and IL-6 were analyzed for allergy. The mast cell degranulation was displayed in HMC-1 mast cells. The activities of janus protein kinase 2 (JAK2), signal transduction and activator of transcription 5 (STAT5) and suppressor of cytokine signaling 3 (SOCS3) were evaluated by Western blot. RESULTS: Treatment with Xinqin resulted in AR symptoms and decreases in levels of histamine, LTD4, IgE, TNF-α, and IL-6 in serum of guinea pig model of AR and in A23187/PMA-stimulated HMC-1 mast cells. Treatment with Xinqin also inhibited cell degranulation in A23187/PMA-stimulated HMC-1 mast cells. The JAK2/STAT5 signaling pathway could play an important role in the anti-allergic activity mediated by Xinqin. CONCLUSIONS: Xinqin exerts the anti-allergic effect by modulating mast cell-mediated allergic responses by down-regulating JAK2/STAT5 signaling pathway. Results from this study provide a mechanistic basis for the application of Xinqin in the treatment of AR.

Mitochondrial translocation of cofilin is required for allyl isothiocyanate-mediated cell death via ROCK1/PTEN/PI3K signaling pathway.

BACKGROUND: Cofilin is a member of the actin depolymerizing factor (ADF)/cofilin family, which regulates actin dynamics. Increasing evidence suggests that mitochondrial translocation of cofilin appears necessary for the regulation of apoptosis. RESULTS: We report that allyl isothiocyanate (AITC) potently induces mitochondria injury and apoptosis. These events were accompanied by a loss of polymerized filamentous actin (F-actin) and increase in unpolymerized globular actin (G-actin). AITC also induces dephosphorylation of cofilin through activation of PP1 and PP2A. Only dephosphorylated cofilin binds to G-actin and translocates to mitochondria during AITC-mediated apoptosis. Mechanistic study revealed that interruption of ROCK1/PTEN/PI3K signaling pathway plays a critical role in AITC-mediated dephosphorylation and mitochondrial translocation of cofilin and apoptosis. Our in vivo study also showed that AITC-mediated inhibition of tumor growth of mouse leukemia xenograft model is in association with dephosphorylation of cofilin. CONCLUSIONS: These findings support a model in which induction of apoptosis by AITC stems primarily from activation of ROCK1 and PTEN, and inactivation of PI3K, leading in turn to activation of PP1 and PP2A, resulting in dephosphorylation of cofilin, which binds to G-actin and translocates to mitochondria, culminating in the dysfunction of mitochondria, release of cytochrome c and apoptosis.