Danshen (Salvia miltiorrhiza) extract inhibits proliferation of breast cancer cells via modulation of Akt activity and p27 level

Danshen is widely used in traditional Chinese medicine, often in combination with other herbs. To check the effect of Danshen on the proliferation of breast cancer cells, Danshen extract was used to treat MCF‐7 and MCF‐7 HER2 cells, the latter of which overexpresses HER2. HER2 is a receptor tyrosine kinase, and is involved in signal transduction pathways leading to tumor cell proliferation. MTT and cell proliferation assays revealed that Danshen strongly inhibited the proliferation of both MCF‐7 vec cells and MCF‐7 HER2 cells. Flow cytometry analyses indicated that Danshen induced cell cycle delay in the G1 phase. HER2 expression was shown to confer resistance to Danshen‐induced inhibition of proliferation and cell cycle delay, suggesting that HER2 is responsible for the resistance to Danshen. Danshen treatment induced the down‐regulation of Akt phosphorylation and an increase in p27 in MCF‐7 vec and MCF‐7 HER2 cells. Nevertheless, MCF‐7 HER2 cells were more resistant to the Danshen‐induced inhibition of Akt phosphorylation and p27 up‐regulation. Copyright © 2009 John Wiley & Sons, Ltd.     

Apoptotic effect of lambertianic acid through AMPK/FOXM1 signaling in MDA‐MB231 breast cancer cells

Though lambertianic acid (LA) was known to exert antitumor effect in liver and prostate cancers, its underlying anticancer mechanism is never reported in breast cancers so far. Thus, in this study, apoptotic mechanism of LA was elucidated in MDA‐MB‐231 breast cancer cells. Here, LA increased cytotoxicity in MCF‐7 and MDA‐MB‐231 cells; enhanced sub‐G1 population, G2/M arrest, and cleaved poly(ADP‐ribose) polymerase; activated phosphorylation of AMP‐activated protein kinase (AMPK)/acetyl‐CoA carboxylase pathway; and also suppressed phosphorylation of AKT and the expression of forkhead box M1 (FOXM1), X‐linked inhibitor of apoptosis protein, B‐cell lymphoma 2, and CyclinB1 in MDA‐MB‐231 cells. Furthermore, AMPK inhibitor compound C reversed the effect of LA on FOXM1, Cyclin B1, and cleaved poly(ADP‐ribose) polymerase in MDA‐MB‐231 cells. Notably, immunoprecipitation revealed that LA disturbed the direct binding of AKT and FOXM1 in MDA‐MB‐231 cells. Overall, these findings suggest that LA‐induced apoptosis is mediated via activation of AMPK and inhibition of AKT/FOXM1 signaling pathway.     

Plumbagin induces apoptosis in Her2-overexpressing breast cancer cells through the mitochondrial-mediated pathway

Breast cancer is the leading cause of death-related cancers in women. Approximately 30% of breast cancers overexpress the Her2 oncogene, which is associated with a poor prognosis and increased resistance to chemotherapy. Plumbagin (1), a constituent of species in the plant genera Drosera and Plumbago, displays antineoplastic activity toward various cancers. The present study was aimed at determining the anticancer potential of 1 toward Her2-overexpressing breast cancer cells and defining the mode of cell death induced in these cells. The results showed that 1 exhibited high antiproliferative activity toward the Her2-overexpressing cell lines SKBR3 and BT474. The antiproliferative activity of 1 was associated with apoptosis-mediated cell death, as revealed by caspase activation and an increase in the sub-G1 fraction of the cell cycle. Compound 1 increased the levels of the proapoptotic Bcl-2 family of proteins and decreased the level of the antiapoptotic Bcl-2 protein in SKBR3 and BT474 cells. Thus, these findings indicate that 1 induces apoptosis in Her2-overexpressing breast cancers through the mitochondrial-mediated pathway and suggest its potential for further investigation for the treatment of Her2-overexpressing breast cancer.     

Activation of AMP‐activated Protein Kinase and Phosphorylation of Glycogen Synthase Kinase3 β Mediate Ursolic Acid Induced Apoptosis in HepG2 Liver Cancer Cells

Despite the antitumour effect of ursolic acid observed in several cancers, the underlying mechanism remains unclear. Thus, in the present study, the roles of AMP‐activated protein kinase (AMPK) and glycogen synthase kinase 3 beta (GSK3β) were examined in ursolic acid induced apoptosis in HepG2 hepatocellular carcinoma cells. Ursolic acid significantly exerted cytotoxicity, increased the sub‐G1 population and the number of ethidium homodimer and terminal deoxynucleotidyl transferase(TdT) mediated dUTP nick end labeling positive cells in HepG2 cells. Also, ursolic acid enhanced the cleavages of poly‐ADP‐ribose polymerase (PARP) and caspase3, attenuated the expression of astrocyte elevated gene (AEG1) and survivin in HepG2 cells. Interestingly, ursolic acid increased the phosphorylation of AMPK and coenzyme A carboxylase and also enhanced phosphorylation of GSK3β at inactive form serine 9, whereas ursolic acid attenuated the phosphorylation of AKT and mTOR in HepG2 cells. Conversely, AMPK inhibitor compound C or GSK3β inhibitor SB216763 blocked the cleavages of PARP and caspase 3 induced by ursolic acid in HepG2 cells. Furthermore, proteosomal inhibitor MG132 suppressed AMPK activation, GSK3β phosphorylation, cleaved PARP and deceased AEG‐1 induced by ursolic acid in HepG2 cells. Overall, our findings suggest that ursolic acid induced apoptosis in HepG2 cells via AMPK activation and GSK3β phosphorylation as a potent chemopreventive agent. Copyright © 2013 John Wiley & Sons, Ltd.     

Reactive Oxygen Species‐Mediated Activation of AMP‐Activated Protein Kinase and c‐Jun N‐terminal Kinase Plays a Critical Role in Beta‐Sitosterol‐Induced Apoptosis in Multiple Myeloma U266 cells

Although beta‐sitosterol has been well known to have anti‐tumor activity in liver, lung, colon, stomach, breast and prostate cancers via cell cycle arrest and apoptosis induction, the underlying mechanism of anti‐cancer effect of beta‐sitosterol in multiple myeloma cells was never elucidated until now. Thus, in the present study, the role of reactive oxygen species (ROS) in association with AMP‐activated protein kinase (AMPK) and c‐Jun N‐terminal kinase (JNK) pathways was demonstrated in beta‐sitosterol‐treated multiple myeloma U266 cells. Beta‐sitosterol exerted cytotoxicity, increased sub‐G₁ apoptotic population and activated caspase‐9 and ‐3, cleaved poly (ADP‐ribose) polymerase (PARP) followed by decrease in mitochondrial potential in U266 cells. Beta‐sitosterol promoted ROS production, activated AMPK, acetyl‐CoA carboxylase (ACC) and JNK in U266 cells. Also, beta‐sitosterol attenuated the phosphorylation of AKT, mammalian target of rapamycin and S6K, and the expression of cyclooxygenase‐2 and VEGF in U266 cells. Conversely, AMPK inhibitor compound C and JNK inhibitor SP600125 suppressed apoptosis induced by beta‐sitosterol in U266 cells. Furthermore, ROS scavenger N‐acetyl L‐cysteine attenuated beta‐sitosterol‐mediated sub‐G₁ accumulation, PARP cleavage, JNK and AMPK activation in U266 cells. Overall, these findings for the first time suggest that ROS‐mediated activation of cancer metabolism‐related genes such as AMPK and JNK plays an important role in beta‐sitosterol‐induced apoptosis in U266 multiple myeloma cells. Copyright © 2013 John Wiley & Sons, Ltd.     

Tanshinone IIA Induces Autophagic Cell Death via Activation of AMPK and ERK and Inhibition of mTOR and p70 S6K in KBM‐5 Leukemia Cells

Although tanshinone IIA (Tan IIA) from Salviae miltiorrhizae was known to induce apoptosis in various cancers, its underlying mechanism of autophagic cell death was not reported yet. Thus, in the present study, the molecular mechanism of autophagic cell death by Tan IIA was investigated in KBM‐5 leukemia cells. Tan IIA significantly increased the expression of microtubule‐associated protein light chain 3 (LC3) II as a hallmark of autophagy in western blotting and immunofluorescence staining. Tan IIA augmented the phosphorylation of adenosine monophosphate‐activated protein kinase (AMPK) and attenuated the phosphorylation of mammalian target of rapamycin (mTOR) and p70 S6K in a dose‐dependent manner. Conversely, autophagy inhibitor 3‐methyladenine partly reversed the cytotoxicity and the phosphorylation of AMPK, mTOR and p70 S6K induced by Tan IIA in KBM‐5 leukemia cells. In addition, Tan IIA dramatically activated the extracellular signal regulated kinase (ERK) signaling pathway including Raf, ERK and p90 RSK in a dose‐dependent and time‐dependent manner. Consistently, ERK inhibitor PD184352 suppressed LC3‐II activation induced by Tan IIA, whereas PD184352 and PD98059 did not affect poly (ADP‐ribose) polymerase cleavage and sub‐G1 accumulation induced by Tan IIA in KBM‐5 leukemia cells. Furthermore, Tan IIA could induce autophagy via LC3‐II activation in various cancer cells such as prostate (PC‐3), multiple myeloma (U266), lung (NCI‐H460), and breast (MDA‐MB‐231) cells. Overall, these findings suggest that Tan IIA induces autophagic cell death via activation of AMPK and ERK and inhibition of mTOR and p70 S6K in KBM‐5 cells as a potent natural compound for leukemia treatment. Copyright © 2013 John Wiley & Sons, Ltd.     

Induction of apoptotic cell death by Pharbitis nil extract in HER2-overexpressing MCF-7 cells

Aim of the study

We performed this study to investigate the anti-cancer activity of Pharbitis nil (PN) ethanol extract which has been used for herbal medicinal treatment against diseases in East Asia.

Materials and methods

We analyzed the effects of PN extract on proliferation of breast cancer cell lines, MCF-7 control vector (vec) and MCF-7 human epidermal growth factor receptor 2 (HER2) cells engineered to overexpress oncogenic HER2 via retroviral infection. We performed the proliferation assay to measure the growth rate of the cells. FACS analysis was used to analyze the cell cycle. Western blot analysis was used to investigate the effect of PN on the level and activation of intracellular molecules.

Results

We found that PN extract inhibited the proliferation of both MCF-7 vec and MCF-7 HER2 cells. This growth inhibition was accompanied with the increase of sub G0/G1 apoptotic fractions. When we check the efficiency of PN on the level of intracellular signaling molecules, we found that PN extract induced the inhibition of phosphorylation of HER2 and its downstream effectors, Akt and extracellular signal-regulated kinases (ERK). Active forms of both Akt and ERK were gradually decreased in PN-treated MCF-7 vec and MCF-7 HER2 cells suggesting that the growth suppressive activity of PN is related to signaling pathway. The level of cyclin D also diminished in PN-treated both cells suggesting that PN may inhibit the growth of MCF-7 vec and MCF-7 HER2 cells by perturbing cell cycle progression. It should be noted that PN decreased the growth rate of both MCF-7 vec and MCF-7 HER2 cells without changing the level and activation of p53.

Conclusion

PN extract suppressed the proliferation rate of HER-2 overexpressing MCF-7 breast cancer cells inducing apoptotic cell death in vitro. Our data demonstrates that PN extracts contain useful anti-tumor activity especially against HER2 overexpressing breast cancer.     

Retracted: Ganoderic acid A alleviates hypoxia‐induced apoptosis,autophagy, and inflammation in rat neural stem cells through the PI3K/AKT/mTOR pathways

Effects of ganoderic acid A (GAA), a lanostane triterpene, on hypoxia‐ischemia encephalopathy (HIE) remain unclear. We aimed to figure out the specific role of GAA in hypoxia‐treated neural stem cells (NSCs) as well as the regulatory mechanisms. Primary rat NSCs were incubated under hypoxia to simulate HIE. Viability and apoptosis of hypoxia‐injured NSCs were measured by cell counting kit‐8 and flow cytometry assays, respectively. Proteins related to apoptosis, autophagy, and the PI3K/AKT/mTOR pathways were evaluated by Western blot analysis. LY294002 and rapamycin were added to inhibit the PI3K/AKT pathway and mTOR pathway, respectively. Enzyme‐linked immunosorbent assay was carried out to test the release of proinflammatory cytokines. We found that hypoxia‐induced decrease of cell viability, increases of apoptotic cells and autophagy, and the release of IL‐6, IL‐1β, and TNF‐α were all attenuated by GAA stimulation. Activation of caspases induced by hypoxia was alleviated by GAA. Furthermore, we found that inhibition of the PI3K/AKT pathway eliminated the effects of GAA on apoptosis and proinflammatory cytokines release in hypoxia‐injured NSCs. Meanwhile, inhibition of the mTOR pathway abrogated the effects of GAA on cell autophagy in hypoxia‐injured NSCs. In conclusion, GAA alleviated hypoxia‐induced injury in NSCs might be through activating the PI3K/AKT and mTOR pathways.     

6-Methoxydihydrosanguinarine induces apoptosis and autophagy in breast cancer MCF-7 cells by accumulating ROS to suppress the PI3K/AKT/mTOR signaling pathway

6-Methoxydihydrosanguinarine (6-MDS) is a natural benzophenanthridine alkaloid extracted from Hylomecon japonica (Thunb.) Prantl. It is the first time to explore the effect and mechanism of 6-MDS in breast cancer. Network pharmacology, molecular docking, and molecular dynamics simulation technology were adopted to identify the potential targets and pathways of 6-MDS in breast cancer. Besides, cell proliferation, apoptosis, and western blotting assays were conducted to investigate the effect of 6-MDS on MCF-7 cells. Network pharmacology, molecular docking, and molecular dynamics simulation results confirmed the effect of 6-MDS on resisting breast cancer via the PI3K/AKT/mTOR signaling pathway. In addition, the functional experiments results demonstrated that 6-MDS inhibited proliferation and induced apoptosis and autophagy. The autophagy inhibitor chloroquine and the silence of Atg5 augmented the effect of 6-MDS on promoting apoptosis. Furthermore, 6-MDS suppressed the PI3K/AKT/mTOR signaling pathway, and the PI3K inhibitor LY294002 enhanced these changes and promoted the 6-MDS pro-apoptotic and autophagy effects. 6-MDS triggered the generation of reactive oxygen species. The pretreatment with antioxidant N-acetyl-L-cysteine reversed the changes induced by 6-MDS, including increases in apoptosis and autophagy and inhibition of the PI3K/AKT/mTOR pathway. In conclusion, 6-MDS induces the apoptosis and autophagy of MCF-7 cells by ROS accumulation to suppress the PI3K/AKT/mTOR signaling pathway.     

Pachastrissamine from Pachastrissa sp. Inhibits Melanoma Cell Growth by Dual Inhibition of Cdk2 and ERK‐mediated FOXO3 Downregulation

Melanoma cells are relatively resistant to apoptosis compared with other tumor cell types, and thus, chemotherapy, radiotherapy and immunotherapy are not effective in treating melanoma. Pachastrissamine (PA) exhibits cytotoxic activity and promotes apoptosis in several cancer cells. However, its specific molecular mechanisms have not been characterized fully. This study investigated the antimelanoma effect of PA, an anhydrophytosphingosine derived from marine sponge, and its underlying molecular mechanisms. The data demonstrated that treatment with PA inhibited the phosphorylation of ERK and subsequent ERK‐mediated FOXO3 phosphorylation in melanoma cells. Interestingly, PA did not inhibit AKT‐mediated FOXO3 phosphorylation. Therefore, it appears that PA‐induced apoptosis results from the inhibition of ERK. Furthermore, intravenous administration of PA was found to suppress melanoma cell growth in a C57BL6 mouse without causing side effects. Additionally, PA inhibited the production of Cdk2, which is involved in cell cycle regulation. Taken together, inhibition of melanoma cell growth by PA is a result of the inhibition of ERK‐mediated FOXO3 downregulation and decreased Cdk2 levels. The results of this study imply that dual inhibition of the ERK pathway and cell cycle progression could be an effective approach to control the growth of melanoma cells. Copyright © 2012 John Wiley & Sons, Ltd.     

Baicalein Triggers Autophagy and Inhibits the Protein Kinase B/Mammalian Target of Rapamycin Pathway in Hepatocellular Carcinoma HepG2 Cells

Baicalein (BA), isolated from the Chinese medicinal herb Scutellariae radix (Huangqin in Chinese), is a flavonoid with various pharmacological effects. Herein, we found that BA only slightly reduced the cell viability on HepG2 cells after 24‐h treatment as determined by 3‐(4, 5‐dimethylthiazol‐2‐yl)‐2, 5‐diphenyl tetrazolium bromide (MTT) assay. However, BA (50 μM) effectively blocked the colony formation. Meanwhile, BA remarkably induced the formation of autophagosomes after 24‐h treatment as determined by immunofluorescence with monodansylcadaverine staining as well as transmission electron microscopy, respectively. Moreover, BA obviously up‐regulated the expression of microtubule‐associated protein 1A/1B‐light chain 3‐II in concentration‐dependent and time‐dependent manners in HepG2 cells. When combined with the autophagy inhibitor chloroquine and BA, the cell viability and colony formation were significantly decreased, indicating that BA triggered protective autophagy, which prevented cell death. Further study showed that BA concentration‐dependently and time‐dependently decreased the expression of p‐AKT (S473), p‐ULK1 (S757) and p‐4EBP1 (T37 and S65), suggesting the involvement of protein kinase B (AKT)/mammalian target of rapamycin (mTOR) in BA‐triggered autophagy. Copyright © 2015 John Wiley & Sons, Ltd.     

Synergistic Effect of TPD7 and Berberine against Leukemia Jurkat Cell Growth through Regulating Ephrin‐B2 Signaling

TPD7, a novel biphenyl urea taspine derivative, and berberine have presented inhibition on VEGFR2 that can be regulated by ephrin‐B2 reverse signaling through interactions with the PDZ domain. The purpose of this study is to investigate the inhibitory effect of the combination of TPD7 and berberine (TAB) on T‐cell acute lymphoblastic leukemia cell growth. TPD7 and berberine together synergistically inhibited the proliferation of Jurkat cells. Also, the combination of TAB induced G₁‐phase cell‐cycle arrest by downregulating the level of cyclin D1, cyclin E, and CDC2. Furthermore, the combination of TAB significantly enhanced apoptosis in Jurkat cells, and the apoptosis most likely resulted from the modulation of the level of Bcl‐2 family members. Most importantly, the concomitant treatment simultaneously regulated the ephrin‐B2 and VEGFR2 signaling, as well as modulated the MEK/ERK and PTEN/PI3K/AKT/mTOR signaling. Therefore, the combination treatment of TAB may be a promising therapeutic method in treating T‐cell acute lymphoblastic leukemia. Copyright © 2017 John Wiley & Sons, Ltd.     

Natural product-derived antitumor compound phenethyl isothiocyanate inhibits mTORC1 activity via TSC2

Phenethyl isothiocyanate (1) is a natural dietary phytochemical with cytostatic, cytotoxic, and antitumor activity. The effects of 1 were investigated on the activity of mTOR, a kinase that enhances the translation of many RNAs encoding proteins critical for cancer cell growth, including the angiogenesis regulator HIF1α. Compound 1 effectively blocked HIF1α RNA translation in MCF7 breast cancer cells, and this was associated with reduced phosphorylation of 4E-BP1 and p70 S6K, well-characterized downstream substrates of the mTOR-containing mTORC1 complex. Compound 1 also inhibited mTORC1 activity in mouse embryonic fibroblasts (MEFs). The 1-mediated inhibition of mTORC1 activity appeared to be independent of the upstream regulators PTEN, AKT, ERK1/2, and AMPK. By contrast, 1-mediated inhibition of mTORC1 activity was dependent on the presence of TSC2, part of a complex that regulates mTORC1 activity negatively. TSC2-deficient MEFs were resistant to 1-mediated inhibition of p70 S6K phosphorylation. TSC2-deficient MEFs were also partially resistant to 1-mediated growth inhibition. Overall, the present results confirm that 1 inhibits mTORC1 activity. This is dependent on the presence of TSC2, and inhibition of mTORC1 contributes to optimal 1-induced growth inhibition. Inhibition of RNA translation may be an important component of the antitumor effects of phenethyl isothiocyanate.     

Retracted: Ganoderic Acid A exerts the cytoprotection against hypoxia‐triggered impairment in PC12 cells via elevating microRNA‐153

Ganoderic Acid A (GAA) is often applied for healing cardiovascular and cerebrovascular ailments, but the influences in cerebral ischemia injury are still hazy. The research delved into the functions of GAA in hypoxia‐triggered impairment in PC12 cells. PC12 cells received hypoxia management for 12 hr, and subsequently, cell viability, migration, apoptosis, and correlative protein levels were assessed. After preprocessing with GAA, above cell behaviors were monitored again. The vector of microRNA (miR)‐153 inhibitor was utilized for PC12 cell transfection to further explore the functions of miR‐153 in hypoxia‐impaired cells. Pathways of phosphatidylinositol 3‐kinase (PI3K)/protein kinase B (AKT) and mammalian target of rapamycin (mTOR) were investigated via executing western blot for uncovering the latent mechanism. Results revealed that hypoxia disposition triggered PC12 cells impairment via restraining cell viability and migration and accelerating apoptosis. However, GAA visibly mollified hypoxia‐provoked impairment in PC12 cells. Interestingly, the enhancement of miR‐153 triggered by GAA was observed in hypoxia‐impaired PC12 cells. After miR‐153 inhibitor transfection, the protective functions of GAA in hypoxia‐impaired PC12 cells were dramatically inversed. Furthermore, GAA caused PI3K/AKT and mTOR activations via enhancement of miR‐153 in hypoxia‐impaired PC12 cells. The findings evinced that GAA exhibited the protective functions in PC12 cells against hypoxia‐evoked impairment through activating PI3K/AKT and mTOR via elevating miR‐153.     

槲皮素对5-FU诱导的结直肠癌SW480细胞耐药及自噬调控机制研究

目的:探讨槲皮素(Que)调控5-氟尿嘧啶(5-FU)诱导的结直肠癌SW480细胞耐药及自噬的作用及机制。方法:采用浓度递增诱导法建立5-FU耐药细胞株SW480/5-FU,MTT法设置Que高、中、低剂量组(10、20、40 μmol/L)和对照组。MTT法检测各组细胞的5-FU耐药性,TUNEL染色法检测细胞凋亡,免疫荧光检测细胞自噬活性,Western blot检测细胞p糖蛋白(Pgp)、多药耐药相关蛋白1(MRP1)、三磷酸腺苷结合转运蛋白G超家族成员2抗体(ABCG2)、LC3Ⅰ、LC3Ⅱ、p62、丝/苏氨酸激酶(AKT)、p-AKT、雷帕霉素靶蛋白(mTOR)和p-mTOR蛋白表达。结果:5-FU抑制SW480/5-FU细胞的半抑制浓度(IC₅₀)明显高于SW480细胞(P<0.05),耐药指数(RI)=13.74。与对照组相比,Que中、高剂量组5-FU抑制SW480/5-FU细胞的IC₅₀明显下降(P<0.05),耐药逆转倍数分别为2.27和4.03。与对照组相比,Que中、高剂量组细胞抑制率、凋亡率和自噬活性明显升高(均P<0.05),蛋白的表达均明显升高(均P<0.05),P-gp、MRP1、ABCG2、p62、p-AKT/AKT和p-mTOR/mTOR蛋白表达明显下降(均P<0.05)。结论:Que可以以剂量依赖性的方式逆转SW480/5-FU细胞的5-FU耐药性,诱导细胞自噬,其作用机制可能与抑制AKT/mTOR的磷酸化有关。     

(−)‐Epigallocatechin‐3‐Gallate Ameliorates Photodynamic Therapy Responses in an In Vitro T Lymphocyte Model

(?)‐Epigallocatechin‐3‐gallate (EGCG), the most abundant polyphenolic constituent in green tea, is known as a powerful antioxidant but concomitantly possesses a prooxidant property. We investigated the effect of EGCG on phloxine B (PhB)‐induced photocytotoxicity in human T lymphocytic leukemia Jurkat cells. EGCG significantly potentiated PhB‐induced photocytotoxic effects, including the inhibition of cell proliferation, DNA fragmentation, and caspase‐3 activity induction in Jurkat cells. Catalase attenuated the enhanced cytotoxicity by EGCG, suggesting the involvement of extracellularly produced hydrogen peroxide. Indeed, EGCG significantly enhanced extracellular hydrogen peroxide formation induced by photo‐irradiated PhB. The EGCG also enhanced intracellular reactive oxygen species accumulation, c‐Jun N ‐terminal kinase (JNK) phosphorylation, and interferon‐γ (IFN‐γ) gene expression, all of which are involved in PhB‐induced apoptosis. Taken together, our data suggest that EGCG is capable of potentiating photodynamic therapy responses, presumably through the intracellular oxidative stress‐sensitive JNK/IFN‐γ pathway by exogenous hydrogen peroxide formation. Copyright © 2014 John Wiley & Sons, Ltd.     

Hypolipogenic effects of Icariside E4 via phosphorylation of AMPK and inhibition of MID1IP1 in HepG2 cells

Though icariside E4 (IE4) is known to have anti-noceptive, anti-oxidant, anti-Alzheimer and anti-inflammatory effects, there was no evidence on the effect of IE4 on lipid metabolism so far. Hence, the hypolipogenic mechanism of IE4 was investigated in HepG2 hepatocellular carcinoma cells (HCCs) in association with MID1 Interacting Protein 1(MID1IP1) and AMPK signaling. Here, IE4 did not show any toxicity in HepG2 cells, but reduced lipid accumulation in HepG2 cells by Oil Red O staining. MID1IP1 depletion decreased the expression of SREBP-1c and fatty acid synthase (FASN) and induced phosphorylation of ACC in HepG2 cells. Indeed, IE4 activated phosphorylation of AMPK and ACC and inhibited the expression of MID1IP1 in HepG2 cells. Furthermore, IE4 suppressed the expression of SREBP-1c, liver X receptor-α (LXR), and FASN for de novo lipogenesis in HepG2 cells. Interestingly, AMPK inhibitor compound C reversed the ability of IE4 to reduce MID1IP1, SREBP-1c, and FASN and activate phosphorylation of AMPK/ACC in HepG2 cells, indicating the important role of AMPK/ACC signaling in IE4-induced hypolipogenic effect. Taken together, these findings suggest that IE4 has hypolipogenic potential in HepG2 cells via activation of AMPK and inhibition of MID1IP1 as a potent candidate for treatment of fatty liver disease.     

Triggering of p38 MAPK and JNK Signaling is Important for Oleanolic Acid‐Induced Apoptosis via the Mitochondrial Death Pathway in Hypertrophic Scar Fibroblasts

Hypertrophic scarring is characterized by collagen overproduction and excessive deposition of extracellular matrix. No consensus arises currently about the best therapeutics to produce complete and permanent improvement of scars with few side effects. In the present study, the mechanism of oleanolic acid (OA)‐induced apoptosis in hypertrophic scar fibroblasts (HSFs) was investigated for the first time. OA activated the protein phosphorylation of p38 MAPK and JNK but not ERK. OA did not antagonize the inhibitory effects of SB203580 on p38 MAPK pathway activity but sharply enhanced JNK phosphorylation when HSFs were pretreated with SB203580. Similarly, the inhibition of JNK signal pathway activation by pretreatment with SP600125 facilitated the protein phosphorylation of p38 MAPK caused by OA. Inhibition of p38 MAPK and/or JNK by inhibitors significantly enhanced cell viability and OA only partially depressed the increased cell viability. Moreover, OA increased Bax translocation, MMP loss, mitochondrial cytochrome c and AIF release, Bax and caspase‐3 protein expression and the ratio of Bax to Bcl‐2, decreased Bcl‐2 protein expression, and elevated the mRNA expression of Apaf‐1, caspase‐9, and capase‐3. These results suggest that OA elicits apoptosis through triggering of p38 MAPK and JNK signaling and activation of the mitochondrial death pathway. OA might be a good and useful natural drug against hypertrophic scars. Copyright © 2014 John Wiley & Sons, Ltd.