Butein induces apoptotic cell death of human cervical cancer cells.

Butein is a chalcone, a flavonoid that is widely biosynthesized in plants. Butein has been identified to possess varied pharmacological activity and is extractable from traditional Chinese medicinal herbs, therefore applicable for disease treatment. Recently, in vitro and in vivo studies have shown that butein may induce apoptotic cell death in various human cancer cells. In this study we investigated the apoptotic effect of butein and the underlying mechanisms in human cervical cancer cells. Two cell lines, C-33A and SiHa cells, were treated with butein at different dosages for different durations. The effect of butein on cell viability was assessed by MTT assay, which revealed that butein exerted cytotoxicity in both cervical cancer cells in a dose- and time-dependent fashion. Apoptotic pathway-related factors in the butein-treated cervical cancer cells were then examined. JC-1 flow cytometry, cytochrome c assay, and caspase activity assays demonstrated that butein disturbed mitochondrial transmembrane potential, and increased cytosolic cytochrome c levels and caspase activities in both cervical cancer cells. Western blot analysis revealed that butein downregulated anti-apoptotic protein Bcl-xL and led to proteolytic cleavage of poly (ADP-ribose) polymerase. In addition, butein decreased expressions of the inhibitor of apoptosis (IAP) proteins, including X-linked IAP, survivin, and cellular IAP-1. The findings of this study suggest that butein can decrease cervical cancer cell viability via a pro-apoptotic effect, which involves inhibition of the IAP proteins and activation of both extrinsic and intrinsic pro-apoptotic pathways. Therefore, butein may be applicable for cervical cancer treatment.

Transcriptome analysis reveals a positive role for nerve growth factor in retinol metabolism in primary rat hepatocytes.

Up-regulation of nerve growth factor (NGF) in parenchymal hepatocytes with cholestatic injury has been previously demonstrated to exert hepatoprotective effects in an autocrine manner; however, the overall impact of NGF up-regulation remains elusive. This study aimed to profile the effects of exogenous NGF on cultured primary rat hepatocytes using transcriptome analysis. Total RNA was isolated from hepatocytes with and without 24 h of NGF exposure, and subjected to RNA enrichment by PCR and RNA sequencing procedures. Comparison of transcriptome profiles between control and NGF-stimulated hepatocytes demonstrated that NGF significantly up-regulated 10 genes and down-regulated 23 genes in hepatocytes. Subsequent KEGG pathway enrichment analysis indicated that NGF significantly affected the retinol metabolism pathway via increased retinol dehydrogenase 16 (RDH16) expression. In a mouse model of bile duct ligation-induced cholestatic liver injury, NGF supplementation significantly enhanced RDH16 expression, whereas administration of anti-NGF neutralizing antibodies prominently decreased RDH16 expression in cholestatic livers, supporting the positive role of NGF in the regulation of RDH16 in diseased livers. In vitro study further demonstrated that NGF triggered de novo synthesis of RDH16 in primary rat hepatocytes, mainly through an NF-κB signaling pathway. In conclusion, this study demonstrates the up-regulation of RDH16 by NGF in cultured rat hepatocytes and mouse cholestatic livers, and provides novel insights on the mechanistic role of NGF in the retinol metabolism of livers.

Free Fatty Acids Induce Autophagy and LOX-1 Upregulation in Cultured Aortic Vascular Smooth Muscle Cells.

Elevation of free fatty acids (FFAs) is known to affect microvascular function and contribute to obesity-associated insulin resistance, hypertension, and microangiopathy. Proliferative and synthetic vascular smooth muscle cells (VSMCs) increase intimal thickness and destabilize atheromatous plaques. This study aimed to investigate whether saturated palmitic acid (PA) and monounsaturated oleic acid (OA) modulate autophagy activity, cell proliferation, and vascular tissue remodeling in an aortic VSMC cell line. Exposure to PA and OA suppressed growth of VSMCs without apoptotic induction, but enhanced autophagy flux with elevation of Beclin-1, Atg5, and LC3I/II. Cotreatment with autophagy inhibitors potentiated the FFA-suppressed VSMC growth and showed differential actions of PA and OA in autophagy flux retardation. Both FFAs upregulated lectin-like oxidized low-density lipoprotein receptor 1 (LOX-1) but only OA increased LDL uptake by VSMCs. Mechanistically, FFAs induced hyperphosphorylation of Akt, ERK1/2, JNK1/2, and p38 MAPK. All pathways, except OA-activated PI3K/Akt cascade, were involved in the LOX-1 upregulation, whereas blockade of PI3K/Akt and MEK/ERK cascades ameliorated the FFA-induced growth suppression on VSMCs. Moreover, both FFAs exhibited tissue remodeling effect through increasing MMP-2 and MMP-9 expression and their gelatinolytic activities, whereas high-dose OA significantly suppressed collagen type I expression. Conversely, siRNA-mediated LOX-1 knockdown significantly attenuated the OA-induced tissue remodeling effects in VSMCs. In conclusion, OA and PA enhance autophagy flux, suppress aortic VSMC proliferation, and exhibit vascular remodeling effect, thereby leading to the loss of VSMCs and interstitial ECM in vascular walls and eventually the instability of atheromatous plaques. J. Cell. Biochem. 118: 1249-1261, 2017. © 2016 Wiley Periodicals, Inc.

High concentration of magnolol induces hepatotoxicity under serum-reduced conditions.

Although magnolol is cytoprotective against warm ischemia/reperfusion injury, its effect on cold preservation has not been fully investigated. This study aimed at examining whether magnolol maintains the liver graft integrity after cold preservation and elucidating the underlying mechanisms in terms of apoptotic signaling under both normothermic and hypothermic conditions. After being preserved in Ringer's lactate (RL) at 4 degrees C for 6h ex vivo, the magnolol-treated grafts demonstrated significantly higher AST, ALT, and LDH levels in perfusates than those from negative controls. TUNEL staining showed no difference in the number of apoptotic nuclei in both groups, whereas a more intense apoptotic signal in magnolol-treated grafts was shown as compared with the controls. In vitro data showed no significant difference in viability of RL-preserved clone-9 hepatocytes between the magnolol-treated and control groups, while magnolol pretreatment at 30min before cold preservation prominently induced hepatocyte cell death. RT-PCR and Western blotting analyses revealed a suppression in Bcl-2, but an up-regulation in Bax expression in clone-9 cells after magnolol treatment. Magnolol suppressed the ratios of NF-kappaB to I-kappaBalpha protein contents and I-kappaBalpha phosphorylation induced by TNF-alpha, and potentiated mitochondrial cytochrome c release and subsequent caspase-3 cleavage. Conversely, caspase-3 inhibitor attenuated magnolol-induced hepatotoxicity. We concluded that magnolol could not protect liver grafts from cold ischemia/reperfusion injury. High concentration of magnolol under serum-reduced conditions attenuates NF-kappaB-mediated signaling and induces intrinsic apoptotic pathway, thereby inducing in vitro hepatotoxicity.