Cinnamaldehyde (Cin), cinnamic acid (Ca) and cinnamyl alcohol (Cal), major constituents of Cinnamomum cassia, have been shown to possess antioxidant, anti-inflammatory, anticancer and other activities. In this study, our aim was to evaluate the antiproliferative activity of these compounds in human hepatoma Hep G2 cells and examine the effects of pifithrin-alpha (PFTα; a specific p53 inhibitor) on their apoptotic signaling transduction mechanism. The antiproliferative activity was measured by XTT assay. Expression of apoptosis-related proteins was detected by western blotting. Results showed that at a concentration of 30 μM, the order of antiproliferative activity in Hep G2 cells was Cin > Ca > Cal. Cin (IC(50) 9.76 ± 0.67 μM) demonstrated an antiproliferative potency as good as 5-fluorouracil (an anti-cancer drug; IC(50) 9.57 ± 0.61 μM). Further studies on apoptotic mechanisms of Cin showed that it downregulated the expression of Bcl-(XL), upregulated CD95 (APO-1), p53 and Bax proteins, as well as cleaving the poly (ADP-ribose) polymerase (PARP) in a time-dependent pattern. PFTα pre-incubation significantly diminished the effect of Cin-induced apoptosis. It markedly upregulated the anti-apoptotic (Bcl-(XL)) expression and downregulated the pro-apoptotic (Bax) expression, as well as effectively blocking the CD95 (APO-1) and p53 expression, and PARP cleavage in Cin-treated cells. This study indicates that Cin was the most potent antiproliferative constituent of C. cassia, and its apoptotic mechanism in Hep G2 cells could be mediated through the p53 induction and CD95 (APO-1) signaling pathways.

The combined cytoactive effects of American ginseng (Panax quinquefolius) and licorice (Glycyrrhiza uralensis) root extracts were investigated in a hepatocarcinoma cell line (Hep-G2). An isobolographic analysis was utilized to express the possibility of synergistic, additive or antagonistic interaction between the two extracts. Both ginseng and licorice roots are widely utilized in traditional Chinese medicine preparations to treat a variety of ailments. However, the effect of the herbs in combination is currently unknown in cultured Hep-G2 cells. Ginseng (GE) and licorice (LE) extracts were both able to reduce cell viability. The LC50 values, after 72 h, were found to be 0.64 ± 0.02 mg/mL (GE) and 0.53 ± 0.02 mg/mL (LE). An isobologram was plotted, which included five theoretical LC50s calculated, based on the fixed fraction method of combination ginseng to licorice extracts to establish a line of additivity. All combinations of GE to LE (1/5, 1/3, 1/2, 2/3, 4/5) produced an effect on Hep-G2 cell viability but they were all found to be antagonistic. The LC50 of fractions 1/3, 1/2, 2/3 were 23%, 21% and 18% above the theoretical LC50. Lactate dehydrogenase release indicated that as the proportion of GE to LE increased beyond 50%, the influence on membrane permeability increased. Cell-cycle analysis showed a slight but significant arrest at the G1 phase of cell cycle for LE. Both GE and LE reduced Hep-G2 viability independently; however, the combinations of both extracts were found to have an antagonistic effect on cell viability and increased cultured Hep-G2 survival.

Among the most common malignancies in humans, liver cancer ranks third in terms of mortality in the world. Seeking new anticancer drugs or adjuvant chemotherapy drugs from natural products has attracted the attention of many researchers. Lycium ruthenicum Murray (LR), a health food and traditional Chinese medicine, exerts extensive pharmacological properties, of which anthocyanins are one of the key active components. In this research, we explored the antitumor activity and autophagy regulation mechanism of anthocyanins from Lycium ruthenicum Murray (ALR) in HepG2 cells. Our results found that ALR profoundly reduced the cell viability, clone formation, migration, and invasion and promoted apoptosis and G2/M phase arrest of HepG2 cells in a dose-dependent pattern. Further studies confirmed that ALR treatment significantly increased the number of autophagic vacuoles and autophagosomes, upregulated the expression of Beclin-1, p62, LC3-II/LC3-I, and p-AMPK, and concomitantly downregulated the expression of p-mTOR. When autophagy was inhibited by 3-methyladenine (3-MA), ALR-induced proliferation inhibition, invasion, and migration capabilities, as well as apoptosis rate and G2/M phase arrest, were all reversed, and the activities of key proteins in the AMPK/mTOR pathway were all constrained. In summary, the results presented here indicate that ALR may be effective as a natural antitumor agent by activating AMPK and inhibiting the mTOR autophagy pathway in HepG2 cells.

'Ki-energy' (life-energy) is believed to increase the immune activity of its practitioners. It has also been shown to cause neuropsychological effects. We undertook this study to obtain objective and scientific evidence as to whether or not a 'Ki-effect' could inhibit the growth of cultured cancer cells. Cultured human liver carcinoma cells, HepG2, were used. A Japanese Ki-expert held his fingers toward the cells in culture dishes for 5 or 10 min. After culturing for 24 h, we measured cell numbers, protein concentration per cell, certain mRNA expressions and the synthesis of regucalcin. The results were compared with those for control cells (non-treated cells). We found that the number of cells in the Ki-exposed groups were less than those in the controls by 30.3 and 40.6% with 5 and 10 min Ki-exposure, respectively. The protein content per cell in the Ki-exposed groups (5 and 10 min) was higher than that in the control groups by 38.8 and 62.9%, respectively. These results were statistically significant. Using RT-PCR, we found that the mRNA expression for c-myc, a tumor stimulator gene, was decreased, while that for regucalcin, which suppresses DNA synthesis, was increased. Our molecular biological studies and mathematical model analysis demonstrated that Ki-energy inhibited cancer cell division. The data also indicate that the Ki-effects involve some form of infrared radiation from the human body. This study suggests the possibility that Ki-energy may be beneficial for cancer patients because it suppresses cancer cell growth, and at the same time, it stimulates immune functions of the patients.

Red raspberry contains a variety of bioactive ingredients and has high edible and medicinal value. Red raspberry extractions (RREs) have strong antioxidant capacity and anticancer ability in vivo and in vitro. This study was to explore the specific mechanism of RREs inhibiting the proliferation of liver cancer HepG2 cells and provide a theoretical basis for the prevention and treatment of liver cancer by RREs. HepG2 cells were cultured in vitro, and MTT assay was adopted to detect the effect of RREs on HepG2 cell activity. Colony formation assay was applied to detect the growth and proliferation of cells, cell apoptosis was detected by flow cytometry, and dichloro-dihydro-fluorescein diacetate (DCFH-DA) assay was adopted to detect the effect of RREs on the production of reactive oxygen species (ROS) in cells. The effect of RREs on cell mitochondrial membrane potential was evaluated by mitochondrial membrane potential assay kit with JC-1 (JC-1 assay), and western blot was used to detect the expression of apoptosis-related proteins (B-cell lymphoma-2 (Bcl-2), Bcl-2-associated x (Bax), and Caspase-3), thus investigating the effect of RREs on the molecular mechanism of HepG2 cell apoptosis. The results showed that RREs could inhibit the proliferation activity of HepG2 cells and promote their apoptosis in a concentration-dependent manner. The level of ROS in HepG2 cells interfered by RREs increased markedly, while the cell mitochondrial membrane potential decreased sharply. As the concentration of HepG2 increased, the mitochondrial membrane potential reduced steeply. Western blot results showed that the expression of apoptosis-related protein Bcl-2 in the RREs treatment group dropped, but the expression of Bax and Caspase-3 rose. In summary, RREs could inhibit the proliferation of liver cancer HepG2 cells and promote their apoptosis. This inhibition might be executed by inducing HepG2 cells to produce ROS, a decrease in Bcl-2/Bax protein ratio, and an obvious reduction in mitochondrial membrane potential.

Elemene, a compound found in an herb used in traditional Chinese medicine, has shown promising anticancer effects against a broad spectrum of tumors. In an in vivo experiment, we found that apatinib, a tyrosine kinase inhibitor that selectively inhibits VEGFR2, combined with elemene injection (Ele) for the treatment of H22 solid tumor in mice resulted in worse effectiveness than apatinib alone. Moreover, Ele could protect HepG2 cells from death induced by serum-free starvation. Further data on the mechanism study revealed that Ele induced protective autophagy and prevented human hepatoma cancer cells from undergoing apoptosis. Proapoptosis effect of Ele was enhanced when proautophagy effect was inhibited by hydroxychloroquine. Above all, Ele has the effect of protecting cancer cells from death either in apatinib induced nutrient deficient environment or in serum-free induced starvation. A combination of elemene injection with autophagy inhibitor might thus be a useful therapeutic option for hepatocellular carcinoma.

The combination of Artemisia iwayomogi and Curcuma longa radix is frequently prescribed for liver diseases in TKM. However, the synergic effects of the two herbs on nonalcoholic steatohepatitis (NASH) have not yet been studied. Therefore, we investigated the anti-NASH effects of the water extract of A. iwayomogi (AI), C. longa radix (CL), and combination of the two herbs (ACE). Hepatic steatosis and NASH were induced in HepG2 cells by treatment with palmitic acid (PA, for 6 h) with/without pretreatment of ACE (25 or 50 μg/mL), AI (50 or 100 μg/mL), CL (50 or 100 μg/mL), curcumin (5 μg/mL), or scopoletin (5 μg/mL). The PA treatment (200 μM) drastically altered intracellular triglyceride levels, total cholesterol, and expression levels of genes related to lipid metabolism (CD36, SREBP1c, PPAR-γ, and PPAR-α), whereas pretreatment with ACE significantly attenuated these alterations. ACE also protected HepG2 cells from PA- (300 μM-) induced endoplasmic reticulum (ER) stress and apoptosis and attenuated the related key molecules including GRP78, eIF2, and CHOP, respectively. In conclusion, we found synergic effects of A. iwayomogi and C. longa on NASH, supporting the clinical potential for fatty liver disorders. In addition, modulation of ER stress-relative molecules would be involved in its underlying mechanism.

Diosgenin, a naturally occurring steroid saponin found abundantly in legumes and yams, is a precursor of various synthetic steroidal drugs. Diosgenin is studied for the mechanism of its action in apoptotic pathway in human hepatocellular carcinoma cells. Based on DAPI staining, diosgenin-treated cells manifested nuclear shrinkage, condensation, and fragmentation. Treatment of HepG2 cells with 40 μM diosgenin resulted in activation of the caspase-3, -8, -9 and cleavage of poly-ADP-ribose polymerase (PARP) and the release of cytochrome c. In the upstream, diosgenin increased the expression of Bax, decreased the expression of Bid and Bcl-2, and augmented the Bax/Bcl-2 ratio. Diosgenin-induced, dose-dependent induction of apoptosis was accompanied by sustained phosphorylation of JNK, p38 MAPK and apoptosis signal-regulating kinase (ASK)-1, as well as generation of the ROS. NAC administration, a scavenger of ROS, reversed diosgene-induced cell death. These results suggest that diosgenin-induced apoptosis in HepG2 cells through Bcl-2 protein family-mediated mitochndria/caspase-3-dependent pathway. Also, diosgenin strongly generated ROS and this oxidative stress might induce apoptosis through activation of ASK1, which are critical upstream signals for JNK/p38 MAPK activation in HepG2 cancer cells.

Hepatocellular carcinoma (HCC) is currently one of the most common tumors, with a high morbidity and mortality rate. HCC induced by persistent hepatitis B virus (HBV) infection is the most common liver cancer subtype at present, and HBV-related HCC is highly malignant and its development mechanism still needs to be explored in depth. This study aimed to explore the molecular mechanism of hsa_circ_0000847 targeting miR-135a-5p (miR-135a) to regulate the proliferation, invasion, and apoptosis of liver cancer cells. The study found that the expression level of hsa_circ_0000847 in liver cancer tissues and cells was significantly increased, while the expression level of miR-135a was significantly decreased. Hsa_circ_0000847 promoted the proliferation of liver cancer cells and elevated the expression of the proliferation-related protein. In addition, hsa_circ_0000847 could promote the invasion of HBV-infected liver cancer cells and inhibit the cell apoptosis of liver cancer cells. At the same time, it significantly promoted the expression of antiapoptotic proteins and inhibited the expression of proapoptotic protein. Interestingly, the dual luciferase experiment proved that hsa_circ_0000847 directly targeted miR-135a. On the other hand, the combined effect of hsa_circ_0000847 and miR-135a further illustrated the effect of hsa_circ_0000847 on the proliferation, invasion, and apoptosis of liver cancer cells. In addition, further experiments have also found that HBV could promote the expression of p-p38, p-ERK, and p-JNK through the hsa_circ_0000847/miR-135a axis, thereby further activating the MAPK pathway. In short, HBV promotes the proliferation and invasion of liver cancer cells and inhibits apoptosis by regulating the hsa_circ_0000847/miR-135a pathway, which provided a theoretical basis for effective treatment of HBV-infected liver cancers.

Resistance of cancer cells to chemotherapy is controlled by the decrease of intracellular drug accumulation, increase of detoxification, and diminished propensity of cancer cells to undergo apoptosis. ATP-binding cassette (ABC) membrane transporters with intracellular metabolic enzymes contribute to the complex and unresolved phenomenon of multidrug resistance (MDR). Natural products as alternative medicine have great potential to discover new MDR inhibitors with diverse modes of action. In this study, we characterized several extracts of traditional Chinese medicine (TCM) plants (N = 16) for their interaction with ABC transporters, cytochrome P3A4 (CYP3A4), and glutathione-S-transferase (GST) activities and their cytotoxic effect on different cancer cell lines. Fallopia japonica (FJ) (Polygonaceae) shows potent inhibitory effect on CYP3A4 P-glycoprotein activity about 1.8-fold when compared to verapamil as positive control. FJ shows significant inhibitory effect (39.81%) compared with the known inhibitor ketoconazole and 100 μg/mL inhibited GST activity to 14 μmol/min/mL. FJ shows moderate cytotoxicity in human Caco-2, HepG-2, and HeLa cell lines; IC50 values were 630.98, 198.80, and 317.37 µg/mL, respectively. LC-ESI-MS were used to identify and quantify the most abundant compounds, emodin, polydatin, and resveratrol, in the most active extract of FJ. Here, we present the prospect of using Fallopia japonica as natural products to modulate the function of ABC drug transporters. We are conducting future study to evaluate the ability of the major active secondary metabolites of Fallopia japonica to modulate MDR and their impact in case of failure of chemotherapy.

In the past few decades, the incidence of liver cancer has been rapidly rising across the world. Rosemary is known to possess antioxidant activity and is used as natural antioxidant food preservative. It is proposed to have anticancer activity in treating different tumor models. In this study, we try to explore the impact of rosemary extracts on upregulating the level of Nrf2 and Nrf2-regulatory proteins, Sestrin2 and MRP2 in HepG2 cells, and to speculate its potential mechanism. The anticancer activity of rosemary extract, including its polyphenolic diterpenes carnosic acid and carnosol, was evaluated to understand the potential effect on HepG2 cells. Rosemary extract, carnosic acid, and carnosol induced the expression of Sestrin2 and MRP2 associate with enhancement of Nrf2 protein level in HepG2 cells, in which carnosic acid showed most obvious effect. Although the activation pathway of Nrf2/ARE was not exactly assessed, it can be assumed that the enhancement of expression of Sestrin2 and MRP2 may result from upregulation of Nrf2.

The global cancer burden remains a serious concern with the alarming incidence of one in eight men and one in eleven women dying in developing countries. This situation is aggravated by the multidrug resistance (MDR) of cancer cells that hampers chemotherapy. In this study, the cytotoxicity of the methanol extract (HRB), fractions (HRBa, HRBb, and HRBa1-5), and compounds from the bark of Hypericum roeperianum (HRB) was evaluated towards a panel of 9 cancer cell lines. The mode of action of the HRB and trichadonic acid (1) was also studied. Column chromatography was applied to isolate the constituents of HRB. The cytotoxicity of botanicals and phytochemicals was evaluated by the resazurin reduction assay (RRA). Caspase-Glo assay was used to evaluate the activity of caspases, and reactive oxygen species (ROS) (H2DCFH-DA) were assessed by flow cytometry. Phytochemicals isolated from HRB were trichadonic acid (1), fridelan-3-one (2), 2-hydroxy-5-methoxyxanthone (3), norathyriol (4), 1,3,5,6-tetrahydroxyxanthone (5), betulinic acid (6), 3'-hydroxymethyl-2'-(4″-hydroxy-3″,5″-dimethoxyphenyl)-5',6':5,6-(6,8-dihydroxyxanthone)-1',4'-dioxane (7), and 3'-hydroxymethyl-2'-(4″-hydroxy-3″,5″-dimethoxyphenyl)-5',6':5,6-(xanthone)-1',4'-dioxane (8). Botanicals HRB, HRBa, HRBa2-4, HRBb, and doxorubicin displayed cytotoxic effects towards the 9 tested cancer cell lines. The recorded IC50 values ranged from 11.43 µg/mL (against the P-glycoprotein (gp)-overexpressing CEM/ADR5000 leukemia cells) to 26.75 µg/mL (against HCT116 (p53+/+) colon adenocarcinoma cells) for the crude extract HRB. Compounds 1, 5, and doxorubicin displayed cytotoxic effects towards the 9 tested cancer cell lines with IC50 values varying from 14.44 µM (against CCRF-CEM leukemia cells) to 44.20 µM (against the resistant HCT116 (p53-/-) cells) for 1 and from 38.46 µM (against CEM/ADR5000 cells) to 112.27 µM (against the resistant HCT116 (p53-/-) cells) for 5. HRB and compound 1 induced apoptosis in CCRF-CEM cells. The apoptotic process was mediated by enhanced ROS production for HRB or via caspases activation and enhanced ROS production for compound 1. This study demonstrated that Hypericum roeperianum is a potential source of cytotoxic phytochemicals such as trichadonic acid and could be further exploited in cancer chemotherapy.

Eucommia ulmoides Oliv. (E. ulmoides Oliv.) and moso bamboo (Phyllostachys pubescens) leaves are used as folk medicines in central-western China to treat diabetes. To investigate the hypoglycemic activity of the effervescent granules prepared using E. ulmoides Oliv. and moso bamboo leaves (EBEG) in HepG2 cells, EBEG were prepared with 5% of each of polysaccharides and chlorogenic acids from moso bamboo and E. ulmoides Oliv. leaves, respectively. HepG2 cells cultured in a high-glucose medium were classified into different groups. The results displayed EBEG-treated cells showed better glucose utilization than the negative controls; thus, the hypoglycemic effect of EBEG was much greater than that of granules prepared using either component alone, thereby indicating that this effect was due to a synergistic action of the components. Further, glucose consumption levels in the cells treated with EBEG (156.35% at 200 μg/mL) and the positive controls (metformin, 162.29%; insulin, 161.52%) were similar. Thus, EBEG exhibited good potential for use as a natural antidiabetic agent. The hypoglycemic effect of EBEG could be due to the synergistic action of polysaccharides from the moso bamboo leaves and chlorogenic acids from E. ulmoides Oliv. leaves via the inhibition of alpha-glucosidase and glucose-6-phosphate displacement enzyme.

Dianthus chinensis L. is used to treat various diseases including cancer; however, the molecular mechanism by which the ethanol extract of Dianthus chinensis L. (EDCL) induces apoptosis is unknown. In this study, the apoptotic effects of EDCL were investigated in human HepG2 hepatocellular carcinoma cells. Treatment with EDCL significantly inhibited cell growth in a concentration- and time-dependent manner by inducing apoptosis. This induction was associated with chromatin condensation, activation of caspases, and cleavage of poly (ADP-ribose) polymerase protein. However, apoptosis induced by EDCL was attenuated by caspase inhibitor, indicating an important role for caspases in EDCL responses. Furthermore, EDCL did not alter the expression of bax in HepG2 cells but did selectively downregulate the expression of bcl-2 and bcl-xl, resulting in an increase in the ratio of bax:bcl-2 and bax:bcl-xl. These results support a mechanism whereby EDCL induces apoptosis through the mitochondrial pathway and caspase activation in HepG2 cells.

Wu-Mei-Wan (WMW) is a Chinese herbal formula used to treat type 2 diabetes. In this study, we aimed to explore the effects and mechanisms of WMW on insulin resistance in HepG2 cells. HepG2 cells were pretreated with palmitate (0.25 mM) to impair the insulin signaling pathway. Then, they were treated with different doses of WMW-containing medicated serum and stimulated with 100 nM insulin. Results showed that palmitate could reduce the glucose consumption rate in HepG2 cells and impair insulin signaling related to phosphorylation of insulin receptor (IR) and insulin receptor substrate-1 (IRS-1), thereby regulating the downstream signaling pathways. However, medicated serum of WMW restored impaired insulin signaling, upregulated the expression of phospho-IR (pIR), phosphatidylinositol 3-kinase p85 subunit, phosphoprotein kinase B, and glucose transporter 4, and decreased IRS serine phosphorylation. In addition, it decreased the expression of interleukin-1β and tumor necrosis factor-α, which are the key proinflammatory cytokines involved in insulin resistance; besides, it reduced the expression of NLRP3 inflammasome. These results suggested that WMW could alleviate palmitate-induced insulin resistance in HepG2 cells via inhibition of NLRP3 inflammasome and reduction of proinflammatory cytokine production.

Hebanthe paniculata roots (formerly Pfaffia paniculata and popularly known as Brazilian ginseng) show antineoplastic, chemopreventive, and antiproliferative properties. Functional properties of these roots and their extracts are usually attributed to the pfaffosidic fraction, which is composed mainly by pfaffosides A-F. However, the therapeutic potential of this fraction in cancer cells is not yet entirely understood. This study aimed to analyze the antitumoral effects of the purified pfaffosidic fraction or saponinic fraction on the human hepatocellular carcinoma HepG2 cell line. Cellular viability, proliferation, and apoptosis were evaluated, respectively, by MTT assay, BrdU incorporation, activated caspase-3 immunocytochemistry, and DNA fragmentation assay. Cell cycle was analyzed by flow cytometry and the cell cycle-related proteins were analyzed by quantitative PCR and Western blot. The cells exposed to pfaffosidic fraction had reduced viability and cellular growth, induced G2/M at 48 h or S at 72 h arrest, and increased sub-G1 cell population via cyclin E downregulation, p27(KIP1) overexpression, and caspase-3-induced apoptosis, without affecting the DNA integrity. Antitumoral effects of pfaffosidic fraction from H. paniculata in HepG2 cells originated by multimechanisms of action might be associated with cell cycle arrest in the S phase, by CDK2 and cyclin E downregulation and p27(KIP1) overexpression, besides induction of apoptosis through caspase-3 activation.

Glossostemon bruguieri (moghat) is used as a nutritive and demulcent drink. This study was performed to investigate the antiproliferative effects of moghat root extract (MRE) and its apoptotic mechanism in hepatocellular carcinoma (HCC) cells, HepG2 and Hep3B. MTT assay, morphological changes, apoptosis enzyme linked immunosorbent assay, caspase and apoptotic activation, flow cytometry, and immunoblot analysis were employed. The IC50 of MRE for HepG2 (910 ± 6 μg/ml) and for Hep3B (1510 ± 5 μg/ml) induced significant growth-inhibitory effects against HCC cells, with no cytotoxic effect on normal hepatocytes. MRE treatment induced apoptotic effects to HepG2 cells in a caspase-dependent manner and via upregulating p53/p21 and PCNA. The upregulation of p21 was controlled by p53 expression in HepG2 but not in Hep3B despite upregulation of Bax protein in both cell lines. Interestingly, p21 may be a remarkable switch to G1 arrest in HepG2 cells, but not in Hep3B cells. In addition, Fas- and mitochondria-mediated pathways were found to be involved in MRE-induced apoptosis in Hep3B cells. The GC-MS analysis of MRE revealed two major constituents of pharmaceutical importance: the flavonoid apigenin (17.04%) and the terpenoid squalene (11.32%). The data presented in this paper introduces G. bruguieri as a promising nontoxic herb with therapeutic potential for HCC. To the authors' knowledge, the present study provides the first report on the anticancer activity of MRE on HCC cells.

Hepatitis B virus (HBV) infection accounts for over a half of cases of hepatocellular carcinoma (HCC), the most frequent malignant tumor of the liver. HBV-encoded X (HBx) plays critical roles in HBV-associated hepatocarcinogenesis. However, it is unclear whether and how HBx regulates the expression of epidermal growth factor receptor (EGFR), an important gene for cell growth. Therefore, the study aimed to investigate the association between HBx and EGFR expression. In this study, we found that HBx upregulates miR-7 expression to target 3'UTR of EGFR mRNA, which in turn results in the reduction of EGFR protein expression in HCC cells. HBx-mediated EGFR suppression renders HCC cells a slow-growth behavior. Deprivation of HBx or miR-7 expression or restoration of EGFR expression can increase the growth rate of HCC cells. Our data showed the miR-7-dependent EGFR suppression by HBx, supporting an inhibitory role of HBx in the cell growth of HCC. These findings not only identify miR-7 as a novel regulatory target of HBx, but also suggest HBx-miR-7-EGFR as a critical signaling in controlling the growth rate of HCC cells.

Hepatocellular carcinoma (HCC) seriously endangers humans. In traditional Chinese medicine, Marsdenia tenacissima (MTE) has anti-inflammatory, antiasthmatic, antihypertensive, and anticancer effects. This study reveals the antiproliferative effect of MTE on the HCC cells in vitro and provides a theoretical basis for the development and clinical application of anti-HCC agents. Methods. MHCC-97H and HepG2 cells were cultured in vitro and exposed to various concentrations and durations of MTE, and an MTT assay was used to detect the effects of MTE on cell proliferation. Transmission electron microscopy revealed the morphological changes in the two cell lines after MTE stimulation. The MTE effects on the apoptosis and cell cycle distribution of the cell lines were detected by flow cytometry. Western blotting and qRT-PCR were used to detect target gene expression at the protein and mRNA levels, respectively. Results. MTE reduced the viability of the MHCC-97H and HepG2 cells in a dose- and time-dependent manners (P < 0.05). Autophagic vesicles and apoptotic bodies were found in the MHCC-97H and HepG2 cells after MTE incubation, and the Annexin V-PI assay showed that the apoptotic rates of the cell lines increased with increasing MTE concentration (P < 0.05). Autophagy inducer rapamycin promoted the MTE-induced apoptotic rates of the cell lines, whereas autophagy inhibitor chloroquine inhibited the apoptotic rates. More cells in the S phase were found in the two cell lines after MTE treatment (P < 0.05). After MTE incubation, MIF, CD47, and beclin-1 protein levels significantly increased. Furthermore, in the MTE group, Akt, mTOR, and caspase3 expressions decreased; however, LC 3 expression increased, which was significantly different from the control group (P < 0.05). Conclusions. MTE inhibited proliferation and induced autophagy, apoptosis, and S phase cell cycle arrest in the MHCC-97H and HepG2 cells. These effects might be related to the activation of MIF and mTOR signaling inhibition.

The present study was conducted to investigate the effect of Sagunja-tang on the lipid related disease in a rat model of menopausal hyperlipidemia and lipid accumulation in methyl-β-cyclodextrin-induced HepG2 cells. In in vivo study using menopausal hyperlipidemia rats, Sagunja-tang reduced retroperitoneal and perirenal fat, serum lipids, atherogenic index, cardiac risk factor, media thickness, and nonalcoholic steatohepatitis score, when compared to menopausal hyperlipidemia control rats. In HepG2 cells, Sagunja-tang significantly decreased the lipid accumulation, total cholesterol levels, and low-density/very-low-density lipoprotein levels. Moreover, Sagunja-tang reversed the methyl-β-cyclodextrin-induced decrease in the protein levels of critical molecule involved in cholesterol synthesis, sterol regulatory element binding protein-2, and low-density lipoprotein receptor and inhibited protein levels of 3-hydroxy-3-methylglutaryl coenzyme A reductase as well as activity. Phosphorylation level of AMP-activated protein kinase was stimulated by Sagunja-tang. These results suggest that Sagunja-tang has effect on inhibiting hepatic lipid accumulation through regulation of cholesterol synthesis and AMPK activity in vitro. These observations support the idea that Sagunja-tang is bioavailable both in vivo and in vitro and could be developed as a preventive and therapeutic agent of hyperlipidemia in postmenopausal females.