Human Embryonic Liver Research Articles

Evaluation of chemical constituents and in vitro antimicrobial, antioxidant and cytotoxicity potential of rhizome of Astilbe rivularis (Bodho-okhati), an indigenous medicinal plant from Eastern Himalayan region of India

BackgroundAstilbe rivularisL. is an indigenous medicinal plant growing in high altitude of Darjeeling Himalayan region of India and Nepal. The plant rhizome has been used traditionally as medicine by local tribes to treat various ailments including infectious and other diseases. The present study aims to evaluate the plant rhizome for chemical composition and in vitro antioxidant, antibacterial and cytotoxic bioactivities.MethodsThe methanolic extract of rhizome was analyzed for phytochemical constituents by biochemical and GC-MS methods. The antibacterial property of the extract was monitored by agar well diffusion assay. Antioxidant potential was assessed by in vitro DPPH and ABTS scavenging assays and scavenging of induced ROS in normal cell line using fluorescent probe 2′, 7′- dichlorofluorescin diacetate. Cytotoxic effect of the extract in cancer and normal cell lines was determined by MTT assay.ResultsRhizome methanolic extract contained terpenoids, flavonoids, tannins, phenols, alkaloids, saponins and reducing sugars. Further analysis of extract by GC-MS showed the presence of nine major constituents belonging to terpenoids and fatty acid groups. The extract had marked in vitro ROS scavenging activity and moderate antibacterial activity against gram positive and gram negative bacteria. It showed cytotoxicity to neuroblastoma (SHSY5Y) cell line with IC50 value < 100 μg ml− 1 but had least damaging effect on normal cells, like human embryonic kidney (HEK-293) and liver (WRL-68) cell lines.ConclusionThe study suggests that Astilbe rivularis has potential as source of new potent antibacterial, antioxidant and anticancer agents. Further studies on purification and characterization of active compounds from Astilbe rivularis and their biological evaluation are highly recommended.

Enhancement of Alcohol Metabolism by Ginseng Berry Extract and its Mixed Herbal Beverages: In vitro and in vivo Experiments

To investigate the enhancement of alcohol metabolism by two ginseng berry (GB) extracts and their two types of mixed herbal beverages through in vitro and in vivo experiments. Two GB extract solutions and their two herbal beverages were evaluated as enhancers of alcohol metabolism in normal human embryonic liver cells (CL-48 cell line) through assays of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) activity. Cytotoxicity was also assessed in the same cell line using an MTT assay. Effects on alcohol metabolism were also observed in vivo through measurement of serum alcohol, acetaldehyde, alanine aminotransferase (ALT), and aspartate aminotransferase (AST) levels in alcohol treated rats. Blood samples were serially collected at 1, 2, 3, 4, 6 and 8 hrs after a single ethanol (EtOH) treatment. A single treatment with the test samples was administered orally 30 minutes after EtOH treatment. GB extract solutions effectively elevated the ADH and ALDH activity observed in vitro, while no treatment- related cytotoxic effects were found with test samples t concentrations up to 100 mg/mL. Significantly lower (p<0.01 or p<0.05) serum alcohol and acetaldehyde content was observed in samples from treated rats than in those from control rats (EtOH only) 1 or 2 hrs after EtOH treatment. In addition, noticeable decreases were observed in serum AST and ALT levels in treated samples 8 hrs after EtOH administration. HM40, an herbal mixture containing GB extract (40 mg/75 mL of ginsenoside Re), showed betters enhancement of alcohol metabolism through ADH/ALDH activation, as well as related hepatoprotective effects. GB extracts effectively enhanced alcohol metabolism without cytotoxicity while also providing possible hepatoprotective effects that could serve as a functional ingredient in anti-hangover alternative therapies. These extracts are expected to be more effective when made into herbal mixture beverages.

Potentiation of β-lactam antibiotics and β-lactam/β-lactamase inhibitor combinations against MDR and XDR Pseudomonas aeruginosa using non-ribosomal tobramycin-cyclam conjugates.

To develop a multifunctional adjuvant molecule that can rescue β-lactam antibiotics and β-lactam/β-lactamase inhibitor combinations from resistance in carbapenem-resistant Pseudomonas aeruginosa clinical isolates. Preparation of adjuvant was guided by structure-activity relationships, following standard protocols. Susceptibility and chequerboard studies were assessed using serial 2-fold dilution assays. Toxicity was evaluated against porcine erythrocytes, human embryonic kidney (HEK293) cells and liver carcinoma (HepG2) cells via MTS assay. Preliminary in vivo efficacy was evaluated using a Galleria mellonella infection model. Conjugation of tobramycin and cyclam abrogates the ribosomal effects of tobramycin but confers a potent adjuvant property that restores full antibiotic activity of meropenem and aztreonam against carbapenem-resistant P. aeruginosa. Therapeutic levels of susceptibility, as determined by CLSI susceptibility breakpoints, were attained in several MDR clinical isolates, and time-kill assays revealed a synergistic dose-dependent pharmacodynamic relationship. A triple combination of the adjuvant with ceftazidime/avibactam (approved), aztreonam/avibactam (Phase III) and meropenem/avibactam enhances the efficacies of β-lactam/β-lactamase inhibitors against recalcitrant strains, suggesting rapid access of the combination to their periplasmic targets. The newly developed adjuvants, and their combinations, were non-haemolytic and non-cytotoxic, and preliminary in vivo evaluation in G. mellonella suggests therapeutic potential for the double and triple combinations. Non-ribosomal tobramycin-cyclam conjugate mitigates the effect of OprD/OprF porin loss in P. aeruginosa and potentiates β-lactam/β-lactamase inhibitors against carbapenem-resistant clinical isolates, highlighting the complexity of resistance to β-lactam antibiotics. Our strategy presents an avenue to further preserve the therapeutic utility of β-lactam antibiotics.

Establishment of a human embryonic stem cell-based liver differentiation model for hepatotoxicity evaluations

The liver is one of the major targets of hormones, including thyroid hormones (THs), and many industrial chemicals, such as endocrine-disrupting chemicals. Those compounds may permeate the placenta barrier and pose a risk for embryonic development. Therefore, it is necessary to assess the toxic effects of those kind of industrial chemicals during liver development. In this study, to mimic liver specification in vitro, we differentiated human embryonic stem cells (ESCs) into functional hepatocyte-like cells. We performed this differentiation process in presence of two THs, triiodothyronine (T3) and thyroxine (T4), with the purpose of identifying biomarkers for toxicity screening. TH exposure (3, 30 and 300 nM) yielded to hepatocytes with impaired glycogen storage ability and abnormal lipid droplets’ accumulation. Global gene expression analysis by RNA-seq identified a number of genes responsible for hepatic differentiation and function which were affected by 30 nM T3 and T4. Those differentially expressed genes were used to assess the potential developmental liver toxicity of two famous environmental pollutants, 2, 2, 4, 4-tetrabromodiphenyl ether (BDE-47) and decabromodiphenyl ether (BDE-209), at 10 nM to 1 μM treatments. Our findings demonstrate that BDE-47 and BDE-209, dysregulated pathways such as “chemical carcinogenesis”, “steroid hormone biosynthesis” and “drug metabolism-cytochrome P450”. Moreover, we were able to identify a set of 17 biomarkers, very useful to predict the potential developmental hepatotoxicity of industrial chemicals.

VE-Cadherin and ACE Co-Expression Marks Highly Proliferative Hematopoietic Stem Cells in Human Embryonic Liver.

Despite advances to engineer transplantable hematopoietic stem and progenitor cells (HSPCs) for research and therapy, an in-depth characterization of the developing human hematopoietic system is still lacking. The human embryonic liver is at the crossroad of several hematopoietic sites and harbors a complex hematopoietic hierarchy, including the first actively dividing HSPCs that will further seed the definitive hematopoietic organs. However, few are known about the phenotypic and functional HSPC organization operating at these stages of development. In this study, using a combination of four endothelial and hematopoietic surface markers, that is, the endothelial-specific marker vascular endothelial-cadherin (Cdh5, CD144), the pan-leukocyte antigen CD45, the hemato-endothelial marker CD34, and the angiotensin-converting enzyme (ACE, CD143), we identified distinct HSPC subsets, and among them, a population co-expressing the four markers that uniquely harbored an outstanding proliferation potential both ex vivo and in vivo. Moreover, we traced back this population to the yolk sac (YS) and aorta-gonad-mesonephros (AGM) sites of hematopoietic emergence. Taken together, our data will help to identify human HSPC self-renewal and amplification mechanisms for future cell therapies.

Numerical areal density of CD34 and CD117 immunoreactive hematopoietic cells in human fetal and embryonic liver

&amp;lt;p&amp;gt;Introduction. Hematopoiesis is the process of developing blood cells and the formed elements of blood. This dynamic process occurs postnatally in the bone marrow, while the prenatal hematopoiesis is scattered over multiple localizations. It begins in the yolk sac, in the intra-embryonic islands of hematopoiesis, and continues in the liver, spleen, and bone marrow, as well as, according to new research, in the placenta. The liver, as the largest hematopoietic organ of intrauterine development, is the source of hematopoietic stem cells. The aim of the study was to identify and quantify hematopoietic cells in human fetal and embryonic liver showing the immunoreactivity of CD34 and CD117 molecules. Methods. 5 human embryonic livers and 25 human fetal livers, 7 to 38 weeks of gestational age, were used in the study. Liver samples were routinely processed to paraffin molds. Tissue sections, 5 &amp;amp;micro;m thick, were stained with hematoxylin and eosin as immunohistochemical detection of CD34 and CD117 antigens. Quantification was performed morphometrically determining numerical areal density. The Chi-square test was used in the statistical analysis. Results. The numerical areal density of CD34 immunoreactive hematopoietic cells in the liver during the embryonic development was 0.35%: 0.30% in the first trimester (including both the embryonic and fetal period), 0.46% in the second trimester and 0.10% in the third trimester. The numerical areal densities of CD117 immunoreactive hematopoietic cells in the liver during the embryonic period were 0.28%: 0.50% in the first trimester (including both the embryonic and fetal periods), 0.11% in the second trimester and 0.09% in the third trimester. Conclusion. Fetal and embryonic liver represents a significant source of CD34 and CD117 immunoreactive hematopoietic cells.&amp;lt;/p&amp;gt;

Bivalent Ligand UDCA-LPE Inhibits Pro-Fibrogenic Integrin Signalling by Inducing Lipid Raft-Mediated Internalization.

Ursodeoxycholyl lysophosphatidylethanolamide (UDCA-LPE) is a synthetic bile acid-phospholipid conjugate with profound hepatoprotective and anti-fibrogenic functions in vitro and in vivo. Herein, we aimed to demonstrate the inhibitory effects of UDCA-LPE on pro-fibrogenic integrin signalling. UDCA-LPE treatment of human embryonic liver cell line CL48 and primary human hepatic stellate cells induced a non-classical internalization of integrin β1 resulting in dephosphorylation and inhibition of SRC and focal adhesion kinase (FAK). Signalling analyses suggested that UDCA-LPE may act as a heterobivalent ligand for integrins and lysophospholipid receptor1 (LPAR1) and co-immunoprecipitation demonstrated the bridging effect of UDCA-LPE on integrin β1 and LPAR1. The disruption of either the UDCA-moiety binding to integrins by RGD-containing peptide GRGDSP or the LPE-moiety binding to LPAR1 by LPAR1 antagonist Ki16425 reversed inhibitory functions of UDCA-LPE. The lack of inhibitory functions of UDCA-PE and UDCA-LPE derivatives (14:0 and 12:0, LPE-moiety containing shorter fatty acid chain) as well as the consistency of the translocation of UDCA-LPE and integrins, which co-fractionated with LPE but not UDCA, suggested that the observed UDCA-LPE-induced translocation of integrins was mediated by LPE endocytic transport pathway.

The cytotoxicity of water-soluble carbon nanotubes on human embryonic kidney and liver cancer cells

The cytotoxicity of water-soluble carbon nanotubes on human embryonic kidney and liver cancer cells

The regulation effect of emodin on human embryonic hepatocyte L02 cell strain Farnesoid X receptor

Objective To investigate the regulation effect of emodin on human embryonic liver L02 cells strain farnesoid X receptor(FXR) pathways. Methods By using Guggulsterones, FXR genes were intervened with in L02 cells as model group, in three different concentrations of emodin (50.0 μmol/L, 25.0 μmol/L, 12.5 μmol/L) of emodin in the model group cells, FXR, small heterodimer parter(SHP), UDP-glucuronosyltransferase 2B4(UGT2B4), bile salt export pump(BSEP) mRNA and protein expressions were detected by real-time fluorescent quantitative PCR and Western blot test. Results (1)The relative expressions of FXR mRNA and protein in the model group (0.240±0.021, 0.385±0.119) decreased significantly than those of control group (1.000±0.088, 1.000±0.223), the differences were statistically significant (t=14.62, 4.21, all P<0.01). Compared with the model group, the relative expressions of FXR mRNA in the high, the middle and the low-dose emodin groups (0.755±0.083, 0.817±0.097, 0.547±0.080) were significantly higher (t=10.42, 10.03, 6.39, all P<0.01). The relative expressions of FXR protein in the medium-dose group (0.865±0.203) increased significantly (t=3.53, P<0.01). The relative expressions of FXR mRNA in the high and the medium-dose groups (0.755±0.083, 0.817±0.097) were higher than those in the low-dose group (0.547±0.080), the differences were statistically significant (t=3.11, 3.70, all P<0.01). (2)The relative expressions of SHP, UGT2B4, BSEP mRNA and protein in the model group (0.148±0.025, 0.205±0.039, 0.184±0.020; 0.458±0.130, 0.255±0.170, 0.303±0.100) were significantly lower than those in the control group (1.000±0.099, 1.000±0.104, 1.000±0.125; 1.000±0.129, 1.000±0.157, 1.000±0.162), the differences were statistically significant (t=14.50, 12.44, 11.19, 5.13, 5.57, 6.33, all P<0.01). The relative expressions of SHP, UGT2B4 and BSEP mRNA in the high, the middle and the low-dose groups (0.610±0.058, 0.514±0.041, 0.707±0.062; 0.755±0.108, 0.800±0.086, 0.727±0.076; 0.470±0.070, 0.582±0.050, 0.500±0.108) were significantly lower than those in the model group (0.148±0.025, 0.205±0.039, 0.184±0.020), the differences were statistically significant (t=12.75, 9.38, 13.94, 9.46, 10.90, 11.96, 7.53, 10.31, 5.00, all P<0.01). The relative expressions of SHP, UGT2B4 and BSEP mRNA in the high and the middle-dose emodin group (0.658±0.091, 0.624±0.113, 0.607±0.097; 0.868±0.194, 0.883±0.099, 0.913±0.131) were significantly higher than those in the low-dose group (0.458±0.130, 0.255±0.170, 0.303±0.100), the differences were statistically significant (t=2.18, 3.13, 3.78, 3.05, 5.53, 6.41, all P<0.01). The relative expression of SHP and BSEP protein in the low-dose group (0.645±0.135, 0.572±0.076) increased, the differences were statistically significant (t=1.73, P<0.05, t=3.72, P<0.01). The relative expression of BSEP protein in the high and the medium-dose groups (0.607±0.097, 0.913±0.131) was significantly higher than those in the low-dose group (0.572±0.076), the differences were statistically significant (t=1.99, 3.90, all P<0.01). The relative expressions of SHP and UGT2B4 mRNA in the high and the low-dose group (0.610±0.058, 0.470±0.070; 0.514±0.041, 0.582±0.050) were significantly lower than those in the medium-dose group (0.800±0.086), the differences were statistically significant (t=3.75, 6.47, 3.83, 3.42, all P<0.01). The expression levels of UGT2B4 and BSEP in the high and the low-dose groups(0.624±0.113, 0.644±0.097; 0.607±0.097, 0.572±0.076) were significantly lower than those in the medium-dose group(0.883±0.099, 0.913±0.131), the differences were statistically significant (t=4.27, 2.98, 6.30, 3.90, all P<0.01). Conclusions Guggulsterones can inhibit FXR and downstream genes SHP, UGT2B4, BSEP expressions in L02, and emodin can enhance FXR gene expression, promote SHP, UGT2B4, BSEP gene expression, inhibit cholestasis pathway, protection of liver cell, which shows a dosage discreapancy. Key words: Youth Fund of Emodin; Human embryonic liver cells L02; Farnesoid X receptor

The cytotoxicity of water-soluble carbon nanotubes on human embryonic kidney and liver cancer cells

The cytotoxicity of polyethylene glycol-modified carbon nanotubes (PEG-CNTs) on human embryonic kidney cells (293T) and hepatoma cells (HepG2) was studied in vitro. The effect of the concentration of PEG-CNTs on the cell activity and survival rate was detected by a colorimetric assay method after exposure for 24, 48 and 72 h. The dead HepG2 cells obtained by incubation in solutions with different concentrations of PEG-CNTs for 24 h can be stained by a dye, propidium iodide, and the cell mortality was determined by flow cytometry. Results show that the toxicity of PEG-CNTs with a good water solubility towards 293T and HepG2 cells increases with increasing PEG-CNT concentration. When the concentration of the PEG-CNTs is less than 100 μg/mL, the toxicity of the PEG-CNTs is Grade I (non-toxic) and when it is greater than 100 μg/mL but less than 200 μg/mL it is Grade II (mildly toxic), according to the ISO2109932-5 cytotoxicity standard. The toxicity grade does not change with increased time.

Human liver segments: role of cryptic liver lobes and vascular physiology in the development of liver veins and left-right asymmetry

Couinaud based his well-known subdivision of the liver into (surgical) segments on the branching order of portal veins and the location of hepatic veins. However, both segment boundaries and number remain controversial due to an incomplete understanding of the role of liver lobes and vascular physiology on hepatic venous development. Human embryonic livers (5–10 weeks of development) were visualized with Amira 3D-reconstruction and Cinema 4D-remodeling software. Starting at 5 weeks, the portal and umbilical veins sprouted portal-vein branches that, at 6.5 weeks, had been pruned to 3 main branches in the right hemi-liver, whereas all (>10) persisted in the left hemi-liver. The asymmetric branching pattern of the umbilical vein resembled that of a “distributing” vessel, whereas the more symmetric branching of the portal trunk resembled a “delivering” vessel. At 6 weeks, 3–4 main hepatic-vein outlets drained into the inferior caval vein, of which that draining the caudate lobe formed the intrahepatic portion of the caval vein. More peripherally, 5–6 major tributaries drained both dorsolateral regions and the left and right ventromedial regions, implying a “crypto-lobar” distribution. Lobar boundaries, even in non-lobated human livers, and functional vascular requirements account for the predictable topography and branching pattern of the liver veins, respectively.

A New Assessment of the Prevalence and Risk Factors of Liver Abnormalities in Hyperthyroidism

Clinical ThyroidologyVol. 29, No. 5 SummariesA New Assessment of the Prevalence and Risk Factors of Liver Abnormalities in HyperthyroidismJacques OrgiazziJacques OrgiazziSearch for more papers by this authorPublished Online:1 May 2017https://doi.org/10.1089/ct.2017;29.186-188AboutSectionsView articleView Full TextPDF/EPUB Permissions & CitationsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail View articleFiguresReferencesRelatedDetailsCited byEstablishment of a human embryonic stem cell-based liver differentiation model for hepatotoxicity evaluationsEcotoxicology and Environmental Safety, Vol. 174 Volume 29Issue 5May 2017 InformationCopyright 2017 American Thyroid Association, Inc.To cite this article:Jacques Orgiazzi.A New Assessment of the Prevalence and Risk Factors of Liver Abnormalities in Hyperthyroidism.Clinical Thyroidology.May 2017.186-188.http://doi.org/10.1089/ct.2017;29.186-188Published in Volume: 29 Issue 5: May 1, 2017PDF download

Naringenin suppresses TPA-induced tumor invasion by suppressing multiple signal transduction pathways in human hepatocellular carcinoma cells

Naringenin, a common dietary flavonoid abundantly present in fruits and vegetables, is believed to possess strong anti-proliferative properties and the ability to induce apoptosis in hepatoma cell lines. However, there are no reports describing its effects on the invasion and metastasis of hepatoma cell lines, and the detailed molecular mechanisms of its effects are still unclear. In this study, we investigated the mechanisms underlying naringenin-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 (HepG2, Huh-7, and HA22T) and murine embryonic liver cells (BNL CL2). Naringenin suppressed MMP-9 transcription by inhibiting activator protein (AP)-1 and nuclear factor-κB (NF-κB) activity. It suppressed TPA-induced AP-1 activity through inhibiting the phosphorylation of the extracellular signal-related kinase (ERK) and c-Jun N-terminal kinase (JNK) signaling pathways, and it suppressed TPA-induced inhibition of NF-κB nuclear translocation through IκB. Additionally, it suppressed TPA-induced activation of ERK/phosphatidylinositol 3-kinase/Akt upstream of NF-κB and AP-1. These data suggest that naringenin suppresses the invasiveness and metastatic potential of hepatocellular carcinoma (HCC) by inhibiting multiple signal transduction pathways.

The relationship between Cd-induced autophagy and lysosomal activation in WRL-68 cells.

This study shows that Cd induces autophagy in the human's embryonic normal liver cell line (WRL-68). The expression of LC3B-II and the mature cathepsin L were analyzed by Western blotting. The autophagosomes and lysosomes were directly visualized by electron microscopy and confocal microscopy analysis in Cd-exposed WRL-68 cells. In this study, we first found that autophagy induced the activation of lysosomal function in WRL-68 cells. The lysosomal activation was markedly decreased when the cells were co-treated with 3-MA (an inhibitor of autophagy). Secondly, we provided the evidence that the activation of lysosomal function depended on autophagosome-lysosome fusion. The colocalization of lysosome-associated membrane protein-2 (LAMP2) and GFP-LC3 was significantly reduced, when they were treated with thapsigargin (an inhibitor of autophagosome-lysosome fusion). We demonstrated that deletion or blockage of the autophagosome-lysosome fusion process effectively diminished lysosomal activation, which suggests that lysosomal activation occurring in the course of autophagy is dependent on autophagosome-lysosome fusion. Thirdly, we provided evidence that the activation of lysosomal function was associated with lysosomal acid. We investigated the relationship between autophagosome-lysosome fusion and pH in acidic compartments by visualizing fusion process in WRL-68 cells. This suggests that increasing pH in acidic compartments in WRL-68 cells inhibits the autophagosome-lysosome fusion. Finally, we found that the activation of lysosomal function was associated with Ca(2+) stores and the intracellular Ca(2+) channels or pumps were possibly pH-dependent.

RNA and DNA binding of inert oligonuclear ruthenium(II) complexes in live eukaryotic cells.

Confocal microscopy was used to study the intracellular localisation of a series of inert polypyridylruthenium(II) complexes with three eukaryotic cells lines - baby hamster kidney (BHK), human embryonic kidney (HEK-293) and liver carcinoma (Hep-G2). Co-staining experiments with the DNA-selective dye DAPI demonstrated that the di-, tri- and tetra-nuclear polypyridylruthenium(II) complexes that are linked by the bis[4(4'-methyl-2,2'-bipyridyl)]-1,12-dodecane bridging ligand ("bb12") showed a high degree of selectivity for the nucleus of the eukaryotic cells. Additional co-localisation experiments with the general nucleic acid stain SYTO 9 indicated that the ruthenium complexes showed a considerable preference for the RNA-rich nucleolus, rather than chromosomal DNA. No significant differences were observed in the intracellular localisation between the ΔΔ and ΛΛ enantiomers of the dinuclear complex. Cytotoxicity assays carried out over 72 hours indicated that the ruthenium complexes, particularly the tri- and tetra-nuclear species, were significantly toxic to the eukaryotic cells. However, when the activity of the least cytotoxic compound (the ΔΔ enantiomer of the dinuclear species) was determined over a 24 hour period, the results indicated that the ruthenium complex was approximately a 100-fold less toxic to liver and kidney cells than to Gram positive bacteria. Circular dichroism (CD) spectroscopy was used to examine the effect of the ΔΔ and ΛΛ enantiomers of the dinuclear complex on the solution conformations of RNA and DNA. The CD experiments indicated that the RNA maintained the A-type conformation, and the DNA the B-type structure, upon binding by the ruthenium complexes.

Genistein inhibits tumor invasion by suppressing multiple signal transduction pathways in human hepatocellular carcinoma cells

BackgroundGenistein (Gen) exhibits anti-mutagenic and anti-metastatic activities in hepatoma cell lines. Gen has suppressive effects on tumor growth and angiogenesis in nude mice. Gen suppresses the enzymatic activity of matrix metalloproteinase (MMP)-9; however, the mechanism underlying its anti-invasive activity on hepatocellular carcinoma (HCC) cells is unclear.MethodsIn this study, the possible mechanisms underlying Gen-mediated reduction of 12-O-Tetradecanoylphorbol-13-acetate (TPA)-induced cell invasion and inhibition of secreted and cytosolic MMP-9 production in human hepatoma cells (HepG2, Huh-7, and HA22T) and murine embryonic liver cells (BNL CL2) were investigated.ResultsGen suppressed MMP-9 transcription by inhibiting activator protein (AP)-1 and nuclear factor-κ B (NF-κB) activity. Gen suppressed TPA-induced AP-1 activity through inhibitory phosphorylation of extracellular signal-related kinase (ERK) and c-Jun N-terminal kinase (JNK) signaling pathways, and TPA-stimulated inhibition of NF-κB nuclear translocation through IκB inhibitory signaling pathways. Moreover, Gen suppressed TPA-induced activation of ERK/phosphatidylinositol 3-kinase/Akt upstream of NF-κB and AP-1.ConclusionsGen and its inhibition of multiple signal transduction pathways can control the invasiveness and metastatic potential of HCC.

Purified vitexin compound 1 inhibits growth and angiogenesis through activation of FOXO3a by inactivation of Akt in hepatocellular carcinoma

Vitexins, isolated from the seeds of the Chinese herb Vitex negundo, is known to exert antitumor activity in cancer xenograft models and cell lines. The aim of the current study was to examine whether the Akt/forkhead box protein O3a (FOXO3a) pathway mediates the biological effects of purified vitexin compound 1 (VB-1) in hepatocellular carcinoma (HCC) cells. The effect of VB-1 on the viability of the HCC cell lines HepG2, Hep3B, Huh-7 and the human embryonic liver cells L-02 was investigated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Growth inhibition was assessed by clonogenic assay, and cell cycle arrest was investigated using flow cytometry. Inhibition of angiogenesis was evaluated using a matrigel in vitro HUVEC tube formation assay. The effects on the Akt/FOXO3a pathway were detected by western blotting. VB-1 suppressed the proliferation of HepG2, Hep3B, Huh-7 cells, but had little effect on L-02 cells. VB-1 inhibited anchorage-dependent and -independent HepG2 cell growth in a concentration-dependent manner by induction of cell cycle arrest at G1/G0. VB-1 also reduced the secretion of vascular endothelial growth factor (VEGF), resulting in the inhibition of endothelial tube formation. Phosphorylated Akt and its downstream effector FOXO3a were downregulated in VB-1-treated HepG2 cells. Knockdown of Akt1 by small interfering RNA (siRNA) enhanced growth inhibition, and silencing FOXO3a by siRNA attenuated this action. VB-1 inhibited growth and induced cell cycle arrest at G1/G0 by regulating the Akt/FOXO3a pathway. The findings suggested that VB-1 is a potentially promising candidate for the treatment of HCC.

Protective effect of blueberry anthocyanins in a CCl4-induced injury model in human embryonic liver cells

This study investigates the possible mechanism of the protective effects of blueberry anthocyanins on human embryonic liver L-02 cells. Results of the WST-8 method showed that different concentrations of blueberry anthocyanins protected the human embryonic liver L-02 cells against CCl4-induced injury in a dose-dependent manner with IC50 at 6.2×g/L. Cell clone formation inhibition assay demonstrated that cell clones increased with increased concentration of anthocyanins. Propidium iodide (PI) staining analysis showed that anthocyanins can decrease cell cycle in the G1 phase. DNA ploidy analysis showed a weakened percentage of hypodiploid cells in a dose-dependent manner. Moreover, Annexin V-FITC/PI staining analysis showed a dose-dependent effect of anthocyanins on late apoptotic and necrotic cells. Western blot assay showed gradually decreased caspase-3 protein expression levels with increased anthocyanin concentration. In summary, these findings provided pharmacological evidence supporting the clinical application and protective effect of blueberry anthocyanins against acute liver injury.

Spiclomazine Induces Apoptosis Associated with the Suppression of Cell Viability, Migration and Invasion in Pancreatic Carcinoma Cells

The effective treatment for pancreatic carcinoma remains critically needed. Herein, this current study showed that spiclomazine treatment caused a reduction in viability in pancreatic carcinoma cell lines CFPAC-1 and MIA PaCa-2 in vitro. It was notable in this regard that, compared with pancreatic carcinoma cells, normal human embryonic kidney (HEK-293) and liver (HL-7702) cells were more resistant to the antigrowth effect of spiclomazine. Biochemically, spiclomazine treatment regulated the expression of protein levels in the apoptosis related pathways. Consistent with this effect, spiclomazine reduced the mitochondria membrane potential, elevated reactive oxygen species, and activated caspase-3/9. In addition, a key finding from this study was that spiclomazine suppressed migration and invasion of cancer cells through down-regulation of MMP-2/9. Collectively, the proposed studies did shed light on the antiproliferation effect of spiclomazine on pancreatic carcinoma cell lines, and further clarified the mechanisms that spiclomazine induced apoptosis associated with the suppression of migration and invasion.

A sorghum xylanase inhibitor-like protein with highly potent antifungal, antitumor and HIV-1 reverse transcriptase inhibitory activities

A 25-kDa protein, with an N-terminal amino acid sequence homologous to that of xylanase inhibitor and designated as xylanase inbibitor-like protein (XILP) was purified from sorghum seeds. The isolation protocol consisted of affinity chromatography, ion exchange chromatography, and gel filtration. XILP inhibited mycelial growth in various phytopathogenic fungi. The antifungal activity was thermostable and pH-stable. XILP inhibited proliferation of various cancer cell lines but did not do so in human embryonic liver (WRL 68) cells. There was no mitogenic activity toward mouse splenocytes. XILP reduced the activity of HIV-1 reverse transcriptase with an IC50 of 11.1μM, but lacked inhibitory activity toward HIV-1 integrase and SARS coronavirus proteinase. In conclusion, sorghum XILP is thermostable and pH stable and exhibits potent antifungal, antiproliferative, and HIV-1 reverse transcriptase inhibitory activities.