G2 Cells Research Articles

Optimization of the extract from flower of Citrus aurantium L. var. amara Engl. and its inhibition of lipid accumulation.

Flower of Citrus aurantium L. var. amara Engl. (CAVA) has been confirmed to have promising anti-obesity effects. However, the regulation of alkaloid extracts from flower of CAVA (Al) on lipid metabolism remain unknown. In this study, Al was optimized by ultrasound-assisted extraction using response surface methodology. The optimal conditions were ultrasonic time 72 min, ethanol concentration 78% and liquid/solid ratio 30 ml/g with the maximum alkaloid yield 5.66%. LC-MS assay indicated that the alkaloid compounds were enriched in Al after optimization. Nine alkaloid compounds were identified in Al by LC-MS assay and stachydrine, caffeine and cathine appeared as the major alkaloid compounds. Bioactivity assay showed that Al treatment significantly increased superoxide dismutase (SOD) activity, and reduced malonaldehyde (MDA) and reactive oxygen species (ROS) levels. Al administration also reversed oleic acid-induced hepatic steatosis in Hep G2 cells by inhibiting the expression of lipogenesis-signaling genes including fatty acid synthase (FAS), peroxisome proliferator-activated receptor subtype γ (PPARγ), uncoupling protein 2 (UCP2), and retinol binding protein (RBP4). However, OA-induced reduction of lipolysis-related gene carnitine palmitoyl transferase 1A (CPT1A) in Hep G2 cells was not improved by Al supplementation. Moreover, the increased SOD activity and decreased MDA and ROS contents were also observed in Caenorhabditis elegans by Al addition. Al intervention exhibited the ability to inhibit lipid accumulation in C. elegans by suppressing expression of lipid metabolism-related genes. These results suggested that the alkaloid extracts from the flower of CAVA showed great potential to regulate lipid metabolism. PRACTICAL APPLICATIONS: The extraction of alkaloid extracts from the flower of CAVA was optimized with a maximum yield of 5.66%. The regulatory effects and mechanisms of Al on lipid metabolism of Hep G2 cells and Caenorhabditis elegans were also investigated. More clinical studies are required to evaluate the potential of using alkaloids from the flower of CAVA as therapeutic agents against lipid metabolic disorders.

Galactomannan-Decorated Lipidic Nanocarrier for Gene Supplementation Therapy in Fabry Disease.

Gene supplementation therapy with plasmid DNA (pDNA) represents one of the most promising strategies for the treatment of monogenic diseases such as Fabry disease (FD). In the present work, we developed a solid lipid nanoparticles (SLN)-based non-viral vector with a size below 100 nm, and decorated with galactomannan (GM) to target the liver as an α-Galactosidase A (α-Gal A) production factory. After the physicochemical characterization of the GM-SLN vector, cellular uptake, transfection efficacy and capacity to increase α-Gal A activity were evaluated in vitro in a liver cell line (Hep G2) and in vivo in an animal model of FD. The vector showed efficient internalization and it was highly efficient in promoting protein synthesis in Hep G2 cells. Additionally, the vector did not show relevant agglutination of erythrocytes and lacked hemolytic activity. After the systemic administration to Fabry mice, it achieved clinically relevant α-Gal A activity levels in plasma, liver, and other organs, importantly in heart and kidneys, two of the most damaged organs in FD. This work shows the potential application of GM-decorated lipidic nanocarries for the treatment of FD by pDNA-based gene augmentation.

Facile synthesis of [ONON] type titanium(IV)bis-chelated complexes in alcoholic solvents and evaluation of anti-tumor activity

Novel anti-tumoral diamino-bis-(phenolato) [ONON] type titanium(IV) complexes stabilized by 2,6-dipicolinic acid were synthesis via an efficient protocol using n-propanol as solvent and H2O for isolation. In total 20 [ONON] type and 2 Salan Ti(IV)bis-chelated complexes were synthesized with yields ranging from 68% to 96%. All reactions could reach to completion in 1.5 min at 80 °C either using Ti(OiPr)4 or TiCl4 as starting materials. Most [ONON] type Ti(IV) complexes exhibit selectively enhanced inhibition activity against Hep G2 cells in comparison with Salan Ti(IV) complexes. Among which, the inhibitory activity of 2 t (IC50: 0.15 ± 0.1 μM) against Hep G2 cells is about 80 times enhanced than that of cisplatin (IC50: 12.4 ± 1.2 μM). The [ONON] type Ti(IV) complexes slowly released nontoxic phenolato ligands in presence of large amount of aqueous media, and a fast cellular uptake process was proposed for these Ti(IV) complexes based on metal uptake analysis. Decagram scale synthesis indicates this facile synthetic methodology can be applied to large scale synthesis.

Synthesis, docking study, and structure-activity relationship of novel niflumic acid derivatives acting as anticancer agents by inhibiting VEGFR or EGFR tyrosine kinase activities

A new series of niflumic acid (NF) derivatives were synthesized by esterification of (NF) to give ester compound 1, which was treated with hydrazine hydrate to produce (NF) hydrazide 2. Hydrazine-1-carboxamide compounds (3A–C), and hydrazine-1-carbothioamide derivatives (4A–D) were synthesized by treatment of (NF) hydrazide with phenyl isocyanate, and phenyl isothiocyanate derivatives, respectively. The cyclization of (4B–D) and (3B) was achieved using NaOH solution to produce 1,2,4-triazole derivatives (5A–C) and 6, respectively. The prepared compounds were characterized using IR, 1HNMR, 13CNMR, and MS (ESI) spectroscopy. A molecular docking study was performed to evaluate the binding affinity of the synthesized compounds against EGFR and VEGFR kinase domains which revealed that compounds 3B, and 4A had the best binding energy (-7.87, and -7.33 kcal/mol, respectively) against VEGFR, while compound 5A had the best binding energy (-7.95 kcal/mol) against EGFR. The biological investigation results indicated that all the tested compounds caused cell killing in the two cancer cell lines (Hep G2 and A549) studied, with compound 4C being the most cytotoxic, as well as being cancer selective. Additionally, compound 4C-treated Hep G2 cells were arrested at the S and G2/M cell cycle phases. Cytotoxicity of compound 4C was attributed to apoptosis as determined by flow cytometry and qRT-PCR results of the apoptosis markers p53, BAX, and caspase-3. Finally, compound 4C inhibited VEGFR kinase activity, while compound 5B inhibited EGFR kinase activity. In conclusion, the novel (NF) derivatives are potent anticancer agents, inhibiting cell proliferation by inhibiting EGFR and VEGFR tyrosine kinase enzymes.

Structure and hypoglycemic effect of a neutral polysaccharide isolated from sea cucumber Stichopus japonicus

In addition to its high nutritious value, sea cucumber has been recognized by folk medicine for a long time. This study investigated the structure and hyperglycemic activity of a neutral polysaccharide (NPsj) from sea cucumber Stichopus japonicus, whose molecular weight was determined as 301.75 kDa by HPGPC method. Monosaccharide composition analysis indicated that NPsj is a glucan. The structure of NPsj was obtained by combining the analysis of methylation analysis, FTIR, NMR, periodate oxidation, Smith degradation and ESI-MS, which is mainly composed of (1 → 4)-α-d-glucoses with β-d-glucose(1→) branches substituted at O-6 every 7–9 of 1,4 linked glucoses. An in vitro insulin resistance Hep G2 cells model and a 3 T3-L1 cells model were established, and the NPsj has significant effect to increase glucose consumption with no toxicity at 10–100 μg/mL. Furthermore, NPsj upregulates the phosphorylation of Akt1 and down-regulated GSK3β, and then reduces the phosphorylation of GS, indicating its mechanism of ameliorating insulin resistance via Akt/GSK3β/GS signaling pathway.

Chemical constituents of the stem of Marsypopetalum modestum and their bioactivities

Phytochemical investigation of Marsypopetalum modestum (Annonaceae) led to the isolation of a new phenylpropanoid glycoside, lyciumphenylpropanoid B (10), along with nine known compounds (1-9) from an aqueous methanolic extract of the stem. Most compounds are reported from this genus for the first time. The structures of the isolated compounds were elucidated using spectroscopic methods including NMR spectroscopy, high-resolution mass spectrometry, and quantum chemical electronic circular dichroism (ECD) calculations. Cytotoxic and antitubercular activities of several isolated compounds were evaluated. Dipyrithione (1) displayed anti-mycobacterial (MIC = 0.23 μM) and cytotoxic (IC50 = 0.8 μM in Hep G2 cells; 4.1 μM in HCT 116 cells) activities. Kelampayoside A (8) showed moderate cytotoxic activity against cancer cells.

Metformin Protects Against Sunitinib-induced Cardiotoxicity: Investigating the Role of AMPK.

Sunitinib is associated with cardiotoxicity through inhibition of AMP-protein kinase (AMPK) signaling. By contrast, the common antidiabetic agent metformin has demonstrated cardioprotection through indirect AMPK activation. In this study, we investigate the effects of metformin during sunitinib-induced cytotoxicity. Left ventricular developed pressure, coronary flow, heart rate, and infarct size were measured in Langendorff-perfused rat hearts treated with 1 µM sunitinib ±50 µM metformin ±1 µM human equilibrative nucleoside transporter inhibitor S-(4-Nitrobenzyl)-6-thioinosine (NBTI). Western blot analysis was performed for p-AMPKα levels. Primary isolated cardiac myocytes from the left ventricular tissue were used to measure live cell population levels. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to assess adjunctive treatment of and metformin in human hepatoma G2 and promyelocytic leukemia (HL-60) cells treated with 0.1-100 µM sunitinib ±50 µM metformin. In the perfused hearts, coadministration of metformin attenuated the sunitinib-induced changes to left ventricular developed pressure, infarct size, and cardiac myocyte population. Western blot analysis revealed a significant decrease in p-AMPKα during sunitinib treatment, which was attenuated after coadministration with metformin. All metformin-induced effects were attenuated, and NBTI was coadministered. The MTT assay demonstrated an increase in the EC50 value during coadministration of metformin with sunitinib compared with sunitinib monotherapy in hepatoma G2 and HL-60 cell lines, demonstrating the impact and complexity of metformin coadministration and the possible role of AMPK signaling. This study highlights the novel cardioprotective properties of metformin and AMPK activation during sunitinib-induced cardiotoxicity when administered together in the Langendorff heart model.

A Naphthalimides‐Based Probe for Sequential Detection of Cu2+ and PPi and Its Application in Cells Imaging

AbstractA novel naphthalimides‐based fluorescent probe (NPHD) selectively detected Cu2+ and PPi through “strong yellow‐weak green‐strong yellow” photonic response in a sequential manner. The formation of NPHD‐Cu2+ resulted in obvious fluorescence decreased and wavelength shift from 595 nm to 525 nm by adding Cu2+ to NPHD solution. The mechanism could be attributed to d→π* electron transfer (ET) from Cu2+ to NPHD. Furthermore, the NPHD‐Cu2+ complex exhibited high selectivity toward PPi through via emission recovery of chemosensor NPHD. The detection limits of NPHD to Cu2+ and NPHD‐Cu2+ to PPi were estimated to be 5.72×10<M‐>8M and 2.39×10<M‐>8M. In addition, NPHD could be employed to detect exogenous Cu2+ and PPi in Hep G2 cells.

Synthesis, docking study, and structure activity relationship of novel anti-tumor 1, 2, 4 triazole derivatives incorporating 2-(2, 3- dimethyl aminobenzoic acid) moiety

A series of 1,2,4 triazole derivatives (H7-12) have been synthesized by reacting an excess of hydrazine hydrate with carbothioamide derivatives (H1-6). The final compounds (HB1-HB6) were synthesized by reacting the triazole derivatives with mefenamic acid using DCC as a coupling agent. The chemical structures were confirmed by FT-IR,1H, and13C-NMR spectra, and some physicochemical properties were determined. The cytotoxicity of the different compounds (HB1-HB6) was evaluated by the MTT assay against two human epithelial cancer cell lines, A549 lung carcinoma and Hep G2 hepatocyte carcinoma, and one normal human cell line WI-38 lung fibroblasts. The mode of cell killing (apoptosis versus necrosis), as well as the effect on cell cycle phases were evaluated via flow cytometry. Additionally, EGFR tyrosine kinase inhibition assay was performed. The results presented in the current study indicate that the six tested compounds exhibited cytotoxicity against both cancer cell lines, and the lowest IC50was achieved with compound HB5 against Hep G2 cancer cells which was found to be highly selective against cancer cells. HB5-treated Hep G2 cells were arrested at the S and G2/M cell cycle phases. Compound HB5 caused cell killing via apoptosis rather than necrosis, and this was achieved by inhibiting EGFR tyrosine kinase activity needed for cell proliferation, and cell cycle progression. In silicopre-ADMET studies confirmed all final compounds don’t cause CNS side effects, with little liver dysfunction effect.

Design, synthesis and antitumor evaluation of novel 1H-indole-2-carboxylic acid derivatives targeting 14-3-3η protein

In this work, a series of novel 1H-indole-2-carboxylic acid derivatives targeting 14-3-3η protein were designed and synthesized for treatment of liver cancer. After structural optimization for several rounds, C11 displayed a relatively better affinity with 14-3-3η, as well as the best inhibitory activities against several typical human liver cancer cell lines, including Bel-7402, SMMC-7721, SNU-387, Hep G2 and Hep 3B cells. Compound C11 also displayed best inhibitory activity against chemotherapy-resistant Bel-7402/5-Fu cells. Besides, C11 was rather safe against hERG and possessed moderate T1/2 and CL values in liver microsomes. In anti-proliferation, trans-well and cell apoptosis assays, C11 also showed its huge potential as a potent antitumor agent. Then, Western blot assay was conducted, following analyzed by molecular docking, the anti-proliferative mechanisms of this small-molecule inhibitor were revealed. Moreover, C11 was demonstrated to induce G1-S phase cell cycle arrest in liver cancer cells.

Semi-Synthetic Analogues of Cryptolepine as a Potential Source of Sustainable Drugs for the Treatment of Malaria, Human African Trypanosomiasis, and Cancer.

The prospect of eradicating malaria continues to be challenging in the face of increasing parasite resistance to antimalarial drugs so that novel antimalarials active against asexual, sexual, and liver-stage malaria parasites are urgently needed. In addition, new antimalarials need to be affordable and available to those most in need and, bearing in mind climate change, should ideally be sustainable. The West African climbing shrub Cryptolepis sanguinolenta is used traditionally for the treatment of malaria; its principal alkaloid, cryptolepine (1), has been shown to have antimalarial properties, and the synthetic analogue 2,7-dibromocryptolepine (2) is of interest as a lead toward new antimalarial agents. Cryptolepine (1) was isolated using a two-step Soxhlet extraction of C. sanguinolenta roots, followed by crystallization (yield 0.8% calculated as a base with respect to the dried roots). Semi-synthetic 7-bromo- (3), 7, 9-dibromo- (4), 7-iodo- (5), and 7, 9-dibromocryptolepine (6) were obtained in excellent yields by reaction of 1 with N-bromo- or N-iodosuccinimide in trifluoroacetic acid as a solvent. All compounds were active against Plasmodia in vitro, but 6 showed the most selective profile with respect to Hep G2 cells: P. falciparum (chloroquine-resistant strain K1), IC50 = 0.25 µM, SI = 113; late stage, gametocytes, IC50 = 2.2 µM, SI = 13; liver stage, P. berghei sporozoites IC50 = 6.13 µM, SI = 4.6. Compounds 3–6 were also active against the emerging zoonotic species P. knowlesi with 5 being the most potent (IC50 = 0.11 µM). In addition, 3–6 potently inhibited T. brucei in vitro at nM concentrations and good selectivity with 6 again being the most selective (IC50 = 59 nM, SI = 478). These compounds were also cytotoxic to wild-type ovarian cancer cells as well as adriamycin-resistant and, except for 5, cisplatin-resistant ovarian cancer cells. In an acute oral toxicity test in mice, 3–6 did not exhibit toxic effects at doses of up to 100 mg/kg/dose × 3 consecutive days. This study demonstrates that C. sanguinolenta may be utilized as a sustainable source of novel compounds that may lead to the development of novel agents for the treatment of malaria, African trypanosomiasis, and cancer.

ITRAQ-Based Quantitative Proteomics Reveals the Energy Metabolism Alterations Induced by Chlorogenic Acid in HepG2 Cells.

Epidemiological studies have suggested that coffee consumption is associated with a decrease in the risk of developing obesity and diabetes; however, the detailed mechanisms underlying these effects of coffee consumption remain poorly understood. In this study, we examined the effects of chlorogenic acid on energy metabolism in vitro. Hepatocellular carcinoma G2 (HepG2) cells were cultured in a medium containing chlorogenic acid. Chlorogenic acid increased the activity of mitochondrial enzymes, including citrate synthase, isocitrate dehydrogenase, and malate dehydrogenase (MDH), which are involved in the tricarboxylic acid (TCA) cycle. Proteome analysis using the isobaric tags for the relative and absolute quantitation (iTRAQ) method revealed the upregulation of proteins involved in the glycolytic system, electron transport system, and ATP synthesis in mitochondria. Therefore, we propose a notable mechanism whereby chlorogenic acid enhances energy metabolism, including the TCA cycle, glycolytic system, electron transport, and ATP synthesis. This mechanism provides important insights into understanding the beneficial effects of coffee consumption.

A Novel Selenium Polysaccharide Alleviates the Manganese (Mn)-Induced Toxicity in Hep G2 Cells and Caenorhabditis elegans.

Manganese (Mn) is now known to have a variety of toxicities, particularly when exposed to it in the workplace. However, there are still ineffective methods for reducing Mn’s hazardous effects. In this study, a new selenium polysaccharide (Se-PCS) was developed from the shell of Camellia oleifera to reduce Mn toxicity in vitro and in vivo. The results revealed that Se-PCS may boost cell survival in Hep G2 cells exposed to Mn and activate antioxidant enzyme activity, lowering ROS and cell apoptosis. Furthermore, after being treated with Se-PCS, Caenorhabditis elegans survived longer under Mn stress. daf-16, a tolerant critical gene, was turned on. Moreover, the antioxidant system was enhanced as the increase in strong antioxidant enzyme activity and high expression of the sod-3, ctl-2, and gst-1 genes. A variety of mutations were also used to confirm that Se-PCS downregulated the insulin signaling pathway. These findings showed that Se-PCS protected Hep G2 cells and C. elegans via the insulin/IGF-1 signaling pathway and that it could be developed into a promising medication to treat Mn toxicity.

Rational deuteration of dronedarone attenuates its toxicity in human hepatic HepG2 cells.

Deuteration is a chemical modification strategy that has recently gained traction in drug development. The replacement of one or more hydrogen atom(s) in a drug molecule with its heavier stable isotope deuterium can enhance its metabolic stability and pharmacokinetic properties. However, it remains uninterrogated if rational deuteration at bioactivation "hot-spots" could attenuate its associated toxicological consequences. Here, our preliminary screening with benzofuran antiarrhythmic agents first revealed that dronedarone and its major metabolite N-desbutyldronedarone elicited a greater loss of viability and cytotoxicity in human hepatoma G2 (HepG2) cells as compared with amiodarone and its corresponding metabolite N-desethylamiodarone. A comparison of dronedarone and its in-house synthesized deuterated analogue (termed poyendarone) demonstrated that deuteration could attenuate its in vitro toxicity in HepG2 cells by modulating the extent of mitochondrial dysfunction, reducing the dissipation of mitochondrial membrane potential, and evoking a distinct apoptotic kinetic signature. Furthermore, although pretreatment with the CYP3A inducer rifampicin or the substitution of glucose with galactose in the growth media significantly augmented the loss of cell viability elicited by dronedarone and poyendarone, a lower loss of cell viability was consistently observed in poyendarone across all concentrations. Taken together, our preliminary investigations suggested that the rational deuteration of dronedarone at its benzofuran ring reduces aberrant cytochrome P450 3A4/5-mediated bioactivation, which attenuated its mitochondrial toxicity in human hepatic HepG2 cells.

Chk1 dynamics in G2 phase upon replication stress predict daughter cell outcome

In human cells, ATR/Chk1 signaling couples S phase exit with the expression of mitotic inducers and prevents premature mitosis upon replication stress (RS). Nonetheless, under-replicated DNA can persist at mitosis, prompting chromosomal instability. To decipher how the DNA replication checkpoint (DRC) allows cells to enter mitosis over time upon RS, we developed a FRET-based Chk1 activity sensor. During unperturbed growth, a basal Chk1 activity level is sustained throughout S phase and relies on replication origin firing. Incremental RS triggers stepwise Chk1 over-activation that delays S-phase, suggesting a rheostat-like role for DRC coupled with the replication machinery. Upon RS, Chk1 is inactivated as DNA replication terminates but surprisingly is reactivated in a subset of G2 cells, which relies on Cdk1/2 and Plk1 and prevents mitotic entry. Cells can override active Chk1 signaling and reach mitosis onset, revealing checkpoint adaptation. Cell division following Chk1 reactivation in G2 results in a p53/p21-dependent G1 arrest, eliminating the daughter cells from proliferation.

Gryllus bimaculatus De Geer hydrolysates alleviate lipid accumulation, inflammation, and endoplasmic reticulum stress in palmitic acid-treated human hepatoma G2 cells

Ethnopharmacological relevanceNonalcoholic fatty liver disease (NAFLD) is one of the most common hepatic diseases closely intertwined with saturated fatty acids intake. Therefore, various studies are being conducted to find natural substances to prevent either the onset or progression of NAFLD. According to traditional medicinal literature, it has been reported that Gryllus bimaculatus De Geer (GB) has systemic detoxifying activity; however, the preventive effects of GB on NAFLD have not been elucidated to date. Aim of studyTo evaluate the potential of GB as a material for the mitigation of NAFLD, we investigated the effects of GB hydrolysates on the hepatic lipid accumulation, inflammation, and endoplasmic reticulum (ER) stress in human hepatoma G2 (Hep G2) cells treated with palmitic acid (PA). MethodsSteamed and dried GB was defatted, pulverized, and then lyophilized following hydrolyzation using Neutrase® (GB-N) or Flavourzyme® (GB-F). Hep G2 cells were incubated with GB-N or GB-F at various concentrations (0, 0.25, 0.5, and 1 mg/mL) for 24 h, and then PA was treated for another 24 h. ResultsThe GB-N and GB-F significantly prevented the PA-induced intracellular lipid accumulation in the human liver cells (p < 0.05). Moreover, the GB-N and GB-F increased the hepatic cellular viability against the PA-treatment (p < 0.05). In addition, the GB-N and GB-F significantly ameliorated the PA-inducible proinflammatory cytokines mRNA expression, such as tumor necrosis factor-α and interleukin-1β, compared to the PA-treated hepatic cells (p < 0.05). Furthermore, the GB-N and GB-F inhibited the PA-inducible lipogenic mRNA expression, such as fatty acid synthase, sterol regulatory element-binding protein 1c, and peroxisome proliferator-activated receptor-γ (p < 0.05). Moreover, the GB-N and GB-F alleviated the ER stress-related mRNA expression, such as glucose regulatory protein 78 and X-box binding protein increased in PA-treated cells (p < 0.05). ConclusionsThese results indicate that GB-N and GB-F could be used as materials to prevent the NAFLD onset or progression with alleviating hepatic lipid accumulation, inflammation, and ER stress.

Cytotoxicity and Apoptosis Induction of 6,7-Dehydroroyleanone from Taiwania cryptomerioides Bark Essential Oil in Hepatocellular Carcinoma Cells.

The objective of the present study is to evaluate the cytotoxicity of Taiwania cryptomerioides essential oil and its phytochemical on the Hep G2 cell line (human hepatocellular carcinoma). Bark essential oil has significant cytotoxicity to Hep G2 cells, and S3 fraction is the most active fraction in cytotoxicity to Hep G2 cells among the six fractions. The diterpenoid quinone, 6,7-dehydroroyleanone, was isolated from the active S3 fraction by bioassay-guided isolation. 6,7-Dehydroroyleanone exhibited significant cytotoxicity in Hep G2 cells, and the efficacy of 6,7-dehydroroyleanone was better than the positive control, etoposide. Apoptosis analysis of Hep G2 cells with different treatments was characterized via flow cytometry to confirm the cell death situation. Etoposide and 6,7-dehydroroyleanone could induce the apoptosis in Hep G2 cells using flow cytometric assay. Results revealed 6,7-dehydroroyleanone from T. cryptomerioides bark essential oil can be a potential phytochemical to develop the anticancer chemotherapeutic agent for the treatment of the human hepatocellular carcinoma.

Incorporation of Low Concentrations of Gold Nanoparticles: Complex Effects on Radiation Response and Fate of Cancer Cells.

(1) Background: In oncology research, a long-standing discussion exists about pros and cons of metal nanoparticle-enhanced radiotherapy and real mechanisms behind the tumor cell response to irradiation (IR) in presence of gold nanoparticles (GNPs). A better understanding of this response is, however, necessary to develop more efficient and safety nanoparticle (NP) types designed to disturb specific processes in tumor cells. (2) Aims and Methods: We combined 3D confocal microscopy and super-resolution single molecule localization microscopy (SMLM) to analyze, at the multiscale, the early and late effects of 10 nm-GNPs on DNA double strand break (DSB) induction and repair in tumor cells exposed to different doses of photonic low-LET (linear energy transfer) radiation. The results were correlated to different aspects of short and long-term cell viability. SkBr3 breast cancer cells (selected for the highest incidence of this cancer type among all cancers in women, and because most breast tumors are treated with IR) were incubated with low concentrations of GNPs and irradiated with 60Co γ-rays or 6 MV X-rays. In numerous post-irradiation (PI) times, ranging from 0.5 to 24 h PI, the cells were spatially (3D) fixed and labeled with specific antibodies against γH2AX, 53BP1 and H3K9me3. The extent of DSB induction, multi-parametric micro- and nano-morphology of γH2AX and 53BP1 repair foci, DSB repair kinetics, persistence of unrepaired DSBs, nanoscale clustering of γH2AX and nanoscale (hetero)chromatin re-organization were measured by means of the mentioned microscopy techniques in dependence of radiation dose and GNP concentration. (3) Results: The number of γH2AX/53BP1 signals increased after IR and an additional increase was observed in GNP-treated (GNP(+)) cells compared to untreated controls. However, this phenomenon reflected slight expansion of the G2-phase cell subpopulation in irradiated GNP(+) specimens instead of enhanced DNA damage induction by GNPs. This statement is further supported by some micro- and nano-morphological parameters of γH2AX/53BP1 foci, which slightly differed for cells irradiated in absence or presence of GNPs. At the nanoscale, Ripley’s distance frequency analysis of SMLM signal coordinate matrices also revealed relaxation of heterochromatin (H3K9me3) clusters upon IR. These changes were more prominent in presence of GNPs. The slight expansion of radiosensitive G2 cells correlated with mostly insignificant but systematic decrease in post-irradiation survival of GNP(+) cells. Interestingly, low GNP concentrations accelerated DSB repair kinetics; however, the numbers of persistent γH2AX/53BP1 repair foci were slightly increased in GNP(+) cells. (4) Conclusions: Low concentrations of 10-nm GNPs enhanced the G2/M cell cycle arrest and the proportion of radiosensitive G2 cells, but not the extent of DNA damage induction. GNPs also accelerated DSB repair kinetics and slightly increased presence of unrepaired γH2AX/53BP1 foci at 24 h PI. GNP-mediated cell effects correlated with slight radiosensitization of GNP(+) specimens, significant only for the highest radiation dose tested (4 Gy).

Glycogen synthase kinase 3β inhibition and insulin-receptor binding enhancement of compounds isolated from wild leafy vegetable Acalypha alnifolia

BackgroundThe traditional information of Acalypha alnifolia is captivate towards its therapeutic efficacy against diabetes along with the scientific confirmation were leads to an instinct to extract phytocompounds and to evaluate antidiabetic-molecular mechanism. MethodsColumn chromatographic compound isolation were adopted to extract the pure compounds from bioactive leaf extract and interactions of the compounds with diabetic metabolic proteins were accomplished through in-silico docking studies. By the direction, the anti-diabetic property was valued by the expressions of Glycogen Synthase Kinase 3β(GSK 3β) and Insulin on insulin-Receptor on Hep G2 cells. ResultsThe flavone compound - 5,7-dihydroxy-2-(4-hydroxyphenyl)-3-methoxy-4H-chromen-4-one and pyridine alkaloid - (2,6-dihydroxypyridin-4-yl)(3,5-dihydroxytetrahydro-2H-pyran-4-yl) acetic acid were isolated. Both the compounds reduced the GSK 3β expression and the flavone contributed also to enhance insulin binding ability expediently. ConclusionsThe results concluded that the compounds from A. alnifolia having potent to maintain glucose homeostasis in liver insulin-metabolism. By exploring pharmaceutical importance of the isolated compounds, this study will be an important breakthrough in developing new drug for Diabetes.

Antibacterial metal-phenolic nanosheets as smart carriers for the controlled release of epirubicin hydrochloride.

Bacterial infections can cause serious complications in cancer treatment and have been proven to weaken therapeutic benefits. Recently, antibacterial nanomaterials that serve as carriers for anticancer drug delivery have been attracting extensive interest due to their combined antimicrobial and anticancer activities. In this study, antibacterial metal-phenolic nanosheets (Cu-TA) were successfully prepared via the self-assembly of the metal-phenolic coordination complexes formed between copper ions and tannic acid, and the structure, morphology, and formation mechanism of Cu-TA nanosheets were explored. The antibacterial activity of Cu-TA nanosheets against both Gram-positive and Gram-negative bacteria was detected using the minimum inhibitory concentration (MIC), zone of inhibition and plate counting methods. The MIC values of both bacterial strains were about 0.4 mg mL-1, and the killing rates of Cu-TA samples were close to 100% at the concentration of 2 and 0.2 mg mL-1 after 12-hour incubation. Epirubicin hydrochloride (EPI) molecules were successfully loaded on the porous Cu-TA nanosheets mainly through the formation of the Cu-EPI chelate complex and strong electrostatic interactions. The Cu-EPI complex and Cu-TA nanosheets could be disassembled under acidic conditions or in the presence of high levels of glutathione (GSH) after uptake by cancer cells, which triggered the unique pH and GSH-responsive controlled release behaviors of EPI and copper ions. The MTT assay results revealed that the presence of bacteria in Hep G2 cells can greatly impair the cell death rate induced by free EPI, but the resultant EPI-loaded Cu-TA nanosheets can significantly enhance cell death both in the presence and absence of bacteria.