Line HepG2 Research Articles

Silver nanoparticles synthesized using aerial part of Achillea fragrantissima and evaluation of their bioactivities

Achillea fragrantissima (A. fragrantissima), a desert plant, is used internally in Arabian traditional medicine to treat inflammatory, spasmodic gastrointestinal disorders, and hepatobiliary diseases. The study focuses on the environmentally friendly production of silver nanoparticles (AgNPs) from the water-based aerial parts of the A. fragrantissima plant and their ability to kill bacteria and cells. Ultraviolet-visible (UV-Vis) spectroscopy, scanning electron microscopy (SEM) with energy dispersive X-ray (EDX) analysis, transmission electron microscopy (TEM), and Fourier transform infrared spectroscopy (FTIR) were used to describe the AgNPs. They were then tested for their ability to fight cancer and bacteria. A change in colour from yellow to brown and a surface plasmon resonance peak at 440 nm, seen with UV-Vis spectroscopy, showed that AgNPs had formed. In a Gas Chromatography-Mass Spectrometry (GC-MS) test of the aerial parts of A. fragrantissima, twenty bioactive components were found. These included isolongifolol and 3E,10Z-Oxacyclotrideca-3,10-diene-2,7-dione, methylbuta-1,3-dienyl)-7-oxabicyclo [4.1.0] heptan-3-ol. The extract exhibited high phenolic and flavonoid content (77.52 ± 1.46 mg GAE/g dry weight and 59 ± 2.17 mg QE/g dry weight, respectively). According to the IC50 values of 17.2 ± 1.18 and 14 ± 2.43 µg/mL, the AgNPs had a lot of power to kill cancer cells from the MCF-7 and HepG2 lines. Some genes that cause cell death (caspase-3, 8, 9, and Bax) were turned on more in the treated cells compared to the control cells that had not been treated. These genes were Bcl-xL and Bcl-2. Additionally, substantial activity against both Gram-positive bacteria and Gram-negative bacteria was found by antibacterial screening. Overall, this study underscores A. fragrantissima’s diverse biological activity and its potential in drug discovery and nanomedicine, promoting the development of natural antibacterial and anticancer therapies.

5149 In-vitro Combo Treatment of Human Liver Cancer cell (Hep-G2) Line with Vitamin-A and RU-486

Abstract Disclosure: P. Ramaraj: None. Our previous in-vitro studies involving vitamins (vitamin-A and vitamin-D3) and steroids (progesterone and RU-486) pointed to a combination of vitamin-A and RU-486 as an efficient combo to decrease human melanoma cell growth. We extended this combo treatment to human liver cancer cell (Hep-G2) line. The aim was to check whether vit-A and RU-486 combo would also decrease human liver cancer cell line growth. Liver cancer (Hep-G2) cells were plated in a 96 well plate overnight. Following day, vit-A and RU-486 were added either alone or in different combinations for 48 hrs. After 48 hrs, cell viability and percentage of cell growth were monitored by the MTT assay. Supernatants were collected for Elisarray and quantitation of cytokine(s) by Elisa. Individual vit-A and RU-486 treatments showed a dose-dependent decrease in cell growth. Though various combinations of vit-A (50, 75, 100 μM) and RU-486 (10, 50 100 μM) also showed a decrease in cell growth, an experimentally significant decrease in cell growth was observed at vit-A 75 μM and RU-486 50 μM combination. This combo resulted in 24% cell growth compared to straight vit-A (75 μM) at 44% cell growth and RU-486 (50 μM) at 35% cell growth. Thus, this combo inhibited liver cancer cell growth also. In the future effect of this combo on normal liver cell line and the cytokine(s) if any, playing a central role in cell growth would be identified by Elisarray and quantitated by Elisa. Presentation: 6/3/2024

5149 In-Vitro Combo Treatment of Human Liver Cancer Cell (Hep-G2) Line With Vitamin-A And RU-486

Abstract Disclosure: P. Ramaraj: None. Our previous in-vitro studies involving vitamins (vitamin-A and vitamin-D3) and steroids (progesterone and RU-486) pointed to a combination of vitamin-A and RU-486 as an efficient combo to decrease human melanoma cell growth. We extended this combo treatment to human liver cancer cell (Hep-G2) line. The aim was to check whether vit-A and RU-486 combo would also decrease human liver cancer cell line growth. Liver cancer (Hep-G2) cells were plated in a 96 well plate overnight. Following day, vit-A and RU-486 were added either alone or in different combinations for 48 hrs. After 48 hrs, cell viability and percentage of cell growth were monitored by the MTT assay. Supernatants were collected for Elisarray and quantitation of cytokine(s) by Elisa. Individual vit-A and RU-486 treatments showed a dose-dependent decrease in cell growth. Though various combinations of vit-A (50, 75, 100 μM) and RU-486 (10, 50 100 μM) also showed a decrease in cell growth, an experimentally significant decrease in cell growth was observed at vit-A 75 μM and RU-486 50 μM combination. This combo resulted in 24% cell growth compared to straight vit-A (75 μM) at 44% cell growth and RU-486 (50 μM) at 35% cell growth. Thus, this combo inhibited liver cancer cell growth also. In the future effect of this combo on normal liver cell line and the cytokine(s) if any, playing a central role in cell growth would be identified by Elisarray and quantitated by Elisa. Presentation: 6/3/2024

Abstract 2308 In-vitro Combo Treatment of Human Liver Cancer Cell (Hep-G2) Line with a Cocktail that had Inhibited Human Melanoma Cell Growth

Abstract 2308 In-vitro Combo Treatment of Human Liver Cancer Cell (Hep-G2) Line with a Cocktail that had Inhibited Human Melanoma Cell Growth

Molecular Profile of the HepG2 Tumor Cell Line

Cell lines are widely used in scientific research due to their accessibility, stability, and functional similarity to the original cells. The HepG2 line, being the fourth most popular cell culture, is often used in toxicological and metabolic studies due to its partial retention of hepatocyte properties.In our study, the molecular portrait of the HepG2 cell culture was constructed for the first time. To build this portrait, we used previously obtained data for a single sample, including results of whole-genome sequencing (WGS), whole-genome bisulfite sequencing (WGBS), transcriptome (RNA-seq), translatome (Polysome-seq), and proteome (LC-MS/MS) profiling. For the assessment of HepG2 cell line heterogeneity, we analyzed whole-genome and transcriptome data published in the NCBI SRA database, as well as proteome research results available in the PRIDE resource.Our study showed that the HepG2 cell line generally demonstrates a high degree of stability at the genomic and transcriptomic levels; however, samples from China require closer attention when transferring the results of transcriptomic and proteomic experiments. The HepG2 genotype is characterized by stable chromosomal rearrangements, such as translocation between the short arms of chromosomes 1p and 21p, tetrasomy of chromosome 20, loss of the short arm of all SAT chromosomes, and the long arm of the Y chromosome. Despite the absence of 1216 protein-coding genes at the genomic level, 12,602 genes are expressed at the transcriptomic level, of which only 10,461 are detected at the translatome level, and only 1027 genes are identified at the proteome level, which is related to the limitations of mass spectrometry sensitivity. As a result of the omics data analysis, we presented a detailed molecular portrait of the HepG2 cell culture, illustrating the omics profile at various levels for each gene.

Effects of Ultrasound Contrast Agent-Encapsulated Paclitaxel Extract on the Proliferation and Invasion Ability of Hepatocellular Carcinoma Cells

Regarding to the limitations of paclitaxel (PTX) in cancer treatment, PTX was extracted from Taxus chinensis and PTX nano-ultrasound contrast agents (NUCA) were prepared to investigate their impacts on proliferation and invasion of hepatocellular carcinoma (HCC) cells (HCCCs). The PTX extract was obtained through extraction and multi-step purification methods using Taxus chinensis as the source material and poly(lactic-co-glycolic acid)-carboxylic acid (PLGA-COOH) as the experimental material. A modified double emulsion solvent evaporation (DESE) method was employed to prepare paclitaxel-loaded PLGA NUCA (PLGA@PTX). The particle size distribution (PSD) and zeta potential (ZP) of PLGA@PTX were identified using a laser particle size (PS) analyzer, while the drug-loading capacity (DLC) and encapsulation efficiency (EE) of PTX in PLGA@PTX NUCA were evaluated using high-performance liquid chromatography (HPLC). The in vitro release rate (IVRR) of PTX from PLGA@PTX NUCA was also analyzed. HepG2 lines, a human HCC cell line, were grouped into four randomly: a blank control group (Blank), a PTX group, a blank nano-contrast agent group without PTX encapsulation (PLGA), and a PTX-loaded NUCA group (PLGA@PTX). In the Blank group, HepG2 lines were cultured conventionally for 12 hours, while PTX or PLGA@PTX was added to the PTX and PLGA@PTX groups, respectively, to achieve a required concentration (10−7 mol/L) of PTX. An equal amount of PLGA nanoparticles was added to the PLGA group. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Transwell were utilized to judge the impacts of PLGA@PTX nanoparticles on proliferation and invasion of HepG2 lines, respectively. Moreover, flow cytometry (FCT) was utilized to examine the influence of PLGA@PTX nanoparticles on cell cycle (CC) and apoptosis of HepG2 lines. The results revealed that the purity of the PTX extract was as high as 99.04±0.92%. The average PS of PLGA@PTX NUCA was (432.79±4.56) nm, with a surface potential of (−10.79±2.28) mV. Furthermore, the EE and DLC were (89.27±2.63) % and (9.03±0.29) %, respectively. The inhibition rate (IR) to HepG2 lines and cell invasion and the apoptotic rate (AR) in the PLGA@PTX group were much higher to those in the PLGA and PTX groups (P <0.01, P <0.05). The ratio of G1/G0 phase in the CC was greatly lower in the PLGA@PTX group to the PLGA and PTX groups, showing obvious differences with (P <0.05), while that of G2/M phase was higher (P <0.05). These findings indicated that the prepared PLGA@PTX NUCA hindered the proliferation and invasion of HepG2 lines and induced CC arrest at the G2/M phase and apoptosis.

ПРОИЗВОДНЫЕ 3-АРИЛИДЕН-2-ОКСИНДОЛА КАК АНАЛОГИ МЕЛАТОНИНА, ОБЛАДАЮЩИЕАНТИОКСИДАНТНЫМИ СВОЙСТВАМИ И СНИЖАЮЩИЕ ВНУТРИГЛАЗНОЕ ДАВЛЕНИЕ

Aim.Glaucoma is a whole group of diseases caused by increased intraocular pressure (IOP), leading to atrophy of the optic nerve and irreversible blindness. Melatonin has a wide range of biological activity, including antioxidant and antiglaucoma, but its use is limited by insufficient stability and duration of action. To search for new, more effective antiglaucoma agents among derivatives of 3-arylidene-2-oxindole, melatonin analogues, a search for quinonoxidoreductase 2 (NQO2) inhibitors was performed, antioxidant activity was determined and the effect of active compounds on intraocular pressure was studied.Materials and methods.The study of inhibitory activity against NQO2 was evaluated kinetically using menadione and N-benzyl-dihydronicotinamide assubstrate and co-substrate, respectively. Antiradical activity was studied for reducing superoxide-dependentluminescence in a reaction medium containing hemoglobin, luminol and hydrogen peroxide using the Infinite M200 PRO microplate reader (Tecan, Austria), as well as for free radical binding tests OH•-, ABTS and DPPH. Cytotoxicity assessment was carried out using a standard MTT test on MCF-7 and HepG2 line cells. Intraocular pressure in laboratory animals was measured using a contactveterinary tonometer Tonovet (Finland).Results.Derivatives of 3-arylidene-2-oxindole have demonstrated significant antioxidant activity. A number of NQO2 inhibitors have also been identified. The most active compound inhibits human NQO2 with an IC50of 0,4 μM. The compound also showed anti-radical activity, binding the superoxide anion to IC50of 6,1 μM, superior to melatonin on both counts, and low cytotoxicity. With topical application of a 0,4% solution, the compound reduced the intraocular pressure of intact rats by (40,9 ± 6,4) %, while not having a systemic hypotensive effect.Conclusion.Derivatives of 3-arylidene-2-oxindole are promising for drug discovery for the treatment of eye diseases associatedwith increased intraocular pressure or oxidative stress, such as glaucoma, uveitis and diabetic retinopathy.

Alkaloids of Dicranostigma franchetianum (Papaveraceae) and Berberine Derivatives as a New Class of Antimycobacterial Agents.

Tuberculosis (TB) is a widespread infectious disease caused by Mycobacterium tuberculosis. The increasing incidence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains has created a need for new antiTB agents with new chemical scaffolds to combat the disease. Thus, the key question is: how to search for new antiTB and where to look for them? One of the possibilities is to search among natural products (NPs). In order to search for new antiTB drugs, the detailed phytochemical study of the whole Dicranostigma franchetianum plant was performed isolating wide spectrum of isoquinoline alkaloids (IAs). The chemical structures of the isolated alkaloids were determined by a combination of MS, HRMS, 1D, and 2D NMR techniques, and by comparison with literature data. Alkaloids were screened against Mycobacterium tuberculosis H37Ra and four other mycobacterial strains (M. aurum, M. avium, M. kansasii, and M. smegmatis). Alkaloids 3 and 5 showed moderate antimycobacterial activity against all tested strains (MICs 15.625–31.25 µg/mL). Furthermore, ten semisynthetic berberine (16a–16k) derivatives were developed and tested for antimycobacterial activity. In general, the derivatization of berberine was connected with a significant increase in antimycobacterial activity against all tested strains (MICs 0.39–7.81 μg/mL). Two derivatives (16e, 16k) were identified as compounds with micromolar MICs against M. tuberculosis H37Ra (MIC 2.96 and 2.78 µM). All compounds were also evaluated for their in vitro hepatotoxicity on a hepatocellular carcinoma cell line (HepG2), exerting lower cytotoxicity profile than their MIC values, thereby potentially reaching an effective concentration without revealing toxic side effects.

Abstract 1662: Orthotopic model of hepatocellular carcinoma (HCC) using Huh-7 and HepG2: Application of micro-CT in detection of cancer progression

Abstract Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer and was listed as the third cause of death by cancer in 2020 (WHO). Hepatocarcinogenesis is a multistep process and to date it is still poorly understood. HCC occurs almost entirely in patients with underlying chronic liver disease and cirrhosis. Pre-clinical animal models play an important role in unveiling cancer biology which can facilitate development and testing of new antineoplastic drugs. The aim of this study was to establish an orthotopic model of HCC utilizing two cell lines Huh-7 and HepG2, which could be effectively used to study efficacy of new potential treatments. In a follow up study, the correlation of tumor marker alpha-fetoprotein (AFP) and tumor growth was investigated. AFP was measured using Quantikine® ELISA kit. For the intrahepatic (ih) inoculation cancer cells were injected into the left lobe of the liver. For the intrasplenic (is) inoculations of HCC cells, the spleen was exposed, and cells were injected into spleen parenchyma along the long axis of the spleen. Afterwards spleen was removed to avoid intrasplenic tumor growth. The animals were weighed two times a week throughout the study and appearance of any clinical signs was closely monitored. The animals were terminated 70 days after cancer cell implantation. The mice did not present with any acute symptoms before the end stage disease. However, prior sacrifice some swelling and darkening of the abdominal area especially in the ih-model was observed. To visualize liver ExiTron Nano 6000 nanoparticle contrast agent was used. The contrast agent was administered intravenously once during the study, prior the first imaging. Disease progression was monitored using Bruker SkyScan 1276 High-Resolution Micro-CT Scanner. The in vivo measurements were performed using one field of view (FOV) and an image pixel size of 18 μm. Each in vivo scanning was completed within 12 minutes and the estimated radiation dose was kept under 500 mGy at each imaging session. The first tumours were detected by µCT between 2-3 weeks depending more on the cancer cell implantation route than the cell line used. The take rate in the liver was 85% in the ih-model, and less than 50% in the is-model. The intrahepatic inoculation led to more uniform and measurable tumor nodules observed in majority of operated animals, whereas the intrasplenic tumors were dispersed throughout with multiple nodules. In addition, a tight correlation between tumor volume and amount of alpha-fetoprotein detected from serum samples was found. In conclusion, an orthotopic model using intrahepatic approach of cancer cells implantation was established. Application of µCT enabled following tumor growth and recognition of cancer spread in the abdomen. Altogether this provides a promising tool contributing to the development of novel therapies targeting HCC in vivo. Citation Format: Justyna Zdrojewska, Mari Suominen, Shirin Katoozi, Katja Fagerlund, Jukka Rissanen, Roger Malerbakken-Bjerke, Jenny Karlsson. Orthotopic model of hepatocellular carcinoma (HCC) using Huh-7 and HepG2: Application of micro-CT in detection of cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1662.

In Silico Analysis and Functional Characterization of Antimicrobial and Insecticidal Vicilin from Moth Bean (Vigna aconitifolia (Jacq.) Marechal) Seeds.

Vicilin has nutraceutical potential and different noteworthy medicative health-promoting biotic diversions, and it is remarkable against pathogenic microorganisms and insects. In this study, Vigna aconitifolia vicilin (VacV) has been identified and characterized from the seed of Vigna aconitifolia (Jacq.) Marechal (Moth beans). LC-MS/MS analysis of VacV provided seven random fragmented sequences comprising 238 residues, showing significant homology with already reported Vigna radiata vicilin (VraV). VacV was purified using ammonium sulfate precipitation (60%) followed by size exclusion chromatography on Hi-Load 16/60 Superdex 200 pg column and anion-exchange chromatography (Hi trap Q FF column). Purified VacV showed a major ~50 kDa band and multiple lower bands on 12% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) under both reduced and non-reduced conditions. After all, a three-dimensional molecular structure of VacV was predicted, which showed β-sheeted molecular conformation similar to crystallographic structure of VraV. All Vicilins from V. aconitifolia and other plants were divided into six sub-groups by phylogenetic analysis, and VacV shared a high degree of similarity with vicilins of Vigna radiata, Pisum sativum, Lupinus albus, Cicer arietinum and Glycine max. Additionally, VacV (20 μg) has significant growth inhibition against different pathogenic bacteria along strong antifungal activity (50 μg). Likewise, VacV (3.0 mg) produced significant growth reduction in Rice Weevil Sitophilus oryzae larvae after 9 days compared with control. Furthermore, by using MMT assay, the cytotoxicity effect of VacV on the growth of HepG2 liver cancerous cells was tested. VacV showed cytotoxicity against the HepG-2 line and the acquired value was 180 µg after 48 h. Finally, we performed molecular docking against caspase-3 protein (PDB ID: 3DEI) for VacV bioactive receptor interface residues. Hence, our results reveal that VacV, has nutraceutical potential and moth beans can be used as a rich resource of functional foods.

Integrin α2 and β1 Cross-Communication with mTOR/AKT and the CDK-Cyclin Axis in Hepatocellular Carcinoma Cells.

Simple SummaryHepatocellular carcinoma (HCC) progression depends on two major processes, tumor growth and invasion. The present study investigated how these events are linked. A panel of HCC cell lines were stimulated with insulin-like growth factor-1 (IGF1) and the biological behavior was evaluated. IGF1 activated the proliferation and invasion cascade by altering the expression level of integrin α subtypes, which were associated with the AKT-mTOR pathway and the CDK-Cyclin axis. We assume that HCC progression is controlled by a fine-tuned network between IGF1 driven integrin signaling, the Akt-mTOR pathway, and the CDK-Cyclin axis. Concerted targeting of these pathways may, therefore, become an innovative option to prevent cancer dissemination.Integrin receptors contribute to hepatocellular carcinoma (HCC) invasion, while AKT-mTOR signaling controls mitosis. The present study was designed to explore the links between integrins and the AKT-mTOR pathway and the CDK-Cyclin axis. HCC cell lines (HepG2, Huh7, Hep3B) were stimulated with soluble collagen or Matrigel to activate integrins, or with insulin-like growth factor 1 (IGF1) to activate AKT-mTOR. HCC growth, proliferation, adhesion, and chemotaxis were evaluated. AKT/mTOR-related proteins, proteins of the CDK-Cyclin axis, focal adhesion kinase (FAK), and integrin-linked kinase (ILK) were determined following IGF1-stimulation or integrin knockdown. Stimulation with collagen or Matrigel increased tumor cell growth and proliferation. This was associated with significant alteration of the integrins α2, αV, and β1. Blockade of these integrins led to cell cycle arrest in G2/M and diminished the number of tumor cell clones. Knocking down the integrins α2 or β1 suppressed ILK, reduced FAK-phosphorylation and diminished AKT/mTOR, as well as the proteins of the CDK-Cyclin axis. Activating the cells with IGF1 enhanced the expression of the integrins α2, αV, β1, activated FAK, and increased tumor cell adhesion and chemotaxis. Blocking the AKT pathway canceled the enhancing effect of IGF on the integrins α2 and β1. These findings reveal that HCC growth, proliferation, and invasion are controlled by a fine-tuned network between α2/β1-FAK signaling, the AKT-mTOR pathway, and the CDK–Cyclin axis. Concerted blockade of the integrin α2/β1 complex along with AKT-mTOR signaling could, therefore, provide an option to prevent progressive dissemination of HCC.

Radiation and chemotherapy variable response induced by tumor cell hypoxia: impact of radiation dose, anticancer drug, and type of cancer

Hypoxia is a condition in which proliferating tumor cells are deprived of oxygen due to limited blood supply from abnormal tumor microvasculature. This study aimed to investigate the molecular changes that occur in tumor cell hypoxia with special emphasis placed on the efficacy of chemotherapeutic and radiation-related effects. Four commercially available chemotherapeutic agents: cisplatin, cyclophosphamide, doxorubicin, and 5-fluorouracil, were tested for their cytotoxic activity on the cancer cell lines PC3 (prostate), HepG2 (liver), and MCF-7 (breast). Tumor cell lines under hypoxia were treated with both IC50 concentrations of the different chemotherapeutic agents and irradiated with 5 and 10 Gy using a 137Cs gamma source. Hypoxia-inducible factor-1α (HIF-1α) protein levels were examined using an ELISA assay. Hypoxic cells showed a significant change in cell viability to all chemotherapeutic agents in comparison to normoxic controls. HepG2 cells were more resistant to the cytotoxic drug doxorubicin compared to other cancer cell lines. The flow cytometric analysis showed that hypoxic cells have lower levels of total apoptotic cell populations (early and late apoptosis) compared to normoxic cells suggesting decreased hypoxia-induced apoptosis in cancer cells. The highest reduction in HIF-1α level was observed in the MCF-7 cell line (95.5%) in response to the doxorubicin treatment combined with 10 Gy irradiation of cells. Chemoradiotherapy could result in minimal as well as a high reduction of HIF-1α based on cell type, type of chemotherapy, and amount of ionizing radiation. This study highlights future research work to optimize a combined chemoradiotherapeutic regime in individual cancer cell hypoxia.

New cobalt(III) complex with triethylenetetramine and 2,2′-bipyridine: synthesis, crystal structure, DNA interaction, hirshfeld surface, DFT analysis, and cytotoxicity

In this study, a new cobalt (III) complex, [Co(trien)(bpy)](ClO4)2Cl (bpy = 2,2-Bipyridine), was successfully synthesized and thoroughly characterized using elemental analyses, IR, UV-visible, Mass spectrum and single-crystal X-ray crystallography. The complex formed in a monoclinic system with space group P21/c, a = 14.6681(4) Å, b = 10.7617(3) Å, c = 15.1670(4) Å, α = 90°, β = 97.75°, and γ = 90°. The distorted octahedral geometry adopted by Co(III) ion coordinated through two N atoms from the 2,2-Bipyridine ligand and four N donor atoms of Triethylenetetramine ligand. UV-Vis absorption, fluorescence spectroscopic, and cyclic voltammetric techniques were used to investigate the binding of this cobalt (III) complex with calf thymus DNA (CT-DNA). Furthermore, the cleavage of pBR322 DNA with cobalt (III) complex was investigated using gel electrophoresis method. The exhibited potent cytotoxic effects against human cell line (HepG2). The primary coordination sphere of Co(III) complex is optimized, structural parameters are calculated and energy gaps of frontier orbital (HOMO-LUMO) have been calculated with B3LYP/6-31G(d,p)/LANL2DZ level of theory in the gaseous phase. The calculated geometric and spectral results closely matched the experimental data. Theoretically calculated molecular orbitals (HOMO-LUMO) and their energies indicate that charge transfer occurs within the complex. NBO and QTAIM analyses were also carried to evaluate occupied and unoccupied localized orbitals and nature of interactions between the ligand and metal ion.

Chemical Characterization of Taif Rose (Rosa damascena Mill var. trigentipetala) Waste Methanolic Extract and Its Hepatoprotective and Antioxidant Effects against Cadmium Chloride (CdCl2)-Induced Hepatotoxicity and Potential Anticancer Activities against Liver Cancer Cells (HepG2)

Taif rose (Rosa damascena Mill) is one of the most important economic products of the Taif Governorate, Saudi Arabia. Cadmium chloride (CdCl2) is a common environmental pollutant that is widely used in industries and essentially induces many toxicities, including hepatotoxicity. In this study, the major compounds in the waste of Taif rose extract (WTR) were identified and chemically and biologically evaluated. GC–MS analysis of WTR indicated the presence of many saturated fatty acids, vitamin E, triterpene, dicarboxylic acid, terpene, linoleic acid, diterpenoid, monoterpenoid, flavonoids, phenylpyrazoles, and calcifediol (vitamin D derivative). The assessment of potential anticancer activity against HepG2 cells proved that WTR had a high cell killing effect with IC50 of 100–150 µg/mL. In addition, WTR successfully induced high cell cycle arrest at G0/G1, S, and G2 phases, significant apoptosis, necrosis, and increased autophagic cell death response in the HepG2 line. For the evaluation of its anti-CdCl2 toxicity, 32 male rats were allocated to four groups: control, CdCl2, WTR, and CdCl2 plus WTR. Hepatic functions and antioxidant biomarkers (SOD, CAT, GRx, GPx, and MDA) were examined. Histological changes and TEM variations in the liver were also investigated to indicate liver status. The results proved that WTR alleviated CdCl2 hepatotoxicity by improving all hepatic vitality markers. In conclusion, WTR could be used as a preventive and therapeutic natural agent for the inhibition of hepatic diseases and the improvement of redox status. Additional in vitro and in vivo studies are warranted.

Synthesis, screening as potential antitumor of new poly heterocyclic compounds based on pyrimidine-2-thiones

BackgroundContinuing our interest in preparing of new heterocyclic compounds and examining their various biological activities, this work was designed to prepare new condensed and non-condensed heterocyclic compounds 9a-c, 10a-c, 11a-c, 13a-c and 14a-c were synthesized starting with pyrimidine-2-thiones 4a-c.ResultsThiazolo[3,2-a]pyrimidines 9a-c were synthesized by S-alkylation of pyrimidine-2-thiones,4a-c, internal cyclization in alkaline medium with ammonia, condensation with benzaldehyde and finally reaction with hydroxylamine hydrochloride.[1,2,4]thiadiazolo[4,5-a]pyrimidines 11a-c were formed by heating of the 4a-c with benzoylcholride to afford 10a-c followed by reaction with sodium hypochlorite, ammonia and sodium hydroxide. Cyclocondensation of 4a-c with ethyl acetoacetate or formic acid yielded pyrazol-3-ones 13a-c or [1,2,4] triazolo[4,3-a]pyrimidines 14a-c, respectively Elements analysis, IR, 1H-NMR, 13C-NMR and mass spectra were used to validate the structures of newly synthesized heterocycles. Screening of the selected compounds 4a, 6a, 7a, 9a, 10a, 13a and 14a against colon carcinoma cell lines (HCT-116) and hepatocellular carcinoma cell lines (HepG-2).ConclusionsElements analysis, IR, 1H-NMR, 13C-NMR and mass spectra were used to validate the structures of newly synthesized heterocycles. Screening of the selected compounds 4a, 6a, 7a, 9a, 10a, 13a and 14a against colon carcinoma cell lines (HCT-116) and hepatocellular carcinoma cell lines (HepG-2) showed that compound 10a exhibited the most cytotoxic, while compounds 4a, 6a and 14a exhibited considerable cytotoxic activity.

Synthesis, and docking studies of novel heterocycles incorporating the indazolylthiazole moiety as antimicrobial and anticancer agents

The current study was directed toward developing a new series of fused heterocycles incorporating indazolylthiazole moiety. The newly synthesized compounds were characterized through elemental analysis and spectral data (IR, 1H-NMR, 13C-NMR, and Mass Spectrometry). The cytotoxic effect of the newly synthesized compounds was evaluated against normal human cells (HFB-4) and cancer cell lines (HepG-2 and Caco-2). Among the synthesized compounds, derivatives 4, and 6 revealed a significant selective antitumor activity, in a dose-dependent manner, against both HepG-2 and Caco-2 cell lines, with lower risk toward HFB-4 cells (normal cells). Derivative 8 revealed the maximum antitumor activity toward both tumor cell lines, with an SI value of about 26 and IC50 value of about 5.9 μg/mL. The effect of these derivatives (8, 4, and 6) upon the expression of 5 tumor regulating genes was studied through quantitative real-time PCR, where its interaction with these genes was simulated through the molecular docking study. Furthermore, the antimicrobial activity results revealed that compounds 2, 7, 8, and 9 have a potential antimicrobial activity, with maximum broad-spectrum activity through compound 3 against the three tested pathogens: Streptococcus mutans, Pseudomonas aeruginosa, and Candida albicans. The newly prepared compounds also revealed anti-biofilm formation activity with maximum activity against Streptococcus mutans, Pseudomonas aeruginosa, and Candida albicans, respectively.

Biomass Accumulation of Gynostemma Pentaphyllum (Thunb.) Makino in Cell Suspension Cultures inhibiting Human Cancer Cell Growth

Gynostemma pentaphyllum (Thunb.) Makino (GpM) is a medicinal plant in traditional medicine throughout Asia for the treatment of several diseases including cancer. GpM plant cell suspension cultures provide a time and cost effective well-controlled means promising a high-yielding biomass production of pharmaceutical compounds. The purpose of the current work is to investigate the effect of GpM cell suspension cultures on human cancer cell lines growth. The biomass was produced by cell suspension culture of GpM callus into 250 mL Erlenmeyer flask containing 50 mL of liquid medium culture. Gypenosides in GpM were confirmed by HPLC. Pharmacological activities of GpM extract were tested on human cancer cell line (HepG2, Huh7, A549 and HL-60) using Sulforhodamine B (SRB) cytotoxicity assay and Tetrazolium (MTT) assay. We successfully produced 5.485 ± 0.223 g GpM fresh biomass per 250 mL Erlenmeyer flask after 18 days culture. Total gypenosides and Rb1 in dry cell suspension were 48.844 ± 3.933 mg/g and 0.041 ± 0.004 mg/g. The crude extract from GpM cell suspension cultures exhibited significant cancerous cell growth inhibition in a dose dependent manner. From the MTT assay and SRB cytotoxicity assay, it is obvious that GpM cell suspension culture extract at 200 μg/mL significantly inhibited the growth of multiple human cancer cells including hepatoma cell lines (HepG2, Huh7), lung carcinoma cell line (A549) and leukemia cell line (HL-60). Anti-cancer cell proliferation properties of GpM cell suspension culture were significantly higher than those of natural plants’ extracts. In this framework, GpM in cell suspension cultures was found to inhibit the proliferation of several human cancer cells. Biomass accumulation of GpM in cell suspension cultures may contribute to the development of efficient strategies for plant-derived anticancer compound production.

Characterization of chikusetsusaponin IV and V induced apoptosis in HepG2 cancer cells

Chikusetsusaponin IV and V (CsIV and CsV), two typical oleanolic acid saponins, are mainly derived from the rhizome of Panax japonicus C.A. Mey. To reveal the anti-cancer effect of CsIV and CsV on liver cancer cells, human hepatic cancer cell lines (HepG2) were exposed to these saponins, and various physiological responses of HepG2 were investigated. HepG2 cells were treated with CsIV, CsV and 5-fluorouracil (5-FU). Cell proliferation was measured by CCK-8 assay. The cell cycle arrest, cell apoptosis and intracellular Ca2+ levels were respectively identified by flow cytometry. The mitochondrial membrane potential was detected by fluorescence microscopy. And, the levels of apoptosis-related proteins were analyzed by western blotting. Both CsIV and CsV were demonstrated to inhibit cell viability, and induce cell cycle arrest and apoptosis of HepG2 in a dose-dependent manner. They also enhanced the intracellular Ca2+ level and decreased the mitochondrial membrane potential in HepG2 cells. Furthermore, p53 and p21 were found up-regulated in HepG2 cells treated by CsIV and CsV. The apoptotic proteins, bax, cytochrome c, cleaved caspase-3/-9, were all found activated in HepG2 cells after CsIV and CsV treatment. The anti-apoptotic protein, bcl-2, was significantly down-regulated in all treated HepG2 cells. Our data demonstrated that CsIV and CsV exerted significant cytotoxic effects on HepG2 cells without affecting normal liver cells. And, these chikusetsusaponins, especially for CsIV, showed a potent effect on promoting cell apoptosis in HepG2 cells, which was associated with the activation of p53-mediated apoptosis pathway.

Probing Fluorinated Motifs onto Dual AChE-MAO B Inhibitors: Rational Design, Synthesis, Biological Evaluation, and Early-ADME Studies.

Bioisosteric H/F or CH2OH/CF2H replacement was introduced in coumarin derivatives previously characterized as dual AChE-MAO B inhibitors to probe the effects on both inhibitory potency and drug-likeness. Along with in vitro screening, we investigated early-ADME parameters related to solubility and lipophilicity (Sol7.4, CHI7.4, log D7.4), oral bioavailability and central nervous system (CNS) penetration (PAMPA-HDM and PAMPA-blood-brain barrier (BBB) assays, Caco-2 bidirectional transport study), and metabolic liability (half-lives and clearance in microsomes, inhibition of CYP3A4). Both specific and nonspecific tissue toxicities were determined in SH-SY5Y and HepG2 lines, respectively. Compound 15 bearing a -CF2H motif emerged as a water-soluble, orally bioavailable CNS-permeant potent inhibitor of both human AChE (IC50 = 550 nM) and MAO B (IC50 = 8.2 nM, B/A selectivity > 1200). Moreover, 15 behaved as a safe and metabolically stable neuroprotective agent, devoid of cytochrome liability.

Recurvataside, a new saponin from aerial parts of Mussaenda recurvata

A new glycoside, recurvataside (1) and six known compounds, quinovic acid (2), quinovic acid 28-O-β-D-glucopyranosyl ester (3), 3-O-β-D-glucopyranosylquinovic acid (4), 3-O-β-D-glucopyranosylquinovic acid 28-O-β-D-glucopyranosyl ester (5), pomolic acid (6), and ursolic acid (7) were isolated from aerial parts of Mussaenda recurvata. The structure of compound 1 was identified from its spectroscopic data and by comparison with the literature. Recurvataside represents the first occurrence of δ-oleanolic acid saponin bearing two D-glucose units at C-3 and C-28 in nature. This is the first time δ-oleanane-type saponin reported in the genus Mussaenda. Compounds 1-7 were evaluated the cytotoxicity against two cancer cell lines MCF-7 and HepG2. Among them, only compound 7 exhibited moderate activity against MCF-7 and HepG2 cell lines with IC50 value of 16.97 ± 1.55 and 20.28 ± 1.00 μM, respectively. Compounds 1–7 were also tested for their inhibitory NO production in LPS-stimulated RAW264.7 cells. Compounds 3, 5, and 7 showed significant reduction of nitrite accumulation in LPS-stimulated RAW 264.7 cells with the IC50 values of 8.81 ± 0.48, 13.42 ± 0.84, and 18.37 ± 0.67 μM, respectively.