Acridine Orange/ethidium Bromide Research Articles

Mucilage-assisted fabrication of molybdenum trioxide nanostructures for photothermal ablation of breast cancer cells.

Nanostructures have been used for various biomedical applications due to their optical, antibacterial, magnetic, antioxidant, and biocompatible properties. Cancer is a prevalent disease that severely threatens human life and health. Thus, innovative and effective therapeutic approaches are urgently required for cancer. Photothermal therapy (PTT) is a promising approach to killing cancer cells. In this investigation, we developed a low-cost, simple, green technique to fabricate molybdenum trioxide nanostructures (MNs) using Opuntia ficus-indica mucilage as a template. Moreover, the MNs were functionalized with folic acid (FA) for cancer PTT. The X-ray diffractometer results revealed that the prepared MNs have an orthorhombic crystal phase. The transmission electron microscope image of MNs shows a flake shape with 20-150nm diameter. The cytotoxicity of MNs and FA-conjugated MNs was studied in vitro. These cell viability assay results suggested that fabricated MoO3 nanostructures reduced 25% of cell viability in MCF-7 cells, even at high doses. However, even with high-dose treatment, FA/MNs do not cause significant cell death. Acridine orange/ethidium bromide (AO/EB) staining revealed DNA and chromatin condensation in MCF-7 cells exposed to MNs. Overall, the in vitro study results suggested that FA/MNs have excellent biocompatibility, which applies to biomedical applications. MNs dispersion temperature gradually increases from 26 to 58°C under 808nm laser irradiation. We found significant mortality rates after NIR irradiation in MNs- or FA/MNs-treated MCF-7 cells. These findings suggest that FA/MNs can be used as an effective photothermal agent to treat breast cancer.

Assessment of anticancer properties of cumin seed (Cuminum cyminum) against bone cancer.

Early-life osteosarcoma is associated with severe morbidity and mortality, particularly affecting young children and adults. The present cancer treatment regimen is exceedingly costly, and medications like ifosfamide, doxorubicin, and cisplatin have unneeded negative effects on the body. With the introduction of hyphenated technology to create medications based on plant molecules, the application of ayurvedic medicine as a new dimension (formulation, active ingredients, and nanoparticles) in the modern period is rapidly growing. The primary source of lead compounds for the development of medications for avariety of ailments is plants and their products. Traditionally, Cuminum cyminum (cumin) has been used as medication to treat a variety of illnesses and conditions. The cumin seed was successfully extracted with solvents Hexane, Chloroform, Methanol, Ethanol and Acetone. Following the solvent extraction, the extract residue was assayed in MG63 cells for their anti-proliferative properties. First, we used the [3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide] (MTT) assay to test the extracted residue's cytotoxicity. The results show that hexane extract Half-maximal inhibitory concentration (IC50 86 µG/mL) effciently inhibits cells by causing programmed cell death. Furthermore, using the Acridine orange/ethidium bromide (AO/EB) staining method, the lactate dehydrogenase assay, and the reactive oxygen species assay using the Dichloro-dihydro-fluorescein diacetate (DCHFDA) staining method, we have demonstrated that the hexane extract causes apoptosis in MG63 cells. Furthermore, flow cytometry research revealed that the hexane extract stops the cell cycle in the S phase. In addition, the hexane extract limits colony formation and the migration potential as shown by the scratch wound healing assay. Furthermore, the extract from cumin seeds exhibits remarkable bactericidal properties against infections that are resistant to drugs. Gas chromatography analysis was used to quantitatively determine the hexane and methanolic extract based on the experimental data. The primary chemical components of the extract are revealed by the study, and these help the malignant cells heal. The present study finds that there is scientific validity in using cumin seeds as a novel method of anticancer therapy after undergoing both intrinsic and extrinsic research.

Norcantharidin-Encapsulated C60-Modified Nanomicelles: A Potential Approach to Mitigate Cytotoxicity in Renal Cells and Simultaneously Enhance Anti-Tumor Activity in Hepatocellular Carcinoma Cells.

The objective of this study was to examine the preparation process of DSPE-PEG-C60/NCTD micelles and assess the impact of fullerenol (C60)-modified micelles on the nephrotoxicity and antitumor activity of NCTD. The micelles containing NCTD were prepared using the ultrasonic method and subsequently optimized and characterized. The cytotoxicity of micelles loaded with NCTD was assessed using the CCK-8 method on human hepatoma cell lines HepG2 and BEL-7402, as well as normal cell lines HK-2 and L02. Acridine orange/ethidium bromide (AO/EB) double staining and flow cytometry were employed to assess the impact of NCTD-loaded micelles on the apoptosis of the HK-2 cells and the HepG2 cells. Additionally, JC-1 fluorescence was utilized to quantify the alterations in mitochondrial membrane potential. The generation of reactive oxygen species (ROS) following micelle treatment was determined through 2',7'-dichlorofluorescein diacetate (DCFDA) staining. The particle size distribution of the DSPE-PEG-C60/NCTD micelles was determined to be 91.57 nm (PDI = 0.231). The zeta potential of the micelles was found to be -13.8 mV. The encapsulation efficiency was measured to be 91.9%. The in vitro release behavior of the micelles followed the Higuchi equation. Cellular experiments demonstrated a notable decrease in the toxicity of the C60-modified micelles against the HK-2 cells, accompanied by an augmented inhibitory effect on cancer cells. Compared to the free NCTD group, the DSPE-PEG-C60 micelles exhibited a decreased apoptosis rate (12%) for the HK-2 cell line, lower than the apoptosis rate observed in the NCTD group (36%) at an NCTD concentration of 75 μM. The rate of apoptosis in the HepG2 cells exhibited a significant increase (49%), surpassing the apoptosis rate observed in the NCTD group (24%) at a concentration of 150 μM NCTD. The HK-2 cells exhibited a reduction in intracellular ROS and an increase in mitochondrial membrane potential (ΔψM) upon exposure to C60-modified micelles compared to the NCTD group. The DSPE-PEG-C60/NCTD micelles, as prepared in this study, demonstrated the ability to decrease cytotoxicity and ROS levels in normal renal cells (HK-2) in vitro. Additionally, these micelles showed an enhanced antitumor activity against human hepatocellular carcinoma cells (HepG2, BEL-7402).

SUSTAINED ANTICANCER EFFECT BY NARINGIN-LOADED ZINC OXIDE NANOPARTICLES IN HUMAN LUNG ADENOCARCINOMA A549 CELLS

Objective: This study aimed to design naringin-loaded zinc oxide nanoparticles (Nar-ZnO NPs) and evaluate the formulated nanoparticles (NPs) for their antioxidant and anticancer potential. Methods: Naringin-loaded zinc oxide nanoparticles (Nar-ZnO NPs) were prepared using a modified sol-gel method with ethylenediaminetetraacetic acid (EDTA) as a capping agent. Subsequently, were characterized using dynamic light scattering (DLS), powder X-ray diffraction (PW-XRD), Fourier transform infrared spectroscopy (FT-IR), high-resolution scanning electron microscopy (HR-SEM), and Energy Dispersive X-ray analysis (EDX). Furthermore, the naringin-loaded zinc oxide nanoparticles (Nar-ZnO NPs) were evaluated for their in vitro free radical scavenging activity using antioxidant assays and inhibition of lipid peroxidation potential using the altered thiobarbituric acid-reactive species (TBARS) test. The cytotoxic effect of naringin-loaded zinc oxide nanoparticles (Nar-ZnO NPs) on the non-transformed Vero cell line and lung cancer A549 cell line was investigated using the (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) MTT assay. Apoptosis study was conducted using the Acridine orange/Ethidium bromide (AO/EB) double staining assay, while propidium iodide (PI) stain was utilized to observe apoptotic morphological changes. Results: The prepared naringin-loaded zinc oxide nanoparticles (Nar-ZnO NPs) were smooth and hexagonal, with an average particle size of 500 nm. The antioxidant assays demonstrated that the naringin-loaded zinc oxide nanoparticles (Nar-ZnO NPs) and ascorbic acid exhibited comparable free radical scavenging and inhibition of lipid peroxidation activity. In MTT assay, the naringin-loaded zinc oxide nanoparticles (Nar-ZnO NPs) displayed IC₅₀ values of 1014.05 µg/ml for Vero cell lines and 317.51 µg/ml for A549 cells, highlighting their influence on cell viability. Remarkably, treatment of A549 cells with the Nar-ZnO NPs resulted in dose-dependent apoptotic morphological changes, as observed through (AO/EB) double staining assay and propidium iodide (PI) stain. Conclusion: The study findings revealed that the naringin-loaded zinc oxide nanoparticles (Nar-ZnO NPs) displayed dose-dependent free radical scavenging activity, significant inhibition of lipid peroxidation, and notable anticancer properties against A549 cells.

Antitumor activity of cordycepin in murine malignant tumor cell line: An in vitro and in silico study

Cordycepin, also known as 3′-deoxyadenosine, is a substance similar to adenosine found in the entomopathogenic fungus Cordyceps militaris. This fungus is recognized for its remarkable therapeutic properties. This study seeks to investigate the antitumor activity of cordycepin in Dalton's lymphoma (DL) malignant cancer cell line using cytotoxicity and apoptosis as key parameters. Molecular docking (MD) and molecular dynamic simulation (MDS) were used to further validate the wet lab findings. Cytotoxicity study revealed that cordycepin induced significant cytotoxicity (P ≤ 0.05) in the DL cell line after 24 h of treatment. The efficacy of cordycepin as an apoptotic agent was substantiated by the highly promising results obtained through the apoptotic assay, utilizing the acridine orange/ethidium bromide (AO/EB) dual staining method. Cordycepin interacted with the active sites of antiapoptotic target proteins (BCL-XL, SURVIVIN, BCL-2, BCL-B, HSP90, and IAP1), as demonstrated by MD study that corroborated the findings of cytotoxicity and apoptotic assays. The comparative examination of docking outcomes indicated that cordycepin exhibits higher affinity for HSP90 in comparison to other targets. Molecular dynamics simulation for 120 ns further validated the findings of MD as cordycepin remained bound with HSP90 throughout the entire simulation time (120 ns) with favourable binding energy.

Mitochondriotropic agents conjugated with NSAIDs through metal ions against breast cancer cells

Two copper(I) polymorphs of formula [Cu(SALH)(TPP)3] (1a and 1b) were prepared by the conjugation of the Non-Steroidal Anti-Inflammatory Drug (NSAID) salicylic acid (SALH2) with the mitochondriotropic agent triphenylphosphine (TPP) via metal ion. For comparison, the isomorph [Ag(SALH)(TPP)3] (2) was prepared. The conjugates 1a, 1b and 2 were characterized by melting point (m.p.), Attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy, Ultraviolet-Visible (UV–Vis) spectroscopy and nuclear magnetic resonance (1H NMR). The crystal structures of 1a, 1b and 2 were confirmed by X-ray diffraction crystallography (XRD). The ex vivo binding affinity of 1–2 towards CT (calf thymus)-DNA was studied by UV, fluorescence, viscosity and DNA Thermal Denaturation studies. Their inhibitory activity against lipoxygenase (LOX) (an enzyme which is mainly located in the mitochondrion) was determined. The in vitro activity of 1–2 was evaluated against human breast cancer cell lines MCF-7 (hormone depended (HD)) and MDA-MB 281 (hormone independent (HI)) cells. Compounds 1–2 inhibit stronger than cisplatin the cancerous cells. The molecular mechanism of action of 1–2 was suspected by the MCF-7 cells morphology and confirmed by DNA fragmentation, Acridine Orange/Ethidium Bromide (AO/EB) Staining and mitochondrial membrane permeabilization tests.

Synthesis, Characterisation, Invitro anticancer activity of Diazo Derivatives of 1,3,4-oxadiazole

This study delves into the comprehensive evaluation of newly synthesized Diazo Derivatives of 1,3,4-oxadiazole compounds, including Aniline, Ortho nitro aniline, Meta nitro aniline, Ortho anisidine, Meta anisidine, 3-chloro aniline, 2,6-dimethyl aniline, 2,5- dicloro aniline, and 3,5- dichloro aniline, aiming to unravel their potential as antioxidants and anticancer agents. The assessment of antioxidant potential involved the utilization of the DPPH (2,2-diphenyl-1-picrylhydrazyl) assay. The cytotoxic activity of these compounds was assessed against MCF-7 breast cancer cells using the MTT assay. Morphological assessments using acridine orange/ethidium bromide (AO/EB) staining. Results were expressed as IC50 values, with 2,5-dichloro aniline and 3-chloro aniline demonstrating remarkable antioxidant activity, each with an IC50 value of 62.5 µg/mL. Several other compounds also exhibited significant antioxidant properties, albeit with slightly higher IC50 values. Notably, Ortho anisidine and Ortho nitro aniline displayed no substantial inhibitory activity against oxidative stress. The cytotoxic activity of these compounds was assessed against MCF-7 breast cancer cells using the MTT assay, with IC50 values ranging from 62.5 to 1000 µg/mL. 2,5-dichloro aniline and 3-chloro aniline emerged as potent inhibitors, both achieving an IC50 value of 62.5 µg/mL. Aniline, Meta nitro aniline, Meta anisidine, 2,6-dimethyl aniline, and 3,5-dichloro aniline exhibited intermediate inhibitory activity, while Ortho anisidine and Ortho nitro aniline showed no significant inhibition. Morphological assessments using acridine orange/ethidium bromide (AO/EB) staining revealed diverse cellular responses induced by 2,5-dichloro aniline and 3-chloro aniline treatment. These changes align with well-established apoptotic processes observed in therapeutically treated cell lines. This research contributes valuable insights into the antioxidant and anticancer potential of Diazo Derivatives of 1,3,4-oxadiazole compounds, with 2,5-dichloro aniline and 3-chloro aniline emerging as promising candidates for further exploration in combatting oxidative stress and breast cancer. These findings underscore the importance of antioxidants in cancer prevention and the potential therapeutic applications of these compounds.
 Keywords: Diazo Derivatives of 1,3,4-oxadiazole, 2,5-dichloro aniline, 3-chloro aniline, DPPH, MTT assay, MCF-7

Ipomoea carnea alkaloid extract vs swainsonine: A comparative study on cytotoxic activity against glial cells

The consumption of Ipomoea carnea produces a neurological syndrome in animals. The toxic principles of I. carnea are the alkaloids swainsonine (SW) and calystegines B1, B2, B3 and C1. In this study, we investigated the cytotoxicity of an alkaloid extract of Ipomoea carnea (AEE) and natural swainsonine (SW) isolated from Astragalus lentiginosus (25–1000 μM of SW) for 48 h in a glioma cell line. Although the natural SW did not induce any changes in cell viability, the AEE exhibited a dose dependent cytotoxic effect and release of lactate dehydrogenase (LDH) indicative of cytolysis. In order to evaluate the morphological changes involved, cells were examined using phase contrast and fluorescence microscopy with acridine orange-ethidium bromide staining. The AEE caused a cell death compatible with necrosis, whereas exposure to 1000 μM of SW resulted in cytoplasmic vacuolation. Immunocytochemical studies revealed that astrocytes treated with 150 μM of AEE from I. carnea or 1000 μM of SW exhibited morphological characteristics of cell activation. These findings suggest that swainsonine would not be the only component present in the AEE of I. carnea responsible for in vitro cytotoxicity. Calystegines might also play a role in acting synergistically and triggering cell death through necrosis.

Hydromethanolic leaves extract of Dalbergia sissoo Roxb. ex DC. induces apoptosis in lung adenocarcinoma cells

Plants have a wide range of active secondary metabolites that are frequently utilized to treat cancer. For the research, Dalbergia sissoo Roxb. ex DC. leaves (DS) were hydromethanolically extracted, and their phytochemical content was determined. Total phenolics and flavonoids were quantified along with the measurement of in vitro antioxidant and cytotoxic activities. The bioactive components in the extract were identified with the use of Gas Chromatography with High-Resolution Mass Spectrometry (GC-HRMS). The crude extract contained 177.500 ± 0.019 mg/ml of flavonoids and 296.122 ± 0.002 mg/ml of Gallic Acid Equivalent (GAE) phenolics. Moreover, plant crude extract showed significant 2,2-diphenyl-1-picrylhydrazyl (DPPH) activity (IC50, 14.06 ± 0.18 μg/ml), 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) [ABTS] activity (IC50, 25.97 ± 1.04 μg/ml), superoxide scavenging activity (IC50, 149.91 ± 0.39 μg/ml), and hydrogen peroxide scavenging activity (IC50, 133.37 ± 2.30 μg/ml). The thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) methods were used to confirm the presence of phenolic compounds. DS effectively scavenges nitric oxide. The crude extract of DS exhibited good cytotoxic effects on lung cancer cells (A549) with an IC50 value of 90.56 ± 2.32 (μg/ml), and less toxicity on normal lung cells (WI-38) with an IC50 value of 381.10 ± 1.58 (μg/ml). IC50 value of methotrexate (the standard drug) was 10.20 ± 1.82 μg/ml on A549 cells and 26.21 ± 1.14 μg/ml on WI-38 cells. Various staining techniques [4, 6-diamidino-2-phenylindole (DAPI), Acridine orange/Ethidium Bromide (AO/EB) staining, Giemsa staining] were used to determine the plausible mechanism of DS to induce apoptosis. The inhibiting mechanism of the crude extract was further demonstrated by the clonogenic assay and qualitative and quantitative measurements of Reactive Oxygen Species (ROS). Real-time polymerase chain reaction (RT-PCR) analysis revealed the induction of apoptosis in A549 cells through the activation of caspases 9 and 3 together with TRAIL receptors. In a nutshell, hydromethanolic extract of DS resulted in distinct apoptotic morphological alterations, ROS production, the initiation of apoptosis via activating the TRAIL receptors, caspase 9 and 3, and the suppression of colony formation in A549 cells.

Identification of highly potent 2,4-diarylaminopyrimidine analogs of a typical piperidinyl-4-ol moiety as promising antitumor ALK inhibitors.

In light of the cocrystal structure of ceritinib with anaplastic lymphoma kinase(ALK)WT protein, a series of novel 2,4-diarylaminopyrimidine analogs(L1-L25) bearing a typical piperidinyl-4-ol moiety were designed and synthesized with improved biological and physicochemical properties. Satisfyingly, most compounds demonstrated moderate to excellent antitumor effects with IC50 values below 5 μM on ALK-positive Karpas299 and H2228 cells. In particular, L6 bearing the 1-(6-methoxy-pyridin-2-yl)-4-(morpholinomethyl)piperidinyl-4-ol moiety was detected as the optimal compound against ALK-dependent cell lines of Karpas299 (0.017 μM) and H2228 cells (0.052 μM), in company with encouraging ALK enzyme inhibition (ALKWT , IC50 = 1.8 nM). In addition, L6 was also capable of inhibiting ALK-resistant mutations, including ALKL1196M (3.9 nM) and ALKG1202R (5.2 nM). Remarkably, L6 typically repressed colony formation and migration of H2228 cells in a dose-dependent manner. Meanwhile, acridine orange-ethidium bromide staining analysis indicated that the proapoptotic effect of L6 was better than that of ceritinib at the same concentration (50 nM). Ultimately, the binding patterns of L6 to ALKWT and ALKG1202R were ideally established, which further confirmed the structural basis in accordance with the structure-activity relationship analysis.

1,2-bis(2,4,6-tribromophenoxy) ethane, a novel brominated flame retardant, disrupts intestinal barrier function via the IRX3/NOS2 axis in rat small intestine

Novel brominated flame retardants are widely used in electronics, textiles, furniture, and other products; they can enter the human body through ingestion and respiration and cause harm to the human body, and have been proven to have potential biological toxicity and accumulation effects. 1,2-bis(2,4,6-tribromophenoxy) ethane (BTBPE) is a widely used novel brominated flame retardant; however, there is a lack of research on its mechanism of toxicity, particularly that of intestinal toxicity. Currently, studies on the functionality of iroquois homeobox 3 (IRX3) are extremely limited. In our study, BTBPE was administered to Sprague-Dawley (SD) rats and rat small intestinal crypt epithelial cells (IEC6 cells) in vivo and in vitro, respectively, and hematoxylin and eosin (HE), immunohistochemical, Alcian blue-periodic acid-Schiff (AB-PAS), CCK8, acridine orange/ethidium bromide (AO/EB), fluorescent probes, qPCR, western blotting, and immunofluorescence analyses were performed. To explore the damage mechanism of BTBPE, we used siRNA to silence IRX3 and iNOs-IN-1 (yeast extract-peptone-wheat; YPW) to inhibit nitric oxide synthase 2 (NOS2). The results showed that BTBPE exposure caused inflammation and necroptosis in the jejunum and ileum, as well as destruction of the tight junctions and mucus layer. Moreover, BTBPE activated the IRX3/NOS2 axis both in vivo and in vitro. Silencing IRX3 or inhibiting NOS2 inhibits necroptosis and restores tight junctions in IEC6 cells. In conclusion, our study found that in the jejunum, ileum, and IEC6 cells, BTBPE exposure caused necroptosis and tight junction destruction by activating the IRX3/NOS2 axis. Blocking the IRX3/NOS2 axis can effectively inhibit necroptosis and restore tight junction. In addition, BTBPE exposure caused inflammation and loss of the mucous layer in the jejunum and ileum. Our study is the first to explore the mechanism of intestinal damage caused by BTBPE exposure and to discover new biological functions regulated by the IRX3/NOS2 axis, providing new research directions for necroptosis and tight junctions.

Antioxidant and anti-HepG2 cell activities of a novel bioactive peptide from cowhide collagen in vitro

In the present study, the antioxidant and anti-human liver cancer (HepG2) cells effects of bioactive peptides from cowhide collagen (BPCC) were evaluated. BPCC exhibited significant scavenging effect on 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals ((60.09 ± 3.51)%), 2,2’-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radicals ((77.40 ± 3.10)%) and hydroxyl radicals ((56.00 ± 2.30)%) as well as strong reducing power (0.320 ± 0.025). Meanwhile, BPCC effectively protected biomacromolecules including proteins, lipids and DNA from oxidative damage induced by Cu2+/H2O2 and 2,2’-azobis(2-methylpropionamidine) dihydrochloride (AAPH). Moreover, BPCC significantly inhibited cell viability of HepG2 cells in a dose-dependent manner with an estimated IC50 of 7.61 mg/mL. The results of 4’,6-diamidino-2-phenylindole (DAPI) and acridine orange/ethidium bromide (AO/EB) staining demonstrated the apoptotic morphological changes and cell mediated death in BPCC treated HepG2 cells. In addition, BPCC induced decrease of mitochondrial membrane potential (MMP) in HepG2 cells. Therefore, the present finding proved that BPCC encompasses significant antioxidant activity and anticancer property on HepG2 cells and can be used as alternative food antioxidants for cancer prevention benefits.

In vitro anticancer efficacy of Cyperus rotundus (L.) on breast adenocarcinoma cells via the induction of DNA fragmentation and apoptosis

Introduction and Aim: Cyperus rotundus (L.), commonly known as nutgrass, has been in use in Indian medicine traditional system against several ailments. This investigation was performed to assess the in vitro antineoplastic activity of extracts prepared from water and ethanol with C. rotundus rhizomes. Materials and Methods: Toxic effects induced by the C. rotundus extracts on MCF-7 breast adenocarcinoma cells was evaluated using Sulphorhodamine- B (SRB) Assay, Trypan blue assay, Hoechst nuclear staining assay, Acridine Orange-Ethidium bromide (AO-EB) differential fluorescence staining assay and DNA fragmentation assays. Results: The results of SRB assay indicated the IC50 value of aqueous extract as 510.887 µg/mL and that of ethanol extract as 122.98 µg/mL. Trypan blue assay also indicated the similar results, wherein ethanol extract exerted higher cytotoxicity compared to the aqueous extract. MCF-7 cells treated with aqueous and ethanol extracts of C. rotundus rhizomes were exposed to Hoechst staining and AO-EB nuclear staining assays, wherein and ethanol extract induced clear apoptotic bodies in the tested cells. Further, the cells were treated with the ethanol extract and DNA fragmentation was confirmed by DNA ladder assay. MCF-7 cells administered with ethanol extract were subjected to DNA fragmentation assay for assessing the apoptosis induction nature of the extract. Conclusion: Results from our study indicate a promising approach into the possibility of developing C. rotundus ethanol extract into an effective anticancer moiety after appropriate pre-clinical and clinical validations.

Screening of Cu4 O3 NPs efficacy and its anticancer potential against cervical cancer.

Cu4 O3 is the least explored copper oxide, and its nanoformulation is anticipated to have important therapeutic potential especially against cancer. The current study aimed to biosynthesize Cu4 O3 nanoparticles(NPs) using an aqueous extract of pumpkin seeds and evaluate its antiproliferative efficacy against cervical cells after screening on different cancer cell lines.The obtained NPs were characterized by different spectroscopic analyses, such as UV-vis, thermogravimetric,energy dispersive X-ray,and Fourier-transform infrared spectroscopy (FTIR). In addition, high-resolution transmission electron microscopes (HR-TEM) were used to observe the morphology of the biosynthesized NPs. The UV-vis spectra showed a peak at around 332 nm, confirming the formation of Cu4 O3 NPs. Moreover, FTIR and TAG analyses identified the presence of various bioactive phytoconstituents that might have worked as capping and stabilization agents and comparative stable NPs at very high temperatures, respectively. The HR-TEM data showed the spherical shape of Cu4 O3 NPs in the range of 100 nm. The Cu4 O3 NPs was screened on three different cancer cell lines viz., Hela, MDA-MB-231, and HCT-116 using cytotoxicity(MTT) reduction assay. In addition, Vero was taken as a normal epithelial (control) cell. The high responsive cell line in terms of least IC50 was further assessed for its anticancer potential using a battery of biological tests, including morphological alterations, induction of apoptosis/ROS generation, regulation of mitochondrial membrane potential (MMP), and suppression of cell adhesion/migration. Vero cells (control) showed a slight decline in % cell viability even at the highest tested Cu4 O3 NPs concentration. However, all the studied cancer cells viz., MDA-MB-231, HCT 116, and HeLa cells showed a dose-dependent decline in cell viability after the treatment with Cu4 O3 NPs with a calculated IC50 value of 10, 11, and 7.2 µg/mL, respectively. Based on the above data, Hela cells were chosen for further studies, that showed induction of apoptosis from 3.5 to 9-folds by three different staining techniques acridine orange/ethidium bromide (AO/EB), 4',6-diamidino-2-phenylindole (DAPI), and propidium iodide (PI). The enhanced production of reactive oxygen species(>3.5-fold), modulation in MMP, and suppression of cell adhesion/migration were observed in the cells treated with Cu4 O3 NPs.The current study obtained the significant antiproliferative potential of Cu4 O3 NPs against the cervical cancer cell line, which needs to be confirmed further in a suitable in vivo model. Based on our results, we also recommend the green-based, eco-friendly, and cost-effective alternative method for synthesizing novel nanoformulation.

Zinc Oxide Nanoparticles Produced Sustainably Utilizing Solanum xanthocarpum Leaf Extracts and Their Anti-Neoplastic Effects on Colon Cancer Cells (HT-29)

Zinc oxide nanoparticles (ZnO-NPs) were synthesized in this study using Solanum xanthocarpum (Sx) leaf extracts via a green method. The nanomaterial, ZnO-NPs-Sx, was prepared using zinc acetate dehydrate solution followed by purification. The nanofabricate size and structural properties were investigated using UV-V Spectroscopy, X-ray diffraction, energy dispersive X-ray, and Fourier transform infrared spectroscopy (FT-IR), as well as scanning electron microscopy (SEM). The anti-neoplastic properties of nanomaterial against the colon cancer cells (HT-29) were measured using MTT assay and the apoptotic biomarkers via staining techniques (i.e., acridine orange/ethidium bromide (AO/EB), 4′,6-diamidino-2-phenylindole (DAPI) and Propidium iodide (PI staining). The nanomaterial induced reactive oxygen species ROS and apoptosis by stimulating cell signaling biomarkers: caspase-3, -8 and -9. Staining showed proportionately higher number of dead cells with nuclear damage and chromatin condensation to confirm the nanomaterial induced apoptosis in HT-29 cells. At low concentrations, the nanomaterial did not elicit adverse cellular changes, and was less toxic towards HT-29 cells. Thus, the bio-synthesized nanofabricate, can be used ZnO-NPs-Sx in nanomedicine to improve current treatments towards cancer cells.

Antiproliferative effect of methanolic extract of Tamarindus indica in C127I cell line

Investigation of natural bioactive compounds that possess antioxidant and anticancer properties have drawn interest in scientific and industrial perspectives. The current study was conducted to assess the antiproliferative action of methanolic extract of seed coat of Tamarindus indica in C127I mouse mammary tumour cell line. 3-(4,5-dimethyl thazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay is performed to assess the cytotoxicity of methanolic extracts of T. indica in the C127I cell line at concentrations of 320, 160, 80, 40, 20, and 10 µg/mL. The half maximal inhibitory concentration (IC50) was then calculated using Graph Pad Prism 5.0. Doxorubicin was used as positive control. After determining IC50, the cells were treated with extract of T. indica at half, IC50 and double IC50 concentrations after being seeded at a concentration of 1x105 cells/mL in 6 well plates. After 24 hours the cells were trypsinised and stained with acridine orange-ethidium bromide (AOEB) to assess the morphology of apoptosis. When the cells were exposed to various doses of the extract, a dose-dependent decrease in cell viability was observed, and the IC50 value of T. indica was estimated to be 13 µg/ml. In the control cells, AO/EB labelling revealed proliferating cells with green fluorescence, but the T. indica extract-treated cells displayed dose dependent shift from orange to red fluorescence. In conclusion, methanolic extract of Tamarindus indica was found to possess antiproliferative effect in C127I cell line comparable to doxorubicin.

Anti–MUC1 nanobody can synergize the Tamoxifen and Herceptin effects on breast cancer cells by inducing ER, PR and HER2 overexpression

IntroductionOne of the most pressing concerns associated with breast cancer-targeted therapies is resistance to Tamoxifen and Herceptin. Such drug resistance is usually characterized by reduced expression of certain cell surface receptors. Some biological regimens can induce perceptible overexpression of these receptors in favor of drug responsiveness. Material and MethodsIn this research, drug-responsive MCF-7 and SKBR-3, along with drug-resistant MCF-7R (Tamoxifen resistant) and JIMT-1 (Herceptin resistant) breast cancer cell lines in 2D and 3D cultures were exposed to anti-MUC1 nanobody and then assessed for their ER, PR, and HER2 gene and protein expression using qRT-PCR and immunofluorescent staining analyses. Cell viability and the synergistic relationships of combination treatments were determined with MTT assay followed by CompuSyn software. Apoptotic cells were evaluated with Annexin V/propidium Iodide (PI) and acridine orange/ethidium bromide (AO/EB) staining. ResultsAnti-MUC1 exposure elevated the expression levels of ER (42 folds), PR (18.5 folds), and HER2 (4.7 folds). As a result of co-treatment, the IC50 levels for Tamoxifen and Herceptin were reduced by up to 10 and 3 folds, respectively. MCF-7R cells responded positively to Tamoxifen, as evidenced by a 5-fold reduction in the IC50 and enhanced apoptosis. ConclusionThe ER, PR, and HER2 overexpression after MUC1 blocking could signal drug hypersensitization and facilitate drug resistance management.

Endoplasmic Reticulum Stress-Mediated Apoptosis Induced by VR12684 Isolated from Mallotus spodocarpus in Cholangiocarcinoma Cell Line.

Cholangiocarcinoma (CCA) is a poor prognosis of a malignant tumor that has been unresponsive to conventional chemotherapeutic agents. Effective and novel therapeutic agents are urgently needed. VR12684 (isolated from Mallotus spodocarpus) has been reported to exhibit growth inhibitory activities in cancer cell lines. The present study investigated the growth inhibitory mechanisms of this compound in a human CCA cell line (KKU-M156). The effects of VR12684 on anti‑proliferation, cell cycle arrest and apoptosis induction in CCA cells were demonstrated by SRB assay, flow cytometry, acridine orange/ethidium bromide (AO/EB) staining and western blot analysis. Treatment with VR12684 decreased cell proliferation in a dose- and time-dependent manner in the KKU-M156 cell line. VR12684 induced cell cycle arrest in the G2 phase in KKU-M156 through down-regulation of cyclin B1 and Cdk1 and up-regulation of p21, p27 and p53 levels. VR12684 induced mitochondria-mediated apoptosis by increasing DNA fragmentation, the Bax/BCL-2 ratio and AIF, and decreasing survivin with subsequent activation of caspase-9 and -3. This compound could induce apoptosis through the endoplasmic reticulum (ER) stress-mediated pathway by up-regulation of GRP78, IRE1α and GADD153 levels leading to down-regulation of Bcl-2 and activation of calpain-1, caspase-7 and -12. These results suggested that VR12684 inhibited KKU-M-156 cell growth by way of cell cycle arrest and induction of apoptosis, at least in part, through the mitochondria- and ER-associated intrinsic pathways. Such compounds warrant evaluation as a candidate for the treatment of human CCA.

Chitosan-encapsulated nickel oxide, tin dioxide, and farnesol nanoparticles: Antimicrobial and anticancer properties in breast cancer cells

Breast cancer is the most frequent cancer in women; however, it is curable in most cases (up to 80 %) when detected and treated at an early non-metastatic stage. Nanotechnology has led to the development of potential chemotherapeutic techniques, particularly for tumor treatment. Nanotechnology has therapeutic and pharmaceutical applications. Chitosan, a natural polymer derived from chitin, has been extensively studied for its potential applications in a wide range of fields. This includes medicine for its anticancer properties. In the present study, Chitosan-encapsulated-NiO-TiO2-Farnesol hybrid nanomaterials (CNTF HNMs) were synthesized and characterized using several techniques, including electron microscopy (TEM, FE-SEM), spectroscopy (UV–visible [UV–Vis], Fourier Transform Infrared [FT-IR] spectroscopy, and photoluminescence [PL]), energy-dispersive X-ray spectroscopy (EDX) composition analysis, X-ray diffraction, and dynamic light scattering (DLS) analyses. With an estimated average crystallite size of 34.8 nm, the face-cantered cubic crystalline structure of the CNTF HNMs is identified. Cell viability assay by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), DAPI (4′,6-diamidino-2-phenylindole) staining, dual AO/EtBr (Acridine Orange/ Ethidium bromide), JC-1 (5,5,6,6′-tetrachloro-1,1′,3,3′ tetraethylbenzimi-dazoylcarbocyanine iodide), DCFH-DA (Dichloro-dihydro-fluorescein diacetate), Annexin V-FITC (Fluorescein isothiocyanate) /PI (Propidium Iodide), and cell cycle study was used to assess the ability of nanoparticles (NPs) to kill MDA-MB-231 cells. The CNTF HNMs had high antibacterial effectiveness against multi-drug resistant extended-spectrum beta-lactamases (ESBL)-producing gram-negative bacterial pathogens and reference strains. The findings suggest that NPs increased the number of reactive oxygen species (ROS), changed the Δψm, and initiated apoptosis. There is enormous potential for CNTF HNMs as both antibacterial and anticancer agents.

CDH1 overexpression sensitizes TRAIL resistant breast cancer cells towards rhTRAIL induced apoptosis.

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is well known for its unique ability to induce apoptosis in cancer cells but not normal cells. However, a subpopulation of cancer cells exist that does not respond to toxic doses of TRAIL. In this study, we aimed to identify key factors regulating TRAIL resistance in breast cancer. rhTRAIL (recombinant human TRAIL) resistant cells (TR) isolated from TRAIL sensitive MDA-MB-231 parental cells (TS) were confirmed using trypan blue assay, cell viability assay and AO/EtBr (acridine orange/ethidium bromide) staining. Microarray was performed followed by analysis using DAVID and Cytoscape bioinformatics software to identify the candidate hub gene. Gene expression of the candidate gene was confirmed using real-time PCR and western blot. Candidate gene was overexpressed via transient transfection to identify its significance in the context of rhTRAIL. Breast cancer patient data was obtained from The Cancer Genome Atlas (TCGA) database. Whole transcriptome analysis identified 4907 differentially expressed genes (DEGs) between TS and TR cells. CDH1 was identified as the candidate hub gene, with 18-degree centrality. We further observed CDH1 protein to be downregulated, overexpression of which increased apoptosis in TR cells after rhTRAIL treatment. TCGA patient data analysis also showed CDH1 mRNA to be low in TRAIL resistant patient group compared to TRAIL sensitive group. CDH1 overexpression sensitizes TR cells towards rhTRAIL induced apoptosis. Therefore, we can hypothesize that CDH1 expression should be taken into account while performing TRAIL therapy in breast cancer.