Acridine Orange/ethidium Bromide Research Articles

Design, synthesis and biological evaluation of novel N-(3-amino-4-methoxyphenyl)acrylamide derivatives as selective EGFRL858R/T790M kinase inhibitors

On the basis of N-(3-amino-4-methoxyphenyl)acrylamide scaffold, a series of novel compounds containing 3-substitutional-1-methyl-1H-indole, 2-substitutional pyrrole or thiophene moieties were synthesized and their in vitro antiproliferation activities against A549 and H1975 cell lines were evaluated. The results indicated that most of the compounds showed moderate to excellent antitumor activities. Especially, compounds 9a (A549 IC50 = 1.96 μM, H1975 IC50 = 0.095 μM), 17i (A549 IC50 = 4.17 μM, H1975 IC50 = 0.052 μM), 17j (A549 IC50 = 1.67 μM, H1975 IC50 = 0.061 μM) exhibited comparable antitumor activities and selectivity ratios compared to the positive control osimertinib (A549 IC50 = 2.91 μM, H1975 IC50 = 0.064 μM). In vitro inhibitory activities against EGFR kinases containing different mutations were also tested. Compound 17i showed remarkable inhibitory activity (with IC50 value of 1.7 nM) to EGFRL858R/T790M kinase and selectivity (22-folds compared to EGFRWT kinase). Furthermore, acridine orange/ethidium bromide (AO/EB) staining assay, cell apoptosis assay, cell cycle distribution assay and wound-healing assay of the compounds 9a and 17i were performed on H1975 cell line. The results showed dose-dependent activities of the induction of apoptosis, G0/G1-phase arrestation and inhibition of migration, which were similar to the positive control osimertinib. Additionally, molecular docking analysis was performed to seek the possible binding mode between the selected compounds (9a, 17i-17j) and EGFRL858R/T790M kinase. The results demonstrated that compound 17i is a promising candidate and worth further study.

Anticancer Activity of Periplanetasin-5, an Antimicrobial Peptide from the Cockroach Periplaneta americana.

Cockroaches live in places where various pathogens exist and thus are more likely to use antimicrobial compounds to defend against pathogen intrusions. We previously performed an in silico analysis of the Periplaneta americana transcriptome and detected periplanetasin-5 using an in silico antimicrobial peptide prediction method. In this study, we investigated whether periplanetasin-5 has anticancer activity against the human leukemia cell line K562. Cell growth and survival of K562 cells treated with periplanetasin-5 were decreased in a dose-dependent manner. By using flow cytometric analysis, acridine orange/ethidium bromide (AO/EB) staining and DNA fragmentation, we found that periplanetasin-5 induced apoptotic and necrotic cell death in leukemia cells. In addition, these events were associated with increased levels of the pro-apoptotic proteins Fas and cytochrome c and reduced levels of the anti-apoptotic protein Bcl-2. Periplanetasin-5 induces the cleavage of pro-caspase-9, pro-caspase-8, pro-caspase-3, and poly (ADP-ribose) polymerase (PARP). The above data suggest that periplanetasin-5 induces apoptosis via both the intrinsic and extrinsic pathways. Moreover, caspase-related apoptosis was further confirmed by using the caspase inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]- fluoromethylketone (Z-VAD-FMK), which reversed the periplanetasin-5-induced reduction in cell viability. In conclusion, periplanetasin-5 caused apoptosis in leukemia cells, suggesting its potential utility as an anticancer therapeutic agent.

Prodigiosin production from Serratia marcescens strain CSK and their antioxidant, antibacterial, cytotoxic effect and in silico study of caspase-3 apoptotic protein.

The present study emphasizes the production and optimization of prodigiosin (PG) pigment from Serratia marcescens strain CSK, which was isolated from Shevaroy Hills, Salem district, Tamil Nadu, India. The response surface methodology analysis was applied for the optimization process of PG production. The maximum production of PG (2950 mg/L) was obtained at pH 7.0 with the addition of tryptophan (4.0 g/L) and sucrose (3.0 g/L) with 60 h of incubation. Further, the PG was characterized using high-performance liquid chromatography, Fourier-transform infrared spectroscopy, and gas chromatography-mass spectrometry. The purified PG exhibited strong antioxidant and antibacterial activities. Also, PG's cytotoxic effects against human breast cancer (MCF-7) cells were observed through acridine orange-ethidium bromide (AO-EB) and Hoechst staining. Molecular dockingstudies revealed that PG could bind positively to the caspase-3 (breast cancer protein 1RE1) binding site with a binding energy score of 17.37 kcal/mol. Overall, the novel PG was found to be an anticancer drug for potential applications in the pharmaceutical industry.

Cytotoxic activity of Indigofera aspalathoides (Vahl.) extracts in cervical cancer (HeLa) cells: Ascorbic acid adjuvant treatment enhances the activity

The folkloric use of Indigofera aspalathoides (Vahl) in anti-inflammatory and anti-cancer treatment dates back to circa 300 BC. However, reports on scientific validation for folkloric usage are not available yet. The objective of the study is to assess the cytotoxic potential of I. aspalathoides and to provide a rationale for its ethnopharmacological use with lime juice (ascorbic acid) as an adjuvant. Using Soxhlet, the active principles of I. aspalathoides were extracted using methanol and ethanol as solvent and studied using gas chromatography coupled with mass spectrometry (GC–MS). We assessed the in vitro antioxidant efficacy by studying the DPPH radical scavenging activity, ferrous reducing antioxidant capacity (FRAC), and ABTS cation radical scavenging activity. Using MTT assay, the cytotoxic activity of SV-ME and EE against human cervical cancer (HeLa) and normal murine fibroblast (L929) cells were evaluated. The cytotoxicity and apoptosis induction was studied by dual acridine orange/ethidium bromide (AO/EB) fluorescent probe and Hoechst dye. Mitochondria membrane potential (Δψm) was assessed using JC1. Ascorbic acid (AA) additive/synergism was studied by co-treating HeLa cells with AA along with SV-ME and SV-EE at IC50 concentration. SV-ME and EE exert higher DPPH radical scavenging at 200 μg/mL. Both the extracts showed effective ABTS radical cation scavenging activity and ferrous reducing antioxidant capacity. Further, SV-ME and SV-EE treatment affected the cell viability and induced apoptosis, reduced cell viability with a concomitant increase in nuclear condensation and DNA fragmentation in a dose-dependent manner in cancer (HeLa) cells, but not in normal (L929) cells. Both the extracts increased the mitochondria membrane potential (Δψm) depolarization and altered the cellular redox state in HeLa cells. In addition, adjuvant ascorbic acid (AA) treatment increased the cytotoxic efficacy of the SV-ME and SV-EE in HeLa cells. I. aspalathoides used in the Siddha medicine for antitumor treatment have a very potent cytotoxic effect against cervical cancer cells (HeLa) in vitro. The cytotoxic and antitumor effect may have plausible involvement of Nrf2-ARE signaling pathway by restoration of cellular redox homeostasis.

Anticancer Activities of <i>Phlogacanthus pulcherrimus </i>T. Anderson Leaves Extract on MCF-7 Breast Cancer Cells

Phlogacanthus pulcherrimus T. Anderson (PPT) is an edible plant found in the northern and northeastern regions of Thailand. There is limited information about the anti-breast cancer activity of the ethanolic leaf extract of PPT. This study aimed to evaluate the effects of an ethanolic leaf extract of PPT on MCF-7 breast cancer cell lines. The biological effects, including cytotoxicity, cell apoptosis, colony formation, reactive oxygen species (ROS) formation and cell migration, were determined by a means of sulforhodamine B (SRB), acridine orange/ethidium bromide (AO/EB) staining, a clonogenic assay, flow cytometry and a scratch wound healing assay, respectively. The results demonstrated that the PPT extract showed cytotoxic on MCF-7 cells in a dose-dependent manner with 50% inhibitory concentration (IC50) values of 119.9 ± 12.1 and 51.3 ± 4.7 μg/mL at 24 h and 48 h incubation, respectively. In addition, the extract exhibited cell apoptosis in a dose-dependent manner when used at a concentration of 50–100 μg/mL and inhibited colony formation with an IC50 value of 26.0 ± 2.0 μg/mL when compared with the control group. The extract induced ROS formation in a dose-dependent manner when used at a concentration of 50–100 μg/mL. The extract suppressed MCF-7 cell migration, with significant effect at 25 μg/mL. These results indicate that PPT ethanolic leaf extract has an anticancer activity against MCF-7 breast cancer cells and may be useful for prevention and treatment of breast cancer.

Anti-colon Cancer Activity of Parkia javanica (Lamk.) Merr. Bark Extract: An In-vitro Study

The Methanolic extract of Parkia javanica bark (MEPJB) was examined in vitro to see whether it has any cytotoxic or chemotherapeutic potential against the colon cancer cell lines HCT116 and SW480. Cell viability or proliferation assay and migration assay were performed, which showed significant results. The bark extract exhibited significant cytotoxic potential against both the colon cancer cell lines, and it also inhibited cell migration in a dose-dependent and time-dependent experiment. The IC₅₀ values of MEPJB on the HCT116 cell line for 24 hours, 48 hours, and 72 hours were 83.33 μg/mL, 37.5 μg/mL, and 34.37 μg/mL, respectively, whereas on the SW480 cell line, the IC₅₀ values were 87.5 μg/mL, 40 μg/mL and 31.25 μg/mL for 24 hours, 48 hours, and 72 hours respectively. However, IC₅₀ doses of MEPJB were inactive against the normal, healthy human fibroblast cell line. Cytotoxicity screening of 5-fluorouracil (5-FU), a commonly used drug in colon cancer treatment, was also done on both the colon cancer cell lines HCT116 and SW480, which showed much higher IC₅₀ than MEPJB after 24 hours, 48 hours, and 72 hours of incubation. MEPJB changed the cell morphology and induced apoptosis in colon cancer cell lines, confirmed by a morphological study of treated and untreated colon cancer cell lines and the acridine orange/ethidium bromide (AO/EB) dual staining procedure, respectively. After 48 hours of incubation with respective IC₅₀ concentrations of MEPJB, the treated colon cancer cells showed early and late apoptotic features. So this current research work indicates that the methanolic extract of P. javanica bark has chemotherapeutic potential against colon cancer cells and promotes apoptosis in cancer cells, which results in cell death.

Secretome of adipose derived stem cells induced apoptosis in anaplastic thyroid carcinoma C-643 cells

Introduction: Previous studies have proven that secretory materials of mesenchymal stem cells (MSCs) (secretome) have remedial properties. Objectives: The main goal of this study was to evaluate the effects of secretome of adipose-derived MSCs on growth and apoptosis in anaplastic thyroid carcinoma (ATC) C-643 cells. Materials and Methods: Initially, thyroid carcinoma cells were exposed to the 25 and 50 μg/mL adipose-derived stem cells (ADSCs) secretome for 24 and 48 hours, to evaluate the proliferation and cytotoxicity of the C-643 cells, MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] test and colony assay was conducted. Acridine orange/ethidium bromide (AO/EB) staining was conducted to evaluate the apoptosis. The expression level of apoptosis-associated genes was determined by real-time polymerase chain reaction (PCR) technique. Results: Cell viability and colony numbers in groups exposed to secretome were significantly lower than the control group. The amount of apoptosis-related genes (Bax/Bcl-2, P53, caspase-3 and caspase-8) expression in secretome-treated groups was more than the control group. Conclusion: The data revealed that ADSCs secretome caused the significant reduction of cell growth and induced apoptosis in C-643 cells.

Preparation and anti-tumor activity of PEG-PCL polymersomes loaded with curcumol derivative in HepG2 cell line

Curcumol is a natural plant product with anti-tumor properties. However, the clinical application of curcumol is limited by its low water solubility. In this study, we synthesized a curcumol derivative 2e (CD-2e) with improved bioavailability and evaluated its cell growth-regulating activities in human hepatoma HepG2 cells, a liver cancer precursor cell line. Using the clone formation assay, we detected that CD-2e inhibits proliferation and activity of HepG2 cells more effectively than curcumol. Flow cytometry analysis indicated that CD-2e can induce HepG2 cell apoptosis and arrests cell cycle in S phase. Acridine orange-ethidium bromide(AO-EB) double staining indicated that CD-2e triggers HepG2 cell apoptosis in a time-dependent manner. Additionally protein expression analysis showed that CD-2e treatment reduced expression levels of c-Jun N-terminal kinase 1 (JNK1) and its phosphorylated form, P-JNK1. PEG-PCL polymersome carrying CD-2e (PEG-PCL-CD-2e) were generated to improve drug delivery. The prepared PEG-PCL-CD-2e had a uniform particle size distribution, with an average particle size of about 190 nm, and good in vitro stability. The drug release and cell uptake experiments showed that PEG-PCL-CD-2e can rapidly release CD-2e in a weak-acid environment in vitro. Pharmacokinetic studies indicated that, compared with free CD-2e, PEG-PCL-CD-2e enhanced Cmax and Area Under Curve (AUC) (0-∞) by about 1.93-fold and 2.45-fold. The T1/2 was prolonged to 2.59 ± 0.03 h.

A SOCT-ISC type photosensitizer coumarin-BODIPY promoted by AIE effect: Mechanism of singlet oxygen generation, simulated PDT in A-549 cells and fluorescence imaging in zebrafish

A spin−orbit charge-transfer intersystem crossing (SOCT-ISC) type photosensitizer coumarin-BODIPY (CM-BDP) was reported in this paper. Due its aggregation induced emission (AIE) effect, CM-BDP could work under high water content condition. Besides that, another important phenomenon was discovered. In DMSO solvent, the fluorescence emission and the singlet oxygen yield of CM-BDP were almost negligible, while, in 90% water content system, the singlet oxygen yield of CM-BDP was up to 20.9% and a strong TICT fluorescence emission at 648 nm was observed, which indicated the AIE effect could also promote the generation of singlet oxygen by preserving the energy. In terms of the mechanism of generating singlet oxygen, the D-A configuration of CM-BDP could facilitate the process of charge separation, as well as the generation of TICT state. Then, the orthogonal configuration of CM-BDP in TICT state and the small △Est between the TICT state and T2 state could promote the SOCT-ISC process effectively. Combining the femtosecond transient absorption spectra and the DFT theoretical calculations, the photophysical process of CM-BDP was proposed in simplified Jablonski diagram. Moreover, the CM-BDP was also applied to intracellular experiment and zebrafish imaging. In the photo/dark toxicity experiment, cell survival rate of dark toxicity experiment was basically higher than 90%, while its decreased significantly under irradiation. The result of acridine orange/ethidium bromide (AO/EB) stain experiment indicated that the CM-BDP could kill A-549 cancer cells effectively under irradiation and the fluorescence imaging in zebrafish proved that CM-BDP could be further applied in organisms.

<p><strong>Apoptosis mediated cytotoxic potential of <em>Erythrina variegata</em> L. stem bark in human breast carcinoma cell lines</strong></p>

Erythrina variegata L. (Fam. Fabaceae) is traditionally known for antitumour, expectorant, febrifuge, antibacterial and antioxidant activities. In the present study, we did cytotoxic evaluation of the methanol extract of E. variegata stem bark (MEV) using 3-(4,5-dimethyl thazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay in MCF-7 and MDA-MB-231 human breast carcinoma cell lines. The cytological and metabolic alterations due to MEV was evaluated using acridine orange/ethidium bromide (AO/EB) dual staining, hoechst 33258 and fluoroprobe, benzimidazol-carbocyanine iodide 5,5',6,6'-tetrachloro-1,1',3,3'-tetra ethyl (JC-1) staining. The antiapoptotic gene and protein, Bcl-2 expression was analysed using real time PCR and Western blotting, respectively. The methanol extract was investigated for the presence of various phytochemicals using gas chromatography high resolution mass spectrometry (GC-HRMS). The results revealed a dose dependent cytotoxic activity in both MDA-MB-231 and MCF-7 breast carcinoma cell lines in vitro. The plant extract was effective in inducing apoptotic cell death via mitochondrial dependent intrinsic pathway and may be considered as a potent source for isolating therapeutic molecules against cancer.

Ozone induces BEL7402 cell apoptosis by increasing reactive oxygen species production and activating JNK.

BackgroundOxidative stress is an important factor in the modulation of both tumorigenesis and anticancer responses. Ozone (O3) is a strong oxidant that causes redox reactions and exerts anticancer effects in various types of cancer cells. However, the pathways involved in O3-induced cell death are not well understood.MethodsIn vitro human hepatocellular carcinoma (HCC) BEL7402 cells were treated with various O3 concentrations to evaluate O3 cytotoxicity by Cell Counting Kit-8 (CCK-8) assay and flow cytometry. The regulatory mechanisms were analyzed by western blot analysis. In vivo, an HCC model was established to evaluate the inhibition of HCC with O3 treatment.ResultsIn vitro cells treated with O3 exhibited a round and small morphology with nuclear shrinkage and fragmentation. The CCK-8 assay confirmed the potent cytotoxic activity of O3 against BEL7402 cells (IC50 value of 5 µg/mL). Acridine orange/ethidium bromide (AO/EB) staining revealed apoptosis of BEL7402 cells after O3 treatment. Flow cytometry analysis showed that S phase cell cycle arrest and apoptosis increased with O3 exposure. In addition, O3 exposure reduced the mitochondrial membrane potential (ΔΨm) and induced reactive oxygen species (ROS) accumulation. Western blot analysis showed that O3 exposure reduced B-cell lymphoma 2 (BCL-2) expression and increased cleaved poly ADP-ribose polymerase (PARP), cytochrome C (Cyt-C), caspase-3, caspase-9, and p-JNK expression. In vivo, treatment with intratumor injection O3 (20 µg/mL) inhibited HCC growth.ConclusionsOverall, our findings showed that O3 induces BEL7402 cell apoptosis via the intrinsic mitochondria-dependent pathway. Therefore, O3 has therapeutic potential for HCC.

Design, synthesis, and biological evaluation of N-(4-substituted)-3-phenylisoxazolo[5,4–d]pyrimidin-4-amine derivatives as apoptosis-inducing cytotoxic agents

A library of new 3-phenylisoxazolo[5,4–d]pyrimidines (8–10) was designed based on a scaffold hybridization technique incorporating the important pharmacophoric features of 4-aminopyrimidine and phenyl isoxazole scaffold which is renowned for its BET inhibition activity. The designed molecules were synthesized and evaluated with the NCI-60 cell line panel. Examination by NCI-60 cell lines at single-dose and the five-dose study showed that compound 10h exhibited promising growth inhibitory effects with GI50 values on various cancer cell lines such as HCT-15 (Colon Cancer)-0.0221 μM, MDA-MB-435 (Melanoma) − 0.0318 μM, SNB-75(CNS Cancer)-0.0263 μM, and MCF7 (Breast Cancer)-0.0372 μM. Further studies to know the mechanism of action of 10h based on the phase-contrast microscopic evaluation, DAPI, acridine orange/ethidium bromide (AO/EB) staining, and annexin V-FITC assays revealed that elevation in the intracellular ROS leads to alteration in mitochondrial membrane potential which in turn induced the apoptosis in BT-474 cancer cells, which could be the plausible mechanism of action for compound 10h.

Tunicamycin-Induced Endoplasmic Reticulum Stress Promotes Breast Cancer Cell MDA-MB-231 Apoptosis through Inhibiting Wnt/β-Catenin Signaling Pathway.

Triple negative breast cancer (TNBC) has significantly threatened human health. Many aspects of TNBC are closely related to Wnt/β-catenin pathway, and cell apoptosis induced by endoplasmic reticulum stress (ER stress) in TNBC may act as a potential target of non-chemotherapy treatment. However, how ER stress interacts with this pathway in TNBC has not yet been understood. Here, the tunicamycin and LiCl have been applied to MDA-MB-231. The related proteins' expression was measured by western blotting. Moreover, acridine orange/ethidium bromide (AO/EB) staining was applied to test the apoptosis degree of the cells, and cell viability was tested by MTT experiment. Then, we found the ER stress and apoptosis degree of MDA-MB-231 were induced after treatment with tunicamycin. Besides, tunicamycin dose dependently inhibited both Wnt/β-catenin pathway and cells viability. Licl, an activator of Wnt/β-catenin signaling pathway, could significantly inhibit cell apoptosis. In conclusion, our study found that the activation of ER stress could promote the MDA-MB-231 apoptosis by repressing Wnt/β-catenin pathway, which provides some promising prospects and basic mechanism to the further research.

Silencing of acetyl-CoA carboxylase-α gene in human gastric cancer cells inhibits proliferation via induction of apoptosis, autophagy and suppression of cell invasion

Purpose: To study the role and therapeutic potential of acetyl-CoA-carboxylase-α (ACC) in the management of gastric cancer.
 Methods: Expression of ACC in gastric cancer cell lines was determined using quantitative real-time polymerase chain reaction (qRT-PCR). Lipofectamine 2000 reagent was used for transfection, while cell viability was determined by MTT assay. Apoptotic cell death was assayed with 4′, 6-diamidino-2- phenylindole (DAPI) and acridine orange/ethidium bromide (AO/EB) staining. The proportion of apoptotic cells was estimated with Annexin V/PI staining. Wound healing and Transwell assays were employed to monitor cell migration and invasion, while protein expression was determined using western blotting.
 Results: The results showed that ACC was significantly enhanced in SNU-1 gastric cancer cells (4.2- fold). Silencing of ACC in SNU-1 gastric cancer cells caused significant decrease in cell proliferation (p < 0.05). Electron microscopy examination showed that ACC silencing triggered autophagic cell death in SNU-1 cells, and increased expression of LC3 II. Results from DAPI and AO/EB assays demonstrated that ACC silencing also promoted apoptosis in SNU-1 gastric cancer cells. Annexin V/PI assay results revealed that apoptotic cell population increased from 2.7 to 13.8 % due to ACC silencing (p < 0.05). Moreover, Bax expression increased, while Bcl-2 expression decreased upon ACC silencing. Transwell assay results indicate that ACC silencing caused marked decrease in the invasion of the SNU-1 cells and downregulation of the expressions of MMP-2 and MMP-9 (p < 0.05).
 Conclusion: ACC is likely to be an important therapeutic target for gastric cancer.

Biocompatible madecassoside encapsulated alginate chitosan nanoparticles, their anti-proliferative activity on C6 glioma cells

Madecassoside (MAD), an ursane type pentacyclic triterpenoid saponin, isolated from Centella asiatica (CA) possesses various pharmacological activities. The aim of this study was to evaluate therapeutic potential of MAD encapsulated alginate chitosan nanoparticles (MACNPs) on C6 glioma cells. Synthesized nanoparticles were characterized by Thermogravimetric analysis (TGA), Differential thermal analysis (DTA), Differential scanning calorimetry (DSC), Scanning electron microscopy (SEM), Transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy (FTIR). Cell viability and compatibility of both MAD and MACNP were observed on primary astrocytes. Cytotoxicity and anti-proliferative effect of MACNPs were studied on C6 glioma cells by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5′-ethynyl-2′-deoxyuridine (Edu) assays. MACNPs showed 90% cytotoxicity, and depicted anti-proliferative activity against C6 cells. Annexin V-Alexa flour-488/propidium iodide (PI) staining followed by flow cytometry revealed that MACNPs induced necrosis in C6 glioma cells. Cell death was also observed using acridine orange/ethidium bromide (AO/EtBr) and Hoechst staining. 2′,7′-Dichlorodihydrofluorescein diacetate (DCFDA) assay by flow cytometry showed an increase in reactive oxygen species (ROS) production in MACNP treated C6 glioma cells. MACNPs showed a greater uptake and distribution in C6 glioma cells and inhibited the proliferation of C6 cells via increased intracellular ROS production. Our findings suggest MACNP as a promising drug delivery carrier for the management of glioma and shed new insights to brain malaises.

Evaluating the genetic basiss of anti-cancer property of Taxol in Saccharomyces cerevisiae model.

Taxol has been regarded as one of the most successful anti-cancer drugs identified from natural sources to date. Although Taxol is known to sensitize cells by stabilizing microtubules, its ability to cause DNA damage in peripheral blood lymphocytes and to induce oxidative stress and apoptosis indicates that Taxol may have other modes of cytotoxic action. This study focuses on identifying the additional targets of Taxol that may contribute to its multifaceted cell killing property, using Saccharomyces cerevisiae. We show that yeast oxidative stress response mutants (sod1Δ, tsa1Δ and cta1Δ) and DNA damage response mutants (mre11∆, sgs1∆ and sub1∆) are highly sensitive to Taxol. Our results also show that Taxol increases the level of reactive oxygen species (ROS) in yeast oxidative stress response mutant strains. Further, 4',6-Diamidino-2'-phenylindole (DAPI) and acridine orange/ethidium bromide (AO/EB) staining show that Taxol induces apoptotic features such as nuclear fragmentation and chromatin condensation in DNA repair mutants. On the whole, our results suggest that Taxol's cytotoxic property is attributed to its multifaceted mechanism of action. Yeast S. cerevisiae anti-oxidant and DNA repair gene mutants are sensitive to Taxol compared to wild-type, suggesting yeast model can be used to identify the genetic targets of anti-cancer drugs.

Assessment of Rhamnus alaternus Leaves Extract: Phytochemical Characterization and Antimelanoma Activity.

Rhamnus alaternus (Rhamnaceae) has been used as a laxative, purgative, diuretic, antihypertensive, and depurative. However, few scientific research studies on its antimelanoma activity have been reported. This study aimed to investigate the in vitro antimelanoma effect of an enriched total oligomer flavonoid (TOF) extract, from R. alaternus, and to identify its phytochemical compounds. The chemical composition of TOF extract was assessed by HPLC-electrospray ionization tandem mass spectrometry (HPLC/ESI-MS2) analysis. Antimelanoma activity was determined on cultured tumor cell B16F10 by the crystal violet assay, the alkaline comet assay, acridine orange/ethidium bromide (AO/EB), annexin V-fluorescein isothiocyanate/ propidium iodide (V-FITC/PI) staining, the cell cycle distribution, and the wound healing assay. Regarding chemical composition, a mixture of quercetin diglucoside, quercetin-3-O-neohesperidoside, kaempferol-3-O-(2G-α-L-rhamnosyl)-rutinoside, rhamnetin hexoside, kaempferol-3-O-rutinoside, rhamnocitrin hexoside, pilosin hexoside, apigenin glucoside, and kaempferol-3-O-glucoside was identified as major phytochemical compounds of the extracts. TOF extract inhibits melanoma B16F10 cell proliferation in dose-dependent manner. The induction of apoptosis was confirmed by comet assay, AO/EB, and annexin V-FITC/PI test. TOF extract could also induce S phase cell cycle, inhibit, and delay the cell migration of B16F10 cells. The findings showed that TOF extract from R. alaternus could be a potentially good candidate for future use in alternative antimelanoma treatments.

A feasible strategy of fabricating hybrid drugs encapsulated polymeric nanoparticles for the treatment of gastric cancer therapy

Chemotherapy of hybrid-drugs has become one of the widely used therapies for the prevention of cancers with promising clinical results. This study aimed to create compostable polymeric nanoparticles (PPNPs) for the codelivery of capsaicin (CAPS) and biotin (BT) and to examine the anticancer activity of the drug delivery mechanism (CAPS/BT@PPNPs) against in vitro and in vivo gastric cancer cells. Because of the insufficiency of double antitumor drugs BT and CAPS compressed in polymer-based bio-degradable nanoparticles, the encapsulation of BT and CAPS antitumor drugs co-loaded with polyethylene glycol and polylactidecoglycolide nanoparticles (PPNPs) is effective. Transmission electron microscopy (TEM) was utilized to investigate the structure of BT@PPNPs, CAPS@PPNPs, and BT/CAPS@PPNPs, as well as their shape and size. Human gastric carcinoma cell lines, such as SGC-791 and NCI-N87, were also induced to apoptosis in vitro by BT/CAPS@PPNPs. Morphologies and cell death were observed using bio-chemical staining methods, AO/EB (acridine orange/ethidium bromide) and Hoechst. Flow cytometry in dual staining was also used to confirm the mechanistic investigations of apoptosis. Overall, this dual drug delivery strategy suggests that BT/CAPS@PPNPs could be used as a new method to increase the efficacy of gastric therapeutics.

Copperpod Plant Synthesized AgNPs Enhance Cytotoxic and Apoptotic Effect in Cancer Cell Lines

The utilization of biological resources on the manufacture of nano silver has attracted the interest of researchers to develop an eco-friendly, cost-effective technology in nanomaterials production. In the present study, plant-mediated silver nanoparticles (AgNPs) were synthesized using aqueous leaf extracts of the Copperpod plant, which was well characterized. The ultraviolet-visible spectrophotometric study showed a maximum absorbance peak at 425 nm, and the observation of transmission electron microscopic features revealed that the nanoparticles size ranged between 20 and 70 nm. The synthesized AgNPs were tested for in vitro cytotoxic effects against cancerous cells, such as HepG2, A549 and MCF-7 cells. The findings showed that the IC50 values of AgNPs against cancerous cells viz., HepG2, MCF-7 and A549 cells, were observed to be 69 µg/mL, 62 µg/mL and 53 µg/mL, respectively. In addition, the apoptosis property was analysed using propidium iodide and acridine orange-ethidium bromide via the DNA fragmentation technique. Thus, the outcomes of the current analysis presume that the plant mediated AgNPs obtained from a synthesized Copperpod plant possess significant anti-cancer properties against various cancerous cells.

Long non-coding RNA H19 regulates proliferation of ovarian granulosa cells via STAT3 in polycystic ovarian syndrome.

IntroductionStudies have shown that long non-coding RNAs (lncRNA) are aberrantly expressed in polycystic ovarian syndrome (PCOS) ovaries and may have a role in PCOS development. In this study, the role and therapeutic implications of lncRNA H19 were investigated in PCOS ovaries and granulosa cells.Material and methodsqRT-PCR was used for expression analysis. Cell Counting Kit 8 (CCK-8) assay was used for cell viability and acridine orange/ethidium bromide (AO/EB) and Annexin V/propidium iodide staining was used to detect apoptosis. All transfections were carried out with Lipofectamine 2000 reagent. Western blot analysis was used for protein expression analysis.ResultsThe expression of lncRNA H19 was remarkably upregulated in the PCOS ovarian tissues as well as the granulosa cells. Suppression of lncRNA H19 expression caused the inhibition of KGN granulosa cell proliferation due to the triggering of apoptosis. Bioinformatic analysis revealed the presence of the GAS binding site for STAT3 in the promoter of lncRNA H19. Silencing of STAT3 suppressed the expression of lncRNA H19 in KGN cells and also halted their growth by triggering apoptosis. Co-transfect experiments revealed that STAT3 and lncRNA H19 silencing cause inhibition of KGN growth synergistically.ConclusionslncRNA H19 regulates the growth of ovarian granulosa cells and might prove to be a therapeutic target for management of PCOS.