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Abstract Background Arsenic in groundwater and its accumulation in plants and animals have assumed a menacing proportion in a large part of West Bengal, India and adjoining areas of Bangladesh. Because of the tremendous magnitude of the problem, there seems to be no way to tackle the problem overnight. Efforts to provide arsenic free water to the millions of people living in these dreaded zones are being made, but are awfully inadequate. In our quest for finding out an easy, safe and affordable means to combat this problem, a homeopathic drug, Arsenicum Album-30, appears to yield promising results in mice. The relative efficacies of two micro doses of this drug, namely, Arsenicum Album-30 and Arsenicum Album-200, in combating arsenic toxicity have been determined in the present study on the basis of some accepted biochemical protocols. Methods Mice were divided into different sets of control (both positive and negative) and treated series (As-intoxicated, As-intoxicated plus drug-fed). Alanine amino transferase (ALT) and aspartate amino transferase (AST) activities and reduced glutathione (GSH) level in liver and blood were analyzed in the different series of mice at six different fixation intervals. Results Both Arsenicum Album-30 and Arsenicum Album-200 ameliorated arsenic-induced toxicity to a considerable extent as compared to various controls. Conclusions The results lend further support to our earlier views that microdoses of potentized Arsenicum Album are capable of combating arsenic intoxication in mice, and thus are strong candidates for possible use in human subjects in arsenic contaminated areas under medical supervision.

Polygala senega is extensively used in traditional systems of medicine against various lung diseases including cancer. In the present study we tested the anticancer potentials of ethanolic extract of roots of P. senega (generally used as a homeopathic drug) in a mammalian model, where mice, in vivo, were treated chronically with benzo[a] pyrene and in vitro where lung adenocarcinoma cell line (A549) were used. We deployed various parameters like cell viability assay, chromatin condensation studies with Hoechst 333258 staining, and maintained suitable controls. To understand the possible signal transduction pathways, expression of various signal proteins such as Aryl Hydrocarbon receptor (AhR), cytochrome P450 (CYP1A1), Bcl-2, proliferating cell nuclear antigen (PCNA), Bax and Caspase-3 was studied. Additionally, reverse transcriptase polymerase chain reaction analysis of AhR, p53, PCNA and ß-actin (housekeeping) genes was made. Immunohistochemical localization of PCNA proteins was also conducted in vivo. Feeding of root extract of P. senega to mice (at the rate of 50 mg/kg and 100 mg/kg bw) chronically treated with the carcinogen (50 mg/kg bw dissolved in olive oil) showed positive modulation in expression of signal proteins. Upregulation of apoptotic signals such as p53, Caspase-3 and Bax, and downregulation of AhR, cytochrome P450 (CYP1A1), Bcl-2 and PCNA were observed. Addition of root extract of Polygala Senega (at doses of 50 µg and 100 µg) into culture medium containing A549 cells induced recovery of decreased cell viability and increased chromatin fragmentation (apoptosis). Therefore, results of both in vivo and in vitro studies scientifically validate its potential use as an anticancer agent, particularly against lung cancer, and provide important information potentially helpful in drug designing. [ABSTRACT FROM AUTHOR]

Homeopathy is a holistic method of treatment that uses ultralow doses of highly diluted natural substances originating from plants, minerals, or animals and is based on the principle of 'like cures like.' Despite being occasionally challenged for its scientific validity and mechanism of action, homeopathy continues to enjoy the confidence of millions of patients around the world who opt for this mode of treatment. Contrary to skeptics' views, research on home-opathy using modern tools mostly tends to support its efficacy and advocates new ideas toward understanding its mechanism of action. As part of a Point-Counterpoint feature, this review and its companion piece in this issue by Moffett et al (Integr Cancer Ther. 2006;5:333-342) are composed of a thesis section, a response section in reaction to the companion thesis, and a rebuttal section to address issues raised in the companion response. [ABSTRACT FROM AUTHOR]

Objectives: This study was conducted to examine whether the potentized homeopathic remedy Carcinosin 200, fed alone and in combination with Chelidonium 200, has differential protective effects against p-dimethylaminoazobenzene (p-DAB)-induced hepatocarcinogenesis in mice. Design: Liver tumors were induced in mice through chronic feeding of p-DAB (initiator) and phenobarbital (PB, promoter). The mice were divided into two subgroups: (1) one was fed potentized Alcohol 200 and served as controls; and (2) the other was fed Carcinosin 200 alone or in combination with Chelidonium 200 and divided into several sets. The relative efficacy of the two potentized remedies, alone or in combination, in combating hepatocarcinogenesis was assessed through several cytogenetical endpoints such as chromosome aberrations, induction of micronuclei, sperm head anomaly, and mitotic index at several intervals of fixation (days 7, 15, 30, 60, 90, and 120). Several toxicity biomarkers such as acid and alkaline phosphatases, glutamate oxaloacetate transaminase, glutamate pyruvate transaminase, and lipid peroxidation activity were also assayed in three organs of treated and control mice. In addition, recovery by the homeopathic drugs, if any, of tissue damage inflicted because of chronic feeding of p-DAB and PB was also assessed by optical, scanning, and transmission electron microscopies of liver done at days 60 and 120. Results: Both Carcinosin 200 and Chelidonium 200 when administered alone show considerable ameliorative effect against p-DAB-induced hepatocarcinogenesis in mice; but the conjoint feeding of these two drugs appears to have had a slightly greater protective effect. Conclusions: These homeopathic remedies have the potential to be used as complementary and alternative medicine in liver cancer therapy, particularly as supporting palliative measures. [ABSTRACT FROM AUTHOR]

The present study tends to evaluate the possible potential of bio-active Morroniside (MOR), against alloxan (ALX)-induced genotoxicity and hyperglycaemia. In silico prediction revealed the interaction of MOR with Poly (ADP-ribose) polymerase (PARP) protein which corroborated well with experimental in vitro L6 cell line and in vivo mice models. Data revealed the efficacy of MOR in the selective activation of PARP protein and modulating other stress proteins NF-κB, and TNF-α to initiate protective potential against ALX-induced genotoxicity and hyperglycaemia. Further, the strong interaction of MOR with CT-DNA (calf thymus DNA) analyzed through CD spectroscopy, UV-Vis study and ITC data revealed the concerted action of bio-factors involved in inhibiting chromosomal aberration and micronucleus formation associated with DNA damage. Finally, MOR does not play any role in microbial growth inhibition which often occurs due to hyperglycemic dysbiosis. Thus, from the overall findings, we may conclude that MOR could be a potential drug candidate for the therapeutic management of induced-hyperglycaemia and genotoxicity.Communicated by Ramaswamy H. Sarma.

Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Garlic ( Allium sativum L.), a popular spice, has been used for decades in treating several medical conditions. Although Allicin, an active ingredient of garlic has been extensively studied on carcinogen-induced hepatotoxicity and oxidative stress in rats ( Rattus norvegicus ), no systematic study on the beneficial effects of generic aged garlic and specific aged garlic extract-Kyolic has been done. The present study involves rats fed chronically with two liver carcinogens, p-dimethylaminoazobenzene and phenobarbital, to produce hepatotoxicity. The aged garlic extract was characterized by UV-spectra, FTIR, HPLC and GC-MS. Biochemical and pathophysiological tests were performed by keeping suitable controls at four fixation intervals, namely, 30, 60, 90, and 120 days, utilizing several widely accepted toxicity biomarkers. Compared to the controls, remarkable elevation in the activities of lactate dehydrogenase, gamma glutamyl transferase and decline in catalase and glucose-6-phosphate dehydrogenase were observed in the carcinogen fed rats. Daily administration of aged garlic extract, could favorably modulate the elevated levels of various toxicity biomarkers including serum triglyceride, creatinine, urea, bilirubin, blood urea nitrogen except total cholesterol. It also altered the levels of blood glucose, HDL-cholesterol, albumin, AST, ALT, and hemoglobin contents in carcinogen intoxicated rats, indicating its protective potential against hepatotoxicity and oxidative stress in the experimental rats. Down-regulation of Bcl-2 and p53 proteins caused cell cycle arrest and apoptosis in garlic fed group. Kyolic exhibited additional benefits by arresting cell viability of cancer cells. This study would thus validate the use of aged garlic extract in the treatment of diseases causing liver toxicity including hepatocarcinoma.

Cisplatin (CDDP) is a potent chemotherapeutic drug, but its severe side-effects often prohibit its use. Combined treatment with CDDP plus Farnesol (FAR) and their co-encapsulated nano form were investigated in in vitro to examine if synergistic cytotoxicity of this combination could reduce unwanted side-effects of CDDP chemotherapy and potentiate CDDP anticancer activity against hepatocellular carcinoma (HCC) cells. After finding combination therapy of CDDP and FAR successfully combat HCC we formulated co-encapsulation of CDDP and FAR within poly(lactic-co-glycolic acid) copolymer (NCDDPFAR) by following the standardized solvent displacement method. NCDDPFAR treatment caused faster drug mobility, sustained particle release, site-specific action and higher percentage of apoptotic death compared with single drug treatment even at relatively low concentrations. Co-encapsulation of two drugs exhibited additive effects against HCC; FAR reduced CDDP-induced glutathione level by increasing expression of CYP2E1 while CDDP directly interacted with DNA; FAR up-regulated the expression of TopII, thereby promoting DNA breaks and escaping DNA repair machinery. Expression pattern of apoptotic genes like p53, Bax, cytochrome c and caspase-3 suggested that NCDDPFAR induced HCC cell death through mitochondrial intrinsic pathway. Administration of NCDDPFAR had better ability of drug carriage and enhanced anticancer potentials against HCC cells.

Background: Cisplatin is a widely-used potent anti-cancer drug having severe side-effects precluding its sustained use., Objectives: Poly (lactide-co-glycolide) (PLGA)-nanoparticles loaded Boldine, an antioxidant ingredient of ethanolic extract of Boldo plant (Peumus boldus) was tested in cancer mice model, Mus musculus to examine if it could reduce unwanted Cisplatin-induced toxicity in normal tissue., Material and Methods: Nano-encapsulation of Boldine was done by following the standardized solvent displacement method. Physico-chemical characterization of PLGA-encapsulated nano-Boldine (NBol) was accomplished through analyses of various spectroscopic techniques. Status of major antioxidant enzymes, functional markers, and lipid peroxidation (LPO) was also determined in certain tissue and serum samples. Percentage of cells undergoing cytotoxic death, Reactive oxygen species (ROS) accumulation and mitochondrial functioning were analyzed in both normal and cancer mice. Nanoscale changes in chromatin organization were assessed by Transmission electron microscopy (TEM). mRNA and protein expressions of Top II, Bax, Bcl-2, Cyt c, caspase 3 were studied by RT-PCR, immunoblot and immunofluorescence., Results: NBol had faster mobility, site-specific action and ability of sustained particle release. NBol readily entered cells, prevented Cisplatin to intercalate with dsDNA resulting in reduction of chromatin condensation, with corresponding changes in ROS levels, mitochondrial functioning and antioxidant enzyme activities, leading to reduction in Deoxyribose nucleic acid (DNA) damage and cytotoxic cell death. Expression pattern of apoptotic genes like Top II, p53, Bax, Bcl-2, cytochrome c and caspase-3 suggested greater cytoprotective potentials of NBol in normal tissues., Conclusions: Compared to Boldine (Bol), NBol had better ability of drug carriage and protective potentials (29.00% approximately) against Cisplatin-induced toxicity. Combinational therapeutic use of PLGA-NBol can reduce unwanted Cisplatin-induced cellular toxicity facilitating use of Cisplatin., (Copyright © 2018 Transdisciplinary University, Bangalore and World Ayurveda Foundation. Published by Elsevier B.V. All rights reserved.)

Removal of bio-accumulated pesticides in edible fish is a global problem. In this study, we tested protective capability of a phytochemical pelargonidin-loaded non-toxic, biodegradable poly-lactide-co-glycolide nano-particles (NPG) against toxicity induced by a pesticide cypermethrin (CM) in a fish model (Oreochromis mossambica) in vivo and also in L6 muscle cell line, in vitro. First we assessed potential sustainable release of nanoparticles following oral administration of NPG to fish, their ability to cross sub-cellular membranes in several tissues and efficacy to cross blood-brain-barrier. Next, protective ability of NPG, if any, against CM in fish was evaluated deploying parameters like % cell viability, DNA damage in muscle cells and modulation of anti-oxidative-enzymes like superoxide dismutase, catalase and lipid peroxidase. Modulation of reactive oxygen species generation, nuclear condensation and alteration in stress related protein signalling cascade were assessed in L6 cells. Results revealed that NPG had nano-size range (~10-12 nm) and negative zeta potential (-17 mV). Bioavailability and distribution of NPG could be followed by spectrophotometric absorbance of pelargonidin at 293 nm from 6 h onward till 24 h in all important tissues including the brain. Thus, 0.5 mg/g b.w. NPG could demonstrate protective ability in CM-intoxicated fish muscle cells in respect of % cell viability, DNA damage and stress related enzymes. Similar alterations could also be found in signalling protein cascade in L6 cells in response to treatment of 5 μg/ml NPG against CM-induced toxicity and depletion of overall ROS generation and nuclear condensation. Therefore, NPG could be used as a potential drug in management of pesticide toxicity in cultured edible fish., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Objectives: To examine if HIV nosode in 30c dilution (HIV 30c) has therapeutic potential against lung cancer cells (A549) as compared to WRL-68 normal cells and to elucidate its possible molecular mechanism of action on DNA replication and apoptosis., Methods: Effects of HIV 30c were thoroughly tested for its possible anticancer potential on A549 cells (lung cancer); WRL-68 normal liver cells served as control. Three doses, one at LD50 and two below LD-50, were used. Proliferation, migration and senescence assays were made and generation of reactive oxygen species (ROS) studied by routine techniques. The ability of HIV 30c to induce apoptosis in A549 cells and its possible signalling pathway were determined using immunoblots of relevant signal proteins and confocal microscopy, including studies on telomerase reverse transcriptase (TERT) and topoisomerase II (Top II) activities, intimately associated with cell division and DNA replication., Results: HIV 30c prevented cancer cell proliferation and migration, induced pre-mature senescence, enhanced pro-apoptotic signal proteins like p53, bax, cytochrome c, caspase-3 and inhibited anti-apoptotic signal proteins Bcl2, TERT and Top II, changed mitochondrial membrane potential and caused externalization of phosphatidyl serine. Thus, it induced apoptosis as also evidenced from increase in cells with distorted membrane morphology, nuclear condensation, DNA fragmentation, and ROS, typical of apoptosis in progress., Conclusion: HIV 30c nosode has therapeutic potential for inducing cytotoxic effects on A549 cells as manifested by changes in nuclear condensation, DNA fragmentation, ROS generation and MMP, and for its inhibitory action on cell proliferation, cell migration, expression of telomerase reverse transcriptase and Top II genes, and increasing expression of pro-apoptotic genes., (Copyright © 2017 The Faculty of Homeopathy. Published by Elsevier Ltd. All rights reserved.)

Nano-encapsulation of several natural products has become an important tool in enhancing the bioavailability of some modern drugs against many diseases. Pelargonidin is an anthocyanidin found in many fruits and vegetables. Pelargonidin is loaded with poly-lactide-co-glycolic-acid, a non-toxic biodegradable polymer, to produce nano-pelargonidin. Size, morphology, zeta potential, and planar uniformity of formulated nano-pelargonidin were determined by atomic force microscopy and dynamic light scattering. The time required for cellular entry, folds of nano-pelargonidin, and drug encapsulation efficiency of poly-lactide-co-glycolic-acid were also ascertained. Relative functional efficacy of nano-pelargonidin and pelargonidin was evaluated by examining markers such as pyruvate kinase, glucokinase, calcium ion level, ATP/ADP ratio, mitochondrial membrane potential, cytosolic release of mitochondrial cytochrome-c, and structural analysis of mitochondrial DNA in controlled and experimental sets of alloxan-induced hyperglycemic L6 cells. Expressions of mitochondrial apoptotic proteins, such as bcl2 and caspase3, and glucose signalling cascades, such as GLUT4, IRS1, IRS2, and PI3, were analyzed. Nano-pelargonidin at a nearly 10-fold reduced dose significantly enhanced protection, presumably due to its smaller size, ability of faster entry, and drug delivery at target-specific sites. Thus, nano-pelargonidin can be used in formulating protective drugs for therapeutic management of mitochondrial dysfunction often encountered in diabetic conditions., (Georg Thieme Verlag KG Stuttgart · New York.)

Chlorophyllin (CHL), a sodium-copper-salt derived from chlorophyll, has been widely used as a food-dye, also reportedly having some anti-cancer effect. We tested if PLGA-loaded CHL (NCHL) could have additional protective abilities through its faster and targeted drug delivery in cancer cells. Physico-chemical characterization of NCHL was done through atomic-force microscopy and UV-spectroscopy. NCHL demonstrated greater ability of drug uptake and strong anti-cancer potentials in non-small cell lung cancer cells, A549, as revealed from data of% cell viability, generation of reactive-oxygen-species and expression of bax, bcl2, caspase3, p53 and cytochrome c proteins. Circular dichroic spectral data indicated strong binding of NCHL with calf-thymus-DNA, causing a conformational/structural change in DNA. Further, NCHL could cross the blood-brain-barrier in mice and showed greater efficacy in recovery process of tissue damage, reduction in chromosomal aberrations and% of micronuclei in co-mutagens (Sodiumarsenite+Benzo[a]Pyrene)-treated mice at a much reduced dose, indicating its use in therapeutic oncology., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Current treatment methods for melanoma have some limitations such as less target-specific action, severe side effects and resistance to drugs. Significant progress has been made in exploring novel drug delivery systems based on suitable biochemical mechanisms using nanoparticles ranging from 10 to 400 nm for drug delivery and imaging, utilizing their enhanced penetration and retention properties. Poly-lactide-co-glycolide (PLGA), a copolymer of poly-lactic acid and poly-glycolic acid, provides an ideally suited performance-based design for better penetration into skin cells, thereby having a greater potential for the treatment of melanoma. Moreover, encapsulation protects the drug from deactivation by biological reactions and interactions with biomolecules, ensuring successful delivery and bioavailability for effective treatment. Controlled and sustained delivery of drugs across the skin barrier that otherwise prohibits entry of larger molecules can be successfully made with adequately stable biocompatible nanocarriers such as PLGA for taking drugs through the small cutaneous pores permitting targeted deposition and prolonged drug action. PLGA is now being extensively used in photodynamic therapy and targeted therapy through modulation of signal proteins and drug-DNA interactions. Recent advances made on these nanomedicines and their advantages in the treatment of skin melanoma are highlighted and discussed in this review., Competing Interests: None.

Objective: Homeopathic nosodes have seldom been scientifically validated for their anticancer effects. This study was conducted to examine if a recently developed hepatitis C nosode has demonstrable anticancer potential in cancer cells in vitro., Methods: Anticancer effects of Hepatitis C 30C (Hep C 30), if any, were initially tested on three cancer cell lines, HepG2 (liver cancer), MCF-7 (breast cancer) and A549 (lung cancer) and one normal liver cell line WRL-68 cells and subsequently a more thorough study using further scientific protocols was undertaken on HepG2 cells (against WRL-68 cells as the normal control) as HepG2 cells showed better anticancer response than the other two. Three doses, one at 50% lethal dose (LD50) and the other two below LD50, were used on HepG2 cells subsequently. Protocols like apoptosis induction and its possible signaling mechanism were deployed using immunoblots of relevant signal proteins and confocal microscopy, with particular reference to telomerase and topoisomerase II (Top II) activities, two strong cancer biomarkers for their direct relationship with divisional activities of cells and DNAs., Results: Hep C 30 induced apoptosis, caused distorted cell morphology typical of apoptotic cells, increased reactive oxygen species generation and produced increased DNA nicks. Further it enhanced pro-apototic signal proteins like Bax, cytochrome c and inhibited anti-apoptotic signal proteins, Bcl-2, cytochrome c and caspase-3, changed mitochondrial membrane potential and caused externalization of phosphatidylserine. The drug also decreased expression of two cancer biomarkers, Top II and telomerase, consistent with its anticancer effect., Conclusion: Hep C 30 has demonstrable anticancer effects against liver cancer cells in vitro.

Alloxan is an environmental food contaminant that causes DNA damage in living cells and induces hyperglycemia. Pelargonidin (PG), an active ingredient found in extract of various fruits and vegetables, has been nanoencapsulated (NPG) with poly-lactide-co-glycolide (PLGA) and tested for efficacy in prevention of alloxan (ALX)-induced DNA damage in L6 cells in vitro. Glucose uptake, reactive oxygen species (ROS) generation, glucose transporter 4, glucokinase levels and mechanism of activation of DNA repair proteins (PARP and p53) have been studied in ALX-induced L6 cells. Drug-DNA interaction has been analyzed using calf thymus DNA as target through circular dichroism and melting temperature profile. NPGs were physico-chemically characterized by standard protocols using dynamic light scattering and transmission electron microscopy. Pre-treatment with both PG and/or NPG was effective in reducing ALX-induced oxidative stress and showed favourable effects for protection against DNA damage by activating DNA repair cascades. Results suggested ∼10-fold increase in efficacy of NPG than PG in prevention of alloxan-induced oxidative stress and DNA damage., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Objectives: This study examined the relative efficacies of a derivative of betulinic acid (dBA) and its poly (lactide- co-glycolide) (PLGA) nano-encapsulated form in A549 lung cancer cells in vivo and in co-mutagen [sodium arsenite (SA) + benzo]undefined[a]pyrene (BaP)]-induced lung cancer in mice in vivo., Methods: dBA was loaded with PLGA nanoparticles by using the standard solvent displacement method. The sizes and morphologies of nano-dBA (NdBA) were determined by using transmission electron microscopy (TEM), and their intracellular localization was verified by using confocal microscopy. The binding and interaction of NdBA with calf thymus deoxyribonucleic acid (CT-DNA) as a target were analyzed by using conventional circular dichroism (CD) and melting temperature (Tm) profile data. Apoptotic signalling cascades in vitro and in vivo were studied by using an enzyme-linked immunosorbent assay (ELISA); the ability of NdBA to cross the blood-brain barrier (BBB) was also examined. The stage of cell cycle arrest was confirmed by using a fluorescence-activated cell-sorting (FACS) data analysis., Results: The average size of the nanoparticles was ~ 110 nm. Confocal microscopy images confirmed the presence of NdBA in the cellular cytoplasm. The bio-physical properties of dBA and NdBA ascertained from the CD and the Tm profiles revealed that NdBA had greater interaction with the target DNA than dBA did. Both dBA and NdBA arrested cell proliferation at G0/G1, NdBA showing the greater effect. NdBA also induced a greater degree of cytotoxicity in A549 cells, but it had an insignificant cytotoxic effect in normal L6 cells. The results of flow cytometric, cytogenetial and histopathological studies in mice revealed that NdBA caused less nuclear condensation and DNA damage than dBA did. TEM images showed the presence of NdBA in brain samples of NdBA fed mice, indicating its ability to cross the BBB., Conclusion: Thus, compared to dBA, NdBA appears to have greater chemoprotective potential against lung cancer.

Objective: To provide in vitro evidence of Psorinum treatment against cancer cells in a controlled study., Methods: Effects of homeopathic Psorinum 6× on cell viability were initially determined in several cancer cell lines, including A549, HepG2 and MCF-7, using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and an ethanol 6× control. The cell line that exhibited highest inhibition was selected and used in the following experiments. A range of Psorinum 6× doses was used to explore treatment effects on cell cycle arrest, cell death (apoptosis), generation of reactive oxygen species (ROS) and change in mitochondrial membrane potential (MMP) using flow cytometry and fluorescence microscopy, respectively. Expression of several signal proteins related to apoptosis and cell survival were quantified with Western blotting and confocal microscopy. Further, circular dichroism (CD) spectroscopy was used to determine possible drug-DNA interactions, as well as the induction of conformational changes., Results: Treatment of cancer cell lines with Psorinum showed greater anticancer effects in A549 cells than in others. In A549 cells Psorinum treatment inhibited cell proliferation at 24 h after treatment, and arrested cell cycle at sub-G1 stage. It also induced ROS generation, MMP depolarization, morphological changes and DNA damage, as well as externalization of phosphatidyl serine. Further, increases in p53 expression, Bax expression, cytochrome c release, along with reduction of Bcl-2 level and caspase-3 activation were observed after Psorinum 6× treatment, which eventually drove A549 cells towards the mitochondria-mediated caspase-3-dependent pathway. CD spectroscopy revealed direct interaction of Psorinum with DNA, using calf thymus-DNA as target., Conclusion: Psorinum 6× triggered apoptosis in A549 cells via both up- and down-regulations of relevant signal proteins, including p53, caspase-3, Bax and Bcl-2.

Objective: Methylation-specific epigenetic process and gene expression profiles of HeLa cells treated with ultra-high dilutions (HDs) of two plant extracts, Hydrastis canadensis (HC-30) and Marsdenia condurango (Condu-30), diluted 1060 times, were analyzed against placebo 30C (Pl-30) for alterations in gene profiles linked to epigenetic modifications., Methods: Separate groups of cells were subjected to treatment of Condu-30, HC-30, and Pl-30 prepared by serial dilutions and succussions. Global microarray data recorded on Affymetrix platform, using 25-mer probes were provided by iLifeDiscoveries, India. Slides were scanned with 3000 7G microarray scanner and raw data sets were extracted from Cel (raw intensity) files. Analyses of global microarray data profile, differential gene expression, fold change and clusters were made using GeneSpring GX12.5 software and standard normalization procedure. Before microarray study, concentration of RNA (ng/μL), RIN value and rRNA ratio for all the samples were analysed by Agilant Bioanalyzer 2100. Reverse transcriptase polymerase chain reaction (RT-PCR) and quantitative RT-PCR were done for analyzing SMAD-4 expression. Fluorescence-activated cell sorting study was further made to elucidate fate of cells at divisional stages. Methylation-specific restriction enzyme assay was conducted for ascertaining methylation status of DNA at specific sites., Results: HDs of HC-30 and Condu-30 differentially altered methylation in specific regions of DNA and expression profiles of certain genes linked to carcinogenesis, as compared to Pl-30. Two separate cut sites were found in genomic DNA of untreated and placebo-treated HeLa cells when digested with McrBC, compared to a single cut observed in Condu-30-treated genomic DNA. SMAD-4 gene expression validated the expression pattern observed in microarray profile. Methylation-specific restriction enzyme assay elucidated differential epigenetic modifications in drug-treated and control cells., Conclusion: HDs triggered epigenetic modifications and alterations in microarray gene expression profiles of many genes associated with carcinogenesis in HeLa cells in vitro.

Probiotic treatment has been gaining attention due to its remarkable effects in alleviating toxicity and carcinogenesis. The novel strain Pediococcus pentosaceus GS4 has been reported for probiotic, survivability in simulated gastrointestinal fluid, and antioxidative and biohydrogenation properties. Therefore, we hypothesize that this specific strain might be able to assuage the effect of azoxymethane (AOM)-induced toxicity in mice. Twenty-eight Swiss albino mice were divided into 4 groups and were studied for 32 weeks. Azoxymethane (10 mg/kg body weight) was administered intraperitoneally twice (0th and 14th days), and probiotic GS4 (1.1 × 10(9) colony-forming unit/mL) was given orally for the respective groups. Mice who served as the normal control received only normal saline. GS4-intervened AOM-induced mice showed marked improvement at the histopathologic level, in the liver and kidney. Moreover, probiotic GS4 intervention in AOM-induced mice exhibited a significant reduction in the liver function biomarker when compared with the AOM-induced mice. Probiotic GS4 intervention reduced the intestinal structural deformities as evident from the elevated brush border membrane-associated disaccharidases (sucrase, lactase) and intestinal alkaline phosphatase activities, which were found disrupted by AOM intoxication. Fecal bacterial load was found to be reduced in AOM-induced mice which were subsequently replenished by the probiotic GS4 intervention as apparent from the enhanced fecal bacterial load. There were no adverse effects observed in the probiotic control group. Conclusively, novel probiotic strain GS4 exhibited safe and beneficial effects against the toxicity threats posed by AOM. Thus, GS4 could be considered as a potential food supplement/additive for therapeutic purposes in gastrointestinal disorders related to inflammation and cancer., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Objectives: Condurango (Gonolobus condurango) extract is used by complementary and alternative medicine (CAM) practitioners as a traditional medicine, including homeopathy, mainly for the treatment of syphilis. Condurango bark extract is also known to reduce tumor volume, but the underlying molecular mechanisms still remain unclear., Methods: Using a cervical cancer cell line (HeLa) as our model, the molecular events behind condurango extract's (CE's) anticancer effect were investigated by using flow cytometry, immunoblotting and reverse transcriptase-polymerase chain reaction (RT-PCR). Other included cell types were prostate cancer cells (PC3), transformed liver cells (WRL-68), and peripheral blood mononuclear cells (PBMCs)., Results: Condurango extract (CE) was found to be cytotoxic against target cells, and this was significantly deactivated in the presence of N-acetyl cysteine (NAC), a scavenger of reactive oxygen species (ROS), suggesting that its action could be mediated through ROS generation. CE caused an increase in the HeLa cell population containing deoxyribonucleic acid (DNA) damage at the G zero/Growth 1 (G0/G1) stage. Further, CE increased the tumor necrosis factor alpha (TNF-α) and the fas receptor (FasR) levels both at the ribonucleic acid (RNA) and the protein levels, indicating that CE might have a cytotoxic mechanism of action. CE also triggered a sharp decrease in the expression of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB ) both at the RNA and the protein levels, a possible route to attenuation of B-cell lymphoma 2 (Bcl-2), and caused an opening of the mitochondrial membrane's permeability transition (MPT) pores, thus enhancing caspase activities., Conclusion: Overall, our results suggest possible pathways for CE mediated cytotoxicity in model cancer cells.

A pyridoxal-based fluorescent probe HL was synthesized for the detection of Cu(2+) in methanol with moderate selectivity. Upon addition of Cu(2+), to the solution of the probe in methanol exhibited a remarkable change in emission at 500 nm. With the limit of detection of 10 μM, the probe could well meet the recommended (less than 32 μM in drinking water) of the World Health Organization (WHO). The intracellular Cu(2+) imaging behaviour of HL was carried out on HeLa cells.

Controlled release of medications remains the most convenient way to deliver drugs. In this study, we precipitated gold nanoparticles with quercetin. We loaded gold-quercetin into poly(DL-lactide-co-glycolide) nanoparticles (NQ) and tested the biological activity of NQ on HepG2 hepatocarcinoma cells to acquire the sustained release property. We determined by circular dichroism spectroscopy that NQ effectively caused conformational changes in DNA and modulated different proteins related to epigenetic modifications and cell cycle control. The mitochondrial membrane potential (MMP), reactive oxygen species (ROS), cell cycle, apoptosis, DNA damage, and caspase 3 activity were analyzed by flow cytometry, and the expression profiles of different anti- and pro-apoptotic as well as epigenetic signals were studied by immunoblotting. A cytotoxicity assay indicated that NQ preferentially killed cancer cells, compared to normal cells. NQ interacted with HepG2 cell DNA and reduced histone deacetylases to control cell proliferation and arrest the cell cycle at the sub-G stage. Activities of cell cycle-related proteins, such as p21(WAF), cdk1, and pAkt, were modulated. NQ induced apoptosis in HepG2 cells by activating p53-ROS crosstalk and induces epigenetic modifications leading to inhibited proliferation and cell cycle arrest.

Background: Marsdenia condurango (condurango) is a tropical woody vine native to South America. Our earlier study was limited to evaluation of anti-cancer potentials of crude condurango extract and its glycoside-rich components in vitro on lung cancer., Objective: This study aims at evaluating the effect of the single isolated active ingredient condurangogenin A (ConA; C32H42O7) on A549, H522 and H460-nonsmall-cell lung cancer cells., Materials and Methods: ConA was isolated by column chromatography and analyzed by mass spectroscopy, Fourier transform infrared spectroscopy and proton-nuclear magnetic resonance. diphenyltetrazolium bromide assays were conducted on three cell-types using 6%-alcohol as control. Critical studies on cellular morphology, cell-cycle regulation, reactive oxygen species, mitochondrial membrane potential, and DNA-damage were made, and expressions of related signaling markers studied., Results: As IC50 doses of ConA proved to be too high and toxic to both A549 and H522 cells, all experimental studies were carried out on H460 cells with the IC50 dose (32 μg/ml - 24 h). Cellular morphology revealed typical apoptotic features after ConA treatment. At early treatment hours (2 h-12 h), maximum cells were arrested at G0/G1 phase that could be correlated with reduced level of cyclin D1-CDK with p21 up-regulation. At 18 h - 24 h, sub G0/G1 cell population was increased gradually, as revealed from cytochrome-c release and caspase-3 activation, further confirming the apoptosis-inducing ability of ConA at later phases. Gradual increase of TUNEL-positive cells with significant modulation of mitochondria-dependent apoptotic markers at longer time-points would establish apoptosis-induction property of ConA, indicating its potential as a strong candidate for anti-cancer drug formulation., Conclusion: Further studies are warranted against other types of cancer cells and animal models before its possible human use.

Objectives: Quercetin, a flavonoid compound, has been reported to induce apoptosis in cancer cells, but its anti-inflammatory effects, which are also closely linked with apoptosis, if any, on non-small-cell lung cancer (NSCLC) have not so far been critically examined. In this study, we tried to determine if quercetin had any demonstrable anti-inflammatory potential, which also could significantly contribute to inducing apoptosis in a NSCLC cell line, A549., Methods: In this context, several assays, including cytotoxicity, flow cytometry and fluorimetry, were done. Gene expression was analyzed by using a western blot analysis., Results: Results revealed that quercetin could induce apoptosis in A549 cells through mitochondrial depolarization by causing an imbalance in B-cell lymphoma 2/ Bcl2 Antagonist X (Bcl2/Bax) ratio and by down-regulating the interleukine-6/signal transducer and activator of transcription 3 (IL-6/STAT3) signaling pathway. An analysis of the data revealed that quercetin could block nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) activity at early hours, which might cause a down-regulation of the IL-6 titer, and the IL-6 expression, in turn, could inhibit p-STAT3 expression. Down-regulation of both the STAT3 and the NF-κB expressions might, therefore, cause down-regulation of Bcl2 activity because both are major upstream effectors of Bcl2. Alteration in Bcl2 responses might result in an imbalance in the Bcl2/Bax ratio, which could ultimately bring about mitochondria mediated apoptosis in A549 cells., Conclusion: Overall, the finding of this study indicates that a quercetin induced anti-inflammatory pathway in A549 cells appeared to make a significant contribution towards induction of apoptosis in NSCLC and, thus, may have a therapeutic use such as a strong apoptosis inducer in cancer cells.

Objective: Chemopreventive approach with natural products, particularly plants and plant-derived ones, is receiving increasing attention for their effective role against cancer without any palpable side effects. In this study, efficacy of ethanolic extract of Ruta graveolens (RG) on skin melanoma cells (A375) in vitro and on 7,12-dimethylbenz(a)anthracene (DMBA)-induced skin cancer in vivo has been tested in Swiss albino mice., Methods: Studies on cell viability, apoptosis and autophagy induction were conducted in vitro. To check apoptosis, assays like alteration in mitochondrial membrane potential, annexin V-fluorescein isothiocyanate/propidium iodide assay and immunoblot were performed. Fluorescence microscopic and immunoblot assays were performed to confirm autophagy induction. The effects of RG were determined by evaluating body weight, tumor incidence, tumor volume and tumor burden in mice. Enzymatic and non-enzymatic antioxidant status was assessed. The role of some relevant signaling proteins was also analyzed., Results: RG caused death of A375 cells through induction of caspase 3-mediated apoptosis and Beclin-1-associated autophagy. Moreover, RG administration (75 mg/kg body weight) which showed no acute or chronic toxicity, showed significant reduction in the skin tumor burden of DMBA-painted mice. RG also demonstrated potent anti-lipid peroxidative and antioxidant functions during the course of skin cancer induction by DMBA., Conclusion: Chemopreventive potential of RG was demonstrated from overall results of this study, indicating its possible use in therapeutic formulation of an effective drug to treat skin cancer.

Background: DNA hypermethylation induces cancer progression involving CpG island of DNA and causes inactivation of tumour suppressor genes. In this study, DNA hypermethylation status of lung cancer and ability of ultra-highly diluted Condurango 30C to modulate DNA methylation were ascertained by analysis of lung cancer-specific tumour suppressor genes in respect to placebo., Materials and Methods: DNA methylation status, if any, was determined by PCR-SSCP analyses in lung cancer-specific tumour suppressor genes (p15, p16 and p53) using H460-NSCLC cell and BaP-induced lung cancer of rats. The ability of Condurango 30C to modulate DNA methylation, if any, was verified against placebo control in blinded manner., Results: Condurango 30C-treated DNA showed significant decrease in band intensity of p15 and p53 genes especially in methylated condition in vitro, at IC50 dose (2.43µl/100µl). SSCP analysis of p15 and p53 genes in Condurango 30C-treated DNA also suggests that Condurango 30C can decrease methylation, in vitro. Inhibition of p15 hypermethylation was observed in post-cancer treatment of rats with Condurango 30C. SSCP results gave a better indication of differences in band position of p15 and p53 in Condurango 30C-treated lung samples., Conclusion: Condurango 30C could trigger epigenetic modification in lung cancer via modulation of DNA hypermethylation.

Increased evidence of chemo-resistance, toxicity and carcinogenicity necessitates search for alternative approaches for determining next generation cancer therapeutics and targets. We therefore tested the efficacy of plant alkaloid berberine on human papilloma virus (HPV) -18 positive cervical cancer cell HeLa systematically-involving certain cellular, viral and epigenetic factors. We observed disruptions of microtubule network and changes in membrane topology due to berberine influx through confocal and atomic force microscopies (AFM). We examined nuclear uptake, internucleosomal DNA damages, mitochondrial membrane potential (MMP) alterations and cell migration assays to validate possible mode of cell death events. Analytical data on interactions of berberine with pBR322 through fourier transform infrared (FTIR) and gel migration assay strengthen berberine׳s biologically significant DNA binding abilities. We measured cellular uptake, DNA ploidy and DNA strand-breaks through fluorescence activated cell sorting (FACS). To elucidate epigenetic modifications, in support of DNA binding associated processes, if any, we conducted methylation-specific restriction enzyme (RE) assay, methylation specific-PCR (MSP) and expression studies of histone proteins. We also analyzed differential interactions and localization of cellular tumor suppressor p53 and viral oncoproteins HPV-18 E6-E7 through siRNA approach. We further made in-silico approaches to determine possible binding sites of berberine on histone proteins. Overall results indicated cellular uptake of berberine through cell membrane depolarization causing disruption of microtubule networks and its biological DNA binding abilities that probably contributed to epigenetic modifications. Results of modulation in p53 and viral oncoproteins HPV-18 E6-E7 by berberine further proved its potential as a promising chemotherapeutic agent in cervical cancer., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Objective: Use of cisplatin, a conventional anticancer drug, is restricted because it generates strong hepatotoxicity by accumulating in liver. Therefore its anticancer potential can only be fully exploited if its own toxicity is considerably reduced. Towards this goal, ethanolic extract of the plant, Boldo (Peumus boldus), known for its antihepatotoxic effects, was used simultaneously with cisplatin, to test its ability to reduce cisplatin's cytotoxicity without affecting its anticancer potential., Methods: The cytotoxicity of Boldo extract (BE) and cisplatin, administered alone and in combination, was determined in three cancer cell lines (A549, HeLa, and HepG2) and in normal liver cells (WRL-68). Drug-DNA interaction, DNA damage, cell cycle, apoptosis, reactive oxygen species (ROS) and mitochondrial membrane potential (MMP, ΔΨ) were also studied. Hepatotoxicity and antioxidant activity levels were determined by alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase and glutathione assays in mice. The cytotoxicity of related proteins was tested by Western blotting., Results: Co-administration of BE and cisplatin increased viability of normal cells, but had no effect on the viability of cancer cells. Boldo protected liver from damage and normalized different antioxidant enzyme levels in vivo and also reduced ROS and re-polarized MMP in vitro. Bax and cytochrome c translocation was reduced with caspase 3 down-regulation. Further, a drug-DNA interaction study revealed that BE reduced cisplatin's DNA-binding capacity, resulting in a reduction in DNA damage., Conclusion: Results indicated that a low dose of BE could be used beneficially in combination with cisplatin to reduce its toxicity without hampering cisplatin's anticancer effect. These findings signify a potential future use of BE in cancer therapy.

A diformyl-p-cresol (DFC)-8-aminoquinoline based dual signaling probe was found to exhibit colorimetric and fluorogenic properties on selective binding towards Mg(2+) and Zn(2+). Turn-on fluorescent enhancements (FE) as high as 40 fold and 53 fold in 9 : 1 MeCN/water (v/v) at pH 7.2 in HEPES buffer for Mg(2+) and Zn(2+), respectively, were observed. The binding constants determined from the fluorescence titration data are: K = (1.52 ± 0.21) × 10(5) M(-1) and (9.34 ± 4.0) × 10(3) M(-2) at n = 1 and 0.5, for Mg(2+) and Zn(2+), respectively. The L : M binding ratios were also determined by Job's method, which support the above findings. This is further substantiated by HRMS analysis. Due to solubility in mixed organo-aqueous solvents as well as cell permeability it could be used for the in vitro/in vivo cell imaging of Mg(2+) and Zn(2+) ions with no or negligible cytotoxicity. This probe could be made selective towards Mg(2+) over Zn(2+) in the presence of TPEN, both under intra- and extracellular conditions and is superior to other Mg(2+) probes which suffer from selectivity of Mg(2+) over Ca(2+) or Zn(2+). Furthermore the dissociation constant (Kd = 6.60 μM) of the Mg(2+)-() complex is far lower than the so far reported Mg(2+) probes which fall in the mM range.

Anti-cancer drugs generally kill cancer cells by apoptosis but fail to do so when they become resistant and escape apoptosis signals. But these resistant cells can still be killed by autophagy. Therefore, drugs having both apoptotic and autophagic abilities are solicited in effective cancer management. In search of such a drug, we examined the efficacy of graveoline, a bioactive compound isolated from Ruta graveolens on skin melanoma A375 cells through the use of specific signaling cascades and their inhibitors. Cytotoxicity of graveoline was tested by conducting MTT assay. Induction of autophagy and apoptosis was checked. Expression of related proteins and their localization were studied by conducting immunoblot assay and through confocal microscopy, respectively. We found graveoline-induced Beclin-1 associated autophagy in A375 cells and 3-methyladenine, an inhibitor of autophagy did not affect apoptosis. Conversely, caspase inhibitor that blocked apoptosis did not affect autophagic cell death, suggesting thereby that these two were independent events. Use of reactive oxygen species (ROS) scavengers inhibited cell death, but blocking autophagy did not affect graveoline-induced ROS generation, suggesting that ROS generation ensued autophagy. Thus, graveoline-induced both apoptotic and autophagic cell death in skin melanoma cells, a desirable quality in effective anti-cancer drug design., (Copyright © 2013 John Wiley & Sons, Ltd.)

Objective: Conium maculatum extract is used as a traditional medicine for cervix carcinoma including homeopathy. However, no systematic work has so far been carried out to test its anti-cancer potential against cervix cancer cells in vitro. Thus, in this study, we investigated whether ethanolic extract of conium is capable of inducing cytotoxicity in different normal and cancer cell lines including an elaborate study in HeLa cells., Materials and Methods: Conium's effects on cell cycle, reactive oxygen species (ROS) accumulation, mitochondrial membrane potential (MMP) and apoptosis, if any, were analyzed through flow cytometry. Whether Conium could damage DNA and induce morphological changes were also determined microscopically. Expression of different proteins related to cell death and survival was critically studied by western blotting and ELISA methods. If Conium could interact directly with DNA was also determined by circular dichroism (CD) spectroscopy., Results: Conium treatment reduced cell viability and colony formation at 48 h and inhibited cell proliferation, arresting cell cycle at sub-G stage. Conium treatment lead to increased generation of reactive oxygen species (ROS) at 24 h, increase in MMP depolarization, morphological changes and DNA damage in HeLa cells along with externalization of phosphatidyl serine at 48 hours. While cytochrome c release and caspase-3 activation led HeLa cells toward apoptosis, down-regulation of Akt and NFkB inhibited cellular proliferation, indicating the signaling pathway to be mediated via the mitochondria-mediated caspase-3-dependent pathway. CD-spectroscopy revealed that Conium interacted with DNA molecule., Conclusion: Overall results validate anti-cancer potential of Conium and provide support for its use in traditional systems of medicine.

Toxic effects of cadmium (Cd) were examined on a cadmium-resistant strain of Aspergillus foetidus isolated from wastewater. The Cd removal potential was analyzed. The results indicated that the strain could tolerate up to 25 mM and 63 mM Cd in liquid and solid Czapek-Dox media, respectively. It efficiently removed Cd from liquid growth media and industrial wastewater by mycelial biosorption. The strain produced oxalic acid for the purpose of Cd bioleaching as confirmed by the presence of cadmium oxalate crystals on the mycelial surface. Intracellular proline contents and the antioxidative enzyme activities increased up to a certain level to detoxify the overproduced free radicals. These data indicate that the strain has inherent mechanisms to grow in Cd contaminated environment, tolerate high Cd doses and high Cd uptake potential which are pre-requisite for acting as a suitable candidate for Cd bioremediation., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Objectives: Condurango is widely used in various systems of complementary and alternative medicines (CAM) against oesophageal and stomach ailments including certain types of cancer. However, until now no systematic study has been conducted to verify its efficacy and dose with proper experimental support. Therefore, we examined if ethanolic extract of Condurango could ameliorate benzo[a]pyrene (BaP)-induced lung cancer in rats, in vivo to validate its use as traditional medicine., Methods: Fifteen male and 15 female Sprague-Dawley (SD) rats were treated with 0.28 mg/kg of Sweet Bee Venom (SBV) (high-dosage group) and the same numbers of male and female SD rats were treated with 0.2 mL/kg of normal saline (control group) for 13 weeks. We selected five male and five female SD rats from the high-dosage group and the same numbers of male and female SD rats from the control group, and we observed these rats for four weeks. We conducted body-weight measurements, ophthalmic examinations, urinalyses and hematology, biochemistry, histology tests., Results: A histological study revealed gradual progress in lung tissue-repair activity in Condurango-fed cancer-bearing rats, showing gradual tissue recovery after three months of drug administration. Condurango has the capacity to generate reactive oxygen species (ROS), which may contribute to a reduction in anti-oxidative activity and to an induction of oxidative stress-mediated cancer cell-death. Condurango-activated pro-apoptotic genes (Bax, caspase-3, caspase-9, p53, cytochrome-c, apaf-1, ICAD and PARP) and down-regulated antiapoptotic-Bcl-2 expression were noted both at mRNA and protein levels. Studies on caspase-3 activation and PARP cleavage by western blot analysis revealed that Condurango induced apoptosis through a caspase-3-dependent pathway., Conclusion: The anticancer efficacy of an ethanolic extract of Condurango for treating BaP-induced lung cancer in rats lends support for its use in various traditional systems of medicine.

We isolated a triterpenoid from an ethanolic extract of Phytolacca decandra and nanoencapsulated it with biodegradable nontoxic polymers of poly(lactide-co-glycolide) to examine if the nanoform of this hitherto unexplored betulinic-acid derivative (NdBA) could produce a stronger anticancer effect by rendering better drug bioavailability and targeted delivery than the nonencapsulated betulinic-acid derivative (dBA). The nanoparticles were characterized with the help of physicochemical and morphological studies involving dynamic light scattering and atomic force microscopy. A549 cancer cells were exposed to NdBA and dBA at the IC50 doses of 50 μg/mL and 100 μg/mL, respectively. Mitochondrial dysfunction-mediated apoptosis was determined by examining the changes in the intracellular calcium content, the reactive oxygen species accumulation, the cytochrome c release, the upregulation of Bcl-2-associated-X protein (Bax) and caspase 3, the downregulation of B cell lymphoma 2, and the mitochondrial membrane potential (ΔΨm) depolarization. Apoptosis was also verified by acridine orange staining observed under fluorescence microscopy and annexin V-fluorescein isothiocyanate/propidium iodide staining through flow cytometric studies. The levels of intracellular adenosine triphosphate/adenosine diphosphate ratio decreased, and the ATPase activity increased more strikingly in A549 cells exposed to NdBA than in A549 cells exposed to dBA. Overall results showed that both drugs directly target the mitochondrial oxidative phosphorylation system, with NdBA having a stronger effect, indicating NdBA to be a better candidate for the development of an anticancer drug for use against lung adenocarcinomas., (Copyright © 2013. Published by Elsevier B.V.)

The effect of substituents on FRET in two newly designed rhodamine-based Hg(2+) ion selective chemosensors (L¹ and L²) has been explored by a systematic experimental and theoretical study. Comparison of these sensors in the analytical study and imaging of Hg(2+) ions in living cells has also been included.

Activation of signal transducer and activator of transcription3 (STAT3) is a hallmark of several types of cancer. Failure to inhibit STAT3 expression by injection of siRNA for STAT3 directly to Balb/c mice led us to adopt alternative means. We formulated nanoparticle-based encapsulation of siRNA (NsiRNA) with polyethylenimine (PEI) and poly(lactide-co-glycolide) (PLGA) and characterized them. The siRNA treated and NsiRNA-treated cells were subjected separately to different assay systems. We also checked if NsiRNA could cross the blood brain barrier (BBB). Cell viability reduced dramatically in A549 cells after NsiRNA administration (23.89% at 24 h), thereby implicating considerable silencing of STAT3 by NsiRNA, but not after siRNA administration. Compared to controls, a significant decrease in expression of IL-6 and the angiogenic factor (VEGF) and increase in Caspase 3 activity was observed with corresponding regression in tumor growth in mice treated with NsiRNA. NsiRNA induced apoptosis of cells and arrested cells at G1/G0 stage, both in vitro and in vivo. Apoptosis was also verified by Annexin-V-FITC/Propidium-iodide staining. NsiRNA could cross blood brain barrier. Overall results revealed PEI-PLGA to be a promising carrier for delivery of siRNA targeting STAT3 expression, which can be utilized as an effective strategy for cancer therapy., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Objective: Oleanolic acid (OA) has been reported to have anticancer effects, but the extent of its cytotoxicity, its ability to interact with nuclear DNA, its action against skin melanoma, as well as the molecular mechanism of its action against cell proliferation and in support of cell death are still unexplored. This led us to examine the efficacy of OA, a bioactive compound isolated from Phytolacca decandra, on these issues in the present investigation., Methods: Studies related to analyses of cell viability, drug-DNA interaction, cell proliferation, cell cycle and epidermal growth factor receptor (EGFR) activity were performed. To investigate whether cells undergo apoptosis, studies like fluorescence microscopy, poly (ADP-ribose) polymerase (PARP) degradation, annexin V-fluorescein isothiocyanate/propidium iodide assay, alteration in mitochondrial membrane potential and activity of some relevant signaling proteins were performed., Results: OA displayed a minimal and negligible cytotoxic effect on normal HaCaT cells (skin keratinocytes) and peripheral blood mononuclear cells but by contrast it reduced A375 cell viability significantly. OA interacted with nuclear DNA quickly after exposure. It acted as an anti-proliferative agent. It suppressed EGFR activity. OA administration led the cells to mitochondria-dependent caspase 3-mediated apoptosis., Conclusion: OA interacts with cellular DNA, inhibits proliferation possibly through modulating EGFR activity and induces mitochondria-dependent caspase 3-mediated apoptosis in A375 cells which would qualify it as a potent anticancer agent.

We report, herein, the development of an easily synthesizable novel dansyl-based turn-on NO sensor L2. The UV-Vis titration data of L2 with Cu(2+) display a gradual increase in absorbance at 418 nm with [Cu(2+)], which were analyzed by using a non-linear least-squares computer-fit program yielding K = (1.16 ± 0.36) × 10(6) M(-1) and n = (1.28 ± 0.03) indicating a 1 : 1 complexation. The ground state geometries of L2 as well as its complex [Cu(L2)Cl](+) (1) were optimized by DFT calculations which showed that in complex 1 the central metal ion is in distorted tetrahedral geometry with bond distances very close to those found in analogous Cu(2+) complexes. The fluorescence of L2 was dramatically quenched (∼60-fold) through complexation with paramagnetic Cu(2+) to form [Cu(L2)Cl](+) in MeCN-H2O (9 : 1, v/v) at pH 7.2 in HEPES buffer, which on further treatment with Angeli's salt (Na2N2O3) restores its fluorescence property by ∼15-fold due to the reduction of Cu(2+) to Cu(+) by NO generated in solution from Na2N2O3. The lifetime measurements displayed a substantial decrease in the lifetime of free ligand L2 (τ0 = 12 ns) on complexation with Cu(2+) (τ0 = 2.1 ns). The detection limit of NO calculated by the 3σ method gives a value of 1.6 nM. The NO induced fluorescence enhancement of [Cu(II)(L2)Cl](+) was due to the reduction of [Cu(II)(L2)Cl](+) (1) to [Cu(I)(L2)](+) (2) and is supported by the disappearance of the d-d transition band at 850 nm as well as the X-band EPR signal of 1. The selective "turn on" fluorogenic behavior of L2 was examined on HeLa cells of human cervical cancer origin by fluorescence microscopy which showed very intense intracellular fluorescence that was strongly suppressed by the addition of Cu(2+) but it regains its fluorescence property on further incubation with Angeli's salt (Na2N2O3). The existence of [Cu(II)(L2)Cl](+) and [Cu(I)(L2)](+) in solution was confirmed by ESI-MS(+) (m/z) analysis. The effect of different biologically relevant cations and anions on the fluorescence property of L2 indicates that it was only the [Cu(II)(L2)Cl](+) which displayed high selectivity for NO, indicating its suitability for intracellular application without much worry about its cytotoxicity in a specified dose.

Objectives: The K-ras gene mutation commonly found in lung adenocarcinomas contributes to their non-invasive expansion. Our main objective here was to develop a chemopreventive agent against K-ras-mutated lung adenocarcinoma cell line like-A549., Materials and Methods: We isolated flavonol from ethanolic leaf extract of Thuja occidentalis, and evaluated its apoptotic potentials on A549 cells. They were treated with 1-10 μg/ml of flavonol and viability was tested retaining normal lung cells L-132 as control. We performed assays such as TUNEL, annexin V, cell-cycle and mitochondrial membrane potentials, by FACS analysis. ROS-mediated oxidative stress and drug-DNA interactions were analysed along with gene expression studies for p53, Bax-Bcl2, cytochrome c, the caspase cascade genes and PARP., Results: Flavonol reduced A549 cell viability in a dose- and time-dependent manner (IC50 value = 7.6 ± 0.05 μg/ml following 48 h incubation) sparing normal L-132 cells. It effected G2-M phase cell cycle arrest and apoptosis, as indicated by progressive increase in the sub-G1, annexin V and TUNEL-positive cell populations. Apoptotic effects appeared to be mitochondria-dependent, caspase-3-mediated, but ROS-independent. Analysis of circular dichroism data revealed that flavonol intercalated with nuclear DNA. In vivo studies on non small cell lung carcinoma (NSCLC)-induced mice confirmed anti-cancer potential of flavonol., Conclusion: Flavonol-induced apoptosis apparently resulted from intercalation of cells' nuclear DNA. Flavonol inhibited growth of induced lung tumours in the mice, indicating its potential as an effective agent against NSCLC., (© 2013 John Wiley & Sons Ltd.)

A new chelating ligand [4-methyl-2,6-bis-(pyridin-2-yl-hydrazonomethyl)-phenol] (1) was prepared by the condensation of 2-hydrazinylpyridine with 2,6-diformyl-p-cresol. Compound 1 exhibits weak fluorescence due to intramolecular photoinduced electron transfer (PET). The sensor (1) demonstrates Zn(2+)-specific emission enhancement due to the “PET off” process through a 1:1 binding mode with the metal ion. The fluorescence quantum yield of chemosensor 1 is only 0.020, and it increases more than 14-fold (0.280) in the presence of one equivalent of the zinc ion. Interestingly, the introduction of other metal ions causes the fluorescence intensity to remain either unchanged or weakened except for Cd(2+). The new sensor showed ‘naked-eye’ detection of Zn(2+) ions: a color change of the solution from colorless to yellow. Ratiometric displacement of Cd(2+) ions from the complex by Zn(2+) ions supports the formation of a more stable sensor–Zn(2+) complex over the sensor–Cd(2+) complex. The experimental findings have been correlated with theoretical results using the B3LYP functional and 6-31G (d, p), LANL2DZ basis set for Cd(2+) (2) and Zn(2+) (3) complexes, respectively, by the Density Functional Theory (DFT) method. Moreover, the ability of probe 1 to sense Zn(2+) within human melanoma cancer cells has been explored, and the Zn(2+)-probing process in living cells was found to be reversible with zinc chelator solution of N,N,N,N-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) or EDTA.

Chemotherapeutic potential of Condurango glycoside-rich components (CGS) was evaluated in NSCLC, in vitro and in BaP-intoxicated rats, in vivo. NSCLC cells were treated with different concentrations of CGS to test their effect on cell viability. Cellular morphology, DNA-damage, AnnexinV-FITC/PI, cell cycle regulation, ROS-accumulation, MMP, and expressions of related signalling genes were critically analysed. 0.22 μg/μl CGS (IC₅₀ dose at 24 h) was selected for the study. CGS-induced apoptosis via DNA damage was evidenced by DNA-ladder formation, increase of AnnexinV-positive cells, cell cycle arrest at subG0/G1 and differential expressions of apoptotic genes. ROS-elevation and MMP-depolarization with significant caspase-3 activation might lead to apoptotic cell death. Anti-proliferative activity was confirmed by EGFR-expression modulation. ROS accumulation and DNA-nick formation with tissue damage-repair activity after post-cancerous CGS treatment, in vivo, supported the in vitro findings. Overall results advocate considerable apoptosis-inducing potential of CGS against NSCLC, validating its use against lung cancer by CAM practitioners., (Copyright © 2013 Elsevier B.V. All rights reserved.)

In recent years, there has been growing interest in cancer prevention by food plants and their products. Although several plant parts have potentials for chemoprevention and other therapeutic use, their molecular mechanisms of action are not always well understood. Extensive research has identified several molecular targets that can potentially be used for the prevention and/or treatment of cancer. In this review, we accumulate evidences of modulating abilities of some dietary plants and their products on several signaling pathways, including the inflammatory and apoptotic ones, which may be targeted for cancer therapy. We have mainly focused on several phytochemicals like resveratrol (red grapes and peanuts), allicin (garlic), lycopene (tomato), indole-3-carbinol (cruciferous vegetables), vitamin C (citrus fruits), [6]-gingerol (ginger), emodin (aloe), natural antioxidant mixture (spinach), beta carotenoids (carrots), sulphoraphane (mustard), ellagic acid (pomegranate), myrecitin (cranberry), carnosol (rosemary), vanillin (vanilla) and eugenol (cloves). They act through one or more signaling pathways like nuclear factor kappa B, cyclooxygenase-2, signal transducer and activator of transcription 3, Akt, mitogen activated protein kinase/extracellular regulated kinase, Bcl-2, caspases, poly (ADP-ribose) polymerase, matrix metalloproteinase 2/9, and cyclin D1. Critical knowledge on these compounds and their signaling pathways may help in formulation of effective anticancer drugs.

Objectives: Whether the ultra-highly-diluted remedies used in homeopathy can effectively bring about modulations of gene expressions through acetylation/deacetylation of histones has not been explored. Therefore, in this study, we pointedly checked if the homeopathically-diluted anti-cancer remedy Condurango 30C (ethanolic extract of Gonolobus condurango diluted 10(-60) times) was capable of arresting the cell cycles in cervical cancer cells HeLa by triggering an epigenetic modification through modulation of the activity of the key enzyme histone deacetylase 2 vis-a-vis the succussed alcohol (placebo) control., Methods: We checked the activity of different signal proteins (like p21(WAF), p53, Akt, STAT3) related to deacetylation, cell growth and differentiation by western blotting and analyzed cell-cycle arrest, if any, by fluorescence activated cell sorting. After viability assays had been performed with Condurango 30C and with a placebo, the activities of histone de-acetylase (HDAC) enzymes 1 and 2 were measured colorimetrically., Results: While Condurango 30C induced cytotoxicity in HeLa cells in vitro and reduced HDAC2 activity quite strikingly, it apparently did not alter the HDAC1 enzyme; the placebo had no or negligible cytotoxicity against HeLa cells and could not alter either the HDAC 1 or 2 activity. Data on p21(WAF), p53, Akt, and STAT3 activities and a cell-cycle analysis revealed a reduction in DNA synthesis and G1-phase cell-cycle arrest when Condurango 30C was used at a 2% dose., Conclusion: Condurango 30C appeared to trigger key epigenetic events of gene modulation in effectively combating cancer cells, which the placebo was unable to do.