Search

Objectives: In homeopathy, it is claimed that more homeopathically-diluted potencies render more protective/curative effects against any disease condition. Potentized forms of Condurango are used successfully to treat digestive problems, as well as esophageal and stomach cancers. However, the comparative efficacies of Condurango 6C and 30C, one diluted below and one above Avogadro’s limit (lacking original drug molecule), respectively, have not been critically analyzed for their cell-killing (apoptosis) efficacy against lung cancer cells in vitro, and signalling cascades have not been studied. Hence, the present study was undertaken. Methods: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylte- trazolium bromide (MTT) assays were conducted on H460-non-small-cell lung cancer (NSCLC) cells by using a succussed ethyl alcohol vehicle (placebo) as a control. Studies on cellular morphology, cell cycle regulation, generation of reactive oxygen species (ROS), changes in mitochondrial membrane potential (MMP), and DNA-damage were made, and expressions of related signaling markers were studied. The observations were done in a “blinded” manner. Results: Both Condurango 6C and 30C induced apoptosis via cell cycle arrest at subG0/G1 and altered expressions of certain apoptotic markers significantly in H460 cells. The drugs induced oxidative stress through ROS elevation and MMP depolarization at 18-24 hours. These events presumably activated a caspase-3-mediated signalling cascade, as evidenced by reverse transcriptase- polymerase chain reaction (RT-PCR), western blot and immunofluorescence studies at a late phase (48 hours) in which cells were pushed towards apoptosis. Conclusion: Condurango 30C had greater apoptotic effect than Condurango 6C as claimed in the homeopathic doctrine.

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.

Triazole motif linked rhodamine derivatives 1 and 2 were synthesized. Chemosensor 1 recognizes both Hg2+ and Pb2+ ions in CH3CN–water (4/1, v/v; 10 μM tris HCl buffer, pH 7.0) both colorimetrically and fluorometrically. In addition, chemosensor 1 is cell permeable and detects intercellular Hg2+ and Pb2+ ions. The aqueous phase recognition of these ions was also studied with a resin bound sensor 2 which exhibits more sensitivity and selectivity towards Hg2+ over Pb2+ ion.

A new water-soluble rhodamine-based Al3+ ion-selective probe (L1) was synthesised and characterized by physico-chemico and spectroscopic tools. In the presence of a large excess of other competing ions, L1 specifically binds Al3+ ions with a concurrent visually observable change from colorless to pink in electronic spectral behavior, making it possible to detect the presence of Al3+ ions with the naked eye. The addition of Al3+ ions to a solution of L1 in HEPES buffer (1 mM, pH 7.4, 2% EtOH) at 25 °C, results in a decrease in the weak fluorescence intensity at λem = 470 nm, while a new peak (at λem = 588 nm) increases gradually through a fluorescence resonance energy transfer process. This ratiometric enhancement helps to detect Al3+ ions at a very low concentration of 33 nM. The detection limit of L1 for Al3+ ions was estimated to be 6.19 × 10−9 M using the 3σ method. This probe is also useful for imaging Al3+ ions in HeLa cells.

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.

A benzimidazole-based compound 1 built on the piperazine motif has been designed and synthesized. The chemosensor 1 is highly selective and sensitive towards ATP over ADP and AMP in CH3CN/H2O (1 : 1, v/v, using 10 mM HEPES buffer, pH 6.4) as evidenced by the significant change in emission. Furthermore, sensor 1 is cell permeable and can detect the presence of ATP in Human cervical cancer cells (HeLa).

A newly designed organic moiety, 5H-5,7a,12-triaza-dibenzo[a,e]azulen-6-one (L) containing a seven membered ring behaves as a hydrogen sulphate ion selective ratiometric chemosensor. The formulation and detailed structural characterisation of L have been established using physico-chemical, spectroscopic tools and single crystal X-ray diffraction study. On additions of hydrogen sulphate ions to the solution of L in HEPES buffer (1 mM; water : ethanol (v/v), 98 : 2) at 25 °C at biological pH, a new fluorescence peak generated at 483 nm was increased with concomitant decrease of the weak fluorescence of L at 430 nm through an isoemissive point at 449 nm due to the selective binding of HSO4− ions with L in a 1 : 1 ratio with a binding constant (K) of 4.13 × 106 M−1, and detects HSO4− ions as low as 5.5 × 10−7 M. The ratiometric enhancement of the fluorescence is based on intermolecular hydrogen bonding assisted chelation enhanced fluorescence (CHEF) process which has been evidenced by 1HNMR titration and supported by theoretical (DFT) calculations. The probe (L) having no cytotoxic effect is also useful for the detection of intracellular HSO4− ion concentrations under a fluorescence microscope.

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.

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.

A new highly sensitive and selective Hg(II) probe, 2-(rhodamine-b-hydrazido)-N-(quinolin-8-yl)acetamide (L1) was developed and characterized. L1 specifically binds to Hg(II) in the presence of a large excess of other competing ions with visually observable changes in both electronic and fluorescence spectral behaviour to make possible the naked eye detection of Hg(II) at a very low level (up to 4.5 × 10−7 M) through a fluorescence resonance energy transfer (FRET) process in HEPES buffer (1 mM, pH 7.4; 2% EtOH) at 25 °C. The theoretical and experimental kinetic study also support the binding of Hg(II) ion to induce the opening of the spirolactam ring in L1 for enabling the FRET process. Further studies reveal that the selective dissociation of the L–Hg complex in the presence of sulphide anions to restore the native structure of L1 is also useful in the detection of sulfide anions with a detection limit of a submicromolar range in the same medium of HEPES buffer (1 mM, pH 7.4; 2% EtOH) at 25 °C. L1 could be employed as a FRET based time dependent reversible chemosensor for imaging Hg(II) in living cells and whole bodies, and also could be used as an imaging probe for the detection of sulfide anions in HeLa cells.

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.

Skin cancer is increasing at an alarming rate and becoming resistant to conventional chemotherapy necessitating improved drug delivery system. We loaded apigenin (Ap), a dietary flavonoid having anti-cancer property, with poly (lactic-co-glycolide) (PLGA) nanoparticles (NAp) to explore if nano-encapsulation could enhance anti-carcinogenic effect against ultra-violet B (UVB) and Benzo(a)pyrene (BaP) induced skin tumor and mitochondrial dysfunction in mice. Particle size, morphology and zeta potential of NAp were determined using dynamic light scattering and atomic force microscopy. Tumor incidence and multiplicity in UVB-BaP induced mice with/without NAp treatment were ascertained and their histolopathological sections and chromosomal aberrations were studied. ROS accumulation and mitochondrial functioning through relevant markers like mitochondrial transmembrane potential were analyzed. Mitochondrial volume changes/swelling, cytochrome c (cyt c) release, mRNA and protein expressions of Apaf-1, bax, bcl-2, cyt c, cleaved caspase-9 and 3 were studied. Results showed that NAp produced better effects than Ap, due to their smaller size, and faster mobility. NAp reduced tissue damage and frequency of chromosomal aberrations, increased ROS accumulation to mediate mitochondrial-apoptosis through modulation of several apoptotic markers and mitochondrial matrix swelling. NAp showed ameliorative potentials in combating skin cancer and therefore has greater prospect of use in therapeutic management of skin cancer.

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 p21WAF, 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 p21WAF, 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.

Chemical fusion of a naturally occurring pharmacophore (pterocarpan) with a fluorophore (acridone) by thermally induced cascade sigmatropic rearrangement leads to the formation of a structurally different class of DNA binding ligands. Competitive binding assay in the presence of a classical minor groove binder shows the possibility of DNA minor groove binding by the ligand 4a. High cell viability of the prepared molecular probes was of utility for nuclear staining as well as cell cycle analysis through fluorescence-activated cell sorting (FACS).

Objective To examine if the ultra-highly diluted homeopathic remedy Thuja 30C can ameliorate benzo(a)pyrene (BaP)-induced DNA damage, stress and viability of perfused lung cells of Swiss albino mice in vitro. Methods Perfused normal lung cells from mice were cultured in 5% Roswell Park Memorial Institute medium and exposed to BaP, a potent carcinogen, at the half maximal inhibitory concentration dose (2.2 μmol/L) for 24 h. Thereafter, the intoxicated cells were either treated with Thuja 30C (used against tumor or cancer) or its vehicle media, succussed alcohol 30C. Relevant parameters of study involving reactive oxygen species (ROS) accumulation, total glutathione (GSH) content, and generations of heat shock protein (hsp)-90 were measured; the cell viability and other test parameters were measured after treatment with either Thuja 30C or its vehicle media. Circular dichroism spectroscopy was performed to examine if Thuja 30C directly interacted with calf thymus DNA as target. For ascertaining if DNA damaged by BaP could be partially repaired and restituted by the remedy, 4′,6-diamidino-2-phenylindole staining was performed. Results Thuja 30C increased cell viability of BaP-intoxicated cells significantly, as compared to drug-untreated or drug-vehicle control. A minimal dose of Thuja 30C significantly inhibited BaP-induced stress level, by down-regulating ROS and hsp-90, and increasing GSH content. Thuja 30C itself had no DNA-damaging effect, and no direct drug-DNA interaction. However, it showed quite striking ability to repair DNA damage caused by BaP. Conclusion Thuja 30C ameliorates BaP-induced toxicity, stress and DNA damage in perfused lung cells of mice and it apparently has no effect on normal lung cells.

A new rhodamine-based dual signaling probe (L3) has been found to display quick responses through visible colorimetric changes as well as fluorogenic properties on selective 1:1 binding to Hg2+, as delineated by absorption and fluorescence titrations as well as by Job′s method and ESI-MS+ studies. The fluorescent probe L3 displays a 252-fold fluorescence enhancement on binding to Hg2+. A Benesi–Hilderband fit of the absorption titration data gives Kd = (32.01 ± 0.74) μM and a 1:1 binding stoichiometry. Owing to its solubility in mixed organic/aqueous solvents as well as its cell permeability, L3 could be used for in vitro/in vivo cell imaging of Hg2+ with no or negligible cytotoxicity. The detection limit of Hg2+ was calculated by the 3σ method to be 9.28 ng L–1. Moreover, the fluorescence of the L3–Hg2+ system was turned off and its red colour disappeared when KI was added. Thus, the “OFF–ON–OFF” fluorescence sensing behaviour of L3 observed in the presence of Hg2+ and I– may find applications in devices with logic gate functions.

OBJECTIVES The present investigation aimed at examining if post-cancer treatment with a potentized homeopathic drug, Condurango 30C, which is generally used to treat oesophageal cancer, could also show an ameliorating effect through apoptosis induction on lung cancer induced by benzo[a]pyrene (BaP) in white rats (Rattus norvegicus). METHODS Lung cancer was induced after four months by chronic feeding of BaP to rats through gavage at a dose of 50 mg/kg body weight for one month. After four months, the lung-cancer-bearing rats were treated with Condurango 30C for the next one (5(th)), two (5(th)-6(th)) and three (5(th)-7(th)) months, respectively, and were sacrificed at the corresponding time- points. The ameliorating effect, if any, after Condurango 30C treatment for the various periods was evaluated by using protocols such as histology, scanning electron microscopy (SEM), annexinV-FITC/PI assay, flow cytometry of the apoptosis marker, DNA fragmentation, reverse transcriptase-polymerase chain reaction (RT-PCR), immunohistochemistry, and western blot analyses of lung tissue samples. RESULTS Striking recovery of lung tissue to a near normal status was noticed after post-cancerous drug treatment, as evidenced by SEM and histology, especially after one and two months of drug treatment. Data from the annexinV-FITC/PI and DNA fragmentation assays revealed that Condurango 30C could induce apoptosis in cancer cells after post-cancer treatment. A critical analysis of signalling cascade, evidenced through a RT-PCR study, demonstrated up-regulation and down-regulation of different pro- and anti-apoptotic genes, respectively, related to a caspase-3-mediated apoptotic pathway, which was especially discernible after one-month and two- month drug treatments. Correspondingly, Western blot and immunohistochemistry studies confirmed the ameliorative potential of Condurango 30C by its ability to down-regulate the elevated epidermal growth factor receptor (EGFR) expression, a hallmark of lung cancer. CONCLUSION The overall result validated a positive effect of Condurango 30C in ameliorating lung cancer through caspase-3-mediated apoptosis induction and EGFR down-regulation.

Poor oral bioavailability of chelidonine, a bio-active ingredient of Chelidonium majus, showing anti-cancer potentials against cancer cells with multidrug resistance, makes its optimal use rather limited. To address this problem, we encapsulated chelidonine in biodegradable poly(lactide-co-glycolide) (PLGA) polymers and evaluated nano-chelidonine's (NCs) anti-cancer efficacy vis-à-vis free chelidonine (FC) against HepG2 cells and also evaluated its bioavailability in mice. Physicochemical characteristics indicated that stable spherical NC were formed in nanometer size range (123±1.15 nm) with good yield (86.34±1.91%), better encapsulation efficiency (82.6±0.574%), negative surface charge (-19.6±2.48 mV) and ability of prolonged and sustained release of chelidonine. Fourier transform infrared analysis revealed that NC resembled similar peaks as that of FC suggesting effective encapsulation in PLGA. NC exhibited rapid cellular uptake and stronger apoptotic effect (∼46.6% reduced IC₅₀ value) than FC, blocking HepG2 cells at G2/M phase. p53, cyclin-D1, Bax, Bcl-2, cytochrome c, Apaf-1, caspase-9 and caspase-3 expressions also corroborated well to suggest greater anticancer potentials of NC. Our in vivo studies demonstrated NC to be more bio-available than FC and showed a better tissue distribution profile without inducing any toxicity (100 mg/kg bw) in mice. Unlike FC, NC could permeate into brain tissue, indicating thereby NC's better potentials for use in therapeutic oncology.

Cancer is a disease that needs a multi-faceted approach from different systems of medicine. The purpose of this study was to evaluate whether homeopathically-potentized ultra-high dilutions of Lycopodium Clavatum (LC-5C and LC-15C, respectively) have any anti-cancer effects on HeLa cells. Cells were exposed to either LC-5C (diluted below Avogadro's limit, i.e., 10−10) or LC-15C (diluted beyond Avogadro's limit, i.e., 10−30) (drug-treated) or to 30% succussed ethanol (“vehicle” of the drug). The drug-induced modulation in the percent cell viability, the onset of apoptosis, and changes in the expressions of Bax, Bcl2, caspase 3, and Apaf proteins in inter-nucleosomal DNA, in mitochondrial membrane potentials and in the release of cytochrome-c were analyzed by utilizing different experimental protocols. Results revealed that administration of LC-5C and LC-15C had little or no cytotoxic effect in normal peripheral blood mononuclear cells, but caused considerable cell death through apoptosis in cancer (HeLa) cells, which was evident from the induction of DNA fragmentation, the increases in the expressions of protein and mRNA of caspase 3 and Bax, and the decreases in the expressions of Bcl2 and Apaf and in the release of cytochrome-c. Thus, the highly-diluted, dynamized homeopathic remedies LC-5C and LC-15C demonstrated their capabilities to induce apoptosis in cancer cells, signifying their possible use as supportive medicines in cancer therapy.

This study tested chemotherapeutic potential of Hydrastis canadensis (HC) extract in HeLa cells in vitro, with emphasis on its drug–DNA interaction and apoptosis induction ability. Nuclear uptake of HC by DAPI, Ao/Eb staining and internucleosomal DNA damage by comet assay was studied through fluorescence microscopy. Possible changes in MMP and apoptotic signalling events were critically analyzed. Cell cycle progression studied through FACS and fragmented DNA through “TUNEL” assay were critically analyzed. RT-PCR studies were conducted for analyzing Cyt-C and Bax translocation in mitochondrial and cytosolic extracts, and Caspase 3 in whole cell lysate. Role of p53-mediated regulation of NF-κβ and TNF-α was elucidated by Western blot analysis. Data of CD and Tm profile of CT-DNA were analyzed. Overall results indicated anti-cancer potential of HC through its ability to induce apoptosis, and interaction with CT-DNA that changed structural conformation of DNA, proving HC to be a promising candidate for chemoprevention.

Objective. Ethanolic extract of G ymnema sylvestre (GS) leaves is used as a potent antidiabetic drug in various systems of alternative medicine, including homeopathy. The present study was aimed at examining if GS also had anticancer potentials, and if it had, to elucidate its possible mechanism of action. Methods. We initially tested possible anticancer potential of GS on A375 cells (human skin melanoma) through MTT assay and determined cytotoxicity levels in A375 and normal liver cells; we then thoroughly studied its apoptotic effects on A375 cells through protocols such as Hoechst 33258, H2DCFDA, and rhodamine 123 staining and conducted ELISA for cytochrome c, caspase 3, and PARP activity levels; we determined the mRNA level expression of cytochrome c, caspase 3, Bcl2, Bax, PARP, ICAD, and EGFR signaling genes through semiquantitative reverse transcriptase polymerase chain reaction and conducted Western blot analysis of caspase 3 and PARP. We also analyzed cell cycle events, determined reactive oxygen species accumulation, measured annexin V-FITC/PI and rhodamine 123 intensity by flow cytometry. Results. Compared with both normal liver cells and drug-untreated A375, the mortality of GS-treated A375 cells increased in a dose-dependent manner. Additionally, GS induced nuclear DNA fragmentation and showed an increased level of mRNA expression of apoptotic signal related genes cytochrome c, caspase 3, PARP, Bax, and reduced expression level of ICAD, EGFR, and the anti-apoptotic gene Bcl2. Conclusion. Overall results indicate GS to have significant anticancer effect on A375 cells apart from its reported antidiabetic effect, indicating possibility of its palliative use in patients with symptoms of both the diseases.

The capability of crude ethanolic extracts of certain medicinal plants like Phytolacca decandra, Gelsemium sempervirens, Hydrastis canadensis and Thuja occidentalis used as homeopathic mother tinctures in precipitating silver nanoparticles from aqueous solution of silver nitrate has been explored. Nanoparticles thus precipitated were characterized by spectroscopic, dynamic light scattering, X-ray diffraction, atomic force and transmission electron microscopic analyses. The drug-DNA interactions of silver nanoparticles were analyzed from data of circular dichroism spectroscopy and melting temperature profiles using calf thymus DNA (CT-DNA) as target. Biological activities of silver nanoparticles of different origin were then tested to evaluate their effective anti-proliferative and anti-bacterial properties, if any, by exposing them to A375 skin melanoma cells and to Escherichia coli C, respectively. Silver nanoparticles showed differences in their level of anti-cancer and anti-bacterial potentials. The nanoparticles of different origin interacted differently with CT-DNA, showing differences in their binding capacities. Particle size differences of the nanoparticles could be attributed for causing differences in their cellular entry and biological action. The ethanolic extracts of these plants had not been tested earlier for their possible efficacies in synthesizing nanoparticles from silver nitrate solution that had beneficial biological action, opening up a possibility of having therapeutic values in the management of diseases including cancer.

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 (LD 50 ) and the other two below LD 50 , 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.

Removal of large sized ovarian cysts by homeopathic treatment is generally not considered as a dependable option vis-a-vis removal via surgery. In this communication, we present three authentic cases of successful removal of big sized ovarian cysts by administration of a single potentized homeopathic remedy, Apis mellifica, selected on the basis of totality of symptoms, and authenticated with the aid of ultrasonographic as well as hormonal studies.

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.

Objective To evaluate the ameliorative efficacy of dihydroxy-isosteviol methyl ester (DIME) of Pulsatilla nigricans extract in arsenic-induced DNA damage in testis cells of mice. Methods The mice were treated with sodium arsenite (SA) solution intragastrically at a dose of 20 mg/kg per day and examined at 30, 60, and 90 d after treatment. We divided SA-intoxicated mice into two sub-groups: one fed with DIME at a dose of 35 mg/kg and the other with 85% alcohol. We analyzed the expressions of apoptotic signal proteins like CYP1A1, p53 and caspase 3, ascertained the level of cellular and DNA damage and estimated the level of testicular-toxicity biomarkers. We studied the interaction of DIME with calf thymus DNA as target through circular dichroism spectra and melting temperature profiles. Results We observed an elevation in all apoptotic and toxicity biomarkers leading to cellular and DNA damage in the SA-intoxicated mice which showed significant inhibition or reversal on administration of DIME. Results also showed that DIME interacted with DNA, bringing in discernible changes in structure and conformation. Conclusion DIME has potentials for therapeutic use in amelioration of arsenic-induced reproductive toxicity.

In South East Asia, groundwater arsenic contamination has become a great menace. Chronic arsenic intoxication leads to a hyperglycemic condition in animals and man. Because of undesirable side-effects and affordability, orthodox medicine, like insulin, is not preferred by many who like natural products instead. Unfortunately, such natural products mostly lack scientific validation. Therefore, we became interested in assessing the efficacy of the ethanolic seed extract of Syzygium jambolanum (SJ), traditionally used against diabetic conditions. We also formulated poly (lactic-co-glycolic) acid (PLGA)-encapsulated nano-SJ (NSJ) and tested whether the ameliorative potentials of SJ could be enhanced by nano-encapsulation. In this study, we conducted both in vitro (in L6 cells) and in vivo (in mice) experiments to assess the relative efficacy of SJ and NSJ. We characterized the physico-chemical features of NSJ by atomic force microscopy and critically analyzed several bio-markers and signal proteins associated with arsenic-induced stress and hyperglycemia. We also determined the relative ameliorative potentials of SJ and NSJ by using standard protocols. NSJ could cross the blood brain barrier in mice. Overall results suggested that NSJ had a greater potential than that of SJ, indicating the possibility of using NSJ in the future drug design and management of arsenic-induced hyperglycemia and stress.

Arsenic contamination has become a menacing health concern, warranting search for new drugs capable of ameliorating its toxicity. Extract of Pulsatilla nigricans is occasionally used as traditional medicine including homeopathy to combat/alleviate toxicity-related symptoms of known or unknown cause. Mice were intoxicated with a sub-lethal dose of sodium arsenite (20mg/kg b.w./day, determined through a range-finding trial) and the effect on testicular toxicity after 30, 60, and 90 days was examined. We observed an increased level of reactive oxygen species, cellular damage in testes of SA-intoxicated mice and further analysed expressions of apoptotic signal proteins and mRNA like Bax, Bcl2 and caspase3. Treatment with EEPN showed significant inhibition/reversal of the arsenic-induced toxic effect in testis and reduced oxidative stress through modulating expressions of signal proteins, thereby inhibiting the progression of events of apoptosis in testis cells and sperm. Therefore, EEPN has potentials for therapeutic use in arsenic- induced reproductive toxicity.

[6]-Gingerol, a pharmacologically important bioactive component of ginger, has been reported to have anti-hyperglycemic, anti-cancer and anti-oxidative properties, but mechanisms through which these are achieved are largely unclear. The present study focuses on apoptosis and autophagy, two key events of anti-cancer activity, in HeLa cells treated with [6]-gingerol. The treated cells showed several morphological changes, including externalization of phosphatidyl serine, degradation of DNA and increase in TUNEL positivity. Furthermore, there was depolarization of mitochondrial membrane potential, providing evidence of mitochondria mediated apoptosis. The expression of caspase 3 and PARP was increased in cells exposed to [6]-gingerol. Circular dichroism study for testing drug-DNA interaction with both calf thymus and nuclear DNA as target revealed that the drug had potential to bind with the nuclear DNA and induce conformational changes of DNA. The over-expression of NFkβ, AKT and Bcl2 genes in cancer cells was down-regulated by [6]-gingerol treatment. On the other hand the expression levels of TNFα, Bax and cytochrome c were enhanced in [6]-gingerol treated cells. Thus, overall results suggest that [6]-gingerol has potential to bind with DNA and induce cell death by autophagy and caspase 3 mediated apoptosis.

Objective To evaluate the role of chelidonine isolated from ethanolic extract of Chelidonium majus in inducing apoptosis in HeLa cells and to assess the main signalling pathways involved. Methods Cells were initially treated with different concentrations of chelidonine for 48 h and the median lethal dose (LD50) value was selected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Morphological analysis of nuclear condensation and DNA damage and fragmentation were measured by 4',6-diamidino-2-phenylindole staining and comet assay. Further, reactive oxygen species (ROS) generation, cell cycle arrest and change in mitochondrial membrane potential were also examined and analyzed by flow cytometry. Evaluation of interaction of drug with CT DNA was investigated by circular dichroism (CD) spectral analysis to find any possible drug-CT DNA interaction. The mRNA and protein expressions of major signal proteins like p38, p53, protein kinase B (AKT), phosphatidylinositol 3-kinases (PI3K), Janus kinase 3 (JAK3), signal transducer and activator of transcription 3 (STAT3) and E6 and E7 oncoproteins as well as the pro-apoptotic genes and antiapoptotic genes were also estimated by reverse transcriptase-polymerase chain reaction and Western blotting. Results Based on LD(50) value (30 μg/mL) of chelidonine, three doses were selected, namely, 22.5 μg/mL (D1), 30.0 μg/mL (D2) and 37.5 μg/mL (D3). Results showed that chelidonine inhibited proliferation and induced apoptosis in HeLa cells through generation of ROS, cell cycle arrest at sub-G1 and G0/G1 stage, change in mitochondrial membrane potential and fragmentation of DNA. Results of CD spectra showed effective interaction between chelidonine and calf thymus DNA. Studies of signalling pathway revealed that chelidonine could efficiently induce apoptosis through up-regulation of expressions of p38, p53 and other pro-apoptotic genes and down-regulation of expressions of AKT, PI3K, JAK3, STAT3, E6, E7 and other antiapoptotic genes. Conclusion Chelidonine isolated from Chelidonium majus efficiently induced apoptosis in HeLa cells through possible alteration of p38-p53 and AKT/PI3 kinase signalling pathways.

We tested relative efficacy of the extract of Phytolacca decandra (PD) and its PLGA nano-encapsulated form (NPD) in mice intoxicated with benzo[a]pyrene (BaP) (25 mg/kg b.w.) plus sodium-arsenite (SA) (10 mg/kg b.w.) and on A549 lung cancer cells in vitro. We characterized nanoparticles by physico-chemical and morphological studies using dynamic light scattering, scanning electron and atomic force microscopies. We also conducted FTIR and (1)H NMR studies to determine if NPD had a co-polymeric nature and analyzed drug-DNA interaction through circular dichroism spectra (CD) and melting temperature profiles (T(m)) taking calf thymus DNA as target. An oral dose of 0.3mg/kg b.w. for NPD and 30 mg/kg b.w. for PD in mice showed chemopreventive effects in regard to DNA fragmentation, comet tail length and toxicity biomarkers like ROS generation, NFκβ, p53, PARP, CYP1A1 and caspase 3. NPD showed greater effects than that by PD. Results of in vivo studies showed similar effects on A549 in regard to cell viability, DAPI and PI staining, Comet tail length, DNA fragmentation. To further confirm the biological molecule present in PD we analyzed its chromatographic fraction through mass spectroscopy, NMR and FT-IR studies and characterized it to be a tri-terpenoid, a derivative of betulinic acid with a molecular formula C(30)H(46)O(2.) Thus, overall results suggest that nano-encapsulation of PD (NPD) increases drug bioavailability and thereby has a better chemo-preventive action against lung cancer in vivo and on A549 cells in vitro than that of PD.

The pyridinium-based fluororeceptor 1 and the sensor bead 2 recognize hydrogen pyrophosphate effectively through the 'Indicator Displacement Assay' (IDA) technique over a series of other anions in aq CH(3)CN (CH(3)CN/H(2)O = 4:1, v/v, pH = 6.5). The sensor bead 2 is also capable of sensing the same anion selectively in pure water.

The purpose of the study was to evaluate whether potentized cholesterinum (Chol) intermittently used with another homeopathic remedy, Natrum Sulphuricum (Nat Sulph) can provide additional benefits in combating hepatotoxicity generated by chronic feeding of carcinogens, p-dimethylaminoazobenzene (p-DAB), and phenobarbital (PB). Mice were categorized into subgroups: normal untreated (Gr-1); normal + alcohol "vehicle" (Alc) (Gr-2), 0.06% p-DAB +0.05% PB (Gr-3), p-DAB+PB+Alc (Gr-4), p-DAB+PB+Nat Sulph-30 (Gr-5), p-DAB+PB+Chol-200 (Gr-6), p-DAB+PB+Nat Sulph-30+Chol-200 (Gr-7), p-DAB+PB+Nat Sulph-200 (Gr-8), and DAB+PB+Nat Sulph-200+Chol-200 (Gr-9). Hepatotoxicity was assessed through biomarkers like aspartate and alanine aminotransferases (AST and ALT), acid and alkaline phosphatases (AcP and AlkP), reduced glutathione content (GSH), glucose 6-phosphate dehydrogenase (G6PD), gamma glutamyl transferase (GGT), lactate dehydrogenase (LDH), and analysis of lipid peroxidation (LPO) at 30, 60, 90, and 120 days and antioxidant biomarkers like superoxide dismutase (SOD), catalase (CAT), and glutathione reductase (GR) were assayed. Electron microscopic studies (scanning and transmission) and gelatin zymography for matrix metalloproteinases were conducted in liver. The feeding of the homeopathic drugs showed intervention in regard to the increased activities of AST, ALT, AcP, AlkP, GGT, LDH, and LPO and decreased activities of G6PD, SOD, CAT, GR, and GSH noted in the intoxicated mice, more appreciable in Groups 7 and 9. Thus, combined therapy provided additional antihepatotoxic and anticancer effects.

Dihydroxy-isosteviol methyl ester (DIME), the principal biological compound isolated from the medicinal plant Pulsatilla nigricans (Fam: Ranunculaceae) having the molecular formula of C21 H34 O3 (molecular weight 334.25), was administered to cervical cancer cells (HeLa) in vitro to evaluate its possible apoptotic (anti-cancer) potentials. We analyzed the expression of p53, Bax, Bcl2, Apaf and caspase 3 signal proteins and analyzed the early apoptotic events in HeLa cells induced by DIME using protocols like Annexin V-FITC and PI staining. DIME caused a significant decrease in cell viability, induced nuclear condensation and inter-nucleosomal DNA fragmentation. We further studied the interaction of DIME with calf thymus DNA as target through circular-dichroism spectra. Results showed that DIME interacted with DNA, bringing indiscernible changes in structure and conformation. Thus, DIME showed its capability to induce apoptosis in cancer cells, signifying its utility in drug design as a possible candidate for chemoprevention.

Arsenic toxicity induces type 2 diabetes via stress mediated pathway. In this study, we attempt to reveal how sodium arsenite (iAs) could induce stress mediated impaired insulin signaling in mice and if an isolated active fraction of ginger, [6]-gingerol could attenuate the iAs intoxicated hyperglycemic condition of mice and bring about improvement in their impaired insulin signaling. [6]-Gingerol treatment reduced elevated blood glucose level and oxidative stress by enhancing activity of super oxide dismutase (SOD), catalase, glutathione peroxidase (GPx) and GSH. [6]-Gingerol also helped in increasing plasma insulin level, brought down after iAs exposure. iAs treatment to primary cell culture of β-cells and hepatocytes in vitro produced cyto-degenerative effect and accumulated reactive oxygen species (ROS) in pancreatic β-cells and hepatocytes of mice. [6]-Gingerol appeared to inhibit/intervene iAs induced cyto-degeneration of pancreatic β-cells and hepatocytes, helped in scavenging the free radicals. The over-expression of TNFα and IL6 in iAs intoxicated mice was down-regulated by [6]-gingerol treatment. iAs intoxication reduced expression levels of GLUT4, IRS-1, IRS-2, PI3K, AKT, PPARγ signaling molecules; [6]-gingerol mediated its action through enhancing the expressions of these signaling molecules, both at protein and mRNA levels. Thus, our results suggest that [6]-gingerol possesses an anti-hyperglycemic property and can improve impaired insulin signaling in arsenic intoxicated mice.

Objective To examine to what degree an ultra-highly diluted homeopathic remedy, Arnica Montana 30C (AM-30C), used in the treatment of shock and injury, can modulate the expression of nucleotide excision repair genes in Escherichia coli exposed to ultraviolet (UV) irradiation. Methods E. coli were cultured to their log phase in a standard Luria-Bertani medium and then exposed to sublethal doses of UV irradiation at 25 and 50 J/m(2) for 22.5 and 45 s, respectively. The UV-exposed bacteria were then supplemented with either AM-30C (drug) or placebo (P-30C). The drug-treated and placebo-treated bacteria were subjected to assay for DNA damage and oxidative stress 90 min after UV exposure. Several protocols like comet assay, gel electrophoresis for DNA ladder and intracellular reactive oxygen species (ROS) generation, and biomarker measurement like superoxide dismutase (SOD), catalase (CAT) and reduced glutathione (GSH) were conducted. The mRNA expressions of the excision repair genes like ultraviolet repair uvrA, B and C genes (or also known as excision repair genes) were estimated by reverse transcription-polymerase chain reaction method. Results The UV-exposed bacteria showed DNA damage and oxidative stress, as revealed by an increase in ROS generation, and a decrease in SOD, CAT and GSH activities. As compared to placebo, the AM-30C-treated bacteria showed less DNA damage and oxidative stress as manifested by a decrease in ROS generation, and an increase in SOD, CAT and GSH activities. AM-30C also up-regulated the expression of repair genes as compared to the control. Conclusion AM-30C helped repair the DNA damage through up-regulation of repair genes and also ameliorated the oxidative stress through the reduction of ROS generation and suitable modulation of anti-oxidative stress enzymes.

Objective To examine if potentiated homeopathic drug Arsenicum Album 30C (Ars Alb 30C) can reduce sodium arsenite-induced toxicity in Escherichia coli. Methods E. coli were exposed to low arsenite insult after they grew up to log phase in standard Luria-Bertani medium. E. coli were treated with 1 or 2 mmol/L sodium arsenite alone (control), or Ars Alb 30C was added to the medium of a subset of sodium arsenite-treated bacteria (drug-treated), or homeopathically agitated alcohol was added to the medium containing a subset of sodium arsenite-treated bacteria (placebo-treated). A sub-set of untreated E. coli served as the negative control. Glucose uptake, specific activities of hexokinase, lipid peroxidase (LPO), superoxide dismutase (SOD) and catalase, intra- and extra-cellular sodium arsenite content, cell growth, cell membrane potential, DNA damage, intracellular reactive oxygen species (ROS), adenosine triphosphate (ATP) and free glutathione content and expressions of arsB and ptsG gene in normal control, sodium arsenite-treated, drug-treated and placebo-treated E. coli were analyzed. Treatments were blinded and randomized. Results In sodium arsenite-treated E. coli, glucose uptake, intracellular ROS, LPO and DNA damage increased along with decrease in the specific activities of hexokinase, SOD and catalase, intracellular ATP and free glutathione contents and cell membrane potential and growth, and there were increases in expression levels of arsB gene and ptsG gene. Ars Alb 30C administration reduced arsenic toxicity in E. coli by inhibiting generation of ROS and increasing tolerance to arsenite toxicity and cell growth. Conclusion Ars Alb 30C ameliorated arsenic toxicity and DNA damage, validating efficacy of ultra-highly diluted remedies used in homeopathy.

Helonias dioica (HD) is a threatened species of herb growing in North America. It is used as a traditional medicine for treating various ailments particularly related to reproductive issues. The root is reported to contain approximately 10% of a saponin (chamaelirin; C36H62O18) apart from certain other fatty acids. As saponins are known to have hypoglycemic effects, we suspected its possible antihyperglycemic potentials. We injected intraperitoneally alloxan (ALX) at the dose of 200 mg/kg body weight (bw) to induce hyperglycemia in mice and tested possible hypoglycemic effects of HD in vivo by deploying two doses (100 and 200 mg/kg bw, respectively). We also tested its effects on the isolated pancreatic islets cells in vitro. We used various standard protocols like reactive oxygen species (ROS) generation and DNA damage, activities of biomarkers like catalase (CAT), superoxide dismutase (SOD), lipid peroxidase (LPO), reduced glutathione (GSH) of the pancreas tissue and glucokinase and glycogen content of the liver of hyperglycemic mice. With a mechanistic approach, we also tracked down the possible signaling pathway involved. We found an elevated level of ROS generation, LPO and overexpression of inducible nitric oxide synthase (iNOS), tumor necrosis factor α (TNF- α), p38 Map kinase (p38 MAPK), nuclear factor (NF)- κβ, interferon gamma (IFN- γ), cytochrome c, caspase 3, poly [ADP ribose] polymerase (PARP) and cyclo oxygenase 2 (COX2) in ALX-induced diabetic mouse. Treatment of hyperglycemic mice with both the doses of HD showed a significant decrease with respect to all these parameters of study. Thus, our results suggest that HD prevents ALX-induced islet cell damage and possesses antihyperglycemic and antioxidative potentials.

A simple salicylaldimine-based receptor equipped with rhodamine moiety (L3) has been designed and synthesized for detection of Al3+ ions, which exhibits high sensitivity over other metal ions in aqueous buffer-methanol solution at physiological pH. It displays quick responses through visible colorimetric as well as fluorogenic changes on 1:1 binding to Al3+, as delineated by absorption and fluorescence titrations and also by Job’s method and ESI-MS+ studies. However, it exhibits calorimetric but not fluorometric responses towards Cu2+ and hence limits its colorimetric application towards the detection of Al3+ in the presence of Cu2+ ions. The properties of the probe like (i) solubility in aqueous buffer-methanol, (ii) cell-permeability and (iii) non-toxic nature towards cell may provide an opportunity for the in vitro/ in vivo bio study. The detection limit of Al3+ calculated by the 3σ method was found to be 24.8 pM. It is also found to be useful in the construction of logic gates; namely ‘OR’ gate (from absorption) and molecular ‘keypad lock’ (from emission) by the two chemical inputs (Al3+ and Cu2+) in the proper sequence of addition. The keypad lock operation is particularly important, as the output of the system depends not only on the proper combination but also on the order of input signals, creating the correct password that can be used to ‘‘open’’ this molecular keypad lock through strong fluorescence emission at 552 nm. Besides, “OFF–ON–OFF” fluorescence behavior observed in the presence of Al3+ and EDTA strengthens the potential applications of the L3–Al3+ system as a device with ‘INHIBIT’ logic gate functions. As a whole, its various logic gate properties may improve its impact for the development of new-generation ‘intelligence’ digital devices. To the best of our knowledge, this is the first report on the fluorescence emission of a rhodamine derivative induced by Al3+ binding with such multiple logic gate operations accompanied by its application to cell-biology.

Crude ethanolic extract ofThuja occidentalis(Fam: Cupressaceae) is used as homeopathic mother tincture (TOΦ) to treat various ailments, particularly moles and tumors, and also used in various other systems of traditional medicine. Anti-proliferative and apoptosis-inducing properties of TOΦ and the thujone-rich fraction (TRF) separated from it have been evaluated for their possible anti-cancer potentials in the malignant melanoma cell line A375. On initial trial byS-diphenyltetrazolium bromide assay, both TOΦ and TRF showed maximum cytotoxic effect on A375 cell line while the other three principal fractions separated by chromatography had negligible or no such effect, because of which only TRF was further characterized and subjected to certain other assays for determining its precise anti-proliferative and apoptotic potentials. TRF was reported to have a molecular formula of C10H16O with a molecular weight of 152. Exposure of TRF ofThuja occidentalisto A375 cellsin vitroshowed more cytotoxic, anti-proliferative and apoptotic effects as compared with TOΦ, but had minimal growth inhibitory responses when exposed to normal cells (peripheral blood mononuclear cell). Furthermore, both TOΦ and TRF also caused a significant decrease in cell viability, induced inter-nucleosomal DNA fragmentation, mitochondrial transmembrane potential collapse, increase in ROS generation, and release of cytochromecand caspase-3 activation, all of which are closely related to the induction of apoptosis in A375 cells. Thus, TRF showed and matched all the anti-cancer responses of TOΦ and could be the main bio-active fraction. The use of TOΦ in traditional medicines against tumors has, therefore, a scientific basis.

Objective We formulated nano-encapsulation of a naturally occurring coumarin-scopoletin (7-hydroxy-6-methoxy coumarin, HMC, C(10)H(8)O(4)), isolated from plant Gelsemium sempervirens having anticancer potentials, with a bio-adhesive agent -polylactic-co-glycolic acid (PLGA) and tested if its cellular uptake, bioavailability and apoptotic (anticancer) potentials could thus be increased vis-a-vis unencapsulated HMC. Methods A375 melanoma cancer cells were used for testing cellular entry and anticancer potentials of HMC and nano-7-hydroxy-6-methoxy coumarin (NHMC) through several standard protocols. Characterization of NHMC was done by dynamic light scattering for determination of particle size, polydispersity index (PDI), and zeta potential. Surface morphology of nanoparticles was determined by scanning electron microscopy and atomic force microscopy. Results HMC was encapsulated with more than 85% entrapment efficiency, the average particle size of NHMC being less than 110 nm and a PDI 0.237, which resulted in enhanced cellular entry and greater bioavailability. NHMC showed a faster cellular uptake (15 min) than its unencapsulated counterpart (30 min). Study of signal molecules through mRNA expressions revealed that NHMC caused down-regulation of cyclin-D1, proliferating cell nuclear antigen (PCNA), survivin and Stat-3, and up-regulation of p53 and caspase-3, that in turn induced a greater number of apoptosis vis-a-vis unencapsulated HMC. Conclusion The formulation yielded small-sized NHMC by biodegradable PLGA that took less time for cellular entry, and caused more apoptosis to cancer cells, but apparently had negligible cytotoxicity against normal skin cells. Nano-encapsulation of bioactive plant ingredients can be a strategy worth trying for designing effective chemopreventive drug products.

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 β-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.

The arsenate tolerance limit in wild-type Aspergillus niger was determined. Because of its high tolerance, toxic effects of arsenate concentrations ranging from 25 to 100 mg/L were investigated in regard to growth, intracellular thiols, proline and malondialdehyde (MDA) contents of wild-type A. niger. Cellular arsenate uptake was analyzed. Activities of catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR) and succinate dehydrogenase (SDH) were assayed. Growth of A. niger increased at 25 mg/L arsenate, and it survived up to 100 mg/L. MDA, intracellular thiol and proline contents increased up to a certain level. Activities of GR, SOD and CAT declined following a rise at low concentration(s); SDH activity decreased gradually with increased arsenate stress. Results indicated that A. niger had high arsenate uptake potential and could tolerate oxidative stress by manipulating its anti-oxidative defense mechanism, a property that may be exploited for removal of arsenate from contaminated aqua-environment.

Several homeopathic remedies, namely, Pulsatilla Nigricans (30th potency), Ceanothus Americanus (both mother tincture and 6th potency) and Ferrum Metallicum (30th potency)selected as per similia principles were administered to 38 thalassemic patients receiving Hydroxyurea (HU) therapy for a varying period of time. Levels of serum ferritin (SF), fetal hemoglobin (HbF), hemoglobin (Hb), platelet count (PC), mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), mean corpuscular hemoglobin (MCH), white blood cell (WBC) count, bilirubin content, alanine amino transferase (ALT), aspartate amino transferase (AST) and serum total protein content of patients were determined before and 3 months after administration of the homeopathic remedies in combination with HU to evaluate additional benefits, if any, derived by the homeopathic remedies, by comparing the data with those of 38 subjects receiving only HU therapy. Preliminary results indicated that there was a significant decrease in the SF and increase in HbF levels in the combined, treated subjects. Although the changes in other parameters were not so significant, there was a significant decrease in size of spleen in most patients with spleenomegaly and improvement in general health conditions along with an increased gap between transfusions in most patients receiving the combined homeopathic treatment. The homeopathic remedies being inexpensive and without any known side-effects seem to have great potentials in bringing additional benefits to thalassemic patients; particularly in the developing world where blood transfusions suffer from inadequate screening and fall short of the stringent safety standards followed in the developed countries. Further independent studies are encouraged.

Mice (Mus musculus) have been used as a model for homeopathy research in relation to cytotoxicity, genotoxicity and carcinogenesis in our laboratory for the last three decades. Initially, anti-radiation activities of several potentized homeopathic drugs were tested against suitable controls by taking into consideration several cytogenetic endpoints. Subsequently, anti-cytotoxic, anti-genotoxic and anti-oxidative stress effects of some homeopathic drugs were tested against several chemical toxic metalloids and metal compounds. Modern techniques including Western blot, immunofluorescence, electron microscopy, UV-spectroscopy, HPLC, FTIR, NMR, RT-PCR etc were deployed to understand the possible mechanisms and pathways of action of potentized homeopathic drugs. We hypothesise that one way by which potentized homeopathic drugs act is through regulatory action on gene expression.

Scopoletin, an alkaloid separated from ethanolic extract of the medicinal plant, Gelsemium sempervirens (Fam: Loganiaceae) has been reported to have anti-cancer potentials. The synthetic coumarin (4-Methyl-7 hydroxy coumarin) derived from resorcinol and ethyl aceto-acetate in presence of concentrated sulphuric acid is structurally close to scopoletin, being a coumarin derivative. Whether this synthetic compound also has anti-cancer potentials has been evaluated in vivo on DMBA (7,12-Dimethylbenz[a]anthracene) induced skin cancer in mice by analyzing results of several cytogenetic endpoints, Comet assay, and fluorescence activated cell sorting (FACS). Further, expressions of signal proteins like Aryl hydrocarbon receptor , p53, PCNA, Akt, Bcl-2, Bcl-xL, Bad, Bax, NF-kappaB Apaf, IL-6, Cytochrome-c, Caspase-3 and Caspase-9 were studied by immunoblot analysis along with histology of skin and immuno-histochemical localization of Aryl hydrocarbon receptor and PCNA in DMBA treated mice vis-a-vis carcinogen treated synthetic coumarin fed mice. Feeding of this synthetic coumarin induced positive modulations in expression of all biomarkers in DMBA administered mice, giving clues on its possible signaling pathway(s) - primarily through down-regulation of Aryl hydrocarbon receptor and PCNA and up-regulation of apoptotic proteins like Bax, Bad, Cytochrome c, Apaf, Caspase-3 and Caspase-9, resulting in an appreciable reduction in growth of papilloma in mice. Therefore, this synthetic coumarin shows promise for use in cancer therapy, particularly in skin cancer.

To examine if a potentized homeopathic drug, Natrum Sulphuricum 200 (Nat Sulph-200) has protective potentials against hepatocarcinogenesis, liver tumors were induced in mice through chronic feeding ofP-dimethylaminoazobenzene (p-DAB; initiator of hepatocarcinogenesis) and phenobarbital (PB; promoter). Mice were divided into five sub-groups: fed normal low protein diet (Gr. I, normal control); fed normal low protein plus alcohol-200 (vehicle of the homeopathic remedy) (Gr. II); fed diet mixed with 0.06% p-DAB plus 0.05% PB (Gr. III); fed diet and carcinogens like Gr.III, plus alcohol 200 (positive control for drug fed mice) (Gr. IV) and fed diet and carcinogens like Gr. III, plus Natrum Sulphuiricum-200 (Gr. V; drug fed). Mice were sacrificed at day 7, 15, 30, 60, 90 and day 120 for study of cytogenetical endpoints like chromosome aberrations (CA), micronuclei (MN), mitotic index (MI) and sperm head anomaly (SHA) and biochemical toxicity parameters like aspartate amino transferase (AST), alanine amino transferase (ALT), acid (AcP) and alkaline (AlkP) phosphatases, lipid peroxidation (LPO) and reduced glutathione (GSH) content. Less number of liver tumors were observed in Gr. V (drug fed) mice. Administration of Nat Sulph 200 reduced genomic damage, activities of AcP, AlkP, AST, ALT, LPO and increased GSH content. Therefore, independent replication of the study by others is encouraged.

The chemical structure of the main fluorescenting compound in the ethanolic extract (mother tincture) of the American yellow jasmine, Gelsemium sempervirens, was determined by employing 1H nuclear magnetic resonance (NMR), 13C NMR, mass spectroscopy, high-performance liquid chromatography (HPLC), correlation spectroscopy (COSY), and Fourier transform infrared (FTIR) spectroscopy analyses. Spectrofluorometric analysis has been made of the mother tincture and its agitated serial dilutions (up to 12th potency) prepared according to a homeopathic procedure in which serial, agitated dilutions were made separately in glass and polypropylene containers. The succussions were made by employing three different modes: hand jerk, sonication, and vortexing. The chemical formula of scopoletin, the main fluorescent compound, was determined to be C10H8O4 having a molecular weight of 192.17. Significant differences were noted between the remedies prepared in the two types of containers. Further, a comparison between any two methods of agitation revealed significant differences in fluorometric data of remedies at certain potency levels. The biological (anticancer) action of the crude extract, the alkaloid scopoletin, and 2C potency of Gelsemium sp were tested in vitro on the HeLa cell line through fluorescence microscopy, the 3(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, and fluorescent activated cell sorting (FACS). The role of nanoparticles presumably derived from the containers, their orientation, and their interaction with the starting substance during the dynamization process initiated by different modes of agitation could possibly be attributed to the differences noted in the fluorometric data of potencies prepared in the two types of containers and among the three different means of succussion tested.

Relative anticancer potentials of six flavonol fractions (F1-F6) isolated from Thuja occidentalis extract were first evaluated against lung cancer cells A549 in vitro, and fraction- F4 showing the maximum protective ability, was then tested in mice intoxicated with Benzo(a)pyrene (BaP), a known carcinogen with major effect on lung, to learn if this could also have ameliorative action against lung toxicity and tissue damage in mice in vivo. Chemical nature of F1-F6 fractions was confirmed with specific flavonol confirmatory test and mass spectral analysis; all fractions were tested for their possible anticancer effects against A549 cells. Results suggested that fraction 4 (F4) had the strongest anticancer effect. When treated to BaP intoxicated mice, F4 induced recovery of damaged lung tissue, presumably through inhibition of ROS generation, and enhanced production of major antioxidant molecules, that in turn blocked PI3K-activated expressions of Akt. Decrease in Bcl2/Bax ratio, over-expression of p53 gene and activation of caspase 3 were observed in tissues of F4 treated mice, further confirming apoptotic cell death as its major target. The F4 fraction of Thuja occidentalis extract showed remarkable apoptotic potential against lung cancer cells and ameliorative ability against BaP induced lung toxicity.