Trypan Blue Exclusion Assay Research Articles

The effect of natural polyphenols Resveratrol, Gallic acid, and Kuromanin chloride on human telomerase reverse transcriptase (hTERT) expression in HepG2 hepatocellular carcinoma: role of SIRT1/Nrf2 signaling pathway and oxidative stress.

There is evidence that low doses or physiological concentrations of certain natural polyphenols enhance the activity of telomerase. However, the precise mechanism by which natural polyphenols regulate telomerase activity remains unclear. Recent research indicates that NF-E2 related factor 2 (Nrf2) and silent information regulator 1 (SIRT1) are involved in human telomerase reverse transcriptase (hTERT) regulation. Thus, in order to better comprehend the mechanism by which polyphenols regulate hTERT, the present study investigated the effects of the natural polyphenols Resveratrol, Gallic acid, and Kuromanin chloride on hTERT, Nrf2, and SIRT1 expression as well as oxidative stress in HepG2 hepatocellular carcinoma. The trypan blue dye exclusion assay was used to assess cell viability. The level of mRNA for hTERT, Nrf2, and SIRT1 was then determined using real-time PCR. A spectrophotometric analysis was conducted to quantify oxidative stress markers. The results demonstrated that Resveratrol induces the expression of hTERT and the SIRT1/Nrf2 pathway in a dose-dependent manner. Gallic acid at concentrations of 10 and 20μM also increased the expression of the hTERT and SIRT1/Nrf2 pathway. Furthermore, dose-dependent overexpression of hTERT and Nrf2 was induced by Kuromanin chloride at 10 and 20µM. Moreover, we found that Resveratrol and Kuromanin chloride ameliorated oxidative stress, whereas Gallic acid exacerbated it. This study demonstrates that low doses of polyphenols (Resveratrol, Gallic acid, and Kuromanin chloride) upregulate the expression of the hTERT gene in the HepG2 hepatocellular carcinoma cell line, possibly via induction of the SIRT1/Nrf2 signaling pathway. Therefore, by targeting this pathway or hTERT, the anti-cancer effect of polyphenols can be enhanced.

Meclofenamic acid inhibits the invasion and migration of LNCaP prostate carcinoma cells by regulating vimentin expression

Objectives: Despite various studies to improve the early detection and development of novel treatment agents, no significant progress has been made in the treatment of prostate cancer. Moreover, the metastasis of prostate carcinoma is a major challenge in treatment modalities. Therefore, in the present study, it was aimed to investigate the effect of meclofenamic acid (MA), a nonsteroidal anti-inflammatory drug, on the migration and invasion of LNCaP prostate carcinoma cells.
 Methods: Firstly, the non-toxic concentrations of MA on LNCaP cells were determined by trypan blue exclusion assay. After that, the effect of MA on migration and invasion was assessed by wound healing assay and matrigel invasion assay, respectively. Finally, the expression level of vimentin, which is a marker for epithelial-to-mesenchymal cell transition, was assessed by western blotting.
 Results: The results of trypan blue exclusion assay showed that 60 and 80 µM concentrations of MA were non-toxic to the cells. The migration rate of non-treated control cells was 44.4%, while it was significantly reduced to 12.9% and 2.9% in 60 and 80 µM MA-treated groups, respectively. When the cells were treated with 80 µM MA, the drug significantly reduced the invasion of LNCaP cells from 52% to 30.3%. Western blot results showed that the level of vimentin expression was significantly decreased in MA-treated cells.
 Conclusion: The study shows for the first time that MA inhibits the invasion and migration of prostate cancer LNCaP cells by decreasing the expression level of vimentin.

Use of gold nanoparticle-silibinin conjugates: A novel approach against lung cancer cells.

Lung cancer presents one of the most challenging carcinomas with meager 5-year survival rates (less than 20%), high metastasis and high recurrence due to chemo- and radio- resistance. An alternative or complementation to existing prognosis modalities is the use of phytochemicals such as silibinin, which targets essential cytokines, angiogenic factors and transcription factors for a profound anti-tumor effect. However, the problems of low solubility in an aqueous physiological environment, poor penetration, high metabolism and rapid systemic clearance limit the therapeutic use of silibinin. Conjugation of gold nanoparticles (GNPs) with silibinin may overcome the above challenges along with distinct advantages of biocompatibility, optical properties for monitoring and causation of cytotoxicity in cancer cells. The current study thus aims to develop silibinin conjugated gold nanoparticles (Sb-GNPs) with pH responsive release in the cancer microenvironment, optimizing several parameters for its higher activity and further evaluate the nanoplatform for their efficacy in inducing cell death in vitro against A549 lung cancer cells. GNPs was synthesized using trisodium citrate dihydrate as the reducing agent and further used for the conjugation of silibinin. The synthesized GNPs were found to be monodispersed and spherical in shape. The silibinin was successfully conjugated with gold nanoparticles and long-term stability of GNPs and Sb-GNPs nanoconjugates in suspension phase was confirmed by FTIR and DLS. Anticancer properties of Sb-GNPs were confirmed by different assay using MTT, Trypan blue dye exclusion assay and cell cycle analysis assay. After conjugation of silibinin with GNPs, the efficacy of silibinin increased 4–5 times in killing the cancer cells. This is the first report on using silibinin gold nanoconjugate system for lung cancer therapy with promising future applications.

Combinatorial Effect of Arsenic and Herbal Compounds in Telomerase-Mediated Apoptosis Induction in Liver Cancer.

Tumour illness and its resistance against existing anticancer therapies pose a serious health concern globally despite the progressive advancement of therapeutic options. The prevailing treatment of HCC using numerous antitumor agents has inflated long-lived complete remissions, but a percentage of individuals still die due to disease recurrence, indicating a need for further exploration of possible anti-tumour regimes. We aim to boost the effectiveness of the HCC treatment by conducting current investigations evaluating the effect of arsenic trioxide (ATO) with different herbal compounds like quercetin and aloe-emodin against liver tumour via inhibition of telomerase, a pro-cancer enzyme. The anticancer activity of ATO with herbal compounds was investigated in human control liver cell line (Wrl-68) and cancer liver cell line (HepG2) at different time intervals. Viability and cytotoxicity in response to combinatorial drugs were assessed in vitro by trypan blue dye exclusion assay and MTT and WST assay. Apoptosis was analysed by annexin V/PI assay, and the expression of telomerase and apoptosis-regulating proteins was evaluated by immunoblotting and qRT-PCR. Arsenic trioxide in combination with quercetin and aloe-emodin reduced cell viability in cancerous cells compared to normal cells by inducing apoptosis, downregulating telomerase and Bcl-2 (anti-apoptotic protein) and upregulating the expression of Bax (pro-apoptotic protein). ATO exhibited significant anticancer effects due to the synergistic effects of quercetin and aloe-emodin in liver tumour cells. The current study data collectively suggest that a successful inhibition of cancer growth by the combination of ATO and tested herbal medicines against liver tumour growth is via the inhibition of telomerase activity.

An in vitro assessment of the thermoreversible PLGA-PEG-PLGA copolymer: Implications for Descemet's membrane endothelial keratoplasty.

To explore the use of a thermoreversible copolymer gel coating to prevent donor tissue scrolling in Descemet's membrane endothelial keratoplasty (DMEK). PLGA-PEG-PLGA triblock copolymer was synthesised via ring opening polymerisation. Two formulations were fabricated and gelation properties characterised using rheological analyses. Endothelial cytotoxicity of the copolymer was assessed using a Trypan Blue exclusion assay. Thickness of the copolymer gel coating on the endothelial surface was analysed using anterior segment optical coherence tomography (OCT) (RTVue-100, Optovue Inc.). Gold nanoparticles were added to the copolymer to aid visualisation using OCT. Prevention of Descemet membrane donor scrolling was represented via a novel, in vitro, immersion of copolymer coated donor graft material. Two different formulations of PLGA-PEG-PLGA copolymer were successfully fabricated and the desired peak gelling temperature of 24°C was achieved by polymer blending. Application of 20%, 30% and 40% (wt/vol) polymer concentrations resulted in a statistically significant increase in polymer thickness on the endothelium (p < 0.001). There was no detectable endothelial cytotoxicity. The polymer was easy to apply to the endothelium and prevented scrolling of the DMEK graft. This PLGA-PEG-PLGA thermoreversible copolymer gel could be exploited as a therapeutic aid for preventing DMEK graft scrolling.

Simvastatin Coadministration Modulates the Electrostatically Driven Incorporation of Doxorubicin into Model Lipid and Cell Membranes.

Understanding the interactions between drugs and lipid membranes is a prerequisite for finding the optimal way to deliver drugs into cells. Coadministration of statins and anticancer agents has been reported to have a positive effect on anticancer therapy. In this study, we elucidate the mechanism by which simvastatin (SIM) improves the efficiency of biological membrane penetration by the chemotherapeutic agent doxorubicin (DOX) in neutral and slightly acidic solutions. The incorporation of DOX, SIM, or a combination of them (DOX:SIM) into selected single-component lipid membranes, zwitterionic unsaturated 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), neutral cholesterol, and negatively charged 1,2-dimyristoyl-sn-glycero-3-phospho-l-serine (DMPS) was assessed using the Langmuir method. The penetration of neutral lipid monolayers by the codelivery of SIM and DOX was clearly facilitated at pH 5.5, which resembles the pH conditions of the environment of cancer cells. This effect was ascribed to partial neutralization of the DOX positive charge as the result of intermolecular interactions between DOX and SIM. On the other hand, the penetration of the negatively charged DMPS monolayer was most efficient in the case of the positively charged DOX. The efficiency of the drug delivery to the cell membranes was evaluated under in vitro conditions using a panel of cancer-derived cell lines (A172, T98G, and HeLa). MTS and trypan blue exclusion assays were performed, followed by confocal microscopy and spheroid culture tests. Cells were exposed to either free drugs or drugs encapsulated in lipid carriers termed cubosomes. We demonstrated that the viability of cancer cells exposed to DOX was significantly impaired in the presence of SIM, and this phenomenon was greatly magnified when DOX and SIM were coencapsulated in cubosomes. Overall, our results confirmed the utility of the DOX:SIM combination delivery, which enhances the interactions between neutral components of cell membranes and positively charged chemotherapeutic agents.

التأثير التآزري المضاد للأورام للثيموكينون مع الريسفيراترول ضد خط خلايا سرطان الرئة البشرية

Introduction: Thymoquinone (TQ) and resveratrol are natural anticancer and chemo-preventive agents through mechanism(s) that are not fully understood. This study aimed to test the anticancer potential activity of the combination of TQ and Resveratrol, and their underlying mechanisms in the human lung cancer cell line, A549 cells. Materials and methods: The A549 cells were treated with the different concentrations (25, 50, 75, 100, or 200 μM) of TQ, resveratrol, and their combination for 24h. The antiproliferative activity against A549 cells was assessed using (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Trypan blue exclusion assay. Apoptosis markers were assessed by using a real-time reverse transcriptase PCR Assay (rRT-PCR). Results : We found that the proliferation of A549 lung cancer cells was inhibited by TQ, and resveratrol and their combination in a concentration-dependent manner, during a defined period. The combination therapy also could inhibit the growth of A549 cells at 25 μM with a 50% inhibitory concentration (IC50) value of 37 ± 3.18μM. These combinations were significantly (P &lt; 0.001) elevated caspase-3, caspase-8, caspase-9, p53, p21, Fas, FasL, and Bax mRNA levels in treating A549 cells compared to the untreated cells. The anticancer effect of this combination is mediated by the activation of the extrinsic and the intrinsic pathways. Conclusion: all these results give a new promising combination with the enhanced anticancer effect.

Atorvastatin attenuates NS1 (Non-structural protein-1) of dengue type-2 serotype-induced expressions of matrix metalloproteinases in HL-60 cells, differentiated to neutrophils: Implications for the immunopathogenesis of dengue viral disease

BackgroundThe dengue is a vector borne viral infection in humans. Bite of mosquito infected with a dengue virus transmits the disease. The neutrophils support more to the innate immune response by switching to infected tissues and triggering immunomodulatory mechanisms including the release of proteases and host defence peptides. MethodsCell viability by MTT and trypan blue dye exclusion assay, bright field microscopy for assessment of cell morphology, cytokines measurements by ELISA, estimation of protein by Bradford assay were done. Assessments of matrix metalloproteinase genes mRNA expressions were done using real-time PCR. ResultsIn the present study, we have for the first time unveiled that, NS1 antigen of dengue type-2 serotype, induce and stimulate the neutrophils cells to express high levels of matrix metalloproteases. NS1 exposure of HL-60 cells differentiated to neutrophils affected cell morphology and in 24 h of exposure. We have demonstrated that, the NS1 antigen has induced MMP-2, MMP-14 and MMP-9 expressions in neutrophils in a 24hrs exposure time. NS1 exposure has also further upregulated MMP-1, MMP-13, and MMP-8 expressions in neutrophils in a 24hrs exposure time. Notably, treatment with atorvastatin concentrations downregulated the expression profile of the all matrix metalloprotease significantly. Importantly, NS1 antigen has significantly increased the IL-6, IL-13 release by the HL,60 cells which was reversed by atorvastatin. On the other hand, NS1 exposure enhanced the mRNA expressions of VEGF-A and VEGF-D which was reversed by atorvastatin. However, we found that, NS1 exposure reduced the mRNA expressions profile of VEGF-C, which was reversed by atorvastatin. ConclusionIn conclusion, we report that, neutrophils associated matrix metalloprotease are involved in the pathogenesis of dengue viral disease. VEGF growth factors may also be released by the neutrophils which may subsequently participate in the endothelial dysfunctions leading to dengue shock syndrome.

Antiamoebic Activity of Imidazothiazole Derivatives against Opportunistic Pathogen Acanthamoeba castellanii.

We examined the antiamoebic effect of several imidazothiazole derivatives on Acanthamoeba castellanii of the T4 genotype. Trypan blue exclusion assays and haemocytometer counting were used to determine the reduction in A. castellanii trophozoite proliferation, in response to treatment with these compounds. To determine the effects of these compounds on host cells, lactate dehydrogenase assay was performed using HeLa cell lines. Amoebicidal assays revealed that the tested compounds at concentrations of 50 µM significantly inhibited amoebae trophozoites compared to controls. Compounds 1m and 1zb showed the highest amoebicidal effects eradicating 70% and 67% of A. castellanii, respectively. The compounds blocked both the encystation and excystation process in A. castellanii. Compounds 1m and 1zb blocked 61% and 55%, respectively, of amoeba binding to human cells. Moreover, the compounds showed minimal cytotoxic effects against host cells and considerably reduced amoeba-mediated host cell death. Overall, our study revealed that compounds 1m and 1zb have excellent antiamoebic potential, and should be considered in the development of curative antiamoebic medications in future studies. Further work is critical to determine the translational value of these findings.

Flower Extracts from Ornamental Plants as Sources of Sunscreen Ingredients: Determination by In Vitro Methods of Photoprotective Efficacy, Antigenotoxicity and Safety.

Plants are sources of sunscreen ingredients that prevent cellular mutations involved in skin cancer and aging. This study investigated the sunscreen properties of the extracts from some ornamental plants growing in Colombia. The UV filter capability of the flower extracts obtained from Rosa centifolia L., Posoqueria latifolia (Rudge) Schult, and Ipomoea horsfalliae Hook. was examined. Photoprotection efficacies were evaluated using in vitro indices such as sun protection factor and critical wavelength. UVB antigenotoxicity estimates measured with the SOS Chromotest were also obtained. Extract cytotoxicity and genotoxicity were studied in human fibroblasts using the trypan blue exclusion and Comet assays, respectively. Major compounds of the promising flower extracts were identified by UHPLC–ESI+–Orbitrap–MS. The studied extracts showed high photoprotection efficacy and antigenotoxicity against UVB radiation, but only the P. latifolia extract showed broad-spectrum photoprotection at non-cytotoxic concentrations. The P. latifolia extract appeared to be safer for human fibroblast cells and the R. centifolia extract was shown to be moderately cytotoxic and genotoxic at the highest assayed concentrations. The I. horsfalliae extract was unequivocally cytotoxic and genotoxic. The major constituents of the promising extracts were as follows: chlorogenic acid, ecdysterone 20E, rhamnetin-rutinoside, cis-resveratrol-diglucoside, trans-resveratrol-diglucoside in P. latifolia; quercetin, quercetin-glucoside, quercetin-3-rhamnoside, kaempferol, kaempferol-3-glucoside, and kaempferol-rhamnoside in R. centifolia. The potential of the ornamental plants as sources of sunscreen ingredients was discussed.

Antitumor immunity and therapeutic properties of marine seaweeds-derived extracts in the treatment of cancer

Marine seaweeds are important sources of drugs with several pharmacological characteristics. The present study aims to evaluate the antitumor and antitumor immunological potentials of the extracts from the brown alga Padinapavonica and the red alga Janiarubens, inhibiting the Egyptian marine coasts. Hep-G2 cell lines were used for assessment of the antitumor efficacy of Padinapavonica and Janiarubens extracts in vitro, while Ehrlich ascites carcinoma (EAC) cells were applied to gain more antitumor immunity and antitumor insights of P.pavonica and J.rubens extracts in vivo. In vitro antitumor potentials of P.pavonica and J.rubens extracts were analyzed against human liver cancer Hep-G2 cells by MTT and trypan blue exclusion assays. In vivo antitumor immunological potentials of P.pavonica and J.rubens extracts at low, high, and prophylactic doses were analyzed by blood counting and flow cytometry in mice challenged with Ehrlich ascites carcinoma (EAC) cells. In vitro results revealed that P.pavonica and J.rubens extracts caused significant decreases in the number and viability of Hep-G2 cells in a dose-dependent manner as compared to untreated Hep-G2 cells or Cisplatin®-treated Hep-G2 cells. In vivo findings showed that P.pavonica and J.rubens extracts at low, high, and prophylactic doses significantly reduced the number and viability of EAC tumor cells accompanied by increases in EAC apoptosis compared to naïve EAC mouse. Additionally, P.pavonica and J.rubens extracts at low and prophylactic doses remarkably increased both the total WBC count and the relative numbers of lymphocytes and decreased the relative numbers of neutrophils and monocytes. Flow cytometric analysis showed that P.pavonica and J.rubens extracts at the treatment and the prophylactic doses resulted in a significant increase in the phenotypic expressions of CD4+ T, CD8+ T, and CD335 cells compared to naïve EAC mouse. Overall, both extracts P.pavonica and J.rubens possess potential antitumor and antitumor immunological effects with less toxicity, opening new approaches for further studies of the chemical and biological mechanisms behind these effects.

The γ-Glutamylcyclotransferase Inhibitor Pro-GA Induces an Antiproliferative Effect Through the Generation of Mitochondrial Reactive Oxygen Species.

γ-Glutamylcyclotransferase (GGCT) is up-regulated in a broad range of cancers, including breast cancer, and GGCT inhibition has been shown to be a promising strategy for therapy. Herein, we evaluated the efficacy and mechanism of action of pro-GA, a GGCT enzymatic inhibitor, in MCF7 breast cancer cells. Proliferation was evaluated by WST-8 and trypan blue dye exclusion assays. Western blot analysis was conducted to examine the expression of cyclin-dependent kinase inhibitors (CDKI), including p21, p27, and p16. Induction of senescence was assessed by senescence-associated β-galactosidase staining. Generation of mitochondrial superoxide reactive oxygen species (ROS) was assessed using flow cytometry. The effect of N-acetylcysteine (NAC) on pro-GA dependent inhibition of proliferation, ROS generation, and senescence was also studied. The efficacy of systemic administration of pro-GA was evaluated in a MCF7 xenograft mouse model. Treatment with pro-GA inhibited proliferation of MCF7 cells, increased CDKI expression and mitochondrial ROS, and induced cellular senescence. We found that cotreatment with NAC restored proliferation in pro-GA treated cells. NAC similarly suppressed CDKI expression, mitochondrial ROS generation, and senescence induced by pro-GA. Furthermore, the systemic administration of pro-GA in an MCF7 xenograft model had significant antitumor effects without toxicity. Pro-GA may be a promising therapeutic agent for the treatment of breast cancer.

Blastocystis hominis undergoing programmed cell death via cytotoxic gamma irradiation

Blastocystis hominis is a worldwide common enteric parasite. Its pathogenic potential has not yet been established, although many reports suggest that it may cause the development of various gastrointestinal symptoms. The aim of the present study is to evaluate the destructive effect of different doses of cytotoxic gamma (γ) irradiation combined with and compared to metronidazole (MTZ) on Blastocystis spp. in vitro. The detection of the parasite in the stool specimen was conventionally done by light microscopic examination of direct smears, cultivation, followed by PCR-sequencing. Evaluation of γ-irradiation and MTZ effects on B. hominis was carried out by trypan blue exclusion assay, caspase activity detection, acridine orange staining, DNA fragmentation assay and transmission electron microscopic (TEM) examination. The current study demonstrated that exposure to γ-irradiation in a dose of 0.5 kGray (kGy) significantly (P < 0.05) reduced the viability of B. hominis subtype 2 by 95.2% compared to the untreated and MTZ-treated parasites (87.1%) after 48 h incubation. Combining the same dose of irradiation (0.5 kGy) with MTZ yielded a viability reduction of 94.2% and 94% after 24 and 48 h respectively, which were statistically significant (P < 0.05) compared to MTZ alone. Moreover, our results showed features of programmed cell death in the form of morphological, biochemical, and molecular changes (TEM abnormalities, caspase-like activity, and DNA fragmentation, respectively) with the high doses of γ-irradiation (0.3 and 0.5 kGy) either singly or combined with MTZ. In conclusion, cytotoxic γ-irradiation plays an important role in the inactivation of Blastocystis spp., so, it can be a promising prophylactic water-sterilizing tool against blastocystosis.

Identification and biotin receptor-mediated activity of a novel seleno-biotin compound that inhibits viability of and induces apoptosis in ovarian cancer cells

A series of seleno-biotin analogs were synthesized and their anticancer activity and mode of action were assessed using ovarian cancer cells. Compound 2, out of the other analogs, in direct comparison to biotin alone, more effectively reduced the cell viability and induced apoptosis in ovarian cancer cell lines in a dose dependent manner as demonstrated by the cell viability assay, trypan blue dye exclusion assay, Annexin V/7-AAD, and Caspase 3/7 apoptosis assays. Furthermore, compound 2 showed efficacy better than 5-fluorouracil (5-FU) and similar to cisplatin, in vitro; notably it was more cytotoxic to drug-resistant Hey A8 cells than cisplatin. The cytotoxicity of compound 2 was primarily mediated by reactive oxygen species (ROS) as demonstrated by DCFDA based ROS estimation. Biotin receptors (BR) saturation and the use of a BR negative cell line showed a significant decline in the cytotoxic ativity of the compound 2, confirming that its activity is BR-mediated. These experiments demonstrated that selenium modified biotin which contains an ester linked redox cycling selenocyanate group has the potential for human therapeutic applications against ovarian and other cancers over-expressing BR.

Anticancer effect of different rhizome extracts of the Lebanese Rheum ribes L. species on colorectal cancer cell lines

IntroductionColorectal cancer is the second leading cause of cancer-related mortality. Rheum ribes L. is one of the most important medicinal plants showing several biological activities. Few studies demonstrated anticancer potential of extracts of Turkish and Iranian Rheum ribes L. species rhizomes. We report for the first time the anticancer activity of different types of extracts of the rhizomes of the Lebanese Rheum ribes L. species against colorectal cancer cell lines. MethodsFive types of extracts were prepared using different solvents and cytotoxicity was assessed on colorectal cancer cell lines. Ethanolic and dichloromethane extracts were chosen for further studies and their effects on cell proliferation (MTT assay and trypan blue exclusion assay), colony forming ability (colony formation in soft agar), migration (scratch wound assay), and adhesion were assessed. ResultsOur results showed that ethanolic and dichloromethane extracts reduce HT-29 and HCT-116 cell viability in a dose and time dependent manner. They are also able to decrease colony-forming ability of these cells in soft agar medium. This anti-proliferative effect was accompanied by an increase in PARP cleavage suggesting a proapoptotic effect. Moreover, these extracts were also able to inhibit HT-29 and HCT-116 cell adhesion and migration suggesting their ability to reduce metastatic dissemination. ConclusionThese findings suggest a high therapeutic potential of the extracts of rhizomes of the Lebanese Rheum ribes L. species in the treatment of colorectal cancer.

Design, Cytotoxicity, and Tumor Targeted Drug Delivery of 5Fluorouracil Encapsulated in pH-Sensitive Co-polymers GG-g-P (HEMA) Conjugate Riboflavin Thin-Film

The study finds the design, cytotoxicity, and tumor-targeted drug delivery of 5Fluorouracil encapsulated in pH-sensitive guar gum grafted polymer. Cancer targeting 5FU loaded GG grafted p(HEMA) conjugated riboflavin thin-film (GG-g-P(HEMA)-RF) was successfully fabricated from guar gum, 2-hydroxy ethyl methacrylate, and riboflavin targeting agents using N, N-methylene bis acrylamide as a crosslinking agent in the presence of tetramethylethylenediamminne (TEMED) initiator Ammonium persulfate act as a catalyst. FT-IR and XRD spectroscopy techniques were used to analyze the structural features of the GG-g-P(HEMA)-RF. The results of SEM revealed that the carrier is of uniform rod shape, with a low porosity index and outstanding performance in phrases of 5FU encapsulation and extended-release ability. Targeted drug delivery strategies have received special attention from the scientific world due to advantages such as more effective therapy and reduction of side effects. Cytotoxicity effects of 5FU loaded GG-g-P(HEMA)-RF against Ehrlich Ascites Carcinoma (EAC) cells in vitro were investigated at different concentrations (0, 25, 50, 100, and 150 μg/mL) using trypan blue exclusion assay. The MTT cytotoxic assay investigated cell viability against EAC experimental models and implied the excellent biocompatibility of the carrier. The results revealed the anti-proliferative effect along with molecular signaling of apoptosis and generated reactive oxygen species (ROS) in Ehrlich Ascites Carcinoma cell lines. The morphological changes of apoptosis were evident in EAC cells and were observed using optical microscopy after staining. DPPH free radical scavenging assay was carried out to determine the radical scavenging activity of the 5FU loaded and unloaded GG-g-P(HEMA)-RF. The 5FU loaded GG-g-P(HEMA)-RF with DNA interaction was examined by electronic and spectroscopic fluorescence methods.

Cytotoxic Effects of the Schweinfurthin Analog 5′-Methylschweinfurthin G in Malignant Plasma Cells

Introduction: Multiple myeloma (MM) is a B plasma cell malignancy currently incurable, and novel therapeutics are needed. Evidences regarding the effect of natural compound schweinfurthins suggest that hematological cancers showed growth inhibitory effects to this family of compounds at single nanomolar concentrations. In this study, we evaluated the cytotoxicity of the schweinfurthin synthetic analog 5′-methylschweinfurthin G (MeSG) in MM cell lines, to better understand the validity of this compound as a therapeutic candidate for further studies in MM. Methods: MeSG toxicity against MM cell lines RPMI-8226, MM.1S, and H-929 was evaluated. Trypan blue exclusion and MTT assays measured cell viability and mitochondrial activity, respectively. Flow cytometry was performed to detect apoptotic mitochondria. Flow cytometry and Western blotting techniques were used to investigate apoptosis and to examine the cell cycle. Western blotting was used to determine AKT activation upon MeSG treatment. Results: We provide evidence that in all MM cells analyzed, MeSG exerts diverse cytotoxic effects. MeSG treatment of MM.1S and H-929, but not in RPMI-8226, causes a loss of mitochondria membrane potential. MeSG causes an arrest in G2/M, especially in RPMI-8226, supported by decreased levels of cyclin-B1 and early increased levels of p21. Finally, there is a diverse response to the MeSG treatment for AKT phosphorylation. MM.1S and H-929 showed a marked decrease in AKT phosphorylation at earlier time points compared to the RPMI-8226 line. Conclusions: MeSG cytotoxicity has been confirmed in all of 3 cell lines studied. Results suggest an early event of increased reactive oxygen species, and/or involvement of cholesterol homeostasis via decreased AKT activation, both of which are currently under investigation.

In vivo and in vitro Study to Evaluate the Anti-osteoporotic Activity of Punica granatum Seed, Bambusa arundinaceae Leaves and Trichosanthes diocio Fruit Ethanolic Extract

Background: In the present study, it was aimed at evaluating the efficacy of ethanolic extract of Punica granatum seed, Bambusa arundinaceae leaves and Trichosanthes diocio fruit on their anti-osteoporotic activity in ovariectomized (OVX) rats and mcf-7 cell line to identify their osteoprotective role. Methods: Thirty six female albino wistar OVX rats were treated with the ethanolic extract of Punica granatum seed, Bambusa arundinaceae leaves and Trichosanthes diocio fruit for 45 days. The ethanolic extracts were given in different concentrations for 24 h and a cytotoxicity study was carried out by trypan blue dye exclusion assay. Result: The ethanolic extract of Punica granatum seed, Bambusa arundinaceae leaves and Trichosanthes diocio fruit treated notably averted an OVX-induced increase in body weight. Moreover, exposure to ethanolic extract of Bambusa arundinacea significantly decreased lipid peroxidation and protein carbonyl formation. Treated mcf-7 cell line viability was also increased. All these finding suggest that the ethanolic extract of Punica granatum seed, Bambusa arundinaceae leaves and Trichosanthes diocio fruit have an important role in reducing osteoporosis.

NAMPT Inhibition Induces Neuroblastoma Cell Death and Blocks Tumor Growth.

High-risk neuroblastoma (NB) portends very poor prognoses in children. Targeting tumor metabolism has emerged as a novel therapeutic strategy. High levels of nicotinamide-adenine-dinucleotide (NAD+) are required for rapid cell proliferation. Nicotinamide phosphoribosyl transferase (NAMPT) is the rate-limiting enzyme for NAD+ salvage and is overexpressed in several cancers. Here, we determine the potential of NAMPT as a therapeutic target for NB treatment. NAMPT inhibition cytotoxicity was determined by trypan blue exclusion and LDH assays. Neuroblastoma stem cell self-renewal was evaluated by neurosphere assay. Protein expression was evaluated via Western blot. The effect of targeting NAMPT in vivo was determined using an NB1691-xenografted mouse model. Robust NAMPT expression was demonstrated in multiple N-MYC amplified, high-risk neuroblastoma cell lines. NAMPT inhibition with STF-118804 (STF) decreased ATP, induced apoptosis, and reduced NB stem cell neurosphere formation. STF treatment down-regulated N-MYC levels and abrogated AKT activation. AKT and glycolytic pathway inhibitors in combination with NAMPT inhibition induced robust, greater-than-additive neuroblastoma cell death. Lastly, STF treatment blocked neuroblastoma tumor growth in mouse xenograft models. NAMPT is a valid therapeutic target as inhibition promoted neuroblastoma cell death in vitro and prevented tumor growth in vivo. Further investigation is warranted to establish this therapy’s role as an adjunctive modality.

Terephthalato and succinato bridged Mn(II) and Zn(II) coordination polymers involving structure-guiding H-bonded tetrameric assemblies: Antiproliferative evaluation and theoretical studies

Two new coordination polymers of Mn(II) and Zn(II) viz. {[Mn(µ-tp)(3-Ampy)2(H2O)2]2(3-Ampy)·2(H2O)}n (1) and {[Zn(µ-succ)(3-Ampy)2]·2H2O}n (2) (tp = terephthalate, 3-Ampy = 3-aminopyridine, succ = succinate) have been synthesized and characterized using elemental and spectroscopic analyses, TGA and single crystal X-ray diffraction techniques. Uncoordinated 3-AMpy and water molecules of the polymeric co-crystal hydrate 1 are involved in the formation of cyclic tetrameric {(3-Ampy)2(H2O)2} assembly which provide rigidity to the structure. The presence of guest 3-Ampy and water molecules in the voids of supramolecular Mn(II) host tetramer in 1 provides additional reinforcement to the crystal structure. The lattice water molecules of 2 are aggregated via non-covalent interactions to form discrete cyclic {(H2O)4} tetrameric clusters in the crystal structure. Density functional theory (DFT), non-covalent index (NCI) plot and quantum theory of atoms in molecules (QTAIM) computational analyses reveal that the supramolecular H-bonded cyclic tetrameric structure-guiding synthon in 1 and the (H2O)4 cyclic water clusters in 2 have significant energetic contribution towards the stability of the polymers. In vitro anticancer activities of the compounds considering trypan blue and apoptosis assays revealed that the compounds induce concentration dependent cytotoxicity in Dalton’s lymphoma (DL) cancer cells with nominal effects in normal peripheral blood mononuclear cells (PBMC) cells. Comparative analysis reveals that compound 2 induced higher apoptosis in DL cells; which is well consistent with the trypan blue exclusion assay. Molecular docking and pharmacophore modelling studies of the compounds reveal that the compound 2 significantly interacts with the antiapoptotic proteins to establish structure activity relationship (SAR).