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Abstract In antimalarial research there are no standard procedures to determine the toxicity of a drug candidate. Among the alternatives available, in vitro cytotoxicity assays are the most widely used to predict toxic effects of future therapeutic products. They have the advantage over the in vivo assays, in that they offer the possibility to restrain the number of experimental variables. The objective of the present study was to compare in vitro cytotoxic methods by testing various compounds currently used to treat malaria against different cell lines. Neutral red (NR) uptake and methylthiazoletetrazolium (MTT) colorimetric in vitro assays were used to determine preliminary toxicity of commercially available antimalarial drugs against tumor and non-tumor cells lines. Toxicity through brine shrimp lethality bioassay and hemolytic activity were also evaluated. Significant differences were observed in the tests measured by NR uptake. The tumor cell lines TOV-21G and HepG2 and non-tumor WI-26VA4 cells showed relatively uniform toxicity results, with TOV-21G being the most sensitive cell tested, presenting the lowest concentration to cause death to 50% of viable cells (CC50) values. The results of this study support the use of TOV-21G, HepG2 and WI-26VA4 cells lines as the choice for cytotoxicity tests to evaluate potential bioactive compounds.
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Objective: To evaluate antioxidant, cytotoxic, and anti-venom capacity of crude bark extracts of Alstonia parvifolia Merr. Methods: Gas chromatography-mass spectrometry (GC-MS) and energy dispersive X-ray analyses were accomplished to characterize the chemical constituents of Alstonia parvifolia. Biochemical characterization was evaluated using an inhibitory phospholipase A 2 (PLA 2) assay, DPPH, and cytotoxicity assays. Using the constituents listed in the GC-MS analyses, molecular docking was conducted to inspect the binding energies between the chosen compounds and selected PLA 2 isoforms. Results: GC-MS analyses showed that the Alstonia parvifolia crude extract consisted predominantly of acetylmarinobufogenin (14.89%), γ-sitosterol (10.44%), 3-O-methyl-D-glucose (5.88%), 3,5-dimethoxy-4-hydroxyphenylacetic acid (5.30%), (2α,5α)-17-methoxyaspidofractinin-3-one (AFM) (4.08%), and 2,3,5,6,7,8,9-heptahydro-1-phenyl-5-(p-chlorophenylimino)-1H-benzo[e] [1],[4] thiazepine (HPT) (1.37%). The principal elemental components of Alstonia parvifolia were Ca (4.012%) and K (1.496%), as exhibited by energy dispersive X-ray examination. Alstonia parvifolia showed significant free radical scavenging ability (IC 50: 0.287 mg/mL) and was non-cytotoxic to normal HDFn cells (IC 50 >100 μg/mL). Moreover, Alstonia parvifolia was favorably cytotoxic to MCF-7 (IC 50: 4.42 μg/mL), followed by H69PR, HT-29, and THP-1, with IC 50 values of 4.94, 5.07, and 6.27 μg/mL, respectively. Alstonia parvifolia also displayed notable inhibition against PLA 2 activity of Naja philippinensis Taylor venom with IC 50 of (15.2 ± 1.8) μg/mL. Docking and cluster analyses projected negative binding energies from AFM (-6.36 to -9.68 kcal/mol), HPT (-7.38 to -9.77 kcal/ mol), and acetylmarinobufogenin (-7.22 to -9.59 kcal/mol). These calculations were for the particular interactions of Alstonia parvifolia constituents to PLA 2 homologues where the utmost affinity was detected in HPT owing to the dipole interactions with amino acid residues. Conclusions: The bark extract of Alstonia parvifolia shows great potential as an anti-venom agent due to its low cytotoxic profile, remarkable PLA 2 inhibition, and docking binding energies between its bioactive constituents and PLA 2 homologues.
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Objective: The evolution of antimicrobial resistance is a universal obstacle that necessities the innovation of more effective and safe antimicrobial alternatives with synergistic properties. The purpose of this study was to investigate the possible improvement of cephalexin antimicrobial treatments by loading into chitosan-based nanoparticles, then evaluate their antibacterial and antibiofilm activities as well as determination of its cytotoxicity. Methods: Chitosan nanoparticles (CSNPs) were prepared by ionic gelation method. Parameters were studied to optimize the particle size of CSNPs including pH, stirring rate, homogenization and ultra-sonication time. Size was measured by transmission electron microscope (TEM) and Zeta sizer, morphology seen by scanning electron microscope (SEM). Entrapment efficiency, drug loading and drug content were calculated. Stability of both plain and loaded chitosan Nano-carriers, Drug release and Kinetics also compatibilities were studied. Antimicrobial activity of CSNPs and cephalexin loaded CSNPs were evaluated against 4 Gram-positive and 4 Gram-negative standard and clinical isolates by microdilution method, also assessment of antibiofilm activity of both formulas was investigated against two biofilm producers clinical isolates by tube assay in addition to determination of their cytotoxicity by MTT(3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Results: Chitosan nanoparticles and its loaded antibiotics proved compatible combination with small Zeta size, suitable Zeta potential, maximum EE% and drug-loading capacity, sustained controlled release properties followed diffusion kinetic model and six month stability studies. Cephalexin loaded CSNPs showed better antimicrobial activity than plain CSNPs. Synergistic effects were found against S. aureus (ATCC 25923), B. subtilis (ATCC 9372), S. epidermidis, E. faecalis, P. aeruginosa (ATCC 29853) in addition to two carbapenem resistant isolates k. pneumoniae and E. coli. Also cephalexin loaded CSNPs exhibited antibiofilm activity against E. faecalis clinical isolate. Even though, cephalexin loaded CSNPs exhibited significant antibacterial activity, it showed less toxicity against mammalian cells, it had IC50 equal to 231.893 and did not exhibit any cytotoxicity against the WI-38 fibroblast cells at concentration 23.4 µg/ml. Conclusion: Cephalexin loaded CSNPs possessed good stability and sustained release effect in addition to its antimicrobial, antibiofilm activities and reduced cytotoxicity.
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Objective: To optimize the preparation method of paclitaxel-oleic acid (PTX-OA)/Brucea javanica oil (BJO) core- matched nanoemulsions (CMNEs). Methods: PTX-OA/BJO was synthesized by esterification of PTX and OA, and determined by HPLC. Ultrasionic emulsification was used to prepare the PTX-OA/BJO CMNEs. The concentration of oil phase (A), quality of polysorbate 80 (B), and ultrasonic power (C) were selected as the significant factors after single-factor experiments and applied in L16(43) orthogonal array design with the average particle size as criterion. In addition, the physicochemical properties and cytotoxicity of PTX-OA/BJO CMNEs were tested. Results: Linear range of PTX-OA was 5-25 μg/mL, Y = 12.709 X + 6.252 0, r = 0.999 5. The optimized conditions of PTX-OA/BJO CMNEs were as follows: The concentration of oil phase was 6.50 mg/mL, the mass ratio of polysorbate 80 to oil phase was 3.5: 6.5 and the ultrasonic power was 120 W. The CMNEs prepared by the optimal conditions showed an entrapment efficiency of (100.6 ± 1.9)% and the nanoscale particle size was (108.7 ± 2.3) nm, PDI was 0.232 ± 0.038. The morphology of CMNEs examined by TEM exhibited a uniform and spherical shape. In vitro drug release was up to 67% after 48 h. After PTX-OA/BJO CMNEs sealed in a bottle were stored at 4 ℃ for 60 h, the average particle size and entrapment efficiency had no significant change. The cytotoxicity in vitro showed that the combined PTX-OA/BJO CMNEs could obviously inhibit the proliferation of HepG-2 cells. Conclusion: The current study demonstrates the feasibility of incorporating PTX-OA and BJO into a single CMNEs for the synergism in cancer therapy. Furthermore, the preparation of PTX-OA/BJO CMNEs has the advantages of practicability and simple operation, as well lays the foundation for the further mechanism research of the combined paclitaxel and BJO in oncotherapy.
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Objective: To evaluate the cytotoxic activities of Premna odorata (P. odorata) leaves and bark, Artocarpus camansi (A. camansi) and Gliricidia sepium against selected human cancer cell lines by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Methods: The crude extracts of P. odorata, A. camansi and Gliricidia sepium were subjected to liquid-liquid partitioning by using hexane and ethyl acetate to separate compounds based on their polarity. The fractions were tested for their cytotoxic activity against human colon cancer cell line (HCT116), breast cancer cell line (MCF-7), lung adenocarcinoma cell line (A549) and Chinese hamster ovary cell line (AA8) by using MTT assay. Results: Based on the standard values of toxicity set by the study of Suffness and Pezzuto, P. odorata leaves and P. odorata bark hexane fractions and A. camansi leaves were all considered highly cytotoxic against the selected human cancer cell lines. P. odorata bark hexane extract exhibited the highest selectivity index for HCT116, MCF-7 and A549 cancer cell lines. Conclusions: The results obtained indicated that P. odorata leaves and bark and A. camansi leaves have excellent cytotoxic activity and warrant further studies to isolate novel compounds for chemotherapeutic use.
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Lung cancer causes 1.4 million deaths worldwide while non-small-cell lung cancer (NSCLC) represents 80-85% of the cases. Cisplatin is a standard chemotherapy against this type of cancer; however, tumor cell resistance to this drug limits its efficacy. Sea anemones produce compounds with pharmacological activities that may be useful for augmenting cisplatin efficacy. This study aimed to evaluate the pharmacological activities of crude venom (CV) from the sea anemone Bunodeopsis globulifera and four derived fractions (F1, F2, F3 and F4) to test their increase efficiency cisplatin cytotoxicity in human lung adenocarcinoma cells. Results Pre-exposure to CV, F1 and F2 fractions increases cisplatin cytotoxicity in human lung adenocarcinoma cells under specific conditions. Exposure to CV at 50 μgmL-1 induced a reduction of approximately 50% in cell viability, while a similar cytotoxic effect was observed when cell culture was exposed to F1 at 25 μgmL -1 or F2 at 50 μgmL-1. The cell culture exposure to F1 (10 μgmL-1) fraction combined with cisplatine (25 μM) provoked a decrease in MTT reduction until 65.57% while F2 (25 μgmL-1) fraction combined with cisplatin (10 μM) provoked a decrease in MTT reduction of 72.55%. Conclusions The F1 fraction had the greatest effect on the lung adenocarcinoma cell line compared with CV and F2. The combination of antineoplastic drugs and sea anemone toxins might allow a reduction of chemotherapeutic doses and thus mitigate side effects.
Subject(s)
Humans , Animals , Adenocarcinoma , Toxins, Biological/analysis , Pharmacology/instrumentation , Lung Neoplasms/pathologyABSTRACT
Background: Lung cancer causes 1.4 million deaths worldwide while non-small-cell lung cancer (NSCLC) represents 80-85% of the cases. Cisplatin is a standard chemotherapy against this type of cancer; however, tumor cell resistance to this drug limits its efficacy. Sea anemones produce compounds with pharmacological activities that may be useful for augmenting cisplatin efficacy. This study aimed to evaluate the pharmacological activities of crude venom (CV) from the sea anemone Bunodeopsis globulifera and four derived fractions (F1, F2, F3 and F4) to test their increase efficiency cisplatin cytotoxicity in human lung adenocarcinoma cells. Results: Pre-exposure to CV, F1 and F2 fractions increases cisplatin cytotoxicity in human lung adenocarcinoma cells under specific conditions. Exposure to CV at 50 μgmL-1 induced a reduction of approximately 50% in cell viability, while a similar cytotoxic effect was observed when cell culture was exposed to F1 at 25 μgmL -1 or F2 at 50 μgmL-1. The cell culture exposure to F1 (10 μgmL-1) fraction combined with cisplatine (25 μM) provoked a decrease in MTT reduction until 65.57% while F2 (25 μgmL-1) fraction combined with cisplatin (10 μM) provoked a decrease in MTT reduction of 72.55%. Conclusions: The F1 fraction had the greatest effect on the lung adenocarcinoma cell line compared with CV and F2. The combination of antineoplastic drugs and sea anemone toxins might allow a reduction of chemotherapeutic doses and thus mitigate side effects.
Subject(s)
Humans , Lung Neoplasms , Cnidarian Venoms/pharmacology , Cnidarian Venoms/therapeutic useABSTRACT
Our study focused on the antioxidant activities of Mosidae leaf ethanol extract (MLE) and included measurements of reducing power, total phenolic compounds, DPPH radical scavenging activity, and hydroxyl radical scavenging activity. In order to determine whether or not MLE evidences any chemopreventive activities, experimental lung metastasis was induced via the i.v. inoculation of colon26-M3.1 carcinoma cells into BALB/c mice. Additionally, we attempted to characterize any possible cytotoxic effects in murine normal splenocytes and tumor cells (B16-BL6 and colon26-M3.1). The total phenolic content and reducing capacity were measured at 39 mg/100 mL and 1.24, respectively, whereas the DPPH and hydroxyl radical scavenging activities of MLE were measured to be 88.89% and 22.10%, respectively. Prophylactic i.v. treatment with MLE resulted in a dose-dependent and significant inhibition of lung metastasis. Specifically, a MLE dose of 200 ug per mouse resulted in an 88.90% inhibition of lung metastasis. For the cytotoxicity assay, MLE doses up to 100 ug/mL were not shown to affect the growth of normal murine splenocytes. Additionally, the survival of normal cells was not affected at MLE doses below 500 ug/mL. However, MLE doses up to 500 ug/mL reduced the percentage of tumor cell growth for B16BL6 (67% alive) and colon26-M3.1 (62% alive) cells.