Isolation Of Cytotoxic Constituent From Bioactivity Guided Fraction Of Alysicarpus Monilifer L. (DC.)

Objective: Alysicarpus monilifer (Family Papilionaceae) has been used in the Indigenous system of medicine in tumor removal. The present study was designed to isolate and identify the constituent responsible for cytotoxic (anti-tumor) effects of the plant Alysicarpus monilifer. Methods: The plant was powdered and extracted to give a methanolic extract. Initially, Hexane, chloroform, ethyl acetate and methanolic fractions of the methanolic extract of the plant were subjected to cytotoxic screening using cell line based assay (MTT assay and NRU assay). The chloroform fraction showed significant cytotoxicity, so it was further subjected to column chromatography, to separate the cytotoxic phytoconstituent. The cell lines selected were breast cancer cells (MCF-7 and MDA-MB-468) and Liver cancer cells (HepG2 and HLE cell). Results were calculated as percentage growth inhibition with respect to untreated (control) cells versus treated cells. Result: A triterpene, Betulinic acid, was isolated from the aerial parts of Alysicarpus monilifer. The cytotoxic activity of the identified compound against MCF-7, MDA-MB-231, HLE and HepG2 cells was also found to be highly significant with 90% growth inhibition. Conclusion: The triterpene was identified to be betulinic acid, to which the cytotoxic activity can be attributed. It is a first report of isolation of betulinic acid from the Alysicarpus species.

Synthesis of Some Novel Tetra Aryl Imidazoles Compounds: Evaluation of In-Vitro Cytotoxicity and Anthelmintic Activity

The imidazole derivatives have potent therapeutic activity against cytotoxicity and parasites. The present study was planned to synthesize novel tetra aryl imidazoles compounds and evaluated for in vitro cytotoxicity and anthelmintic activity. Firstly, 2-amine-4-chloro pyridine was condensed with substituted benzaldehyde to give corresponding Schiff’s base. These Schiff’s bases further on treatment with ammonium acetic acid derivation and isatin yielded comparing novel tetra aryl imidazoles. The synthesized compounds were examined for in-vitro cytotoxicity and anthelmintic activity. The discoveries showed that all the synthesized novel substituted imidazoles have moderate to great anthelmintic action. They additionally had critical in-vitro cytotoxicity against HEp2 cell lines (Human larynx malignancy cell line) against standard utilizing 5-fluorouracil. The compounds 1b, 2b, 4b, 6b, and 8b had higher anthelmintic action contrasted with standard mebendazole. The synthesized compounds 1b, 2b and 8b had noteworthy in-vitro cytotoxicity against HEp2 cell lines.

In vitro biological activities of silver nanoparticles synthesized from Scedosporium sp. isolated from soil

One of the important fields in nanotechnology is the development of an environment friendly method for the synthesis of nanoparticles. Many approaches show that microorganisms are the most reliable tools for biosynthesis of nanoparticles compared to physical and chemical methods. In our study, fungi have been exploited for extracellular production of metal nanoparticles. It was observed that in Scedosporium, silver ions are reduced to silver nanoparticles, which was confirmed by UV-visible spectrophotometry and AFM. Optimization studies showed that as the concentration of AgNO3 used for synthesis increased, particles' size also increased. Size of the particles at different concentrations of AgNO3 was observed to be 79-107 nm with particles being ellipsoidal to spherical in shape. Silver nanoparticles synthesized from 2.0 mM silver nitrate, showed maximum antimicrobial activity compared to all antibiotics tested including synergistic effects. In vitro cytotoxicity of silver nanoparticles against MCF 7 and PC 3 showed that as the concentration of silver nanoparticles increased, a decrease in the percentage cell viability was observed with IC50 values being 60.09 and 57.43 µg/ml respectively. Therefore, through this study, it could be said that extracellular synthesis of silver nanoparticles from Scedosporium was simple, ecofriendly, proving excellent antimicrobial and anticancer agents.

Study on the in vitro Cell Uptake and Toxicity of Resibufogenin-loaded PLGA-TPGS Nanoparticles / 中国药房

OBJECTIVE:To study the in vitro uptake of Resibufogenin(RBG)lactic acid glycolic acid copolymer-water solu-ble vitamin E (PLGA-TPGS) in human liver cancer HepG2 cells,mouse ascites-type lymphatic metastasis of tumor HCa-F cells, and the toxicity on HepG2 cells. METHODS:RCPTN loading RBG and coumarin-6(C6)were prepared. Fluorescent inverted mi-croscope was used to observe the in vitro uptake by RCPTN HepG2,HCa-F cells. It was divided into negative control group,blank PLGA-TPGS nanoparticles(EPTN)group,5-fluorouracil solution(FS)group,RBG solution(RS)group,RBG/PLGA nanoparti-cles(RPN)group and RPTN group. WST-1 was conducted to investigate the optical density at 450 nm wavelength of HepG2 cells after 24,48,72 h incubated by FS,RS,RPN and RPTN with different final concentrations (1.25,2.5,5,10,20 μg/mL);the cell viability (CV) and half inhibitory concentration (IC50) were calculated. RESULTS:RCPTN distributed around the nucleus of HepG2,HCa-F cells. CV was decreased by RBG concentration increased in RPN group and RPTN group,and decreased by time prolonged;compared with FS group,CV in RPTN group was decreased(PFS>RPN>RPTN;IC50 incubated by RPN and RPTN for 48,72 h was obviously less than that of FS and RS(P<0.05 or P<0.01). CONCLUSIONS:RPTN can deliver RBG in-to HepG2,HCa-F cells,showing inhibition effect on HepG2 cells which is stronger than RPN,RS and FS.

Nanoencapsulação de um peptídeo isolado de anuros: citotoxicidade in vitro em células tumorais humanas / Nanoencapsulation of a peptide isolated from anurans: in vitro cytotoxicity in human tumor cells

A terapia antineoplásica tradicional apresenta algumas limitações que podem ser superadas através da utilização dos lipossomas. Estes nanossistemas de carreamento possibilitam o direcionamento de fármacos e reduzem efeitos secundários. Alguns peptídeos catiônicos sintetizados na pele de anuros apresentam atividade citotóxica seletiva (microorganismos e/ou tumores). Neste sentido, espécies do gênero Phyllomedusa secretam as dermaseptinas. Objetivou-se neste estudo, avaliar a citotoxicidade in vitro da dermaseptina 01 (DS 01) livre e encapsulada em lipossomas unilamelares pequenos (SUVs) em células tumorais humanas. Os lipossomas foram preparados pelo método de hidratação do filme lipídico seguido de sonicação. Foram produzidas formulações neutras e catiônicas, convencionais e furtivas. A citotoxicidade foi analisada em células tumorais de pulmão (NCI-H292), cólon (HT-29) e laringe (HEp-2), pelo ensaio de redução do sal tetrazólio (MTT) em placas de 96 poços. Os lipossomas foram submetidos a testes de estabilidade acelerada e em longo prazo. Em NCI-H292, a DS 01 livre apresentou efeito citostático médio de 35,6%. A encapsulação do peptídeo em lipossomas convencionais neutros aumentou o efeito, ao contrário dos furtivos. Para HT-29 e HEp-2, a DS 01 livre inibiu o crescimento celular em aproximadamente 50%, em média. A encapsulação em lipossomas catiônicos potencializou o efeito; os lipossomas convencionais inibiram na faixa de 80% e os furtivos, mais que 95% para as duas linhagens celulares. A DS 01, um peptídeo catiônico antimicrobiano, apresentou efeito citotóxico in vitro para células tumorais humanas que foi potencializado com a nanoencapsulação...

The conventional anticancer therapies show some limitations that can be overcome by using liposomes. This type of nanocarriers allows preferential targeting of drugs and reduces undesirable secondary effects. Some cationic peptides synthesized by the skin of anurans exhibit selective cytotoxicity (to pathogens and/or tumors). Species of Phyllomedusa secrete dermaseptins (DSs). The aim of this study was to assess the in vitro cytotoxicity of free and small unilamellar vesicle (SUV)-encapsulated dermaseptin 01 (DS 01) in various human tumor cells. Liposomes were prepared by lipid film hydration followed by sonication. Neutral and cationic, conventional and stealth liposomes were produced. Cytotoxicity was analyzed in lung (NCI-H292), colon (HT-29) and larynx (HEp-2) cancer cells, by the tetrazolium reduction method (MTT) carried out in 96-well microplates. Liposomes were tested for accelerated and long-term stability. In NCI-H292, free DS 01 showed a medium cytostatic effect of 36.5%. The conventional neutral liposome encapsulation of peptide increased this effect, whereas the stealth did not. In HT-29 and HEp-2, free DS 01 inhibited the cell growth by approximately 50% on average. The cationic liposome encapsulation was synergic, inhibition being about 80% in conventional and higher than 95% in stealth liposomes for both celllines. Thus, DS 01, an antimicrobial cationic peptide, showed in vitro cytotoxicity to human tumor cells that was potentiated by nanoencapsulation...

Biocompatibility of a non-woven silk fibroin and poly L-lactic acid net and evaluation on its safety / 吉林大学学报(医学版)

Objective To study the biocompatibility of silk fibroin/poly L-lactic acid (SF/PLLA) non-woven network,a kind of new composite tissue engineering nanomaterials,and to explore its possibility as the biological implant materials.Methods The PLLA non-woven network was prepared by electrostatic spinning.Physiological saline as control,the leaching solution was prepared and injected into the mice,then the mice were observed for 2 weeks.The materials were implanted into the back of the mice,and 3-0 suture was used as control.Tissues were collected at 1,2,3,and 4 weeks after operation,dyed by HE staining and then the photos were taken.The tissue reactions in experimental group and control group were observed.The rabbit knee joint cartilage cells were cultured,and then subculture cells were seeded to the surface of materials.After cultured invitro,the adhesion and growth of the cells were observed with inverted optical microscope.The bioactivities of the rabbit knee joint cartilage cells in negative control group(DMEM culture media),experimental group(DMEM containing materials) and positive control group(DMEM containing phenol solution)were determined by MTT assay after cocultured for 24 and 48 h.Results After injection,the body status of the mice in experimental group was the same to the control group.There were little fibroblasts was and a little of lymphocytes and macrophage cells in the materials which were implanted into the back of the mice at the beginning.Then the number of the fibroblasts was increased, but the number of the lymphocytes and macrophage cells did not change obviously.The materials degraded slowly, and the material degraded obviously at 4 weeks.The inflammation of tissue around the material reduced gradually from the 2nd week.The inflammation of tissue around the material was the same to the suture,and sometimes was slighter than the suture.After sed for 24 h,there were cells attaching to the fibers of the material.More and more cells attached to the fibers.The reasult of MTT assay showed that the cytotoxicities in experimental groups were all on LevelⅠ at 24 and 48 h.Except for positive control group,the A values were increased in other groups with the extended response time.At the same time,there was no significant difference in cytotoxicity between experimental group and negative control group(P>0.05)and the A value in experimental group was higher than that in positive control group(P<0.01).Conclusion The SF/PLLA non-woven network scaffold material has good biological compatibility and safety,it could be used as implant material in tissue engineering.

Caracterización del perfil quimiosensibilidad y del estado de ampliación génica de un panel de líneas celulares de origen tumoral pulmonar / Characterization of chemosensitivity profile gene amplification status of a panel of lung cancer cell lines

En este trabajo se presentan los resultados de un análisis de amplificación génica y de sensibilidad a fármacos antineoplásicos, realizado para un panel de líneas celulares de origen tumoral pulmonar. Para los ensayos de quimiosensibilidad las células fueron tratadas durante 48 h con concentraciones variables de taxol, cisplatino, doxorrubicina y 5-fluoracilo. La citotoxicidad de los fármacos se cuantificó usando el ensayo de reducción de resazurina y se reportó en valores de concentración inhibitoria 50 (CI50). Para los análisis de amplificación génica se emplearon sondas TaqMan® dirigidas contra los genes AKT2, PIK3CA, ERBB2, EGFR, c-REL y genes de la familia MYC. El número de copias para cada gen fue calculado usando el método de doble delta Ct, empleando ACTB como gen de referencia y la línea MRC-5 como muestra control. Los resultados mostraron que la viabilidad de todas las líneas celulares se afectó por el tratamiento con taxol, cisplatino y doxorrubicina, pero no con el tratamiento con 5-fluoracilo. Las CI50 calculadas se ubicaron entre 0,38 ± 0,03 µM y 111,3 ± 3,58 µM, siendo el taxol y la doxorrubicina los fármacos más potentes. Del panel evaluado las células NCI-H292 resultaron ser las más sensibles y las células LSPG8G las más resistentes a los fármacos. Interesantemente en las células NCI-H292 ningún gen se encontró amplificado; por el contrario en las células LSPG8G los genes cMYC, MYCN, MYCL y AKT2 mostraron un aumento en el número de copias con respecto al de las células control. Estos resultados sugieren que eventos de amplificación génica podrían contribuir con el fenómeno de quimioresistencia en líneas celulares de cáncer de pulmón, sin embargo otros estudios deben realizarse para confirmar esta hipótesis.

In this paper, we show results of anticancer drug sensitivity assays and studies of gen amplification performed for a panel of lung cancer cell lines. For the chemosensitivity assays the cells were treated for 48 h with different concentrations of taxol, cisplatin, doxorubicin and 5-fluorouracil. The cytotoxic effect of each drug was determined using the resazurin reduction assay and reported in terms of inhibitory concentration 50 (IC50). For the analysis of gene amplification we used TaqMan® probes designed against AKT2, PIK3CA, ERBB2, EGFR, REL and MYC family members. Copy number for each gene was calculated using the delta-delta-CT method, employing ACTB as reference gen and MRC-5 cell line as control sample. In the chemosensitivity assays, we observed a clear decrease in cell viability in the cells treated with taxol, cisplatin and doxorubicin but not in the cells treated with 5-fluorouracil. IC50 values ranging between 0,38± 0,03 µM and 111,3 ±3,58 µM, being the taxol and doxorubicin the most potent drugs. NCI-H292 cell line was the most sensitivity and LSPG8G cell line was the most resistant. Interestingly, NCI-H292 cells did not show increase in the copy numbers for the gene evaluated, in contrast, we observed changes in the gene dosage for cMYC, MYCN, MYCL and AKT2 in LSPG8G cells. These results suggest that gene amplification could contribute to drug resistance in lung cancer cell lines; however, more studies are needed to confirm this hypothesis.

Design and evaluation of liposomal delivery system for L-Asparaginese.

Despite of its effective anti-tumour activity,L-Asparaginase has limited clinical application due to the high rate of clinical hypersensitivity. In an attempt to develop a liposomal drug delivery for L-Asparaginase, enzyme loaded liposomes were formulated using soy lecithin, cholesterol and charge inducers by thin film hydration method. The effect of various components of the liposomes including the concentration of lecithin and cholesterol with or without the charge inducers on the entrapment efficiency and short term invitro cytotoxicity study was systematically investigated. The average particle sizes of the vesicles were found to be 43.2, 35.6 and 65.8 μm respectively for neutral, positive and negative liposomes. The percentage of drug loading was found to be 1.95, 2.39 and 2.35 % respectively for neutral, positive and negative liposomes.The invitro release study of L-Asparaginase was carried out using normal saline as dissolution medium and the release was found to be 86.88, 78.29 and 82.04 % respectively for neutral, positive and negative liposomes. The release of LAsparaginase from liposomes was followed first order kinetics obeying non-Fickian diffusion. A short term cytotoxicity study was carried out using Ehrlich Ascites Carcinoma cells (EAC cells) which revealed that the cytotoxicity concentration CTC50 for pure drug was found to be 64 mcg as compared to liposomal formulation of 50 mcg.