Polybia occidentalis and Polybia fastidiosa venom: a cytogenotoxic approach of effects on human and vegetal cells.

The venoms of wasps are a complex mixture of biologically active compounds, such as low molecular mass compounds, peptides, and proteins. The aim of the study was to evaluate the action of wasp venoms, Polybia occidentalis and Polybia fastidiosa, on the DNA of human leukocytes and on the cell cycle and genetic material of the plant model Lactuca sativa L. (lettuce). The cultured leukocytes were treated with the venoms and then evaluated by the comet assay. On another assay, seeds were exposed to a venom solution; the emitted roots were collected and the occurrence of cell cycle alterations (CCAs) and DNA fragmentation were evaluated by agarose gel electrophoresis and TUNEL assay. The results demonstrated that the venom of both wasps induces several CCAs and reduces the mitotic index (MI) on treated cells. They induced damage on human leukocytes DNA. High frequencies of fragments were observed in cells exposed to P. occidentalis venom, while those exposed to P. fastidiosa showed a high frequency of non-oriented chromosome. Both venoms induced the occurrence of various condensed nuclei (CN). This alteration is an excellent cytological mark to cell death (CD). Additionally, CD was evidenced by positive signals in TUNEL assay, by DNA fragmentation in agarose gel electrophoresis with vegetal cells, and by DNA fragmentation of the human leukocytes evaluated. Furthermore, human leukocytes exposed to the venom of P. fastidiosa had high rate of damage. The data demonstrate that both vegetal and human cells are adequate to evaluate the genotoxicity induced by venoms.

Radiotherapy resistance in chondrosarcoma cells; a possible correlation with alterations in cell cycle related genes.

BACKGROUND: Conventional chondrosarcomas are malignant cartilage tumors considered radioresistant. Nevertheless, retrospective series show a small but significant survival benefit for patients with locally advanced disease treated with radiotherapy. And, in daily practice when considered inoperable their irradiation is an accepted indication for proton beam radiotherapy. Therefore, we investigated the sensitivity of chondrosarcoma cell lines and -tissue samples towards radiotherapy and screened for biomarkers to identify predictors of radiosensitivity. METHODS: Proliferation and clonogenic assays were performed in chondrosarcoma cell lines after γ-radiation in combination with mutant IDH1 inhibitor AGI-5198. In addition, glutathione levels were measured using mass spectrometry. Chondrosarcoma tumor explants were irradiated after which γ-H2AX foci were counted. Mutation analysis was performed using the Ion AmpliSeq™ Cancer Hotspot Panel and immunohistochemical staining's were performed for P-S6, LC-3B, P53, Bcl-2, Bcl-xl and Survivin. Results were correlated with the number of γ-H2AX foci. RESULTS: Chondrosarcoma cell lines were variably γ-radiation resistant. No difference in radiosensitivity, nor glutathione levels was observed after treatment with AGI-5198. Irradiated chondrosarcoma patient tissue presented a variable increase in γ-H2AX foci compared to non-radiated tissue. Samples were divided into two groups, high and low radioresistant, based on the amount of γ-H2AX foci. All four highly resistant tumors exhibited mutations in the pRb pathway, while none of the less radioresistant tumors showed mutations in these genes. CONCLUSIONS: Chondrosarcoma cell lines as well as primary tumors are variably radioresistant, particularly in case of a defective Rb pathway. Whether selection for radiotherapy can be based upon an intact Rb pathway should be further investigated.

Effects of long exposure to spent potliner on seeds, root tips, and meristematic cells of Allium cepa L.

Spent potliner (SPL) is a solid waste generated in the aluminum mining and processing industry. It is sometimes dumped into the environment and leach in contact with water, thereupon affecting living beings, which are likely to be exposed to the waste for long periods. Considering this, we aimed to evaluate the effects of extended exposure to SPL through bioassays using Allium cepa as plant model system. Seeds of A. cepa were either directly exposed to SPL (continuous exposure) or first germinated in water and then exposed to SPL (discontinuous exposure). The germination rate was determined from 24 to 192 h of exposure. The maximum effects of SPL on germination were observed after 96 h in both exposure approaches. For the parameter root elongation, the discontinuous treatment was more efficient in demonstrating differences among the applied SPL concentrations (60% of reduction). Microscopic analysis was carried out in root tip cells discontinuously exposed to SPL for 96 h. A mitodepressive effect was observed (above 50%), as well as increased rate of chromosome abnormalities (up to 100-fold) and induction of cell death. The consequences of exposure to SPL for longer periods are discussed.

Graphene oxide and reduced graphene oxide induced neural pheochromocytoma-derived PC12 cell lines apoptosis and cell cycle alterations via the ERK signaling pathways.

Given the novel applications of graphene materials in biomedical and electronics industry, the health hazards of these particles have attracted extensive worldwide attention. Although many studies have been performed on graphene material-induced toxic effects, toxicological data for the effect of graphene materials on the nervous system are lacking. In this study, we focused on the biological effects of graphene oxide (GO) and reduced graphene oxide (rGO) materials on PC12 cells, a type of traditional neural cell line. We found that GO and rGO exerted significant toxic effects on PC12 cells in a dose- and time-dependent manner. Moreover, apoptosis appeared to be a response to toxicity. A potent increase in the number of PC12 cells at G0/G1 phase after GO and rGO exposure was detected by cell cycle analysis. We found that phosphorylation levels of ERK signaling molecules, which are related to cell cycle regulation and apoptosis, were significantly altered after GO and rGO exposure. In conclusion, our results show that GO has more potent toxic effects than rGO and that apoptosis and cell cycle arrest are the main toxicity responses to GO and rGO treatments, which are likely due to ERK pathway regulation.

Rapid Assessment of Genotoxicity by Flow Cytometric Detection of Cell Cycle Alterations.

Flow cytometry is a convenient method for the determination of genotoxic effects of environmental pollution and can reveal genotoxic compounds in unknown environmental mixtures. It is especially suitable for the analyses of large numbers of samples during monitoring programs. The speed of detection is one of the advantages of this technique which permits the acquisition of 104-105 cells per sample in 5 min. This method can rapidly detect cell cycle alterations resulting from DNA damage. The outcome of such an analysis is a diagram of DNA content across the cell cycle which indicates cell proliferation, G2 arrests, G1 delays, apoptosis, and ploidy.Here, we present the flow cytometric procedure for rapid assessment of genotoxicity via detection of cell cycle alterations. The described protocol simplifies the analysis of genotoxic effects in marine environments and is suitable for monitoring purposes. It uses marine mussel cells in the analysis and can be adapted to investigations on a broad range of marine invertebrates.

Phytotoxicity and cytotoxicity of Lepidaploa rufogrisea (Asteraceae) extracts in the plant model Lactuca sativa (Asteraceae) / Fitotoxicidad y citotoxicidad de los extractos de Lepidaploa rufogrisea (Asteraceae) en la planta modelo Lactuca sativa (Asteraceae)

AbstractLepidaploa rufogrisea is a promising candidate in the search of natural herbicides, with compounds (sesquiterpene lactones-glaucolide B) which have been reported to present phytotoxic potential against weeds. Thus, this work aimed to accomplish the phytochemical investigation of L. rufogrisea as well as the evaluation of its biological effects on the plant model Lactuca sativa. To this end, different extracts (aqueous, hydroalcoholic and hexane) of L. rufogrisea at different concentrations [0.25, 0.5, 1.0, 2.0 and 4.0 % (m/v)] were evaluated from macroscopic (percentage of germinated seeds, germination speed index and root length) and microscopic analysis (mitotic index and cell cycle alterations) in plantlets (150 per treatment) and meristematic cells (10 000 per treatment) of L. sativa var. Cristina. The extracts decreased the germination speed index and also the root growth of L. sativa plantlets. Cell proliferation was not significantly affected by the extracts. However, some chromosome and nuclear alterations in the mitotic spindle were observed, due to the interference of allelochemicals present in L. rufogrisea. Moreover, the plant model L. sativa demonstrated to be efficient for this kind of studies.

ResumenLepidaploa rufogrisea es una candidata prometedora en la búsqueda de herbicidas naturales, con compuestos (lactano sesquiterpeno- glaucolíde B) los cuales han sido reportados por presentar un potencial fitotóxico contra las malas hierbas. Por lo tanto, este trabajo tuvo como objetivo lograr la investigación fitoquímica de L. rufogrisea, así como también la evaluación de sus efectos biológicos sobre el modelo de planta Lactuca sativa L. Para esto, diferentes extractos (acuosos, hidroalcohólicos y hexanos) de L. rufogrisea fueron evaluados. El porcentaje de semillas germinadas, el índice de velocidad de germinación y la longitud de raíz de plántulas de L. sativa var. Cristina fueron evaluadas, así como también la citotoxicidad de los extractos en las células meristemáticas. Los extractos disminuyeron el índice de velocidad de germinación y también el crecimiento de las raíces de las plántulas de L. sativa. La proliferación celular no se vio significativamente afectada por los extractos. Sin embargo, se observaron alteraciones cromosómicas y nucleares, debido a la interferencia de los aleloquímicos presentes en L. rufogrisea en el huso mitótico. Además, el modelo de planta L. sativa demostró ser eficiente para tales estudios.

Antitumor potential of the myotoxin BthTX-I from Bothrops jararacussu snake venom: evaluation of cell cycle alterations and death mechanisms induced in tumor cell lines.

BACKGROUND: Phospholipases A2 (PLA2s) are abundant components of snake venoms that have been extensively studied due to their pharmacological and pathophysiological effects on living organisms. This study aimed to assess the antitumor potential of BthTX-I, a basic myotoxic PLA2 isolated from Bothrops jararacussu venom, by evaluating in vitro processes of cytotoxicity, modulation of the cell cycle and induction of apoptosis in human (HL-60 and HepG2) and murine (PC-12 and B16F10) tumor cell lines. METHODS: The cytotoxic effects of BthTX-I were evaluated on the tumor cell lines HL-60 (promyelocytic leukemia), HepG2 (human hepatocellular carcinoma), PC-12 (murine pheochromocytoma) and B16F10 (murine melanoma) using the MTT method. Flow cytometry technique was used for the analysis of cell cycle alterations and death mechanisms (apoptosis and/or necrosis) induced in tumor cells after treatment with BthTX-I. RESULTS: It was observed that BthTX-I was cytotoxic to all evaluated tumor cell lines, reducing their viability in 40 to 50 %. The myotoxin showed modulating effects on the cell cycle of PC-12 and B16F10 cells, promoting delay in the G0/G1 phase. Additionally, flow cytometry analysis indicated cell death mainly by apoptosis. B16F10 was more susceptible to the effects of BthTX-I, with ~40 % of the cells analyzed in apoptosis, followed by HepG2 (~35 %), PC-12 (~25 %) and HL-60 (~4 %). CONCLUSIONS: These results suggest that BthTX-I presents antitumor properties that may be useful for developing new therapeutic strategies against cancer.

Antitumor potential of the myotoxin BthTX-I from Bothrops jararacussu snake venom: evaluation of cell cycle alterations and death mechanisms induced in tumor cell lines

Background Phospholipases A 2 (PLA 2 s) are abundant components of snake venoms that have been extensively studied due to their pharmacological and pathophysiological effects on living organisms. This study aimed to assess the antitumor potential of BthTX-I, a basic myotoxic PLA 2isolated from Bothrops jararacussu venom, by evaluating in vitro processes of cytotoxicity, modulation of the cell cycle and induction of apoptosis in human (HL-60 and HepG2) and murine (PC-12 and B16F10) tumor cell lines. Methods The cytotoxic effects of BthTX-I were evaluated on the tumor cell lines HL-60 (promyelocytic leukemia), HepG2 (human hepatocellular carcinoma), PC-12 (murine pheochromocytoma) and B16F10 (murine melanoma) using the MTT method. Flow cytometry technique was used for the analysis of cell cycle alterations and death mechanisms (apoptosis and/or necrosis) induced in tumor cells after treatment with BthTX-I. Results It was observed that BthTX-I was cytotoxic to all evaluated tumor cell lines, reducing their viability in 40 to 50 %. The myotoxin showed modulating effects on the cell cycle of PC-12 and B16F10 cells, promoting delay in the G0/G1 phase. Additionally, flow cytometry analysis indicated cell death mainly by apoptosis. B16F10 was more susceptible to the effects of BthTX-I, with ~40 % of the cells analyzed in apoptosis, followed by HepG2 (~35 %), PC-12 (~25 %) and HL-60 (~4 %). Conclusions These results suggest that BthTX-I presents antitumor properties that may be useful for developing new therapeutic strategies against cancer.(AU)

Antitumor potential of the myotoxin BthTX-I from Bothrops jararacussu snake venom: evaluation of cell cycle alterations and death mechanisms induced in tumor cell lines

Background Phospholipases A 2 (PLA 2 s) are abundant components of snake venoms that have been extensively studied due to their pharmacological and pathophysiological effects on living organisms. This study aimed to assess the antitumor potential of BthTX-I, a basic myotoxic PLA 2isolated from Bothrops jararacussu venom, by evaluating in vitro processes of cytotoxicity, modulation of the cell cycle and induction of apoptosis in human (HL-60 and HepG2) and murine (PC-12 and B16F10) tumor cell lines. Methods The cytotoxic effects of BthTX-I were evaluated on the tumor cell lines HL-60 (promyelocytic leukemia), HepG2 (human hepatocellular carcinoma), PC-12 (murine pheochromocytoma) and B16F10 (murine melanoma) using the MTT method. Flow cytometry technique was used for the analysis of cell cycle alterations and death mechanisms (apoptosis and/or necrosis) induced in tumor cells after treatment with BthTX-I. Results It was observed that BthTX-I was cytotoxic to all evaluated tumor cell lines, reducing their viability in 40 to 50 %. The myotoxin showed modulating effects on the cell cycle of PC-12 and B16F10 cells, promoting delay in the G0/G1 phase. Additionally, flow cytometry analysis indicated cell death mainly by apoptosis. B16F10 was more susceptible to the effects of BthTX-I, with ~40 % of the cells analyzed in apoptosis, followed by HepG2 (~35 %), PC-12 (~25 %) and HL-60 (~4 %). Conclusions These results suggest that BthTX-I presents antitumor properties that may be useful for developing new therapeutic strategies against cancer.