Influence of chromium (III), cobalt (II) and their mixtures on cell metabolic activity

Chromium (III) and cobalt (II) are necessary elements required for the proper functioning of the organism, but their excess can cause toxic effects. They are the basic components of implants and are also commonly used in medicine as components of dietary supplements, vitamin and mineral products and energy drinks. The aim of this study was to investigate the effect of cobalt (II) and chromium (III) and their combination on BJ cells. In the study, BJ cells were exposed to CoCl2 or CrCl3 at concentrations ranging from 100 to 1400 µM, and the cytotoxicity of chromium (III) and cobalt (II) and their mixtures was assessed by MTT reduction, LDH release and NRU assays. The outcome of this work reveals the cytotoxic effects of chromium (III) and cobalt (II) and their mixtures on BJ cells. In the cytotoxicity assays, at low concentrations of CoCl2 and CrCl3, stimulation of cell proliferation was observed. In higher concentrations, the cell viability decreased for the tested line in all the assays. During the simultaneous incubation of fibroblasts with 200 µM of CrCl3 and 1000 µM of CoCl2, antagonism was observed: chromium (III) at the concentration of 200 µM induced protection from cobalt (II) toxicity; in the case of interaction of chromium chloride at 1000 µm and cobalt chloride at 200 µM, the protective effect of CrCl3 on CoCl2 was not observed. In the latter case, synergism between these elements was noted. Our work indicates that cobalt (II) and chromium (III) show cytotoxic properties. These metals have a destructive effect on the cell membrane, lysosomes and mitochondria, which leads to disorders of cell metabolism.

Nanocarriers of Miconazole or Fluconazole: Effects on Three-Species Candida Biofilms and Cytotoxic Effects In Vitro

The contribution of different Candida species in oral fungal infections has stimulated the search for more effective therapies. This study assessed the antibiofilm effects of nanocarriers of miconazole (MCZ) or fluconazole (FLZ) on Candida biofilms, and their cytotoxic effects on murine fibroblasts. Three-species biofilms (Candida albicans/Candida glabrata/Candida tropicalis) were formed on 96-well plates, and they were treated with nanocarriers (iron oxide nanoparticles coated with chitosan—“IONPs-CS”) of MCZ or FLZ at 39/78/156 µg/mL; antifungals alone at 156 µg/mL and artificial saliva were tested as positive and negative controls, respectively. Biofilms were analyzed by colony forming units (CFU), biomass, metabolic activity, and structure/viability. The cytotoxicity (L929 cells) of all treatments was determined via 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) reduction assay. Data were submitted to one- or two-way ANOVA, followed by Tukey’s or Fisher LSD’s tests (p < 0.05). IONPs-CS-MCZ at 78 µg/mL promoted similar antibiofilm and cytotoxic effects compared with MCZ at 156 µg/mL. In turn, IONPs-CS-FLZ at 156 µg/mL was overall the most effective FLZ antibiofilm treatment, surpassing the effects of FLZ alone; this nanocarrier was also less cytotoxic compared with FLZ alone. It can be concluded that both nanocarriers are more effective alternatives to fight Candida biofilms compared with their respective positive controls in vitro, being a promising alternative for the treatment of oral fungal infections.

Volatile oils from Philodendron meridionale Buturi & Sakur: chemical composition and in vitro effect against Ctenocephalides felis and leukemia cells

Essential oils are an important natural source of pesticides and can be suitable used in ectoparasite control. The goal of this paper was to investigate the chemical composition of the volatile oils obtained from leaves and stems of Philodendron meridionale Buturi & Sakur by gas chromatography/mass spectrometry and to evaluate their insecticidal effect against Ctenocephalides felis Bouché adult insects and their cytotoxicity on human Jurkat leukemic T-cells by 3-(4,5-dimethylthiazol-2-yl)-2,5diphenyltetrazoluim bromide (MTT) reduction method. The main compound was ent-kaur-16-ene at concentrations of 20.78 and 22.46% for volatile oils from leaves and stems, respectively. These volatile oils provided insecticidal effect against C. felis with an average knockdown time of 9.61 and 7.35 min for fumigant test performed with the leaves and stems samples, respectively. Considering the topical application, these P. meridionale volatile oils extracted from leaves and stems demonstrated average knockdown values of 14.70 and 20.0 min, respectively. A very low cytotoxic effect of 9.48 and 11.32% was observed for P. meridionale volatile oils from leaves and stems even at the highest tested concentration of 400 μg.mL-1, respectively. Therefore, it is possible to propose a biological selectivity to the volatile oils under study, which guarantees, at the same time, effect against this insect with no evident damage to human biological system.

Apamin-induced alterations in J774 1.6 macrophage metabolism

ABSTRACT Among the immune system cells, macrophages have an important role. Apamin, a bee venom constituent, is important in the defense of these insects. Thus, we aimed to evaluate the metabolism of J774 1.6 macrophage cell line when exposed to isolated and purified apamin, using cytotoxicity tests by MTT reduction and analysis by flow cytometry (apoptosis / necrosis, production of reactive oxygen species (ROS), membranous lipoperoxidation (LPO), electrical potential of the mitochondrial membrane (mMP) and DNA fragmentation). None of the tested concentrations (10 to 100μg/mL) were cytotoxic according to MTT reductions. Apoptosis rates decreased at concentrations of 2.5, 5.0, and 10.0μg/mL (P<0.05), while necrosis rates increased (P<0.05). However, rates of healthy cells at the highest tested concentration (10μg/mL) did not differ from control (P>0.05). Apamin did not alter ROS, LPO, or DNA fragmentation. Therefore, all analyzed concentrations (1.25 to 10μg/mL) decreased mMP. Such decrease in apoptosis might be due to a suppression of mitochondrial pro-apoptotic messengers, as this peptide causes no oxidative stress, lipid peroxidation, and DNA damage. Highly sensitive techniques are majorly important for proper interpretation of cellular toxicity mechanisms, combined with routine laboratory methods.

Novel Colloidal Nanocarrier of Cetylpyridinium Chloride: Antifungal Activities on Candida Species and Cytotoxic Potential on Murine Fibroblasts

Nanocarriers have been used as alternative tools to overcome the resistance of Candida species to conventional treatments. This study prepared a nanocarrier of cetylpyridinium chloride (CPC) using iron oxide nanoparticles (IONPs) conjugated with chitosan (CS), and assessed its antifungal and cytotoxic effects. CPC was immobilized on CS-coated IONPs, and the nanocarrier was physico-chemically characterized. Antifungal effects were determined on planktonic cells of Candida albicans and Candida glabrata (by minimum inhibitory concentration (MIC) assays) and on single- and dual-species biofilms of these strains (by quantification of cultivable cells, total biomass and metabolic activity). Murine fibroblasts were exposed to different concentrations of the nanocarrier, and the cytotoxic effect was evaluated by MTT reduction assay. Characterization methods confirmed the presence of a nanocarrier smaller than 313 nm. IONPs-CS-CPC and free CPC showed the same MIC values (0.78 µg mL−1). CPC-containing nanocarrier at 78 µg mL−1 significantly reduced the number of cultivable cells for all biofilms, surpassing the effect promoted by free CPC. For total biomass, metabolic activity, and cytotoxic effects, the nanocarrier and free CPC produced statistically similar outcomes. In conclusion, the IONPs-CS-CPC nanocarrier was more effective than CPC in reducing the cultivable cells of Candida biofilms without increasing the cytotoxic effects of CPC, and may be a useful tool for the treatment of oral fungal infections.

Sequenced Combinations of Cisplatin and Selected Phytochemicals towards Overcoming Drug Resistance in Ovarian Tumour Models

In the present study, cisplatin, artemisinin, and oleanolic acid were evaluated alone, and in combination, on human ovarian A2780, A2780ZD0473R, and A2780cisR cancer cell lines, with the aim of overcoming cisplatin resistance and side effects. Cytotoxicity was assessed by MTT reduction assay. Combination index (CI) values were used as a measure of combined drug effect. MALDI TOF/TOF MS/MS and 2-DE gel electrophoresis were used to identify protein biomarkers in ovarian cancer and to evaluate combination effects. Synergism from combinations was dependent on concentration and sequence of administration. Generally, bolus was most synergistic. Moreover, 49 proteins differently expressed by 2 ≥ fold were: CYPA, EIF5A1, Op18, p18, LDHB, P4HB, HSP7C, GRP94, ERp57, mortalin, IMMT, CLIC1, NM23, PSA3,1433Z, and HSP90B were down-regulated, whereas hnRNPA1, hnRNPA2/B1, EF2, GOT1, EF1A1, VIME, BIP, ATP5H, APG2, VINC, KPYM, RAN, PSA7, TPI, PGK1, ACTG and VDAC1 were up-regulated, while TCPA, TCPH, TCPB, PRDX6, EF1G, ATPA, ENOA, PRDX1, MCM7, GBLP, PSAT, Hop, EFTU, PGAM1, SERA and CAH2 were not-expressed in A2780cisR cells. The proteins were found to play critical roles in cell cycle regulation, metabolism, and biosynthetic processes and drug resistance and detoxification. Results indicate that appropriately sequenced combinations of cisplatin with artemisinin (ART) and oleanolic acid (OA) may provide a means to reduce side effects and circumvent platinum resistance.

Cytotoxicity of Kelakai (Stenochlaena palustris) Extract to MCF-7 Breast Cancer Cell

Breast cancer is reported to rank fifth among all types of cancer with a case of death of 6.6%. In the Central Kalimantan region, early examinations of cancers including breast cancer were carried out and 247 tumors in the breast were identified (1.76%). Kelakai (Stenochlaena palustris) as one of the typical plants of Kalimantan which contain natural chemical constituents has been reported to be effective as an anti-inflammatory and antioxidant, so that with this potential can be developed to overcome diseases associated with it, one of them is breast cancer. This study aimed to examine the cytotoxicity of MCF-7 breast cancer cells using ethanol extract. The cytotoxicity assay of kelakai extract against MCF-7 cells conducted in vitro by the MTT reduction method. The variation of concentration used is 1000; 500; 250; 125; 62.5; 31.5; and 15.625 µg/ml, doxorubicin as a positive control was performed in a concentration of 1 μg / ml. The results of the cytotoxicity assay showed that the kelakai extract had a toxic effect on MCF-7 cells with an IC50 value of 493.57 µg / ml.

Patterns of MTT reduction in mammalian spermatozoa

MTT is widely used in biology as a probe for cell viability by virtue of its ability to generate deposits of insoluble formazan at sites of intense oxidoreductase activity. This response is generally held to reflect mitochondrial redox activity; however, extra-mitochondrial MTT reduction has also been recorded in certain cell types. Given this background, we set out to determine the major sites of formazan deposition in mammalian spermatozoa. In the mouse, most MTT reduction took place within the extensive mitochondrial gyres, with a single minor site of formazan deposition on the sperm head. By contrast, human spermatozoa generally displayed small disorganized midpieces exhibiting moderate MTT reduction activity accompanied by a major extra-mitochondrial formazan deposit on various locations in the sperm head from the neck to the anterior acrosome. Equine spermatozoa presented a combination of these two patterns, with major formazan deposition in the mitochondria accompanied by an extra-mitochondrial formazan deposit in around 20% of cells. The functionality of human spermatozoa was positively associated with the presence of an extra-mitochondrial formazan granule. Subsequent studies indicated that this extra-mitochondrial activity was suppressed by the presence of diphenylene iodonium, zinc, 2-deoxyglucose, co-enzyme Q, an SOD mimetic and NADPH oxidase inhibitors. We conclude that the pattern of MTT reduction to formazan by spermatozoa is species specific and conveys significant information about the relative importance of mitochondrial vs extra-mitochondrial redox activity that, in turn, defines the functional qualities of these cells.

Sequenced Combinations of Cisplatin and Selected Phytochemicals Towards Overcoming Drug Resistance in Ovarian Tumour Models

Background: In the present study, cisplatin, artemisinin and oleanolic acid were evaluated alone and in combination, on human ovarian A2780, A2780ZD0473R and A2780cisR cancer cell lines with aim of overcoming cisplatin resistance and side effects. Methods: Cytotoxicity was assessed by MTT reduction assay. CI values were used as a measure of combined drug effect. MALDI TOF/TOF MS/MS and 2-DE gel electrophoresis were used to identify protein biomarkers in ovarian cancer and to evaluate combination effects. Results: Synergism from combinations was dependent on concentration and sequence of administration. Generally, bolus was most synergistic. 49 proteins differently expressed by 2 &ge; fold were: CYPA, EIF5A1, Op18, p18, LDHB, P4HB, HSP7C, GRP94, ERp57, mortalin, IMMT, CLIC1, NM23, PSA3,1433Z, and HSP90B were down-regulated, whereas hnRNPA1, hnRNPA2/B1, EF2, GOT1, EF1A1, VIME, BIP, ATP5H, APG2, VINC, KPYM, RAN, PSA7, TPI, PGK1, ACTG and VDAC1 were up-regulated, while TCPA, TCPH, TCPB, PRDX6, EF1G, ATPA, ENOA, PRDX1, MCM7, GBLP, PSAT, Hop, EFTU, PGAM1, SERA and CAH2 were not-expressed in A2780cisR cells. The proteins were found to play critical roles in cell cycle regulation, metabolism and biosynthetic processes and drug resistance and detoxification. Conclusion: Results indicate that appropriately sequenced combinations of cisplatin with ART and OA may provide a means to reduce side effects and circumvent platinum resistance.

Semi-pilot production of xanthan gum using nejayote as culture medium substrate

Background: Xanthan gum is an industrialized polysaccharide produced by Xanthomonas genus. Alternative carbon sources for Xanthomonas culturing may help reducing its production cost. Nejayote, a residue from maize nixtamalization process, is an alternative culture medium substrate to produce xanthan. In this study, industrial and semi-industrial nejayote alone or in combination with supplements, were tested for xanthan production by Xantomonas campestris (1 × 108 cells/mL inoculum), using YGC medium as control, in 100 mL and 1 L volumes. Cellular growth was determined by the colorimetric MTT reduction assay (OD 540 ). Xanthan pyruvate and acetyl groups from nejayote plus supplement cultures in the bioreactor were evaluated (OD 570). Results: Industrial nejayote steadily produced up to 4 g/L xanthan, as compared with YGC medium control, which increased its production over time up to 9.3 g/L at 96 h. Cellular activity assay revealed that the highest values after 24 h (3.88 and 2.71 OD 540 for YGC and industrial nejayote, respectively). Nejayote supplemented with MgSO4 •7 H2O resulted in the highest xanthan production (10.8 g/L), but low cell growth (3.6 and 1.82 OD 540 for YGC and nejayote plus supplement, respectively), after 96 h of culture. Furthermore, gum yield reached up to 6 g/L and 1.9 OD 540, after 96 h of nejayote plus supplement culture, using a 14 L bioreactor. Xanthan pyruvate and acetyl groups from nejayote plus supplement cultures in the bioreactor resulted in similar amounts (0.107 and 0.130 OD 570 , respectively), compared with a commercial biopolymer (0.148 and 0.127 OD 570 respectively). Conclusions: X. campestris effectively grew and produced industrial-quality xanthan gum in nejayote substrate-containing culture medium, thus providing an inexpensive alternative for bioremediation.