Adulterated dietary supplements commercialized in Brazil: development of a screening method and a preliminary study of cytotoxicity.

The high consumption of dietary supplements was a fundamental driver for the creation of the regulatory framework by the Brazilian governmental authorities. However, the regulatory agencies lack official low-cost methodologies to evaluate the quality of food supplements. A preliminary screening method by HPLC-DAD was proposed and validated for screening and quantification of adulterants in dietary supplements. The limits of detection and quantification were <0.11 and 0.37 µg.g-1, respectively. The method was applied for the investigation of ten unauthorized substances (spironolactone, hydrochlorothiazide, furosemide, clenbuterol, testosterone, testosterone propionate, yohimbine, vardenafil, tadalafil, and sildenafil) with a time of analysis of <5 min. Sixteen percent of the 44 samples analyzed had at least one adulterant at or above therapeutic concentrations. Subsequently, in vitro evaluations were performed of the potential cytotoxicity to evaluate the cell viability, DNA damage, determination of nitric oxide levels, and quantification of reactive oxygen species. Despite the necessity of further studies, the results indicate a relationship between the presence of adulterants in food supplements and a potential cytotoxic effect.

Impact of CAD/CAM burs lifetime on surface properties of dental zirconia: Implications for biocompatibility of custom abutments.

OBJECTIVE: To evaluate the effect of the deterioration of computer aided design/computer aided manufacturing (CAD/CAM) burs during zirconia milling, on surface roughness, contact angle, and fibroblast viability. MATERIALS AND METHODS: Ceramic blocks were milled and 75 ceramic disks (8 × 1.5 mm) made and allocated into three groups (n = 25): G1-brand new 2L and 1L burs, G2-2L bur at the end of lifetime and brand new 1L bur and G3-both burs at the end of their lifetimes. Roughness (Ra, Rq, and Rz) was evaluated using a 3D optical profilometer, the contact angle by the sessile drop method and the cell viability of the mouse NIH/3T3 fibroblast, using the Alamar Blue assay at intervals of 24, 48, and 72 h (ISO 10993-5). Data were analyzed by one-way ANOVA and Kruskal-Wallis tests (p ≤ 0.05). RESULTS: Roughness increased as the burs deteriorated and G3 (0.27 ± 0.04) presented a higher value for Ra (p < 0.001). The highest contact angle was observed in G3 (86.2 ± 2.66) when compared with G1 (63.7 ± 12.49) and G2 (75.3 ± 6.36) (p < 0.001). Alamar Blue indicated an increase in cell proliferation, with no significant differences among the groups at 24 and 72 h (p > 0.05). CONCLUSIONS: The deterioration of the burs increased the surface roughness and decreased the wettability, but did not interfere in cell viability and proliferation. CLINICAL SIGNIFICANCE: The use of custom zirconia abutments represents an effective strategy for single crowns restorations. Our findings suggest that these abutments can be efficiently milled using CAD/CAM burs within their recommended lifetime.

Ascorbic acid and ascorbyl palmitate-loaded liposomes: Development, characterization, stability evaluation, in vitro security profile, antimicrobial and antioxidant activities.

The objective of this work was to prepare and characterize liposomes containing co-encapsulated ascorbic acid (AA) and ascorbyl palmitate (AP), as well as to evaluate their stability, cytotoxicity, antioxidant, and antimicrobial activity. Through the pre-formulation studies, it was possible to improve the formulation, as leaving it more stable and with a greater antioxidant activity, resulting in a formulation designated LIP-AAP, with 161 nm vesicle size, 0.215 polydispersity index, -31.7 mV zeta potential, and pH of 3.34. Encapsulation efficiencies were 37% for AA and 79% for AP, and the content was 1 mg/mL for each compound. The optimized liposomes demonstrated stability under refrigeration for 60 days, significant antioxidant activity (31.4 µMol of TE/mL), and non-toxicity, but no antimicrobial effects against bacteria and fungi were observed. These findings confirm that the co-encapsulated liposomes are potent, stable antioxidants that maintain their physical and chemical properties under optimal storage conditions.

Supercritical Carbon Dioxide Extraction of Coumarins from the Aerial Parts of Pterocaulon polystachyum.

Pterocaulon polystachyum is a species of pharmacological interest for providing volatile and non-volatile extracts with antifungal and amebicidal properties. The biological activities of non-volatile extracts may be related to the presence of coumarins, a promising group of secondary metabolites. In the present study, leaves and inflorescences previously used for the extraction of essential oils instead of being disposed of were subjected to extraction with supercritical CO2 after pretreatment with microwaves. An experimental design was followed to seek the best extraction condition with the objective function being the maximum total extract. Pressure and temperature were statistically significant factors, and the optimal extraction condition was 240 bar, 60 °C, and pretreatment at 30 °C. The applied mathematical models showed good adherence to the experimental data. The extracts obtained by supercritical CO2 were analyzed and the presence of coumarins was confirmed. The extract investigated for cytotoxicity against bladder tumor cells (T24) exhibited significant reduction in cell viability at concentrations between 6 and 12 µg/mL. The introduction of green technology, supercritical extraction, in the exploration of P. polystachyum as a source of coumarins represents a paradigm shift with regard to previous studies carried out with this species, which used organic solvents. Furthermore, the concept of circular bioeconomy was applied, i.e., the raw material used was the residue of a steam-distillation process. Therefore, the approach used here is in line with the sustainable exploitation of native plants to obtain extracts rich in coumarins with cytotoxic potential against cancer cells.

Amantadine mitigates the cytotoxic and genotoxic effects of doxorubicin in SH-SY5Y cells and reduces its mutagenicity.

Amantadine (AMA) is a useful drug in neuronal disorders, but few studies have been performed to access its toxicological profile. Conversely, doxorubicin (Dox) is a well-known antineoplastic drug that has shown neurotoxic effects leading to cognitive impairment. The aims of this study are to evaluate the cytotoxic, genotoxic, and mutagenic effects of AMA, as well as its possible protective actions against deleterious effects of Dox. The Salmonella/microsome assay was performed to assess mutagenicity while cytotoxicity and genotoxicity were evaluated in SH-SY5Y cells using MTT and comet assays. Possible modulating effects of AMA on the cytotoxicity, genotoxicity, and mutagenicity induced by Dox were evaluated through cotreatment procedures. Amantadine did not induce mutations in the Salmonella/microsome assay and decreased Dox-induced mutagenicity in the TA98 strain. AMA reduced cell viability and induced DNA damage in SH-SY5Y cells. In cotreatment with Dox, AMA attenuated the cytotoxicity of Dox and showed an antigenotoxic effect. In conclusion, AMA does not induce gene mutations, although it has shown a genotoxic effect. Furthermore, AMA decreases frameshift mutations induced by Dox as well as the cytotoxic and genotoxic effects of Dox in SH-SY5Y cells, suggesting that AMA can interfere with Dox mutagenic activity and attenuate its neurotoxic effects.

In vitro assessment of the genotoxic and cytotoxic effects of clarified açai (Euterpe oleracea MART) extract in a gastric cancer cell line (AGP01 cells).

Açaí (Euterpe oleracea MART) is a fruit of great importance for the Amazon region in nutritional, cultural and socioeconomic terms. In recent years, açaí has been the subject of several studies due to its beneficial properties for health, including effects against tumor cells. Therefore, the present work aimed to evaluate in vitro the genotoxic and cytotoxic effects of the clarified extract of açaí juice in a human metastatic gastric cancer cell line (AGP01 cells). For comparison purposes, a non-transformed cell line of African green monkey renal epithelial cells (VERO cells) was used. The viability assay by resazurin reduction, the comet assay, the determination of cell death by differential fluorescent dyes and the wound healing migration assay were performed. A reduction in viability was observed only in the AGP01 line within 72 h. There was no genotoxic damage or cell death (through apoptosis or necrosis) in any of the cell lines. However, açaí extract induced motility reduction in both cell lines. The reduction in cell viability and the induction of the anti-migratory effect in the AGP01 cell line opens perspectives for exploring the potential of açaí as an adjuvant in the treatment of gastric cancer.

Effect of Rapid High-Intensity Light-Curing on Increasing Transdentinal Temperature and Cell Viability: An In Vitro Study.

BACKGROUND: This study investigated effects of rapid high-intensity light-curing (3 s) on increasing transdentinal temperature and cell viability. METHODS: A total of 40 dentin discs (0.5 mm) obtained from human molars were prepared, included in artificial pulp chambers (4.5 × 5 mm), and subjected to four light-curing protocols (n = 5), with a Valo Grand light curing unit: (i) 10 s protocol with a moderate intensity of 1000 mW/cm2 (Valo-10 s); (ii) 3 s protocol with a high intensity of 3200 mW/cm2 (Valo-3 s); (iii) adhesive system + Filtek Bulk-Fill Flow bulk-fill composite resin in 10 s (FBF-10 s); (iv) adhesive system + Tetric PowerFlow bulk-fill composite resin in 3 s (TPF-3 s). Transdentinal temperature changes were recorded with a type K thermocouple. Cell viability was assessed using the MTT assay. Data were analyzed using one-way ANOVA and Tukey tests for comparison between experimental groups (p < 0.05). RESULTS: The 3 s high-intensity light-curing protocol generated a higher temperature than the 10 s moderate-intensity standard (p < 0.001). The Valo-10 s and Valo-3 s groups demonstrated greater cell viability than the FBF-10s and TPF-3 s groups and statistical differences were observed between the Valo-3 s and FBF-10 s groups (p = 0.023) and Valo-3 s and TPF-3 s (p = 0.025), with a potential cytotoxic effect for the FBF-10 s and TPF-3 s groups. CONCLUSIONS: The 3 s rapid high-intensity light-curing protocol of bulk-fill composite resins caused a temperature increase greater than 10 s and showed cell viability similar to and comparable to the standard protocol.

Performance of ß-carotene-loaded nanostructured lipid carriers under dynamic in vitro digestion system: Influence of the emulsifier type.

A better understanding of how emulsifier type could differently influence the behavior of nanostructured lipid carriers (NLC) under the gastrointestinal digestion process, as well as at the cellular level, is of utmost importance for the NLC-based formulations' optimization and risk assessment in the food field. In this study, NLC composed by fully hydrogenated soybean and high-oleic sunflower oils were prepared using soy lecithin (NLC Lß) or Tween 80 (NLC Tß) as an emulsifier. ß-Carotene was entrapped within NLC developed as a promising strategy to overcome ß-carotene's low bioavailability and stability. The effect of emulsifier type on the digestibility of ß-carotene-loaded NLC was evaluated using an in vitro dynamic digestion model mimicking peristalsis motion. The influence of ß-carotene-loaded NLC on cell viability was assessed using Caco-2 cells in vitro. NLC Tß remained stable in the gastric compartment, presenting particle size (PS) similar to the initial NLC (PS: 245.68 and 218.18 nm, respectively), while NLC Lß showed lower stability (PS > 1000 nm) in stomach and duodenum phases. NLC Tß also provided high ß-carotene protection and delivery capacity (i.e., ß-carotene bioaccessibility increased 10-fold). Based on the results of digestion studies, NLC Tß has shown better physical stability during the passage through the in vitro dynamic gastrointestinal system than NLC Lß. Moreover, the developed NLC did not compromise cell viability up to 25 µg/mL of ß-carotene. Thus, the NLC developed proved to be a biocompatible structure and able to incorporate and protect ß-carotene for further food applications. PRACTICAL APPLICATION: The findings of this study hold significant implications for industrial applications in terms of developing nanostructured lipid carriers from natural raw materials widely available and used to produce other lipid-based products in the food industry, as an alternative to synthetic ones. In this respect, the ß-carotene-loaded NLC developed in this study would find a great industrial application in the food industry, which is in constant search to develop functional foods capable of increasing the bioavailability of bioactive compounds.

In vitro toxicity of latex, its terpenoidal fractions and isolated phorbol esters from Euphorbia umbellata (Pax) Bruyns on monocytic and melanoma cells.

In Brazil, latex from Euphorbia umbellata (African milk tree) has been increasingly used in folk medicine to treat several types of cancer, including melanoma. The effect of lyophilized latex (LL), its hydroethanolic extract (E80), triterpene (F-TRI)- and diterpene (F-DIT)-enriched fractions, along with six isolated phorbol esters from LL and phorbol 12-myristate 13-acetate (PMA) on J774A.1, THP-1, SK-MEL-28, and B16-F10 cell line viability were evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) method. The compounds were identified by 2D-NMR and HRESIMS. The effect of the LL, extract and fractions on cell viability was also assessed through a resazurin reduction assay. At 100 µg/ml, LL, and its fractions moderately inhibited J774A.1 (37.5-59.5%) and THP-1 (12.6-43.6%) metabolism. LL (IC50 70 µg/ml) and F-TRI (IC50 68 µg/ml) were barely more effective against B16-F10 cells, and only F-TRI exerted an inhibitory effect on SK-MEL-28 cells (IC50 66-75 µg/ml). The samples did not effectively inhibit THP-1 growth (IC50 69-87 µg/ml, assessed by MTT). B16-F10 was susceptible to PMA (IC50 53 µM) and two 12-phenylacetate esters (IC50 56-60 µM), while SK-MEL-28 growth was inhibited (IC50 58 µM) by one of these kinds of esters with an additional 4ß-deoxy structure. Synagrantol A (IC50 39 µM) was as effective as PMA (IC50 47 µM) in inhibiting J774A.1 growth in a dose-dependent manner. Furthermore, an in silico study with target receptors indicated a high interaction of the compounds with the PKC proteins. These results provide useful knowledge on the effect of tigliane-type diterpenes on tumor cell from the perspective of medicinal chemistry.

Effect of Cannabis sativa L. extracts, phytocannabinoids and their acetylated derivates on the SHSY-5Y neuroblastoma cells' viability and caspases 3/7 activation.

BACKGROUND: There is a need for novel treatments for neuroblastoma, despite the emergence of new biological and immune treatments, since refractory pediatric neuroblastoma is still a medical challenge. Phyto cannabinoids and their hemisynthetic derivatives have shown evidence supporting their anticancer potential. The aim of this research was to examine Phytocannabinoids or hemisynthetic cannabinoids, which reduce the SHSY-5Y, neuroblastoma cell line's viability. METHODS: Hexane and acetyl acetate extracts were produced starting with Cannabis sativa L. as raw material, then, 9-tetrahidrocannabinol, its acid counterpart and CBN were isolated. In addition, acetylated derivatives of THC and CBN were synthesized. The identification and purity of the chemicals was determined by High Performance Liquid Chromatography and 1H y 13C Magnetic Nuclear Resonance. Then, the capacity to affect the viability of SHSY-5Y, a neuroblastoma cell line, was examined using the resazurin method. Finally, to gain insight into the mechanism of action of the extracts, phytocannabinoids and acetylated derivatives on the examined cells, a caspase 3/7 determination was performed on cells exposed to these compounds. RESULTS: The structure and purity of the isolated compounds was demonstrated. The extracts, the phytocannabinoids and their acetylated counterparts inhibited the viability of the SHSY 5Y cells, being CBN the most potent of all the tested molecules with an inhibitory concentration of 50 percent of 9.5 µM. CONCLUSION: Each of the evaluated molecules exhibited the capacity to activate caspases 3/7, indicating that at least in part, the cytotoxicity of the tested phytocannabinoids and their hemi-synthetic derivatives is mediated by apoptosis.

A three-dimensional cell culture approach to investigate cytotoxic effects and production of inflammatory mediators by epoxy resin-based and calcium silicate-based endodontic sealer.

OBJECTIVES: The aim of the present study was to assess the cytocompatibility of epoxy resin-based AH Plus Jet (Dentsply De Trey, Konstanz, Germany), Sealer Plus (MK Life, Porto Alegre, Brazil), calcium silicate-based Bio-C Sealer (Angelus, Londrina, PR, Brazil), Sealer Plus BC (MK Life) and AH Plus BC (Dentsply) through a tridimensional (3D) culture model of human osteoblast-like cells. METHODS: Spheroids of MG-63 cells were produced and exposed to fresh root canal sealers extracts by 24 h, and the cytotoxicity was assessed by the Lactate Dehydrogenase assay (LDH). The distribution of dead cells within the microtissue was assessed by fluorescence microscopy, and morphological effects were investigated by histological analysis. The secreted inflammatory mediators were detected in cell supernatants through flow luminometry (XMap Luminex). RESULTS: Cells incubated with AH Plus Jet, AH Plus BC, Sealer Plus BC and Bio-C Sealer extracts showed high rates of cell viability, while the Sealer Plus induced a significant reduction of cell viability, causing reduction on the spheroid structure. Sealer Plus and Seaker Plus BC caused alterations on 3D microtissue morphology. The AH Plus BC extract was associated with the downregulation of secretion of pro-inflammatory cytokines IL-5, IL-7, IP-10 and RANTES. CONCLUSIONS: The new AH Plus BC calcium silicate-based endodontic sealer did not reduce cell viability in vitro, while led to the downregulation of pro-inflammatory cytokines. CLINICAL SIGNIFICANCE: Choosing the appropriate endodontic sealer is a crucial step. AH Plus BC demonstrated high cell viability and downregulation of pro-inflammatory cytokines, appearing reliable for clinical use, while Sealer Plus presented lower cytocompatibility.

Modulation of heat shock protein expression and cytokine levels in MCF-7 cells through photodynamic therapy.

In this study, we assess the impact of photodynamic therapy (PDT) using aluminum phthalocyanine tetrasulfonate (AlPcS4) on the viability and cellular stress responses of MCF-7 breast cancer cells. Specifically, we investigate changes in cell viability, cytokine production, and the expression of stress-related genes. Experimental groups included control cells, those treated with AlPcS4 only, light-emitting diode (LED) only, and combined PDT. To evaluate these effects on cell viability, cytokine production, and the expression of stress-related genes, techniques such as 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay, enzyme-linked immunosorbent assays (ELISA), and real-time quantitative PCR (RT‒qPCR) were employed. Our findings reveal how PDT with AlPcS4 modulates mitochondrial activity and cytokine responses, shedding light on the cellular pathways essential for cell survival and stress adaptation. This work enhances our understanding of PDT's therapeutic potential and mechanisms in treating breast cancer.

Toxicity of REEs, Th, and U: A Biodisponibility, Cytotoxicity, and Bioaccumulation Assessment in Marine Sediment.

In this work, bioaccessibility tests for rare earth elements (REEs), Th, and U in marine sediment were carried out, in addition to complementary tests for cytotoxicity and bioaccumulation for the elements La, Ce, Eu, and Gd. The evaluation of human health risk through dermal absorption and oral ingestion was performed using the hazard quotient (HQ). According to the gastric digestion simulation (SBET), it was observed that the elements Ce and Nd exhibited higher absorption capacities in the human body (> 2 µg g-1). La and Sc presented intermediate concentrations (close to 1 µg g-1), while the remaining elements displayed concentrations below 0.5 µg g-1. In the gastrointestinal digestion extraction stage (PBET), all the elements maintained a similar absorption capacity to that observed in SBET, except for the absorption of Y which increased. The results of the bioaccumulation test conducted with fibroblast cells (L929) indicated that La and Eu had a 25% probability of intracellular accumulation. The cell viability test, with exposure to a standard REEs, Th, and U solution in 2% v v-1 HNO3 medium (until 100 µg mL-1) and an aqueous solution of La2O3, Gd(NO3)3, Ce(NO3)3, and Eu2O3 (until 1000 µg mL-1), did not demonstrate cytotoxic effects on fibroblast cells. Considering the ingestion hazard quotient (HQing) and dermal hazard quotient (HQderm) obtained, it was suggested that there is no significant risk of non-carcinogenic effects (< 1). However, they had higher HQing values compared to HQderm, indicating that REEs pose more significant risk to human health through oral ingestion absorption than dermal absorption.

Development of a Resveratrol Nanoformulation for the Treatment of Diabetic Retinopathy.

Diabetic retinopathy (RD) is a microvascular disease that can cause the formation of fragile neovessels, increasing the risk of hemorrhages and leading to vision loss. Current therapies are based on the intravitreal injection of anti-VEGF (vascular endothelial growth factor), which is invasive and can cause secondary effects. The development of new treatments that complement the current therapies is necessary to improve the patient's outcomes. Nanostructured formulations offer several advantages regarding drug delivery and penetration. In this research, a resveratrol nanosuspension (RSV-NS) was prepared and characterized using dynamic light scattering, field emission scanning electron microscopy, and infrared spectroscopy. The RSV-NS had an average particle size of 304.0 ± 81.21 nm with a PDI of 0.225 ± 0.036, and a spherical-like morphology and uniform particle distribution. Cell viability, proliferation, and migration were tested on endothelial cells (HMRECs). RSV-NS in a concentration of less than 18.75 µM did not have a cytotoxic effect on HMRECs. Likewise, proliferation and migration were significantly reduced compared to the unstimulated control at 37.5 µM. The RSV-NS did not present cytotoxic effects but decreased cell proliferation and migration, indicating that it could provide an important contribution to future medical implementations and could have a high potential to treat this disease.

Oxidative damage analysis and cell viability of Drosophila melanogaster exposed to three different endodontic sealers: an in vivo and ex vivo study.

The aim of this work was to evaluate the toxicological action of AH Plus (AHP), Bio-C Sealer (BCS), and EndoSequence BC Sealer (ESB), using Drosophila melanogaster as the model organism performing in vivo and ex vivo analysis. D. melanogaster were exposed for 10 days to three concentrations (5 mg/ml, 10 mg/ml, and 20 mg/ml) of AHP, BCS, and ESB sealers mixed with 10 ml of standard diet. During this period, the mortality of flies was evaluated. On the 11th day, the locomotor activity test was performed and the flies were euthanized for oxidative damage analysis (reactive species and lipid peroxidation) and cell viability (resazurin reduction). For the mortality curves evaluation, the log-rank test (Mantel-Cox) was used. For the analysis of other data, a one-way analysis of variance (ANOVA) was applied, followed by Tukey's post hoc test (α = 0.05). Regarding mortality, there were no significant differences. The locomotor activity was reduced, mainly in the two highest concentrations of AHP and BCS. Besides, reactive species generation was bigger in the AHP 20 mg/ml group. AHP induced a lipid peroxidation increase in all three concentrations tested, when compared to other sealers. Considering cell viability, the two highest concentrations of AHP reduced this parameter; while in other sealers, viability was reduced only in the highest concentration. AHP showed changes in oxidative markers that led to greater damage to the flies.

Chemical profile and biological properties of the Piper corcovadense C.DC. essential oil.

The essential oil from Piper corcovadense D.DC. (EOPc), an important plant belonging to the Piperaceae family, which is commonly found in the northern region of Brazil and poorly explored scientifically, was used in this study. Thus, the EOPc was characterized chemically by Gas Chromatography/Mass Spectrometry (GC/MS) and the antioxidant and antimicrobial activities and their potential effects on cutaneous melanoma (SK-MEL-28) and healthy peripheral blood mononuclear (PBMC) cells were determined. The major compounds identified in the EOPc were: trans-sesquisabinene hydrate, trans-caryophyllene, ß-pinene, trans-ß-farnesene, 14-hydroxycaryophyllene, limonene and p-cymene. The EOPc demonstrated antioxidant activity as evaluated by Folin-Ciocalteu reagent (FC) reducing capacity, DPPH, and ABTS methods. The values found were respectively 5.41 ± 0.17 mg GAE mL-1 (GAE: Gallic acid equivalent), 2.88 ± 0.17 µmol TE mL-1 (TE: Trolox equivalent) and 6.26 ± 0.02 µmol TE mL-1. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined for different bacterial strains. The EOPc at a concentration of 2.61 µg mL-1 exhibited both bactericidal and bacteriostatic properties against Escherichia coli. The EOPc showed potential antitumor activity as it reduced the cell viability of human cutaneous melanoma cells SK-MEL-28. Besides, the EOPc did not exhibit cytotoxic activity against healthy PBMCs, indicating that it does not harm healthy cells at the tested concentrations. The EOPc increased the levels of ROS at concentrations of 250 µg mL-1. The EOPc also did not stimulate the mobilization of endogenous antioxidant defenses, as assessed by total thiol (PSH) and non-protein thiols (NPSH). Thus, the study suggests that the EOPc has antioxidant and antimicrobial properties due to the presence of specific compounds. It also exhibits antitumor potential against cutaneous melanoma cells while showing no cytotoxicity to healthy PBMCs. It directly influenced ROS levels at the highest tested concentration in the cells, suggesting an antitumor effect related to the intrinsic apoptosis pathway. Nevertheless, while the study has initial findings, the results are promising and indicate an attractive biological potential of P. corcovadense, mainly in human cutaneous melanoma cells.

Combined 6-benzylaminopurine and H2O2 stimulate the astaxanthin biosynthesis in Xanthophyllomyces dendrorhous.

Astaxanthin is one of the most attractive carotenoids due to its high antioxidant activity and beneficial biological properties, while Xanthophyllomyces dendrorhous is one of its main microbial sources. Since astaxanthin is synthesized as a response to oxidative stress, several oxidative agents have been evaluated to increase X. dendrorhous astaxanthin yields. However, the extent of the stimulation is determined by the cellular damage caused by the applied oxidative agent. Phytohormones have also been reported as stimulants of astaxanthin biosynthesis acting directly on its metabolic pathway and indirectly promoting cellular resistance to reactive oxygen species. We reasoned that both oxidative agents and phytohormones lead to increased astaxanthin synthesis, but the latter could mitigate the drawbacks of the former. Thus, here, the stimulation on astaxanthin biosynthesis, as well as the cellular and transcriptional responses of wild type X. dendrorhous to phytohormones (6-benzylaminopurine, 6-BAP; abscisic acid, ABA; and indole-3-acetic acid, IAA), and oxidative agents (glutamate, menadione, H2O2, and/or Fe2+) were evaluated as a single or combined treatments. ABA and 6-BAP were the best individual stimulants leading to 2.24- and 2.60-fold astaxanthin biosynthesis increase, respectively. Nevertheless, the effect of combined 6-BAP and H2O2 led to a 3.69-fold astaxanthin synthesis increase (0.127 ± 0.018 mg astaxanthin/g biomass). Moreover, cell viability (> 82.75%) and mitochondrial activity (> 82.2%) remained almost intact in the combined treatment (6-BAP + H2O2) compared to control (< 52.17% cell viability; < 85.3% mitochondrial activity). On the other hand, mRNA levels of hmgR, idi, crtYB, crtR, and crtS, genes of the astaxanthin biosynthetic pathway, increased transiently along X. dendrorhous fermentation due to stimulations assayed in this study. KEY POINTS: • Combined 6-BAP and H2O2 is the best treatment to increase astaxanthin yields in X. dendrorhous. • 6-BAP preserves cell integrity under oxidative H2O2 stress conditions. • 6-BAP and H2O2 increase transcriptional responses of hmgR, idi, and crt family genes transiently.

κ-Carrageenan/sericin polymer matrix modified with different crosslinking agents and thermal crosslinking: Improved release profile of mefenamic acid.

The crosslinking of the polymer matrix with compatible macromolecules results in a three-dimensional network structure that offers an enhancement in the controlled release properties of the material. In this sense, this work aimed to improve the release profile of mefenamic acid (MAC) through crosslinking strategies. κ-Carrageenan/sericin crosslinked blend was obtained by covalent and thermal crosslinking and the different formulations were characterized. The gastroresistant potential and release profile were evaluated in the dissolution assay. The effect and characterization of the particles were investigated. Multiple units presented high entrapment efficiency (94.11-104.25), high drug loading (36.50-47.50 %) and adequate particle size (1.34-1.57 mm) with rough surface and visually spherical shape. The Weibull model showed that drug release occurred by relaxation, erosion and Fickian diffusion. Material stability and absence of MAC -polymer interactions were demonstrated by FTIR and thermogravimetric analysis. DSC showed a stable character of MAC in the drug-loaded beads. Moreover, the application studies of κ-Car/Ser/carboxymethylcellulose in the in vitro intestine mode showed that the crosslinked blend increased cell viability (>85 %), while free MAC exhibited a cytotoxic effect. Finally, the crosslinked k-Car/Ser blend MAC -loaded showed promising properties of a sustained release form of anti-inflammatory drug.

Effect of hydrofluoric acid concentration and aging on the bond strength ceramics to a resin cement

Abstract This study evaluated the influence of hydrofluoric acid (HF) concentration and thermal cycling on the microshear bond strength (µSBS) of a resin luting agent to IPS e.max® CAD and Rosetta® SM. Ceramic specimens (12.0 x 14.0 x 1.5mm) were randomized into 8 groups (n=10) according to HF concentration, commercial brand, and aging. Immediately after polishing, and etching, all specimens were silanized and a layer of adhesive was applied. A PVS mold of 3 mm thickness and 10mm diameter with (four) 1.0mm holes was fabricated, placed on each specimen, and then filled with a resin luting agent. Half of the specimens were subjected to the µSBS test using an Instron at a speed of 1.0 mm/min, following a 24-hour storage in deionized water at 37ºC. The remaining specimens were subjected to thermal cycling (5ºC-55ºC, 30 seconds per bath) and µSBS. The data were evaluated utilizing a three-way ANOVA and Tukey's post-hoc test (α=0.05). Significant differences were found for HF concentration and aging (p<0.0001). No significant difference in µSBS was found for commercial brands (p=0.085). The interaction between brand and HF concentration (p=0.358), brand and aging (p=0.135), and HF concentration and aging (p=0.138) were not statistically significant. The triple interaction among these factors was not statistically significant (p=0.610). In conclusion, the bond strength is affected by the HF concentration. No statistical difference was observed between the two ceramics. Thermal cycling significantly reduced µSBS.

Resumo Este estudo avaliou a influência da concentração do ácido fluorídrico (AF) e da ciclagem térmica na resistência de união ao microcisalhamento (RUµC) de um cimento resinoso para IPS e.max® CAD e Rosetta® SM. Espécimes cerâmicos (12,0 x 14,0 x 1,5mm) foram divididos em 8 grupos (n=10) de acordo com concentração do HF, marca comercial e envelhecimento. Imediatamente após o polimento e condicionamento ácido, todos os espécimes foram silanizados e uma camada de adesivo foi aplicada. Um molde PVS de 3 mm de espessura e 10 mm de diâmetro com (quatro) orifícios de 1,0 mm foi confeccionado, colocado em cada espécime e preenchido com o cimento resinoso. Metade dos espécimes foi submetida ao teste RUµC na Instron a velocidade de 1,0 mm/min, após 24 horas de armazenamento em água deionizada a 37ºC. Os espécimes restantes foram submetidos a ciclagem térmica (5ºC-55ºC, 30 segundos por banho) e a RUµC. Os dados foram avaliados por ANOVA de três fatores e ao teste post-hoc de Tukey (α=0,05). Diferenças significativas foram encontradas para concentração de HF e envelhecimento (p<0,0001). Nenhuma diferença significativa na RUµC foi encontrada para cada marca comercial (p=0,085). A interação entre marca comercial e a concentração do HF (p=0,358), marca comercial e envelhecimento (p=0,135) e concentração do HF e envelhecimento (p=0,138) não foram estatisticamente significativas. A tripla interação entre esses fatores não foi estatisticamente significativa (p=0,610). Concluindo, a resistência de união é afetada pela concentração de HF. Não foi observada diferença estatística entre as duas cerâmicas. A ciclagem térmica reduziu significativamente a resistência de união ao microcisalhamento.

Cytotoxic effects on human dental pulp stem Cells after exposure to adhesive bonding agents

Abstract Studies regarding cytotoxic effects attributed to the use of adhesive bonding agents on pulp tissue are not conclusive. To point out whether these materials are safe for clinical use, in vivo exposure of dental pulp to adhesive bonding agents was simulated using an experimental setup in which Human Dental Pulp Stem Cells (hDPSC) are exposed to the action of two kinds of adhesives: self-etching adhesives and two-step bonding agents through a dentine barrier. Cytotoxic effects on these cells were evaluated by MTT assay protocol and fluorescence microscopy, and their results were contrasted to those obtained through Raman spectra taken on single hDPSCs. Overall, no significant cytotoxic effects were observed by combining all the techniques, and cell viability close to 90% was achieved for a dentine barrier of at least 1 mm thick. Moreover, Raman spectroscopy was able to detect structural DNA damage in some dental pulp cells when exposed to two-step bonding agents, suggesting that this technique could be considered a complementary tool with the potential to evaluate cell toxicity beyond cell viability.

Resumo Os estudos sobre os efeitos citotóxicos atribuídos ao uso de agentes de união adesivo no tecido pulpar não são conclusivos. Para determinar se esses materiais são seguros para uso clínico, a exposição in vivo da polpa dentária a agentes de união adesiva foi simulada por meio de uma configuração experimental na qual as células-tronco da polpa dentária humana (hDPSC) são expostas à ação de dois tipos de adesivos: adesivos autocondicionantes e agentes de união de duas etapas por meio de uma barreira de dentina. Os efeitos citotóxicos nessas células foram avaliados pelo protocolo de ensaio MTT e microscopia de fluorescência, e seus resultados foram contrastados com os obtidos por meio de espectros Raman obtidos em hDPSCs individuais. De modo geral, não foram observados efeitos citotóxicos significativos com a combinação de todas as técnicas, e a viabilidade celular próxima a 90% foi obtida para uma barreira de dentina de pelo menos 1 mm de espessura. Além disso, a espectroscopia Raman foi capaz de detectar danos estruturais ao DNA em algumas células da polpa dentária quando expostas a agentes de colagem de duas etapas, sugerindo que essa técnica poderia ser considerada uma ferramenta complementar com potencial para avaliar a toxicidade celular além da viabilidade celular.