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ItemArtigo Photochemical outcomes triggered by gold shell-isolated nanorods on bioinspired nanoarchitectonics for bacterial membranes(Elsevier, 2023-08-19) Almeida Junior, Alexandre Mendes de; Moreira, Lucas Gontijo; Camacho, Sabrina Aléssio; Ferreira, Fabiana; Conceição, Katia da; Tada, Dayane Batista; Aoki, Pedro Henrique Benites; UNIFESPBoosted by the indiscriminate use of antibiotics, multidrug-resistance (MDR) demands new strategies to combat bacterial infections, such as photothermal therapy (PTT) based on plasmonic nanostructures. PTT efficiency relies on photoinduced damage caused to the bacterial machinery, for which nanostructure incorporation into the cell envelope is key. Herein, we shall unveil the binding and photochemical mechanisms of gold shell-isolated nanorods (AuSHINRs) on bioinspired bacterial membranes assembled as Langmuir and Langmuir-Schaefer (LS) monolayers of DOPE, Lysyl-PG, DOPG and CL. AuSHINRs incorporation expanded the isotherms, with stronger effect on the anionic DOPG and CL. Indeed, FTIR of LS films revealed more modifications for DOPG and CL owing to stronger attractive electrostatic interactions between anionic phosphates and the positively charged AuSH INRs, while electrostatic repulsions with the cationic ethanolamine (DOPE) and lysyl (Lysyl-PG) polar groups might have weakened their interactions with AuSHINRs. No statistical difference was observed in the surface area of irradiated DOPE and Lysyl-PG monolayers on AuSHINRs, which is evidence of the restricted nano structures insertion. In contrast, irradiated DOPG monolayer on AuSHINRs decreased 4.0 % in surface area, while irradiated CL monolayer increased 3.7 %. Such results agree with oxidative reactions prompted by ROS gener ated by AuSHINRs photoactivation. The deepest AuSHINRs insertion into DOPG may have favored chain cleavage while hydroperoxidation is the mostly like outcome in CL, where AuSHINRs are surrounding the polar groups. Furthermore, preliminary experiments on Escherichia coli culture demonstrated that the electrostatic interactions with AuSHINRs do not inhibit bacterial growth, but the photoinduced effects are highly toxic, resulting in microbial inactivation.ItemArtigo Unveiling the mechanisms underlying photothermal efficiency of gold shell-isolated nanoparticles (AuSHINs) on ductal mammary carcinoma cells (BT-474)(Elsevier, 2023-07-19) Kobal, Mirella Boaro; Camacho, Sabrina Aléssio; Moreira, Lucas Gontijo; Toledo, Karina Alves; Tada, Dayane Batista; Aoki, Pedro Henrique Benites; UNIFESPGold nanoparticles are valuable photothermal agents owing to their efficient photothermal conversion, photo bleaching resistance, and potential surface functionalization. Herein, we combined bioinspired membranes with in vitro assays to elicit the molecular mechanisms of gold shell-isolated nanoparticles (AuSHINs) on ductal mammary carcinoma cells (BT-474). Langmuir and Langmuir-Schaefer (LS) films were handled to build bio membranes from BT-474 lipid extract. AuSHINs incorporation led to surface pressure-area (π-A) isotherms expansion, increasing membrane flexibility. Fourier-transform infrared spectroscopy (FTIR) of LS multilayers revealed electrostatic AuSHINs interaction with head portions of BT-474 lipid extract, causing lipid chain disorganization. Limited AuSHINs insertion into monolayer contributed to hydroperoxidation of the unsaturated lipids upon irradiation, consistently with the surface area increments of ca. 2.0%. In fact, membrane disruption of irradiated BT-474 cells containing AuSHINs was confirmed by confocal microscopy and LDH leakage, with greater damage at 2.2 × 10^(13) AuSHINs/mL. Furthermore, the decrease in nuclei dimensions indicates cell death through photoinduced damage.ItemArtigo Modulating photochemical reactions in Langmuir monolayers of Escherichia coli lipid extract with the binding mechanisms of eosin decyl ester and toluidine blue-O photosensitizers(2021-05) Moreira, Lucas Gontijo; Almeida Junior, Alexandre Mendes; Nield, Tyler [UNESP]; Camacho, Sabrina Aléssio; Aoki, Pedro Henrique Benites; Universidade Estadual Paulista (Unesp)Photodynamic damage to the cell envelope can inactivate microorganisms and may be applied to combat super-resistance phenomenon, empowered by the indiscriminate use of antibiotics. Efficiency in microbial inactivation is dependent on the incorporation of photosensitizers (PS) into the bacterial membranes to trigger oxidation reactions under illumination. Herein, Langmuir monolayers of Escherichia coli lipid extract were built to determine the binding mechanisms and oxidation outcomes induced by eosin decyl ester (EosDEC) and toluidine blue-O (TBO) PSs. Surface-pressure isotherms of the E. coli monolayers were expanded upon EosDEC and TBO, suggesting incorporation of both PSs. Fourier-transform infrared spectroscopy (FTIR) of Langmuir-Schaefer (LS) films reveled that the EosDEC and TBO binding mechanisms are dominated by electrostatic interactions with the anionic polar groups, with limited penetration into the chains. Light-irradiation reduced the relative area of E. coli monolayer on TBO, indicating an increased loss of material to the subphase owing to the chain cleavage, generated by contact-dependent reactions with excited states of TBO. In contrast, the increased relative area of E. coli monolayers containing EosDEC suggests lipid hydroperoxidation, which is PS contact-independent. Even considering a small chain penetration, the saturated EosDEC may have partitioned towards saturated reach domains, avoiding direct contact with membrane unsaturations.ItemArtigo Chain cleavage of bioinspired bacterial membranes photoinduced by eosin decyl ester(ACS Publications, 2020-07-16) Moreira, Lucas Gontijo; Almeida Junior, Alexandre Mendes; Camacho, Sabrina Alessio [UNESP]; Estevão, Bianca Martins; Oliveira Junior, Osvaldo Novais; Aoki, Pedro Henrique Benites [UNESP]; Universidade Estadual Paulista (Unesp)Photodynamic therapy (PDT) is promising for bacterial inactivation since cellular internalization of photosensitizers (PS) is not crucial for the treatment effectiveness. Photoinduced damage in the lipid envelope may already induce microbial inactivation, which requires PS capable of easily penetrating into the membrane. Herein, we report on the insertion of the PS eosin decyl ester (EosDec) into Langmuir films of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), 1,2-dioleoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (DOPG), and cardiolipin (CLP) used as mimetic systems of bacterial membranes. Surface pressure isotherms and polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS) indicated that the hydrophobic nature of EosDec favored deeper penetration in all the phospholipid monolayers. The incorporation of EosDec led to monolayer expansion, especially in the anionic DOPG and CLP owing to repulsive electrostatic interactions, and induced disorder in the lipid chains. Light irradiation of DOPE, DOPG, and CLP monolayers containing EosDec increased the rate of material loss to the subphase, which is attributed to cleavage of lipid chains triggered by contact-dependent reactions between excited states of EosDec and lipid unsaturations. The latter is key for membrane permeabilization and efficiency in microbial inactivation.ItemArtigo Molecular-level effects on cell membrane models to explain the phototoxicity of gold shell-isolated nanoparticles to cancer cells(Elsevier, 2020-10) Camacho, Sabrina Aléssio; Kobal, Mirella Boaro [UNESP]; Almeida Junior, Alexandre Mendes; Toledo, Karina Alves de [UNESP]; Oliveira Junior, Osvaldo Novais de; Aoki, Pedro Henrique Benites [UNESP]; Universidade Estadual Paulista (Unesp)Metallic nanoparticles are promising agents for photothermal cancer therapy (PTT) owing to their photostability and efficient light-to-heat conversion, but their possible aggregation remains an issue. In this paper, we report on the photoinduced heating of gold shell-isolated nanoparticles (AuSHINs) in in vitro experiments to kill human oropharyngeal (HEp-2) and breast (BT-474 and MCF-7) carcinoma cells, with cell viability reducing below 50 % with 2.2 × 1012 AuSHINs/mL and 6 h of incubation. This toxicity to cancer cells is significantly higher than in previous works with gold nanoparticles. Considering the AuSHINs dimensions we hypothesize that cell uptake is not straightforward, and the mechanism of action involves accumulation on phospholipid membranes as the PTT target for photoinduced heating and subsequent generation of reactive oxygen species (ROS). Using Langmuir monolayers as simplified membrane models, we confirmed that AuSHINs have a larger effect on 1,2-dioleoyl-sn-glycero-3-phospho-l-serine (DOPS), believed to represent cancer cell membranes, than on 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) taken as representative of healthy eukaryotic cells. In particular, data from polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS) revealed an increased conformational order of DOPS tails due to the stronger adsorption of AuSHINs. Furthermore, light irradiation reduced the stability of AuSHINs containing DOPC and DOPS monolayers owing to oxidative reactions triggered by ROS upon photoinduced heating. Compared to DOPC, DOPS lost nearly twice as much material to the subphase, which is consistent with a higher rate of ROS formation in the vicinity of the DOPS monolayer.ItemArtigo The protective effect of Artepillin C against lipid oxidation on simplified model membranes(Elsevier, 2021-02-15) Pazin, Wallance Moreira [UNESP]; Ruiz, Gilia Cristine Marques [UNESP]; Ito, Amando Siuiti; Constantino, Carlos José Leopoldo [UNESP]; Aoki, Pedro Henrique Benites [UNESP]; Santos, Marcelo Jose dos; Universidade Estadual Paulista (Unesp)Brazilian green propolis is a well-known therapeutic product, commonly used in folk medicine. Artepillin C is the major compound of propolis and has received considerable attention owing to its lipophilic affinity and antioxidant activity, enabling the use against lipid oxidation caused by free radicals, which is a first step before degenerative diseases. The protective effect of Artepillin C against lipid oxidation was evaluated here on simplified models of lipid membranes based on Langmuir monolayers and giant unilamellar vesicles (GUVs) formed of DOPC under oxidative stress induced by the photoactivated erythrosin. Our findings show that Artepillin C donates a hydrogen atom of the phenolic hydroxyl group to the lipid radical, avoiding the formation of truncated aldehyde lipids, playing a role in the termination step of lipid peroxidation generating hydroperoxides. The affinity of Artepillin C for lipid structures, together with its antioxidant potential, preclude the lipid peroxidation caused by reactive species.