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Made available in DSpace on 2020-12-12T01:08:56Z (GMT). No. of bitstreams: 0 Previous issue date: 2020-06-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Modifications on shear stress-based mechanical forces are associated with pathophysiological susceptibility and their effect on endothelial cells (EC) needs to be better addressed looking for comprehending the cellular and molecular mechanisms. This prompted us to better evaluate the effects of shear stress in human primary venous EC obtained from the umbilical cord, using an in vitro model to mimic the laminar blood flow, reaching an intensity 1–4 Pa. First, our data shows there is a significant up-expression of phosphatidylinositol 3-kinase (PI3K) in shear-stressed cells culminating downstream with an up-phosphorylation of AKT and up-expression of MAPK-ERK, concomitant to a dynamic cytoskeleton rearrangement upon integrin subunits (α4 and ß 3) requirements. Importantly, the results show there is significant involvement of nitric oxide synthase (eNOS), nNOS, and vascular endothelial growth factors receptor 2 (VEGFR2) in shear-stressed EC, while cell cycle-related events seem to being changed. Additionally, although diminution of 5-hydroxymethylcytosine in shear-stressed EC, suggesting a global repression of genes transcription, the promoters of PI3K and eNOS genes were significantly hydroxymethylated corroborating with their respective transcriptional profiles. Finally, to better address, the pivotal role of PI3K in shear-stressed EC we have revisited these biological issues by wortmannin targeting PI3K signaling and the data shows a dependency of PI3K signaling in controlling the expression of VGFR1, VGFR2, VEGF, and eNOS, once these genes were significantly suppressed in the presence of the inhibitor, as well as transcripts from Ki67 and CDK2 genes. Finally, our data still shows a coupling between PI3K and the epigenetic landscape of shear-stressed cells, once wortmannin promotes a significant suppression of ten-11 translocation 1 (TET1), TET2, and TET3 genes, evidencing that PI3K signaling is a necessary upstream pathway to modulate TET-related genes. In this study we determined the major mechanotransduction pathway by which blood flow driven shear stress activates PI3K which plays a pivotal role on guaranteeing endothelial cell phenotype and vascular homeostasis, opening novel perspectives to understand the molecular basis of pathophysiological disorders related with the vascular system. Bioassays and Cell Dynamics Laboratory Department of Chemistry and Biochemistry Bioscience Institute UNESP Department of Biology Dental School University of Taubaté Bioassays and Cell Dynamics Laboratory Department of Chemistry and Biochemistry Bioscience Institute UNESP FAPESP: 2014/22689-3 FAPESP: 2016/22270-8

Made available in DSpace on 2021-06-25T10:20:57Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-01-01 Although osseointegration and clinical success of titanium (Ti)-implanted materials depend on neovascularization in the reactional peri-implant tissue, very little has been achieved considering the Ti-molecules release on the behavior of endothelial cells. To address this issue, we challenged endothelial cells (HUVECs) with Ti-enriched medium obtained from two types of commercial titanium surfaces [presenting or not dual-acid etching (DAE)] up to 72 h to allow molecular machinery analysis. Our data show that the Ti-enriched medium provokes significant stimulus of angiogenesis-related machinery in endothelial cells by upexpressing VEGFR1, VEGFR2, VEGF, eNOS, and iNOS genes, while the PI3K/Akt signaling pathway was also significantly enhanced. As PI3K/AKT signaling was related to angiogenesis in response to vascular endothelial growth factor (VEGF), we addressed the importance of PI3K/Akt upon Ti-enriched medium responses by concomitantly treating the cells with wortmannin, a well-known PI3K inhibitor. Wortmannin suppressed the angiogenic factors, because VEGF, VEGFR1, and eNOS genes were downregulated in those cells, highlighting the importance of PI3K/AKT signaling on driving angiogenic phenotype and angiogenesis performance within the peri-implant tissue reaction. In conjunction, these data reinforce that titanium-implantable devices modify the metabolism of surrounding cells, such as endothelial cells, probably coupling osteogenesis and angiogenesis processes in peri-implant tissue and then contributing to successfully osseointegration of biomedical titanium-based devices. [Figure not available: see fulltext.]. Institute of Biosciences of Botucatu Department of Chemical and Biological Sciences UNESP – São Paulo State University Institute of Biosciences of Botucatu Department of Chemical and Biological Sciences UNESP – São Paulo State University

Made available in DSpace on 2019-10-06T15:57:24Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-12-15 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) We therefore investigated whether there is synergism between triiodothyronine (T3) hormone and trophic molecules released from mechanically-stressed endothelial cells (EC-enriched medium) in osteogenic phenotype by mapping classical repertory of genes. Although there are studies reporting the efficiency of T3 hormone on bone cells, it is scarce considering their effect in conjunction with other physiologically active molecules, such as those released by the active endothelial cells. To address this issue, human bone marrow-derived mesenchymal stem cells (hBMSCs) were treated with EC-enriched medium subjected to shear-stress up to 72 h in vitro, in conjunction or not with T3 hormone. Although our results found an important synergism considering these parameters on modulating key bone-related gene markers, such as on the alkaline phosphatase (ALP) behavior (at both mRNA and protein content), contributing for osteoblast differentiation, important genes such as OSTERIX and RUNX2 were significantly down-expressed, while a over-expression of RANKL was found when the conjunction effect of T3 and endothelial paracrine signaling was considered. In addition, T3 hormone over expressed both OCT4 and NANOG genes in a DNA epigenetic-independent manner. However, we observed a dynamic reprogramming of DNMT1, DNMT3A, DNMT3B and TET1, important DNA-related epigenetic markers. Specifically, T3 hormone alone up-modulated TET2 transcripts profile. Complimentarily, expression of microRNA (miRs) processing-related genes also was modulated, as well as miR-10b, miR-22, miR-21, miR-143 and miR-145 transcriptional related profiles. Altogether, our results suggested a positive effect of mechanically-stressed endothelial cells-induced paracrine signaling on T3 hormone-obtaining osteogenic phenotype, contributing to understanding the paradoxal effect of T3 hormone on the bone physiology. Department of Chemistry and Biochemistry São Paulo State University (UNESP) Institute of Biosciences, Campus Botucatu Electron Microscopy Center São Paulo State University (UNESP) Institute of Biosciences, Campus Botucatu Área de Periodontia Departamento de Prótese e Periodontia Faculdade de Odontologia de Piracicaba Universidade de Campinas Área de Epigenética Faculdade de Odontologia Universidade Paulista Department of Chemistry and Biochemistry São Paulo State University (UNESP) Institute of Biosciences, Campus Botucatu Electron Microscopy Center São Paulo State University (UNESP) Institute of Biosciences, Campus Botucatu FAPESP: 2014/22689-3 FAPESP: 2016/01139-0 FAPESP: 2016/10392-1

Shear stress changes are associated with a repertory of signaling cascade modulating vascular phenotype. As shear stress-related tensional forces might be associated with pathophysiological susceptibility, a more comprehensive molecular map needs to be addressed. Thus, we subjected human umbilical vein endothelial cells (HUVECs) to a circuit of different tensional forces in vitro considering the following three groups: (a) physiological blood flow shear stress condition (named Normo), (b) a hypertensive blood flow shear stress (named Hyper), and (c) these hyper-stressed cells were returned to Normo condition (named Return). The samples were properly collected to allow different methodologies analysis. Our data showed a pivotal involvement of c-Src on driving the mechanotransduction cascade by modulating signaling related with adhesion, survival (PI3K/Akt) and proliferative phenotype. Moreover, c-Src seems to develop important role during extracellular matrix remodeling. Additionally, proteomic analysis showed strong involvement of heat shock protein 70 (HSP70) in the hypertensive-stressed cells; it being significantly decreased in return phenotype. This result prompted us to investigate 20S proteasome as an intracellular proteolytic alternative route to promote the turnover of those proteins. Surprisingly, our data reveled significant overexpression of sets of proteasome subunit α-type (PSMA) and β-type (PSMB) genes. In conjunction, our data showed c-Src as a pivotal protein to drive mechanotransduction in endothelial cells in a HSP70-dependent turnover scenario. Because shear patterns is associated with pathophysiological changes, such as atherosclerosis and hypertension, these results paved new road to understand the molecular mechanism on driving mechanotransduction in endothelial cells and, if drugable, these targets must be considered within pharmacological treatment optimization.

The aim of this study was to show that abundant and inexpensive plant oils can be biotransformed to increase biological activity (antioxidant, antimicrobial and cytotoxicity) through hydrolysis reaction catalysed by lipases. We tested homemade and commercial lipases through the biotransformation of nine different plant oils in forty different combinations. First, the chemical composition of the samples was investigated. Thereafter, biological tests were conducted to assess the antioxidant and antimicrobial activity of the sampled biotransformation products, as well as analyzed their influence on the viability of healthy and cancer cells. Summarising, sunflower, corn and olive oils modified by orange waste‐obtained lipases presented the most promising results, reaching up to 90% of antioxidant activity increase and significant growth inhibition of bacteria colonies belonging to genera Escherichia, Listeria, Staphylococcus, Pseudomonas and Salmonella. In addition, those compounds affected human oral squamous carcinoma cells. The bioconversion of plant oils through lipases improves their biological properties and might be an option for biotechnological application.

Made available in DSpace on 2018-12-11T17:36:31Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-06-15 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) In this study, the effect of molybdenum content in Ti-15Zr-based alloys (wt%) was analyzed in terms of crystalline structure, microstructure, selected mechanical properties, and cytotoxicity. The samples were produced by argon arc-melting followed by hot rolling and heat treatment processes. The crystalline structure and microstructure were dependent of both alloying elements (zirconium and molybdenum). Ti-15Zr alloy displayed only laths of α′ phase, while the alloys up to Ti-15Zr-10Mo exhibited different proportions of α′ α” and β phases. Molybdenum content higher than 12.5 wt% fully stabilized the β phase. Vickers microhardness values of Ti-15Zr-Mo alloys were higher than those of CP-Ti due to solid solution and phase precipitation strengthening. Young's modulus values of Ti-15Zr-Mo alloys were lower than those of CP-Ti due to β phase stabilization. Cytotoxicity levels of Ti-15Zr-Mo alloys were within a tolerable range for biomedical purposes. In addition, we observed molybdenum content in Ti-15Zr-based alloys promoted an increase on pre-osteoblast adhesion up to 3 h of adhesion's time. Thus, Ti-15Zr-15Mo alloy presented better combination of properties than some traditional metallic biomaterials. IFSP – Federal Institute of Education Science and Technology UNESP – Univ Estadual Paulista Laboratório de Anelasticidade e Biomateriais USP - Universidade de São Paulo Faculdade de Odontologia Departamento de Ciências Biológicas UNESP - Univ Estadual Paulista Departamento de Química e Bioquímica IBTN-Br - Institute of Biomaterials Tribocorrosion and Nanomedicine – Brazilian Branch UNESP – Univ Estadual Paulista Laboratório de Anelasticidade e Biomateriais UNESP - Univ Estadual Paulista Departamento de Química e Bioquímica FAPESP: #2010/20440-7 FAPESP: #2012/22742-6 FAPESP: #2014/06.221-1 FAPESP: #2014/22689-3 FAPESP: #2015/00851-6 CNPq: #307.279/2013-8 CNPq: #481.313/2012-5

Background Biofunctionalization of titanium surfaces can improve host responses, especially considering the time for osteointegration and patient recovery. This prompted us to modify titanium surfaces with alendronate and albumin and to investigate the behavior of osteoblasts on these surfaces. Methods The biofunctionalization of titanium surfaces was characterized using classical physicochemical approaches and later used to challenge pre-osteoblast cells up to 24 h. Then their viability and molecular behavior were investigated using mitochondrial dehydrogenase activity and RTq-PCR technologies, respectively. Potential stimulus of extracellular remodeling was also investigated by zymography. Results Our data indicates a differential behavior of cells responding to the surfaces, considering the activity of mitochondrial dehydrogenases. Molecularly, the differential expression of genes related with cell adhesion highlighted the importance of Integrin-β1, Fak, and Src. These 3 genes were significantly decreased in response to titanium surfaces modified with alendronate, but this behavior was reverted when alendronate was associated with albumin. Alendronate-modified surfaces promoted a significant increase on ECM remodeling, as well as culminating with greater gene activity related to the osteogenic phenotype (Runx2, Alp, Bsp). Conclusion Altogether, our study found interesting osteogenic behavior of cells in response to alendronate and albumin surfaces, which indicates the need for in vivo analyses to better consider these surfaces before clinical trials within the biomedical field., Materials science; Cell biology; Implants; Biocompatibility; Trace elements; Osseointegration; Bisphosphonate; Albumin.

Made available in DSpace on 2021-06-25T10:36:45Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-04-16 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Shrimp trawling is an important socio-economic activity; however, the bycatch can be problematic to the environment. Thus, the present study investigated potential uses of the bycatch to generate value-added products. The biological activity of the protein hydrolysates obtained from the two most abundant fish species (Micropogonias furnieri and Paralonchurus brasiliensis) was evaluated. Muscle and skin samples of both species were hydrolyzed using two enzymes, Alcalase 2.4 L® or Protamex®. The in vitro antioxidant capacity against peroxyl radicals, DPPH, and sulfhydryl groups were analyzed. Cell viability, Western Blotting, Zymogram, and Real-time PCR analyses were performed. The results showed that the hydrolysates have antioxidant activity and no effect on cell viability at doses lower than 16 mg/mL. In addition, they can modulate extracellular remodelling and intracellular pathways related to cell adhesion. Thus, the hydrolysis of the fish bycatch allows the release of bioactive peptides with potential use in the food industry. Aquaculture Center São Paulo State University (UNESP), Campus Jaboticabal Marine Station of Aquaculture Aquaculture postgraduate Oceanography Institute Federal University of Rio Grande (FURG) Rio Grande Institute of Biological Sciences (ICB) Federal University of Rio Grande (FURG) Department of Chemistry and Biochemistry Bioscience Institute São Paulo State University (UNESP), Campus Botucatu Aquaculture Center São Paulo State University (UNESP), Campus Jaboticabal Department of Chemistry and Biochemistry Bioscience Institute São Paulo State University (UNESP), Campus Botucatu

Made available in DSpace on 2018-12-11T17:12:50Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-10-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) The reciprocal and adaptive interactions between cells and substrates governing morphological transitions in the osteoblast compartment remain largely obscure. Here we show that osteoblast cultured in basement membrane matrix (Matrigel™) exhibits significant morphological changes after ten days of culture, and we decided to exploit this situation to investigate the molecular mechanisms responsible for guiding osteoblast morphological transitions. As almost all aspects of cellular physiology are under control of kinases, we generated more or less comprehensive cellular kinome profiles employing PepChip peptide arrays that contain over 1000 consensus substrates of kinase peptide. The results obtained were used to construct interactomes, and these revealed an important role for FoxO in mediating morphological changes of osteoblast, which was validated by Western blot technology when FoxO was significantly up-expressed in response to Matrigel™. As FoxO is a critical protein in canonical hedgehog signalling, we decided to explore the possible involvement of hedgehog signalling during osteoblast morphological changes. It appeared that osteoblast culture in Matrigel™ stimulates release of a substantial amounts Shh while concomitantly inducing upregulation of the expression of the bona fide hedgehog target genes Gli-1 and Patched. Functional confirmation of the relevance of these results for osteoblast morphological transitions came from experiments in which Shh hedgehog signalling was inhibited using the well-established pathway inhibitor cyclopamine (Cyc). In the presence of Cyc, culture of osteoblasts in Matrigel™ is not capable of inducing morphological changes but appears to provoke a proliferative response as evident from the upregulation of Cyclin D3 and cdk4. The most straightforward interpretation of our results is that hedgehog signalling is both necessary and sufficient for membrane matrix-based morphological transitions. Lab. de Bioensaios e Dinâmica Celular Depto de Química e Bioquímica Instituto de Biociências Universidade Estadual Paulista - UNESP, campus Botucatu Laboratory of Bioassays and Signal Transduction Departamento de Bioquímica Instituto de Biologia Universidade Estadual de Campinas (Unicamp), 6109 Instituto Nacional de Metrologia Normalização e Qualidade Industrial (INMETRO) Life Sciences Applied Metrology (Dimav)/Bioengineering Group Erasmus MC Cancer Institute Erasmus MC Erasmus University of Rotterdam Lab. de Bioensaios e Dinâmica Celular Depto de Química e Bioquímica Instituto de Biociências Universidade Estadual Paulista - UNESP, campus Botucatu FAPESP: 2014/22689-3 CNPq: 301966/2015-0 CNPq: 477452/2012-4

Made available in DSpace on 2019-10-06T16:06:56Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-02-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Although layered double hydroxides (LDH) have been listed as promising nanomaterials in human healthcare, very little has been achieved on osteoblast inflammatory signaling. Thus, osteoblasts were challenged with two LDHs (Mg 2 Al-Cl and Zn 2 Al-Cl, at 0.002 mg/mL) up to 24 h, establishing an acute inflammatory mechanism, as well as identifying whether Sonic hedgehog (Shh) signaling has an influence. Functional experiments were performed by previously treating (2 h) semiconfluent osteoblast cultures with cyclopamine molecule (cyc), a widely used Shh inhibitor. Considering inflammasome complex, the asc1 gene was significantly up-expressed in response to Zn 2 Al-Cl - LDHs, as well as the nrlp3 gene. By treating the osteoblast with cyc, the asc1 gene presented an even higher profile. Our results found a down-modulation of major pro-inflammatory cytokines-related genes, when tnfα and il1ß were significantly down-modulated in response to LDHs. Conversely, anti-inflammatory cytokines were up-modulated considering the same experimental procedures. Except the il6, the other il13, il10, and tgfß genes were up modulated. Additionally, Shh signaling seems to modulate this repertory as both the il13 and il10 genes were significantly up-modulated when the Shh signaling was inhibited. Altogether, our results reveal for the first time the exigency of Shh-dependent anti-inflammatory signals in LDH-induced osteoblast responses. Lab. de Bioensaios e Dinâmica Celular Departamento de Química e Bioquímica Instituto de Biociências Universidade Estadual Paulista - UNESP campus Botucatu Lab. de Engenharia Celular Hemocentro Hospital das Clínicas Faculdade de Medicina de Botucatu Departamento de Cirurgia Universidade Federal de São Paulo-UNIFESP, Rua Botucatu 740 Departamento de Química Fundamental Instituto de Química Universidade de São Paulo-USP, Av. Prof. Lineu Prestes 748 Lab. de Bioensaios e Dinâmica Celular Departamento de Química e Bioquímica Instituto de Biociências Universidade Estadual Paulista - UNESP campus Botucatu Lab. de Engenharia Celular Hemocentro Hospital das Clínicas Faculdade de Medicina de Botucatu FAPESP: 2011/50318-1 FAPESP: 2014/22689-3

Made available in DSpace on 2018-12-11T17:21:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-12-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Titanium is widely used for biomedical applications, but little information is being delivered regarding the cellular/molecular mechanisms explaining their efficacy, mainly considering the effects of the Ti-released trace elements on pre-osteoblasts. We addressed this issue by investigating decisive intracellular signal transduction able to modulate cytoskeleton rearrangement, proliferative phenotype and extracellular matrix (ECM) remodeling. We considered titanium grades IV and V, submitted or not to dual acid-etching (w/DAE or wo/DAE, respectively). Our results showed there is no cytotoxicity, preserving AKT involvement. Additionally, Ti-enriched medium promoted a diminution of the downstream signaling upon integrin activation (phosphorylating Rac1 and cofilin), guaranteeing a dynamic cytoskeleton rearrangement. Moreover, the low profile of ECM remodeling obtained in response to trace molecules released by Ti-based devices seems contributing to the osteoblast performance in mediating extracellular support to cell anchorage. This hypothesis was validated by the up-expression of ß1-integrin, src and Focal adhesion kinase (fak) genes, mainly in response to titanium grade V. Proliferative phenotype showed an unbalance between cyclin-dependent kinases (CDKs) and p15INK4b/p21Cip1. In conjunction, we showed for the first time that trace elements from Ti-based biomedical devices provoke important modulation of the osteoblast biology, driving cell anchoring, viability, and proliferative phenotype. Certainly, these biological outcomes compromise implant osseointegration. Dept. of Chemistry and Biochemistry Bioscience Institute São Paulo State University UNESP Campus Botucatu Electron Microscopy Center IBB UNESP Dept. of Chemistry and Biochemistry Bioscience Institute São Paulo State University UNESP Campus Botucatu Electron Microscopy Center IBB UNESP CNPq: # 477452/2012-4 FAPESP: #2014/22689-3 CNPq: #301966/2015-0

Made available in DSpace on 2019-10-06T16:27:49Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-07-01 Angiogenesis is a relevant mechanism to be considered for the success of bone healing, even considering endosseous implantable devices, providing adequate delivery of substances necessaries for the cell viability and bone de novo deposition. Within of the repertory of metal-based implantable alloys, cobalt-chromium (CoCr) has emerged with very interesting properties for biomedical applications. Additionally, we have shown that released molecules from implants devices are able to modulate cells away and because that we hypothesized these released molecules might act on endothelial cells. In order to better address this issue, we investigated the effect of Co-Cr-enriched medium on endothelial cells (HUVECs), considering a biological model subjecting those cells to shear-stress to partially mimic the physiological environment and further allow investigating intracellular pathways responsible to drive cytoskeletal rearrangement, cell viability and extracellular matrix (ECM) remodeling processes. Considering the analysis of the metalloproteinases (MMPs) activities, our data indicates an intense ECM remodeling in response to CoCr-enriched medium suggesting some role on angiogenesis once ECM remodeling is prerequisite to cell growth. This was better addressed by revealing its involvement on modifying both mRNA expression and protein levels of members of the MAPK family. Additionally, the expression of CDK4 gene was modulated within the cell response to Co-Cr-enriched medium, while the modulation in the expression of P15 and P21 indicates an important regulatory mechanism required. Overall, our results demonstrate that trace of CoCr elements triggers decisive intracellular signaling in shear-stressed endothelial cells, suggesting influence on angiogenesis-related mechanism and they bring novel insights to explain the biological activity of CoCr as it has been emerged as interesting biomedical materials within the medical and dentistry fields. Department of Chemistry and Biochemistry Bioscience Institute Sao Paulo State University UNESP, Campus Botucatu, Botucatu Electron Microscopy Center IBB UNESP Department of Chemistry and Biochemistry Bioscience Institute Sao Paulo State University UNESP, Campus Botucatu, Botucatu Electron Microscopy Center IBB UNESP

Lipases are an economic important group of biocatalysts that can be produced by some fungal under solid-state fermentation. Orange wastes are source of lipases and potential substrates for lipases production. This work assessed 19 fugal strains cultivated in Citrus sinensis cv. Hamlin orange wastes (peel, frit and core) for production of lipases in order to generate compounds with antioxidant, antimicrobial and cytotoxic properties. Fifteen of those fungi grew and produced lipases, mainly the Aspergillus brasiliensis [National Institute of Quality Control (INCQS) 40036]/frit system, which showed 99.58 U/g total lipase. The substrate with the highest production of lipase was frit with 26.67 and 78.91 U/g of total lipases produced on average by the 15 microorganisms. Aspergillus niger 01/frit (33.53 U/g) and Aspergillus niger (INCQS 40015)/frit (34.76 U/g) systems showed the highest specificity values in all the herein tested synthetic substrates with 4, 12 and 16 carbons. Analysis of the fatty acid profile of hydrolysis products obtained in the most prominent systems applied to corn and sunflower oils showed: palmitic acid, linoleic acid, oleic acid, and stearic acid. These acids showed antioxidant capacity of up to 58% DPPH (2,2-diphenyl-1-pierylhydrazyl) radical reduction and antibacterial activity against Escherichia coli, Listeria monocytogenes, Pseudomonas aureginosa, Salmonella Enteritidis and Staphylococcus aureus, as well as cytotoxicity to SCC9 cells (squamous cancer cells).

Made available in DSpace on 2018-12-11T17:21:58Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-10-01 Background: The placenta is a multifunctional organ that can suffer with imbalances between pro- and antioxidant molecules, contributing for inflammatory imbalance. The inflammation generated by oxidative stress may induce inflammasome activation, an essential complex for pro-inflammatory cytokine production. Objective: The aim of this study was to evaluate whether hydrogen peroxide (H2O2) mediated oxidative stress induces inflammasome activation on placental explants. Study design: Tissue cultures of placental explants obtained from normotensive pregnant women were performed in different concentrations of H2O2. Gene expressions of NLRP3, caspase-1, IL-1β TNF-α and IL-10 were evaluated by qPCR. Superoxide dismutase (SOD), catalase, Heat shock protein 70 (Hsp70), Caspase-1, TNF-α IL-1β IL-10 and human Chorionic Gonadotropin (hCG) were determined by ELISA. Results: Concentrations of catalase, Hsp70, hCG and SOD were higher in cultures with 100 and 1000 µM H2O2 compared to controls. Gene and protein expressions of TNF-α and IL-1β were elevated in cultures with 1000 μM H2O2 compared to controls. This concentration led to inflammasome activation, by increasing gene expressions of NLRP3, caspase-1 and IL-1β. In contrast, gene and protein expressions of IL-10 were reduced at 100 and 1000 μM H2O2. Protein expression of caspase-1 was higher in cultures of 100 μM H2O2 compared to controls. Treatment with Glybenclamide at 200 μM was used to prevent NLRP3 inflammasome activation. This concentration reduced protein expression of caspase-1 compared to culture with only H2O2 and control cultures. Conclusions: Our results confirm that H2O2 induces oxidative stress on placental explants and demonstrate that cell responses to this stress involve inflammasome activation. Department of Gynaecology and Obstetrics Medical School Botucatu Sao Paulo State University (Unesp) Department of Microbiology and Immunology Institute of Biosciences Botucatu Sao Paulo State University (Unesp) Department of Gynaecology and Obstetrics Medical School Botucatu Sao Paulo State University (Unesp) Department of Microbiology and Immunology Institute of Biosciences Botucatu Sao Paulo State University (Unesp)

Made available in DSpace on 2018-12-11T17:19:11Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-03-01 Over the last several years, we have focused on the importance of intracellular signaling pathways in dynamically governing the biointerface between pre-osteoblast and surface of biomaterial. Thus, this study investigates the molecular hallmarks involved in the pre-osteoblast relationship with different topography considering Machined (Mc), Dual Acid-Etching (DAE), and nano hydroxyapatite-blasted (nHA) groups. There was substantial differences in topography of titanium surface, considering Atomic Force Microscopy and water contact angle (Mc = 81.41 ± 0.01; DAE = 97.18 ± 0.01; nHA = 40.95 ± 0.02). Later, to investigate their topography differences on biological responses, pre-osteoblast was seeded on the different surfaces and biological samples were collected after 24 h (to consider adhesion signaling) and 10 days (to consider differentiation signaling). Preliminary results evidenced significant differences in morphological changes of pre-osteoblasts mainly resulting from the interaction with the DAE and nHA, distinguishing cellular adaptation. These results pushed us to analyze activation of specific genes by exploring qPCR technology. In sequence, we showed that Src performs crucial roles during cell adhesion and later differentiation of the pre-osteoblast in relationship with titanium-based biomaterials, as our results confirmed strong feedback of the Src activity on the integrin-based pathway, because integrin-ß1 (∼5-fold changes), FAK (∼12-fold changes), and Src (∼3.5-fold changes) were significantly up-expressed when Src was chemically inhibited by PP1 (5 μM). Moreover, ECM-related genes were rigorously reprogrammed in response to the different surfaces, resulting on Matrix Metalloproteinase (MMP) activities concomitant to a significant decrease of MMP inhibitors. In parallel, we showed PP1-based Src inhibition promotes a significant increase of MMP activity. Taking all our results into account, we showed for the first time nano hydroxyapatite-blasted titanium surface creates a biointerface able to govern Src-dependent osteoblast metabolism as pre-requisite to ECM remodeling Department of Chemistry and Biochemistry Bioscience Institute São Paulo State University UNESP Campus Botucatu Electron Microscopy Center IBB UNESP Department of Chemistry and Biochemistry Bioscience Institute São Paulo State University UNESP Campus Botucatu Electron Microscopy Center IBB UNESP

Made available in DSpace on 2018-12-11T17:34:38Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-10-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Cooking oil waste leads to well-known environmental impacts and its bioremediation by lipase-based enzymatic activity can minimize the high cytotoxic potential. In addition, they are among the biocatalysts most commercialized worldwide due to the versatility of reactions and substrates. However, although lipases are able to process cooking oil wastes, the products generated from this process do not necessarily become less toxic. Thus, the aim of the current study is to analyze the bioremediation of lipase-catalyzed cooking oil wastes, as well as their effect on the cytotoxicity of both the oil and its waste before and after enzymatic treatment. Thus, assessed the post-frying modification in soybean oil and in its waste, which was caused by hydrolysis reaction catalyzed by commercial and home-made lipases. The presence of lipases in the extracts obtained from orange wastes was identified by zymography. The profile of the fatty acid esters formed after these reactions was detected and quantified through gas chromatography and fatty acids profile compared through multivariate statistical analyses. Finally, the soybean oil and its waste, with and without enzymatic treatment, were assessed for toxicity in cytotoxicity assays conducted in vitro using fibroblast cell culture. The soybean oil wastes treated with core and frit lipases through transesterification reaction were less toxic than the untreated oils, thus confirming that cooking oil wastes can be bioremediated using orange lipases. Chemistry and Biochemistry Department Institute of Biosciences São Paulo State University (UNESP) Agronomic Institute (IAC) CEP Chemistry and Biochemistry Department Institute of Biosciences São Paulo State University (UNESP) FAPESP: 2014/ 22689-3 FAPESP: 2014/10962-7 FAPESP: 2015/01753-8