Leticia Labriola - Academia.edu (original) (raw)
Papers by Leticia Labriola
Photochemical & Photobiological Sciences
Pancreatic ductal adenocarcinomas (PDAC) are the fourth leading cause of death due to neoplasms. ... more Pancreatic ductal adenocarcinomas (PDAC) are the fourth leading cause of death due to neoplasms. In view of the urgent need of effective treatments for PDAC, photodynamic therapy (PDT) appears as a promising alternative. However, its efficacy against PDAC and the mechanisms involved in cell death induction remain unclear. In this study, we set out to evaluate PDT's cytotoxicity using methylene blue (MB) as a photosensitizer (PS) (MB-PDT) and to evaluate the contribution of necroptosis in its effect in human PDAC cells. Our results demonstrated that MB-PDT induced significant death of different human PDAC models presenting two different susceptibility profiles. This effect was independent of MB uptake or its subcellular localization. We found that the ability of triggering necroptosis was determinant to increase the treatment efficiency. Analysis of single cell RNA-seq data from normal and neoplastic human pancreatic tissues showed that specific necroptosis proteins RIPK1, RIPK3 and MLKL presented significant higher expression levels in cells displaying a transformed phenotype providing further support to the use of approaches that activate necroptosis, like MB-PDT, as useful adjunct to surgery of PDAC to tackle the problem of microscopic residual disease as well as to minimize the chance of local and metastatic recurrence.
Research article Generation and characterization of human insulin-releasing cell lines
Photodiagnosis and Photodynamic Therapy, 2015
s / Photodiagnosis and Photodynamic Therapy 12 (2015) 325–375 349 treatment outcome as an attempt... more s / Photodiagnosis and Photodynamic Therapy 12 (2015) 325–375 349 treatment outcome as an attempt to predict the PDT efficiency. The clinical study is being conducted at Hospital Amaral Carvalho in Jahu, and 100 lesions of the basal cell carcinoma types treated with PDT using Aminolevulinic Acid with photosensitizer. The thermal changes of the lesions are registeredwith a thermal imager (Fluke® FLK-Ti400) during four phases: lesion initial, lesion curetted, after the drug incubation and after the illumination in the two sessions. The temperature variation was compared with the result of the tissue biopsies after 30 days. Significant results are being collect in order to better understand the PDT response. http://dx.doi.org/10.1016/j.pdpdt.2015.07.095 Unveiling the molecular mechanisms involved in the cytotoxicity induced by photodynamic therapy in human breast cancer cells A.F. dos Santos, L.F. Terra, R.A.M. Wailemann, T.C. Oliveira, M.S. Baptista, L. Labriola Biochemistry Department, Chemistry Institute, University of Sao Paulo (USP), Sao Paulo, Brazil The efficacy of photodynamic therapy (PDT) on mammary tumors as well as the mechanisms of cell death remains unclear. We evaluated the cytotoxicity of PDT by using methylene blue (MB-PDT) in two (2D) and three (3D) dimension cultures of human breast tumor cells and the death pathways involved. Invasive, non-invasive and non-tumorigenic cells were incubated with different MB concentrations and irradiated ( =640nm) at 4.5 J/cm2. Fluorescence microscopy was used to evaluate cell viability and death. Autophagy was evaluated also by Western blotting. The role of autophagy was investigated using chloroquine (CQ) and LY294002 (LY) as inhibitors and rapamycin (RAPA) as activator. MB-PDT increased cell death of both tumoral cell lines in 2D or 3D culture, but was significantly lower in non-tumorigenic cells despite the higher intracellular concentration ofMB. No dark effect and no morphological signs of apoptotic were observed, but there were autophagy increase. After cotreatmentwith CQor LY the cytotoxicity was increased and with RAPA it was lowered. We showed that MB-PDT induced selective cell death in a model that recapitulates the morphology of glandular epithelium, and that autophagy is mainly related with cytoprotection. Our observations indicate that PDT is an effective therapy displaying minimal side effects. http://dx.doi.org/10.1016/j.pdpdt.2015.07.096 Comparison of the response and mechanisms of MAL-PDT of different squamous carcinoma cell lines Daniela Leon1, Ramon Silva1, Natalia Inada2, Cristina Kurachi2, Aleida Vivallo1, Priscilla Brebi1, Juan Carlos Roa3 1 Departamento de Anatomia Patologica, Laboratorio de Patologia Molecular BIOREN-CEGIN, Universidad de La Frontera, Chile 2 Sao Carlos Institute of Physics, University of Sao Paulo, Brazil 3 Departamento de Patologia, Facultad de Medicina, Pontificia Universidad Catolica de Chile, Chile Introduction: Non-melanoma skin cancer (NMSC) is the most common neoplasia worldwide and can be treated with Photodynamic Therapy (PDT). NMSC has good outcome after PDT, however, recurrent lesions has been reported. The aim of this study was to compare the effect of PDT on two non-melanoma skin cancer cell lines. Methodology: A keratinocytes cell line and two squamous carcinoma cell lines, A-431 and HSC-1, were used. PDT was carried outusingmethyl aminolevulinate (MAL) as aprecursor of Protoporphyrin IX. All cell lineswere incubatedwith distinct concentrations ofMAL and irradiatedwith different red light fluences. Cell viability wasevaluated24hafter PDTbyMTT, TrypanBluedyeexclusionand flow cytometry assays. In addition, ROS production was measured by flow cytometry. Results: A-431 cells were more resistant to PDT cytotoxic effect than HSC-1 cells compared the same condition of treatment. Meanwhile, keratinocyteswere themost sensitive to PDT. ROS production after PDT varied among the skin cell type. Conclusion: Cells from squamous skin cancer have a different response to the same PDT conditions.Wewill investigate the resistant phenotype of those cells for improving the effect of PDT on them. http://dx.doi.org/10.1016/j.pdpdt.2015.07.097 Penetration depth of the 664-nm semiconductor laser light with talaporfin sodium into human brain tissue with glioma Soko Ikuta1, Yuki Kawase2, Yoshihiro Muragaki1,3, Takashi Maruyama1,3, Masayuki Nitta3, Taiichi Saito3, Hiroshi Iseki1 1 Institute of Advanced Biomedical Engineering & Science, Tokyo Women’s Medical University, Japan 2 Panasonic Healthcare Co., Ltd., Japan 3 Department of Neurosurgery, Tokyo Women’s Medical University, Japan Glioma cell has characteristicswhich invasively spread into normal brain tissue. Talaporfin-PDT is assumed to be a promising to affect infiltrating glioma cells where the extirpation cavity, but there is no data showed about its optical penetration depth and cellular cytotoxicity in human brain tissue. This study evaluated these affections with…
Brazilian Journal of Medical and Biological Research, 2019
The scientific publication landscape is changing quickly, with an enormous increase in options an... more The scientific publication landscape is changing quickly, with an enormous increase in options and models. Articles can be published in a complex variety of journals that differ in their presentation format (online-only or in-print), editorial organizations that maintain them (commercial and/or society-based), editorial handling (academic or professional editors), editorial board composition (academic or professional), payment options to cover editorial costs (open access or pay-to-read), indexation, visibility, branding, and other aspects. Additionally, online submissions of non-revised versions of manuscripts prior to seeking publication in a peer-reviewed journal (a practice known as pre-printing) are a growing trend in biological sciences. In this changing landscape, researchers in biochemistry and molecular biology must rethink their priorities in terms of scientific output dissemination. The evaluation processes and institutional funding for scientific publications should also be revised accordingly. This article presents the results of discussions within the Department of Biochemistry, University of São Paulo, on this subject.
En trabajos previos hemos demostramos la existencia de interacciones bi-direccionales entre las v... more En trabajos previos hemos demostramos la existencia de interacciones bi-direccionales entre las vías de los progestá- genos y de la heregulina (HRG) en cáncer mamario. Encontramos que los progestágenos regulan la actividad y expresión del ErbB-2 y de la HRG. Describimos que la interacción entre la vía de la progesterona y de la HRG ocurre a nivel del PR que es activado transcripcionalmente por la HRG. Además encontramos que los progestágenos inducen la activación transcripcional de la proteína transductora de señales y activadora de la transcripción 3 (Stat3), que es un requisito para el crecimiento inducido por progestágenos en cáncer mamario. Demostramos que Stat3 es un punto de convergencia entre las vías de PR y de HRG/ErbB-2 en cáncer de mama, ya que la HRG, a través del ErbB-2, induce la activación de Stat3 mediante la integración del PR como molécula señalizadora. En línea con estos resultados, describimos la función de Stat3 como coactivador del PR activado por progesterona ...
Fil: Elizalde, Patricia Virginia. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Ins... more Fil: Elizalde, Patricia Virginia. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Instituto de Biologia y Medicina Experimental. Fundacion de Instituto de Biologia y Medicina Experimental. Instituto de Biologia y Medicina Experimental; Argentina
Journal of Physics: Materials, 2021
Photodynamic therapy (PDT) has been clinically applied to cure various diseases including cancer.... more Photodynamic therapy (PDT) has been clinically applied to cure various diseases including cancer. Indeed, photophrin (porfimer sodium, Axcan Pharma, Montreal, Canada), a heterogenous mixture of porphyrins, was the first photosensitizer (PS) approved for the treatment of human bladder cancer in 1993 in Canada. Over the past 10 years the use of PDT in the treatment of benign and malignant lesions has increased dramatically. However, PDT is still considered as an adjuvant strategy due to its limitations, primarily including low tissue penetration by light and inaccurate lesion selectivity by the PSs. To overcome this scenario, new technologies and approaches including nanotechnology have been incorporated into the concept of PS formulations as PS delivery systems, as PSs per se or as energy transducers. The ideal nanophotosensitizer (NPS) for cancer therapy should possess the following characteristics: biocompatibility and biodegradability without toxicity, stability in physiological c...
Journal of Proteomics, 2021
A new method to probe the conformational changes of glycoproteins on a systems-wide scale, termed... more A new method to probe the conformational changes of glycoproteins on a systems-wide scale, termed limited deglycosylation assay (LDA), is described. The method measures the differential rate of deglycosylation of N-glycans on natively folded proteins by the common peptide:N-glycosidase F (PNGase F) enzyme which in turn informs on their spatial presentation and solvent exposure on the protein surface hence ultimately the glycoprotein conformation. LDA involves 1) protein-level N-deglycosylation under native conditions, 2) trypsin digestion, 3) glycopeptide enrichment, 4) peptide-level N-deglycosylation and 5) quantitative MS-based analysis of formerly N-glycosylated peptides (FNGPs). LDA was initially developed and the experimental conditions optimized using bovine RNase B and fetuin. The method was then applied to glycoprotein extracts from LLC-MK2 epithelial cells upon treatment with dithiothreitol to induce endoplasmic reticulum stress and promote protein misfolding. Data from the LDA and 3D structure analysis showed that glycoproteins predominantly undergo structural changes in loops/turns upon ER stress as exemplified with detailed analysis of ephrin-A5, GALNT10, PVR and BCAM. These results show that LDA accurately reports on systems-wide conformational changes of glycoproteins induced under controlled treatment regimes. Thus, LDA opens avenues to study glycoprotein structural changes in a range of other physiological and pathophysiological conditions relevant to acute and chronic diseases. SIGNIFICANCE: We describe a novel method termed limited deglycosylation assay (LDA), to probe conformational changes of glycoproteins on a systems-wide scale. This method improves the current toolbox of structural proteomics by combining site and conformational-specific PNGase F enzymatic activity with large scale quantitative proteomics. X-ray crystallography, nuclear magnetic resonance spectroscopy and cryoEM techniques are the major techniques applied to elucidate macromolecule structures. However, the size and heterogeneity of the oligosaccharide chains poses several challenges to the applications of these techniques to glycoproteins. The LDA method presented here, can be applied to a range of pathophysiological conditions and expanded to investigate PTMs-mediated structural changes in complex proteomes.
Cell Death & Disease, 2020
Lack of effective treatments for aggressive breast cancer is still a major global health problem.... more Lack of effective treatments for aggressive breast cancer is still a major global health problem. We have previously reported that photodynamic therapy using methylene blue as photosensitizer (MB-PDT) massively kills metastatic human breast cancer, marginally affecting healthy cells. In this study, we aimed to unveil the molecular mechanisms behind MB-PDT effectiveness and specificity towards tumor cells. Through lipidomics and biochemical approaches, we demonstrated that MB-PDT efficiency and specificity rely on polyunsaturated fatty acid-enriched membranes and on the better capacity to deal with photo-oxidative damage displayed by non-tumorigenic cells. We found out that, in tumorigenic cells, lysosome membrane permeabilization is accompanied by ferroptosis and/or necroptosis. Our results also pointed at a cross-talk between lysosome-dependent cell death (LDCD) and necroptosis induction after photo-oxidation, and contributed to broaden the understanding of MB-PDT-induced mechanism...
Islets, 2011
Background: Many studies have evaluated whether there are characteristics related to pancreas don... more Background: Many studies have evaluated whether there are characteristics related to pancreas donors and the islet isolation process that can influence pancreatic islet yield. However, this analysis has not yet been performed in Brazil, one of the world leaders in whole pancreas organ transplantation (WOPT), where pancreas allocation for pancreatic islet transplantation (PIT) has no officially defined criteria. Definition of parameters that would predict the outcome of islet isolation from local pancreas donors would be useful for defining allocation priority in Brazil. Objective: To analyze the relationship between multiple donor-related and islet isolation variables with the total number of isolated pancreatic islet equivalents (IEQ) in a Brazilian sample of pancreas donors. Methods: Several variables were analyzed in 74 pancreata relative to the outcome of total IEQs obtained at the end of the process. Results: In univariate analysis, body mass index (BMI) (p = 0.003), the presence of fatty infiltrates in the pancreas as observed during harvesting (p = 0.042) and pancreas digestion time (p = 0.046) were identified as variables related to a greater IEQ yield. In a multivariate analysis a statistically significant contribution to the variability of islet yield was found only for the BMI (p = 0.017). A ROC curve defined a BMI = 30 as a cutoff point, with pancreata from donors with BMI > 30 yielding more islets than donors with BMI < 30 (p < 0.001). Conclusion: These data reinforce the importance of the donor BMI as a defining parameter for successful islet isolation and establishes this variable as a potential pancreas allocation criterion in Brazil, where there is unequal competition for good quality organs between WOPT and PIT.
Journal of Endocrinology, 2004
Strategies to differentiate progenitor cells into β cells in vitro have been considered as an alt... more Strategies to differentiate progenitor cells into β cells in vitro have been considered as an alternative to increase β cell availability prior to transplantation. It has recently been suggested that nestin-positive cells could be multipotential stem cells capable of expressing endocrine markers upon specific stimulation; however, this issue still remains controversial. Here, we characterized short- and long-term islet cell cultures derived from three different human islet preparations, with respect to expression of nestin and islet cell markers, using confocal microscopy and semi-quantitative RT-PCR. The number of nestin-positive cells was found to be strikingly high in long-term cultures. In addition, a large proportion (49.7%) of these nestin-positive cells, present in long-term culture, are shown to be proliferative, as judged by BrdU incorporation. The proportion of insulin-positive cells was found to be high in short-term (up to 28 days) cultures and declined thereafter, when ...
ß cell replacement with either pancreas or islet transplantation has progressed immensely over th... more ß cell replacement with either pancreas or islet transplantation has progressed immensely over the last decades with current 1and 5-year insulin independence rates of ~85% and ~50%, respectively. Recent advances are largely attributed to improvements in immunosuppressive regimen, donor selection and surgical technique. However, both strategies are compromised by a scarce donor source. Xenotransplantation provides a potential solution by providing a theoretically unlimited supply of islets, but clinical application has been limited by concerns for a potent immune response against xenogeneic tissue. ß cell clusters derived from embryonic or induced pluripotent stem (iPS) cells represent another promising unlimited source of insulin producing cells, but clinical application is pending further advances in the function of the ß cell like clusters. Exciting developments and rapid progress in all areas of ß cell replacement prompted a lively debate by members of the young investigator comm...
![Research paper thumbnail of [Islet transplantation as a clinical tool: present state and future perspectives]](https://attachments.academia-assets.com/78958565/thumbnails/1.jpg)
](Arquivos brasileiros de endocrinologia e metabologia, 2009
Islet transplant is an innovative treatment for type 1 diabetic patients, which still lies betwee... more Islet transplant is an innovative treatment for type 1 diabetic patients, which still lies between experimental and approved transplant therapy. Islet cells are seeded in a non-physiological territory where an uncertain fraction will be able to adapt and survive. Thus, the challenge lies in improving the whole procedure, employing the tools of cell biology, immunology and laboratory techniques, in order to reach the results obtained with whole organ transplant. This review describes the procedure, its progress to the present methodology and clinical results obtained. Future perspectives of islet transplantation in the light of recent biotechnological advances are also focused.
Background: Metastasis is the main factor responsible for death in breast cancer patients. Matrix... more Background: Metastasis is the main factor responsible for death in breast cancer patients. Matrix metalloproteinases (MMPs) and their inhibitors, known as tissue inhibitors of MMPs (TIMPs), and the membraneassociated MMP inhibitor (RECK), are essential for the metastatic process. We have previously shown a positive correlation between MMPs and their inhibitors expression during breast cancer progression; however, the molecular mechanisms underlying this coordinate regulation remain unknown. In this report, we investigated whether TGF-b1 could be a common regulator for MMPs, TIMPs and RECK in human breast cancer cell models. Methods: The mRNA expression levels of TGF-b isoforms and their receptors were analyzed by qRT-PCR in a panel of five human breast cancer cell lines displaying different degrees of invasiveness and metastatic potential. The highly invasive MDA-MB-231 cell line was treated with different concentrations of recombinant TGF-b1 and also with pharmacological inhibitors...
Photochemical & Photobiological Sciences
Pancreatic ductal adenocarcinomas (PDAC) are the fourth leading cause of death due to neoplasms. ... more Pancreatic ductal adenocarcinomas (PDAC) are the fourth leading cause of death due to neoplasms. In view of the urgent need of effective treatments for PDAC, photodynamic therapy (PDT) appears as a promising alternative. However, its efficacy against PDAC and the mechanisms involved in cell death induction remain unclear. In this study, we set out to evaluate PDT's cytotoxicity using methylene blue (MB) as a photosensitizer (PS) (MB-PDT) and to evaluate the contribution of necroptosis in its effect in human PDAC cells. Our results demonstrated that MB-PDT induced significant death of different human PDAC models presenting two different susceptibility profiles. This effect was independent of MB uptake or its subcellular localization. We found that the ability of triggering necroptosis was determinant to increase the treatment efficiency. Analysis of single cell RNA-seq data from normal and neoplastic human pancreatic tissues showed that specific necroptosis proteins RIPK1, RIPK3 and MLKL presented significant higher expression levels in cells displaying a transformed phenotype providing further support to the use of approaches that activate necroptosis, like MB-PDT, as useful adjunct to surgery of PDAC to tackle the problem of microscopic residual disease as well as to minimize the chance of local and metastatic recurrence.
Research article Generation and characterization of human insulin-releasing cell lines
Photodiagnosis and Photodynamic Therapy, 2015
s / Photodiagnosis and Photodynamic Therapy 12 (2015) 325–375 349 treatment outcome as an attempt... more s / Photodiagnosis and Photodynamic Therapy 12 (2015) 325–375 349 treatment outcome as an attempt to predict the PDT efficiency. The clinical study is being conducted at Hospital Amaral Carvalho in Jahu, and 100 lesions of the basal cell carcinoma types treated with PDT using Aminolevulinic Acid with photosensitizer. The thermal changes of the lesions are registeredwith a thermal imager (Fluke® FLK-Ti400) during four phases: lesion initial, lesion curetted, after the drug incubation and after the illumination in the two sessions. The temperature variation was compared with the result of the tissue biopsies after 30 days. Significant results are being collect in order to better understand the PDT response. http://dx.doi.org/10.1016/j.pdpdt.2015.07.095 Unveiling the molecular mechanisms involved in the cytotoxicity induced by photodynamic therapy in human breast cancer cells A.F. dos Santos, L.F. Terra, R.A.M. Wailemann, T.C. Oliveira, M.S. Baptista, L. Labriola Biochemistry Department, Chemistry Institute, University of Sao Paulo (USP), Sao Paulo, Brazil The efficacy of photodynamic therapy (PDT) on mammary tumors as well as the mechanisms of cell death remains unclear. We evaluated the cytotoxicity of PDT by using methylene blue (MB-PDT) in two (2D) and three (3D) dimension cultures of human breast tumor cells and the death pathways involved. Invasive, non-invasive and non-tumorigenic cells were incubated with different MB concentrations and irradiated ( =640nm) at 4.5 J/cm2. Fluorescence microscopy was used to evaluate cell viability and death. Autophagy was evaluated also by Western blotting. The role of autophagy was investigated using chloroquine (CQ) and LY294002 (LY) as inhibitors and rapamycin (RAPA) as activator. MB-PDT increased cell death of both tumoral cell lines in 2D or 3D culture, but was significantly lower in non-tumorigenic cells despite the higher intracellular concentration ofMB. No dark effect and no morphological signs of apoptotic were observed, but there were autophagy increase. After cotreatmentwith CQor LY the cytotoxicity was increased and with RAPA it was lowered. We showed that MB-PDT induced selective cell death in a model that recapitulates the morphology of glandular epithelium, and that autophagy is mainly related with cytoprotection. Our observations indicate that PDT is an effective therapy displaying minimal side effects. http://dx.doi.org/10.1016/j.pdpdt.2015.07.096 Comparison of the response and mechanisms of MAL-PDT of different squamous carcinoma cell lines Daniela Leon1, Ramon Silva1, Natalia Inada2, Cristina Kurachi2, Aleida Vivallo1, Priscilla Brebi1, Juan Carlos Roa3 1 Departamento de Anatomia Patologica, Laboratorio de Patologia Molecular BIOREN-CEGIN, Universidad de La Frontera, Chile 2 Sao Carlos Institute of Physics, University of Sao Paulo, Brazil 3 Departamento de Patologia, Facultad de Medicina, Pontificia Universidad Catolica de Chile, Chile Introduction: Non-melanoma skin cancer (NMSC) is the most common neoplasia worldwide and can be treated with Photodynamic Therapy (PDT). NMSC has good outcome after PDT, however, recurrent lesions has been reported. The aim of this study was to compare the effect of PDT on two non-melanoma skin cancer cell lines. Methodology: A keratinocytes cell line and two squamous carcinoma cell lines, A-431 and HSC-1, were used. PDT was carried outusingmethyl aminolevulinate (MAL) as aprecursor of Protoporphyrin IX. All cell lineswere incubatedwith distinct concentrations ofMAL and irradiatedwith different red light fluences. Cell viability wasevaluated24hafter PDTbyMTT, TrypanBluedyeexclusionand flow cytometry assays. In addition, ROS production was measured by flow cytometry. Results: A-431 cells were more resistant to PDT cytotoxic effect than HSC-1 cells compared the same condition of treatment. Meanwhile, keratinocyteswere themost sensitive to PDT. ROS production after PDT varied among the skin cell type. Conclusion: Cells from squamous skin cancer have a different response to the same PDT conditions.Wewill investigate the resistant phenotype of those cells for improving the effect of PDT on them. http://dx.doi.org/10.1016/j.pdpdt.2015.07.097 Penetration depth of the 664-nm semiconductor laser light with talaporfin sodium into human brain tissue with glioma Soko Ikuta1, Yuki Kawase2, Yoshihiro Muragaki1,3, Takashi Maruyama1,3, Masayuki Nitta3, Taiichi Saito3, Hiroshi Iseki1 1 Institute of Advanced Biomedical Engineering & Science, Tokyo Women’s Medical University, Japan 2 Panasonic Healthcare Co., Ltd., Japan 3 Department of Neurosurgery, Tokyo Women’s Medical University, Japan Glioma cell has characteristicswhich invasively spread into normal brain tissue. Talaporfin-PDT is assumed to be a promising to affect infiltrating glioma cells where the extirpation cavity, but there is no data showed about its optical penetration depth and cellular cytotoxicity in human brain tissue. This study evaluated these affections with…
Brazilian Journal of Medical and Biological Research, 2019
The scientific publication landscape is changing quickly, with an enormous increase in options an... more The scientific publication landscape is changing quickly, with an enormous increase in options and models. Articles can be published in a complex variety of journals that differ in their presentation format (online-only or in-print), editorial organizations that maintain them (commercial and/or society-based), editorial handling (academic or professional editors), editorial board composition (academic or professional), payment options to cover editorial costs (open access or pay-to-read), indexation, visibility, branding, and other aspects. Additionally, online submissions of non-revised versions of manuscripts prior to seeking publication in a peer-reviewed journal (a practice known as pre-printing) are a growing trend in biological sciences. In this changing landscape, researchers in biochemistry and molecular biology must rethink their priorities in terms of scientific output dissemination. The evaluation processes and institutional funding for scientific publications should also be revised accordingly. This article presents the results of discussions within the Department of Biochemistry, University of São Paulo, on this subject.
En trabajos previos hemos demostramos la existencia de interacciones bi-direccionales entre las v... more En trabajos previos hemos demostramos la existencia de interacciones bi-direccionales entre las vías de los progestá- genos y de la heregulina (HRG) en cáncer mamario. Encontramos que los progestágenos regulan la actividad y expresión del ErbB-2 y de la HRG. Describimos que la interacción entre la vía de la progesterona y de la HRG ocurre a nivel del PR que es activado transcripcionalmente por la HRG. Además encontramos que los progestágenos inducen la activación transcripcional de la proteína transductora de señales y activadora de la transcripción 3 (Stat3), que es un requisito para el crecimiento inducido por progestágenos en cáncer mamario. Demostramos que Stat3 es un punto de convergencia entre las vías de PR y de HRG/ErbB-2 en cáncer de mama, ya que la HRG, a través del ErbB-2, induce la activación de Stat3 mediante la integración del PR como molécula señalizadora. En línea con estos resultados, describimos la función de Stat3 como coactivador del PR activado por progesterona ...
Fil: Elizalde, Patricia Virginia. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Ins... more Fil: Elizalde, Patricia Virginia. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Instituto de Biologia y Medicina Experimental. Fundacion de Instituto de Biologia y Medicina Experimental. Instituto de Biologia y Medicina Experimental; Argentina
Journal of Physics: Materials, 2021
Photodynamic therapy (PDT) has been clinically applied to cure various diseases including cancer.... more Photodynamic therapy (PDT) has been clinically applied to cure various diseases including cancer. Indeed, photophrin (porfimer sodium, Axcan Pharma, Montreal, Canada), a heterogenous mixture of porphyrins, was the first photosensitizer (PS) approved for the treatment of human bladder cancer in 1993 in Canada. Over the past 10 years the use of PDT in the treatment of benign and malignant lesions has increased dramatically. However, PDT is still considered as an adjuvant strategy due to its limitations, primarily including low tissue penetration by light and inaccurate lesion selectivity by the PSs. To overcome this scenario, new technologies and approaches including nanotechnology have been incorporated into the concept of PS formulations as PS delivery systems, as PSs per se or as energy transducers. The ideal nanophotosensitizer (NPS) for cancer therapy should possess the following characteristics: biocompatibility and biodegradability without toxicity, stability in physiological c...
Journal of Proteomics, 2021
A new method to probe the conformational changes of glycoproteins on a systems-wide scale, termed... more A new method to probe the conformational changes of glycoproteins on a systems-wide scale, termed limited deglycosylation assay (LDA), is described. The method measures the differential rate of deglycosylation of N-glycans on natively folded proteins by the common peptide:N-glycosidase F (PNGase F) enzyme which in turn informs on their spatial presentation and solvent exposure on the protein surface hence ultimately the glycoprotein conformation. LDA involves 1) protein-level N-deglycosylation under native conditions, 2) trypsin digestion, 3) glycopeptide enrichment, 4) peptide-level N-deglycosylation and 5) quantitative MS-based analysis of formerly N-glycosylated peptides (FNGPs). LDA was initially developed and the experimental conditions optimized using bovine RNase B and fetuin. The method was then applied to glycoprotein extracts from LLC-MK2 epithelial cells upon treatment with dithiothreitol to induce endoplasmic reticulum stress and promote protein misfolding. Data from the LDA and 3D structure analysis showed that glycoproteins predominantly undergo structural changes in loops/turns upon ER stress as exemplified with detailed analysis of ephrin-A5, GALNT10, PVR and BCAM. These results show that LDA accurately reports on systems-wide conformational changes of glycoproteins induced under controlled treatment regimes. Thus, LDA opens avenues to study glycoprotein structural changes in a range of other physiological and pathophysiological conditions relevant to acute and chronic diseases. SIGNIFICANCE: We describe a novel method termed limited deglycosylation assay (LDA), to probe conformational changes of glycoproteins on a systems-wide scale. This method improves the current toolbox of structural proteomics by combining site and conformational-specific PNGase F enzymatic activity with large scale quantitative proteomics. X-ray crystallography, nuclear magnetic resonance spectroscopy and cryoEM techniques are the major techniques applied to elucidate macromolecule structures. However, the size and heterogeneity of the oligosaccharide chains poses several challenges to the applications of these techniques to glycoproteins. The LDA method presented here, can be applied to a range of pathophysiological conditions and expanded to investigate PTMs-mediated structural changes in complex proteomes.
Cell Death & Disease, 2020
Lack of effective treatments for aggressive breast cancer is still a major global health problem.... more Lack of effective treatments for aggressive breast cancer is still a major global health problem. We have previously reported that photodynamic therapy using methylene blue as photosensitizer (MB-PDT) massively kills metastatic human breast cancer, marginally affecting healthy cells. In this study, we aimed to unveil the molecular mechanisms behind MB-PDT effectiveness and specificity towards tumor cells. Through lipidomics and biochemical approaches, we demonstrated that MB-PDT efficiency and specificity rely on polyunsaturated fatty acid-enriched membranes and on the better capacity to deal with photo-oxidative damage displayed by non-tumorigenic cells. We found out that, in tumorigenic cells, lysosome membrane permeabilization is accompanied by ferroptosis and/or necroptosis. Our results also pointed at a cross-talk between lysosome-dependent cell death (LDCD) and necroptosis induction after photo-oxidation, and contributed to broaden the understanding of MB-PDT-induced mechanism...
Islets, 2011
Background: Many studies have evaluated whether there are characteristics related to pancreas don... more Background: Many studies have evaluated whether there are characteristics related to pancreas donors and the islet isolation process that can influence pancreatic islet yield. However, this analysis has not yet been performed in Brazil, one of the world leaders in whole pancreas organ transplantation (WOPT), where pancreas allocation for pancreatic islet transplantation (PIT) has no officially defined criteria. Definition of parameters that would predict the outcome of islet isolation from local pancreas donors would be useful for defining allocation priority in Brazil. Objective: To analyze the relationship between multiple donor-related and islet isolation variables with the total number of isolated pancreatic islet equivalents (IEQ) in a Brazilian sample of pancreas donors. Methods: Several variables were analyzed in 74 pancreata relative to the outcome of total IEQs obtained at the end of the process. Results: In univariate analysis, body mass index (BMI) (p = 0.003), the presence of fatty infiltrates in the pancreas as observed during harvesting (p = 0.042) and pancreas digestion time (p = 0.046) were identified as variables related to a greater IEQ yield. In a multivariate analysis a statistically significant contribution to the variability of islet yield was found only for the BMI (p = 0.017). A ROC curve defined a BMI = 30 as a cutoff point, with pancreata from donors with BMI > 30 yielding more islets than donors with BMI < 30 (p < 0.001). Conclusion: These data reinforce the importance of the donor BMI as a defining parameter for successful islet isolation and establishes this variable as a potential pancreas allocation criterion in Brazil, where there is unequal competition for good quality organs between WOPT and PIT.
Journal of Endocrinology, 2004
Strategies to differentiate progenitor cells into β cells in vitro have been considered as an alt... more Strategies to differentiate progenitor cells into β cells in vitro have been considered as an alternative to increase β cell availability prior to transplantation. It has recently been suggested that nestin-positive cells could be multipotential stem cells capable of expressing endocrine markers upon specific stimulation; however, this issue still remains controversial. Here, we characterized short- and long-term islet cell cultures derived from three different human islet preparations, with respect to expression of nestin and islet cell markers, using confocal microscopy and semi-quantitative RT-PCR. The number of nestin-positive cells was found to be strikingly high in long-term cultures. In addition, a large proportion (49.7%) of these nestin-positive cells, present in long-term culture, are shown to be proliferative, as judged by BrdU incorporation. The proportion of insulin-positive cells was found to be high in short-term (up to 28 days) cultures and declined thereafter, when ...
ß cell replacement with either pancreas or islet transplantation has progressed immensely over th... more ß cell replacement with either pancreas or islet transplantation has progressed immensely over the last decades with current 1and 5-year insulin independence rates of ~85% and ~50%, respectively. Recent advances are largely attributed to improvements in immunosuppressive regimen, donor selection and surgical technique. However, both strategies are compromised by a scarce donor source. Xenotransplantation provides a potential solution by providing a theoretically unlimited supply of islets, but clinical application has been limited by concerns for a potent immune response against xenogeneic tissue. ß cell clusters derived from embryonic or induced pluripotent stem (iPS) cells represent another promising unlimited source of insulin producing cells, but clinical application is pending further advances in the function of the ß cell like clusters. Exciting developments and rapid progress in all areas of ß cell replacement prompted a lively debate by members of the young investigator comm...
Arquivos brasileiros de endocrinologia e metabologia, 2009
Islet transplant is an innovative treatment for type 1 diabetic patients, which still lies betwee... more Islet transplant is an innovative treatment for type 1 diabetic patients, which still lies between experimental and approved transplant therapy. Islet cells are seeded in a non-physiological territory where an uncertain fraction will be able to adapt and survive. Thus, the challenge lies in improving the whole procedure, employing the tools of cell biology, immunology and laboratory techniques, in order to reach the results obtained with whole organ transplant. This review describes the procedure, its progress to the present methodology and clinical results obtained. Future perspectives of islet transplantation in the light of recent biotechnological advances are also focused.
Background: Metastasis is the main factor responsible for death in breast cancer patients. Matrix... more Background: Metastasis is the main factor responsible for death in breast cancer patients. Matrix metalloproteinases (MMPs) and their inhibitors, known as tissue inhibitors of MMPs (TIMPs), and the membraneassociated MMP inhibitor (RECK), are essential for the metastatic process. We have previously shown a positive correlation between MMPs and their inhibitors expression during breast cancer progression; however, the molecular mechanisms underlying this coordinate regulation remain unknown. In this report, we investigated whether TGF-b1 could be a common regulator for MMPs, TIMPs and RECK in human breast cancer cell models. Methods: The mRNA expression levels of TGF-b isoforms and their receptors were analyzed by qRT-PCR in a panel of five human breast cancer cell lines displaying different degrees of invasiveness and metastatic potential. The highly invasive MDA-MB-231 cell line was treated with different concentrations of recombinant TGF-b1 and also with pharmacological inhibitors...