Tiago Fernandes | PUC Minas (original) (raw)

Papers by Tiago Fernandes

Research paper thumbnail of Synthesis and catalytic properties of manganese(II) and oxovanadium(IV) complexes anchored to mesoporous MCM-41

Microporous and Mesoporous Materials, 2008

with the 1,4-diazabutadiene (DAB) ligands Ph-DAB-(CH 2 ) 3 R [R = Si(OEt) 3 (1a); H (1b)] (L) lea... more with the 1,4-diazabutadiene (DAB) ligands Ph-DAB-(CH 2 ) 3 R [R = Si(OEt) 3 (1a); H (1b)] (L) leads to complexes of the type [Mn II (acac) 2 L] and [V IV O(acac)L]Cl in good yields. These complexes were characterised by spectroscopic techniques, and magnetic measurements showed that in the case of the manganese the metal was reduced during the reaction with the nitrogen ligand.

Research paper thumbnail of Synthesis and catalytic properties of manganese(II) and oxovanadium(IV) complexes anchored to mesoporous MCM-41

Microporous and Mesoporous Materials, 2008

with the 1,4-diazabutadiene (DAB) ligands Ph-DAB-(CH 2 ) 3 R [R = Si(OEt) 3 (1a); H (1b)] (L) lea... more with the 1,4-diazabutadiene (DAB) ligands Ph-DAB-(CH 2 ) 3 R [R = Si(OEt) 3 (1a); H (1b)] (L) leads to complexes of the type [Mn II (acac) 2 L] and [V IV O(acac)L]Cl in good yields. These complexes were characterised by spectroscopic techniques, and magnetic measurements showed that in the case of the manganese the metal was reduced during the reaction with the nitrogen ligand.

Research paper thumbnail of Improving communication skills of children with ASDs through interaction with virtual characters

This article presents the LIFEisGAME project, a serious game that will help children with ASDs to... more This article presents the LIFEisGAME project, a serious game that will help children with ASDs to recognize and express emotions through facial expressions. The game design tackles the main experiential learning cycle of emotion recognition: watch and recognize, learn by doing, recognize and mimic, generalize or knowledge transfer to real life. We briefly describe the technology behind the character animation pipeline centered on the creation of a generic rig. Then, we detail the facial expression analyzer that uses Active Appearance Models. Last, we describe the user study experiment using game mode “recognize the expression”.

Research paper thumbnail of Improving communication skills of children with ASDs through interaction with virtual characters

This article presents the LIFEisGAME project, a serious game that will help children with ASDs to... more This article presents the LIFEisGAME project, a serious game that will help children with ASDs to recognize and express emotions through facial expressions. The game design tackles the main experiential learning cycle of emotion recognition: watch and recognize, learn by doing, recognize and mimic, generalize or knowledge transfer to real life. We briefly describe the technology behind the character animation pipeline centered on the creation of a generic rig. Then, we detail the facial expression analyzer that uses Active Appearance Models. Last, we describe the user study experiment using game mode “recognize the expression”.

Research paper thumbnail of On-Chip, Cell-Based Microarray Immunofluorescence Assay for High-Throughput Analysis of Target Proteins

Analytical Chemistry, 2008

We have developed an immunofluorescence-based assay for high-throughput analysis of target protei... more We have developed an immunofluorescence-based assay for high-throughput analysis of target proteins on a threedimensional cellular microarray platform. This process integrates the use of three-dimensional cellular microarrays, which should better mimic the cellular microenvironment, with sensitive immunofluorescence detection and provides quantitative information on cell function. To demonstrate this assay platform, we examined the accumulation of the r subunit of the hypoxia-inducible factor (HIF-1r) after chemical stimulation of human pancreatic tumor cells encapsulated in 3D alginate spots in volumes as low as 60 nL. We also tested the effect of the known dysregulator of HIF-1r, 2-methoxyestradiol (2ME2), on the levels of HIF-1r using a dual microarray stamping technique. This chip-based in situ Western immunoassay protocol was able to provide quantitative information on cell function, namely, the cellular response to hypoxia mimicking conditions and the reduction of HIF-1r levels after cell treatment with 2ME2. This system is the first to enable high-content screening of cellular protein levels on a 3D human cell microarray platform.

Research paper thumbnail of Three-dimensional cell culture microarray for high-throughput studies of stem cell fate

Biotechnology and Bioengineering, 2010

We have developed a novel three-dimensional (3D) cellular microarray platform to enable the rapid... more We have developed a novel three-dimensional (3D) cellular microarray platform to enable the rapid and efficient tracking of stem cell fate and quantification of specific stem cell markers. This platform consists of a miniaturized 3D cell culture array on a functionalized glass slide for spatially addressable high-throughput screening. A microarray spotter was used to deposit cells onto a modified glass surface to yield an array consisting of cells encapsulated in alginate gel spots with volumes as low as 60 nL. A method based on an immunofluorescence technique scaled down to function on a cellular microarray was also used to quantify specific cell marker protein levels in situ. Our results revealed that this platform is suitable for studying the expansion of mouse embryonic stem (ES) cells as they retain their pluripotent and undifferentiated state. We also examined neural commitment of mouse ES cells on the microarray and observed the generation of neuroectodermal precursor cells characterized by expression of the neural marker Sox-1, whose levels were also measured in situ using a GFP reporter system. In addition, the high-throughput capacity of the platform was tested using a dual-slide system that allowed rapid screening of the effects of tretinoin and fibroblast growth factor-4 (FGF-4) on the pluripotency of mouse ES cells. This high-throughput platform is a powerful new tool for investigating cellular mechanisms involved in stem cell expansion and differentiation and provides the basis for rapid identification of signals and conditions that can be used to direct cellular responses. Biotechnol. Bioeng. 2010; 106: 106–118. © 2010 Wiley Periodicals, Inc.

Research paper thumbnail of Stem cell cultivation in bioreactors

Biotechnology Advances, 2011

Cell-based therapies have generated great interest in the scientific and medical communities, and... more Cell-based therapies have generated great interest in the scientific and medical communities, and stem cells in particular are very appealing for regenerative medicine, drug screening and other biomedical applications. These unspecialized cells have unlimited self-renewal capacity and the remarkable ability to produce mature cells with specialized functions, such as blood cells, nerve cells or cardiac muscle. However, the actual number of cells that can be obtained from available donors is very low. One possible solution for the generation of relevant numbers of cells for several applications is to scale-up the culture of these cells in vitro. This review describes recent developments in the cultivation of stem cells in bioreactors, particularly considerations regarding critical culture parameters, possible bioreactor configurations, and integration of novel technologies in the bioprocess development stage. We expect that this review will provide updated and detailed information focusing on the systematic production of stem cell products in compliance with regulatory guidelines, while using robust and cost-effective approaches.

Research paper thumbnail of Kinetic and metabolic analysis of mouse embryonic stem cell expansion under serum-free conditions

Biotechnology Letters, 2010

There is a need for a deeper understanding of the biochemical events affecting embryonic stem (ES... more There is a need for a deeper understanding of the biochemical events affecting embryonic stem (ES) cell culture by analyzing the expansion of mouse ES cells in terms of both cell growth and metabolic kinetics. The influence of the initial cell density on cell expansion was assessed. Concomitantly, the biochemical profile of the culture was evaluated, which allowed measuring the consumption of important substrates, such as glucose and glutamine, and the production of metabolic byproducts, like lactate. The results suggest a more efficient cell metabolism in serum-free conditions and a preferential use of glutaminolysis as an energy source during cell expansion at low seeding densities. This work contributes to the development of fully-controlled bioprocesses to produce relevant numbers of ES cells for cell therapies and high-throughput drug screening.

Research paper thumbnail of Effect of hypoxia on proliferation and neural commitment of embryonic stem cells at different stages of pluripotency

We have studied the effect of low oxygen levels (2% O2), or hypoxia, in the expansion and neural ... more We have studied the effect of low oxygen levels (2% O2), or hypoxia, in the expansion and neural commitment of mouse embryonic stem (ES) cells. When ES cells were maintained in culture with leukemia inhibitory factor (LIF), cell proliferation was reduced at low oxygen levels and a simultaneous reduction in cell viability was also observed. Morphological changes and different cell cycle patterns also occurred, suggesting some early differentiation under hypoxic conditions. However, when cells were maintained in a ground state of pluripotency, by inhibition of autocrine FGF4/ERK and GSK3 signaling, hypoxia did not affect cell proliferation, and did not induce early differentiation. Nevertheless, during neural commitment, low oxygen tension exerted a positive effect on early differentiation of ES cells, resulting in a faster commitment towards neural progenitors. Overall our results demonstrate the need to specifically regulate the oxygen content, especially hypoxia, along with other culture conditions, when developing new strategies for ES cell expansion and/or controlled differentiation.

Research paper thumbnail of High-throughput cellular microarray platforms: applications in drug discovery, toxicology and stem cell research

Trends in Biotechnology, 2009

Cellular microarrays are powerful experimental tools for high-throughput screening of large numbe... more Cellular microarrays are powerful experimental tools for high-throughput screening of large numbers of test samples. Miniaturization increases assay throughput while reducing reagent consumption and the number of cells required, making these systems attractive for a wide range of assays in drug discovery, toxicology, stem cell research and potentially therapy. Here, we provide an overview of the emerging technologies that can be used to generate cellular microarrays, and we highlight recent significant advances in the field. This emerging and multidisciplinary approach offers new opportunities for the design and control of stem cells in tissue engineering and cellular therapies and promises to expedite drug discovery in the biotechnology and pharmaceutical industries.

Research paper thumbnail of Different stages of pluripotency determine distinct patterns of proliferation, metabolism, and lineage commitment of embryonic stem cells under hypoxia

Stem Cell Research, 2010

Oxygen tension is an important component of the stem cell microenvironment. Herein, we have studi... more Oxygen tension is an important component of the stem cell microenvironment. Herein, we have studied the effect of low oxygen levels (2% O 2 ), or hypoxia, in the expansion of mouse embryonic stem (ES) cells. In the presence of leukemia inhibitory factor (LIF), cell proliferation was reduced under hypoxia and a simultaneous reduction in cell viability was also observed. Morphological changes and different cell cycle patterns were observed, suggesting some early differentiation under hypoxic conditions. However, when cells were maintained in a ground state of pluripotency, by inhibition of autocrine FGF4/ERK and GSK3 signaling, hypoxia did not affect cell proliferation, and did not induce early differentiation. As expected, there was an increase in lactate-specific production rate and a significant increase in the glucose consumption under hypoxic conditions. Nevertheless, during neural commitment, low oxygen tension exerted a positive effect on early differentiation of ground-state ES cells, resulting in a faster commitment toward neural progenitors. Overall our results demonstrate the need to specifically regulate the oxygen content, especially hypoxia, along with other culture conditions, when developing new strategies for ES cell expansion and/or controlled differentiation.

Research paper thumbnail of Synthesis and catalytic properties of manganese(II) and oxovanadium(IV) complexes anchored to mesoporous MCM-41

Microporous and Mesoporous Materials, 2008

with the 1,4-diazabutadiene (DAB) ligands Ph-DAB-(CH 2 ) 3 R [R = Si(OEt) 3 (1a); H (1b)] (L) lea... more with the 1,4-diazabutadiene (DAB) ligands Ph-DAB-(CH 2 ) 3 R [R = Si(OEt) 3 (1a); H (1b)] (L) leads to complexes of the type [Mn II (acac) 2 L] and [V IV O(acac)L]Cl in good yields. These complexes were characterised by spectroscopic techniques, and magnetic measurements showed that in the case of the manganese the metal was reduced during the reaction with the nitrogen ligand.

Research paper thumbnail of Synthesis and catalytic properties of manganese(II) and oxovanadium(IV) complexes anchored to mesoporous MCM-41

Microporous and Mesoporous Materials, 2008

with the 1,4-diazabutadiene (DAB) ligands Ph-DAB-(CH 2 ) 3 R [R = Si(OEt) 3 (1a); H (1b)] (L) lea... more with the 1,4-diazabutadiene (DAB) ligands Ph-DAB-(CH 2 ) 3 R [R = Si(OEt) 3 (1a); H (1b)] (L) leads to complexes of the type [Mn II (acac) 2 L] and [V IV O(acac)L]Cl in good yields. These complexes were characterised by spectroscopic techniques, and magnetic measurements showed that in the case of the manganese the metal was reduced during the reaction with the nitrogen ligand.

Research paper thumbnail of Improving communication skills of children with ASDs through interaction with virtual characters

This article presents the LIFEisGAME project, a serious game that will help children with ASDs to... more This article presents the LIFEisGAME project, a serious game that will help children with ASDs to recognize and express emotions through facial expressions. The game design tackles the main experiential learning cycle of emotion recognition: watch and recognize, learn by doing, recognize and mimic, generalize or knowledge transfer to real life. We briefly describe the technology behind the character animation pipeline centered on the creation of a generic rig. Then, we detail the facial expression analyzer that uses Active Appearance Models. Last, we describe the user study experiment using game mode “recognize the expression”.

Research paper thumbnail of Improving communication skills of children with ASDs through interaction with virtual characters

This article presents the LIFEisGAME project, a serious game that will help children with ASDs to... more This article presents the LIFEisGAME project, a serious game that will help children with ASDs to recognize and express emotions through facial expressions. The game design tackles the main experiential learning cycle of emotion recognition: watch and recognize, learn by doing, recognize and mimic, generalize or knowledge transfer to real life. We briefly describe the technology behind the character animation pipeline centered on the creation of a generic rig. Then, we detail the facial expression analyzer that uses Active Appearance Models. Last, we describe the user study experiment using game mode “recognize the expression”.

Research paper thumbnail of On-Chip, Cell-Based Microarray Immunofluorescence Assay for High-Throughput Analysis of Target Proteins

Analytical Chemistry, 2008

We have developed an immunofluorescence-based assay for high-throughput analysis of target protei... more We have developed an immunofluorescence-based assay for high-throughput analysis of target proteins on a threedimensional cellular microarray platform. This process integrates the use of three-dimensional cellular microarrays, which should better mimic the cellular microenvironment, with sensitive immunofluorescence detection and provides quantitative information on cell function. To demonstrate this assay platform, we examined the accumulation of the r subunit of the hypoxia-inducible factor (HIF-1r) after chemical stimulation of human pancreatic tumor cells encapsulated in 3D alginate spots in volumes as low as 60 nL. We also tested the effect of the known dysregulator of HIF-1r, 2-methoxyestradiol (2ME2), on the levels of HIF-1r using a dual microarray stamping technique. This chip-based in situ Western immunoassay protocol was able to provide quantitative information on cell function, namely, the cellular response to hypoxia mimicking conditions and the reduction of HIF-1r levels after cell treatment with 2ME2. This system is the first to enable high-content screening of cellular protein levels on a 3D human cell microarray platform.

Research paper thumbnail of Three-dimensional cell culture microarray for high-throughput studies of stem cell fate

Biotechnology and Bioengineering, 2010

We have developed a novel three-dimensional (3D) cellular microarray platform to enable the rapid... more We have developed a novel three-dimensional (3D) cellular microarray platform to enable the rapid and efficient tracking of stem cell fate and quantification of specific stem cell markers. This platform consists of a miniaturized 3D cell culture array on a functionalized glass slide for spatially addressable high-throughput screening. A microarray spotter was used to deposit cells onto a modified glass surface to yield an array consisting of cells encapsulated in alginate gel spots with volumes as low as 60 nL. A method based on an immunofluorescence technique scaled down to function on a cellular microarray was also used to quantify specific cell marker protein levels in situ. Our results revealed that this platform is suitable for studying the expansion of mouse embryonic stem (ES) cells as they retain their pluripotent and undifferentiated state. We also examined neural commitment of mouse ES cells on the microarray and observed the generation of neuroectodermal precursor cells characterized by expression of the neural marker Sox-1, whose levels were also measured in situ using a GFP reporter system. In addition, the high-throughput capacity of the platform was tested using a dual-slide system that allowed rapid screening of the effects of tretinoin and fibroblast growth factor-4 (FGF-4) on the pluripotency of mouse ES cells. This high-throughput platform is a powerful new tool for investigating cellular mechanisms involved in stem cell expansion and differentiation and provides the basis for rapid identification of signals and conditions that can be used to direct cellular responses. Biotechnol. Bioeng. 2010; 106: 106–118. © 2010 Wiley Periodicals, Inc.

Research paper thumbnail of Stem cell cultivation in bioreactors

Biotechnology Advances, 2011

Cell-based therapies have generated great interest in the scientific and medical communities, and... more Cell-based therapies have generated great interest in the scientific and medical communities, and stem cells in particular are very appealing for regenerative medicine, drug screening and other biomedical applications. These unspecialized cells have unlimited self-renewal capacity and the remarkable ability to produce mature cells with specialized functions, such as blood cells, nerve cells or cardiac muscle. However, the actual number of cells that can be obtained from available donors is very low. One possible solution for the generation of relevant numbers of cells for several applications is to scale-up the culture of these cells in vitro. This review describes recent developments in the cultivation of stem cells in bioreactors, particularly considerations regarding critical culture parameters, possible bioreactor configurations, and integration of novel technologies in the bioprocess development stage. We expect that this review will provide updated and detailed information focusing on the systematic production of stem cell products in compliance with regulatory guidelines, while using robust and cost-effective approaches.

Research paper thumbnail of Kinetic and metabolic analysis of mouse embryonic stem cell expansion under serum-free conditions

Biotechnology Letters, 2010

There is a need for a deeper understanding of the biochemical events affecting embryonic stem (ES... more There is a need for a deeper understanding of the biochemical events affecting embryonic stem (ES) cell culture by analyzing the expansion of mouse ES cells in terms of both cell growth and metabolic kinetics. The influence of the initial cell density on cell expansion was assessed. Concomitantly, the biochemical profile of the culture was evaluated, which allowed measuring the consumption of important substrates, such as glucose and glutamine, and the production of metabolic byproducts, like lactate. The results suggest a more efficient cell metabolism in serum-free conditions and a preferential use of glutaminolysis as an energy source during cell expansion at low seeding densities. This work contributes to the development of fully-controlled bioprocesses to produce relevant numbers of ES cells for cell therapies and high-throughput drug screening.

Research paper thumbnail of Effect of hypoxia on proliferation and neural commitment of embryonic stem cells at different stages of pluripotency

We have studied the effect of low oxygen levels (2% O2), or hypoxia, in the expansion and neural ... more We have studied the effect of low oxygen levels (2% O2), or hypoxia, in the expansion and neural commitment of mouse embryonic stem (ES) cells. When ES cells were maintained in culture with leukemia inhibitory factor (LIF), cell proliferation was reduced at low oxygen levels and a simultaneous reduction in cell viability was also observed. Morphological changes and different cell cycle patterns also occurred, suggesting some early differentiation under hypoxic conditions. However, when cells were maintained in a ground state of pluripotency, by inhibition of autocrine FGF4/ERK and GSK3 signaling, hypoxia did not affect cell proliferation, and did not induce early differentiation. Nevertheless, during neural commitment, low oxygen tension exerted a positive effect on early differentiation of ES cells, resulting in a faster commitment towards neural progenitors. Overall our results demonstrate the need to specifically regulate the oxygen content, especially hypoxia, along with other culture conditions, when developing new strategies for ES cell expansion and/or controlled differentiation.

Research paper thumbnail of High-throughput cellular microarray platforms: applications in drug discovery, toxicology and stem cell research

Trends in Biotechnology, 2009

Cellular microarrays are powerful experimental tools for high-throughput screening of large numbe... more Cellular microarrays are powerful experimental tools for high-throughput screening of large numbers of test samples. Miniaturization increases assay throughput while reducing reagent consumption and the number of cells required, making these systems attractive for a wide range of assays in drug discovery, toxicology, stem cell research and potentially therapy. Here, we provide an overview of the emerging technologies that can be used to generate cellular microarrays, and we highlight recent significant advances in the field. This emerging and multidisciplinary approach offers new opportunities for the design and control of stem cells in tissue engineering and cellular therapies and promises to expedite drug discovery in the biotechnology and pharmaceutical industries.

Research paper thumbnail of Different stages of pluripotency determine distinct patterns of proliferation, metabolism, and lineage commitment of embryonic stem cells under hypoxia

Stem Cell Research, 2010

Oxygen tension is an important component of the stem cell microenvironment. Herein, we have studi... more Oxygen tension is an important component of the stem cell microenvironment. Herein, we have studied the effect of low oxygen levels (2% O 2 ), or hypoxia, in the expansion of mouse embryonic stem (ES) cells. In the presence of leukemia inhibitory factor (LIF), cell proliferation was reduced under hypoxia and a simultaneous reduction in cell viability was also observed. Morphological changes and different cell cycle patterns were observed, suggesting some early differentiation under hypoxic conditions. However, when cells were maintained in a ground state of pluripotency, by inhibition of autocrine FGF4/ERK and GSK3 signaling, hypoxia did not affect cell proliferation, and did not induce early differentiation. As expected, there was an increase in lactate-specific production rate and a significant increase in the glucose consumption under hypoxic conditions. Nevertheless, during neural commitment, low oxygen tension exerted a positive effect on early differentiation of ground-state ES cells, resulting in a faster commitment toward neural progenitors. Overall our results demonstrate the need to specifically regulate the oxygen content, especially hypoxia, along with other culture conditions, when developing new strategies for ES cell expansion and/or controlled differentiation.