In both embryonal carcinoma (EC) and embryonic stem (ES) cells, the differentiation pathway enter... more In both embryonal carcinoma (EC) and embryonic stem (ES) cells, the differentiation pathway entered after treatment with retinoic acid (RA) varies as it is based upon different conditions of culture. This study employs mouse EC cells P19 to investigate the effects of serum on RA-induced neural differentiation occurring in a simplified monolayer culture. Cell morphology and expression of lineage-specific molecular markers document that, while non-neural cell types arise after treatment with RA under serum-containing conditions, in chemically defined serum-free media RA induces massive neural differentiation in concentrations of 10(-9) M and higher. Moreover, not only neural (Mash-1) and neuroectodermal (Pax-6), but also endodermal (GATA-4, alpha-fetoprotein) genes are expressed at early stages of differentiation driven by RA under serum-free conditions. Furthermore, as determined by the luciferase reporter assay, the presence or absence of the serum does not affect the activity of th...
In both embryonal carcinoma (EC) and embryonic stem (ES) cells, the differentiation pathway enter... more In both embryonal carcinoma (EC) and embryonic stem (ES) cells, the differentiation pathway entered after treatment with retinoic acid (RA) varies as it is based upon different conditions of culture. This study employs mouse EC cells P19 to investigate the effects of serum on RA-induced neural differentiation occurring in a simplified monolayer culture. Cell morphology and expression of lineage-specific molecular markers document that, while non-neural cell types arise after treatment with RA under serum-containing conditions, in chemically defined serum-free media RA induces massive neural differentiation in concentrations of 10(-9) M and higher. Moreover, not only neural (Mash-1) and neuroectodermal (Pax-6), but also endodermal (GATA-4, alpha-fetoprotein) genes are expressed at early stages of differentiation driven by RA under serum-free conditions. Furthermore, as determined by the luciferase reporter assay, the presence or absence of the serum does not affect the activity of th...
Extracellular matrix (ECM) forms an essential part of tissue microenvironment. Compositional and ... more Extracellular matrix (ECM) forms an essential part of tissue microenvironment. Compositional and structural properties of ECM modulate behaviour of cells, including their differentiation, proliferation, and turnover. Here we aimed at detailed characterization of the nanostructure of lung ECM and evaluation of its bioactivity on cells grown in vitro. Mouse lungs were decellularized using 0.2 % sodium dodecyl sulphate, hypotonic solutions, and DNase. Morphological analysis of the resulting ECM scaffolds was performed by means of transmission and scanning electron microscopy. The ECM scaffolds retained 3D architecture of the main lung anatomical regions: the alveolar region and the blood/ airway network. The region-specific ECM nanotopology and organization of ECM macromolecules such as fibres of collagen, elastin, and fibrillin were characterized. The lung ECM scaffolds were also homogenized and applied as a supplement to growth medium on primary lung cells in vitro to test the bioact...
Extracellular matrix (ECM) forms an essential part of tissue microenvironment. Compositional and ... more Extracellular matrix (ECM) forms an essential part of tissue microenvironment. Compositional and structural properties of ECM modulate behaviour of cells, including their differentiation, proliferation, and turnover. Here we aimed at detailed characterization of the nanostructure of lung ECM and evaluation of its bioactivity on cells grown in vitro. Mouse lungs were decellularized using 0.2 % sodium dodecyl sulphate, hypotonic solutions, and DNase. Morphological analysis of the resulting ECM scaffolds was performed by means of transmission and scanning electron microscopy. The ECM scaffolds retained 3D architecture of the main lung anatomical regions: the alveolar region and the blood/ airway network. The region-specific ECM nanotopology and organization of ECM macromolecules such as fibres of collagen, elastin, and fibrillin were characterized. The lung ECM scaffolds were also homogenized and applied as a supplement to growth medium on primary lung cells in vitro to test the bioact...
Checkpoint-mediated dependency of tumor cells can be deployed to selectively kill them without su... more Checkpoint-mediated dependency of tumor cells can be deployed to selectively kill them without substantial toxicity to normal cells. Specifically, loss of CHK1, a serine threonine kinase involved in the surveillance of the G2/M checkpoint in the presence of replication stress inflicted by DNA damaging drugs, has been reported to dramatically influence the viability of tumor cells. CHK1's pivotal role in maintaining genomic stability offers attractive opportunity for increasing the selectivity, effectivity and reduced toxicity of chemotherapy. Some recently identified CHK1 inhibitors entered clinical trials in combination with DNA antimetabolites. Herein, we report synthesis and profiling of MU380, a non-trivial analog of clinically profiled compound SCH900776 possessing the highly unusual N-trifluoromethylpyrazole motif, which was envisioned not to undergo metabolic oxidative dealkylation and thereby provide greater robustness to the compound. MU380 is a selective and potent inh...
During heart morphogenesis, cardiac chambers arise by differential expansion of regions of the pr... more During heart morphogenesis, cardiac chambers arise by differential expansion of regions of the primitive cardiac tube. This process is under the control of specific transcription factors such as Tbx5 and dHAND. To gain insight into the cellular mechanisms that underlie cardiogenesis, we have used a retrospective clonal approach based on the spontaneous recombination of an nlaacZ reporter gene targeted to the murine alpha-cardiac actin locus. We show that clonal growth of myocardial cells is oriented. At embryonic day (E) 10.5, the shape of clones is characteristic of a given cardiac region and reflects its morphology. This is already detectable in the primitive cardiac tube at E8.5, and is maintained after septation at E14.5 with additional modulations. The clonal analysis reveals new subdivisions of the myocardium, including an interventricular boundary region. Our results show that the myocardium, from the time of its formation, is a polarized and regionalized tissue and point to ...
The delivery of newly synthesized soluble lysosomal hydrolases to the endosomal system is essenti... more The delivery of newly synthesized soluble lysosomal hydrolases to the endosomal system is essential for lysosome function and cell homeostasis. This process relies on the proper trafficking of the mannose 6-phosphate receptors (MPRs) between the trans-Golgi network, endosomes and the plasma membrane. Many transmembrane proteins regulating diverse biological processes ranging from virus production to the development of multi-cellular organisms also use these pathways. To explore how cell signaling modulates MPR trafficking, we used high throughput RNA interference to target the human kinome and phosphatome. Using high content image analysis, we identified 127 kinases and phosphatases belonging to different signaling networks that regulate MPR trafficking and/or the dynamic states of the subcellular compartments encountered by the MPRs. Our analysis maps the MPR trafficking pathways based on enzymes regulating phosphatidylinositol phosphate metabolism. Furthermore, it reveals how cell...
In both embryonal carcinoma (EC) and embryonic stem (ES) cells, the differentiation pathway enter... more In both embryonal carcinoma (EC) and embryonic stem (ES) cells, the differentiation pathway entered after treatment with retinoic acid (RA) varies as it is based upon different conditions of culture. This study employs mouse EC cells P19 to investigate the effects of serum on RA-induced neural differentiation occurring in a simplified monolayer culture. Cell morphology and expression of lineage-specific molecular markers document that, while non-neural cell types arise after treatment with RA under serum-containing conditions, in chemically defined serum-free media RA induces massive neural differentiation in concentrations of 10(-9) M and higher. Moreover, not only neural (Mash-1) and neuroectodermal (Pax-6), but also endodermal (GATA-4, alpha-fetoprotein) genes are expressed at early stages of differentiation driven by RA under serum-free conditions. Furthermore, as determined by the luciferase reporter assay, the presence or absence of the serum does not affect the activity of th...
In both embryonal carcinoma (EC) and embryonic stem (ES) cells, the differentiation pathway enter... more In both embryonal carcinoma (EC) and embryonic stem (ES) cells, the differentiation pathway entered after treatment with retinoic acid (RA) varies as it is based upon different conditions of culture. This study employs mouse EC cells P19 to investigate the effects of serum on RA-induced neural differentiation occurring in a simplified monolayer culture. Cell morphology and expression of lineage-specific molecular markers document that, while non-neural cell types arise after treatment with RA under serum-containing conditions, in chemically defined serum-free media RA induces massive neural differentiation in concentrations of 10(-9) M and higher. Moreover, not only neural (Mash-1) and neuroectodermal (Pax-6), but also endodermal (GATA-4, alpha-fetoprotein) genes are expressed at early stages of differentiation driven by RA under serum-free conditions. Furthermore, as determined by the luciferase reporter assay, the presence or absence of the serum does not affect the activity of th...
Extracellular matrix (ECM) forms an essential part of tissue microenvironment. Compositional and ... more Extracellular matrix (ECM) forms an essential part of tissue microenvironment. Compositional and structural properties of ECM modulate behaviour of cells, including their differentiation, proliferation, and turnover. Here we aimed at detailed characterization of the nanostructure of lung ECM and evaluation of its bioactivity on cells grown in vitro. Mouse lungs were decellularized using 0.2 % sodium dodecyl sulphate, hypotonic solutions, and DNase. Morphological analysis of the resulting ECM scaffolds was performed by means of transmission and scanning electron microscopy. The ECM scaffolds retained 3D architecture of the main lung anatomical regions: the alveolar region and the blood/ airway network. The region-specific ECM nanotopology and organization of ECM macromolecules such as fibres of collagen, elastin, and fibrillin were characterized. The lung ECM scaffolds were also homogenized and applied as a supplement to growth medium on primary lung cells in vitro to test the bioact...
Extracellular matrix (ECM) forms an essential part of tissue microenvironment. Compositional and ... more Extracellular matrix (ECM) forms an essential part of tissue microenvironment. Compositional and structural properties of ECM modulate behaviour of cells, including their differentiation, proliferation, and turnover. Here we aimed at detailed characterization of the nanostructure of lung ECM and evaluation of its bioactivity on cells grown in vitro. Mouse lungs were decellularized using 0.2 % sodium dodecyl sulphate, hypotonic solutions, and DNase. Morphological analysis of the resulting ECM scaffolds was performed by means of transmission and scanning electron microscopy. The ECM scaffolds retained 3D architecture of the main lung anatomical regions: the alveolar region and the blood/ airway network. The region-specific ECM nanotopology and organization of ECM macromolecules such as fibres of collagen, elastin, and fibrillin were characterized. The lung ECM scaffolds were also homogenized and applied as a supplement to growth medium on primary lung cells in vitro to test the bioact...
Checkpoint-mediated dependency of tumor cells can be deployed to selectively kill them without su... more Checkpoint-mediated dependency of tumor cells can be deployed to selectively kill them without substantial toxicity to normal cells. Specifically, loss of CHK1, a serine threonine kinase involved in the surveillance of the G2/M checkpoint in the presence of replication stress inflicted by DNA damaging drugs, has been reported to dramatically influence the viability of tumor cells. CHK1's pivotal role in maintaining genomic stability offers attractive opportunity for increasing the selectivity, effectivity and reduced toxicity of chemotherapy. Some recently identified CHK1 inhibitors entered clinical trials in combination with DNA antimetabolites. Herein, we report synthesis and profiling of MU380, a non-trivial analog of clinically profiled compound SCH900776 possessing the highly unusual N-trifluoromethylpyrazole motif, which was envisioned not to undergo metabolic oxidative dealkylation and thereby provide greater robustness to the compound. MU380 is a selective and potent inh...
During heart morphogenesis, cardiac chambers arise by differential expansion of regions of the pr... more During heart morphogenesis, cardiac chambers arise by differential expansion of regions of the primitive cardiac tube. This process is under the control of specific transcription factors such as Tbx5 and dHAND. To gain insight into the cellular mechanisms that underlie cardiogenesis, we have used a retrospective clonal approach based on the spontaneous recombination of an nlaacZ reporter gene targeted to the murine alpha-cardiac actin locus. We show that clonal growth of myocardial cells is oriented. At embryonic day (E) 10.5, the shape of clones is characteristic of a given cardiac region and reflects its morphology. This is already detectable in the primitive cardiac tube at E8.5, and is maintained after septation at E14.5 with additional modulations. The clonal analysis reveals new subdivisions of the myocardium, including an interventricular boundary region. Our results show that the myocardium, from the time of its formation, is a polarized and regionalized tissue and point to ...
The delivery of newly synthesized soluble lysosomal hydrolases to the endosomal system is essenti... more The delivery of newly synthesized soluble lysosomal hydrolases to the endosomal system is essential for lysosome function and cell homeostasis. This process relies on the proper trafficking of the mannose 6-phosphate receptors (MPRs) between the trans-Golgi network, endosomes and the plasma membrane. Many transmembrane proteins regulating diverse biological processes ranging from virus production to the development of multi-cellular organisms also use these pathways. To explore how cell signaling modulates MPR trafficking, we used high throughput RNA interference to target the human kinome and phosphatome. Using high content image analysis, we identified 127 kinases and phosphatases belonging to different signaling networks that regulate MPR trafficking and/or the dynamic states of the subcellular compartments encountered by the MPRs. Our analysis maps the MPR trafficking pathways based on enzymes regulating phosphatidylinositol phosphate metabolism. Furthermore, it reveals how cell...
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Papers by Milan Esner