Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 50
Filtrar
Mais filtros

Base de dados
Tipo de documento
Intervalo de ano de publicação
1.
Cell ; 162(2): 403-411, 2015 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-26165941

RESUMO

Small molecules that interfere with microtubule dynamics, such as Taxol and the Vinca alkaloids, are widely used in cell biology research and as clinical anticancer drugs. However, their activity cannot be restricted to specific target cells, which also causes severe side effects in chemotherapy. Here, we introduce the photostatins, inhibitors that can be switched on and off in vivo by visible light, to optically control microtubule dynamics. Photostatins modulate microtubule dynamics with a subsecond response time and control mitosis in living organisms with single-cell spatial precision. In longer-term applications in cell culture, photostatins are up to 250 times more cytotoxic when switched on with blue light than when kept in the dark. Therefore, photostatins are both valuable tools for cell biology, and are promising as a new class of precision chemotherapeutics whose toxicity may be spatiotemporally constrained using light.


Assuntos
Antimitóticos/química , Morte Celular , Microtúbulos/efeitos dos fármacos , Mitose , Estilbenos/química , Animais , Antimitóticos/toxicidade , Linhagem Celular Tumoral , Citoesqueleto/química , Humanos , Luz , Camundongos , Polimerização , Estilbenos/toxicidade
2.
Bioessays ; 45(4): e2200186, 2023 04.
Artigo em Inglês | MEDLINE | ID: mdl-36871153

RESUMO

Unraveling molecular and functional heterogeneity of niche cells within the developing endoderm could resolve mechanisms of tissue formation and maturation. Here, we discuss current unknowns in molecular mechanisms underlying key developmental events in pancreatic islet and intestinal epithelial formation. Recent breakthroughs in single-cell and spatial transcriptomics, paralleled with functional studies in vitro, reveal that specialized mesenchymal subtypes drive the formation and maturation of pancreatic endocrine cells and islets via local interactions with epithelium, neurons, and microvessels. Analogous to this, distinct intestinal niche cells regulate both epithelial development and homeostasis throughout life. We propose how this knowledge can be used to progress research in the human context using pluripotent stem cell-derived multilineage organoids. Overall, understanding the interactions between the multitude of microenvironmental cells and how they drive tissue development and function could help us make more therapeutically relevant in vitro models.


Assuntos
Endoderma , Pâncreas , Humanos , Diferenciação Celular/fisiologia , Homeostase , Intestinos
3.
Cell Mol Life Sci ; 80(11): 335, 2023 Oct 26.
Artigo em Inglês | MEDLINE | ID: mdl-37882878

RESUMO

Muscleblind-like splicing regulators (MBNLs) activate or repress the inclusion of alternative splicing (AS) events, enabling the developmental transition of fetal mRNA splicing isoforms to their adult forms. Herein, we sought to elaborate the mechanism by which MBNLs mediate AS related to biological processes. We evaluated the functional role of DEAD-box (DDX) RNA helicases, DDX5 and DDX17 in MBNL-dependent AS regulation. Whole-transcriptome analysis and validation approaches revealed a handful of MBNLs-dependent AS events to be affected by DDX5 and DDX17 in mostly an opposite manner. The opposite expression patterns of these two groups of factors during muscle development and coordination of fetal-to-adult splicing transition indicate the importance of these proteins at early stages of development. The identified pathways of how the helicases modulate MBNL splicing activity include DDX5 and DDX17-dependent changes in the ratio of MBNL splicing isoforms and most likely changes in accessibility of MBNL-binding sites. Another pathway involves the mode of action of the helicases independent of MBNL activity. These findings lead to a deeper understanding of the network of interdependencies between RNA-binding proteins and constitute a valuable element in the discussion on developmental homeostasis and pathological states in which the studied protein factors play a significant role.


Assuntos
Processamento Alternativo , RNA Helicases , Processamento Alternativo/genética , RNA Helicases/genética , Splicing de RNA , Isoformas de Proteínas/genética , Sítios de Ligação/genética
4.
Eur J Immunol ; 52(7): 1158-1170, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35389516

RESUMO

The contribution of low-affinity T cells to autoimmunity in the context of polyclonal T-cell responses is understudied due to the limitations in their capture by tetrameric reagents and low level of activation in response to antigenic stimulation. As a result, low-affinity T cells are often disregarded as nonantigen-specific cells irrelevant to the immune response. Our study aimed to assess how the level of self-antigen reactivity shapes T-cell lineage and effector responses in the context of spontaneous tissue-specific autoimmunity observed in NOD mice. Using multicolor flow cytometry in combination with Nur77GFP reporter of TCR signaling, we identified a dormant population of T cells that infiltrated the pancreatic islets of prediabetic NOD mice, which exhibited reduced levels of self-tissue reactivity based on expression of CD5 and Nur77GFP . We showed that these CD5low T cells had a unique TCR repertoire and exhibited low activation and minimal effector function; however, induced rapid diabetes upon transfer. The CD4+ CD5low T-cell population displayed transcriptional signature of central memory T cells, consistent with the ability to acquire effector function post-transfer. Transcriptional profile of CD5low T cells was similar to T cells expressing a low-affinity TCR, indicating TCR affinity to be an important factor in shaping CD5low T-cell phenotype and function at the tissue site. Overall, our study suggests that autoimmune tissue can maintain a reservoir of undifferentiated central memory-like autoreactive T cells with pathogenic effector potential that might be an important source for effector T cells during long-term chronic autoimmunity.


Assuntos
Diabetes Mellitus Tipo 1 , Animais , Linfócitos T CD4-Positivos , Camundongos , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T/genética
5.
Angew Chem Int Ed Engl ; 61(48): e202210220, 2022 11 25.
Artigo em Inglês | MEDLINE | ID: mdl-36048143

RESUMO

The natural product jasplakinolide is widely used to stabilize F-actin. Based on extensive structure-activity relationship studies, we have developed a new generation of photoswitchable jasplakinolides that feature rationally designed red-shifted azobenzene photoswitches. Our lead compound, nOJ, can be activated with longer wavelengths in the visible range (e.g. 440-475 nm) and rapidly returns to its inactive state through thermal relaxation. nOJ enables the reversible control of F-actin dynamics, as shown through live-cell imaging, cell migration, and cell proliferation assays. Short, local irradiation with blue light resulted in highly localized and reversible actin aggregation with subcellular precision. Our optical tool can be useful in diverse fields to study actin dynamics with excellent spatiotemporal resolution.


Assuntos
Actinas , Depsipeptídeos , Citoesqueleto de Actina , Depsipeptídeos/farmacologia , Movimento Celular
6.
J Am Chem Soc ; 142(20): 9240-9249, 2020 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-32388980

RESUMO

Cell-permeable photoswitchable small molecules, termed optojasps, are introduced to optically control the dynamics of the actin cytoskeleton and cellular functions that depend on it. These light-dependent effectors were designed from the F-actin-stabilizing marine depsipeptide jasplakinolide by functionalizing them with azobenzene photoswitches. As demonstrated, optojasps can be employed to control cell viability, cell motility, and cytoskeletal signaling with the high spatial and temporal resolution that light affords. Optojasps can be expected to find applications in diverse areas of cell biological research. They may also provide a template for photopharmacology targeting the ubiquitous actin cytoskeleton with precision control in the micrometer range.


Assuntos
Actinas/química , Compostos Azo/química , Depsipeptídeos/química , Bibliotecas de Moléculas Pequenas/química , Compostos Azo/síntese química , Conformação Molecular , Processos Fotoquímicos , Bibliotecas de Moléculas Pequenas/síntese química
7.
J Immunol ; 200(3): 909-914, 2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29282307

RESUMO

Regulatory T cells (Tregs) use a distinct TCR repertoire and are more self-reactive compared with conventional T cells. However, the extent to which TCR affinity regulates the function of self-reactive Tregs is largely unknown. In this study, we used a two-TCR model to assess the role of TCR affinity in Treg function during autoimmunity. We observed that high- and low-affinity Tregs were recruited to the pancreas and contributed to protection from autoimmune diabetes. Interestingly, high-affinity cells preferentially upregulated the TCR-dependent Treg functional mediators IL-10, TIGIT, GITR, and CTLA4, whereas low-affinity cells displayed increased transcripts for Areg and Ebi3, suggesting distinct functional profiles. The results of this study suggest mechanistically distinct and potentially nonredundant roles for high- and low-affinity Tregs in controlling autoimmunity.


Assuntos
Autoimunidade/imunologia , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/prevenção & controle , Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Anfirregulina/biossíntese , Animais , Antígeno CTLA-4/biossíntese , Adesão Celular/imunologia , Proteína Relacionada a TNFR Induzida por Glucocorticoide/biossíntese , Interleucina-10/biossíntese , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Antígenos de Histocompatibilidade Menor/biossíntese , Pâncreas/citologia , Pâncreas/imunologia , Receptores de Citocinas/biossíntese , Receptores Imunológicos/biossíntese
8.
Nature ; 491(7426): 765-8, 2012 Nov 29.
Artigo em Inglês | MEDLINE | ID: mdl-23041930

RESUMO

One goal of regenerative medicine, to use stem cells to replace cells lost by injury or disease, depends on producing an excess of the relevant cell for study or transplantation. To this end, the stepwise differentiation of stem cells into specialized derivatives has been successful for some cell types, but a major problem remains the inefficient conversion of cells from one stage of differentiation to the next. If specialized cells are to be produced in large numbers it will be necessary to expand progenitor cells, without differentiation, at some steps of the process. Using the pancreatic lineage as a model for embryonic-stem-cell differentiation, we demonstrate that this is a solvable problem. Co-culture with organ-matched mesenchyme permits proliferation and self-renewal of progenitors, without differentiation, and enables an expansion of more than a million-fold for human endodermal cells with full retention of their developmental potential. This effect is specific both to the mesenchymal cell and to the progenitor being amplified. Progenitors that have been serially expanded on mesenchyme give rise to glucose-sensing, insulin-secreting cells when transplanted in vivo. Theoretically, the identification of stage-specific renewal signals can be incorporated into any scheme for the efficient production of large numbers of differentiated cells from stem cells and may therefore have wide application in regenerative biology.


Assuntos
Técnicas de Cocultura/métodos , Células-Tronco Embrionárias/citologia , Endoderma/citologia , Mesoderma/citologia , Pâncreas/citologia , Animais , Diferenciação Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Células-Tronco Embrionárias/efeitos dos fármacos , Células-Tronco Embrionárias/metabolismo , Endoderma/efeitos dos fármacos , Endoderma/metabolismo , Glucose/farmacologia , Humanos , Insulina/metabolismo , Masculino , Camundongos , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/efeitos dos fármacos , Células-Tronco Pluripotentes/metabolismo
9.
J Hepatol ; 65(2): 325-33, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27117591

RESUMO

BACKGROUND & AIMS: Pediatric liver cancer is a rare but serious disease whose incidence is rising, and for which the therapeutic options are limited. Development of more targeted, less toxic therapies is hindered by the lack of an experimental animal model that captures the heterogeneity and metastatic capability of these tumors. METHODS: Here we established an orthotopic engraftment technique to model a series of patient-derived tumor xenograft (PDTX) from pediatric liver cancers of all major histologic subtypes: hepatoblastoma, hepatocellular cancer and hepatocellular malignant neoplasm. We utilized standard (immuno) staining methods for histological characterization, RNA sequencing for gene expression profiling and genome sequencing for identification of druggable targets. We also adapted stem cell culturing techniques to derive two new pediatric cancer cell lines from the xenografted mice. RESULTS: The patient-derived tumor xenografts recapitulated the histologic, genetic, and biological characteristics-including the metastatic behavior-of the corresponding primary tumors. Furthermore, the gene expression profiles of the two new liver cancer cell lines closely resemble those of the primary tumors. Targeted therapy of PDTX from an aggressive hepatocellular malignant neoplasm with the MEK1 inhibitor trametinib and pan-class I PI3 kinase inhibitor NVP-BKM120 resulted in significant growth inhibition, thus confirming this PDTX model as a valuable tool to study tumor biology and patient-specific therapeutic responses. CONCLUSIONS: The novel metastatic xenograft model and the isogenic xenograft-derived cell lines described in this study provide reliable tools for developing mutation- and patient-specific therapies for pediatric liver cancer. LAY SUMMARY: Pediatric liver cancer is a rare but serious disease and no experimental animal model currently captures the complexity and metastatic capability of these tumors. We have established a novel animal model using human tumor tissue that recapitulates the genetic and biological characteristics of this cancer. We demonstrate that our patient-derived animal model, as well as two new cell lines, are useful tools for experimental therapies.


Assuntos
Neoplasias Hepáticas , Animais , Carcinoma Hepatocelular , Linhagem Celular Tumoral , Criança , Modelos Animais de Doenças , Xenoenxertos , Humanos , Camundongos , Transplante de Neoplasias , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Development ; 140(3): 675-86, 2013 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-23293299

RESUMO

Embryonic stem (ES) cells hold great promise with respect to their potential to be differentiated into desired cell types. Of interest are organs derived from the definitive endoderm, such as the pancreas and liver, and animal studies have revealed an essential role for Nodal in development of the definitive endoderm. Activin A is a related TGFß member that acts through many of the same downstream signaling effectors as Nodal and is thought to mimic Nodal activity. Detailed characterization of ES cell-derived endodermal cell types by gene expression analysis in vitro and functional analysis in vivo reveal that, despite their similarity in gene expression, Nodal and Activin-derived endodermal cells exhibit a distinct difference in functional competence following transplantation into the developing mouse embryo. Pdx1-expressing cells arising from the respective endoderm populations exhibit extended differences in their competence to mature into insulin/c-peptide-expressing cells in vivo. Our findings underscore the importance of functional cell-type evaluation during stepwise differentiation of stem cells.


Assuntos
Células-Tronco Embrionárias/metabolismo , Endoderma/citologia , Proteína Nodal/metabolismo , Animais , Técnicas de Cultura de Células/métodos , Diferenciação Celular , Células Cultivadas , Meios de Cultura/metabolismo , Técnicas de Cultura Embrionária , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/efeitos dos fármacos , Endoderma/metabolismo , Feminino , Imunofluorescência , Perfilação da Expressão Gênica , Proteínas de Fluorescência Verde/metabolismo , Proteínas HMGB/genética , Proteínas HMGB/metabolismo , Humanos , Subunidades beta de Inibinas/metabolismo , Subunidades beta de Inibinas/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Proteína Nodal/farmacologia , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia , Fatores de Transcrição SOXF/genética , Fatores de Transcrição SOXF/metabolismo
11.
J Virol ; 88(16): 8956-70, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24872589

RESUMO

Human T-cell leukemia virus types 3 and 4 (HTLV-3 and HTLV-4) are recently isolated retroviruses. We have previously characterized HTLV-3- and HTLV-4-encoded antisense genes, termed APH-3 and APH-4, respectively, which, in contrast to HBZ, the HTLV-1 homologue, do not contain a typical bZIP domain (M. Larocque É Halin, S. Landry, S. J. Marriott, W. M. Switzer, and B. Barbeau, J. Virol. 85:12673-12685, 2011, doi:10.1128/JVI.05296-11). As HBZ differentially modulates the transactivation potential of various Jun family members, the effect of APH-3 and APH-4 on JunD-, c-Jun-, and JunB-mediated transcriptional activation was investigated. We first showed that APH-3 and APH-4 upregulated the transactivation potential of all tested Jun family members. Using an human telomerase catalytic subunit (hTERT) promoter construct, our results also highlighted that, unlike HBZ, which solely modulates hTERT expression via JunD, both APH-3 and APH-4 acted positively on the transactivation of the hTERT promoter mediated by tested Jun factors. Coimmunoprecipitation experiments demonstrated that these Jun proteins interacted with APH-3 and APH-4. Although no activation domain was identified for APH proteins, the activation domain of c-Jun was very important in the observed upregulation of its activation potential. We further showed that APH-3 and APH-4 required their putative bZIP-like domains and corresponding leucine residues for interaction and modulation of the transactivation potential of Jun factors. Our results demonstrate that HTLV-encoded antisense proteins behave differently, and that the bZIP-like domains of both APH-3 and APH-4 have retained their interaction potential for Jun members. These studies are important in assessing the differences between HBZ and other antisense proteins, which might further contribute to determining the role of HBZ in HTLV-1-associated diseases. IMPORTANCE HBZ, the antisense transcript-encoded protein from HTLV-1, is now well recognized as a potential factor for adult T-cell leukemia/lymphoma development. In order to better appreciate the mechanism of action of HBZ, comparison to antisense proteins from other HTLV viruses is important. Little is known in relation to the seemingly nonpathogenic HTLV-3 and HTLV-4 viruses, and studies of their antisense proteins are limited to our previously reported study (M. Larocque É Halin, S. Landry, S. J. Marriott, W. M. Switzer, and B. Barbeau, J. Virol. 85:12673-12685, 2011, doi:10.1128/JVI.05296-11). Here, we demonstrate that Jun transcription factors are differently affected by APH-3 and APH-4 compared to HBZ. These intriguing findings suggest that these proteins act differently on viral replication but also on cellular gene expression, and that highlighting their differences of action might lead to important information allowing us to understand the link between HTLV-1 HBZ and ATL in infected individuals.


Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , DNA Antissenso/genética , Vírus Linfotrópico T Tipo 3 Humano/genética , Vírus Linfotrópico T Tipo 3 Humano/metabolismo , Proteínas Proto-Oncogênicas c-jun/metabolismo , Ativação Transcricional/genética , Animais , Células COS , Linhagem Celular , Linhagem Celular Tumoral , Chlorocebus aethiops , Deltaretrovirus/genética , Deltaretrovirus/metabolismo , Células HEK293 , Células HeLa , Humanos , Regiões Promotoras Genéticas/genética , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas c-jun/genética , Telomerase/genética , Telomerase/metabolismo , Transcrição Gênica/genética , Regulação para Cima/genética , Proteínas Virais/genética , Proteínas Virais/metabolismo
12.
Carcinogenesis ; 34(11): 2664-72, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23784080

RESUMO

Telomerase activity in cancer cells is dependent on the transcriptional regulation of the human telomerase reverse transcriptase (hTERT) gene, encoding the catalytic subunit of human telomerase. We have shown previously that HTLV-1 basic leucine zipper (HBZ), a viral regulatory protein encoded by the human retrovirus, human T-cell leukemia virus, type 1 (HTLV-1) cooperates with JunD to enhance hTERT transcription in adult T-cell leukemia (ATL) cells. Menin, the product of the tumor-suppressor MEN-1 gene, also interacts with JunD, represses its transcriptional activity and downregulates telomerase expression. The main objective of this study was to examine how menin and HBZ get involved in the regulation of hTERT transcription. In this study, we report that JunD and menin form a repressor complex of hTERT transcription in HBZ-negative cells. Conversely, in HBZ-positive cells, the formation of a JunD/HBZ/menin ternary complex and the recruitment of p300 histone acetyl transferase activity by HBZ lead to a decreased activity of the JunD-menin suppressor unit that correlates with the activation of hTERT transcription. Silencing HBZ or menin expression in ATL cells confirms that these proteins are differentially involved in telomerase regulation. These results propose that HBZ, by impeding the tumor-suppressor activity of menin, functions as a leukemogenic cofactor to upregulate gene transcription and promote JunD-mediated leukemogenesis.


Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Proteína p300 Associada a E1A/metabolismo , Regulação Neoplásica da Expressão Gênica , Leucemia de Células T/patologia , Proteínas Proto-Oncogênicas c-jun/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Telomerase/genética , Proteínas Virais/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/antagonistas & inibidores , Fatores de Transcrição de Zíper de Leucina Básica/genética , Western Blotting , Proliferação de Células , Imunoprecipitação da Cromatina , Proteína p300 Associada a E1A/genética , Células HeLa , Humanos , Imunoprecipitação , Leucemia de Células T/genética , Leucemia de Células T/metabolismo , Regiões Promotoras Genéticas/genética , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-jun/genética , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Proteínas dos Retroviridae , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Telomerase/metabolismo , Transcrição Gênica , Células Tumorais Cultivadas , Proteínas Virais/antagonistas & inibidores , Proteínas Virais/genética
13.
bioRxiv ; 2023 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-37808662

RESUMO

Cell proliferation is a ubiquitous process required for organismal development and homeostasis. However, individuals with partial loss-of-function variants in DNA replicative helicase components often present with immunodeficiency due to specific loss of natural killer (NK) cells. Such lineage-specific disease phenotypes raise questions on how the proliferation is regulated in cell type-specific manner. We aimed to understand NK cell-specific proliferative dynamics and vulnerability to impaired helicase function using iPSCs from individuals with NK cell deficiency (NKD) due to hereditary compound heterozygous GINS4 variants. We observed and characterized heterogeneous cell populations that arise during the iPSC differentiation along with NK cells. While overall cell proliferation decreased with differentiation, early NK cell precursors showed a short burst of cell proliferation. GINS4 deficiency induced replication stress in these early NK cell precursors, which are poised for apoptosis, and ultimately recapitulate the NKD phenotype.

14.
Biomolecules ; 13(1)2023 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-36671537

RESUMO

Apart from chaperoning, disulfide bond formation, and downstream processing, the molecular sequence of proinsulin folding is not completely understood. Proinsulin requires proline isomerization for correct folding. Since FK506-binding protein 2 (FKBP2) is an ER-resident proline isomerase, we hypothesized that FKBP2 contributes to proinsulin folding. We found that FKBP2 co-immunoprecipitated with proinsulin and its chaperone GRP94 and that inhibition of FKBP2 expression increased proinsulin turnover with reduced intracellular proinsulin and insulin levels. This phenotype was accompanied by an increased proinsulin secretion and the formation of proinsulin high-molecular-weight complexes, a sign of proinsulin misfolding. FKBP2 knockout in pancreatic ß-cells increased apoptosis without detectable up-regulation of ER stress response genes. Interestingly, FKBP2 mRNA was overexpressed in ß-cells from pancreatic islets of T2D patients. Based on molecular modeling and an in vitro enzymatic assay, we suggest that proline at position 28 of the proinsulin B-chain (P28) is the substrate of FKBP2's isomerization activity. We propose that this isomerization step catalyzed by FKBP2 is an essential sequence required for correct proinsulin folding.


Assuntos
Células Secretoras de Insulina , Proinsulina , Proinsulina/metabolismo , Dobramento de Proteína , Retículo Endoplasmático/metabolismo , Células Secretoras de Insulina/metabolismo , Chaperonas Moleculares/metabolismo , Prolina/metabolismo , Proteínas de Ligação a Tacrolimo/genética , Proteínas de Ligação a Tacrolimo/metabolismo , Insulina/metabolismo
15.
J Cell Biol ; 177(1): 151-62, 2007 Apr 09.
Artigo em Inglês | MEDLINE | ID: mdl-17403932

RESUMO

Wound healing of the skin is a crucial regenerative process in adult mammals. We examined wound healing in conditional mutant mice, in which the c-Met gene that encodes the receptor of hepatocyte growth factor/scatter factor was mutated in the epidermis by cre recombinase. c-Met-deficient keratinocytes were unable to contribute to the reepithelialization of skin wounds. In conditional c-Met mutant mice, wound closure was slightly attenuated, but occurred exclusively by a few (5%) keratinocytes that had escaped recombination. This demonstrates that the wound process selected and amplified residual cells that express a functional c-Met receptor. We also cultured primary keratinocytes from the skin of conditional c-Met mutant mice and examined them in scratch wound assays. Again, closure of scratch wounds occurred by the few remaining c-Met-positive cells. Our data show that c-Met signaling not only controls cell growth and migration during embryogenesis but is also essential for the generation of the hyperproliferative epithelium in skin wounds, and thus for a fundamental regenerative process in the adult.


Assuntos
Proteínas Proto-Oncogênicas c-met/fisiologia , Fenômenos Fisiológicos da Pele , Cicatrização/fisiologia , Animais , Comunicação Autócrina , Células Cultivadas , Fator de Crescimento de Hepatócito/metabolismo , Integrases/metabolismo , Queratinócitos/citologia , Queratinócitos/metabolismo , Queratinócitos/fisiologia , Camundongos , Camundongos Endogâmicos , Mutação , Proteínas Proto-Oncogênicas c-met/genética , Transdução de Sinais , Cicatrização/genética
16.
Front Cell Dev Biol ; 10: 913305, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35769263

RESUMO

SARS-CoV-2, a newly emerged virus described for the first time in late 2019, affects multiple organs in humans, including the pancreas. Here, we present the bilateral link between the pathophysiology of diabetes and COVID-19, with diabetes being COVID-19 comorbidity, and a complication of SARS-CoV-2 infection. Analysis of clinical data indicates that patients with chronic conditions like diabetes are at increased risk of severe COVID-19, hospitalization, ICU admission, and death compared to the healthy subjects. Further, we show that SARS-CoV-2 infection might be also associated with the development of new-onset diabetes and diabetic ketoacidosis. We then discuss the options for studying SARS-CoV-2 infection in pancreatic settings, including the use of human pluripotent stem cell-derived pancreatic organoids. Further, we review the presence of SARS-CoV-2 receptors in different pancreatic cell types and the infection efficiency based on pancreatic sections from COVID-19 patients and primary human islet in vitro studies. Finally, we discuss the impact of SARS-CoV-2 infection on human pancreatic cell homeostasis, focusing on ß-cells.

17.
Cells ; 11(10)2022 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-35626685

RESUMO

SMER28 (Small molecule enhancer of Rapamycin 28) is an autophagy-inducing compound functioning by a hitherto unknown mechanism. Here, we confirm its autophagy-inducing effect by assessing classical autophagy-related parameters. Interestingly, we also discovered several additional effects of SMER28, including growth retardation and reduced G1 to S phase progression. Most strikingly, SMER28 treatment led to a complete arrest of receptor tyrosine kinase signaling, and, consequently, growth factor-induced cell scattering and dorsal ruffle formation. This coincided with a dramatic reduction in phosphorylation patterns of PI3K downstream effectors. Consistently, SMER28 directly inhibited PI3Kδ and to a lesser extent p110γ. The biological relevance of our observations was underscored by SMER28 interfering with InlB-mediated host cell entry of Listeria monocytogenes, which requires signaling through the prominent receptor tyrosine kinase c-Met. This effect was signaling-specific, since entry of unrelated, gram-negative Salmonella Typhimurium was not inhibited. Lastly, in B cell lymphoma cells, which predominantly depend on tonic signaling through PI3Kδ, apoptosis upon SMER28 treatment is profound in comparison to non-hematopoietic cells. This indicates SMER28 as a possible drug candidate for the treatment of diseases that derive from aberrant PI3Kδ activity.


Assuntos
Fosfatidilinositol 3-Quinases , Serina-Treonina Quinases TOR , Autofagia , Fosfatidilinositol 3-Quinases/metabolismo , Receptores Proteína Tirosina Quinases , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo
18.
iScience ; 25(7): 104594, 2022 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-35756892

RESUMO

Recent studies showed that SARS-CoV-2 can infect adult human pancreas and trigger pancreatic damage. Here, using human fetal pancreas samples and 3D differentiation of human pluripotent cells into pancreatic endocrine cells, we determined that SARS-CoV-2 receptors ACE2, TMPRSS2, and NRP1 are expressed in precursors of insulin-producing pancreatic ß-cells, rendering them permissive to SARS-CoV-2 infection. We also show that SARS-CoV-2 enters and undergoes efficient replication in human multipotent pancreatic and endocrine progenitors in vitro. Moreover, we investigated mechanisms by which SARS-CoV-2 enters pancreatic cells, and found that ACE2 mediates the entry, while NRP1 and TMPRSS2 do not. Surprisingly, we found that in pancreatic progenitors, SARS-CoV-2 enters cells via cathepsin-dependent endocytosis, which is a different route than in respiratory tract. Therefore, pancreatic spheroids might serve as a model to study candidate drugs for endocytosis-mediated viral entry inhibition and to investigate whether SARS-CoV-2 infection may affect pancreas development, possibly causing lifelong health consequences.

19.
Nat Commun ; 13(1): 1952, 2022 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-35414140

RESUMO

In vitro derivation of pancreatic ß-cells from human pluripotent stem cells holds promise as diabetes treatment. Despite recent progress, efforts to generate physiologically competent ß-cells are still hindered by incomplete understanding of the microenvironment's role in ß-cell development and maturation. Here, we analyze the human mesenchymal and endothelial primary cells from weeks 9-20 fetal pancreas and identify a time point-specific microenvironment that permits ß-cell differentiation. Further, we uncover unique factors that guide in vitro development of endocrine progenitors, with WNT5A markedly improving human ß-cell differentiation. WNT5A initially acts through the non-canonical (JNK/c-JUN) WNT signaling and cooperates with Gremlin1 to inhibit the BMP pathway during ß-cell maturation. Interestingly, we also identify the endothelial-derived Endocan as a SST+ cell promoting factor. Overall, our study shows that the pancreatic microenvironment-derived factors can mimic in vivo conditions in an in vitro system to generate bona fide ß-cells for translational applications.


Assuntos
Pâncreas , Via de Sinalização Wnt , Proteínas Morfogenéticas Ósseas/metabolismo , Diferenciação Celular , Humanos , MAP Quinase Quinase 4/metabolismo , Pâncreas/metabolismo , Via de Sinalização Wnt/fisiologia , Proteína Wnt-5a/genética , Proteína Wnt-5a/metabolismo
20.
Nat Chem Biol ; 5(4): 258-65, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19287398

RESUMO

Stepwise differentiation from embryonic stem cells (ESCs) to functional insulin-secreting beta cells will identify key steps in beta-cell development and may yet prove useful for transplantation therapy for diabetics. An essential step in this schema is the generation of pancreatic progenitors--cells that express Pdx1 and produce all the cell types of the pancreas. High-content chemical screening identified a small molecule, (-)-indolactam V, that induces differentiation of a substantial number of Pdx1-expressing cells from human ESCs. The Pdx1-expressing cells express other pancreatic markers and contribute to endocrine, exocrine and duct cells, in vitro and in vivo. Further analyses showed that (-)-indolactam V works specifically at one stage of pancreatic development, inducing pancreatic progenitors from definitive endoderm. This study describes a chemical screening platform to investigate human ESC differentiation and demonstrates the generation of a cell population that is a key milepost on the path to making beta cells.


Assuntos
Carcinógenos/farmacologia , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/efeitos dos fármacos , Indóis/farmacologia , Células Secretoras de Insulina/citologia , Lactamas/farmacologia , Animais , Biomarcadores , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Células-Tronco Embrionárias/fisiologia , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Transativadores/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA