Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 114
Filtrar
1.
Proc Natl Acad Sci U S A ; 118(6)2021 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-33526658

RESUMO

Chemotaxis, the guided motion of cells by chemical gradients, plays a crucial role in many biological processes. In the social amoeba Dictyostelium discoideum, chemotaxis is critical for the formation of cell aggregates during starvation. The cells in these aggregates generate a pulse of the chemoattractant, cyclic adenosine 3',5'-monophosphate (cAMP), every 6 min to 10 min, resulting in surrounding cells moving toward the aggregate. In addition to periodic pulses of cAMP, the cells also secrete phosphodiesterase (PDE), which degrades cAMP and prevents the accumulation of the chemoattractant. Here we show that small aggregates of Dictyostelium can disperse, with cells moving away from instead of toward the aggregate. This surprising behavior often exhibited oscillatory cycles of motion toward and away from the aggregate. Furthermore, the onset of outward cell motion was associated with a doubling of the cAMP signaling period. Computational modeling suggests that this dispersal arises from a competition between secreted cAMP and PDE, creating a cAMP gradient that is directed away from the aggregate, resulting in outward cell motion. The model was able to predict the effect of PDE inhibition as well as global addition of exogenous PDE, and these predictions were subsequently verified in experiments. These results suggest that localized degradation of a chemoattractant is a mechanism for morphogenesis.

2.
PLoS One ; 16(1): e0245924, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33481950

RESUMO

SARS-CoV-2 is a single stranded RNA (ssRNA) virus and contains GU-rich sequences distributed abundantly in the genome. In COVID-19, the infection and immune hyperactivation causes accumulation of inflammatory immune cells, blood clots, and protein aggregates in lung fluid, increased lung alveolar wall thickness, and upregulation of serum cytokine levels. A serum protein called serum amyloid P (SAP) has a calming effect on the innate immune system and shows efficacy as a therapeutic for fibrosis in animal models and clinical trials. Here we show that aspiration of the GU-rich ssRNA oligonucleotide ORN06 into mouse lungs induces all of the above COVID-19-like symptoms. Men tend to have more severe COVID-19 symptoms than women, and in the aspirated ORN06 model, male mice tended to have more severe symptoms than female mice. Intraperitoneal injections of SAP starting from day 1 post ORN06 aspiration attenuated the ORN06-induced increase in the number of inflammatory cells and formation of clot-like aggregates in the mouse lung fluid, reduced ORN06-increased alveolar wall thickness and accumulation of exudates in the alveolar airspace, and attenuated an ORN06-induced upregulation of the inflammatory cytokines IL-1ß, IL-6, IL-12p70, IL-23, and IL-27 in serum. SAP also reduced D-dimer levels in the lung fluid. In human peripheral blood mononuclear cells, SAP attenuated ORN06-induced extracellular accumulation of IL-6. Together, these results suggest that aspiration of ORN06 is a simple model for both COVID-19 as well as cytokine storm in general, and that SAP is a potential therapeutic for diseases with COVID-19-like symptoms and/or a cytokine storm.


Assuntos
/tratamento farmacológico , Síndrome da Liberação de Citocina/tratamento farmacológico , Pneumonia/tratamento farmacológico , Componente Amiloide P Sérico/uso terapêutico , Animais , /patologia , Síndrome da Liberação de Citocina/complicações , Síndrome da Liberação de Citocina/patologia , Modelos Animais de Doenças , Feminino , Humanos , Pulmão/efeitos dos fármacos , Pulmão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pneumonia/complicações , Pneumonia/patologia , Componente Amiloide P Sérico/administração & dosagem
3.
Artigo em Inglês | MEDLINE | ID: mdl-33268492

RESUMO

Polyphosphate is a linear chain of phosphate residues and is present in organisms ranging from bacteria to humans. Pathogens such as Mycobacterium tuberculosis accumulate polyphosphate, and reduced expression of the polyphosphate kinase that synthesizes polyphosphate decreases their survival. How polyphosphate potentiates pathogenicity is poorly understood. Escherichia coli K-12 do not accumulate detectable levels of extracellular polyphosphate and have poor survival after phagocytosis by Dictyostelium discoideum or human macrophages. In contrast, Mycobacterium smegmatis and Mycobacterium tuberculosis accumulate detectable levels of extracellular polyphosphate, and have relatively better survival after phagocytosis by D. discoideum or macrophages. Adding extracellular polyphosphate increased E. coli survival after phagocytosis by D. discoideum and macrophages. Reducing expression of polyphosphate kinase 1 in M. smegmatis reduced extracellular polyphosphate and reduced survival in D. discoideum and macrophages, and this was reversed by the addition of extracellular polyphosphate. Conversely, treatment of D. discoideum and macrophages with recombinant yeast exopolyphosphatase reduced the survival of phagocytosed M. smegmatis or M. tuberculosis D. discoideum cells lacking the putative polyphosphate receptor GrlD had reduced sensitivity to polyphosphate and, compared to wild-type cells, showed increased killing of phagocytosed E. coli and M. smegmatis Polyphosphate inhibited phagosome acidification and lysosome activity in D. discoideum and macrophages and reduced early endosomal markers in macrophages. Together, these results suggest that bacterial polyphosphate potentiates pathogenicity by acting as an extracellular signal that inhibits phagosome maturation.

4.
PLoS One ; 15(12): e0244762, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33378413

RESUMO

Non-alcoholic fatty liver disease (NAFLD) is associated with obesity and type 2 diabetes and is characterized by the accumulation of fat in the liver (steatosis). NAFLD can transition into non-alcoholic steatohepatitis (NASH), with liver cell injury, inflammation, and an increased risk of fibrosis. We previously found that injections of either 1866, a synthetic ligand for the lectin receptor CD209, or DANA, a sialidase inhibitor, can inhibit inflammation and fibrosis in multiple animal models. The methionine and choline-deficient (MCD) diet is a model of NASH which results in the rapid induction of liver steatosis and inflammation. In this report, we show that for C57BL/6 mice on a MCD diet, injections of both 1866 and DANA reversed MCD diet-induced decreases in white fat, decreases in adipocyte size, and white fat inflammation. However, these effects were not observed in type 2 diabetic db/db mice on a MCD diet. In db/db mice on a MCD diet, 1866 decreased liver steatosis, but these effects were not observed in C57BL/6 mice. There was no correlation between the ability of 1866 or DANA to affect steatosis and the effects of these compounds on the density of liver macrophage cells expressing CLEC4F, CD64, F4/80, or Mac2. Together these results indicate that 1866 and DANA modulate adipocyte size and adipose tissue macrophage populations, that 1866 could be useful for modulating steatosis, and that changes in the local density of 4 different liver macrophages cell types do not correlate with effects on liver steatosis.


Assuntos
Tecido Adiposo/efeitos dos fármacos , Moléculas de Adesão Celular/agonistas , Lectinas Tipo C/agonistas , Fígado/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Ácido N-Acetilneuramínico/análogos & derivados , Neuraminidase/antagonistas & inibidores , Receptores de Superfície Celular/agonistas , Tecido Adiposo/metabolismo , Animais , Deficiência de Colina/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Modelos Animais de Doenças , Fígado Gorduroso/tratamento farmacológico , Fígado Gorduroso/metabolismo , Inflamação/metabolismo , Fígado/metabolismo , Macrófagos/metabolismo , Camundongos , Ácido N-Acetilneuramínico/farmacologia , Ácido N-Acetilneuramínico/uso terapêutico , Neuraminidase/metabolismo , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/metabolismo
5.
J Pharmacol Exp Ther ; 2020 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-33144389

RESUMO

The active form of transforming growth factor-ß1 (TGF-ß1) plays a key role in potentiating fibrosis. TGF-ß1 is sequestered in an inactive state by a latency associated glycopeptide (LAP). Sialidases (also called neuraminidases) cleave terminal sialic acids from glycoconjugates. The sialidase NEU3 is upregulated in fibrosis, and mice lacking Neu3 show attenuated bleomycin-induced increases in active TGF-ß1 in the lungs, and attenuated pulmonary fibrosis. Here we observe that recombinant human NEU3 upregulates active human TGF-ß1 by releasing active TGF-ß1 from its latent inactive form by desialylating LAP. Based on the proposed mechanism of action of NEU3, we hypothesized that compounds with a ring structure resembling picolinic acid might be transition state analogues and thus possible NEU3 inhibitors. Some compounds in this class showed nanomolar IC50s for recombinant human NEU3 releasing active human TGF-ß1 from the latent inactive form. The compounds given as daily 0.1 mg/kg - 1 mg/kg injections starting at day 10 strongly attenuated lung inflammation, lung TGF-ß1 upregulation, and pulmonary fibrosis at day 21 in a mouse bleomycin model of pulmonary fibrosis. These results suggest that NEU3 participates in fibrosis by desialylating LAP and releasing TGF-ß1, and that the new class of NEU3 inhibitors are potential therapeutics for fibrosis. Significance Statement The extracellular sialidase NEU3 appears to be a key driver of pulmonary fibrosis. The significance of this report is that 1) we show the mechanism (NEU3 desialylates the LAP protein that keeps the profibrotic cytokine TGF-ß1 in an inactive state, causing TGF-ß1 release), 2) we then use the predicted NEU3 mechanism to create nM IC50 NEU3 inhibitors, and 3) we show that these new NEU3 inhibitors are potent therapeutics in a mouse model of pulmonary fibrosis.

6.
Am J Pathol ; 2020 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-33039353

RESUMO

High-fat diet (HFD)-induced inflammation and steatosis of adipose tissue and liver are associated with a variety of serious health risks. Sialic acids are found as the distal terminal sugar on glycoproteins, which are removed by sialidases (neuraminidases). In humans and mice, pulmonary fibrosis is associated with up-regulation of sialidases, and injections of sialidase inhibitors attenuate bleomycin-induced pulmonary fibrosis. Sialidase levels are altered in obese rodents and humans. This report shows that for mice on an HFD, injections of the sialidase inhibitor N-acetyl-2,3-dehydro-2-deoxyneuraminic acid inhibit weight gain, reduce steatosis, and decrease adipose tissue and liver inflammation. Compared with control, mice lacking the sialidase neuraminidase 3 have reduced HFD-induced adipose tissue and liver inflammation. These data suggest that sialidases promote adipose and liver inflammation in response to a high-fat diet.

7.
bioRxiv ; 2020 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-32869032

RESUMO

SARS-CoV-2 is a single stranded RNA (ssRNA) virus and contains GU-rich sequences distributed abundantly in the genome. In COVID-19, the infection and immune hyperactivation causes accumulation of inflammatory immune cells, blood clots, and protein aggregates in lung fluid, increased lung alveolar wall thickness, and upregulation of serum cytokine levels. A serum protein called serum amyloid P (SAP) has a calming effect on the innate immune system and shows efficacy as a therapeutic for fibrosis in animal models and clinical trials. In this report, we show that aspiration of the GU-rich ssRNA oligonucleotide ORN06 into mouse lungs induces all of the above COVID-19-like symptoms. Men tend to have more severe COVID-19 symptoms than women, and in the aspirated ORN06 model, male mice tended to have more severe symptoms than female mice. Intraperitoneal injections of SAP starting from day 1 post ORN06 aspiration attenuated the ORN06-induced increase in the number of inflammatory cells and formation of clot-like aggregates in the mouse lung fluid, reduced ORN06-increased alveolar wall thickness and accumulation of exudates in the alveolar airspace, and attenuated an ORN06-induced upregulation of the inflammatory cytokines IL-1ß, IL-6, IL-12p70, IL-23, and IL-27 in serum. Together, these results suggest that aspiration of ORN06 is a simple model for both COVID-19 as well as cytokine storm in general, and that SAP is a potential therapeutic for diseases with COVID-19-like symptoms as well as diseases that generate a cytokine storm.

8.
Immunohorizons ; 4(6): 352-362, 2020 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-32576593

RESUMO

Pulmonary fibrosis is a chronic and generally fatal disorder characterized by progressive formation of scar-like tissue in the lungs. Sialic acids are often found as the terminal sugar on extracellular glycoconjugates such as protein glycosylations. Sialidases, also known as neuraminidases, desialylate glycoconjugates. Serum amyloid P (SAP), a pentameric serum glycoprotein that has two sialic acids on each polypeptide, inhibits the differentiation of monocytes into fibrocytes and promotes human PBMCs to accumulate high extracellular levels of IL-10. When SAP is desialylated with sialidase, the effects of SAP on fibrocyte differentiation and IL-10 accumulation are strongly inhibited. Intriguingly, in patients with pulmonary fibrosis, there are increased levels of sialidase activity in the bronchoalveolar lavage fluid, increased levels of sialidases in the lungs, and decreased levels of SAP in the sera. To elucidate the role of SAP desialylation in idiopathic pulmonary fibrosis (IPF) pathogenesis, we purified SAP from the serum of IPF patients and healthy controls and measured the extent of sialylation and bioactivity of the purified SAP. We find that some IPF patients have abnormally high levels of the sialidase NEU3 in their sera and that the SAP in the sera of IPF patients has an abnormally high extent of desialylation and an abnormally low ability to inhibit fibrocyte differentiation and induce extracellular IL-10 accumulation by PBMC. These results suggest that SAP desialylation may play a role in IPF pathogenesis and that inhibiting NEU3 could be a potential therapeutic target for IPF.

9.
Adv Exp Med Biol ; 1224: 79-85, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32036606

RESUMO

Tumors have long been compared to chronic wounds that do not heal, since they share many of the same molecular and cellular processes. In normal wounds, healing processes lead to restoration of cellular architecture, while in malignant tumors, these healing processes become dysregulated and contribute to growth and invasion of neoplastic cells into the surrounding tissues. Fibrocytes are fibroblast-like cells that differentiate from bone marrow-derived CD14+ circulating monocytes and aid wound healing. Although most monocytes will differentiate into macrophages after extravasating into a tissue, signals present in a wound environment can cause some monocytes to differentiate into fibrocytes. The fibrocytes secrete matrix proteins and inflammatory cytokines, activate local fibroblasts to proliferate and increase extracellular matrix production, and promote angiogenesis, and because fibrocytes are contractile, they also help wound contraction. There is now emerging evidence that fibrocytes are present in the tumor microenvironment, attracted by the chronic tissue damage and cytokines from both cancer cells and other immune cells. Fibrocytes may aid in the survival and spread of neoplastic cells, so these wound-healing cells may be a promising target for anticancer research in future studies.


Assuntos
Fibroblastos/patologia , Neoplasias/patologia , Microambiente Tumoral , Fibroblastos/efeitos dos fármacos , Humanos , Neoplasias/tratamento farmacológico , Cicatrização/efeitos dos fármacos
10.
Exp Lung Res ; 46(3-4): 75-80, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32102576

RESUMO

Purpose: We previously found extensive desialylation of glycoconjugates and upregulation of the sialidase enzyme NEU3 in fibrotic lesions in human and mouse lungs. However, studies using microarray analysis of whole lung tissue mRNA and single cell RNA-seq found no significant difference in levels of NEU3 mRNA between IPF patients and controls. This study aimed to elucidate how NEU3 was upregulated in fibrotic lungs.Materials and methods: Transforming growth factor-ß1 (TGF-ß1), a key driver of fibrosis, was added to A549 human alveolar basal epithelial adenocarcinoma cells and human small airway epithelial cells (HSAEpC). NEU3 expression in A549 cells and HSAEpC was detected by immunofluorescence staining. NEU3 translation and degradation were assessed by polysome profiling (polysomes efficiently translate mRNAs; monosomes poorly translate mRNAs) and cycloheximide chase after treating cells with or without TGF-ß1 for 48 h.Results: TGF-ß1 increased NEU3 expression and secretion in A549 cells and HSAEpC but did not change total (nuclear + cytosolic) NEU3 mRNA levels. TGF-ß1 decreased the degradation rate of NEU3 in A549 cells. TGF-ß1 decreased NEU3 mRNA levels in monosomes and increased NEU3 mRNA level in polysomes.Conclusion: TGF-ß1 upregulates levels of NEU3 in epithelial cells by both decreasing NEU3 degradation and by increasing the translation of NEU3 mRNA, explaining the apparent paradox of high levels of NEU3 protein in pulmonary fibrosis without a concomitant increase in the expression of NEU3 mRNA.

11.
Biotechniques ; 68(3): 163-165, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31973564

RESUMO

Shotgun expression of antisense cDNA, where each transformed cell expresses a different antisense cDNA, has been used for mutagenesis and gene identification in Dictyostelium discoideum. However, the method has two limitations. First, there were too few clones in the shotgun antisense cDNA library to have an antisense cDNA for every gene in the genome. Second, the unequal transcription level of genes resulted in many antisense cDNAs in the library for some genes but relatively few antisense cDNAs for other genes. Here we report an improved method for generating a larger antisense cDNA library with a reduced percentage of cDNA clones from highly prevalent mRNAs and demonstrate its utility by screening for signal transduction pathway components in D. discoideum.

12.
Am J Physiol Lung Cell Mol Physiol ; 318(1): L165-L179, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31617733

RESUMO

Pulmonary fibrosis involves the formation of inappropriate scar tissue in the lungs, but what drives fibrosis is unclear. Sialidases (also called neuraminidases) cleave terminal sialic acids from glycoconjugates. In humans and mice, pulmonary fibrosis is associated with desialylation of glycoconjugates and upregulation of sialidases. Of the four mammalian sialidases, we previously detected only NEU3 in the bronchoalveolar lavage fluid from mice with bleomycin-induced pulmonary fibrosis. In this report, we show that NEU3 upregulates extracellular accumulation of the profibrotic cytokines IL-6 and IL-1ß, and IL-6 upregulates NEU3 in human peripheral blood mononuclear cells, suggesting that NEU3 may be part of a positive feedback loop potentiating fibrosis. To further elucidate the role of NEU3 in fibrosis, we used bleomycin to induce lung fibrosis in wild-type C57BL/6 and Neu3-/- mice. At 21 days after bleomycin, compared with male and female C57BL/6 mice, male and female Neu3-/- mice had significantly less inflammation, less upregulation of other sialidases and the profibrotic cytokine active transforming growth factor ß1, and less fibrosis in the lungs. Our results suggest that NEU3 participates in fibrosis and that NEU3 could be a target to develop treatments for fibrosis.


Assuntos
Neuraminidase/metabolismo , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/fisiopatologia , Animais , Bleomicina/farmacologia , Líquido da Lavagem Broncoalveolar , Linhagem Celular , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Inflamação/metabolismo , Inflamação/fisiopatologia , Interleucina-1beta/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/fisiologia , Pulmão/metabolismo , Pulmão/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ácidos Siálicos/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologia
13.
Int J Dev Biol ; 63(8-9-10): 395-405, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31840778

RESUMO

In the last few decades, we have learned a considerable amount about how eukaryotic cells communicate with each other, and what it is the cells are telling each other. The simplicity of Dictyostelium discoideum, and the wide variety of available tools to study this organism, makes it the equivalent of a hydrogen atom for cell and developmental biology. Studies using Dictyostelium have pioneered a good deal of our understanding of eukaryotic cell communication. In this review, we will present a brief overview of how Dictyostelium cells use extracellular signals to attract each other, repel each other, sense their local cell density, sense whether the nearby cells are starving or stressed, count themselves to organize the formation of structures containing a regulated number of cells, sense the volume they are in, and organize their multicellular development. Although we are probably just beginning to learn what the cells are telling each other, the elucidation of Dictyostelium extracellular signals has already led to the development of possible therapeutics for human diseases.


Assuntos
Comunicação Celular , Fatores Quimiotáticos/metabolismo , Dictyostelium/fisiologia , Adenina/análogos & derivados , Adenina/farmacologia , Trifosfato de Adenosina/metabolismo , Amônia/farmacologia , Meios de Cultivo Condicionados , AMP Cíclico/metabolismo , Dictyostelium/genética , Morfogênese , Policetídeos/metabolismo , Polifosfatos/metabolismo
14.
Am J Pathol ; 189(12): 2400-2413, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31539521

RESUMO

High-fat diet (HFD)-induced inflammation is associated with a variety of health risks. The systemic pentraxin serum amyloid P (SAP) inhibits inflammation. SAP activates the high-affinity IgG receptor Fcγ receptor I (FcγRI; CD64) and the lectin receptor dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN; CD209). Herein, we show that for mice on an HFD, injections of SAP and a synthetic CD209 ligand (1866) reduced HFD-increased adipose and liver tissue inflammation, adipocyte differentiation, and lipid accumulation in adipose tissue. HFD worsened glucose tolerance test results and caused increased adipocyte size; for mice on an HFD, SAP improved glucose tolerance test results and reduced adipocyte size. Mice on an HFD had elevated serum levels of IL-1ß, IL-23, interferon (IFN)-ß, IFN-γ, monocyte chemoattractant protein 1 [MCP-1; chemokine (C-C motif) ligand 2 (CCL2)], and tumor necrosis factor-α. SAP reduced serum levels of IL-23, IFN-ß, MCP-1, and tumor necrosis factor-α, whereas 1866 reduced IFN-γ. In vitro, SAP, but not 1866, treated cells isolated from white fat tissue (stromal vesicular fraction) produced the anti-inflammatory cytokine IL-10. HFD causes steatosis, and both SAP and 1866 reduced it. Conversely, compared with control mice, SAP knockout mice fed on a normal diet had increased white adipocyte cell sizes, increased numbers of inflammatory cells in adipose and liver tissue, and steatosis; and these effects were exacerbated on an HFD. SAP and 1866 may inhibit some, but not all, of the effects of a high-fat diet.


Assuntos
Tecido Adiposo/patologia , Moléculas de Adesão Celular/metabolismo , Dieta Hiperlipídica/efeitos adversos , Fígado Gorduroso/prevenção & controle , Hepatite/prevenção & controle , Lectinas Tipo C/metabolismo , Obesidade/complicações , Receptores de Superfície Celular/metabolismo , Componente Amiloide P Sérico/metabolismo , Tecido Adiposo/metabolismo , Animais , Moléculas de Adesão Celular/genética , Fígado Gorduroso/etiologia , Fígado Gorduroso/patologia , Hepatite/etiologia , Hepatite/patologia , Resistência à Insulina , Lectinas Tipo C/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Superfície Celular/genética , Componente Amiloide P Sérico/genética
15.
Comput Struct Biotechnol J ; 17: 684-688, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31303972

RESUMO

Microbial communities are the simplest possible model of multicellular tissues, allowing studies of cell-cell interactions to be done with as few extraneous factors as possible. For instance, the eukaryotic microbe Dictyostelium discoideum proliferates as single cells, and when starved, the cells aggregate together and form structures of ~20,000 cells. The cells use a variety of signals to direct their movement, inform each other of their local cell density and whether they are starving, and organize themselves into groups of ~20,000 cells. Mathematical models and computational approaches have been a key check on, and guide of, the experimental work. In this minireview, I will discuss diffusion calculations and Monte Carlo simulations that were used for Dictyostelium studies that offer general paradigms for several aspects of cell-cell communication. For instance, computational work showed that diffusible secreted cell-density sensing (quorum) factors can diffuse away so quickly from a single cell that the local concentration will not build up to incorrectly cause the cell to sense that it is in the presence of a high density of other cells secreting that signal. In another example, computation correctly predicted a mechanism that allows a group of cells to break up into subgroups. These are thus some examples of the power and necessity of computational work in biology.

16.
J Immunol ; 203(2): 493-499, 2019 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-31160533

RESUMO

Fibrocytes are monocyte-derived fibroblast like cells that participate in wound healing, but little is known about what initiates fibrocyte differentiation. Blood platelets contain 60-100-mer polymers of phosphate groups called polyphosphate, and when activated, platelets induce blood clotting (the first step in wound healing) in part by the release of polyphosphate. We find that activated platelets release a factor that promotes fibrocyte differentiation. The factor is abolished by treating the crude platelet factor with the polyphosphate-degrading enzyme polyphosphatase, and polyphosphate promotes fibrocyte differentiation. Macrophages and recruited neutrophils also potentiate wound healing, and polyphosphate also promotes macrophage differentiation and induces chemoattraction of neutrophils. In support of the hypothesis that polyphosphate is a signal that affects leukocytes, we observe saturable binding of polyphosphate to these cells. Polyphosphate also inhibits leukocyte proliferation and proteasome activity. These results suggest new roles for extracellular polyphosphate as a mediator of wound healing and inflammation and also provide a potential link between platelet activation and the progression of fibrosing diseases.


Assuntos
Diferenciação Celular/fisiologia , Proliferação de Células/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Leucócitos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Polifosfatos/farmacologia , Coagulação Sanguínea/efeitos dos fármacos , Coagulação Sanguínea/fisiologia , Fibroblastos/metabolismo , Humanos , Leucócitos/metabolismo , Macrófagos/metabolismo , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Neutrófilos/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Cicatrização/fisiologia
17.
mBio ; 10(2)2019 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-30862745

RESUMO

In patients with invasive fungal diseases, there is often little cellular inflammatory response. We tested the idea that binding of the human constitutive plasma protein serum amyloid P component (SAP) (also called PTX2) to Candida albicans dampens the innate immune response to this fungus. Many pathogenic fungi have cell surface amyloid-like structures important for adhesion and biofilm formation. Human SAP bound to fungi that expressed functional cell surface amyloid, but SAP had minimal binding to fungi with reduced expression of cell surface amyloid. In the absence of SAP, phagocytosis of fungi by human macrophages was potentiated by expression of amyloid on the fungi. SAP binding to fungi inhibited their phagocytosis by macrophages. Macrophages pretreated with SAP displayed reduced fungal phagocytosis, reduced secretion of inflammatory cytokines (IFN-γ, IL-6, and TNF-α), and increased secretion of the anti-inflammatory cytokine IL-10. SAP bound to fungi or added to the medium upregulated the expression of the anti-inflammatory receptor CD206 on macrophages. These findings suggest that SAP bound to amyloid-like structures on fungal cells dampens the host cellular immune response in fungal diseases such as invasive candidiasis.IMPORTANCE Macrophages are a key part of our innate immune system and are responsible for recognizing invading microbes, ingesting them, and sending appropriate signals to other immune cells. We have found that human macrophages can recognize invading yeast pathogens that have a specific molecular pattern of proteins on their surfaces: these proteins have structures similar to the structures of amyloid aggregates in neurodegenerative diseases like Alzheimer's disease. However, this surface pattern also causes the fungi to bind a serum protein called serum amyloid P component (SAP). In turn, the SAP-coated yeasts are poorly recognized and seldom ingested by the macrophages, and the macrophages have a more tolerant and less inflammatory response in the presence of SAP. Therefore, we find that surface structures on the yeast can alter how the macrophages react to invading microbes.


Assuntos
Candida albicans/imunologia , Candidíase/microbiologia , Citocinas/metabolismo , Proteínas Fúngicas/metabolismo , Macrófagos/imunologia , Fagocitose/efeitos dos fármacos , Componente Amiloide P Sérico/metabolismo , Candidíase/imunologia , Células Cultivadas , Humanos , Macrófagos/efeitos dos fármacos
18.
Mol Biol Cell ; 30(9): 1118-1128, 2019 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-30785840

RESUMO

Five or more orthophosphates bound together by high-energy phosphoanhydride bonds are highly ubiquitous inorganic molecules called polyphosphate. Polyphosphate acts as a signaling molecule eliciting a number of responses in eukaryotic cells, but the mechanisms mediating these effects are poorly understood. Proliferating Dictyostelium discoideum cells accumulate extracellular polyphosphate. At extracellular concentrations similar to those observed in stationary phase cells, polyphosphate inhibits proteasome activity and proliferation, and induces aggregation. Here we identify GrlD as a putative G protein-coupled receptor that mediates binding of extracellular polyphosphate to the cell surface. Cells lacking GrlD do not respond to polyphosphate-induced proteasome inhibition, aggregation, or proliferation inhibition. Polyphosphate also elicits differential effects on cell-substratum adhesion and cytoskeletal F-actin levels based on nutrient availability, and these effects were also mediated by GrlD. Starving cells also accumulate extracellular polyphosphate. Starved cells treated with exopolyphosphatase failed to aggregate effectively, suggesting that polyphosphate also acts as a signaling molecule during starvation-induced development of Dictyostelium. Together, these results suggest that a eukaryotic cell uses a G protein-coupled receptor to mediate the sensing and response to extracellular polyphosphate.


Assuntos
Dictyostelium/metabolismo , Polifosfatos/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Membrana Celular/metabolismo , Dictyostelium/fisiologia , Receptores Acoplados a Proteínas-G/fisiologia , Transdução de Sinais/fisiologia
19.
Mol Biol Cell ; 30(2): 242-255, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30462573

RESUMO

Eukaryotic chemoattraction signal transduction pathways, such as those used by Dictyostelium discoideum to move toward cAMP, use a G protein-coupled receptor to activate multiple conserved pathways such as PI3 kinase/Akt/PKB to induce actin polymerization and pseudopod formation at the front of a cell, and PTEN to localize myosin II to the rear of a cell. Relatively little is known about chemorepulsion. We previously found that AprA is a chemorepellent protein secreted by Dictyostelium cells. Here we used 29 cell lines with disruptions of cAMP and/or AprA signal transduction pathway components, and delineated the AprA chemorepulsion pathway. We find that AprA uses a subset of chemoattraction signal transduction pathways including Ras, protein kinase A, target of rapamycin (TOR), phospholipase A, and ERK1, but does not require the PI3 kinase/Akt/PKB and guanylyl cyclase pathways to induce chemorepulsion. Possibly as a result of not using the PI3 kinase/Akt/PKB pathway and guanylyl cyclases, AprA does not induce actin polymerization or increase the pseudopod formation rate, but rather appears to inhibit pseudopod formation at the side of cells closest to the source of AprA.


Assuntos
Quimiotaxia , Dictyostelium/citologia , Pseudópodes/metabolismo , Actinas/metabolismo , Canais de Cálcio/metabolismo , Polaridade Celular , AMP Cíclico/metabolismo , Citoesqueleto/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Guanilato Ciclase/metabolismo , Modelos Biológicos , Mutação/genética , Miosina Tipo II/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosfolipases/metabolismo , Proteína Quinase C/metabolismo , Transporte Proteico , Proteínas de Protozoários/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Transdução de Sinais , Proteínas ras/metabolismo
20.
J Immunol ; 202(1): 239-248, 2019 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-30510066

RESUMO

The movement of neutrophils between blood and tissues appears to be regulated by chemoattractants and chemorepellents. Compared with neutrophil chemoattractants, relatively little is known about neutrophil chemorepellents. Slit proteins are endogenously cleaved into a variety of N- and C-terminal fragments, and these fragments are neuronal chemorepellents and inhibit chemoattraction of many cell types, including neutrophils. In this report, we show that the ∼140-kDa N-terminal Slit2 fragment (Slit2-N) is a chemoattractant and the ∼110-kDa N-terminal Slit2 fragment (Slit2-S) is a chemorepellent for human neutrophils. The effects of both Slit2 fragments were blocked by Abs to the Slit2 receptor Roundabout homolog 1 or the Slit2 coreceptor Syndecan-4. Slit2-N did not appear to activate Ras but increased phosphatidylinositol 3,4,5-triphosphate levels. Slit2-N-induced chemoattraction was unaffected by Ras inhibitors, reversed by PI3K inhibitors, and blocked by Cdc42 and Rac inhibitors. In contrast, Slit2-S activated Ras but did not increase phosphatidylinositol 3,4,5-triphosphate levels. Slit2-S-induced chemorepulsion was blocked by Ras and Rac inhibitors, not affected by PI3K inhibitors, and reversed by Cdc42 inhibitors. Slit2-N, but not Slit2-S, increased neutrophil adhesion, myosin L chain 2 phosphorylation, and polarized actin formation and single pseudopods at the leading edge of cells. Slit2-S induced multiple pseudopods. These data suggest that Slit2 isoforms use similar receptors but different intracellular signaling pathways and have different effects on the cytoskeleton and pseudopods to induce neutrophil chemoattraction or chemorepulsion.


Assuntos
Citoesqueleto de Actina/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neutrófilos/fisiologia , Isoformas de Proteínas/metabolismo , Pseudópodes/metabolismo , Anticorpos Bloqueadores/metabolismo , Orientação de Axônios , Adesão Celular , Células Cultivadas , Quimiotaxia/genética , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/imunologia , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/imunologia , Fosfatos de Fosfatidilinositol/metabolismo , Ligação Proteica , Isoformas de Proteínas/genética , Pseudópodes/ultraestrutura , Transdução de Sinais , Sindecana-4/metabolismo , Proteína cdc42 de Ligação ao GTP/metabolismo , Proteínas ras/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...