Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 14.020
Filtrar
1.
Nat Commun ; 11(1): 4477, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32901019

RESUMO

Individual cells detach from cohesive ensembles during development and can inappropriately separate in disease. Although much is known about how cells separate from epithelia, it remains unclear how cells disperse from clusters lacking apical-basal polarity, a hallmark of advanced epithelial cancers. Here, using live imaging of the developmental migration program of Drosophila primordial germ cells (PGCs), we show that cluster dispersal is accomplished by stabilizing and orienting migratory forces. PGCs utilize a G protein coupled receptor (GPCR), Tre1, to guide front-back migratory polarity radially from the cluster toward the endoderm. Posteriorly positioned myosin-dependent contractile forces pull on cell-cell contacts until cells release. Tre1 mutant cells migrate randomly with transient enrichment of the force machinery but fail to separate, indicating a temporal contractile force threshold for detachment. E-cadherin is retained on the cell surface during cell separation and augmenting cell-cell adhesion does not impede detachment. Notably, coordinated migration improves cluster dispersal efficiency by stabilizing cell-cell interfaces and facilitating symmetric pulling. We demonstrate that guidance of inherent migratory forces is sufficient to disperse cell clusters under physiological settings and present a paradigm for how such events could occur across development and disease.


Assuntos
Drosophila melanogaster/embriologia , Células Germinativas Embrionárias/fisiologia , Animais , Animais Geneticamente Modificados , Fenômenos Biomecânicos , Padronização Corporal/fisiologia , Caderinas/metabolismo , Adesão Celular/fisiologia , Movimento Celular/fisiologia , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/fisiologia , Células Germinativas Embrionárias/citologia , Microscopia de Fluorescência por Excitação Multifotônica , Miosina Tipo II/metabolismo , Transdução de Sinais , Análise de Célula Única , Proteínas rho de Ligação ao GTP/metabolismo
2.
Medicine (Baltimore) ; 99(39): e22172, 2020 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-32991410

RESUMO

Osteoporosis is a severe chronic skeletal disorder that increases the risks of disability and mortality; however, the mechanism of this disease and the protein markers for prognosis of osteoporosis have not been well characterized. This study aims to characterize the imbalanced serum proteostasis, the disturbed pathways, and potential serum markers in osteoporosis by using a set of bioinformatic analyses. In the present study, the large-scale proteomics datasets (PXD006464) were adopted from the Proteome Xchange database and processed with MaxQuant. The differentially expressed serum proteins were identified. The biological process and molecular function were analyzed. The protein-protein interactions and subnetwork modules were constructed. The signaling pathways were enriched. We identified 209 upregulated and 230 downregulated serum proteins. The bioinformatic analyses revealed a highly overlapped functional protein classification and the gene ontology terms between the upregulated and downregulated protein groups. Protein-protein interactions and pathway analyses showed a high enrichment in protein synthesis, inflammation, and immune response in the upregulated proteins, and cell adhesion and cytoskeleton regulation in the downregulated proteins. Our findings greatly expand the current view of the roles of serum proteins in osteoporosis and shed light on the understanding of its underlying mechanisms and the discovery of serum proteins as potential markers for the prognosis of osteoporosis.


Assuntos
Mineração de Dados/métodos , Osteoporose/sangue , Proteoma/fisiologia , Biomarcadores , Adesão Celular/fisiologia , Biologia Computacional , Citoesqueleto/metabolismo , Regulação para Baixo , Humanos , Mediadores da Inflamação/metabolismo , Mapas de Interação de Proteínas/fisiologia , Proteômica , Regulação para Cima
3.
Int J Nanomedicine ; 15: 5061-5072, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32764936

RESUMO

Purpose: Zirconia is one of the most promising implant materials due to its favorable physical, mechanical and biological properties. However, until now, we know little about the mechanism of osseointegration on zirconia. The purpose of this study is to evaluate the effect of Syndecan (Sdc) on osteoblastic cell (MC3T3-E1) adhesion and proliferation onto zirconia materials. Materials and Methods: The mirror-polished disks 15 mm in diameter and 1.5 mm in thick of commercial pure titanium (CpTi), 3mol% yttria-stabilized tetragonal zirconia polycrystalline (3Y-TZP) and nano-zirconia (NanoZr) are used in this study. MC3T3-E1 cells were seeded onto specimen surfaces and subjected to RNA interference (RNAi) for Syndecan-1, Syndecan-2, Syndecan-3, and Syndecan-4. At 48h post-transfection, the cell morphology, actin cytoskeleton, and focal adhesion were observed using scanning electron microscopy or laser scanning confocal fluorescence microscopy. At 24h and 48h post-transfection, cell counting kit-8 (CCK-8) assay was used to investigate cell proliferation. Results: The cell morphology of MC3T3-E1 cells on CpTi, 3Y-TZP, and NanoZr changed into abnormal shape after gene silencing of Syndecan. Among the Syndecan family, Sdc-2 is responsible for NanoZr-specific morphology regulation, via maintenance of cytoskeletal conformation without affecting cellular attachment. According to CCK-8 assay, Sdc-2 affects the osteoblastic cell proliferation onto NanoZr. Conclusion: Within the limitation of this study, we suggest that Syndecan affects osteoblastic cell adhesion on CpTi, 3Y-TZP, and NanoZr. Sdc-2 might be an important heparin-sensitive cell membrane regulator in osteoblastic cell adhesion, specifically on NanoZr, through the organization of actin cytoskeleton and affects osteoblastic cell proliferation.


Assuntos
Osseointegração/fisiologia , Osteoblastos/citologia , Osteoblastos/fisiologia , Sindecanas/metabolismo , Citoesqueleto de Actina/efeitos dos fármacos , Animais , Adesão Celular/fisiologia , Membrana Celular/efeitos dos fármacos , Proliferação de Células/fisiologia , Células Cultivadas , Camundongos , Microscopia Eletrônica de Varredura , Osseointegração/genética , Propriedades de Superfície , Sindecana-2/genética , Sindecana-2/metabolismo , Sindecanas/genética , Titânio/química , Ítrio/química , Zircônio/química
4.
Gene ; 760: 144965, 2020 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-32687948

RESUMO

OBJECTIVE: Immunologic contact urticaria (ICU) is an immediate response of wheal caused by various contactants in vulnerable individuals, with undefined pathogenesis. METHODS: In the present study, we aim to explore the effects of intercellular cell adhesion molecule-1 (ICAM-1) gene silencing by RNA inference (RNAi) on vascular endothelial cells (VECs) adhesion molecule expression and cell-cell adhesion in ICU mice. Sixty BALB/c mice were selected, among which 48 mice were used for establishment of ICU models. VECs from normal and ICU mice were grouped into different groups. Expressions of ICAM-1, eosinophilic cationic protein (ECP), total immunologlobulin E (tIgE), L-selectin (CD62L), integrin, alpha L (CD11a) in tissues and cells were evaluate by RT-qPCR and western blotting. Cell proliferation was evaluated by MTT assay and EdU staining and cell adhesive function by cell-cell adhesion assay. RESULTS: Compared with normal mice, ICU mice had increased expressions of ICAM-1, ECP, tIgE, CD62L, and CD11a.ICAM-1 gene silencing decreased expressions of ECP, tIgE, CD62L, and CD11a, enhanced cell proliferation, and more activity in cell adhesion. CONCLUSION: These findings indicate that RNAi-mediated gene silencing of ICAM-1 may decrease VECs adhesion expression and reduce cell-cell adhesion in mice with ICU.


Assuntos
Células Endoteliais/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Urticária/genética , Animais , Adesão Celular/fisiologia , Células Cultivadas , Células Endoteliais/patologia , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Feminino , Inativação Gênica , Molécula 1 de Adesão Intercelular/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Interferência de RNA , Urticária/metabolismo , Urticária/patologia
5.
Nat Commun ; 11(1): 2694, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32483155

RESUMO

Toxin complex (Tc) toxins are virulence factors of pathogenic bacteria. Tcs are composed of three subunits: TcA, TcB and TcC. TcA facilitates receptor-toxin interaction and membrane permeation, TcB and TcC form a toxin-encapsulating cocoon. While the mechanisms of holotoxin assembly and pore formation have been described, little is known about receptor binding of TcAs. Here, we identify heparins/heparan sulfates and Lewis antigens as receptors for different TcAs from insect and human pathogens. Glycan array screening reveals that all tested TcAs bind negatively charged heparins. Cryo-EM structures of Morganella morganii TcdA4 and Xenorhabdus nematophila XptA1 reveal that heparins/heparan sulfates unexpectedly bind to different regions of the shell domain, including receptor-binding domains. In addition, Photorhabdus luminescens TcdA1 binds to Lewis antigens with micromolar affinity. Here, the glycan interacts with the receptor-binding domain D of the toxin. Our results suggest a glycan dependent association mechanism of Tc toxins on the host cell surface.


Assuntos
Toxinas Bacterianas/toxicidade , Adesão Celular/efeitos dos fármacos , Adesão Celular/fisiologia , Polissacarídeos/metabolismo , Animais , Toxinas Bacterianas/química , Toxinas Bacterianas/farmacocinética , Sítios de Ligação , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Células HEK293 , Heparina/química , Heparina/metabolismo , Humanos , Insetos/microbiologia , Antígenos CD15/química , Antígenos CD15/metabolismo , Modelos Moleculares , Simulação de Acoplamento Molecular , Morganella morganii/patogenicidade , Photorhabdus/patogenicidade , Polissacarídeos/química , Xenorhabdus/patogenicidade
6.
Am J Pathol ; 190(9): 1931-1942, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32526166

RESUMO

Pancreatic cancer has a dismal prognosis, and there is no targeted therapy against this malignancy. The neuronal membrane protein sortilin is emerging as a regulator of cancer cell development, but its expression and impact in pancreatic cancer are unknown. This study found that sortilin expression was higher in pancreatic cell lines versus normal pancreatic ductal epithelial cells, as shown by Western blot analysis and mass spectrometry. The increased sortilin level in pancreatic cancer cells was confirmed by immunohistochemistry in a series of 99 human pancreatic adenocarcinomas versus 48 normal pancreatic tissues (P = 0.0014). Sortilin inhibition by siRNA and the pharmacologic inhibitor AF38469 strongly reduced the adhesion and invasion of pancreatic cancer cells without affecting cell survival and viability. Sortilin inhibition also decreased the phosphorylation of the focal adhesion kinase in Tyr925. Together, these data show that sortilin contributes to pancreatic cancer invasion and could eventually be targeted in therapy.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Neoplasias Pancreáticas/patologia , Adesão Celular/fisiologia , Movimento Celular/fisiologia , Humanos , Invasividade Neoplásica/patologia , Neoplasias Pancreáticas/metabolismo
7.
Life Sci ; 254: 117780, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32407844

RESUMO

AIMS: In vivo studies suggest a positive influence of fresh frozen plasma (FFP) on endothelial properties and vascular barrier function, leading to improved outcomes in animal sepsis models as well as in major abdominal surgery. However, those effects are incompletely described. It was our aim to evaluate in vitro effects of FFP on endothelial key functions and to identify underlying mechanisms. MATERIALS AND METHODS: Human pulmonary microvascular endothelial cells (HPMECs) were prestimulated with LPS, followed by incubation with FFP. Permeability for FITC-dextran was assessed, and intercellular gap formation was visualized. NF-κB nuclear translocation and expression of pro-inflammatory, pro-adhesion, and leakage-related genes were evaluated, and monocyte adhesion to ECs was assessed. Intracellular cAMP levels as well as phosphorylation of functional proteins were analyzed. In patients undergoing major abdominal surgery, Syndecan-1 serum levels were assessed prior to and following FFP transfusion. KEY FINDINGS: Post-incubation of HPMVECs with FFP increased intracellular cAMP levels that had been decreased by preceding LPS stimulation. On one hand, this reduced endotoxin-mediated upregulation of IL-8, ICAM-1, VCAM-1, VEGF, and ANG-2. Impaired phosphorylation of functional proteins was restored, and intercellular cohesion and barrier function were rescued. On the other hand, NF-κB nuclear translocation as well as monocyte adhesion was markedly increased by the combination of LPS and FFP. Syndecan-1 serum levels were lower in surgery patients that were transfused with FFP compared to those that were not. SIGNIFICANCE: Our data provide evidence for a differential modulation of crucial endothelial properties by FFP, potentially mediated by elevation of intracellular cAMP levels.


Assuntos
Células Endoteliais/fisiologia , Endotélio Vascular/metabolismo , Plasma/fisiologia , Idoso , Adesão Celular/fisiologia , Permeabilidade da Membrana Celular/fisiologia , Células Cultivadas , AMP Cíclico/metabolismo , Dextranos/metabolismo , Fluoresceína-5-Isotiocianato/análogos & derivados , Fluoresceína-5-Isotiocianato/metabolismo , Junções Comunicantes/fisiologia , Humanos , Lipopolissacarídeos , Pessoa de Meia-Idade , Monócitos/fisiologia , NF-kappa B/metabolismo , Fosforilação , Sindecana-1/sangue
8.
PLoS One ; 15(5): e0232432, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32365067

RESUMO

CR3 and CR4, the leukocyte specific ß2-integrins, involved in cellular adherence, migration and phagocytosis, are often assumed to have similar functions. Previously however, we proved that under physiological conditions CR4 is dominant in the adhesion to fibrinogen of human monocyte-derived macrophages (MDMs) and dendritic cells (MDDCs). Here, using inflammatory conditions, we provide further evidence that the expression and function of CR3 and CR4 are not identical in these cell types. We found that LPS treatment changes their expression differently on MDMs and MDDCs, suggesting a cell type specific regulation. Using mAb24, specific for the high affinity conformation of CD18, we proved that the activation and recycling of ß2-integrins is significantly enhanced upon LPS treatment. Adherence to fibrinogen was assessed by two fundamentally different approaches: a classical adhesion assay and a computer-controlled micropipette, capable of measuring adhesion strength. While both receptors participated in adhesion, we demonstrated that CR4 exerts a dominant role in the strong attachment of MDDCs. Studying the formation of podosomes we found that MDMs retain podosome formation after LPS activation, whereas MDDCs lose this ability, resulting in a significantly reduced adhesion force and an altered cellular distribution of CR3 and CR4. Our results suggest that inflammatory conditions reshape differentially the expression and role of CR3 and CR4 in macrophages and dendritic cells.


Assuntos
Células Dendríticas/imunologia , Inflamação/imunologia , Integrina alfaXbeta2/imunologia , Antígeno de Macrófago 1/imunologia , Macrófagos/imunologia , Podossomos/imunologia , Anticorpos Bloqueadores/imunologia , Antígenos CD18/imunologia , Adesão Celular/imunologia , Adesão Celular/fisiologia , Diferenciação Celular/imunologia , Movimento Celular/imunologia , Movimento Celular/fisiologia , Células Dendríticas/patologia , Células Dendríticas/fisiologia , Fibrinogênio/imunologia , Humanos , Técnicas In Vitro , Inflamação/patologia , Lipopolissacarídeos/imunologia , Macrófagos/patologia , Macrófagos/fisiologia , Fagocitose/imunologia , Fagocitose/fisiologia , Podossomos/patologia
9.
Nat Commun ; 11(1): 1606, 2020 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-32231207

RESUMO

Tetraspanins play critical roles in various physiological processes, ranging from cell adhesion to virus infection. The members of the tetraspanin family have four membrane-spanning domains and short and large extracellular loops, and associate with a broad range of other functional proteins to exert cellular functions. Here we report the crystal structure of CD9 and the cryo-electron microscopic structure of CD9 in complex with its single membrane-spanning partner protein, EWI-2. The reversed cone-like molecular shape of CD9 generates membrane curvature in the crystalline lipid layers, which explains the CD9 localization in regions with high membrane curvature and its implications in membrane remodeling. The molecular interaction between CD9 and EWI-2 is mainly mediated through the small residues in the transmembrane region and protein/lipid interactions, whereas the fertilization assay revealed the critical involvement of the LEL region in the sperm-egg fusion, indicating the different dependency of each binding domain for other partner proteins.


Assuntos
Tetraspanina 29/química , Tetraspanina 29/fisiologia , Animais , Antígenos CD/química , Adesão Celular/fisiologia , Microscopia Crioeletrônica , Cristalografia por Raios X , Feminino , Fertilização/fisiologia , Humanos , Masculino , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Modelos Moleculares , Conformação Proteica , Tetraspanina 29/genética
10.
Invest Ophthalmol Vis Sci ; 61(3): 44, 2020 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-32232343

RESUMO

Purpose: To determine the composition of extracellular matrix (ECM) proteins secreted by a conjunctival epithelial cell line and to identify components that aid conjunctival epithelial cell culture. Methods: Human conjunctival epithelial cell line (HCjE-Gi) cells were cultured in serum-free media and their ECM isolated using ammonium hydroxide. Growth characteristics were evaluated for fresh HCjE-Gi cells plated onto ECMs obtained from 3- to 28-day cell cultures. Mass spectrometry was used to characterize the ECM composition over 42 culture days. Cell adhesion and growth on pre-adsorbed fibronectin and α-2-HS-glycoprotein (α-2-HS-GP) were investigated. Results: Day 3 ECM provided the best substrate for cell growth compared to ECM obtained from 5- to 28-day cell cultures. Mass spectrometry identified a predominantly laminin 332 matrix throughout the time course, with progressive changes to matrix composition over time: proportional decreases in matrix-bound growth factors and increases in proteases. Fibronectin and α-2-HS-GP were 5- and 200-fold enriched as a proportion of the early ECM relative to the late ECM, respectively. Experiments on these proteins in isolation demonstrated that fibronectin supported rapid cell adhesion, whereas fibronectin and α-2-HS-GP both supported enhanced cell growth compared to tissue culture polystyrene. Conclusions: These data reveal α-2-HS-GP as a candidate protein to enhance the growth of conjunctival epithelial cells and raise the possibility of exploiting these findings for targeted improvement to synthetic tissue engineered conjunctival substrates.


Assuntos
Túnica Conjuntiva/metabolismo , Proteínas da Matriz Extracelular/metabolismo , alfa-2-Glicoproteína-HS/metabolismo , Biomarcadores/metabolismo , Adesão Celular/fisiologia , Contagem de Células , Linhagem Celular , Proliferação de Células/fisiologia , Túnica Conjuntiva/citologia , Meios de Cultura Livres de Soro , Células Epiteliais/metabolismo , Matriz Extracelular/metabolismo , Fibronectinas/metabolismo , Humanos , Espectrometria de Massas
11.
J Neurosci ; 40(17): 3360-3373, 2020 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-32265259

RESUMO

The Drosophila nervous system is ensheathed by a layer of outer glial cells, the perineurial glia, and a specialized extracellular matrix, the neural lamella. The function of perineurial glial cells and how they interact with the extracellular matrix are just beginning to be elucidated. Integrin-based focal adhesion complexes link the glial membrane to the extracellular matrix, but little is known about integrin's regulators in the glia. The transmembrane Ig domain protein Basigin/CD147/EMMPRIN is highly expressed in the perineurial glia surrounding the Drosophila larval nervous system. Here we show that Basigin associates with integrin at the focal adhesions to uphold the structure of the glia-extracellular matrix sheath. Knockdown of Basigin in perineurial glia using RNAi results in significant shortening of the ventral nerve cord, compression of the glia and extracellular matrix in the peripheral nerves, and reduction in larval locomotion. We determined that Basigin is expressed in close proximity to integrin at the glial membrane, and that expression of the extracellular integrin-binding domain of Basigin is sufficient to rescue peripheral glial compression. We also found that a reduction in expression of integrin at the membrane rescues the ventral nerve cord shortening, peripheral glial compression, and locomotor phenotypes, and that reduction in the integrin-binding protein Talin can partially rescue glial compression. These results identify Basigin as a potential negative regulator of integrin in the glia, supporting proper glial and extracellular matrix ensheathment of the nervous system.SIGNIFICANCE STATEMENT The glial cells and extracellular matrix play important roles in supporting and protecting the nervous system, but the interactions between these components have not been well characterized. Our study identified expression of a conserved Ig superfamily protein, Basigin, at the glial membrane of Drosophila where it associates with the integrin-based focal adhesion complexes to ensure proper ensheathment of the CNS and PNS. Loss of Basigin in the glia results in an overall compression of the nervous system due to integrin dysregulation, which causes locomotor defects in the animals. This underlies the importance of glia-matrix communication for structural and functional support of the nervous system.


Assuntos
Proteínas de Drosophila/metabolismo , Integrinas/metabolismo , Glicoproteínas de Membrana/metabolismo , Neuroglia/metabolismo , Nervos Periféricos/metabolismo , Animais , Adesão Celular/fisiologia , Drosophila melanogaster , Matriz Extracelular/metabolismo , Larva/metabolismo , Locomoção/fisiologia , Neuroglia/citologia , Nervos Periféricos/citologia , Interferência de RNA
12.
Lab Invest ; 100(7): 916-927, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32238905

RESUMO

Synovial mesenchymal stem cells (MSCs) are an attractive cell source for transplantation because of their high chondrogenic potential, especially in areas like the meniscus of the knee. A synovial MSC suspension placed onto the meniscus for 10 min promoted healing of repaired meniscal tears that generally do not heal. Here, we quantified the proportion of human synovial MSCs that adhered to a porcine abraded meniscus, clarified their morphological changes, and revealed the mechanism by which the synovial MSCs adhered to the meniscus. The numbers of adhering cells at immediately after 10, 60 min and 6, 24 h after suspension placement were calculated. The meniscus surface was examined by scanning electron microscopy, and 50 cells were randomly selected at each time period, classified, and quantified for each of the six donors. Approximately 28% of the synovial MSCs immediately adhered to the meniscus after placement and the proportion of adhered cells increased further with time. All cells maintained a round shape for 60 min, and then transformed to a mixture of round and semi-flattened cells. By 24 h, flattened cells covered the meniscus. Microspikes were observed in 36% of the floating synovial MSCs and in 76% of the cells on the meniscus shortly after placement on the meniscus, then the proportion of cells with pseudopodia increased. The bleb-dominant cell proportion significantly decreased, and the smooth-dominant cell proportion increased within 60 min. Microspikes or the bodies of synovial MSCs were trapped by meniscal fibers immediately after placement. The proportion of adhered cells increased with time, and the cell morphology changed dynamically for 24 h as the synovial MSCs adhered to the meniscus. The MSCs in the round morphological state had a heterogeneous morphology. The microspikes, and the subsequent development of pseudopodia, may play an important role in adhesion onto the meniscus.


Assuntos
Adesão Celular/fisiologia , Menisco/metabolismo , Células-Tronco Mesenquimais , Membrana Sinovial/citologia , Idoso , Idoso de 80 Anos ou mais , Animais , Células Cultivadas , Feminino , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/fisiologia , Pessoa de Meia-Idade , Suínos
13.
Inflamm Res ; 69(7): 631-633, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32303780

RESUMO

OBJECTIVE AND DESIGN: CHRFAM7A is a unique human gene that encodes a dominant negative inhibitor of the α7 nicotinic acetylcholine receptor. We have recently shown that CHRFAM7A is expressed in human leukocytes, increases cel-cell adhesion, and regulates the expression of genes associated with leukocyte migration. MATERIAL: Human THP-1, RAW264.7 and HEK293 cells. METHODS: Cell migration, cell proliferation and colony formation in soft agar to compare the biological activity of vector vs. CHRFAM7A-transduced cells. RESULTS: We show that gene delivery of CHRFAM7A into the THP-1 human monocytic cell line reduces cell migration, reduces chemotaxis to monocyte chemoattractant protein, and reduces colony formation in soft agar. CONCLUSION: Taken together, the findings demonstrate that CHRFAM7A regulates the biological activity of monocytes/macrophages to migrate and undergo anchorage-independent growth in vitro.


Assuntos
Receptor Nicotínico de Acetilcolina alfa7/antagonistas & inibidores , Animais , Adesão Celular/fisiologia , Movimento Celular/genética , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Expressão Gênica , Regulação da Expressão Gênica , Células HEK293 , Humanos , Leucócitos , Macrófagos/fisiologia , Camundongos , Monócitos/fisiologia , Células RAW 264.7 , Células-Tronco/fisiologia , Células THP-1 , Transdução Genética , Receptor Nicotínico de Acetilcolina alfa7/genética , Receptor Nicotínico de Acetilcolina alfa7/fisiologia
14.
J Surg Oncol ; 121(7): 1084-1089, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32153051

RESUMO

BACKGROUND AND OBJECTIVES: The aim of this study was to evaluate whether the amount of signet ring cells (SRCs) affects clinicopathological characteristics and prognosis of poorly cohesive (PC) gastric tumours. STUDY DESIGN: One hundred seventy-three patients with PC tumours treated at three European centres from 2004 to 2014 were reclassified in three categories: (a) pure SRC cancers (SRC1) (≥90% SRCs); (b) PC carcinoma with SRC component (SRC2) (>10%, <90% SRCs); (c) PC carcinoma not otherwise specified (SRC3) (≤10% SRCs). RESULTS: The percentage of SRCs was inversely related to the pT stage (Spearman's ρ = -0.174, P < .001) and the number of positive nodes coded as a continuous variable (P = .009). Five year cancer-related survival was significantly higher (58%, 95% confidence interval [CI]: 36%-75%) in SRC1 compared with SRC2 (39%, 95% CI: 28%-50%) and SRC3 (38%, 95% CI: 22%-53%), (P = .048). In multivariable analysis, the impact of PC categories on cancer-related survival was significant when controlling for sex, age, pT, pN, and curativity (hazard ratio [HR] of sSRC2 vs SRC1 = 2.08, 95% CI: 1.01-4.29, P = .046; HR of SRC3 vs SRC1 = 2.38, 95% CI: 1.05-5.41, P = .039). CONCLUSION: The percentage of SRCs was inversely related to tumour aggressiveness, with long-term survival significantly higher in SRC1 compared with SRC2 and SRC3 tumours.


Assuntos
Carcinoma de Células em Anel de Sinete/patologia , Neoplasias Gástricas/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células em Anel de Sinete/mortalidade , Adesão Celular/fisiologia , Europa (Continente)/epidemiologia , Feminino , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Estudos Retrospectivos , Neoplasias Gástricas/mortalidade
15.
Life Sci ; 249: 117518, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32147432

RESUMO

AIMS: The objectives of the present study were to investigate the mechanisms of Ninj-1 regulation in TNFα-activated human endothelial cells (HEC), and to test if Amlodipine (AML) ameliorates the inflammatory stress by decreasing Ninj-1 expression. MAIN METHODS: TNFα-activated HEC with/without AML (0.1 µM and 1 µM) were used. TNFα-receptor 1 (TNFR1) was silenced and inhibitors for oxidative stress (N-acetyl cysteine), endoplasmic reticulum stress (salubrinal, 4-phenyl butyric acid), or NF-kB (Bay 11-7085) and p38 MAPK (SB203580) were used. Levels of Ninj-1, TNFR1, monocyte adhesion, endoplasmic reticulum stress (ERS) sensors, NADPH oxidase- and mitochondria-derived oxidative species were evaluated. KEY FINDINGS: The novel findings that we report here are: (i) silencing the endothelial TNFR1 leads to decreased Ninj-1 expression and diminished monocyte adhesion; (ii) increased oxidative stress, ERS and NF-kB activation enhance Ninj-1 expression and monocyte adhesion; (iii) up-regulation of endothelial Ninj-1 expression stimulates monocytes adhesion to TNFα - activated HEC; (iv) AML diminishes monocyte adhesion by reducing Ninj-1 expression through mechanisms involving the decrease of NADPH oxidase and mitochondria-dependent oxidative stress, ERS and NF-kB. In addition, AML alleviates apoptosis by reducing the pro-apoptotic CHOP expression and re-establishing the mitochondrial transmembrane potential. SIGNIFICANCE: The results of the present study suggest that Ninj-1 and the proteins involved in its regulation can be considered therapeutic targets for the alleviation of inflammation- dependent disorders. In addition, we demonstrate that some of the benefic effects of AML can be achieved through regulation of Ninj-1.


Assuntos
Anlodipino/farmacologia , Moléculas de Adesão Celular Neuronais/fisiologia , Adesão Celular/fisiologia , Monócitos/citologia , Fatores de Crescimento Neural/fisiologia , Receptores Tipo I de Fatores de Necrose Tumoral/fisiologia , Fator de Necrose Tumoral alfa/fisiologia , Regulação para Cima , Vasodilatadores/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Inativação Gênica , Células Endoteliais da Veia Umbilical Humana , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , NADPH Oxidases/metabolismo , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral/genética
16.
PLoS One ; 15(3): e0229170, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32210425

RESUMO

Surface protein CD20 serves as the critical target of immunotherapy in various B-cell malignancies for decades, however its biological function and regulation remain largely elusive. Better understanding of CD20 function may help to design improved rational therapies to prevent development of resistance. Using CRISPR/Cas9 technique, we have abrogated CD20 expression in five different malignant B-cell lines. We show that CD20 deletion has no effect upon B-cell receptor signaling or calcium flux. Also B-cell survival and proliferation is unaffected in the absence of CD20. On the contrary, we found a strong defect in actin cytoskeleton polymerization and, consequently, defective cell adhesion and migration in response to homeostatic chemokines SDF1α, CCL19 and CCL21. Mechanistically, we could identify a reduction in chemokine-triggered PYK2 activation, a calcium-activated signaling protein involved in activation of MAP kinases and cytoskeleton regulation. These cellular defects in consequence result in a severely disturbed homing of B cells in vivo.


Assuntos
Actinas/metabolismo , Antígenos CD20/fisiologia , Linfócitos B/fisiologia , Leucemia de Células B/patologia , Linfoma de Células B/patologia , Receptores de Antígenos de Linfócitos B/metabolismo , Animais , Antígenos CD20/genética , Antígenos CD20/metabolismo , Linfócitos B/patologia , Adesão Celular/fisiologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Técnicas de Silenciamento de Genes , Humanos , Leucemia de Células B/metabolismo , Linfoma de Células B/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Polimerização , Multimerização Proteica/fisiologia , Transdução de Sinais/imunologia
17.
PLoS One ; 15(3): e0230380, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32163511

RESUMO

Epidermal morphogenesis and hair follicle (HF) development depend on the ability of keratinocytes to adhere to the basement membrane (BM) and migrate along the extracellular matrix. Integrins are cell-matrix receptors that control keratinocyte adhesion and migration, and are recognized as major regulators of epidermal homeostasis. How integrins regulate the behavior of keratinocytes during epidermal morphogenesis remains insufficiently understood. Here, we show that α-parvin (α-pv), a focal adhesion protein that couples integrins to actin cytoskeleton, is indispensable for epidermal morphogenesis and HF development. Inactivation of the murine α-pv gene in basal keratinocytes results in keratinocyte-BM detachment, epidermal thickening, ectopic keratinocyte proliferation and altered actin cytoskeleton polarization. In vitro, α-pv-null keratinocytes display reduced adhesion to BM matrix components, aberrant spreading and stress fibers formation, and impaired directed migration. Together, our data demonstrate that α-pv controls epidermal homeostasis by facilitating integrin-mediated adhesion and actin cytoskeleton organization in keratinocytes.


Assuntos
Membrana Basal/metabolismo , Epiderme/crescimento & desenvolvimento , Folículo Piloso/metabolismo , Queratinócitos/metabolismo , Proteínas dos Microfilamentos/fisiologia , Morfogênese/fisiologia , Actinas/metabolismo , Animais , Membrana Basal/citologia , Adesão Celular/fisiologia , Movimento Celular/fisiologia , Células Cultivadas , Adesões Focais/metabolismo , Integrinas/metabolismo , Queratinócitos/citologia , Camundongos , Camundongos Transgênicos
19.
Biochem Soc Trans ; 48(1): 83-93, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-32065228

RESUMO

Spatiotemporal control of integrin-mediated cell adhesion to the extracellular matrix (ECM) is critical for physiological and pathological events in multicellular organisms, such as embryonic development, angiogenesis, platelet aggregation, leukocytes extravasation, and cancer cell metastatic dissemination. Regulation of integrin adhesive function and signaling relies on the modulation of both conformation and traffic. Indeed, integrins exist in a dynamic equilibrium between a bent/closed (inactive) and an extended/open (active) conformation, respectively endowed with low and high affinity for ECM ligands. Increasing evidence proves that, differently to what hypothesized in the past, detachment from the ECM and conformational inactivation are not mandatory for integrin to get endocytosed and trafficked. Specific transmembrane and cytosolic proteins involved in the control of ECM proteolytic fragment-bound active integrin internalization and recycling exist. In the complex masterplan that governs cell behavior, active integrin traffic is key to the turnover of ECM polymers and adhesion sites, the polarized secretion of endogenous ECM proteins and modifying enzymes, the propagation of motility and survival endosomal signals, and the control of cell metabolism.


Assuntos
Endocitose/fisiologia , Integrinas/química , Integrinas/metabolismo , Transporte Proteico/fisiologia , Animais , Adesão Celular/fisiologia , Membrana Celular/metabolismo , Movimento Celular , Endossomos/metabolismo , Matriz Extracelular/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Humanos , Ligantes , Conformação Proteica
20.
Nat Commun ; 11(1): 1071, 2020 02 26.
Artigo em Inglês | MEDLINE | ID: mdl-32103027

RESUMO

The adhesion of soft connective tissues (tendons, ligaments, and cartilages) on bones in many animals can maintain high toughness (∽800 J m-2) over millions of cycles of mechanical loads. Such fatigue-resistant adhesion has not been achieved between synthetic hydrogels and engineering materials, but is highly desirable for diverse applications such as artificial cartilages and tendons, robust antifouling coatings, and hydrogel robots. Inspired by the nanostructured interfaces between tendons/ligaments/cartilages and bones, we report that bonding ordered nanocrystalline domains of synthetic hydrogels on engineering materials can give a fatigue-resistant adhesion with an interfacial fatigue threshold of 800 J m-2, because the fatigue-crack propagation at the interface requires a higher energy to fracture the ordered nanostructures than amorphous polymer chains. Our method enables fatigue-resistant hydrogel coatings on diverse engineering materials with complex geometries. We further demonstrate that the fatigue-resistant hydrogel coatings exhibit low friction and low wear against natural cartilages.


Assuntos
Cartilagem/fisiologia , Hidrogéis/química , Teste de Materiais , Tíbia/fisiologia , Engenharia Tecidual/métodos , Animais , Adesão Celular/fisiologia , Galinhas , Modelos Moleculares , Propriedades de Superfície
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA