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1.
J Immunol ; 209(1): 136-144, 2022 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-35715008

RESUMO

Neutrophil granulocytes are the first and robust responders to the chemotactic molecules released from an inflamed acidic tissue. The aim of this study was to elucidate the role of microenvironmental pH in neutrophil chemotaxis. To this end, we used neutrophils from male C57BL/6J mice and combined live cell imaging chemotaxis assays with measurements of the intracellular pH (pHi) in varied extracellular pH (pHe). Observational studies were complemented by biochemical analyses of leukotriene B4 (LTB4) production and activation of the Cdc42 Rho GTPase. Our data show that pHi of neutrophils dose-dependently adapts to a given pH of the extracellular milieu. Neutrophil chemotaxis toward C5a has an optimum at pHi ∼7.1, and its pHi dependency is almost parallel to that of LTB4 production. Consequently, a shallow pHe gradient, resembling that encountered by neutrophils during extravasation from a blood vessel (pH ∼7.4) into the interstitium (pH ∼7.2), favors chemotaxis of stimulated neutrophils. Lowering pHe below pH 6.8, predominantly affects neutrophil chemotaxis, although the velocity is largely maintained. Inhibition of the Na+/H+ exchanger 1 (NHE1) with cariporide drastically attenuates neutrophil chemotaxis at the optimal pHi irrespective of the high LTB4 production. Neutrophil migration and chemotaxis are almost completely abrogated by inhibiting LTB4 production or blocking its receptor (BLT1). The abundance of the active GTP-bound form of Cdc42 is strongly reduced by NHE1 inhibition or pHe 6.5. In conclusion, we propose that the pH dependence of neutrophil chemotaxis toward C5a is caused by a pHi-dependent production of LTB4 and activation of Cdc42. Moreover, it requires the activity of NHE1.


Assuntos
Leucotrieno B4 , Neutrófilos , Animais , Quimiotaxia , Quimiotaxia de Leucócito , Concentração de Íons de Hidrogênio , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/fisiologia
2.
J Cell Biol ; 221(8)2022 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-35731205

RESUMO

Migrating cells must interpret chemical gradients to guide themselves within tissues. A long-held principle is that gradients guide cells via reorientation of leading-edge protrusions. However, recent evidence indicates that protrusions can be dispensable for locomotion in some contexts, raising questions about how cells interpret endogenous gradients in vivo and whether other mechanisms are involved. Using laser wound assays in zebrafish to elicit acute endogenous gradients and quantitative analyses, we demonstrate a two-stage process for leukocyte chemotaxis in vivo: first a "search" phase, with stimulation of actin networks at the leading edge, cell deceleration, and turning. This is followed by a "run" phase, with fast actin flows, cell acceleration, and persistence. When actin dynamics are perturbed, cells fail to resolve the gradient, suggesting that pure spatial sensing of the gradient is insufficient for navigation. Our data suggest that cell contractility and actin flows provide memory for temporal sensing, while expansion of the leading edge serves to enhance gradient sampling.


Assuntos
Actinas , Quimiotaxia de Leucócito , Leucócitos , Peixe-Zebra , Animais , Leucócitos/citologia
3.
Nature ; 607(7919): 585-592, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35732737

RESUMO

The regenerative potential of mammalian peripheral nervous system neurons after injury is critically limited by their slow axonal regenerative rate1. Regenerative ability is influenced by both injury-dependent and injury-independent mechanisms2. Among the latter, environmental factors such as exercise and environmental enrichment have been shown to affect signalling pathways that promote axonal regeneration3. Several of these pathways, including modifications in gene transcription and protein synthesis, mitochondrial metabolism and the release of neurotrophins, can be activated by intermittent fasting (IF)4,5. However, whether IF influences the axonal regenerative ability remains to be investigated. Here we show that IF promotes axonal regeneration after sciatic nerve crush in mice through an unexpected mechanism that relies on the gram-positive gut microbiome and an increase in the gut bacteria-derived metabolite indole-3-propionic acid (IPA) in the serum. IPA production by Clostridium sporogenes is required for efficient axonal regeneration, and delivery of IPA after sciatic injury significantly enhances axonal regeneration, accelerating the recovery of sensory function. Mechanistically, RNA sequencing analysis from sciatic dorsal root ganglia suggested a role for neutrophil chemotaxis in the IPA-dependent regenerative phenotype, which was confirmed by inhibition of neutrophil chemotaxis. Our results demonstrate the ability of a microbiome-derived metabolite, such as IPA, to facilitate regeneration and functional recovery of sensory axons through an immune-mediated mechanism.


Assuntos
Indóis , Regeneração Nervosa , Propionatos , Cicatrização , Animais , Axônios/efeitos dos fármacos , Axônios/fisiologia , Quimiotaxia de Leucócito , Clostridium/metabolismo , Jejum , Gânglios Espinais/metabolismo , Microbioma Gastrointestinal , Indóis/sangue , Indóis/metabolismo , Indóis/farmacologia , Camundongos , Compressão Nervosa , Fatores de Crescimento Neural/metabolismo , Regeneração Nervosa/efeitos dos fármacos , Neutrófilos/citologia , Neutrófilos/imunologia , Propionatos/sangue , Propionatos/metabolismo , Propionatos/farmacologia , Recuperação de Função Fisiológica , Nervo Isquiático/lesões , Análise de Sequência de RNA , Cicatrização/efeitos dos fármacos
4.
Int J Mol Sci ; 23(12)2022 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-35742962

RESUMO

Proinflammatory chemokine ligand 26 (CCL26, eotaxin-3) mediates transendothelial cell migration of eosinophils by binding and activating the G-protein-coupled (GPC) chemokine receptor 3 on the surface of eosinophilic cells. Here we have investigated the role of glycosaminoglycans (GAGs) as potential co-receptors in the process of CCL26-induced eosinophil chemotaxis. For this purpose, we have first identified the GAG-binding site of CCL26 by a site-directed mutagenesis approach in the form of an alanine screening. A panel of GAG-binding-deficient mutants has been designed, generated, and analyzed with respect to their binding affinities to heparan sulphate (HS) by isothermal fluorescence titration studies. This showed that basic amino acids in the α-helical part of CCL26 are strongly involved in GAG-binding. In chemotaxis experiments, we found that decreased GAG-binding affinity correlated with decreased chemotactic activity, which indicates an involvement of GAGs in eosinophil migration. This was further proven by the negative impact of heparinase III treatment and, independently, by the incubation of eosinophils with an anti heparan sulfate antibody. We finally investigated eosinophils' proteoglycan (PG) expression patterns by real-time PCR, which revealed the highest expression level for serglycin. Including an anti-serglycin antibody in CCL26-induced eosinophil migration experiments reduced the chemotaxis of these immune cells, thereby proving the dependence of eosinophil mobilization on the proteoglycan serglycin.


Assuntos
Quimiotaxia , Eosinófilos , Quimiocina CCL26 , Quimiocinas CC/metabolismo , Quimiotaxia de Leucócito , Glicosaminoglicanos/metabolismo , Heparitina Sulfato/metabolismo , Proteoglicanas/metabolismo
5.
Clin Immunol ; 238: 108994, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35390547

RESUMO

Chemotaxis is the directed movement of neutrophils towards an infected site. This physiological process can be reproduced using a modified Boyden chamber, such as the Transwell® support. Different techniques can be used to count neutrophils after migration to the lower chamber of the holder. The present study supports the use of an optimized Transwell® assay coupled with a flow cytometry-based method (Sysmex XN-9000) to detect chemotaxis abnormalities. A reference interval of neutrophil's chemotaxis was determined as part of this work. A first step involves the extraction of neutrophils from whole blood. The migration of neutrophils from the upper to the lower support chamber is subsequently directed by a chemoattractant gradient using N-formyl-l-Methionyl-l-Leucyl-l-Phenylalanine (fMLP). Neutrophils collected in the lower chamber are finally counted by flow cytometry. The original protocol was optimized through the comparison of different parameters. The use of Polymorphprep®, in the extraction of neutrophils, showed an improvement of the neutrophils yield of 1.65 times (57.5% of recovery) compared to the extraction using the Ficoll-Hypaque® gradient. A solution containing 5% of Bovin Serum Albumin (BSA) was used to suspend the extracted neutrophils, stabilize their viability and preserve their integrity. The mechanical agitation of the Transwell® permeable supports during migration did not show an increase in neutrophil yield. A migration time of 1 h 30 was identified as the best time for collecting the largest number of neutrophils after migration. Finally, we demonstrated that scraping the bottom of the well after migration improved neutrophil collection from the lower chamber by 1.9-fold compared to a non-scraping method. In conclusion, our results support the use of Polymorphprep® and a 5% BSA solution in the suspension, without agitation of the medium. An incubation time of 1 h 30 was identified as optimal for neutrophil migration through the chamber. Scraping the bottom after neutrophil migration improved neutrophil collection yield. Normal adult values were obtained with directed migration equal to 32.4% ±13.41% on 15 men and 18 women.


Assuntos
Quimiotaxia , Neutrófilos , Adulto , Quimiotaxia de Leucócito/fisiologia , Feminino , Citometria de Fluxo , Humanos , Masculino , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Neutrófilos/fisiologia
6.
J Exp Med ; 219(3)2022 03 07.
Artigo em Inglês | MEDLINE | ID: mdl-35195682

RESUMO

Leukocyte trafficking between blood and tissues is an essential function of the immune system that facilitates humoral and cellular immune responses. Within tissues, leukocytes perform surveillance and effector functions via cell motility and migration toward sites of tissue damage, infection, or inflammation. Neurotransmitters that are produced by the nervous system influence leukocyte trafficking around the body and the interstitial migration of immune cells in tissues. Neural regulation of leukocyte dynamics is influenced by circadian rhythms and altered by stress and disease. This review examines current knowledge of neuro-immune interactions that regulate leukocyte migration and consequences for protective immunity against infections and cancer.


Assuntos
Leucócitos/imunologia , Neuroimunomodulação/imunologia , Movimento Celular/imunologia , Quimiotaxia de Leucócito/imunologia , Ritmo Circadiano/imunologia , Humanos , Modelos Imunológicos , Modelos Neurológicos , Vias Neurais/imunologia , Sistema Nervoso Simpático/imunologia , Microambiente Tumoral/imunologia
7.
Elife ; 112022 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-35112667

RESUMO

Infection is a major co-morbidity that contributes to impaired healing in diabetic wounds. Although impairments in diabetic neutrophils have been blamed for this co-morbidity, what causes these impairments and whether they can be overcome, remain largely unclear. Diabetic neutrophils, isolated from diabetic individuals, exhibit chemotaxis impairment but this peculiar functional impairment has been largely ignored because it appears to contradict the clinical findings which blame excessive neutrophil influx as a major impediment to healing in chronic diabetic ulcers. Here, we report that exposure to glucose in diabetic range results in impaired chemotaxis signaling through the formyl peptide receptor (FPR) in neutrophils, culminating in reduced chemotaxis and delayed neutrophil trafficking in the wound of Leprdb (db/db) type two diabetic mice, rendering diabetic wound vulnerable to infection. We further show that at least some auxiliary receptors remain functional under diabetic conditions and their engagement by the pro-inflammatory cytokine CCL3, overrides the requirement for FPR signaling and substantially improves infection control by jumpstarting the neutrophil trafficking toward infection, and stimulates healing in diabetic wound. We posit that CCL3 may have therapeutic potential for the treatment of diabetic foot ulcers if it is applied topically after the surgical debridement process which is intended to reset chronic ulcers into acute fresh wounds.


Assuntos
Quimiotaxia de Leucócito/imunologia , Diabetes Mellitus Experimental/imunologia , Neutrófilos/patologia , Receptores de Formil Peptídeo/genética , Transdução de Sinais/imunologia , Cicatrização/imunologia , Infecção dos Ferimentos/microbiologia , Animais , Quimiocina CCL3/imunologia , Complicações do Diabetes/microbiologia , Glucose/administração & dosagem , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/imunologia , Receptores de Formil Peptídeo/imunologia , Infecção dos Ferimentos/tratamento farmacológico , Infecção dos Ferimentos/etiologia
8.
J Leukoc Biol ; 111(6): 1175-1184, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35100458

RESUMO

Following injury and infection, neutrophils are guided to the affected site by chemoattractants released from injured tissues and invading microbes. During this process (chemotaxis), neutrophils must integrate multiple chemical signals, while also responding to physical constraints and prioritizing their directional decisions to generate an efficient immune response. In some clinical conditions, human neutrophils appear to lose the ability to chemotax efficiently, which may contribute both directly and indirectly to disease pathology. Here, a range of microfluidic designs is utilized to test the sensitivity of chemotaxing neutrophils to various perturbations, including binary decision-making in the context of channels with different chemoattractant gradients, hydraulic resistance, and angle of approach. Neutrophil migration in long narrow channels and planar environments is measured. Conditions in which neutrophils are significantly more likely to choose paths with the steepest chemoattractant gradient and the most direct approach angle, and find that migration efficiency across planar chambers is inversely correlated with chamber diameter. By sequential measurement of neutrophil binary decision-making to different chemoattractant gradients, or chemotactic index in sequential planar environments, data supporting a model of biased random walk for neutrophil chemotaxis are presented.


Assuntos
Quimiotaxia , Neutrófilos , Movimento Celular/fisiologia , Fatores Quimiotáticos/farmacologia , Quimiotaxia/fisiologia , Quimiotaxia de Leucócito/fisiologia , Humanos , Neutrófilos/fisiologia
9.
Cancer Lett ; 529: 112-125, 2022 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-34999169

RESUMO

Though circulating monocytes are the main source of tumour-associated macrophages (TAMs), the regulatory mechanisms of their recruitment to tumours and further differentiation remain unclear. In the present study, we observed a significant decrease in CXCR2 expression in classical circulating monocytes of patients with colorectal cancer (CRC), particularly those in the late TNM stage. The percentage of CXCR2+ monocytes was negatively associated with systemic inflammatory markers and positively associated with intratumoural immunocyte infiltration. The pro-inflammatory cytokine IFN-γ, which was overexpressed in patients with CRC, down-regulated CXCR2 expression of monocytes/TAMs by promoting GRK-2 expression. In vitro, inhibition of CXCR2 signalling in monocytes led to impaired chemotaxis to the tumour cell line supernatant and lower responsiveness to lipopolysaccharide (LPS) stimulation. Finally, monocytes from patients with CRC with decreased CXCR2 expression showed distinct phenotypes and functions after differentiating into CRC cell line-educated TAMs, including expression of co-stimulatory factors and secretion profile, than those from healthy controls. GRK-2 inhibitor altered the functional characteristics of TAMs. In summary, our findings suggest that CXCR2 expression on circulating monocytes reflects CRC stages and is an important factor determining TAM composition in the tumour microenvironment.


Assuntos
Quimiotaxia de Leucócito/genética , Monócitos/metabolismo , Receptores de Interleucina-8B/genética , Macrófagos Associados a Tumor/imunologia , Macrófagos Associados a Tumor/metabolismo , Idoso , Idoso de 80 Anos ou mais , Biomarcadores , Linhagem Celular Tumoral , Quimiotaxia de Leucócito/imunologia , Neoplasias Colorretais/etiologia , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Feminino , Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Monócitos/imunologia , Estadiamento de Neoplasias , Receptores de Interleucina-8B/metabolismo , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia
10.
Sci Rep ; 12(1): 685, 2022 01 13.
Artigo em Inglês | MEDLINE | ID: mdl-35027618

RESUMO

Low-density neutrophils (LDNs) have been described in tumors and various autoimmune diseases, where they exhibit immune dysfunction and alter disease progression. Nevertheless, LDNs have been rarely reported in sepsis. We studied sepsis patients admitted to the intensive care unit. Wright-Giemsa stain assay and Transmission electron microscopy were performed to detect the morphology of neutrophils. Flow cytometry was used to analyze the number and function of LDNs. Concentration of cytokines was measured using ELISA. Neutrophil chemotaxis was examined using an under-agarose chemotaxis model. We found that LDNs were significantly elevated in patients with sepsis. Phenotypes and morphological characteristics suggest that LDNs may be formed by mixtures of neutrophils at various maturation stages. In vitro experiments showed that LDN formation was closely associated with neutrophil degranulation. We preliminarily discussed changes in immune function in LDNs. Compared with high-density neutrophils, expression levels of CXC chemokine receptor 4 on LDN surfaces were increased, phagocytotic capacity was decreased, and life span was prolonged. The chemotactic ability of LDNs was significantly reduced, possibly related to the increased expression of P2X1. These data suggest that LDNs are essential components of neutrophils in sepsis. To clarify the source and dysfunction mechanism of LDN in sepsis may be helpful for the diagnosis and treatment of sepsis in the future.


Assuntos
Neutrófilos/patologia , Neutrófilos/fisiologia , Sepse/sangue , Adulto , Idoso , Degranulação Celular , Quimiotaxia de Leucócito , Citocinas/sangue , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Neutrófilos/imunologia , Fagocitose , Receptores Purinérgicos P2X1/metabolismo , Sepse/diagnóstico
11.
Cell Death Dis ; 13(1): 33, 2022 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-35013119

RESUMO

Aged microglia display augmented inflammatory activity after neural injury. Although aging is a risk factor for poor outcome after brain insults, the precise impact of aging-related alterations in microglia on neural injury remains poorly understood. Microglia can be eliminated via pharmacological inhibition of the colony-stimulating factor 1 receptor (CSF1R). Upon withdrawal of CSF1R inhibitors, microglia rapidly repopulate the entire brain, leading to replacement of the microglial compartment. In this study, we investigated the impact of microglial replacement in the aged brain on neural injury using a mouse model of intracerebral hemorrhage (ICH) induced by collagenase injection. We found that replacement of microglia in the aged brain reduced neurological deficits and brain edema after ICH. Microglial replacement-induced attenuation of ICH injury was accompanied with alleviated blood-brain barrier disruption and leukocyte infiltration. Notably, newly repopulated microglia had reduced expression of IL-1ß, TNF-α and CD86, and upregulation of CD206 in response to ICH. Our findings suggest that replacement of microglia in the aged brain restricts neuroinflammation and brain injury following ICH.


Assuntos
Envelhecimento/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Hemorragia Cerebral/tratamento farmacológico , Microglia/efeitos dos fármacos , /tratamento farmacológico , Envelhecimento/patologia , Aminopiridinas/administração & dosagem , Aminopiridinas/farmacologia , Animais , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/patologia , Encéfalo/imunologia , Encéfalo/patologia , Lesões Encefálicas/tratamento farmacológico , Lesões Encefálicas/etiologia , Lesões Encefálicas/imunologia , Lesões Encefálicas/patologia , Morte Celular/efeitos dos fármacos , Hemorragia Cerebral/complicações , Hemorragia Cerebral/imunologia , Hemorragia Cerebral/patologia , Quimiotaxia de Leucócito/efeitos dos fármacos , Modelos Animais de Doenças , Camundongos , Microglia/imunologia , Microglia/patologia , /imunologia , Pirróis/administração & dosagem , Pirróis/farmacologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores
12.
Infect Immun ; 90(1): e0051621, 2022 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-34662214

RESUMO

Salmonella invades and disrupts gut epithelium integrity, creating an infection-generated electric field that can drive directional migration of macrophages, a process called galvanotaxis. Phagocytosis of bacteria reverses the direction of macrophage galvanotaxis, implicating a bioelectrical mechanism to initiate life-threatening disseminations. The force that drives direction reversal of macrophage galvanotaxis is not understood. One hypothesis is that Salmonella can alter the electrical properties of the macrophages by modifying host cell surface glycan composition, which is supported by the fact that cleavage of surface-exposed sialic acids with a bacterial neuraminidase severely impairs macrophage galvanotaxis, as well as phagocytosis. Here, we utilize N-glycan profiling by nanoLC-chip QTOF mass cytometry to characterize the bacterial neuraminidase-associated compositional shift of the macrophage glycocalyx, which revealed a decrease in sialylated and an increase in fucosylated and high mannose structures. The Salmonella nanH gene, encoding a putative neuraminidase, is required for invasion and internalization in a human colonic epithelial cell infection model. To determine whether NanH is required for the Salmonella infection-dependent direction reversal, we constructed and characterized a nanH deletion mutant and found that NanH is partially required for Salmonella infection in primary murine macrophages. However, compared to wild type Salmonella, infection with the nanH mutant only marginally reduced the cathode-oriented macrophage galvonotaxis, without canceling direction reversal. Together, these findings strongly suggest that while neuraminidase-mediated N-glycan modification impaired both macrophage phagocytosis and galvanotaxis, yet to be defined mechanisms other than NanH may play a more important role in bioelectrical control of macrophage trafficking, which potentially triggers dissemination.


Assuntos
Quimiotaxia de Leucócito/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Neuraminidase/metabolismo , Infecções por Salmonella/imunologia , Infecções por Salmonella/metabolismo , Salmonella/fisiologia , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Feminino , Interações Hospedeiro-Patógeno/imunologia , Masculino , Camundongos , Modelos Biológicos , Mutação , Fagocitose/imunologia , Polissacarídeos/metabolismo , Infecções por Salmonella/microbiologia , Ácidos Siálicos/metabolismo , Virulência
13.
Cardiovasc Res ; 118(1): 169-183, 2022 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-33560342

RESUMO

AIMS: Myocardial infarction (MI) is the most common cause of heart failure (HF) worldwide. G protein-coupled receptor kinase 5 (GRK5) is upregulated in failing human myocardium and promotes maladaptive cardiac hypertrophy in animal models. However, the role of GRK5 in ischemic heart disease is still unknown. In this study, we evaluated whether myocardial GRK5 plays a critical role post-MI in mice and included the examination of specific cardiac immune and inflammatory responses. METHODS AND RESULTS: Cardiomyocyte-specific GRK5 overexpressing transgenic mice (TgGRK5) and non-transgenic littermate control (NLC) mice as well as cardiomyocyte-specific GRK5 knockout mice (GRK5cKO) and wild type (WT) were subjected to MI and, functional as well as structural changes together with outcomes were studied. TgGRK5 post-MI mice showed decreased cardiac function, augmented left ventricular dimension and decreased survival rate compared to NLC post-MI mice. Cardiac hypertrophy and fibrosis as well as fetal gene expression were increased post-MI in TgGRK5 compared to NLC mice. In TgGRK5 mice, GRK5 elevation produced immuno-regulators that contributed to the elevated and long-lasting leukocyte recruitment into the injured heart and ultimately to chronic cardiac inflammation. We found an increased presence of pro-inflammatory neutrophils and macrophages as well as neutrophils, macrophages and T-lymphocytes at 4-days and 8-weeks respectively post-MI in TgGRK5 hearts. Conversely, GRK5cKO mice were protected from ischemic injury and showed reduced early immune cell recruitment (predominantly monocytes) to the heart, improved contractility and reduced mortality compared to WT post-MI mice. Interestingly, cardiomyocyte-specific GRK2 transgenic mice did not share the same phenotype of TgGRK5 mice and did not have increased cardiac leukocyte migration and cytokine or chemokine production post-MI. CONCLUSIONS: Our study shows that myocyte GRK5 has a crucial and GRK-selective role on the regulation of leucocyte infiltration into the heart, cardiac function and survival in a murine model of post-ischemic HF, supporting GRK5 inhibition as a therapeutic target for HF.


Assuntos
Quimiotaxia de Leucócito , Quinase 5 de Receptor Acoplado a Proteína G/metabolismo , Insuficiência Cardíaca/enzimologia , Leucócitos/metabolismo , Infarto do Miocárdio/enzimologia , Miócitos Cardíacos/enzimologia , Função Ventricular Esquerda , Animais , Citocinas/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Quinase 5 de Receptor Acoplado a Proteína G/genética , Insuficiência Cardíaca/imunologia , Insuficiência Cardíaca/patologia , Insuficiência Cardíaca/fisiopatologia , Mediadores da Inflamação/metabolismo , Leucócitos/imunologia , Camundongos Knockout , Contração Miocárdica , Infarto do Miocárdio/imunologia , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Miócitos Cardíacos/imunologia , Miócitos Cardíacos/patologia , Transdução de Sinais , Volume Sistólico , Transcriptoma , Pressão Ventricular
14.
Fertil Steril ; 117(1): 160-168, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34656305

RESUMO

OBJECTIVE: To characterize T lymphocyte infiltration and programmed cell death protein-1 (PD-1)/programmed death-ligand 1 (PD-L1) expression in early-stage endometriosis-associated ovarian cancer (EAOC), ovarian endometriosis (OE), atypical endometriosis (AE), and deep endometriosis (DE). DESIGN: Case-control, retrospective study. SETTING: Research University Hospital. PATIENT(S): A total of 362 patients with a histologic diagnosis of EAOC, OE, AE, or DE were identified between 2000 and 2019 from Fondazione Policlinico Universitario Agostino Gemelli IRCCS and Gemelli Molise SpA tissue data banks. A 1:1 propensity score-matched method yielded matched pairs of 55 subjects with EAOC, 55 patients with OE, 12 patients with AE, and 42 patients with DE, resulting in no differences in family history of cancer, parity, and use of oral contraceptives. INTERVENTION(S): Immunohistochemistry assays using the following primary antibodies: CD3+; CD4+; CD8+; PD-1; and PD-L1. MAIN OUTCOME MEASURE(S): To characterize T lymphocyte infiltration and PD-1/PD-L1 expression in 4 different endometriosis-related diseases. RESULT(S): Endometriosis-associated ovarian cancer cases displayed significantly higher levels of PD-1/PD-L1 expression compared with all other endometriosis-related diseases (vs. OE vs. AE vs. DE). Moreover, a significantly lower count of infiltrating T lymphocytes was observed in EAOC cases compared with OE ones. Finally, one-third of OE cases showed a cancer-like PD-1/PD-L1 expression profile. CONCLUSION(S): Endometriosis-associated ovarian cancer is characterized by higher levels of PD-1/PD-L1 expression compared with benign endometriosis-related diseases. This profile was found in one-third of clinically benign cases, suggesting that it develops early in the carcinogenesis process.


Assuntos
Endometriose/complicações , Neoplasias Ovarianas/etiologia , Receptor de Morte Celular Programada 1/metabolismo , Linfócitos T/patologia , Adulto , Antígeno B7-H1/metabolismo , Carcinoma Epitelial do Ovário/etiologia , Carcinoma Epitelial do Ovário/imunologia , Carcinoma Epitelial do Ovário/metabolismo , Estudos de Casos e Controles , Movimento Celular/imunologia , Quimiotaxia de Leucócito/fisiologia , Endometriose/imunologia , Endometriose/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Itália , Pessoa de Meia-Idade , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/metabolismo , Estudos Retrospectivos , Linfócitos T/fisiologia
15.
J Leukoc Biol ; 112(2): 273-278, 2022 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34939227

RESUMO

Increased levels of the anti-inflammatory peptide Catestatin (CST), a cleavage product of the pro-hormone chromogranin A, correlate with less severe outcomes in hypertension, colitis, and diabetes. However, it is unknown how CST reduces the infiltration of monocytes and macrophages (Mϕs) in inflamed tissues. Here, it is reported that CST blocks leukocyte migration toward inflammatory chemokines. By in vitro and in vivo migration assays, it is shown that although CST itself is chemotactic, it blocks migration of monocytes and neutrophils to inflammatory attracting factor CC-chemokine ligand 2 (CCL2) and C-X-C motif chemokine ligand 2 (CXCL2). Moreover, it directs CX3 CR1+ Mϕs away from pancreatic islets. These findings suggest that the anti-inflammatory actions of CST are partly caused by its regulation of chemotaxis.


Assuntos
Quimiotaxia de Leucócito , Quimiotaxia , Anti-Inflamatórios/farmacologia , Quimiocina CCL2/farmacologia , Quimiocinas/farmacologia , Cromogranina A/farmacologia , Ligantes , Neutrófilos , Fragmentos de Peptídeos , Peptídeos/farmacologia
16.
Front Immunol ; 12: 775447, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34858434

RESUMO

CD11d/CD18 is the most recently discovered and least understood ß2 integrin. Known CD11d adhesive mechanisms contribute to both extravasation and mesenchymal migration - two key aspects for localizing peripheral leukocytes to sites of inflammation. Differential expression of CD11d induces differences in monocyte/macrophage mesenchymal migration including impacts on macrophage sub-set migration. The participation of CD11d/CD18 in leukocyte localization during atherosclerosis and following neurotrauma has sparked interest in the development of CD11d-targeted therapeutic agents. Whereas the adhesive properties of CD11d have undergone investigation, the signalling pathways induced by ligand binding remain largely undefined. Underlining each adhesive and signalling function, CD11d is under unique transcriptional control and expressed on a sub-set of predominately tissue-differentiated innate leukocytes. The following review is the first to capture the nearly three decades of CD11d research and discusses the emerging role of CD11d in leukocyte migration and retention during the progression of a staged immune response.


Assuntos
Antígenos CD11/genética , Antígenos CD18/genética , Quimiotaxia de Leucócito/genética , Quimiotaxia de Leucócito/imunologia , Regulação da Expressão Gênica , Cadeias alfa de Integrinas/genética , Leucócitos/fisiologia , Animais , Antígenos CD11/química , Antígenos CD11/metabolismo , Antígenos CD18/química , Antígenos CD18/metabolismo , Suscetibilidade a Doenças , Desenvolvimento de Medicamentos , Humanos , Cadeias alfa de Integrinas/química , Cadeias alfa de Integrinas/metabolismo , Linfopoese/genética , Terapia de Alvo Molecular , Especificidade de Órgãos/genética , Fagocitose/genética , Fagocitose/imunologia , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Processamento de Proteína Pós-Traducional , Relação Estrutura-Atividade , Fatores de Transcrição
17.
Front Immunol ; 12: 789454, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34868077

RESUMO

Herpes simplex virus type-1 (HSV-1) ocular infection is one of the leading causes of infectious blindness in developed countries. The resultant herpetic keratitis (HK) is caused by an exacerbated reaction of the adaptive immune response that persists beyond virus clearance causing substantial damage to the cornea. Intramuscular immunization of mice with the HSV-1(VC2) live-attenuated vaccine strain has been shown to protect mice against lethal ocular challenge. Herein, we show that following ocular challenge, VC2 vaccinated animals control ocular immunopathogenesis in the absence of neutralizing antibodies on ocular surfaces. Ocular protection is associated with enhanced intracorneal infiltration of γδ T cells compared to mock-vaccinated animals. The observed γδ T cellular infiltration was inversely proportional to the infiltration of neutrophils, the latter associated with exacerbated tissue damage. Inhibition of T cell migration into ocular tissues by the S1P receptors agonist FTY720 produced significant ocular disease in vaccinated mice and marked increase in neutrophil infiltration. These results indicate that ocular challenge of mice immunized with the VC2 vaccine induce a unique ocular mucosal response that leads into the infiltration of γδ T cells resulting in the amelioration of infection-associated immunopathogenesis.


Assuntos
Quimiotaxia de Leucócito , Córnea/imunologia , Vacinas contra o Vírus do Herpes Simples/administração & dosagem , Herpesvirus Humano 1/imunologia , Linfócitos Intraepiteliais/imunologia , Ceratite Herpética/prevenção & controle , Vacinação , Animais , Córnea/patologia , Córnea/virologia , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Vacinas contra o Vírus do Herpes Simples/imunologia , Herpesvirus Humano 1/patogenicidade , Interações Hospedeiro-Patógeno , Injeções Intramusculares , Linfócitos Intraepiteliais/virologia , Ceratite Herpética/imunologia , Ceratite Herpética/patologia , Ceratite Herpética/virologia , Linfangiogênese , Camundongos Endogâmicos BALB C , Neovascularização Patológica , Infiltração de Neutrófilos , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/imunologia
18.
Front Immunol ; 12: 779085, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34880871

RESUMO

Strict control of B lymphocyte development is required for the ability to mount humoral immune responses to diverse foreign antigens while remaining self-tolerant. In the bone marrow, B lineage cells transit through several developmental stages in which they assemble a functional B cell receptor in a stepwise manner. The immunoglobulin heavy chain gene is rearranged at the pro-B stage. At the large pre-B stage, cells with a functional heavy chain expand in response to signals from IL-7 and the pre-BCR. Cells then cease proliferation at the small pre-B stage and rearrange the immunoglobulin light chain gene. The fully formed BCR is subsequently expressed on the surface of immature B cells and autoreactive cells are culled by central tolerance mechanisms. Once in the periphery, transitional B cells develop into mature B cell subsets such as marginal zone and follicular B cells. These developmental processes are controlled by transcription factor networks, central to which are IRF4 and IRF8. These were thought to act redundantly during B cell development in the bone marrow, with their functions diverging in the periphery where IRF4 limits the number of marginal zone B cells and is required for germinal center responses and plasma cell differentiation. Because of IRF4's unique role in mature B cells, we hypothesized that it may also have functions earlier in B cell development that cannot be compensated for by IRF8. Indeed, we find that IRF4 has a unique role in upregulating the pre-B cell marker CD25, limiting IL-7 responsiveness, and promoting migration to CXCR4 such that IRF4-deficient mice have a partial block at the pre-B cell stage. We also find that IRF4 acts in early transitional B cells to restrict marginal zone B cell development, as deletion of IRF4 in mature B cells with CD21-cre impairs plasma cell differentiation but has no effect on marginal zone B cell numbers. These studies highlight IRF4 as the dominant IRF family member in early B lymphopoiesis.


Assuntos
Proliferação de Células , Fatores Reguladores de Interferon/metabolismo , Linfopoese , Células Precursoras de Linfócitos B/metabolismo , Receptores de Complemento 3d/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Quimiocina CXCL12/farmacologia , Quimiotaxia de Leucócito , Regulação da Expressão Gênica no Desenvolvimento , Fatores Reguladores de Interferon/genética , Interleucina-7/farmacologia , Linfopoese/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Células Precursoras de Linfócitos B/efeitos dos fármacos , Células Precursoras de Linfócitos B/imunologia , Receptores de Complemento 3d/genética , Transdução de Sinais
19.
Bull Exp Biol Med ; 172(2): 263-269, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34855085

RESUMO

Activation and migration of donor T cells to the host target organs are critical mechanisms in the pathogenesis of graft-versus-host disease (GVHD). The role of monocyte chemoattractant protein-1 (MCP-1/CCL2) and its receptor CCR2 in the recruitment of T cells during immune or inflammatory response is also well known. For elucidation of the mechanism of the therapeutic effect of human bone marrow derived-mesenchymal stem cells (MSC) in GVHD, we studied the effect of these cells on migration of activated donor T cells through the CCL2-CCR2 axis in vitro. MSC were expanded from donors' bone marrow mononuclear cells. After co-culturing of IL-2-activated T cells with allogeneic MSC at different ratios, the levels of CCL2 in supernatants were measured by ELISA, and CCR2 expression in CD4+/CD8+ T cells subsets were detected by flow cytometry. The effect of MSC on the migration of activated T cells in the Transwell system was studied in the absence or presence of CCL2. Our results show that CCL2 levels in supernatants of co-cultures were significantly higher than in MSC monoculture and this increase depended on the number of MSC. MSC inhibited proliferation of T cells, but did not change the percentages of CD4+ and CD8+ T cells subsets. MSC can up-regulate the CCR2 expression in CD8+ subsets rather than in CD4+ subsets; MSC enhanced migration of IL-2-activated T cells to CCL2 by increasing the expression of CCR2. The data demonstrate that MSC can enhance chemotaxis of cytokine-activated T cells through the CCL2-CCR2 axis in vitro.


Assuntos
Quimiotaxia de Leucócito/fisiologia , Células-Tronco Mesenquimais/fisiologia , Linfócitos T/fisiologia , Adulto , Diferenciação Celular/imunologia , Células Cultivadas , Quimiocina CCL2/fisiologia , Técnicas de Cocultura , Humanos , Imunofenotipagem , Ativação Linfocitária , Células-Tronco Mesenquimais/citologia , Receptores CCR2/fisiologia , Transdução de Sinais , Linfócitos T/imunologia
20.
Bull Exp Biol Med ; 172(2): 137-142, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34855095

RESUMO

3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) along with their blood lipid-lowering effect exhibit anti-inflammatory and immunomodulatory activity. We studied the effects of long-term (72-h or longer) exposure of human T lymphocytes in culture to atorvastatin and rosuvastatin (5-80 nM) on their functional activity. Treatment with statins inhibited PHA/IL-2-induced proliferation of CD4+ T lymphocytes isolated from the peripheral blood of healthy donors. This was accompanied by a decrease in the relative content of cells expressing active caspase-3. Addition of mevalonate or fetal bovine serum simultaneously with statins restored proliferative activity of cells. Culturing of CD4+ T lymphocytes with statins in the presence of IL-2 did not significantly affect the expression of chemokine receptors CCR4, CCR5, CXCR3, and CXCR4. Pretreatment with statins suppressed spontaneous and SDF-1-stimulated migration of CD4+ T lymphocytes, but little changed the content of intracellular phosphorylated protein kinases Akt, p38 and p42/44 (ERK1/2). The cellular effects of "lipophilic" atorvastatin were observed at lower concentrations compared to "hydrophilic" rosuvastatin.


Assuntos
Linfócitos T CD4-Positivos/efeitos dos fármacos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Atorvastatina/farmacologia , Linfócitos T CD4-Positivos/fisiologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Quimiotaxia de Leucócito/efeitos dos fármacos , Humanos , Receptores CCR4/metabolismo , Receptores CCR5/metabolismo , Receptores CXCR3/metabolismo , Receptores CXCR4/metabolismo , Rosuvastatina Cálcica/farmacologia , Transdução de Sinais/efeitos dos fármacos
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