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1.
Physiol Rep ; 9(5): e14796, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33687143

RESUMO

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), continues to be a world-wide pandemic with overwhelming socioeconomic impact. Since inflammation is one of the major causes of COVID-19 complications, the associated molecular mechanisms have been the focus of many studies to better understand this disease and develop improved treatments for patients contracting SARS-CoV-2. Among these, strong emphasis has been placed on pro-inflammatory cytokines, associating severity of COVID-19 with so-called "cytokine storm." More recently, peptide bradykinin, its dysregulated signaling or "bradykinin storm," has emerged as a primary mechanism to explain COVID-19-related complications. Unfortunately, this important development may not fully capture the main molecular players that underlie the disease severity. To this end, in this focused review, several lines of evidence are provided to suggest that in addition to bradykinin, two closely related vasoactive peptides, substance P and neurotensin, are also likely to drive microvascular permeability and inflammation, and be responsible for development of COVID-19 pathology. Furthermore, based on published experimental observations, it is postulated that in addition to ACE and neprilysin, peptidase neurolysin (Nln) is also likely to contribute to accumulation of bradykinin, substance P and neurotensin, and progression of the disease. In conclusion, it is proposed that "vasoactive peptide storm" may underlie severity of COVID-19 and that simultaneous inhibition of all three peptidergic systems could be therapeutically more advantageous rather than modulation of any single mechanism alone.


Assuntos
Bradicinina/metabolismo , Neprilisina/metabolismo , Neurotensina/metabolismo , Substância P/metabolismo , Animais , /patologia , Citocinas/metabolismo , Humanos , Microvasos/metabolismo , Microvasos/patologia
2.
Physiol Rep ; 9(3): e14726, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33523608

RESUMO

Corona virus disease 2019 (COVID-19) causes symptoms from multiple organs after infection by severe acute respiratory syndrome corona virus 2 (SARS CoV-2). They range from early, low blood oxygen levels (hypoxemia) without breathlessness ("silent hypoxia"), delirium, rashes, and loss of smell (anosmia), to persisting chest pain, muscle weakness and -pain, fatigue, confusion, memory problems and difficulty to concentrate ("brain fog"), mood changes, and unexpected onset of hypertension or diabetes. SARS CoV-2 affects the microcirculation, causing endothelial cell swelling and damage (endotheliitis), microscopic blood clots (microthrombosis), capillary congestion, and damage to pericytes that are integral to capillary integrity and barrier function, tissue repair (angiogenesis), and scar formation. Similar to other instances of critical illness, COVID-19 is also associated with elevated cytokine levels in the systemic circulation. This review examines how capillary damage and inflammation may contribute to these acute and persisting COVID-19 symptoms by interfering with blood and tissue oxygenation and with brain function. Undetectable by current diagnostic methods, capillary flow disturbances limit oxygen diffusion exchange in lungs and tissue and may therefore cause hypoxemia and tissue hypoxia. The review analyzes the combined effects of COVID-19-related capillary damage, pre-existing microvascular changes, and upstream vascular tone on tissue oxygenation in key organs. It identifies a vicious cycle, as infection- and hypoxia-related inflammation cause capillary function to deteriorate, which in turn accelerates hypoxia-related inflammation and tissue damage. Finally, the review addresses the effects of low oxygen and high cytokine levels in brain tissue on neurotransmitter synthesis and mood. Methods to assess capillary functions in human organs and therapeutic means to protect capillary functions and stimulate capillary bed repair may prove important for the individualized management of COVID-19 patients and targeted rehabilitation strategies.


Assuntos
/complicações , Microvasos/patologia , Consumo de Oxigênio , Oxigênio/metabolismo , Animais , /patologia , Humanos , Inflamação , Microvasos/metabolismo , Microvasos/virologia , Oxigênio/sangue , /patogenicidade
3.
Cells ; 10(2)2021 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-33578631

RESUMO

Clinical manifestations of coronavirus disease 2019 (COVID-19) in pregnant women are diverse, and little is known of the impact of the disease on placental physiology. Severe acute respiratory syndrome coronavirus (SARS-CoV-2) has been detected in the human placenta, and its binding receptor ACE2 is present in a variety of placental cells, including endothelium. Here, we analyze the impact of COVID-19 in placental endothelium, studying by immunofluorescence the expression of von Willebrand factor (vWf), claudin-5, and vascular endothelial (VE) cadherin in the decidua and chorionic villi of placentas from women with mild and severe COVID-19 in comparison to healthy controls. Our results indicate that: (1) vWf expression increases in the endothelium of decidua and chorionic villi of placentas derived from women with COVID-19, being higher in severe cases; (2) Claudin-5 and VE-cadherin expression decrease in the decidua and chorionic villus of placentas from women with severe COVID-19 but not in those with mild disease. Placental histological analysis reveals thrombosis, infarcts, and vascular wall remodeling, confirming the deleterious effect of COVID-19 on placental vessels. Together, these results suggest that placentas from women with COVID-19 have a condition of leaky endothelium and thrombosis, which is sensitive to disease severity.


Assuntos
/complicações , Placenta/irrigação sanguínea , Placenta/patologia , Complicações Cardiovasculares na Gravidez/etiologia , Complicações Infecciosas na Gravidez/etiologia , Trombose/etiologia , Adulto , Antígenos CD/análise , /virologia , Caderinas/análise , Claudina-5/análise , Endotélio/irrigação sanguínea , Endotélio/patologia , Endotélio/virologia , Feminino , Humanos , Recém-Nascido , Microvasos/patologia , Microvasos/virologia , Gravidez , Complicações Cardiovasculares na Gravidez/patologia , Complicações Cardiovasculares na Gravidez/virologia , Complicações Infecciosas na Gravidez/patologia , Complicações Infecciosas na Gravidez/virologia , Trombose/patologia , Trombose/virologia , Adulto Jovem , Fator de von Willebrand/análise
4.
Medicine (Baltimore) ; 100(3): e23985, 2021 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-33545987

RESUMO

PURPOSE: To assess expression levels of Ephrin type-A receptor 2 (EphA2), vascular endothelial growth factor (VEGF), and von Willebrand factor (vWF), and assess their potentials as prognostic biomarkers to predict the risk of poor survival in patients with primary lower grade glioma. METHOD: The study included75 patients with histopathologically confirmed primary glioma (World Health Organization Grade IV). All patients underwent combined surgery and postoperative radiotherapy for the management of primary glioma. Immuno-histochemical analysis was performed to evaluate expression levels ofEphA2 and VEGF. Evaluation of tumor microvessel density was also performed at angiogenesis hot spots due to tumor growth. Main outcomes of the study were the prognostic efficiencies of EphA2, VEGF, and vWF in primary low-grade glioma, as well as whether their expression levels were associated with cancer progression. RESULTS: Of the patients with glioma, 67% had very strong expression of EphA2. Overall survival was inversely correlated with the expression of EphA2. Regarding VEGF expression, 38 patients (51%) had strong expression, 29 patients (39%) had weak expression, and 8 patients (11%) had no expression. Strong VEGF expression was associated with poor prognosis and poor survival. CONCLUSION: EphA2, VEGF, and vWF could be considered prognostic markers for assessment of primary glioma.


Assuntos
Neoplasias do Sistema Nervoso Central/mortalidade , Efrina-A2/metabolismo , Glioma/mortalidade , Fator A de Crescimento do Endotélio Vascular/metabolismo , Fator de von Willebrand/metabolismo , Adulto , Idoso , Biomarcadores Tumorais/metabolismo , Sistema Nervoso Central/irrigação sanguínea , Neoplasias do Sistema Nervoso Central/genética , Feminino , Glioma/genética , Humanos , Imuno-Histoquímica , Masculino , Microvasos/patologia , Pessoa de Meia-Idade , Neovascularização Patológica , Prognóstico
5.
Methods Mol Biol ; 2206: 151-178, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32754817

RESUMO

This protocol focuses on the quantitative description of the angioarchitecture of experimental tumor xenografts. This semiautomatic analysis is carried out on functional vessels and microvessels acquired by confocal imaging and processed into progressively reconstructed angioarchitectures following a caliber-classification step. The protocol can be applied also to the quantification of pathological angioarchitectures other than tumor grafts as well as to the microvasculature of physiological tissue samples.


Assuntos
Microscopia Confocal/métodos , Microvasos/patologia , Neoplasias/patologia , Neovascularização Patológica/patologia , Animais , Xenoenxertos/patologia , Humanos , Camundongos
6.
Microvasc Res ; 133: 104093, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33007316

RESUMO

Acute respiratory distress syndrome (ARDS) is a rapidly progressive disease with unknown pathogenesis. Damage of pulmonary microvascular endothelial cells (PMVECs) caused by inflammatory storm caused by cytokines such as TNF-α is the potential pathogenesis of ARDS. In this study, we examined the role of ezrin and Rac1 in TNF-α-related pathways, which regulates the permeability of PMVECs. Primary rat pulmonary microvascular endothelial cells (RPMVECs) were isolated and cultured. RPMVECs were treated with rat TNF-α (0, 1, 10, 100 ng/ml), and the cell activity of each group was measured using a CCK8 kit. The integrity of endothelial barrier was measured by transendothelial resistance (TEER) and FITC-BSA flux across RPMVECs membranes. Pulldown assay and Western blot was used to detect the activity of RAS-associated C3 botulinum toxin substrate 1 (Rac1) and Ezrin phosphorylation. Short hairpin RNA (shRNA) targeting ezrin and Rac1 was utilized to evaluate the effect of RPMVECs permeability and related pathway. The effects of ezrin and Rac1 on cytoskeleton were confirmed by immunofluorescence. Our results revealed that active Rac1 was essential for protecting the RPMVEC barrier stimulated by TNF-α, while active ezrin could partially destroy the PMVEC barrier by reducing Rac1 activity and regulating the subcellular structure of the cytoskeleton. These findings may be used to create new therapeutic strategies for targeting Rac1 in the treatment of ARDS.


Assuntos
Permeabilidade Capilar/efeitos dos fármacos , Proteínas do Citoesqueleto/metabolismo , Citoesqueleto/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Pulmão/irrigação sanguínea , Microvasos/efeitos dos fármacos , Fator de Necrose Tumoral alfa/toxicidade , Animais , Células Cultivadas , Proteínas do Citoesqueleto/genética , Citoesqueleto/metabolismo , Citoesqueleto/patologia , Impedância Elétrica , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Masculino , Microvasos/metabolismo , Microvasos/patologia , Fosforilação , Ratos Sprague-Dawley , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/metabolismo
7.
Microvasc Res ; 133: 104098, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33075405

RESUMO

During diabetes mellitus, advanced glycation end-products (AGEs) are major contributors to the development of alterations in cerebral capillaries, leading to the disruption of the blood-brain barrier (BBB). Consequently, this is often associated with an amplified oxidative stress response in microvascular endothelial cells. As a model to mimic brain microvasculature, the bEnd.3 endothelial cell line was used to investigate cell barrier function. Cells were exposed to native bovine serum albumin (BSA) or modified BSA (BSA-AGEs). In the presence or absence of the antioxidant compound, N-acetyl-cysteine, cell permeability was assessed by FITC-dextran exclusion, intracellular free radical formation was monitored with H2DCF-DA probe, and mitochondrial respiratory and redox parameters were analyzed. We report that, in the absence of alterations in cell viability, BSA-AGEs contribute to an increase in endothelial cell barrier permeability and a marked and prolonged oxidative stress response. Decreased mitochondrial oxygen consumption was associated with these alterations and may contribute to reactive oxygen species production. These results suggest the need for further research to explore therapeutic interventions to restore mitochondrial functionality in microvascular endothelial cells to improve brain homeostasis in pathological complications associated with glycation.


Assuntos
Encéfalo/irrigação sanguínea , Permeabilidade Capilar/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Produtos Finais de Glicação Avançada/toxicidade , Microvasos/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Soroalbumina Bovina/toxicidade , Animais , Linhagem Celular , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Camundongos , Microvasos/metabolismo , Microvasos/patologia , Mitocôndrias/metabolismo , Mitocôndrias/patologia
9.
Radiat Res ; 194(6): 625-635, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-33348373

RESUMO

Persistent vasculature abnormalities contribute to an altered CNS microenvironment that further compromises the integrity of the blood-brain barrier and exposes the brain to a host of neurotoxic conditions. Standard radiation therapy at conventional (CONV) dose rate elicits short-term damage to the blood-brain barrier by disrupting supportive cells, vasculature volume and tight junction proteins. While current clinical applications of cranial radiotherapy use dose fractionation to reduce normal tissue damage, these treatments still cause significant complications. While dose escalation enhances treatment of radiation-resistant tumors, methods to subvert normal tissue damage are clearly needed. In this regard, we have recently developed a new modality of irradiation based on the use of ultra-high-dose-rate FLASH that does not induce the classical pathogenic patterns caused by CONV irradiation. In previous work, we optimized the physical parameters required to minimize normal brain toxicity (i.e., FLASH, instantaneous intra-pulse dose rate, 6.9 · 106 Gy/s, at a mean dose rate of 2,500 Gy/s), which we then used in the current study to determine the effect of FLASH on the integrity of the vasculature and the blood-brain barrier. Both early (24 h, one week) and late (one month) timepoints postirradiation were investigated using C57Bl/6J female mice exposed to whole-brain irradiation delivered in single doses of 25 Gy and 10 Gy, respectively, using CONV (0.09 Gy/s) or FLASH (>106 Gy/s). While the majority of changes found one day postirradiation were minimal, FLASH was found to reduce levels of apoptosis in the neurogenic regions of the brain at this time. At one week and one month postirradiation, CONV was found to induce vascular dilation, a well described sign of vascular alteration, while FLASH minimized these effects. These results were positively correlated with and temporally coincident to changes in the immunostaining of the vasodilator eNOS colocalized to the vasculature, suggestive of possible dysregulation in blood flow at these latter times. Overall expression of the tight junction proteins, occludin and claudin-5, which was significantly reduced after CONV irradiation, remained unchanged in the FLASH-irradiated brains at one and four weeks postirradiation. Our data further confirm that, compared to isodoses of CONV irradiation known to elicit detrimental effects, FLASH does not damage the normal vasculature. These data now provide the first evidence that FLASH preserves microvasculature integrity in the brain, which may prove beneficial to cognition while allowing for better tumor control in the clinic.


Assuntos
Indução Enzimática/efeitos da radiação , Óxido Nítrico Sintase Tipo III/biossíntese , Radioterapia/métodos , Junções Íntimas/efeitos da radiação , Vasodilatação/efeitos da radiação , Animais , Apoptose/efeitos da radiação , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Microvasos/enzimologia , Microvasos/patologia , Microvasos/efeitos da radiação
10.
Curr Neurovasc Res ; 17(5): 784-792, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33355054

RESUMO

A pericyte-centered theory suggesting that embolisms occurring within the microvasculature of a neurovascular unit that can result in either parenchymal hemorrhage or intravascular congestion is presented here. Dysfunctional microvascular pericytes are characterized by their location in the neurovascular unit, either on the arteriole or venule side. Pathophysiological and pathological changes caused by coronavirus disease 2019 (COVID-19) include pulmonary hypertension, edema, focal hemorrhage, microvascular congestion, and thrombosis. In this paper, the application of the pericytes-centered hypothesis to COVID-19 has been presented by proposing the concept of a pulmonary neurovascular unit (pNVU). The application of this concept implies that human lungs contain approximately 300 million pNVUs. This concept of existing local regulation of microvascular blood flow is supported by the observation of pathophysiology in pulmonary embolism and in acute high-altitude illness. The autonomic control seen in these three disease states matches blood flow with oxygen supply in each pNVU to maintain physiological blood oxygen saturation level. This paper illustrates how the malfunction of microvascular pericytes may cause focal hemorrhage, edema or microvascular congestion and thrombosis. A bypass existing in each pNVU would autonomically deviate blood flow from a COVID-19-affected pNVU to other healthy pNVUs. This action would prevent systemically applied medicines from reaching the therapeutic threshold in COVID-19-affected pNVUs. While testing this hypothesis with experimental evidence is urgently needed, supporting therapy aimed at improving microcirculation or rebuilding the physiological function of microvascular pericytes is recommended as a potentially effective treatment of COVID 19.


Assuntos
Barreira Hematoencefálica/metabolismo , Encéfalo/metabolismo , Acoplamento Neurovascular/fisiologia , Pericitos/metabolismo , Animais , Barreira Hematoencefálica/patologia , Encéfalo/patologia , Humanos , Microcirculação/fisiologia , Microvasos/metabolismo , Microvasos/patologia , Pericitos/patologia
11.
Nat Commun ; 11(1): 5778, 2020 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-33188196

RESUMO

Breakdown of vascular barriers is a major complication of inflammatory diseases. Anucleate platelets form blood-clots during thrombosis, but also play a crucial role in inflammation. While spatio-temporal dynamics of clot formation are well characterized, the cell-biological mechanisms of platelet recruitment to inflammatory micro-environments remain incompletely understood. Here we identify Arp2/3-dependent lamellipodia formation as a prominent morphological feature of immune-responsive platelets. Platelets use lamellipodia to scan for fibrin(ogen) deposited on the inflamed vasculature and to directionally spread, to polarize and to govern haptotactic migration along gradients of the adhesive ligand. Platelet-specific abrogation of Arp2/3 interferes with haptotactic repositioning of platelets to microlesions, thus impairing vascular sealing and provoking inflammatory microbleeding. During infection, haptotaxis promotes capture of bacteria and prevents hematogenic dissemination, rendering platelets gate-keepers of the inflamed microvasculature. Consequently, these findings identify haptotaxis as a key effector function of immune-responsive platelets.


Assuntos
Plaquetas/patologia , Vasos Sanguíneos/patologia , Quimiotaxia , Inflamação/patologia , Pneumonia/sangue , Complexo 2-3 de Proteínas Relacionadas à Actina/metabolismo , Adulto , Animais , Movimento Celular , Microambiente Celular , Modelos Animais de Doenças , Fibrinogênio/metabolismo , Humanos , Lipopolissacarídeos , Lesão Pulmonar/microbiologia , Lesão Pulmonar/patologia , Staphylococcus aureus Resistente à Meticilina/fisiologia , Camundongos Endogâmicos C57BL , Microvasos/patologia , Pneumonia/microbiologia , Pseudópodes/metabolismo
12.
Neurol Sci ; 41(12): 3401-3404, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33034804

RESUMO

We describe the case of a COVID-19 patient with severely impaired consciousness after sedation hold, showing magnetic resonance imaging (MRI) findings of (i) acute bilateral supratentorial ischemic lesions involving the fronto-parietal white matter and the corpus callosum and (ii) multiple diffuse susceptibility weighted imaging (SWI) hypointense foci, infra and supratentorial, predominantly bithalamic, suggestive of microhemorrhage or alternatively microthrombi. Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) RNA was detected in the cerebrospinal fluid. Our findings suggest the occurrence of vascular damage, predominantly involving microvessels. The underlying mechanisms, which include direct and indirect penetration of the virus to the central nervous system and systemic cardiorespiratory complications, are yet to be elucidated, and a direct correlation with SARS-CoV-2 infection remains uncertain.


Assuntos
Isquemia Encefálica/patologia , Isquemia Encefálica/virologia , Infecções por Coronavirus/complicações , Microvasos/patologia , Pneumonia Viral/complicações , Idoso , Betacoronavirus , Diabetes Mellitus , Evolução Fatal , Humanos , Hipertensão/complicações , Masculino , Pandemias
13.
Medicine (Baltimore) ; 99(36): e21883, 2020 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-32899018

RESUMO

To investigate the expression and clinical significance of aquaporin-1 (AQP1), vascular endothelial growth factor (VEGF) and microvessel density (MVD) in gastric cancer.A total of 79 gastric cancer patients who were admitted into Beijing Chao-Yang Hospital from January, 2018 to December, 2019 were involved in this study. Tumor specimens and para-cancerous normal tissues (> 2 cm away from the tumor) of all the enrolled patients were collected. Immunohistochemistry were performed to identify the expression of AQP1, VEGF, and MVD and the correlation between AQP1, VEGF, MVD, and clinicopathological parameters was analyzed.The expression of AQP1, VEGF and MVD in gastric cancer tissue was increased significantly compared with those in para-cancerous tissue (P < .05). AQP1, VEGF, and MVD were closely correlated with gastric cancer differentiation, lymph node metastasis, vascular tumor thrombosis and clinical stage (P < .05). Spearman correlation analysis showed that AQP1 was positively associated with VEGF expression (r = 0.497, P < .05). MVD was enhanced in VEGF or AQP1 positive cancer tissues compared with that in VEGF or AQP1 negative tissue (P < .05).Synergistic effect among AQP1, VEGF, and MVD is involved in occurrence and development of gastric cancer.


Assuntos
Aquaporina 1/metabolismo , Microvasos/patologia , Neoplasias Gástricas/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Estudos Retrospectivos , Neoplasias Gástricas/patologia , Neoplasias Gástricas/cirurgia
14.
Medicine (Baltimore) ; 99(38): e22126, 2020 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-32957332

RESUMO

INTRODUCTION: Coronary microvascular disease (CMVD) can affect the structure, function, and metabolism of the heart, and has an important impact on the occurrence, development and prognosis of coronary artery disease (CAD). Shexiang Tongxin dropping pill (STDP) can dilate blood vessels, alleviate inflammation, reduce endothelial damage, and improve coronary microvascular function in mice with myocardial infarction. This study aims to assess the impact of STDP on stable coronary artery disease (SCAD) patients with normal FFR and CMVD. METHODS AND ANALYSIS: This is a single-center, prospective randomized trial that will enroll 64 SCAD patients, CAD with normal FFR and CMVD. Patients will be randomly divided into study group and control group in a 1:1 fashion. On the basis of conventional drug treatment, the former will receive STDP while the latter will not. The follow-up period of the subjects is 12 months, and clinical follow-up will be conducted before discharge, 30 days, 3 months, 6 months, and 12 months after procedure to complete the detection of relevant indicators. The primary endpoint is the change of index of microcirculatory resistance (ΔIMR) at 12-month follow-up. DISCUSSION: The present study will be the first randomized control study to evaluate the efficacy and safety of STDP on SCAD patients, CAD with normal FFR and CMVD, which will provide a broader idea and more experimental basis for improving the treatment of CMVD. TRIAL REGISTRATION: This is a protocol for the randomized clinical trial which has been registered in the Chinese clinical Trial Registry with an identifier: ChiCTR2000032429.


Assuntos
Doença da Artéria Coronariana/tratamento farmacológico , Doença da Artéria Coronariana/patologia , Medicamentos de Ervas Chinesas , Microvasos/patologia , Circulação Coronária , Reserva Fracionada de Fluxo Miocárdico , Humanos , Microcirculação , Estudos Prospectivos , Ensaios Clínicos Controlados Aleatórios como Assunto
15.
PLoS One ; 15(8): e0237883, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32866169

RESUMO

Although whole-genome sequencing has provided novel insights into Neisseria meningitidis, many open reading frames have only been annotated as hypothetical proteins with unknown biological functions. Our previous genetic analyses revealed that the hypothetical protein, NMB1345, plays a crucial role in meningococcal infection in human brain microvascular endothelial cells; however, NMB1345 has no homology to any identified protein in databases and its physiological function could not be elucidated using pre-existing methods. Among the many biological technologies to examine transient protein-protein interaction in vivo, one of the developed methods is genetic code expansion with non-canonical amino acids (ncAAs) utilizing a pyrrolysyl-tRNA synthetase/tRNAPyl pair from Methanosarcina species: However, this method has never been applied to assign function-unknown proteins in pathogenic bacteria. In the present study, we developed a new method to genetically incorporate ncAAs-encoded photocrosslinking probes into N. meningitidis by utilizing a pyrrolysyl-tRNA synthetase/tRNAPyl pair and elucidated the biological function(s) of the NMB1345 protein. The results revealed that the NMB1345 protein directly interacts with PilE, a major component of meningococcal pili, and further physicochemical and genetic analyses showed that the interaction between the NMB1345 protein and PilE was important for both functional pilus formation and meningococcal infectious ability in N. meningitidis. The present study using this new methodology for N. meningitidis provides novel insights into meningococcal pathogenesis by assigning the function of a hypothetical protein.


Assuntos
Aminoácidos/genética , Reagentes para Ligações Cruzadas/metabolismo , Luz , Neisseria meningitidis/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/isolamento & purificação , Encéfalo/irrigação sanguínea , Endocitose , Células Endoteliais/microbiologia , Fímbrias Bacterianas/metabolismo , Humanos , Microvasos/patologia , Mutação/genética , Plasmídeos/genética
16.
Cancer Imaging ; 20(1): 60, 2020 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-32811570

RESUMO

BACKGROUND: To investigate the potential value of volumetric iodine quantification using preoperative dual-energy computed tomography (DECT) for predicting microvascular invasion (MVI) of hepatocellular carcinoma (HCC). METHODS: This retrospective study included patients with single HCC treated through surgical resection who underwent preoperative DECT. Quantitative DECT features, including normalized iodine concentration (NIC) to the aorta and mixed-energy CT attenuation value in the arterial phase, were three-dimensionally measured for peritumoral and intratumoral regions: (i) layer-by-layer analysis for peritumoral layers (outer layers 1 and 2; numbered in close order from the tumor boundary) and intratumoral layers (inner layers 1 and 2) with 2-mm layer thickness and (ii) volume of interest (VOI)-based analysis with different volume coverage (tumor itself; VOIO1, tumor plus outer layer 1; VOIO2, tumor plus outer layers 1 and 2; VOII1, tumor minus inner layer 1; VOII2, tumor minus inner layers 1 and 2). In addition, qualitative CT features, including peritumoral enhancement and tumor margin, were assessed. Qualitative and quantitative CT features were compared between HCC patients with and without MVI. Diagnostic performance of DECT parameters of layers and VOIs was assessed using receiver operating characteristic curve analysis. RESULTS: A total of 36 patients (24 men, mean age 59.9 ± 8.5 years) with MVI (n = 14) and without MVI (n = 22) were included. HCCs with MVI showed significantly higher NICs of outer layer 1, outer layer 2, VOIO1, and VOIO2 than those without MVI (P = 0.01, 0.04, 0.02, 0.02, respectively). Among the NICs of layers and VOIs, the highest area under the curve was obtained in outer layer 1 (0.747). Qualitative features, including peritumoral enhancement and tumor margin, and the mean CT attenuation of each layer and each VOI were not significantly different between HCCs with and without MVI (both P >  0.05). CONCLUSIONS: Volumetric iodine quantification of peritumoral and intratumoral regions in arterial phase using DECT may help predict the MVI of HCC.


Assuntos
Carcinoma Hepatocelular/diagnóstico por imagem , Tomografia Computadorizada de Feixe Cônico/métodos , Neoplasias Hepáticas/diagnóstico por imagem , Imagem Radiográfica a Partir de Emissão de Duplo Fóton/métodos , Adulto , Idoso , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/cirurgia , Tomografia Computadorizada de Feixe Cônico/normas , Feminino , Humanos , Iodo , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/cirurgia , Masculino , Microvasos/patologia , Pessoa de Meia-Idade , Invasividade Neoplásica , Período Pré-Operatório , Imagem Radiográfica a Partir de Emissão de Duplo Fóton/normas
17.
EBioMedicine ; 58: 102925, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32745993

RESUMO

BACKGROUND: Coronavirus induced disease 2019 (COVID-19) can be complicated by severe organ damage leading to dysfunction of the lungs and other organs. The processes that trigger organ damage in COVID-19 are incompletely understood. METHODS: Samples were donated from hospitalized patients. Sera, plasma, and autopsy-derived tissue sections were examined employing flow cytometry, enzyme-linked immunosorbent assays, and immunohistochemistry. PATIENT FINDINGS: Here, we show that severe COVID-19 is characterized by a highly pronounced formation of neutrophil extracellular traps (NETs) inside the micro-vessels. Intravascular aggregation of NETs leads to rapid occlusion of the affected vessels, disturbed microcirculation, and organ damage. In severe COVID-19, neutrophil granulocytes are strongly activated and adopt a so-called low-density phenotype, prone to spontaneously form NETs. In accordance, markers indicating NET turnover are consistently increased in COVID-19 and linked to disease severity. Histopathology of the lungs and other organs from COVID-19 patients showed congestions of numerous micro-vessels by aggregated NETs associated with endothelial damage. INTERPRETATION: These data suggest that organ dysfunction in severe COVID-19 is associated with excessive NET formation and vascular damage. FUNDING: Deutsche Forschungsgemeinschaft (DFG), EU, Volkswagen-Stiftung.


Assuntos
Infecções por Coronavirus/patologia , Armadilhas Extracelulares/metabolismo , Microvasos/patologia , Neutrófilos/metabolismo , Pneumonia Viral/patologia , Trombose/metabolismo , Células Cultivadas , Infecções por Coronavirus/complicações , Infecções por Coronavirus/metabolismo , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Humanos , Microvasos/metabolismo , Neutrófilos/patologia , Pandemias , Pneumonia Viral/complicações , Pneumonia Viral/metabolismo , Trombose/etiologia , Trombose/patologia
18.
Toxicol Lett ; 333: 211-221, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32841740

RESUMO

Bothrops (lance-head pit vipers) venoms are rich in weaponised metalloprotease enzymes (SVMP). These toxic enzymes are structurally diverse and functionally versatile. Potent coagulotoxicity is particularly important for prey capture (via stroke-induction) and relevant to human clinical cases (due to consumption of clotting factors including the critical depletion of fibrinogen). In this study, three distinct isoforms of P-III class SVMPs (IC, IIB and IIC), isolated from Bothrops neuwiedi venom, were evaluated for their differential capacities to affect hemostasis of prey and human plasma. Furthermore, we tested the relative antivenom neutralisation of effects upon human plasma. The toxic enzymes displayed differential procoagulant potency between plasma types, and clinically relevant antivenom efficacy variations were observed. Of particular importance was the confirmation the antivenom performed better against prothrombin activating toxins than Factor X activating toxins, which is likely due to the greater prevalence of the former in the immunising venoms used for antivenom production. This is clinically relevant as the enzymes displayed differential potency in this regard, with one (IC) in particular being extremely potent in activating Factor X and thus was correspondingly poorly neutralised. This study broadens the current understanding about the adaptive role of the SVMPs, as well as highlights how the functional diversity of SVMP isoforms can influence clinical outcomes. Key Contribution: Our findings shed light upon the hemorrhagic and coagulotoxic effects of three SVMPs of the P-III class, as well as the coagulotoxic effects of SVMPs on human, avian and amphibian plasmas. Antivenom neutralised prothrombin-activating isoforms better than Factor X activating isoforms.


Assuntos
Antivenenos/farmacologia , Coagulação Sanguínea/efeitos dos fármacos , Hemorragia/prevenção & controle , Metaloproteases/toxicidade , Venenos de Serpentes/enzimologia , Animais , Bothrops , Feminino , Hemorragia/sangue , Hemorragia/induzido quimicamente , Hemorragia/fisiopatologia , Humanos , Microscopia Intravital , Masculino , Metaloproteases/química , Camundongos , Microcirculação/efeitos dos fármacos , Microvasos/diagnóstico por imagem , Microvasos/efeitos dos fármacos , Microvasos/patologia , Isoformas de Proteínas
19.
PLoS One ; 15(7): e0234568, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32658932

RESUMO

INTRODUCTION: The aim of this study was to define histo-morphological stroma characteristics by analyzing stromal components, and to evaluate their impact on local and systemic tumor spread and overall survival in pancreatic ductal adenocarcinoma (PDAC). METHODS AND MATERIALS: Patients who underwent oncologic resections with curative intent for PDAC were identified from a prospectively maintained database. Histological specimens were re-evaluated for morphological stroma features as stromal fibers, fibroblast morphology, stroma matrix density, microvessel density and distribution of immune cell populations. RESULTS: A total of 108 patients were identified undergoing curative resection for PDAC in the period from 2011-2016. 33 (30.6%) patients showed parallel alignment of stroma fibers while 75 (69.4%) had randomly oriented stroma fibers. As compared to parallel alignment, random orientation of stroma fibers was associated with larger tumor size (median 3.62 cm vs. median 2.87cm, p = 0.037), nodal positive disease (76.0% vs. 54.5%, p = 0.040), higher margin positive resection rates (41.9% vs. 15.2%, p = 0.008) and a trend for higher rates of T3/4 tumors (33.3% vs. 15.2%, p = 0.064). In univariate analysis, patients with parallel alignment of stroma fibers had improved overall survival rates as compared to patients with random orientation of stroma fibers (42 months vs. 22 months, p = 0.046). The combination of random orientation of stroma fibers and low microvessel density was associated with impaired overall survival rates (16 months vs. 36 months, p = 0.019). A high CD4/CD3 ratio (16 months vs. 33 months, p = 0.040) and high stromal density of CD163 positive cells were associated with reduced overall survival (27 months vs. 34 months, p = 0.039). In multivariable analysis, the combination of random orientation of stroma fibers and low microvessel density (HR 1.592, 95%CI 1.098-2.733, p = 0.029), high CD4/CD3 ratio (HR 2.044, 95%CI 1.203-3.508, p = 0.028) and high density of CD163 positive cells (HR 1.596, 95%CI 1.367-1.968, p = 0.036) remained independent prognostic factors. CONCLUSION: Alignment of stroma fibers and microvessel density are simple histomorphological features serving as surrogate markers of local tumor progression dissemination and surgical resectability and determine prognosis in PDAC patients. High CD4/CD3 ratio and CD163 positive cell counts determine poor prognosis.


Assuntos
Carcinoma Ductal Pancreático/mortalidade , Carcinoma Ductal Pancreático/patologia , Microvasos/patologia , Células Estromais/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/patologia , Prognóstico , Taxa de Sobrevida
20.
Microvasc Res ; 131: 104036, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32603698

RESUMO

OBJECTIVE: The role of nailfold capillaroscopy (NC) in common non-rheumatic conditions has not been systematically reported. The aim of this review is to outline NC features observed in frequent non-rheumatic conditions, providing a practical tool to support rheumatologists for the interpretation of capillaroscopic abnormalities in patients with no established connective tissue disease (CTD). METHODS: We undertook a systematic search in PubMed and Web of Science databases. Studies reporting adults or children with common non-rheumatic diseases or conditions in which quantitative and/or qualitative assessment of morphological nailbed capillary findings was obtained, were included. The presence of a control group composed by subjects not affected by the studied condition and direct comparison of findings between groups were needed. RESULTS: We included 25 articles. Diabetes mellitus (11 studies), glaucoma (7 studies) and essential hypertension (3 studies) were the most represented diseases. Reduced capillary density, tortuosity, dilated capillaries, microhaemorrhages, ramified capillaries and avascular areas can be observed in diabetic patients. Association was reported between poor glycaemic control or longer duration of diabetes, or presence of microvascular complications as retinopathy and neuropathy, and more severe capillaroscopic abnormalities. Decreased capillary density, tortuosity, microhaemorrhages, dilated capillaries, avascular areas and ramifications might also be present in glaucoma, while in essential hypertension a reduced capillary density might be expected. CONCLUSION: Abnormal capillaroscopic findings are not uncommon even in individuals with no CTD. Therefore, presence of comorbidities known to potentially affect the microvascular array should always be investigated in patients undergoing NC and the interpretation of findings might be weighted accordingly.


Assuntos
Angiopatias Diabéticas/patologia , Hipertensão Essencial/patologia , Glaucoma/patologia , Angioscopia Microscópica , Microvasos/patologia , Doenças Reumáticas/patologia , Diagnóstico Diferencial , Feminino , Humanos , Masculino , Valor Preditivo dos Testes
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