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1.
JIMD Rep ; 62(1): 15-21, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34765393

RESUMO

Acid sphingomyelinase deficiency (ASMD) is a lysosomal storage disease (LSD) in which sphingomyelin accumulates due to deficient acid sphingomyelinase. In the chronic visceral subtype, organ manifestations are generally limited to the spleen, liver, and lungs. We report a male patient with the chronic visceral subtype who developed proteinuria and renal insufficiency at the age of 49. In renal tissue, foam cells were observed in the glomeruli as well as sphingomyelin accumulation within podocytes, mesangial cells, endothelial cells, and tubular epithelial cells. Although macrophages are the primary storage cells in both ASMD and Gaucher disease, comparison to the histopathological findings in Gaucher and Fabry disease revealed a diffuse storage pattern in multiple renal cell types, closer resembling the pattern found in Fabry disease.

2.
Lancet Digit Health ; 2021 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-34794930

RESUMO

BACKGROUND: Histopathological assessment of transplant biopsies is currently the standard method to diagnose allograft rejection and can help guide patient management, but it is one of the most challenging areas of pathology, requiring considerable expertise, time, and effort. We aimed to analyse the utility of deep learning to preclassify histology of kidney allograft biopsies into three main broad categories (ie, normal, rejection, and other diseases) as a potential biopsy triage system focusing on transplant rejection. METHODS: We performed a retrospective, multicentre, proof-of-concept study using 5844 digital whole slide images of kidney allograft biopsies from 1948 patients. Kidney allograft biopsy samples were identified by a database search in the Departments of Pathology of the Amsterdam UMC, Amsterdam, Netherlands (1130 patients) and the University Medical Center Utrecht, Utrecht, Netherlands (717 patients). 101 consecutive kidney transplant biopsies were identified in the archive of the Institute of Pathology, RWTH Aachen University Hospital, Aachen, Germany. Convolutional neural networks (CNNs) were trained to classify allograft biopsies as normal, rejection, or other diseases. Three times cross-validation (1847 patients) and deployment on an external real-world cohort (101 patients) were used for validation. Area under the receiver operating characteristic curve (AUROC) was used as the main performance metric (the primary endpoint to assess CNN performance). FINDINGS: Serial CNNs, first classifying kidney allograft biopsies as normal (AUROC 0·87 [ten times bootstrapped CI 0·85-0·88]) and disease (0·87 [0·86-0·88]), followed by a second CNN classifying biopsies classified as disease into rejection (0·75 [0·73-0·76]) and other diseases (0·75 [0·72-0·77]), showed similar AUROC in cross-validation and deployment on independent real-world data (first CNN normal AUROC 0·83 [0·80-0·85], disease 0·83 [0·73-0·91]; second CNN rejection 0·61 [0·51-0·70], other diseases 0·61 [0·50-0·74]). A single CNN classifying biopsies as normal, rejection, or other diseases showed similar performance in cross-validation (normal AUROC 0·80 [0·73-0·84], rejection 0·76 [0·66-0·80], other diseases 0·50 [0·36-0·57]) and generalised well for normal and rejection classes in the real-world data. Visualisation techniques highlighted rejection-relevant areas of biopsies in the tubulointerstitium. INTERPRETATION: This study showed that deep learning-based classification of transplant biopsies could support pathological diagnostics of kidney allograft rejection. FUNDING: European Research Council; German Research Foundation; German Federal Ministries of Education and Research, Health, and Economic Affairs and Energy; Dutch Kidney Foundation; Human(e) AI Research Priority Area of the University of Amsterdam; and Max-Eder Programme of German Cancer Aid.

3.
Front Immunol ; 12: 723967, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34552589

RESUMO

Bruton's tyrosine kinase (Btk) is a cytoplasmic kinase expressed in B cells and myeloid cells. It is essential for B cell development and natural antibody-mediated host defense against bacteria in humans and mice, but little is known about the role of Btk in innate host defense in vivo. Previous studies have indicated that lack of (natural) antibodies is paramount for impaired host defense against Streptococcus (S.) pneumoniae in patients and mice with a deficiency in functional Btk. In the present study, we re-examined the role of Btk in B cells and myeloid cells during pneumococcal pneumonia and sepsis in mice. The antibacterial defense of Btk-/- mice was severely impaired during pneumococcal pneumosepsis and restoration of natural antibody production in Btk-/- mice by transgenic expression of Btk specifically in B cells did not suffice to protect against infection. Btk-/- mice with reinforced Btk expression in MhcII+ cells, including B cells, dendritic cells and macrophages, showed improved antibacterial defense as compared to Btk-/- mice. Bacterial outgrowth in Lysmcre-Btkfl/Y mice was unaltered despite a reduced capacity of Btk-deficient alveolar macrophages to respond to pneumococci. Mrp8cre-Btkfl/Y mice with a neutrophil specific paucity in Btk expression, however, demonstrated impaired antibacterial defense. Neutrophils of Mrp8cre-Btkfl/Y mice displayed reduced release of granule content after pulmonary installation of lipoteichoic acid, a gram-positive bacterial cell wall component relevant for pneumococci. Moreover, Btk deficient neutrophils showed impaired degranulation and phagocytosis upon incubation with pneumococci ex vivo. Taken together, the results of our study indicate that besides regulating B cell-mediated immunity, Btk is critical for regulation of myeloid cell-mediated, and particularly neutrophil-mediated, innate host defense against S. pneumoniae in vivo.

4.
Vox Sang ; 2021 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-34396543

RESUMO

BACKGROUND AND OBJECTIVES: Transfusion-associated circulatory overload (TACO) is the primary cause of transfusion-related mortality. Speed and volume of transfusion are major risk factors. The aim of this study was to investigate the interaction of red blood cell (RBC) transfusion speed and volume on the development of TACO. MATERIALS AND METHODS: A validated model for TACO in anaemic Lewis rats with an acute myocardial infarction was used. The effect on pulmonary hydrostatic pressure of one, two or four units of packed RBCs transfused in either 30 or 60 min was evaluated (3.3-26.6 ml·kg-1 ·hr-1 ). Pulmonary capillary pressure was measured as left ventricular end-diastolic pressure (LVEDP). Cardiac stress biomarkers atrial natriuretic-peptide (ANP) and N-terminal pro-brain natriuretic peptide (NT-proBNP) were measured 1-h post-transfusion. RESULTS: Thirty animals were included (n = 5 per group). Transfusion of RBCs increased LVEDP in a volume-dependent manner (ΔLVEDP [mmHg]: -0.95, +0.50, +6.26, p < 0.001). Fast transfusion increased overall ΔLVEDP by +3.5 mmHg and up to +11.8 mmHg in the four units' group (p = 0.016). Doubling transfusion speed increased ΔLVEDP more than doubling volume in the larger volume groups. No difference in ANP or NT-proBNP were seen in high transfusion volume or groups. CONCLUSION: Transfusion volume dose-dependently increased LVEDP, with speed of transfusion rapidly elevating LVEDP at higher transfusion volumes. ANP and NT-proBNP were not impacted by transfusion volume or speed in this model. TACO is seen as purely volume overload, however, this study emphasizes that limiting transfusion speed, as a modifiable risk factor, might aid in preventing TACO.

5.
Mediators Inflamm ; 2021: 9958281, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34393650

RESUMO

Hypoxia-inducible factor- (HIF-) 1α has been implicated in the ability of cells to adapt to alterations in oxygen levels. Bacterial stimuli can induce HIF1α in immune cells, including those of myeloid origin. We here determined the role of myeloid cell HIF1α in the host response during pneumonia and sepsis caused by the common human pathogen Klebsiella pneumoniae. To this end, we generated mice deficient for HIF1α in myeloid cells (LysM-cre × Hif1α fl/fl) or neutrophils (Mrp8-cre × Hif1α fl/fl) and infected these with Klebsiella pneumoniae via the airways. Myeloid, but not neutrophil, HIF1α-deficient mice had increased bacterial loads in the lungs and distant organs after infection as compared to control mice, pointing at a role for HIF1α in macrophages. Myeloid HIF1α-deficient mice did not show increased bacterial growth after intravenous infection, suggesting that their phenotype during pneumonia was mediated by lung macrophages. Alveolar and lung interstitial macrophages from LysM-cre × Hif1α fl/fl mice produced lower amounts of the immune enhancing cytokine tumor necrosis factor upon stimulation with Klebsiella, while their capacity to phagocytose or to produce reactive oxygen species was unaltered. Alveolar macrophages did not upregulate glycolysis in response to lipopolysaccharide, irrespective of HIF1α presence. These data suggest a role for HIF1α expressed in lung macrophages in protective innate immunity during pneumonia caused by a common bacterial pathogen.

6.
Biomolecules ; 11(8)2021 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-34439745

RESUMO

CCAAT/enhancer-binding protein delta (C/EBPδ) is a transcription factor involved in apoptosis and proliferation, which is downregulated in pancreatic ductal adenocarcinoma (PDAC) cells. Loss of nuclear C/EBPδ in PDAC cells is associated with decreased patient survival and pro-tumorigenic properties in vitro. Interestingly however, next to C/EBPδ expression in tumor cells, C/EBPδ is also expressed by cells constituting the tumor microenvironment and by cells comprising the organs and parenchyma. However, the functional relevance of systemic C/EBPδ in carcinogenesis remains elusive. Here, we consequently assessed the potential importance of C/EBPδ in somatic tissues by utilizing an orthotopic pancreatic cancer model. In doing so, we show that genetic ablation of C/EBPδ does not significantly affect primary tumor growth but has a strong impact on metastases; wildtype mice developed metastases at multiple sites, whilst this was not the case in C/EBPδ-/- mice. In line with reduced metastasis formation in C/EBPδ-/- mice, C/EBPδ-deficiency also limited tumor cell dissemination in a specific extravasation model. Tumor cell extravasation was dependent on the platelet-activating factor receptor (PAFR) as a PAFR antagonist inhibited tumor cell extravasation in wildtype mice but not in C/EBPδ-/- mice. Overall, we show that systemic C/EBPδ facilitates pancreatic cancer metastasis, and we suggest this is due to C/EBPδ-PAFR-dependent tumor cell extravasation.


Assuntos
Proteína delta de Ligação ao Facilitador CCAAT/biossíntese , Metástase Neoplásica , Neoplasias Pancreáticas/metabolismo , Animais , Apoptose , Proteína delta de Ligação ao Facilitador CCAAT/genética , Linhagem Celular Tumoral , Proliferação de Células , Proteínas de Ligação a DNA/metabolismo , Mineração de Dados , Progressão da Doença , Regulação da Expressão Gênica , Humanos , Camundongos , Transplante de Neoplasias , Neoplasias Pancreáticas/patologia , Glicoproteínas da Membrana de Plaquetas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Análise Serial de Tecidos , Transativadores/metabolismo , Microambiente Tumoral
7.
Intensive Care Med Exp ; 9(1): 23, 2021 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-33997943

RESUMO

BACKGROUND: Hemorrhagic shock is associated with acute kidney injury and increased mortality. Targeting the endothelial angiopoietin/Tie2 system, which regulates endothelial permeability, previously reduced hemorrhagic shock-induced vascular leakage. We hypothesized that as a consequence of vascular leakage, renal perfusion and function is impaired and that activating Tie2 restores renal perfusion and function. METHODS: Rats underwent 1 h of hemorrhagic shock and were treated with either vasculotide or PBS as control, followed by fluid resuscitation for 4 h. Microcirculatory perfusion was measured in the renal cortex and cremaster muscle using contrast echography and intravital microscopy, respectively. Changes in the angiopoietin/Tie2 system and renal injury markers were measured in plasma and on protein and mRNA level in renal tissue. Renal edema formation was determined by wet/dry weight ratios and renal structure by histological analysis. RESULTS: Hemorrhagic shock significantly decreased renal perfusion (240 ± 138 to 51 ± 40, p < 0.0001) and cremaster perfusion (12 ± 2 to 5 ± 2 perfused vessels, p < 0.0001) compared to baseline values. Fluid resuscitation partially restored both perfusion parameters, but both remained below baseline values (renal perfusion 120 ± 58, p = 0.08, cremaster perfusion 7 ± 2 perfused vessels, p < 0.0001 compared to baseline). Hemorrhagic shock increased circulating angiopoietin-1 (p < 0.0001), angiopoietin-2 (p < 0.0001) and soluble Tie2 (p = 0.05), of which angiopoietin-2 elevation was associated with renal edema formation (r = 0.81, p < 0.0001). Hemorrhagic shock induced renal injury, as assessed by increased levels of plasma neutrophil gelatinase-associated lipocalin (NGAL: p < 0.05), kidney injury marker-1 (KIM-1; p < 0.01) and creatinine (p < 0.05). Vasculotide did not improve renal perfusion (p > 0.9 at all time points) or reduce renal injury (NGAL p = 0.26, KIM-1 p = 0.78, creatinine p > 0.9, renal edema p = 0.08), but temporarily improved cremaster perfusion at 3 h following start of fluid resuscitation compared to untreated rats (resuscitation + 3 h: 11 ± 3 vs 8 ± 3 perfused vessels, p < 0.05). CONCLUSION: Hemorrhagic shock-induced renal impairment cannot be restored by standard fluid resuscitation, nor by activation of Tie2. Future treatment strategies should focus on reducing angiopoietin-2 levels or on activating Tie2 via an alternative strategy.

8.
Clin Kidney J ; 14(3): 855-862, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33777368

RESUMO

Background: In systemic amyloidosis, the kidney is frequently affected and renal involvement has a major impact on survival. Renal involvement is clinically characterized by decreased estimated glomerular filtration rate (eGFR) and proteinuria. The two most common renal amyloidosis types are light chain-related amyloidosis (AL) and serum amyloid A (AA) amyloidosis. Standardized histopathological scoring of amyloid deposits is crucial to assess disease progression. Therefore, we aimed to validate the proposed scoring system from Rubinstein et al. (Novel pathologic scoring tools predict end-stage kidney disease in light chain (AL) amyloidosis. Amyloid 2017; 24: 205-211) in an independent patient cohort. Methods: We attempt to reproduce the scoring system, consisting of an amyloid score (AS) and a composite scarring injury score (CSIS), in a multicentre AL and AA case series. Additionally, we analysed all renal amyloidosis kidney biopsies performed in the Netherlands between 1993 and 2012. Results: Similar to the original study, AS and CSIS correlated to eGFR (r = -0.45, P = 0.0061 and r = -0.60, P < 0.0001, respectively) but not to proteinuria at diagnosis. Furthermore, AS, but not CSIS, was associated with renal outcome. The scoring system was not reproducible in AA patients. The median incidence rate for renal amyloidosis in the Netherlands was 2.3 per million population per year, and increased during the study period. Conclusions: In our AL case series and the original study, AS and CSIS were correlated to eGFR but not to proteinuria, and AS correlated with renal outcome. Overall, we regard this scoring system as competent for standardized histopathological assessment of amyloid deposits burden and thereby disease advancement in renal biopsies.

9.
Clin Kidney J ; 14(1): 358-365, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33564439

RESUMO

Background: Diabetic nephropathy (DN) is a major complication of diabetes and the main cause of end-stage renal disease. Extracellular vesicles (EVs) are small cell-derived vesicles that can alter disease progression by microRNA (miRNA) transfer. Methods: In this study, we aimed to characterize the cellular origin and miRNA content of EVs in plasma samples of type 2 diabetes patients at various stages of DN. Type 2 diabetes patients were classified in three groups: normoalbuminuria, microalbuminuria and macroalbuminuria. The concentration and cellular origin of plasma EVs were measured by flow cytometry. A total of 752 EV miRNAs were profiled in 18 subjects and differentially expressed miRNAs were validated. Results: Diabetic patients with microalbuminuria and/or macroalbuminuria showed elevated concentrations of total EVs and EVs from endothelial cells, platelets, leucocytes and erythrocytes compared with diabetic controls. miR-99a-5p was upregulated in macroalbuminuric patients compared with normoalbuminuric and microalbuminuric patients. Transfection of miR-99a-5p in cultured human podocytes downregulated mammalian target of rapamycin (mTOR) protein expression and downregulated the podocyte injury marker vimentin. Conclusions: Type 2 diabetes patients with microalbuminuria and macroalbuminuria display differential EV profiles. miR-99a-5p expression is elevated in EVs from macroalbuminuria and mTOR is its validated mRNA target.

10.
Am J Respir Crit Care Med ; 203(2): 175-184, 2021 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-32721210

RESUMO

Rationale: Bronchial thermoplasty (BT) is a bronchoscopic treatment for severe asthma targeting airway smooth muscle (ASM). Observational studies have shown ASM mass reduction after BT, but appropriate control groups are lacking. Furthermore, as treatment response is variable, identifying optimal candidates for BT treatment is important.Objectives: First, to assess the effect of BT on ASM mass, and second, to identify patient characteristics that correlate with BT response.Methods: Patients with severe asthma (n = 40) were randomized to immediate (n = 20) or delayed (n = 20) BT treatment. Before randomization, clinical, functional, blood, and airway biopsy data were collected. In the delayed control group, reassessment, including biopsies, was performed after 6 months of standard clinical care, followed by BT. In both groups, post-BT data including biopsies were obtained after 6 months. ASM mass (% positive desmin or α-smooth muscle actin area in the total biopsy) was calculated with automated digital analysis software. Associations between baseline characteristics and Asthma Control Questionnaire and Asthma Quality of Life Questionnaire (AQLQ) improvement were explored.Measurements and Main Results: Median ASM mass decreased by >50% in the immediate BT group (n = 17) versus no change in the delayed control group (n = 19) (P = 0.0004). In the immediate group, Asthma Control Questionnaire scores improved with -0.79 (interquartile range [IQR], -1.61 to 0.02) compared with 0.09 (IQR, -0.25 to 1.17) in the delayed group (P = 0.006). AQLQ scores improved with 0.83 (IQR, -0.15 to 1.69) versus -0.02 (IQR, -0.77 to 0.75) (P = 0.04). Treatment response in the total group (n = 35) was positively associated with serum IgE and eosinophils but not with baseline ASM mass.Conclusions: ASM mass significantly decreases after BT when compared with a randomized non-BT-treated control group. Treatment response was associated with serum IgE and eosinophil levels but not with ASM mass.


Assuntos
Asma/cirurgia , Brônquios/cirurgia , Termoplastia Brônquica , Músculo Liso/cirurgia , Adolescente , Adulto , Idoso , Remodelação das Vias Aéreas , Asma/diagnóstico , Asma/patologia , Asma/fisiopatologia , Biópsia , Brônquios/patologia , Broncoscopia , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Músculo Liso/patologia , Índice de Gravidade de Doença , Resultado do Tratamento , Adulto Jovem
11.
Intensive Care Med Exp ; 8(Suppl 1): 42, 2020 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-33336308

RESUMO

INTRODUCTION: In hemorrhaging trauma patients, the endothelium is activated, resulting in excessive endothelial synthesis of von Willebrand Factor (vWF), which may enhance micro-thrombi formation, resulting in obstruction of the microcirculation and endothelial injury, aggravating bleeding, as well as contributing to organ failure. Under normal conditions, vWF is cleaved by the metalloprotease ADAMTS-13. After trauma, ADAMTS-13 levels are reduced. OBJECTIVES: To assess whether recombinant human ADAMTS-13 inhibits endothelial injury and organ failure in a rat trauma-transfusion model. METHODS: Blood products were prepared from syngeneic rat blood according to blood bank standards. Polytrauma was induced in rats by crush injury to the intestines and liver and by fracture of the femur. The rats were hemorrhaged until a mean arterial pressure (MAP) of 40 mmHg was reached. Rats were randomized to receive transfusion of RBCs, FFPs, and platelets in a 1:1:1 ratio to achieve a MAP of 70 mmHg, with or without the addition of ADAMTS-13 (50 µg/kg). Blood samples were assessed for biochemistry and rotational thromboelastometry (ROTEM). Syndecan-1 and VE-cadherin levels were measured as a reflection of endothelial integrity. The amount of leakage of dextran-FITC from the vascular system to the parenchyma in lungs was quantified. To assess inflammation, IL-6 and IL-8 levels were determined. Organ damage was assessed by histopathology. RESULTS: All rats were severely shocked, with no significant differences in shock parameters between groups. Rats treated with ADAMTS-13 showed signs of a more effective shock reversal (higher blood pressure, lower lactate levels) compared to controls. Also, ROTEM parameters of clot formation in rats receiving ADAMTS-13 improved compared to controls, which was mainly platelet-dependent. Syndecan-1 levels relative to baseline trended to be lower in ADAMTS-13 treated rats compared to controls (107 vs 149%, p = 0.08). ADAMTS-13 reduced albuminuria (1.7 vs 4.4 g/L, p < 0.01) and organ-specific inflammation (pulmonary IL-6 243 vs 369 pg/mL, p = 0.08; splenic IL-6 253 vs 307, p = 0.03) compared to controls, but did not improve histopathological scores. CONCLUSIONS: The use of ADAMTS-13 in a rat trauma-transfusion model improves parameters of shock, platelet-driven coagulation, endothelial damage, and organ inflammation. These results suggest that ADAMTS-13 is important in mediating outcome of trauma. Whether ADAMTS-13 can be used as a therapeutic adjunct to treat bleeding trauma patients remains to be determined.

12.
Lancet Microbe ; 1(7): e290-e299, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33015653

RESUMO

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) targets multiple organs and causes severe coagulopathy. Histopathological organ changes might not only be attributable to a direct virus-induced effect, but also the immune response. The aims of this study were to assess the duration of viral presence, identify the extent of inflammatory response, and investigate the underlying cause of coagulopathy. Methods: This prospective autopsy cohort study was done at Amsterdam University Medical Centers (UMC), the Netherlands. With informed consent from relatives, full body autopsy was done on 21 patients with COVID-19 for whom autopsy was requested between March 9 and May 18, 2020. In addition to histopathological evaluation of organ damage, the presence of SARS-CoV-2 nucleocapsid protein and the composition of the immune infiltrate and thrombi were assessed, and all were linked to disease course. Findings: Our cohort (n=21) included 16 (76%) men, and median age was 68 years (range 41-78). Median disease course (time from onset of symptoms to death) was 22 days (range 5-44 days). In 11 patients tested for SARS-CoV-2 tropism, SARS-CoV-2 infected cells were present in multiple organs, most abundantly in the lungs, but presence in the lungs became sporadic with increased disease course. Other SARS-CoV-2-positive organs included the upper respiratory tract, heart, kidneys, and gastrointestinal tract. In histological analyses of organs (sampled from nine to 21 patients per organ), an extensive inflammatory response was present in the lungs, heart, liver, kidneys, and brain. In the brain, extensive inflammation was seen in the olfactory bulbs and medulla oblongata. Thrombi and neutrophilic plugs were present in the lungs, heart, kidneys, liver, spleen, and brain and were most frequently observed late in the disease course (15 patients with thrombi, median disease course 22 days [5-44]; ten patients with neutrophilic plugs, 21 days [5-44]). Neutrophilic plugs were observed in two forms: solely composed of neutrophils with neutrophil extracellular traps (NETs), or as aggregates of NETs and platelets.. Interpretation: In patients with lethal COVID-19, an extensive systemic inflammatory response was present, with a continued presence of neutrophils and NETs. However, SARS-CoV-2-infected cells were only sporadically present at late stages of COVID-19. This suggests a maladaptive immune response and substantiates the evidence for immunomodulation as a target in the treatment of severe COVID-19. Funding: Amsterdam UMC Corona Research Fund.

13.
Cancers (Basel) ; 12(9)2020 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-32906832

RESUMO

CCAAT/enhancer-binding protein δ (C/EBPδ) is a transcription factor involved in growth arrest and differentiation, which has consequently been suggested to harbor tumor suppressive activities. However, C/EBPδ over-expression correlates with poor prognosis in glioblastoma and promotes genomic instability in cervical cancer, hinting at an oncogenic role of C/EBPδ in these contexts. Here, we explore the role of C/EBPδ in pancreatic cancer. We determined C/EBPδ expression in biopsies from pancreatic cancer patients using public gene-expression datasets and in-house tissue microarrays. We found that C/EBPδ is highly expressed in healthy pancreatic ductal cells but lost in pancreatic ductal adenocarcinoma. Furthermore, loss of C/EBPδ correlated with increased lymph node involvement and shorter overall survival in pancreatic ductal adenocarcinoma patients. In accordance with this, in vitro experiments showed reduced clonogenic capacity and proliferation of pancreatic ductal adenocarcinoma cells following C/EBPδ re-expression, concurrent with decreased sphere formation capacity in soft agar assays. We thus report a previously unrecognized but important tumor suppressor role of C/EBPδ in pancreatic ductal adenocarcinoma. This is of particular interest since only few tumor suppressors have been identified in the context of pancreatic cancer. Moreover, our findings suggest that restoration of C/EBPδ activity could hold therapeutic value in pancreatic ductal adenocarcinoma, although the latter claim needs to be substantiated in future studies.

14.
FASEB J ; 34(10): 13750-13761, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32856376

RESUMO

Diabetic nephropathy (DN) is a major complication of diabetes and is associated with high risk for cardiovascular mortality, which is partially related to elevated platelet activity. Platelets are also active players in inflammation and fibrosis. In this study, we examine the effect of ticagrelor-induced platelet inhibition on the development of DN. DN was induced by unilateral nephrectomy followed by streptozotocin injections for 5 days. Mice received ticagrelor (300 mg/kg) or vehicle every other day, for 16 weeks. Experimental groups: non-diabetic control, diabetic control, non-diabetic ticagrelor, and diabetic ticagrelor. Ticagrelor treatment in diabetic mice lowered urinary albumin excretion, it prevented diabetes-induced mesangial matrix expansion, podocyte effacement, and glomerular endothelial cell injury, which includes loss of endothelial fenestrations, ICAM-1 expression, and PECAM expression. In addition, ticagrelor treatment prevented collagen IV deposition and macrophage infiltration in the tubulointerstitium and these diabetic mice showed lower systemic and tubular inflammation and tubular apoptosis. This tubular protection is likely to be a result of protection to the glomerular endothelium by ticagrelor, which reduces albuminuria and albumin toxicity to the tubules and reduced tubular and interstitial inflammation and fibrosis. In conclusion, ticagrelor-induced platelet inhibition protects against renal injury in diabetic mice, likely by protecting the glomerular endothelial cells.


Assuntos
Nefropatias Diabéticas/tratamento farmacológico , Inibidores da Agregação Plaquetária/uso terapêutico , Ticagrelor/uso terapêutico , Animais , Apoptose , Colágeno/metabolismo , Nefropatias Diabéticas/etiologia , Células Endoteliais/efeitos dos fármacos , Molécula 1 de Adesão Intercelular/metabolismo , Túbulos Renais/efeitos dos fármacos , Túbulos Renais/metabolismo , Túbulos Renais/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Inibidores da Agregação Plaquetária/administração & dosagem , Inibidores da Agregação Plaquetária/farmacologia , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Podócitos/efeitos dos fármacos , Ticagrelor/administração & dosagem , Ticagrelor/farmacologia
15.
Intensive Care Med Exp ; 8(1): 47, 2020 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-32840685

RESUMO

BACKGROUND: The microbiome has emerged as an important player in the pathophysiology of a whole spectrum of diseases that affect the critically ill. We hypothesized that differences in microbiota composition across vendors can influence murine models of pulmonary lipopolysaccharide (LPS) inflammation and Gram-negative pneumonia. METHODS: A multi-vendor approach was used with genetically similar mice derived from three different vendors (Janvier, Envigo, Charles River). This model was employed to study the effect on the host response to a pulmonary LPS challenge (1 µg Klebsiella pneumoniae LPS, intranasal), as well as experimental K. pneumoniae infection (ATCC43816, 1 × 104 CFU, intranasal). RESULTS: Gut microbiota analysis revealed profound intervendor differences in bacterial composition as shown by beta diversity and at various taxonomic levels. Tumor necrosis factor (TNF)-α and interleukin (IL)-6 release in lung and bronchoalveolar lavage fluid (BALF) were determined 6 and 24 h after intranasal treatment with LPS. No differences were found between the groups, with the exception for Envigo, showing a higher level of TNFα in lung and BALF at 6 h compared to Janvier and Charles River. In another set of experiments, mice from different vendors were subjected to a clinically relevant model of Gram-negative pneumonia (K. pneumoniae). At 12 and 36 h post-infection, no intervendor differences were found in bacterial dissemination, or TNFα and IL-6 levels in the lungs. In line, markers for organ failure did not differ between groups. CONCLUSIONS: Although there was a marked variation in the gut microbiota composition of mice from different vendors, the hypothesized impact on our models of pulmonary inflammation and severe pneumonia was limited. This is of significance for experimental settings, showing that differences in gut microbiota do not have to lead to differences in outcome.

16.
J Infect Dis ; 2020 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-32648919

RESUMO

BACKGROUND: Liver kinase B1 (LKB1) has been studied extensively as a tumor suppressor gene (Stk11) in the context of cancer. We hypothesized that myeloid LKB1 plays a role in innate immunity during pneumonia. METHODS: Mice deficient for LKB1 in myeloid cells (LysM-cre x Stk11fl/fl ) or neutrophils (Mrp8-cre x Stk11fl/fl) were infected with Klebsiellapneumoniae via the airways. LysM-cre x Stk11fl/fl mice were also intranasally challenged with lipopolysaccharide (LPS). RESULTS: Myeloid, but not neutrophil LKB1 deficient mice had increased bacterial loads in lungs from 6 to 40 hours after infection as compared to control mice, pointing at a role for LKB1 in macrophages. Myeloid LKB1 deficiency was associated with reduced cytokine release into the airways upon local LPS instillation. The number of classical (SiglecFhighCD11bneg) alveolar macrophages (AMs) was reduced by approximately 50% in the lungs of myeloid LKB1 deficient mice, which was not caused by increased cell death or reduced proliferation. Instead, myeloid LKB1 deficient mice had AMs with a 'non-classical' (SiglecFlowCD11bpos) phenotype. AMs did not upregulate glycolysis in response to LPS, irrespective of LKB1 presence. CONCLUSION: Myeloid LKB1 is important for local host defense during Klebsiella pneumonia by maintaining adequate AM numbers in the lung.

17.
Clin Immunol ; 218: 108522, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32682923

RESUMO

Human hematopoiesis is critically dependent on the transcription factor GATA2. Patients with GATA2 deficiency typically present with myelodysplastic syndrome, reduced numbers of monocytes, NK cells and B cells, and/or opportunistic infections. Here, we present two families that harbor distinct GATA2 mutations with highly variable onset and course of disease. We discuss the use of allogeneic hematopoietic cell transplantation in these patients, especially as treatment for pulmonary alveolar proteinosis.


Assuntos
Deficiência de GATA2/terapia , Transplante de Células-Tronco Hematopoéticas , Proteinose Alveolar Pulmonar/terapia , Adolescente , Adulto , Aloenxertos , Feminino , Fator de Transcrição GATA2/genética , Humanos , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Mutação , Adulto Jovem
18.
Transpl Int ; 33(9): 1116-1127, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32480425

RESUMO

The impact of primary cytomegalovirus infection (pCMV) on renal allograft function and histology is controversial. We evaluated the influence on incidence of acute rejection, allograft loss, allograft function and interstitial fibrosis/tubular atrophy (IF/TA). Retrospective case-control study, recipients transplanted between 2000 and 2014. Risk of acute rejection and allograft loss for those who experienced pCMV infection compared with those who did not, within an exposure period of two months after transplantation. Besides, its influence on allograft function and histology at one to three years after transplantation. Of 113 recipients experienced pCMV infection, 306 remained CMV seronegative. pCMV infection in the exposure period could not be proven as increasing the risk for acute rejection [HR = 2.18 (95% CI 0.80-5.97) P = 0.13] or allograft loss [HR = 1.11 (95%CI 0.33-3.72) P = 0.87]. Combination of pCMV infection and acute rejection posed higher hazard for allograft loss than acute rejection alone [HR = 3.69 (95% CI 1.21-11.29) P = 0.02]. eGFR(MDRD) values did not significantly differ at years one [46 vs. 50], two [46 vs. 51] and three [46 vs. 52]. No association between pCMV infection and IF/TA could be demonstrated [OR = 2.15 (95%CI 0.73-6.29) P = 0.16]. pCMV infection was not proven to increase the risk for acute rejection or allograft loss. However, it increased the risk for rejection-associated allograft loss. In remaining functioning allografts, it was not significantly associated with decline in function nor with presence of IF/TA.


Assuntos
Infecções por Citomegalovirus , Transplante de Rim , Estudos de Casos e Controles , Infecções por Citomegalovirus/epidemiologia , Rejeição de Enxerto/etiologia , Humanos , Transplante de Rim/efeitos adversos , Estudos Retrospectivos , Fatores de Risco
19.
Shock ; 54(6): 794-801, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32195920

RESUMO

BACKGROUND: Septic patients are often anemic, requiring red blood cell (RBC) transfusions. However, RBC transfusions are associated with organ injury. The mechanisms of RBC-induced organ injury are unknown, but increased clearance of donor RBCs from the circulation with trapping in the organs could play a role. We hypothesized that washing of RBCs prior to transfusion may reduce clearance and trapping of donor cells and thereby reduce organ injury. METHODS: Sprague-Dawley rats were inoculated intratracheally with 10 colony-forming units (CFU) of Streptococcus pneumoniae or vehicle as a control and transfused with either a washed or standard (non-washed) biotinylated RBC transfusion from syngeneic rats. Controls received saline. Blood samples were taken directly after transfusion and at 24 h to calculate the 24 h post transfusion recovery (PTR). After sacrifice, flow cytometry was used to detect donor RBCs in organs and blood. The organs were histologically scored by a pathologist and CFUs in the lung and blood were counted. RESULTS: The 24h-PTR was similar between healthy and pneumoseptic rats after a standard transfusion. In healthy rats, a washed transfusion resulted in a higher PTR and less accumulation of donor RBCs in the organs compared with a standard transfusion. However, during pneumonia, this effect of washing was not seen. Transfusion did not further augment lung injury induced by pneumonia, but washing decreased bacterial outgrowth in the lungs associated with reduced lung injury. CONCLUSION: In healthy recipients, washing increased 24h-PTR of donor RBCs and decreased trapping in organs. In pneumoseptic rats, washing reduced bacterial outgrowth and lung injury, but did not improve PTR.


Assuntos
Preservação de Sangue , Transfusão de Eritrócitos , Eritrócitos/metabolismo , Pneumonia Pneumocócica , Sepse , Streptococcus pneumoniae/metabolismo , Animais , Masculino , Pneumonia Pneumocócica/metabolismo , Pneumonia Pneumocócica/microbiologia , Pneumonia Pneumocócica/terapia , Ratos , Ratos Sprague-Dawley , Sepse/metabolismo , Sepse/microbiologia , Sepse/terapia
20.
Platelets ; 31(3): 383-391, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31364433

RESUMO

The pathophysiology of renal ischemia/reperfusion (I/R) injury is characterized by excessive activation of inflammation and coagulation processes followed by abnormal renal tissue repair, resulting in renal injury and function loss. Platelets are important actors in these processes, however to what extent platelets contribute to the pathophysiology of renal I/R injury still needs to be elucidated. In the current study, we treated wild-type mice with a platelet depleting antibody, which caused thrombocytopenia. We then investigated the role of platelets during the pathophysiology of renal I/R by subjecting control wild-type mice with normal platelet counts and thrombocytopenic wild-type mice to renal I/R injury. Our results showed that in the early phase of renal I/R injury, thrombocytopenia 24 hours after ischemia reperfusion does not influence renal injury, neutrophil infiltration and accumulation of inflammatory chemokines (e.g. keratinocyte chemoattractant, monocyte chemoattractant protein 1, tumor necrosis factor alpha). In the recovery and regeneration phase of I/R injury, respectively 5 and 10 days post-ischemia, thrombocytopenia did also not affect the accumulation of intra-renal neutrophils and macrophages, renal injury, and renal fibrosis. Together, these results imply that lowering platelet counts do not impact the pathogenesis of I/R injury in mice.


Assuntos
Injúria Renal Aguda/etiologia , Injúria Renal Aguda/patologia , Traumatismo por Reperfusão/complicações , Trombocitopenia/complicações , Animais , Biomarcadores , Modelos Animais de Doenças , Imuno-Histoquímica , Imunofenotipagem , Masculino , Camundongos , Traumatismo por Reperfusão/etiologia , Trombocitopenia/etiologia
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