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1.
ChemMedChem ; 2021 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-34582626

RESUMO

Bruton's tyrosine kinase (BTK) is a member of the Tec kinase family that is expressed in cells of hematopoietic lineage. Evidence has shown that inhibition of BTK has clinical benefit for the treatment of a wide array of autoimmune and inflammatory diseases. Previously we reported the discovery of a novel nicotinamide selectivity pocket (SP) series of potent and selective covalent irreversible BTK inhibitors. The top molecule 1 of that series strongly inhibited CYP2C8 (IC50 =100 nM), which was attributed to the bridged linker group. However, our effort on the linker replacement turned out to be fruitless. With the study of the X-ray crystal structure of compound 1, we envisioned the opportunity of removal of this liability via transposition of the linker moiety in 1 from C6 to C5 position of the pyridine core. With this strategy, our optimization led to the discovery of a novel series, in which the top molecule 18 A displayed reduced CYP inhibitory activity and good potency. To further explore this new series, different warheads besides acrylamide, for example cyanamide, were also tested. However, this effort didn't lead to the discovery of molecules with better potency than 18 A. The loss of potency in those molecules could be related to the reduced reactivity of the warhead or reversible binding mode. Further profiling of 18 A disclosed that it had a strong hERG (human Ether-a-go-go Related Gene) inhibition, which could be related to the phenoxyphenyl group.

2.
Vaccines (Basel) ; 9(8)2021 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-34452033

RESUMO

The aim of this prospective study was to assess lymphocyte proliferative and cytokine response prior to and following tick-borne encephalitis (TBE) immunization among patients after allogeneic hematopoietic stem cell transplantation (HSCT). Seventeen adult patients 11-13 months after HSCT and eight unvaccinated healthy adults received up to three TBE vaccinations. Following in vitro stimulation with TBE-antigen, lymphocyte proliferation and cytokine secretion (IL-2, IL-10, IL-13, TNF-alpha, IFN-gamma, GM-CSF) were analyzed by thymidine incorporation assay and the Luminex system. Ten patients (59%) showed significant baseline TBE-specific lymphocyte proliferation (stimulation index (SI) > 3) prior to vaccination, but none of the unvaccinated controls (p = 0.002). All patients with a TBE-specific antibody response after two vaccinations (at least 2-fold increase of neutralization test titers) exhibited a strong TBE-specific lymphocyte proliferative response at baseline (SI > 10). Patients with sibling donors had a significantly stronger baseline TBE-specific lymphocyte proliferative and IL-13 cytokine response than patients with unrelated donors (p < 0.05). In conclusion, a relevant proportion of patients showed TBE-specific lymphocyte proliferative and cytokine responses prior to vaccination after HSCT, which predicted the humoral response to the vaccine. Patients with vaccinated sibling donors were more likely to elicit a cellular immune response than patients with unrelated donors of unknown vaccination status.

3.
Sci Rep ; 11(1): 13592, 2021 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-34193912

RESUMO

With global demand for SARS-CoV-2 testing ever rising, shortages in commercially available viral transport media pose a serious problem for laboratories and health care providers. For reliable diagnosis of SARS-CoV-2 and other respiratory viruses, executed by Real-time PCR, the quality of respiratory specimens, predominantly determined by transport and storage conditions, is crucial. Therefore, our aim was to explore the reliability of minimal transport media, comprising saline or the CDC recommended Viral Transport Media (HBSS VTM), for the diagnosis of SARS-CoV-2 and other respiratory viruses (influenza A, respiratory syncytial virus, adenovirus, rhinovirus and human metapneumovirus) compared to commercial products, such as the Universal Transport Media (UTM). We question the assumptions, that the choice of medium and temperature for storage and transport affect the accuracy of viral detection by RT-PCR. Both alternatives to the commercial transport medium (UTM), HBSS VTM or saline, allow adequate detection of SARS-CoV-2 and other respiratory viruses, regardless of storage temperatures up to 28 °C and storage times up to 28 days. Our study revealed the high resilience of SARS-CoV-2 and other respiratory viruses, enabling proper detection in clinical specimens even after long-time storage at high temperatures, independent of the transport medium's composition.


Assuntos
Teste de Ácido Nucleico para COVID-19/métodos , COVID-19/diagnóstico , Meios de Cultura/química , Preservação Biológica/métodos , SARS-CoV-2/genética , Manejo de Espécimes/métodos , Virologia/métodos , Temperatura Baixa , Humanos , Reagentes de Laboratório/química , Reprodutibilidade dos Testes , Fatores de Tempo
4.
Liver Int ; 41(11): 2622-2634, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34268869

RESUMO

BACKGROUND AND AIMS: Despite vaccination recommendations, hepatitis B (HBV) and D (HDV) coinfections are common in HIV+individuals. METHODS: HBV immunization status (anti-HBs) as well as HBV (HBsAg/HBV-DNA) and HDV (anti-HDV) coinfection rates were assessed in 1870 HIV+individuals at HIV diagnosis (baseline, BL) and last follow-up (FU). RESULTS: Sixty-eight (3.6%) HIV patients were never tested for HBV. At BL, 89/1802 (4.9%) HIV patients were HBV coinfected. Four hundred and fifteen (23.0%) showed virological HBV clearance [HBsAg(-)/anti-HBc(+)/anti-HBs(+)] and 210 (11.7%) presented with anti-HBc(+) only. Seven hundred and ten (39.4%) were HBV naïve [HBsAg(-)/anti-HBs(-)/anti-HBc(-)/HBV-DNA(-)], but only 378 (21.0%) received vaccinations with detectable anti-HBs(+) titres. Among the 89 HBV/HIV-coinfected patients, only 52 (58.4%) were tested for HDV: 11/49 (22.4%) had anti-HDV(+) and 3/12 (25.0%) showed HDV-RNA viraemia. During a median FU of 6.5 (IQR 7.2) years, 44 (4.6%) of the 953 retested BL HBV-negative patients acquired new HBV infection (including 15/304, 4.9% of vaccinated patients). Of the 89 patients, 22 (24.7%) patients cleared their HBsAg, resulting in 60/1625 (3.7%) HIV/HBV individuals at FU: 34 (56.7%) showed HBV-DNA suppression and 15 (25.0%) were HBV viraemic, while 12/89 (13.5%) remained without a FU test. Vaccinations induced anti-HBs(+) in 137 of the retested 649 (21.1%) BL HBV-naïve patients. CONCLUSION: HBV testing is well established among Viennese HIV+patients with HBV coinfection rates around 4%-5%. HBV vaccinations are insufficiently implemented since anti-HBs titres were detected in only 21.1% of HBV-naive HIV(+) patients and new HBV infections occurred in previously vaccinated patients. HDV testing is not systematically performed despite up to 25% of HIV/HBV patients may show HDV coinfection.


Assuntos
Coinfecção , Infecções por HIV , Hepatite B , Coinfecção/epidemiologia , Infecções por HIV/complicações , Infecções por HIV/epidemiologia , Hepatite B/epidemiologia , Antígenos de Superfície da Hepatite B , Vírus da Hepatite B , Humanos
5.
BMC Med Imaging ; 21(1): 107, 2021 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-34238246

RESUMO

BACKGROUND: To develop a regression neural network for the reconstruction of lesion probability maps on Magnetic Resonance Fingerprinting using echo-planar imaging (MRF-EPI) in addition to [Formula: see text], [Formula: see text], NAWM, and GM- probability maps. METHODS: We performed MRF-EPI measurements in 42 patients with multiple sclerosis and 6 healthy volunteers along two sites. A U-net was trained to reconstruct the denoised and distortion corrected [Formula: see text] and [Formula: see text] maps, and to additionally generate NAWM-, GM-, and WM lesion probability maps. RESULTS: WM lesions were predicted with a dice coefficient of [Formula: see text] and a lesion detection rate of [Formula: see text] for a threshold of 33%. The network jointly enabled accurate [Formula: see text] and [Formula: see text] times with relative deviations of 5.2% and 5.1% and average dice coefficients of [Formula: see text] and [Formula: see text] for NAWM and GM after binarizing with a threshold of 80%. CONCLUSION: DL is a promising tool for the prediction of lesion probability maps in a fraction of time. These might be of clinical interest for the WM lesion analysis in MS patients.

6.
Bioorg Med Chem ; 40: 116163, 2021 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-33932711

RESUMO

Bruton's tyrosine kinase (BTK) is a cytoplasmic, non-receptor tyrosine kinase member of the TEC family of tyrosine kinases. Pre-clinical and clinical data have shown that targeting BTK can be used for the treatment for B-cell disorders. Here we disclose the discovery of a novel imidazo[4,5-b]pyridine series of potent, selective reversible BTK inhibitors through a rational design approach. From a starting hit molecule 1, medicinal chemistry optimization led to the development of a lead compound 30, which exhibited 58 nM BTK inhibitory potency in human whole blood and high kinome selectivity. Additionally, the compound demonstrated favorable pharmacokinetics (PK), and showed potent dose-dependent efficacy in a rat CIA model.


Assuntos
Tirosina Quinase da Agamaglobulinemia/antagonistas & inibidores , Descoberta de Drogas , Imidazóis/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Tirosina Quinase da Agamaglobulinemia/metabolismo , Relação Dose-Resposta a Droga , Humanos , Imidazóis/síntese química , Imidazóis/química , Estrutura Molecular , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Piridinas/síntese química , Piridinas/química , Relação Estrutura-Atividade
7.
Medicine (Baltimore) ; 100(12): e25170, 2021 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-33761694

RESUMO

RATIONALE: The immunologic syndrome induced by severe acute coronavirus disease 2019 (COVID-19) is yet not fully understood. Typical patterns of clinical and laboratory features match secondary hemophagocytic lymphohistiocytosis (sHLH). However, the optimal approach to COVID-19 patients testing positive for sHLH is still unclear. PATIENT CONCERNS: Three patients with COVID-19 are reviewed. All showed hyperinflammation and cytokine storm, necessitating intensive care treatment including mechanical ventilation. DIAGNOSIS: Secondary hemophagocytic lymphohistiocytosis due to severe COVID-19; diagnosed via HScore. INTERVENTIONS: A treatment regimen of methylprednisolone, pentaglobin, and anakinra was developed and administered. OUTCOMES: One patient survived the ICU stay. Two other patients, in whom sHLH was diagnosed too late, deceased. LESSONS: A routine screening of COVID-19 patients for secondary HLH by using the HScore is feasible; especially those patients deteriorating clinically with no sufficient response to shock management might be at particular high risk. A stepwise therapeutic approach comprising corticosteroids, immunoglobulins and anakinra, accompanied by immunoadsorption, may dampen cytokine storm effects, and potentially reduce mortality.


Assuntos
COVID-19/complicações , Linfo-Histiocitose Hemofagocítica/tratamento farmacológico , Linfo-Histiocitose Hemofagocítica/etiologia , Idoso , Anti-Inflamatórios/uso terapêutico , COVID-19/fisiopatologia , COVID-19/terapia , Terapia Combinada , Cuidados Críticos , Síndrome da Liberação de Citocina/tratamento farmacológico , Diagnóstico Tardio , Evolução Fatal , Feminino , Humanos , Imunoglobulina A/uso terapêutico , Imunoglobulina M/uso terapêutico , Imunoglobulinas Intravenosas/uso terapêutico , Imunossupressores/uso terapêutico , Proteína Antagonista do Receptor de Interleucina 1/uso terapêutico , Linfo-Histiocitose Hemofagocítica/diagnóstico , Masculino , Metilprednisolona/uso terapêutico , Pessoa de Meia-Idade , Respiração Artificial , SARS-CoV-2
8.
Magn Reson Med ; 86(1): 471-486, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33547656

RESUMO

PURPOSE: To develop an accelerated postprocessing pipeline for reproducible and efficient assessment of white matter lesions using quantitative magnetic resonance fingerprinting (MRF) and deep learning. METHODS: MRF using echo-planar imaging (EPI) scans with varying repetition and echo times were acquired for whole brain quantification of T 1 and T 2 ∗ in 50 subjects with multiple sclerosis (MS) and 10 healthy volunteers along 2 centers. MRF T 1 and T 2 ∗ parametric maps were distortion corrected and denoised. A CNN was trained to reconstruct the T 1 and T 2 ∗ parametric maps, and the WM and GM probability maps. RESULTS: Deep learning-based postprocessing reduced reconstruction and image processing times from hours to a few seconds while maintaining high accuracy, reliability, and precision. Mean absolute error performed the best for T 1 (deviations 5.6%) and the logarithmic hyperbolic cosinus loss the best for T 2 ∗ (deviations 6.0%). CONCLUSIONS: MRF is a fast and robust tool for quantitative T 1 and T 2 ∗ mapping. Its long reconstruction and several postprocessing steps can be facilitated and accelerated using deep learning.


Assuntos
Aprendizado Profundo , Substância Branca , Encéfalo/diagnóstico por imagem , Humanos , Processamento de Imagem Assistida por Computador , Imageamento por Ressonância Magnética , Espectroscopia de Ressonância Magnética , Imagens de Fantasmas , Reprodutibilidade dos Testes , Substância Branca/diagnóstico por imagem
9.
IEEE Trans Biomed Eng ; 68(5): 1518-1526, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33275574

RESUMO

OBJECTIVE: Three-dimensional (3D) blood vessel structure information is important for diagnosis and treatment in various clinical scenarios. We present a fully automatic method for the extraction and differentiation of the arterial and venous vessel trees from abdominal contrast enhanced computed tomography (CE-CT) volumes using convolutional neural networks (CNNs). METHODS: We used a novel ratio-based sampling method to train 2D and 3D versions of the U-Net, the V-Net and the DeepVesselNet. Networks were trained with a combination of the Dice and cross entropy loss. Performance was evaluated on 20 IRCAD subjects. Best performing networks were combined into an ensemble. We investigated seven different weighting schemes. Trained networks were additionally applied to 26 BTCV cases to validate the generalizability. RESULTS: Based on our experiments, the optimal configuration is an equally weighted ensemble of 2D and 3D U- and V-Nets. Our method achieved Dice similarity coefficients of 0.758 ± 0.050 (veins) and 0.838 ± 0.074 (arteries) on the IRCAD data set. Application to the BTCV data set showed a high transfer ability. CONCLUSION: Abdominal vascular structures can be segmented more accurately using ensembles than individual CNNs. 2D and 3D networks have complementary strengths and weaknesses. Our ensemble of 2D and 3D U-Nets and V-Nets in combination with ratio-based sampling achieves a high agreement with manual annotations for both artery and vein segmentation. Our results surpass other state-of-the-art methods. SIGNIFICANCE: Our segmentation pipeline can provide valuable information for the planning of living donor organ transplantations.


Assuntos
Redes Neurais de Computação , Tomografia Computadorizada por Raios X , Abdome/diagnóstico por imagem , Artérias , Humanos , Processamento de Imagem Assistida por Computador
10.
Eur J Clin Microbiol Infect Dis ; 40(2): 335-344, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32940811

RESUMO

To explore the epidemiology and clinical course of hepatitis A virus (HAV) infections at the Vienna General Hospital. We retrospectively identified patients who were tested positive for HAV-IgM at the Vienna General Hospital form Q1/2008 to Q3/2018. Our definition of severe HAV infection was AST and/or ALT > 5 × above the upper limit of normal (ULN); and liver dysfunction as (i) hepatic encephalopathy or ammonia > 100 µmol/L, (ii) coagulopathy with INR > 1.5, or (iii) jaundice with bilirubin > 5 mg/dL. A total of 578 HAV-IgM (+) were identified, including 31 (5.4%) and 38 (6.6%) without and with liver dysfunction, respectively. A proportional increase in severe HAV cases with and without liver dysfunction occurred in 2016/2017 with (21.5% (vs. 8.0% in the years before; p < 0.001). Thirty-seven (53.6%) patients with severe HAV were hospitalized, 6 (9%) required ICU support, and one patient received liver transplantation within 30 days. Patients with severe HAV and liver dysfunction were more often male (60.5 vs. 43.1%, p = 0.055) and younger (31.5 vs. 63 years, p < 0.001) as compared with other HAV-IgM (+) cases. The observed increase of severe HAV infections in Vienna in 2017 among young males, coincided with a multinational HAV outbreak among MSM. Our data suggests a higher likelihood of severe courses of hepatitis A in MSM. Vaccination against HAV should be recommended for risk groups.


Assuntos
Surtos de Doenças , Hepatite A/epidemiologia , Adulto , Áustria/epidemiologia , Feminino , Vírus da Hepatite A Humana/isolamento & purificação , Hospitais Gerais , Hospitais Urbanos , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Fatores de Risco , Minorias Sexuais e de Gênero
11.
FEBS J ; 288(2): 640-662, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32386462

RESUMO

Nuclear factor 'κ-light-chain-enhancer' of activated B cells (NF-κB) signaling is a signaling pathway used by most immune cells to promote immunostimulatory functions. Recent studies have indicated that regulatory T cells (Treg) differentially integrate TCR-derived signals, thereby maintaining their suppressive features. However, the role of NF-κB signaling in the activation of human peripheral blood (PB) Treg has not been fully elucidated so far. We show that the activity of the master transcription factor forkhead box protein 3 (FOXP3) attenuates p65 phosphorylation and nuclear translocation of the NF-κB proteins p50, p65, and c-Rel following activation in human Treg. Using pharmacological and genetic inhibition of canonical NF-κB signaling in FOXP3-transgenic T cells and PB Treg from healthy donors as well as Treg from a patient with a primary NFKB1 haploinsufficiency, we validate that Treg activation and suppressive capacity is independent of NF-κB signaling. Additionally, repression of residual NF-κB signaling in Treg further enhances interleukin-10 (IL-10) production. Blockade of NF-κB signaling can be exploited for the generation of in vitro induced Treg (iTreg) with enhanced suppressive capacity and functional stability. In this respect, dual blockade of mammalian target of rapamycin (mTOR) and NF-κB signaling was accompanied by enhanced expression of the transcription factors FOXP1 and FOXP3 and demethylation of the Treg-specific demethylated region compared to iTreg generated under mTOR blockade alone. Thus, we provide first insights into the role of NF-κB signaling in human Treg. These findings could lead to strategies for the selective manipulation of Treg and the generation of improved iTreg for cellular therapy.


Assuntos
Fatores de Transcrição Forkhead/imunologia , Haploinsuficiência/imunologia , Subunidade p50 de NF-kappa B/imunologia , Linfócitos T Reguladores/imunologia , Serina-Treonina Quinases TOR/imunologia , Fator de Transcrição RelA/imunologia , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Transporte Ativo do Núcleo Celular/imunologia , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/imunologia , Núcleo Celular/metabolismo , Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica , Humanos , Interleucina-10/genética , Interleucina-10/imunologia , Ativação Linfocitária , Subunidade p50 de NF-kappa B/deficiência , Subunidade p50 de NF-kappa B/genética , Fosforilação/efeitos dos fármacos , Cultura Primária de Células , Proteínas Repressoras/genética , Proteínas Repressoras/imunologia , Transdução de Sinais , Sirolimo/farmacologia , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/efeitos dos fármacos , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/genética , Tiazóis/farmacologia , Fator de Transcrição RelA/antagonistas & inibidores , Fator de Transcrição RelA/genética
12.
GMS J Med Educ ; 37(7): Doc66, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33364345

RESUMO

Objective: During the early Covid 19 pandemic, undergraduate medical teaching of pediatric medicine had to be switched to online teaching at the Hanover Medical School (MHH). The aim was to develop an online module together with students. Methodology: In a multi-stage process, a working group consisting of lecturers and students developed the concept and implemented it. Afterwards the online module was evaluated. Results: The conceptualization process and the implementation of the module together with students can be represented as a modified PDCA cycle (Plan-Do-Check-Act). We showed that including students in the development of an online module is helpful in times of limited resources e.g. such as personnel and time. Conclusion: The cooperation between students and lecturers is suitable for developing and implementing an online module in a short time. In the future, in addition to joint conceptualization phases, digital elements (e.g. preparatory webinars) for the module itself in attendance phases should be retained.


Assuntos
COVID-19/epidemiologia , Instrução por Computador/métodos , Educação à Distância/organização & administração , Docentes de Medicina/organização & administração , Pediatria/educação , Estudantes de Medicina , Educação de Graduação em Medicina/organização & administração , Humanos , Pandemias , SARS-CoV-2
13.
Nat Commun ; 11(1): 6341, 2020 12 11.
Artigo em Inglês | MEDLINE | ID: mdl-33311468

RESUMO

Mutations in the RNA-binding protein Fused in Sarcoma (FUS) cause early-onset amyotrophic lateral sclerosis (ALS). However, a detailed understanding of central RNA targets of FUS and their implications for disease remain elusive. Here, we use a unique blend of crosslinking and immunoprecipitation (CLIP) and NMR spectroscopy to identify and characterise physiological and pathological RNA targets of FUS. We find that U1 snRNA is the primary RNA target of FUS via its interaction with stem-loop 3 and provide atomic details of this RNA-mediated mode of interaction with the U1 snRNP. Furthermore, we show that ALS-associated FUS aberrantly contacts U1 snRNA at the Sm site with its zinc finger and traps snRNP biogenesis intermediates in human and murine motor neurons. Altogether, we present molecular insights into a FUS toxic gain-of-function involving direct and aberrant RNA-binding and strengthen the link between two motor neuron diseases, ALS and spinal muscular atrophy (SMA).


Assuntos
Esclerose Amiotrófica Lateral/metabolismo , RNA Nuclear Pequeno/metabolismo , Proteína FUS de Ligação a RNA/genética , Proteína FUS de Ligação a RNA/metabolismo , Ribonucleoproteína Nuclear Pequena U1/metabolismo , Esclerose Amiotrófica Lateral/genética , Animais , Linhagem Celular , Predisposição Genética para Doença/genética , Humanos , Camundongos , Camundongos Knockout , Modelos Moleculares , Neurônios Motores/metabolismo , Mutação , Domínios e Motivos de Interação entre Proteínas , RNA Nuclear Pequeno/química , Proteína FUS de Ligação a RNA/química , Ribonucleoproteína Nuclear Pequena U1/química
14.
Int J Mol Sci ; 21(21)2020 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-33143232

RESUMO

The purpose of this study was to investigate the tissue regenerating and biomechanical properties of processed eggshell membrane powder (PEP) for use in 3D-scaffolds. PEP is a low-cost, natural biomaterial with beneficial bioactive properties. Most importantly, this material is available as a by-product of the chicken egg processing (breaking) industry on a large scale, and it could have potential as a low-cost ingredient for therapeutic scaffolds. Scaffolds consisting of collagen alone and collagen combined with PEP were produced and analyzed for their mechanical properties and the growth of primary fibroblasts and skeletal muscle cells. Mechanical testing revealed that a PEP/collagen-based scaffold increased the mechanical hardness of the scaffold compared with a pure collagen scaffold. Scanning electron microscopy (SEM) demonstrated an interconnected porous structure for both scaffolds, and that the PEP was evenly distributed in dense clusters within the scaffold. Fibroblast and skeletal muscle cells attached, were viable and able to proliferate for 1 and 2 weeks in both scaffolds. The cell types retained their phenotypic properties expressing phenotype markers of fibroblasts (TE7, alpha-smooth muscle actin) and skeletal muscle (CD56) visualized by immunostaining. mRNA expression of the skeletal muscle markers myoD, myogenin, and fibroblasts marker (SMA) together with extracellular matrix components supported viable phenotypes and matrix-producing cells in both types of scaffolds. In conclusion, PEP is a promising low-cost, natural biomaterial for use in combination with collagen as a scaffold for 3D-tissue engineering to improve the mechanical properties and promote cellular adhesion and growth of regenerating cells.


Assuntos
Materiais Biocompatíveis/química , Casca de Ovo/química , Matriz Extracelular/química , Fibroblastos/citologia , Músculo Esquelético/citologia , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Bovinos , Células Cultivadas , Humanos , Pós/química
15.
J Exp Bot ; 71(22): 7146-7159, 2020 12 31.
Artigo em Inglês | MEDLINE | ID: mdl-32911544

RESUMO

The number of flowers and seed-bearing structures formed by the inflorescence meristem and the formation of ovules in the female reproductive part of the flowers are important yield-related traits of crop plants. It has been shown that cytokinin is a pivotal factor regulating these traits. Here, we explore the impact of mutation of CYTOKININ OXIDASE/DEHYDROGENASE (CKX) genes encoding cytokinin-degrading enzymes on these yield-related traits in oilseed rape (Brassica napus L.). We describe the identification of four BnCKX3 and two BnCKX5 genes as regulators of reproductive development in the allotetraploid B. napus. RNA-seq analysis and in situ hybridization showed expression of these genes in reproductive organs. Loss-of-function mutants for each of these CKX gene copies were identified by targeting induced local lesions in genomes (TILLING) and combined by crossing. Sextuple ckx3 ckx5 mutants showed an increased cytokinin concentration and larger and more active inflorescence meristems. They also produced up to 72% more flowers with gynoecia containing 32% more ovules and up to 54% more pods on the main stem. The weight of seeds harvested from the main stem of plants grown in the greenhouse or in the field was increased by 20-32%. Our results show that cytokinin regulates inflorescence meristem and placenta activity in oilseed rape. The work demonstrates the potential to achieve yield enhancement in a dicot crop plant by modulating the cytokinin status through mutagenesis of specific CKX genes.


Assuntos
Brassica napus , Brassica napus/genética , Citocininas , Inflorescência/genética , Meristema/genética , Sementes/genética
16.
J Clin Oncol ; 38(30): 3547-3554, 2020 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-32795227

RESUMO

PURPOSE: To analyze the prevalence of SARS-CoV-2 infection in patients with cancer in hospital care after implementation of institutional and governmental safety measurements. METHODS: Patients with cancer routinely tested for SARS-CoV-2 RNA by nasal swab and real-time polymerase chain reaction between March 21 and May 4, 2020, were included. The results of this cancer cohort were statistically compared with the SARS-CoV-2 prevalence in the Austrian population as determined by a representative nationwide random sample study (control cohort 1) and a cohort of patients without cancer presenting to our hospital (control cohort 2). RESULTS: A total of 1,688 SARS-CoV-2 tests in 1,016 consecutive patients with cancer were performed. A total of 270 of 1,016 (26.6%) of the patients were undergoing active anticancer treatment in a neoadjuvant/adjuvant and 560 of 1,016 (55.1%) in a palliative setting. A total of 53 of 1,016 (5.2%) patients self-reported symptoms potentially associated with COVID-19. In 4 of 1,016 (0.4%) patients, SARS-CoV-2 was detected. At the time of testing at our department, all four SARS-CoV-2-positive patients were asymptomatic, and two of them had recovered from symptomatic COVID-19. Viral clearance was achieved in three of the four patients 14-56 days after testing positive. The estimated odds ratio of SARS-CoV-2 prevalence between the cancer cohort and control cohort 1 was 1.013 (95% CI, 0.209 to 4.272; P = 1), and between control cohort 2 and the cancer cohort it was 18.333 (95% CI, 6.056 to 74.157). CONCLUSION: Our data indicate that continuation of active anticancer therapy and follow-up visits in a large tertiary care hospital are feasible and safe after implementation of strict population-wide and institutional safety measures during the current COVID-19 pandemic. Routine SARS-CoV-2 testing of patients with cancer seems advisable to detect asymptomatic virus carriers and avoid uncontrolled viral spread.


Assuntos
Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/diagnóstico , Neoplasias/virologia , Pneumonia Viral/diagnóstico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , COVID-19 , Estudos de Coortes , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias/tratamento farmacológico , Pandemias , SARS-CoV-2 , Centros de Atenção Terciária , Adulto Jovem
17.
Virulence ; 11(1): 964-967, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32726172

RESUMO

Currently, testing for coronavirus is performed with time and personnel consuming PCR assays. The aim of this study was to evaluate the sensitivity, specificity and capacity of a fully automated, random access high-throughput real-time PCR-based diagnostic platform for the detection of SARS-CoV-2. The NeuMoDx N96 system displayed an equal or better detection rate for SARS-CoV-2 compared with the LightCycler 480II system and showed a specificity of 100%. The median PCR run time for all 28 PCR runs was 91 (IQR 84-97) minutes. The capacity of the NeuMoDx N96 could easily surpass the capacity of most currently used molecular test systems and significantly reduce the turn-around time.


Assuntos
Betacoronavirus/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala/métodos , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real/métodos , Betacoronavirus/genética , Sequenciamento de Nucleotídeos em Larga Escala/instrumentação , Sequenciamento de Nucleotídeos em Larga Escala/normas , Humanos , RNA Viral/genética , Reação em Cadeia da Polimerase em Tempo Real/instrumentação , Reação em Cadeia da Polimerase em Tempo Real/normas , Reprodutibilidade dos Testes , SARS-CoV-2 , Sensibilidade e Especificidade , Fatores de Tempo
18.
FASEB J ; 34(6): 8367-8384, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32319705

RESUMO

The ectonucleotidase CD39 on human regulatory T-cells (Treg) is an important immune regulator which is dysregulated in autoimmune diseases and cancer immunosuppression. We here define that CD39 expression on Treg is independent of the Treg-specific transcription factors FOXP3 and HELIOS and promoted by canonical TGF-b- and mTOR-signaling. Furthermore, the TGF-b mediated upregulation of CD39 is counteracted by reactive oxygen species (ROS)-driven autophagy. In line, CD39+ peripheral blood Treg constitute a distinct lineage with low autophagic flux and absent ROS production. Patients with rare genetic defects in autophagy show supraphysiological levels of CD39+ Treg, validating our observations in vivo. These biological processes rely on a distinct transcriptional program with CD39+ Treg expressing low levels of two genes with putative involvement in autophagy, NEFL and PLAC8. Furthermore, the TGF-b downstream transcription factor SOX4 is selectively upregulated in CD39+ Treg. Overexpression of SOX4 in Treg strongly increases CD39 expression, while Crispr/Cas9-mediated knockout of SOX4 in Treg has the opposing effect. Thus, we identify a crucial role of SOX4 in immune regulation and provide new insights involving the interplay of tolerogenic cues and autophagy in Treg.


Assuntos
Apirase/imunologia , Espécies Reativas de Oxigênio/imunologia , Fatores de Transcrição SOXC/imunologia , Linfócitos T Reguladores/imunologia , Fator de Crescimento Transformador beta/imunologia , Adulto , Células Cultivadas , Feminino , Humanos , Tolerância Imunológica/imunologia , Fatores Imunológicos/imunologia , Masculino , Transdução de Sinais/imunologia
19.
Liver Int ; 40(4): 787-796, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32017359

RESUMO

BACKGROUND & AIMS: Human immunodeficiency virus (HIV)/hepatitis C virus (HCV) coinfection is common in people who inject drugs (PWIDs). Recently, 'high-risk' behaviour among men who have sex with men (MSM) has emerged as another main route of HCV transmission. We analysed temporal trends in HCV epidemiology in a cohort of Viennese HIV+ patients. METHODS: Hepatitis C virus parameters were recorded at HIV diagnosis (baseline [BL]) and last visit (follow-up [FU]) for all HIV+ patients attending our HIV clinic between January 2014 and December 2016. Proportions of HIV+ patients with anti-HCV(+) and HCV viraemia (HCV-RNA(+)) at BL/FU were assessed and stratified by route of transmission. RESULTS: In all, 1806/1874 (96.4%) HIV+ patients were tested for HCV at BL. Anti-HCV(+) was detected in 93.2% (276/296) of PWIDs and in 3.7% (31/839) of MSM. After a median FU of 6.9 years, 1644 (91.0%) patients underwent FU HCV-testing: 167 (90.3%) of PWIDs and 49 (6.7%) of MSM showed anti-HCV(+). Among 208 viraemic HCV-RNA(+) patients at BL, 30 (14.4%) had spontaneously cleared HCV, 76 (36.5%) achieved treatment-induced eradication and 89 (42.8%) remained HCV-RNA(+) at last FU. Among 1433 initially HCV-naive patients, 45 (3.5%) acquired de-novo HCV infection (11.1% PWIDs/80.0% MSM; incidence rate (IR) 0.004%; 95% confidence interval [CI] 0.0%-0.022%) and 14 had HCV reinfections (85.7% PWIDs/14.3% other; IR 0.001%; 95% CI 0.0%-0.018%) during a median FU of 6.7 years (interquartile range 7.4). CONCLUSION: Hepatitis C virus testing was successfully implemented in the Viennese HIV(+) patients. Anti-HCV(+) prevalence remained stable in HIV+ PWIDs but almost doubled in HIV+ MSM. De-novo HCV infection occurred mostly in MSM, while HCV reinfections were mainly observed in PWIDs. HCV treatment uptake was suboptimal with 42.8% remaining HCV-RNA(+) at FU.


Assuntos
Infecções por HIV , Hepatite C , Minorias Sexuais e de Gênero , Infecções por HIV/diagnóstico , Infecções por HIV/tratamento farmacológico , Infecções por HIV/epidemiologia , Hepacivirus , Hepatite C/diagnóstico , Hepatite C/epidemiologia , Homossexualidade Masculina , Humanos , Masculino , Prevalência
20.
Liver Int ; 40(2): 393-404, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31967400

RESUMO

BACKGROUND & AIMS: The loss-of-function rs72613567 T > TA-variant in the 17ß-hydroxysteroid dehydrogenase 13 (HSD17B13) gene might protect from alcoholic and non-alcoholic fatty liver disease (ALD/NAFLD) and associated fibrosis/cirrhosis. We investigated the impact of the T > TA-variant on hepatic decompensation and mortality and investigated its implications on retinol and sex steroid metabolism in patients who had already developed advanced chronic liver disease (ACLD). METHODS: Retrospective analysis in prospectively characterized patients with viral hepatitis- and ALD/NAFLD-induced portal hypertension (hepatic venous pressure gradient (HVPG) ≥ 6 mmHg) diagnosed at the Medical University of Vienna. RESULTS: Among 487 patients who were followed longitudinally, 166 (34%) were heterozygous and 24 (5%) were homozygous for the 'protective' TA-allele. Patients harbouring at least one TA-allele had a lower MELD (9 (8-12) vs 10 (8-13) points; P = .003) and showed a trend towards lower HVPG (16 ± 6 vs 17 ± 7 mmHg; P = .067). Interestingly, in competing risk analyses adjusted for age, HVPG and MELD, harbouring the TA-allele was associated with numerically increased risks for mortality (adjusted subdistribution hazard ratio (aSHR): 1.3 (95% confidence interval (95% CI): 0.888-1.91); P = .18), liver-related death (aSHR: 1.34 (95% CI: 0.9-1.98); P = .15) and hepatic decompensation (aSHR: 1.29 (95% CI: 0.945-1.77); P = .11). This might be explained by trends towards worse outcomes (eg liver-related death: aSHR: 1.64 (95% CI: 0.95-2.84); P = .076) in patients with viral hepatitis-induced ACLD. In a cross-sectional analysis of 211 additional patients, serum retinol levels were comparable between HSD17B13 genotypes, but in males, serum testosterone levels numerically decreased with an increasing number of TA-alleles. CONCLUSION: In patients with viral hepatitis- and ALD-induced portal hypertension, the T > TA-variant was not protective of hepatic decompensation and mortality. Further studies should investigate the pathophysiological mechanisms underlying the effects of HSD17B13 genotype at different stages of liver disease.


Assuntos
17-Hidroxiesteroide Desidrogenases/genética , Hipertensão Portal , Estudos Transversais , Genótipo , Humanos , Hipertensão Portal/genética , Hipertensão Portal/mortalidade , Cirrose Hepática/genética , Masculino , Estudos Retrospectivos
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