RESUMO
Metabolic dysfunction-associated steatotic liver disease (MASLD), formerly known as non-alcoholic fatty liver disease (NAFLD), encompasses a progressive spectrum of liver conditions, ranging from steatosis to metabolic dysfunction-associated steatohepatitis, characterised by hepatocellular death and inflammation, potentially progressing to cirrhosis and/or liver cancer. In both experimental and human MASLD, necroptosis-a regulated immunogenic necrotic cell death pathway-is triggered, yet its exact role in disease pathogenesis remains unclear. Noteworthy, necroptosis-related signalling pathways are emerging as key players in metabolic reprogramming, including lipid and mitochondrial metabolism. Additionally, metabolic dysregulation is a well-established contributor to MASLD development and progression. This review explores the intricate interplay between cell metabolism and necroptosis regulation and its impact on MASLD pathogenesis. Understanding these cellular events may offer new insights into the complexity of MASLD pathophysiology, potentially uncovering therapeutic opportunities and unforeseen metabolic consequences of targeting necroptosis.
Assuntos
Necroptose , Hepatopatia Gordurosa não Alcoólica , Humanos , Necroptose/fisiologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Animais , Transdução de Sinais/fisiologiaRESUMO
Conventional plastics are being slowly replaced by biodegradable ones to prevent plastic pollution. However, in the natural environment, the biodegradation of plastics is usually slow or incomplete due to unfavorable conditions and leads to faster micro-bioplastic formation. Many analytical methods were developed to determine microplastics, but micro-bioplastics are still overlooked. This work presents a simple method for determining poly-3-hydroxybutyrate and polylactic acid micro-bioplastics in soil based on the thermogravimetry-mass spectrometry analysis of low molecular gases evolved during pyrolysis. For the method development, model soils containing different soil organic carbon contents were spiked with micro-bioplastics. Specific gaseous pyrolysis products of the analytes were identified, while the ratio of their amounts appeared to be constant above the level of detection of the suggested method. The constant ratio was explained as a lower soil influence on the evolution of the gaseous product, and it was suggested as an additional identification parameter. The advantages of the presented method are no sample pretreatment, presumably no need for an internal standard, low temperature needed for the transfer of gaseous products and the possibility of using its principles with other, cheaper detectors. The method can find application in the verification of biodegradation tests and in the monitoring of soils after the application of biodegradable products.
Assuntos
Plásticos , Solo , Biodegradação Ambiental , Carbono , Gases , Plásticos/química , Poliésteres , Solo/químicaRESUMO
Acquired epidermodysplasia verruciformis is a rare disease. It can develop in immunocompromised patients due to infection with human papillomaviruses. Because such patients are at high risk of developing cutaneous squamous cell carcinoma, timely diagnosis and regular monitoring of the patient is essential. Here we present the case of a 46-year-old male patient with acquired epidermodysplasia verruciformis occurring 5 years after a kidney transplantation. A skin biopsy detected human papillomavirus genotype 20 with low oncogenic potential. Accordingly, a follow-up interval of 1 year was determined. He was instructed to follow strict photoprotection and to visit earlier if atypical lesions appeared. Overall, our case emphasizes the consideration of possible squamous cell carcinoma in such patients and the importance of appropriate preventive measures.
Assuntos
Carcinoma de Células Escamosas , Epidermodisplasia Verruciforme , Transplante de Rim , Neoplasias Cutâneas , Epidermodisplasia Verruciforme/etiologia , Epidermodisplasia Verruciforme/patologia , Humanos , Transplante de Rim/efeitos adversos , Masculino , Pessoa de Meia-Idade , Papillomaviridae/genética , Neoplasias Cutâneas/etiologiaRESUMO
The cGAS-STING pathway is known for its role in sensing cytosolic DNA introduced by a viral infection, bacterial invasion or tumorigenesis. Free DNA is recognized by the cyclic GMP-AMP synthase (cGAS) catalyzing the production of 2',3'-cyclic guanosine monophosphate-adenosine monophosphate (2',3'-cGAMP) in mammals. This cyclic dinucleotide acts as a second messenger, activating the stimulator of interferon genes (STING) that finally triggers the transcription of interferon genes and inflammatory cytokines. Due to the therapeutic potential of this pathway, both the production and the detection of cGAMP via fluorescent moieties for assay development is of great importance. Here, we introduce the paralleled synthetic access to the intrinsically fluorescent, cyclic dinucleotides 2'3'-cth GAMP and 3'3'-cth GAMP based on phosphoramidite and phosphate chemistry, adaptable for large scale synthesis. We examine their binding properties to murine and human STING and confirm biological activity including interferon induction by 2'3'-cth GAMP in THP-1 monocytes. Two-photon imaging revealed successful cellular uptake of 2'3'-cth GAMP in THP-1 cells.
Assuntos
Proteínas de Membrana , Nucleotidiltransferases , Animais , DNA/metabolismo , Fosfatos de Dinucleosídeos , Humanos , Interferons , Mamíferos/genética , Mamíferos/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Nucleotidiltransferases/metabolismo , Sistemas do Segundo MensageiroRESUMO
The cyclic dinucleotide second messenger c-di-AMP is a major player in regulation of potassium homeostasis and osmolyte transport in a variety of bacteria. Along with various direct interactions with proteins such as potassium channels, the second messenger also specifically binds to transcription factors, thereby altering the processes in the cell on the transcriptional level. We here describe the structural and biochemical characterization of BusR from the human pathogen Streptococcus agalactiae. BusR is a member of a yet structurally uncharacterized subfamily of the GntR family of transcription factors that downregulates transcription of the genes for the BusA (OpuA) glycine-betaine transporter upon c-di-AMP binding. We report crystal structures of full-length BusR, its apo and c-di-AMP bound effector domain, as well as cryo-EM structures of BusR bound to its operator DNA. Our structural data, supported by biochemical and biophysical data, reveal that BusR utilizes a unique domain assembly with a tetrameric coiled-coil in between the binding platforms, serving as a molecular ruler to specifically recognize a 22 bp separated bipartite binding motif. Binding of c-di-AMP to BusR induces a shift in equilibrium from an inactivated towards an activated state that allows BusR to bind the target DNA, leading to transcriptional repression.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , DNA Bacteriano/genética , Fosfatos de Dinucleosídeos/metabolismo , Regulação Bacteriana da Expressão Gênica/genética , Sistemas do Segundo Mensageiro/genética , Streptococcus agalactiae/genética , Transporte Biológico/genética , Cristalografia por Raios X , Proteínas de Ligação a DNA/genética , Potássio/metabolismo , Domínios Proteicos/genética , Fatores de Transcrição/genéticaRESUMO
All living cells use cyclic nucleotides as second messengers for signal sensing and transduction. Cyclic di-3',5'-adenosine monophosphate (c-di-AMP) is primarily involved in the control of bacterial and euryarcheal osmoadaptation and is produced by diadenylate cyclases from two molecules of ATP. Specific phosphodiesterases hydrolyze c-di-AMP to the linear phosphoadenylate adenosine 5'-pApA or to AMP. Different methods including high-performance liquid chromatography (HPLC), thin-layer chromatography (TLC) and ion exchange chromatography (IEX) can be used to determine activities of c-di-AMP-synthesizing and degrading enzymes. Here, we describe in detail the TLC and IEX methods adapted for characterization of the diadenylate cyclase DisA and the phosphodiesterase AtaC from Streptomyces venezuelae. TLC allows quick and easy separation of radioactive-labeled substrates and products, while IEX avoids utilization of potentially hazardous radioactive substrates and can be used as a good substitute if an HPLC system is not available. Unlike in TLC assays, samples cannot be analyzed in parallel by using the IEX assay, thus it is more time consuming.
RESUMO
Antibiotic-producing Streptomyces use the diadenylate cyclase DisA to synthesize the nucleotide second messenger c-di-AMP, but the mechanism for terminating c-di-AMP signaling and the proteins that bind the molecule to effect signal transduction are unknown. Here, we identify the AtaC protein as a c-di-AMP-specific phosphodiesterase that is also conserved in pathogens such as Streptococcus pneumoniae and Mycobacterium tuberculosis AtaC is monomeric in solution and binds Mn2+ to specifically hydrolyze c-di-AMP to AMP via the intermediate 5'-pApA. As an effector of c-di-AMP signaling, we characterize the RCK_C domain protein CpeA. c-di-AMP promotes interaction between CpeA and the predicted cation/proton antiporter, CpeB, linking c-di-AMP signaling to ion homeostasis in Actinobacteria. Hydrolysis of c-di-AMP is critical for normal growth and differentiation in Streptomyces, connecting ionic stress to development. Thus, we present the discovery of two components of c-di-AMP signaling in bacteria and show that precise control of this second messenger is essential for ion balance and coordinated development in Streptomyces.
Assuntos
Fosfatos de Dinucleosídeos/metabolismo , Diester Fosfórico Hidrolases/metabolismo , Streptomyces/metabolismo , Monofosfato de Adenosina/metabolismo , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica/genética , Hidrólise , Mycobacterium tuberculosis/metabolismo , Sistemas do Segundo Mensageiro , Transdução de Sinais/fisiologia , Streptococcus pneumoniae/metabolismoRESUMO
Ballast Water Managements Systems (BWMS) installed on vessels may use Active Substances (AS) to inactivate organisms. This paper provides new insights in the global issue of noxious Disinfection By-Products (DBP) produced with primarily oxidant-based BWMS, and the risk assessment for workers, including port State control officers, while performing tasks on a vessel that involve exposure to treated ballast water. The Joint Group of Experts on the Scientific Aspects of Marine Environmental Protection - Ballast Water Working Group (GESAMP-BWWG) plays a role in the certification process of BWMS that make use of AS evaluating potential negative effects. All BWMS that passed GESAMP-BWWG Final Approval until mid 2019 were analyzed providing an overview of chemicals in the treated ballast water before and after neutralization. The ballast tank cleaning scenario, the sampling scenario, and the ballast tank inspection scenario all showed elevated human health risks using the Derived Minimal Effect Levels approach. The most critical exposure occurs in the ballast tank cleaning scenario through the inhalation of volatile DBP, such as tribromomethane. This substance may cause acute effects such as headache, dizziness and also has carcinogenic properties. The two risk reducing options available in the GESAMP-BWWG Tier 2 calculations were compared, one being mitigation measures such as protective gloves and coveralls, the other option is taking into account a time correction factor. The results showed that the trihalomethanes in air are most problematic, however, there is a possibility that the calculated values may be overestimated as generally worst case assumptions were used.
Assuntos
Conservação dos Recursos Naturais/métodos , Desinfecção/métodos , Exposição Ocupacional/análise , Medição de Risco/métodos , Navios , Poluentes Químicos da Água/análise , Humanos , Trialometanos/análise , Trialometanos/toxicidade , Poluentes Químicos da Água/toxicidade , Purificação da Água/métodosRESUMO
Laparoscopic pancreatoduodenectomy (LPD) is increasingly performed worldwide as an alternative to the open conventional method, with the aim of advancing recovery. Because of its recent introduction, little is known about late surgical complications from this procedure. Here, we describe a case of a 76-year-old woman who underwent LPD with end-to-side gastrojejunostomy and presented with pain in the right upper quadrant of the abdomen 6 months thereafter. CT showed signs of an internal herniation. Elective diagnostic laparoscopy demonstrated a hernia of the efferent loop in Petersen's space. The loop was repositioned and Petersen's space was closed. The patient was discharged on the same day. Apart from a short readmission for nausea and vomiting, which were managed conservatively, the patient recovered without any further complications. Internal herniation should be considered in patients with abdominal complaints following LPD.
Assuntos
Hérnia Abdominal/diagnóstico , Hérnia Abdominal/etiologia , Pancreaticoduodenectomia/efeitos adversos , Idoso , Feminino , Humanos , Laparoscopia , Radiografia AbdominalRESUMO
Ballast water managements systems (BWMS) installed on vessels may use active substances to inactivate organisms. This paper provides new insights in the global issue of noxious disinfection by-products (DBP) discharge with ballast water, and the related risk assessment for human health. The GESAMP ballast water working group plays a role in the certification process of BWMS that make use of active substances evaluating potential negative effects. We analyzed all BWMS that passed GESAMP final approval over a decade until 2017 providing an overview of chemicals in the discharged ballast water generated by BWMS. We used these data to calculate the chemical load humans may be exposed to for two different commercial ports (Koper, Slovenia and Hamburg, Germany). None of the chemicals in this study reached levels of concern that would indicate a risk for humans after exposure to chemicals present in the discharged ballast water. Nevertheless, although this exposure only adds to a lesser degree to the overall exposure to disinfection by-products, some chemicals, such as tribromomethane, have carcinogenic properties. In case studies we show which chemicals have the largest contribution to the aggregated exposure of humans. We note that tribromomethane, despite its low bio-concentration factor (BCF), may accumulate in fat, when fish are continuously exposed to DBPs during low-level chlorination. Since this figure would give a higher value for the internal dose for tribromomethane from seafood consumption than the current BCF in the GISIS database, the calculated value may underestimate the contribution of tribromomethane, and possibly also other DBPs.
Assuntos
Desinfetantes/toxicidade , Exposição Ambiental/análise , Medição de Risco/métodos , Água do Mar/análise , Navios , Eliminação de Resíduos Líquidos/métodos , Poluentes Químicos da Água/toxicidade , Animais , Desinfetantes/análise , Desinfecção/métodos , Exposição Ambiental/efeitos adversos , Peixes , Alemanha , Halogenação , Humanos , Água do Mar/química , Eslovênia , Poluentes Químicos da Água/análiseRESUMO
OBJECTIVES: Presymptomatic detection of patients with rare diseases (RD), defined by a population frequency less than 1 : 2,000, is the task of newborn screening (NBS). In the Czech Republic (CZ), currently eighteen RD are screened: phenylketonuria/hyperphenylalaninemia (PKU/HPA), congenital hypothyroidism (CH), congenital adrenal hyperplasia (CAH), cystic fibrosis (CF), medium chain acyl-CoA dehydrogenase deficiency (MCADD), long chain 3-hydroxyacyl-CoA dehydrogenase deficiency (LCHADD), very long chain acyl-CoA dehydrogenase deficiency (VLCADD), carnitine palmitoyl transferase I and II deficiency (CPTID, CPTIID), carnitine-acylcarnitine translocase deficiency (CACTD), maple syrup urine disease (MSUD), glutaric aciduria type I (GA I), isovaleryl-CoA dehydrogenase deficiency (IVA), argininemia (ARG), citrullinemia (CIT), biotinidase deficiency (BTD), cystathionine beta-synthase-deficient homocystinuria (CBSD HCU), and methylenetetrahydrofolate reductase deficiency homocystinuria (MTHFRD HCU). The aim was to analyze the prevalence of RD screened by NBS in CZ. METHODS: We examined the NBS programme in CZ from 1 January 2010 to 31 December 2017, which covered 888,891 neonates. Dried blood spots were primarily analyzed using fluorescence immuno-assay, tandem mass spectrometry and fluorimetry. RESULTS: The overall prevalence of RD among the neonate cohort was 1 : 1,043. Individually, 1 : 2,877 for CH, 1 : 5,521 for PKU/HPA, 1 : 6,536 for CF (1 : 5,887 including false negative patients), 1 : 12,520 for CAH, 1 : 22,222 for MCADD, 1 : 80,808 for LCHADD, 1 : 177,778 for GA I, 1 : 177,778 for IVA, 1 : 222,223 for VLCADD, 1 : 296,297 for MSUD, 1 : 8,638 for BTD, and 1 : 181,396 for CBSD HCU. CONCLUSIONS: The observed prevalence of RD, based on NBS, corresponds to that expected, more precisely it was higher for BTD and lower for MSUD, IVA, CBSD HCU, MCADD and VLCADD. Early detection of rare diseases by means of NBS is an effective secondary prevention tool.
Assuntos
Triagem Neonatal/métodos , Doenças Raras/epidemiologia , Biomarcadores/sangue , República Tcheca/epidemiologia , Fluorometria , Humanos , Recém-Nascido , Doenças Raras/sangue , Espectrometria de Massas em TandemRESUMO
Methods for analysis of microplastic in soils are still being developed. In this study, we evaluated the potential of a soil universal model method (SUMM) based on thermogravimetry (TGA) for the identification and quantification of microplastics in standard loamy sand. Blank and spiked soils (with amounts of one of four microplastic types) were analyzed by TGA. For each sample, thermal mass losses (TML) in 10⯰C intervals were extracted and used for further analysis. To explain and demonstrate the principles of SUMM, two scenarios were discussed. The first refers to a rare situation in which an uncontaminated blank of investigated soil is available and TML of spiked and blank soils are subtracted. The results showed that the investigated microplastics degraded in characteristic temperature areas and differences between spiked and blank soils were proportional to the microplastics concentrations. The second scenario reflects the more common situation where the blank is not available and needs to be replaced by the previously developed interrelationships representing soil universal models. The models were consequently subtracted from measured TML. Sparse principal component analysis (sPCA) identified 8 of 14 modeled differences between measured TMLs and the universal model as meaningful for microplastics discrimination. Calibrating various microplastics concentrations with the first principal component extracted from sPCA resulted in linear fits and limits of detection in between environmentally relevant microplastics concentrations. Even if such an approach using calculated standards still has limitations, the SUMM shows a certain potential for a fast pre-screening method for analysis of microplastics in soils.
Assuntos
Modelos Químicos , Plásticos/química , Poluentes do Solo/análise , Solo/química , Plásticos/análise , Polietileno/análise , Polietilenotereftalatos/análise , Poliestirenos/análise , Cloreto de Polivinila/análise , Análise de Componente Principal , TemperaturaRESUMO
Neonates with low birthweight (LBW) represent a vulnerable population. This retrospective study analyzed the birth frequency of diseases detected by neonatal screening (NBS) in normal and LBW neonates in the Czech Republic. Between years 2002 and 2016, the number of screened disorders in the Czech Republic gradually increased from two to 13. Prevalence of screened diseases was calculated for cohorts ranging from 777,100 to 1,277,283 neonates stratified by birthweight. Odds ratio of the association of LBW with each disease was calculated and statistical significance was evaluated using the chi-square test or Fisher's exact test, as appropriate. Three diseases were associated with higher risk of prevalence in LBW neonates, namely congenital hypothyroidism (OR 2.50, CI 1.92; 3.25), cystic fibrosis (OR 2.44, CI 1.51; 3.94), and long chain 3-hydroxyacyl-CoA dehydrogenase deficiency (LCHADD) (OR 7.74, CI 2.18; 27.42).Conclusion: Although the underlying mechanisms are not well understood, results can be hypothesized that LBW (respectively prematurity) may lead to the secondary and often transitory hypothyroidism while cystic fibrosis and LCHADD may manifest already prenatally and result into preterm birth and LBW. What is Known: ⢠The percentage of low birthweight (LBW) neonates in the Czech Republic has been increasing. ⢠Previously published studies reported positive association between LBW and congenital hypothyroidism and cystic fibrosis. What is New: ⢠The association between LCHADD and LBW has not yet been described. ⢠LBW can be the first manifestation of cystic fibrosis and LCHADD.
Assuntos
Doenças do Recém-Nascido/epidemiologia , Triagem Neonatal/normas , Peso ao Nascer , República Tcheca/epidemiologia , Humanos , Recém-Nascido de Baixo Peso , Recém-Nascido , Prevalência , Estudos RetrospectivosRESUMO
The use of plastic materials in daily life, industry, and agriculture can cause soil pollution with plastic fragments down to the micrometer scale, i.e., microplastics. Quantitative assessment of microplastics in soil has been limited so far. Until now, microplastic analyses in soil require laborious sample cleanup and are mostly restricted to qualitative assessments. In this study, we applied thermogravimetry-mass spectrometry (TGA-MS) to develop a method for the direct quantitative analysis of poly(ethylene terephthalate) (PET) without further sample pretreatment. For this, soil samples containing 1.61 ± 0.15 wt % organic matter were spiked with 0.23-4.59 wt % PET bottle recyclate microplastics. dl-Cysteine was used as the internal standard (IS). Sample mixtures were pyrolyzed with a 5 K min-1 ramp (40-1000 °C), while sample mass loss and MS signal intensity of typical PET pyrolysis products were recorded. We found MS signal intensities linearly responding to microplastic concentrations. The most-promising results were obtained with the IS-corrected PET pyrolysis product vinylbenzene/benzoic acid ( m/ z = 105, adj. R2 = 0.987). The limits of detection and quantification were 0.07 and 1.72 wt % PET, respectively. Our results suggest that TGA-MS can be an easy and viable complement to existing methods such as pyrolysis or thermogravimetry-thermal desorption assays followed by gas chromatography/mass spectrometry detection or to spectral microscopy techniques.
RESUMO
The concentration of messenger molecules in bacterial cells needs to be tightly regulated. This can be achieved by either controlling the synthesis rate, degradation, or export by specific transporters, respectively. The regulation of the essential second messenger c-di-AMP is achieved by modulation of the diadenylate cyclase activity as well as by specific phosphodiesterases that hydrolyze c-di-AMP in the cell. We provide here structural and biochemical data on the DHH-type phosphodiesterase TmPDE (TM1595) from Thermotoga maritima. Our analysis shows that TmPDE is preferentially degrading linear dinucleotides, such as 5'-pApA, 5'-pGpG, and 5'-pApG, compared with cyclic dinucleotide substrates. The high-resolution structural data provided here describe all steps of the PDE reaction: the ligand-free enzyme, two substrate-bound states, and three post-reaction states. We can furthermore show that Pde2 from Streptococcus pneumoniae shares both structural features and substrate specificity based on small-angle X-ray scattering data and biochemical assays.