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1.
Med Hypotheses ; 147: 110485, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33450625

RESUMO

Corona Virus Disease 2019 (COVID-19) has emerged as a pandemic leading to unprecedented disruption of global health and economy. Transmembrane protease serine 2 (TMPRSS2) has been found to be critical in priming the viral spike protein and the host ACE2 receptor before the virus enters into the host cell. Recent studies have experimentally demonstrated that Alpha 1 antitrypsin (encoded by SERPINA1 gene) is an inhibitor of TMPRSS2 and provided support to the already approved therapy as a candidate for COVID-19. Interestingly Alpha 1 antitrypsin deficiency is common among Europeans. Here we have provided in silico evidence that Alpha 1 antitrypsin can interact with TMPRSS2 and both of them are co-expressed in the human liver and lung. We then analyzed the gnomAD dataset to show that Europeans and Latinos have a substantially higher carrier frequency of Alpha 1 Antitrypsin Deficiency (~12%) compared to other large ethnicities. Therefore, we hypothesize that Alpha 1 antitrypsin deficiency might be a risk factor for severe infection with SARS-CoV-2. We propose Alpha 1 antitrypsin status as a potential prognostic predictor of COVID-19 outcome.


Assuntos
/genética , Genoma Humano , Deficiência de alfa 1-Antitripsina/genética , Comorbidade , Grupo com Ancestrais do Continente Europeu , Feminino , Hispano-Americanos , Humanos , Inflamação , Fígado/metabolismo , Pulmão/metabolismo , Masculino , Modelos Teóricos , Prognóstico , Fatores de Risco , Serina Endopeptidases/genética , alfa 1-Antitripsina/genética
2.
Nat Commun ; 11(1): 6371, 2020 12 11.
Artigo em Inglês | MEDLINE | ID: mdl-33311470

RESUMO

Genetic mutations predispose the serine protease inhibitor α1-antitrypsin to misfolding and polymerisation within hepatocytes, causing liver disease and chronic obstructive pulmonary disease. This misfolding occurs via a transiently populated intermediate state, but our structural understanding of this process is limited by the instability of recombinant α1-antitrypsin variants in solution. Here we apply NMR spectroscopy to patient-derived samples of α1-antitrypsin at natural isotopic abundance to investigate the consequences of disease-causing mutations, and observe widespread chemical shift perturbations for methyl groups in Z AAT (E342K). By comparison with perturbations induced by binding of a small-molecule inhibitor of misfolding we conclude that they arise from rapid exchange between the native conformation and a well-populated intermediate state. The observation that this intermediate is stabilised by inhibitor binding suggests a paradoxical approach to the targeted treatment of protein misfolding disorders, wherein the stabilisation of disease-associated states provides selectivity while inhibiting further transitions along misfolding pathways.


Assuntos
Espectroscopia de Ressonância Magnética/métodos , Deficiência de alfa 1-Antitripsina/genética , Deficiência de alfa 1-Antitripsina/metabolismo , alfa 1-Antitripsina/química , alfa 1-Antitripsina/genética , alfa 1-Antitripsina/metabolismo , Predisposição Genética para Doença/genética , Glicoproteínas , Humanos , Modelos Moleculares , Medicina Molecular , Mutação , Agregação Patológica de Proteínas , Conformação Proteica , Proteínas Recombinantes , Inibidores de Serino Proteinase/química
3.
Clin Nephrol ; 94(6): 297-306, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32909541

RESUMO

We report a unique case of an immunosuppressed 67-year-old female with homozygous Z-allele mutation A1AT deficiency and liver transplant with baseline chronic kidney disease (CKD) stage IIIa with creatinine of 1 mg/dL and glomerular filtration rate (GFR) of 49 mL/min/1.73m2 ~ 6 months before the presentation. She presented with COVID-19 mediated hypoxic respiratory failure complicated by AKI requiring provisional renal replacement therapy with recovery of kidney function with a new baseline of creatinine of 1.6 - 1.8 mg/dL with GFR of 31 mL/min/1.73m2.


Assuntos
Lesão Renal Aguda , Infecções por Coronavirus , Transplante de Fígado , Pandemias , Pneumonia Viral , Insuficiência Renal Crônica/complicações , Deficiência de alfa 1-Antitripsina/complicações , Idoso , Betacoronavirus , Feminino , Taxa de Filtração Glomerular , Humanos , Hospedeiro Imunocomprometido , Diálise Renal , alfa 1-Antitripsina/genética
4.
FASEB J ; 34(11): 14160-14165, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32960480

RESUMO

Infection rates, severity, and fatalities due to COVID-19, the pandemic mediated by SARS-CoV-2, vary greatly between countries. With few exceptions, these are lower in East and Southeast Asian and Sub-Saharan African countries compared with other regions. Epidemiological differences may reflect differences in border closures, lockdowns, and social distancing measures taken by each county, and by cultural differences, such as common use of face masks in East and Southeast Asian countries. The plasma serine protease inhibitor alpha-1 antitrypsin was suggested to protect from COVID-19 by inhibiting TMPRSS2, a cell surface serine protease essential for the SARS-CoV-2 cell entry. Here, we present evidence that population differences in alpha-1 antitrypsin deficiency allele frequencies may partially explain national differences in the COVID-19 epidemiology. Our study compared reported national estimates for the major alpha-1 antitrypsin deficiency alleles PiZ and PiS (SERPINA1 rs28929474 and rs17580, respectively) with the Johns Hopkins University Coronavirus Resource Center dataset. We found a significant positive correlation (R = .54, P = 1.98e-6) between the combined frequencies of the alpha-1 antitrypsin PiZ and PiS deficiency alleles in 67 countries and their reported COVID-19 mortality rates. Our observations suggest that alpha-1 antitrypsin deficiency alleles may contribute to national differences in COVID-19 infection, severity, and mortality rates. Population-wide screening for carriers of alpha-1 antitrypsin deficiency alleles should be considered for prioritizing individuals for stricter social distancing measures and for receiving a SARS-CoV-2 vaccine once it becomes available.


Assuntos
Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/genética , Pneumonia Viral/epidemiologia , Pneumonia Viral/genética , alfa 1-Antitripsina/genética , Infecções por Coronavirus/sangue , Infecções por Coronavirus/metabolismo , Frequência do Gene , Humanos , Inflamação/metabolismo , Pandemias , Pneumonia Viral/sangue , Pneumonia Viral/metabolismo , Polimorfismo de Nucleotídeo Único , Serina Endopeptidases/metabolismo , alfa 1-Antitripsina/sangue , alfa 1-Antitripsina/metabolismo , Deficiência de alfa 1-Antitripsina/metabolismo
6.
Gene ; 762: 144974, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-32707305

RESUMO

BACKGROUND: There exists considerable evidence conforming that autophagy may play an important role in the biological process of breast cancer. This study aimed to construct and evaluate a novel autophagy-related gene signature as a potential prognostic factor and therapeutic target in breast cancer patients based on high-throughput sequencing datasets. MATERIALS & METHODS: Autophagy-related genes obtained from the Human Autophagy Database and high-sequencing data obtained from The Cancer Genome Atlas (TCGA) were analyzed to identify differential expressed genes (DEGs) between tumor and normal tissues. Then GO and KEGG analysis were performed to explore potential biological and pathological functions of DEGs. Autophagy-related prognostic genes were identified by univariate COX regression analysis. Subsequently stepwise model selection using the Alkaike information criterion (AIC) and multivariate COX regression model was performed to construct autophagy-related gene signature. Then patients were divided into high- and low-risk groups based on the risk score identified by the autophagy-related gene signature. Multivariate COX regression model and stratification analysis were used to specify the prognostic value of this gene signature in whole cohort and various subgroups. T-test and ANOVA analysis were used to compare the expression differences of continuous variables (5 prognostic genes and risk score) in binary and multiple category groups respectively. Kaplan-Meier analysis, log-rank tests and the area under receiver operating characteristic (ROC) curve (AUC) were conducted to validate the accuracy and precise of the autophagy-related gene signature based on GSE20685 and GSE21653 datasets. RESULTS: We profiled autophagy-related DEGs in normal and breast tumor tissues. GO and KEGG analysis indicated that autophagy-related DEGs might participate in breast cancer occurrence, development and drug resistance. Then we identified five autophagy-related genes (EIF4EBP1, ATG4A, BAG1, MAP1LC3A and SERPINA1) that had significantly prognostic values for breast cancer. Autophagy-related gene signature was constructed and patients were divided into high- and low- risk groups based on their risk score. Patients in the high-risk group tended to have shorter overall survival (OS) and relapse-free survival (RFS) times than those in the low-risk group (OS: HR = 1.620, 95%CIs: 1.345-1.950; P < 0.001; RFS: HR = 1.487, 95%CIs: 1.248-1.771, P < 0.001). Autophagy-related gene signature had significant prognostic value in stratified subgroups especially in advanced breast cancer subgroups (T3-4; N2-3; stage III-IV). Its prognostic value was further confirmed in two GEO validation datasets (GSE20685: P = 6.795e-03; GSE21653: P = 1.383e-03). Finally, association analysis between clinicopathological factors and gene signature showed the risk score was higher in patients with ER/PR negative, higher clinical stage or T stage (P < 0.01). CONCLUSION: We established and confirmed a novel autophagy-related gene signature for patients with breast cancer that had independent survival prognostic value especially in advanced breast cancer subgroups. Our research might promote the molecular mechanism study of autophagy-related genes in breast cancer.


Assuntos
Autofagia , Biomarcadores Tumorais/genética , Neoplasias da Mama/genética , Transcriptoma , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Relacionadas à Autofagia/genética , Proteínas Relacionadas à Autofagia/metabolismo , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Cisteína Endopeptidases/genética , Cisteína Endopeptidases/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Humanos , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , alfa 1-Antitripsina/genética , alfa 1-Antitripsina/metabolismo
8.
Am J Respir Cell Mol Biol ; 63(4): 444-451, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32515985

RESUMO

Alpha-1 antitrypsin deficiency (AATD) is an inherited condition characterized by reduced levels of serum AAT due to mutations in the SERPINA1 (Serpin family A member 1) gene. The Pi*S (Glu264Val) is one of the most frequent deficient alleles of AATD, showing high incidence in the Iberian Peninsula. Herein, we describe two new alleles carrying an S mutation but producing a null phenotype: QOVigo and QOAachen. The new alleles were identified by sequencing the SERPINA1 gene in three patients who had lower AAT serum levels than expected for the initial genotype. These alleles are the result of combined mutations in cis in a PI*S allele. Sequencing detected the S mutation in cis with Tyr138Cys (S+Tyr138Cys) in two patients, whereas a third one had the S mutation in cis with Pro391Thr variant (S+Pro391Thr). When expressed in a cellular model, these variants caused strong AAT polymerization and very low AAT secretion to almost undetectable levels. The isoelectric focusing method for plasma AAT phenotyping did not show AAT protein encoded by the novel mutant alleles, behaving as null. We called these alleles PI*S-plus because the S variant was phased with another variant conferring more aggressive characteristics to the allele. The current data demonstrate that the clinical variability observed in AATD can be explained by additional genetic variation, such as dual cis-acting variants in the SERPINA1 gene. The possible existence of other unrevealed variants combined in the PI*S alleles should be considered to improve the genetic diagnosis of the patients.


Assuntos
Mutação/genética , Deficiência de alfa 1-Antitripsina/genética , alfa 1-Antitripsina/genética , Adulto , Alelos , Análise Mutacional de DNA/métodos , Feminino , Frequência do Gene/genética , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo
9.
PLoS One ; 15(5): e0233640, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32453766

RESUMO

Understanding the coagulation process is critical to developing treatments for trauma and coagulopathies. Clinical studies on tranexamic acid (TXA) have resulted in mixed reports on its efficacy in improving outcomes in trauma patients. The largest study, CRASH-2, reported that TXA improved outcomes in patients who received treatment prior to 3 hours after the injury, but worsened outcomes in patients who received treatment after 3 hours. No consensus has been reached about the mechanism behind the duality of these results. In this paper we use a computational model for coagulation and fibrinolysis to propose that deficiencies or depletions of key anti-fibrinolytic proteins, specifically antiplasmin, a1-antitrypsin and a2-macroglobulin, can lead to worsened outcomes through urokinase-mediated hyperfibrinolysis.


Assuntos
Transtornos da Coagulação Sanguínea/tratamento farmacológico , Ácido Tranexâmico/uso terapêutico , Ativador de Plasminogênio Tipo Uroquinase/genética , Ferimentos e Lesões/tratamento farmacológico , Antifibrinolíticos/uso terapêutico , Coagulação Sanguínea/genética , Transtornos da Coagulação Sanguínea/sangue , Transtornos da Coagulação Sanguínea/genética , Transtornos da Coagulação Sanguínea/patologia , Simulação por Computador , Fibrina/genética , Tempo de Lise do Coágulo de Fibrina , Fibrinolisina/genética , Fibrinólise/efeitos dos fármacos , Hemorragia/sangue , Hemorragia/tratamento farmacológico , Hemorragia/genética , Humanos , Proteínas de Membrana/genética , Mortalidade , alfa 2-Macroglobulinas Associadas à Gravidez/genética , Trombina/genética , Trombina/metabolismo , Ferimentos e Lesões/sangue , Ferimentos e Lesões/genética , Ferimentos e Lesões/patologia , alfa 1-Antitripsina/genética
10.
Int J Mol Sci ; 21(3)2020 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-32012925

RESUMO

Cystic fibrosis (CF) is an autosomal recessive genetic disorder arising from mutations to the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Disruption to normal ion homeostasis in the airway results in impaired mucociliary clearance, leaving the lung more vulnerable to recurrent and chronic bacterial infections. The CF lung endures an excess of neutrophilic inflammation, and whilst neutrophil serine proteases are a crucial part of the innate host defence to infection, a surplus of neutrophil elastase (NE) is understood to create a net destructive effect. Alpha-1 antitrypsin (A1AT) is a key antiprotease in the control of NE protease activity but is ineffective in the CF lung due to the huge imbalance of NE levels. Therapeutic strategies to boost levels of protective antiproteases such as A1AT in the lung remain an attractive research strategy to limit the damage from excess protease activity. microRNAs are small non-coding RNA molecules that bind specific cognate sequences to inhibit expression of target mRNAs. The inhibition of miRNAs which target the SERPINA1 (A1AT-encoding gene) mRNA represents a novel therapeutic approach for CF inflammation. This could involve the delivery of antagomirs that bind and sequester the target miRNA, or target site blockers that bind miRNA recognition elements within the target mRNA to prevent miRNA interaction. Therefore, miRNA targeted therapies offer an alternative strategy to drive endogenous A1AT production and thus supplement the antiprotease shield of the CF lung.


Assuntos
Fibrose Cística/genética , MicroRNAs/genética , alfa 1-Antitripsina/genética , Antagomirs/farmacologia , Antagomirs/uso terapêutico , Fibrose Cística/metabolismo , Fibrose Cística/terapia , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Humanos , Elastase de Leucócito/metabolismo , MicroRNAs/antagonistas & inibidores , Terapia de Alvo Molecular , Regulação para Cima , alfa 1-Antitripsina/metabolismo
11.
PLoS One ; 15(1): e0228331, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31990955

RESUMO

Gastric cancer (GC) is a significant cancer-related cause of death worldwide. The most used chemotherapeutic regimen in GC is based on platinum drugs such as cisplatin (CDDP). However, CDDP resistance reduces advanced GC survival. In vitro drug-resistant cell model would help in the understanding of molecular mechanisms underlying this drug-resistance phenomenon. The aim of this study was to characterize new models of CDDP-resistant GC cell lines (AGS R-CDDP and MKN-28 R-CDDP) obtained through a stepwise increasing drug doses method, in order to understand the molecular mechanisms underlying chemoresistance as well as identify new therapeutic targets for the treatment of GC. Cell viability assays, cell death assays and the expression of resistance molecular markers confirmed that AGS R-CDDP and MKN-28 R-CDDP are reliable CDDP-resistant models. RNA-seq and bioinformatics analyses identified a total of 189 DEGs, including 178 up-regulated genes and 11 down-regulated genes, associated mainly to molecular functions involved in CDDP-resistance. DEGs were enriched in 23 metabolic pathways, among which the most enriched was the inflammation mediated by chemokine and cytokine signaling pathway. Finally, the higher mRNA expression of SERPINA1, BTC and CCL5, three up-regulated DEGs associated to CDDP resistance found by RNA-seq analysis was confirmed. In summary, this study showed that AGS R-CDDP and MKN-28 R-CDDP are reliable models of CDDP resistance because resemble many of resistant phenotype in GC, being also useful to assess potential therapeutic targets for the treatment of gastric cancers resistant to CDDP. In addition, we identified several DEGs associated with molecular functions such as binding, catalytic activity, transcription regulator activity and transporter activity, as well as signaling pathways associated with inflammation process, which could be involved in the development of CDDP resistance in GC. Further studies are necessary to clarify the role of inflammatory processes in GC resistant to CDDP and these models could be useful for these purposes.


Assuntos
Resistencia a Medicamentos Antineoplásicos , Perfilação da Expressão Gênica/métodos , Redes Reguladoras de Genes , Neoplasias Gástricas/genética , Idoso , Betacelulina/genética , Linhagem Celular Tumoral , Quimiocina CCL5/genética , Cisplatino , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Pessoa de Meia-Idade , Modelos Biológicos , Análise de Sequência de RNA , Neoplasias Gástricas/tratamento farmacológico , alfa 1-Antitripsina/genética
12.
Am J Respir Crit Care Med ; 201(5): 505-506, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31810377
13.
J Biotechnol ; 307: 87-97, 2020 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-31697975

RESUMO

Alpha-1-antitrypsin (A1AT) is an abundant serum inhibitor of serine proteases. A1AT deficiency is a common genetic disorder which is currently treated with augmentation therapies. These treatments involve weekly injections of patients with purified plasma-derived A1AT. Such therapies can be extremely expensive and rely on plasma donors. Hence, large-scale production of recombinant A1AT (rA1AT) could greatly benefit these patients, as it could decrease the cost of treatments, reduce biosafety concerns and ensure quantitative and qualitative controls of the protein. In this report, we sought to produce α2,6-sialylated rA1AT with our cumate-inducible stable CHO pool expression system. Our different CHO pools could reach volumetric productivities of 1,2 g/L. The human α2,6-sialyltransferase was stably expressed in these cells in order to mimic elevated α2,6-sialylation levels of native A1AT protein. Sialylation of the recombinant protein was stable over the duration of the fed-batch production phase and was higher in a pool where cells were sorted and enriched by FACS based on cell-surface α2,6-sialylation. Addition of ManNAc to the cell culture media during production enhanced both α2,3 and α2,6 A1AT sialylation levels whereas addition of 2F-peracetylfucose potently inhibited fucosylation of the protein. Finally, we demonstrated that rA1AT proteins exhibited human neutrophil elastase inhibitory activities similar to the commercial human plasma-derived A1AT.


Assuntos
Elastase de Leucócito/antagonistas & inibidores , Sialiltransferases/metabolismo , alfa 1-Antitripsina/metabolismo , Animais , Medicamentos Biossimilares/metabolismo , Células CHO , Cricetulus , Humanos , Proteínas Recombinantes , Sialiltransferases/genética , alfa 1-Antitripsina/genética
14.
Hepatol Int ; 14(1): 127-137, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31832977

RESUMO

BACKGROUND AND AIMS: Alpha-1 antitrypsin (AAT) is a product of SERPINA1 gene mainly expressed by hepatocytes. Clinically relevant mutations in the SERPINA1 gene, such as Z (Glu342Lys), results in an expression of misfolded AAT protein having high propensity to polymerize, accumulate in hepatocytes and thus to enhance a risk for hepatocyte damage and subsequent liver disease. So far, the relationship between the Z-AAT accumulation and liver cell damage remains not completely understood. We present three-dimensional organoid culture systems, as a novel tool for modeling Z-AAT-related liver diseases. METHODS: We have established liver organoids from liver biopsies of patients with homozygous (ZZ) and heterozygous (MZ) deficiency and normal (MM) genotypes of AAT. The features of these organoid models were characterized by analyzing AAT protein secretion and intracellular aggregation in MZ and ZZ genotypes as well as SERPINA1 expression in differentiated cultures. RESULTS: Transcriptional analysis of differentiated organoid cultures by RNA-Seq showed hepatocyte-specific gene expression profile. Genes, such as ALB, APOB, CYP3A4 and SERPINA1, were validated and confirmed through quantitative-PCR analysis. The organoids from MZ and ZZ cases showed intracellular aggregation and lower secretion of AAT protein, and lower expression of ALB and APOB, as typically seen in hepatocytes from Z-AAT deficiency patients. Furthermore, organoids responded to external stimulus. Treatment with oncostatin M, a well-known inducer of SERPINA1, increased expression of the full-length transcripts (AAT-1C) as well as the short transcript of AAT (AAT-ST1C4). CONCLUSIONS: Liver organoid model recapitulates the key features of Z-AAT deficiency and provides a useful tool for disease modeling.


Assuntos
Cirrose Hepática , Modelos Teóricos , Organoides , Deficiência de alfa 1-Antitripsina , alfa 1-Antitripsina/genética , Humanos
15.
Am J Respir Crit Care Med ; 201(5): 540-554, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31661293

RESUMO

Rationale: The role of PI (protease inhibitor) type Z heterozygotes and additional rare variant genotypes in the gene encoding alpha-1 antitrypsin, SERPINA1 (serpin peptidase inhibitor, clade A, member 1), in determining chronic obstructive pulmonary disease risk and severity is controversial.Objectives: To comprehensively evaluate the effects of rare SERPINA1 variants on lung function and emphysema phenotypes in subjects with significant tobacco smoke exposure using deep gene resequencing and alpha-1 antitrypsin concentrations.Methods: DNA samples from 1,693 non-Hispanic white individuals, 385 African Americans, and 90 Hispanics with ≥20 pack-years smoking were resequenced for the identification of rare variants (allele frequency < 0.05) in 16.9 kB of SERPINA1.Measurements and Main Results: White PI Z heterozygotes confirmed by sequencing (MZ; n = 74) had lower post-bronchodilator FEV1 (P = 0.007), FEV1/FVC (P = 0.003), and greater computed tomography-based emphysema (P = 0.02) compared with 1,411 white individuals without PI Z, S, or additional rare variants denoted as VR. PI Z-containing compound heterozygotes (ZS/ZVR; n = 7) had lower FEV1/FVC (P = 0.02) and forced expiratory flow, midexpiratory phase (P = 0.009). Nineteen white heterozygotes for five non-S/Z coding variants associated with lower alpha-1 antitrypsin had greater computed tomography-based emphysema compared with those without rare variants. In African Americans, a 5' untranslated region insertion (rs568223361) was associated with lower alpha-1 antitrypsin and functional small airway disease (P = 0.007).Conclusions: In this integrative deep sequencing study of SERPINA1 with alpha-1 antitrypsin concentrations in a heavy smoker and chronic obstructive pulmonary disease cohort, we confirmed the effects of PI Z heterozygote and compound heterozygote genotypes. We demonstrate the cumulative effects of multiple SERPINA1 variants on alpha-1 antitrypsin deficiency, lung function, and emphysema, thus significantly increasing the frequency of SERPINA1 variation associated with respiratory disease in at-risk smokers.


Assuntos
Pulmão/fisiopatologia , Doença Pulmonar Obstrutiva Crônica/genética , Enfisema Pulmonar/genética , Fumar/epidemiologia , alfa 1-Antitripsina/genética , Adulto , Afro-Americanos , Idoso , Idoso de 80 Anos ou mais , Grupo com Ancestrais do Continente Europeu , Feminino , Volume Expiratório Forçado , Genótipo , Heterozigoto , Hispano-Americanos , Humanos , Focalização Isoelétrica , Masculino , Fluxo Máximo Médio Expiratório , Pessoa de Meia-Idade , Fenótipo , Polimorfismo Genético , Doença Pulmonar Obstrutiva Crônica/epidemiologia , Doença Pulmonar Obstrutiva Crônica/fisiopatologia , Enfisema Pulmonar/epidemiologia , Enfisema Pulmonar/metabolismo , Enfisema Pulmonar/fisiopatologia , Tomografia Computadorizada por Raios X , Capacidade Vital , alfa 1-Antitripsina/metabolismo
16.
Int J Chron Obstruct Pulmon Dis ; 14: 2535-2542, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31819391

RESUMO

Purpose: Alpha-1-antitrypsin deficiency (AATD) is a rare hereditary condition resulting from the mutations in the SERPINA1 (serine protease inhibitor) gene and is characterized by low circulating levels of the alpha-1 antitrypsin (AAT) protein. The traditional algorithm for laboratory testing of AATD involves the analysis of AAT concentrations (nephelometry), phenotyping (isoelectric focusing, IEF), and genotyping (polymerase chain reaction, PCR); in selected cases, full sequencing of the SERPINA1 gene can be undertaken. New technologies arise that may make diagnosis easier and faster. Methods: We developed and evaluated a new diagnostic algorithm based on Luminex xMAP (multi-analyte profiling) technology using Progenika A1AT Genotyping Test. In an initial learning phase, 1979 samples from individuals suspected of having AATD were examined by both, a traditional and a "new" algorithm. In a second phase, 1133 samples were analyzed with the Luminex xMAP only. Results: By introducing a Luminex xMAP based algorithm, we were able to simultaneously identify 14 mutations in SERPINA1 gene (instead of two- S and Z-by using our old algorithm). Although the quantity of IEF assays remained unchanged, the nephelometric measurements and sequencing were reduced by 79% and 63.4%, respectively. Conclusion: The new method is convenient, fast and user-friendly. The application of the Luminex xMAP technology can simplify and shorten the diagnostic workup of patients with suspected AATD.


Assuntos
Análise Mutacional de DNA/métodos , Mutação , Deficiência de alfa 1-Antitripsina/diagnóstico , alfa 1-Antitripsina/genética , Predisposição Genética para Doença , Humanos , Medições Luminescentes , Reação em Cadeia da Polimerase , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Estudos Retrospectivos , Fatores de Tempo , Fluxo de Trabalho , Deficiência de alfa 1-Antitripsina/enzimologia , Deficiência de alfa 1-Antitripsina/genética
17.
Cells ; 8(11)2019 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-31717526

RESUMO

The presence of the homozygous 'Pi*Z' variant of alpha-1 antitrypsin (AAT) ('Pi*ZZ' genotype) predisposes to liver fibrosis development, but the role of iron metabolism in this process remains unknown. Therefore, we assessed iron metabolism and variants in the Homeostatic Iron Regulator gene (HFE) as the major cause of hereditary iron overload in a large cohort of Pi*ZZ subjects without liver comorbidities. The human cohort comprised of 409 Pi*ZZ individuals and 254 subjects without evidence of an AAT mutation who were recruited from ten European countries. All underwent a comprehensive work-up and transient elastography to determine liver stiffness measurements (LSM). The corresponding mouse models (Pi*Z overexpressors, HFE knockouts, and double transgenic [DTg] mice) were used to evaluate the impact of mild iron overload on Pi*Z-induced liver injury. Compared to Pi*Z non-carriers, Pi*ZZ individuals had elevated serum iron, transferrin saturation, and ferritin levels, but relevant iron overload was rare. All these parameters were higher in individuals with signs of significant liver fibrosis (LSM ≥ 7.1 kPa) compared to those without signs of significant liver fibrosis. HFE knockout and DTg mice displayed similar extent of iron overload and of fibrosis. Loss of HFE did not alter the extent of AAT accumulation. In Pi*ZZ individuals, presence of HFE mutations was not associated with more severe liver fibrosis. Taken together, Pi*ZZ individuals display minor alterations in serum iron parameters. Neither mild iron overload seen in these individuals nor the presence of HFE mutations (C282Y and H63D) constitute a major contributor to liver fibrosis development.


Assuntos
Proteína da Hemocromatose/deficiência , Homozigoto , Sobrecarga de Ferro/genética , Cirrose Hepática/etiologia , Mutação , alfa 1-Antitripsina/genética , Animais , Modelos Animais de Doenças , Suscetibilidade a Doenças , Sobrecarga de Ferro/complicações , Sobrecarga de Ferro/metabolismo , Sobrecarga de Ferro/patologia , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Camundongos , Camundongos Knockout , Camundongos Transgênicos
18.
Sci Rep ; 9(1): 17293, 2019 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-31754242

RESUMO

A possible role for calcium signalling in the autosomal dominant form of dementia, familial encephalopathy with neuroserpin inclusion bodies (FENIB), has been proposed, which may point towards a mechanism by which cells could sense and respond to the accumulation of mutant serpin polymers in the endoplasmic reticulum (ER). We therefore explored possible defects in Ca2+-signalling, which may contribute to the pathology associated with another serpinopathy, α1-antitrypsin (AAT) deficiency. Using CHO K1 cell lines stably expressing a wild type human AAT (MAAT) and a disease-causing polymer-forming variant (ZAAT) and the truncated variant (NHK AAT), we measured basal intracellular free Ca2+, its responses to thapsigargin (TG), an ER Ca2+-ATPase blocker, and store-operated Ca2+-entry (SOCE). Our fura2 based Ca2+ measurements detected no differences between these 3 parameters in cell lines expressing MAAT and cell lines expressing ZAAT and NHK AAT mutants. Thus, in our cell-based models of α1-antitrypsin (AAT) deficiency, unlike the case for FENIB, we were unable to detect defects in calcium signalling.


Assuntos
Sinalização do Cálcio/genética , Cálcio/metabolismo , Epilepsias Mioclônicas/genética , Transtornos Heredodegenerativos do Sistema Nervoso/genética , alfa 1-Antitripsina/metabolismo , Animais , Células CHO , Cricetulus , Epilepsias Mioclônicas/patologia , Transtornos Heredodegenerativos do Sistema Nervoso/patologia , Humanos , Microscopia Confocal , Mutação , Imagem Óptica , alfa 1-Antitripsina/genética
19.
J Gastrointestin Liver Dis ; 28(3): 297-302, 2019 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-31517326

RESUMO

BACKGROUND AND AIMS: Two single nucleotide polymorphisms (SNPs) in SERPINA1 (Pi*Z rs28929474 and Pi*S rs17580) are risk factors for developing liver cirrhosis. A recent study identified a common SNP in HSD17B13 (rs72613567) that conferred protection from chronic liver disease. The aim of the present study was to test these associations in a cohort of Lithuanian patients with liver fibrosis or cirrhosis. METHODS: The study included 302 patients with cirrhosis, 127 patients with liver fibrosis (METAVIR stages I-III) and 548 controls, all from Lithuania. SNPs were genotyped by quantitative PCR, using TaqMan allelic discrimination assays. Adjusted p value of ≤ 0.016 was considered significant. RESULTS: Genotype distributions of SERPINA1 and HSD17B13 SNPs were in Hardy-Weinberg equilibrium. SERPINA1 Pi*Z was not associated with liver fibrosis or cirrhosis. HSD17B13 rs10433937 (in high linkage disequilibrium with rs72613567; r 2 =0.96) also showed no overall association with liver disease, but the GG- genotype was associated with reduced risk of liver fibrosis (aOR 0.37, p=0.03). SERPINA1 Pi*S was associated with higher risk of developing hepatic fibrosis (aOR 3.42, p=0.001) and cirrhosis (aOR 2.59, p=0.02). CONCLUSIONS: We found that SERPINA1 Pi*S variant conferred an increased risk of developing liver fibrosis, while SERPINA1 Pi*Z and HSD17B13 rs10433937 were not associated with liver fibrosis or cirrhosis of different aetiology.


Assuntos
17-Hidroxiesteroide Desidrogenases/genética , Cirrose Hepática/genética , Polimorfismo de Nucleotídeo Único , alfa 1-Antitripsina/genética , Adulto , Idoso , Alelos , Estudos de Casos e Controles , Estudos de Coortes , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Cirrose Hepática/etiologia , Masculino , Pessoa de Meia-Idade , Fatores de Risco
20.
J Card Surg ; 34(11): 1215-1219, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31523846

RESUMO

AIM: Atrial septal aneurysm (ASA) is one of the congenital heart defects. The underlying pathophysiology of ASA has not been fully understood yet. Alpha-1 antitrypsin (A1AT) is a serine protease inhibitor glycoprotein, which is held responsible from tissue wall proteolysis if it is deficient in the body. The aim of this study was to investigate A1AT serum levels and the rs1303 (Pi*M3) variant in A1AT gene in patients with ASA. MATERIAL AND METHODS: Thirty patients (7 male and 23 female) with isolated ASA and 33 patients (11 male and 22 female) with normal atrial septum on echocardiography were included in this study. A1AT serum levels of study patients were measured quantitatively by the enzyme-linked immune sorbent assay (ELISA) method. The A1AT gene mutation rs1303 was analyzed by genotyping, which is performed on genomic DNA extracted from circulating mononuclear blood cells. Single-nucleotide polymorphism was evaluated on polymerase chain reaction using commercial kits. RESULTS: A1AT serum levels were not statistically different among patients with and without ASA (9.52 ± 4.33 µg/mL vs 9.83 ± 5.27 µg/mL, respectively, P = .80). A1AT homozygote mutation (PiM3M3) was significantly higher in the ASA group than the control group (21 vs 11, OR (95% CI): 6.68 [2.09-21.40], P = .001). A1AT serum levels were similar among patients with normal A1AT allele (PiMM), homozygote variant (PiM3M3), and heterozygote variant (PiMM3) (P = .79). CONCLUSION: This preliminary study revealed that homozygote A1AT rs1303 (PiM3M3) variant is significantly higher in patients with isolated ASA and may be associated with ASA development. Large scale comprehensive studies are needed to validate these results.


Assuntos
Aneurisma Cardíaco/genética , Septos Cardíacos , alfa 1-Antitripsina/genética , Estudos de Associação Genética , Átrios do Coração , Humanos
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