RESUMEN
Recent studies have revealed an unexplored population of long cell-free DNA (cfDNA) molecules in human plasma using long-read sequencing technologies. However, the biological properties of long cfDNA molecules (>500 bp) remain largely unknown. To this end, we have investigated the origins of long cfDNA molecules from different genomic elements. Analysis of plasma cfDNA using long-read sequencing reveals an uneven distribution of long molecules from across the genome. Long cfDNA molecules show overrepresentation in euchromatic regions of the genome, in sharp contrast to short DNA molecules. We observe a stronger relationship between the abundance of long molecules and mRNA gene expression levels, compared with short molecules (Pearson's r = 0.71 vs. -0.14). Moreover, long and short molecules show distinct fragmentation patterns surrounding CpG sites. Leveraging the cleavage preferences surrounding CpG sites, the combined cleavage ratios of long and short molecules can differentiate patients with hepatocellular carcinoma (HCC) from non-HCC subjects (AUC = 0.87). We also investigated knockout mice in which selected nuclease genes had been inactivated in comparison with wild-type mice. The proportion of long molecules originating from transcription start sites are lower in Dffb-deficient mice but higher in Dnase1l3-deficient mice compared with that of wild-type mice. This work thus provides new insights into the biological properties and potential clinical applications of long cfDNA molecules.
Asunto(s)
Carcinoma Hepatocelular , Ácidos Nucleicos Libres de Células , Neoplasias Hepáticas , Humanos , Animales , Ratones , Ácidos Nucleicos Libres de Células/genética , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , ADN/genética , Genómica , Ratones Noqueados , Endodesoxirribonucleasas/genéticaRESUMEN
The nascent polypeptide-associated complex (NAC) is a conserved ribosome-associated protein biogenesis factor. Whether NAC exerts chaperone activity and whether this function is restricted to de novo protein synthesis is unknown. Here, we demonstrate that NAC directly exerts chaperone activity toward structurally diverse model substrates including polyglutamine (PolyQ) proteins, firefly luciferase, and Aß40. Strikingly, we identified the positively charged ribosome-binding domain in the N terminus of the ßNAC subunit (N-ßNAC) as a major chaperone entity of NAC. N-ßNAC by itself suppressed aggregation of PolyQ-expanded proteins in vitro, and the positive charge of this domain was critical for this activity. Moreover, we found that NAC also exerts a ribosome-independent chaperone function in vivo. Consistently, we found that a substantial fraction of NAC is non-ribosomal bound in higher eukaryotes. In sum, NAC is a potent suppressor of aggregation and proteotoxicity of mutant PolyQ-expanded proteins associated with human diseases like Huntington's disease and spinocerebellar ataxias.
Asunto(s)
Péptidos beta-Amiloides/genética , Chaperonas Moleculares/genética , Agregación Patológica de Proteínas/genética , Péptidos beta-Amiloides/química , Sitios de Unión/genética , Humanos , Enfermedad de Huntington/genética , Enfermedad de Huntington/patología , Luciferasas/química , Luciferasas/genética , Chaperonas Moleculares/química , Péptidos/química , Péptidos/genética , Unión Proteica/genética , Biosíntesis de Proteínas/genética , Dominios Proteicos/genética , Pliegue de Proteína , Ribosomas/genética , Ataxias Espinocerebelosas/genética , Ataxias Espinocerebelosas/patologíaRESUMEN
Cell-free DNA (cfDNA) fragmentation is nonrandom, at least partially mediated by various DNA nucleases, forming characteristic cfDNA end motifs. However, there is a paucity of tools for deciphering the relative contributions of cfDNA cleavage patterns related to underlying fragmentation factors. In this study, through non-negative matrix factorization algorithm, we used 256 5' 4-mer end motifs to identify distinct types of cfDNA cleavage patterns, referred to as "founder" end-motif profiles (F-profiles). F-profiles were associated with different DNA nucleases based on whether such patterns were disrupted in nuclease-knockout mouse models. Contributions of individual F-profiles in a cfDNA sample could be determined by deconvolutional analysis. We analyzed 93 murine cfDNA samples of different nuclease-deficient mice and identified six types of F-profiles. F-profiles I, II, and III were linked to deoxyribonuclease 1 like 3 (DNASE1L3), deoxyribonuclease 1 (DNASE1), and DNA fragmentation factor subunit beta (DFFB), respectively. We revealed that 42.9% of plasma cfDNA molecules were attributed to DNASE1L3-mediated fragmentation, whereas 43.4% of urinary cfDNA molecules involved DNASE1-mediated fragmentation. We further demonstrated that the relative contributions of F-profiles were useful to inform pathological states, such as autoimmune disorders and cancer. Among the six F-profiles, the use of F-profile I could inform the human patients with systemic lupus erythematosus. F-profile VI could be used to detect individuals with hepatocellular carcinoma, with an area under the receiver operating characteristic curve of 0.97. F-profile VI was more prominent in patients with nasopharyngeal carcinoma undergoing chemoradiotherapy. We proposed that this profile might be related to oxidative stress.
Asunto(s)
Ácidos Nucleicos Libres de Células , Humanos , Ratones , Animales , Ácidos Nucleicos Libres de Células/genética , Desoxirribonucleasas/genética , Ratones Noqueados , Endonucleasas/genética , Fragmentación del ADN , Endodesoxirribonucleasas/genéticaRESUMEN
Cell-free DNA (cfDNA) fragmentation patterns contain important molecular information linked to tissues of origin. We explored the possibility of using fragmentation patterns to predict cytosine-phosphate-guanine (CpG) methylation of cfDNA, obviating the use of bisulfite treatment and associated risks of DNA degradation. This study investigated the cfDNA cleavage profile surrounding a CpG (i.e., within an 11-nucleotide [nt] window) to analyze cfDNA methylation. The cfDNA cleavage proportion across positions within the window appeared nonrandom and exhibited correlation with methylation status. The mean cleavage proportion was â¼twofold higher at the cytosine of methylated CpGs than unmethylated ones in healthy controls. In contrast, the mean cleavage proportion rapidly decreased at the 1-nt position immediately preceding methylated CpGs. Such differential cleavages resulted in a characteristic change in relative presentations of CGN and NCG motifs at 5' ends, where N represented any nucleotide. CGN/NCG motif ratios were correlated with methylation levels at tissue-specific methylated CpGs (e.g., placenta or liver) (Pearson's absolute r > 0.86). cfDNA cleavage profiles were thus informative for cfDNA methylation and tissue-of-origin analyses. Using CG-containing end motifs, we achieved an area under a receiver operating characteristic curve (AUC) of 0.98 in differentiating patients with and without hepatocellular carcinoma and enhanced the positive predictive value of nasopharyngeal carcinoma screening (from 19.6 to 26.8%). Furthermore, we elucidated the feasibility of using cfDNA cleavage patterns to deduce CpG methylation at single CpG resolution using a deep learning algorithm and achieved an AUC of 0.93. FRAGmentomics-based Methylation Analysis (FRAGMA) presents many possibilities for noninvasive prenatal, cancer, and organ transplantation assessment.
Asunto(s)
Ácidos Nucleicos Libres de Células , Neoplasias Hepáticas , Embarazo , Femenino , Humanos , Ácidos Nucleicos Libres de Células/genética , Biomarcadores de Tumor/genética , Metilación de ADN , Neoplasias Hepáticas/genética , Epigénesis Genética , ADN/genética , Citosina , Guanina , Nucleótidos , FosfatosRESUMEN
Urinary cell-free DNA (ucfDNA) is a potential biomarker for bladder cancer detection. However, the biological characteristics of ucfDNA are not well understood. We explored the roles of deoxyribonuclease 1 (DNASE1) and deoxyribonuclease 1-like 3 (DNASE1L3) in the fragmentation of ucfDNA using mouse models. The deletion of Dnase1 in mice (Dnase1-/-) caused aberrations in ucfDNA fragmentation, including a 24-fold increase in DNA concentration, and a 3-fold enrichment of long DNA molecules, with a relative decrease of fragments with thymine ends and reduction of jaggedness (i.e., the presence of single-stranded protruding ends). In contrast, such changes were not observed in mice with Dnase1l3 deletion (Dnase1l3-/-). These results suggested that DNASE1 was an important nuclease contributing to the ucfDNA fragmentation. Western blot analysis revealed that the concentration of DNASE1 protein was higher in urine than DNASE1L3. The native-polyacrylamide gel electrophoresis zymogram showed that DNASE1 activity in urine was higher than that in plasma. Furthermore, the proportion of ucfDNA fragment ends within DNase I hypersensitive sites (DHSs) was significantly increased in Dnase1-deficient mice. In humans, patients with bladder cancer had lower proportions of ucfDNA fragment ends within the DHSs when compared with participants without bladder cancer. The area under the curve (AUC) for differentiating patients with and without bladder cancer was 0.83, suggesting the analysis of ucfDNA fragmentation in the DHSs may have potential for bladder cancer detection. This work revealed the intrinsic links between the nucleases in urine and ucfDNA fragmentomics.
Asunto(s)
Ácidos Nucleicos Libres de Células , Neoplasias de la Vejiga Urinaria , Animales , Ácidos Nucleicos Libres de Células/genética , ADN/genética , Desoxirribonucleasa I/genética , Desoxirribonucleasa I/metabolismo , Endodesoxirribonucleasas/genética , Endonucleasas , Humanos , Ratones , Ratones Noqueados , Neoplasias de la Vejiga Urinaria/genéticaRESUMEN
The effects of DNASE1L3 or DNASE1 deficiency on cell-free DNA (cfDNA) methylation were explored in plasma of mice deficient in these nucleases and in DNASE1L3-deficient humans. Compared to wild-type cfDNA, cfDNA in DNASE1L3-deficient mice was significantly hypomethylated, while cfDNA in DNASE1-deficient mice was hypermethylated. The cfDNA hypomethylation in DNASE1L3-deficient mice was due to increased fragmentation and representation from open chromatin regions (OCRs) and CpG islands (CGIs). These findings were absent in DNASE1-deficient mice, demonstrating the preference of DNASE1 to cleave in hypomethylated OCRs and CGIs. We also observed a substantial decrease of fragment ends at methylated CpGs in the absence of DNASE1L3, thereby demonstrating that DNASE1L3 prefers to cleave at methylated CpGs. Furthermore, we found that methylation levels of cfDNA varied by fragment size in a periodic pattern, with cfDNA of specific sizes being more hypomethylated and enriched for OCRs and CGIs. These findings were confirmed in DNASE1L3-deficient human cfDNA. Thus, we have found that nuclease-mediated cfDNA fragmentation markedly affects cfDNA methylation level on a genome-wide scale. This work provides a foundational understanding of the relationship between methylation, nuclease biology, and cfDNA fragmentation.
Asunto(s)
Ácidos Nucleicos Libres de Células , Fragmentación del ADN , Endodesoxirribonucleasas , Animales , Ácidos Nucleicos Libres de Células/genética , Ácidos Nucleicos Libres de Células/metabolismo , Cromatina , Islas de CpG/genética , Metilación de ADN , Endodesoxirribonucleasas/genética , Humanos , RatonesRESUMEN
Cell-free DNA (cf.DNA) is a powerful noninvasive biomarker for cancer and prenatal testing, and it circulates in plasma as short fragments. To elucidate the biology of cf.DNA fragmentation, we explored the roles of deoxyribonuclease 1 (DNASE1), deoxyribonuclease 1 like 3 (DNASE1L3), and DNA fragmentation factor subunit beta (DFFB) with mice deficient in each of these nucleases. By analyzing the ends of cf.DNA fragments in each type of nuclease-deficient mice with those in wild-type mice, we show that each nuclease has a specific cutting preference that reveals the stepwise process of cf.DNA fragmentation. Essentially, we demonstrate that cf.DNA is generated first intracellularly with DFFB, intracellular DNASE1L3, and other nucleases. Then, cf.DNA fragmentation continues extracellularly with circulating DNASE1L3 and DNASE1. With the use of heparin to disrupt the nucleosomal structure, we also show that the 10 bp periodicity originates from the cutting of DNA within an intact nucleosomal structure. Altogether, this work establishes a model of cf.DNA fragmentation.
Asunto(s)
Ácidos Nucleicos Libres de Células/metabolismo , Cromatina/metabolismo , Fragmentación del ADN , Desoxirribonucleasa I/fisiología , Desoxirribonucleasas/fisiología , Endodesoxirribonucleasas/fisiología , Nucleosomas/metabolismo , Proteínas de Unión a Poli-ADP-Ribosa/fisiología , Animales , Ácidos Nucleicos Libres de Células/genética , Cromatina/genética , Femenino , Masculino , Ratones , Ratones Noqueados , Nucleosomas/genéticaRESUMEN
Plasma DNA fragmentomics is an emerging area in cell-free DNA diagnostics and research. In murine models, it has been shown that the extracellular DNase, DNASE1L3, plays a role in the fragmentation of plasma DNA. In humans, DNASE1L3 deficiency causes familial monogenic systemic lupus erythematosus with childhood onset and anti-dsDNA reactivity. In this study, we found that human patients with DNASE1L3 disease-associated gene variations showed aberrations in size and a reduction of a "CC" end motif of plasma DNA. Furthermore, we demonstrated that DNA from DNASE1L3-digested cell nuclei showed a median length of 153 bp with CC motif frequencies resembling plasma DNA from healthy individuals. Adeno-associated virus-based transduction of Dnase1l3 into Dnase1l3-deficient mice restored the end motif profiles to those seen in the plasma DNA of wild-type mice. Our findings demonstrate that DNASE1L3 is an important player in the fragmentation of plasma DNA, which appears to act in a cell-extrinsic manner to regulate plasma DNA size and motif frequency.
Asunto(s)
ADN/genética , Endodesoxirribonucleasas/genética , Lupus Eritematoso Sistémico/genética , Mutación , Animales , Estudios de Casos y Controles , ADN/sangre , Fragmentación del ADN , Dependovirus/genética , Dependovirus/metabolismo , Modelos Animales de Enfermedad , Endodesoxirribonucleasas/deficiencia , Endodesoxirribonucleasas/metabolismo , Terapia Genética , Vectores Genéticos/química , Vectores Genéticos/metabolismo , Humanos , Lupus Eritematoso Sistémico/enzimología , Lupus Eritematoso Sistémico/patología , Ratones , Ratones Transgénicos , Especificidad por Sustrato , Transducción GenéticaRESUMEN
Cell-free DNA in plasma has been used for noninvasive prenatal testing and cancer liquid biopsy. The physical properties of cell-free DNA fragments in plasma, such as fragment sizes and ends, have attracted much recent interest, leading to the emerging field of cell-free DNA fragmentomics. However, one aspect of plasma DNA fragmentomics as to whether double-stranded plasma molecules might carry single-stranded ends, termed a jagged end in this study, remains underexplored. We have developed two approaches for investigating the presence of jagged ends in a plasma DNA pool. These approaches utilized DNA end repair to introduce differential methylation signals between the original sequence and the jagged ends, depending on whether unmethylated or methylated cytosines were used in the DNA end-repair procedure. The majority of plasma DNA molecules (87.8%) were found to bear jagged ends. The jaggedness varied according to plasma DNA fragment sizes and appeared to be in association with nucleosomal patterns. In the plasma of pregnant women, the jaggedness of fetal DNA molecules was higher than that of the maternal counterparts. The jaggedness of plasma DNA correlated with the fetal DNA fraction. Similarly, in the plasma of cancer patients, tumor-derived DNA molecules in patients with hepatocellular carcinoma showed an elevated jaggedness compared with nontumoral DNA. In mouse models, knocking out of the Dnase1 gene reduced jaggedness, whereas knocking out of the Dnase1l3 gene enhanced jaggedness. Hence, plasma DNA jagged ends represent an intrinsic property of plasma DNA and provide a link between nuclease activities and the fragmentation of plasma DNA.
Asunto(s)
Ácidos Nucleicos Libres de Células/sangre , Fragmentación del ADN , Metilación de ADN/genética , ADN/sangre , ADN/genética , Animales , Carcinoma Hepatocelular/genética , Ácidos Nucleicos Libres de Células/genética , Reparación del ADN por Unión de Extremidades/genética , Endodesoxirribonucleasas/genética , Femenino , Humanos , Neoplasias Hepáticas/genética , Ratones , Ratones Noqueados , Nucleosomas/genética , EmbarazoRESUMEN
BACKGROUND: Nuclear-derived cell-free DNA (cfDNA) molecules in blood plasma are nonrandomly fragmented, bearing a wealth of information related to tissues of origin. DNASE1L3 (deoxyribonuclease 1 like 3) is an important player in shaping the fragmentation of nuclear-derived cfDNA molecules, preferentially generating molecules with 5 CC dinucleotide termini (i.e., 5 CC-end motif). However, the fragment end properties of microbial cfDNA and its clinical implication remain to be explored. METHODS: We performed end motif analysis on microbial cfDNA fragments in plasma samples from patients with sepsis. A sequence context-based normalization method was used to minimize the potential biases for end motif analysis. RESULTS: The end motif profiles of microbial cfDNA appeared to resemble that of nuclear cfDNA (Spearman correlation coefficient: 0.82, P value 0.001). The CC-end motif was the most preferred end motif in microbial cfDNA, suggesting that DNASE1L3 might also play a role in the fragmentation of microbe-derived cfDNA in plasma. Of note, differential end motifs were present between microbial cfDNA originating from infection-causing pathogens (enriched at the CC-end) and contaminating microbial DNA potentially derived from reagents or the environment (nearly random). The use of fragment end signatures allowed differentiation between confirmed pathogens and contaminating microbes, with an area under the receiver operating characteristic curve of 0.99. The performance appeared to be superior to conventional analysis based on microbial cfDNA abundance alone. CONCLUSIONS: The use of fragmentomic features could facilitate the differentiation of underlying contaminating microbes from true pathogens in sepsis. This work demonstrates the potential usefulness of microbial cfDNA fragmentomics in metagenomics analysis.
Asunto(s)
Ácidos Nucleicos Libres de Células , Sepsis , Humanos , ADN/genética , Sepsis/diagnóstico , Fragmentación del ADNRESUMEN
Juvenile myelomonocytic leukemia (JMML) is an aggressive myeloproliferative neoplasia that lacks effective targeted chemotherapies. Clinically, JMML manifests as monocytic leukocytosis, splenomegaly with consequential thrombocytopenia. Most commonly, patients have gain-of-function (GOF) oncogenic mutations in PTPN11 (SHP2), leading to Erk and Akt hyperactivation. Mechanism(s) involved in co-regulation of Erk and Akt in the context of GOF SHP2 are poorly understood. Here, we show that Bruton's tyrosine kinase (BTK) is hyperphosphorylated in GOF Shp2-bearing cells and utilizes B cell adaptor for PI3K to cooperate with p110δ, the catalytic subunit of PI3K. Dual inhibition of BTK and p110δ reduces the activation of both Erk and Akt. In vivo, individual targeting of BTK or p110δ in a mouse model of human JMML equally reduces monocytosis and splenomegaly; however, the combined treatment results in a more robust inhibition and uniquely rescues anemia and thrombocytopenia. RNA-seq analysis of drug-treated mice showed a profound reduction in the expression of genes associated with leukemic cell migration and inflammation, leading to correction in the infiltration of leukemic cells in the lung, liver, and spleen. Remarkably, in a patient derived xenograft model of JMML, leukemia-initiating stem and progenitor cells were potently inhibited in response to the dual drug treatment.
Asunto(s)
Leucemia Mielomonocítica Juvenil , Trombocitopenia , Agammaglobulinemia Tirosina Quinasa/genética , Animales , Humanos , Leucemia Mielomonocítica Juvenil/genética , Leucemia Mielomonocítica Juvenil/metabolismo , Leucemia Mielomonocítica Juvenil/terapia , Ratones , Fosfatidilinositol 3-Quinasas , Proteínas Proto-Oncogénicas c-akt/metabolismo , Esplenomegalia/genética , Células Madre/metabolismoRESUMEN
As the coronavirus disease 2019 (COVID-19) pandemic evolves, much evidence implicates the heart as a critical target of injury in patients. The mechanism(s) of cardiac involvement has not been fully elucidated, although evidence of direct virus-mediated injury, thromboembolism with ischemic complications, and cytokine storm has been reported. We examined suggested mechanisms of COVID-19-associated heart failure in 21 COVID-19-positive decedents, obtained through standard autopsy procedure, compared to clinically matched controls and patients with various etiologies of viral myocarditis. We developed a custom tissue microarray using regions of pathological interest and interrogated tissues via immunohistochemistry and in situ hybridization. Severe acute respiratory syndrome coronavirus 2 was detected in 16/21 patients, in cardiomyocytes, the endothelium, interstitial spaces, and percolating adipocytes within the myocardium. Virus detection typically corresponded with troponin depletion and increased cleaved caspase-3. Indirect mechanisms of injury-venous and arterial thromboses with associated vasculitis including a mixed inflammatory infiltrate-were also observed. Neutrophil extracellular traps (NETs) were present in the myocardium of all COVID-19 patients, regardless of injury degree. Borderline myocarditis (inflammation without associated myocyte injury) was observed in 19/21 patients, characterized by a predominantly mononuclear inflammatory infiltrate. Edema, inflammation of percolating adipocytes, lymphocytic aggregates, and large septal masses of inflammatory cells and platelets were observed as defining features, and myofibrillar damage was evident in all patients. Collectively, COVID-19-associated cardiac injury was multifactorial, with elevated levels of NETs and von Willebrand factor as defining features of direct and indirect viral injury.
Asunto(s)
COVID-19 , Miocarditis , Autopsia , COVID-19/complicaciones , Humanos , Inflamación , Miocitos CardíacosRESUMEN
BACKGROUND: Jagged ends of plasma DNA are a recently recognized class of fragmentomic markers for cell-free DNA, reflecting the activity of nucleases. A number of recent studies have also highlighted the importance of jagged ends in the context of pregnancy and oncology. However, knowledge regarding the generation of jagged ends is incomplete. METHODS: Jaggedness of plasma DNA was analyzed based on Jag-seq, which utilized the differential methylation signals introduced by the DNA end-repair process. We investigated the jagged ends in plasma DNA using mouse models by deleting the deoxyribonuclease 1 (Dnase1), DNA fragmentation factor subunit beta (Dffb), or deoxyribonuclease 1 like 3 (Dnase1l3) gene. RESULTS: Aberrations in the profile of plasma DNA jagged ends correlated with the type of nuclease that had been genetically deleted, depending on nucleosomal structures. The deletion of Dnase1l3 led to a significant reduction of jaggedness for those plasma DNA molecules involving more than 1 nucleosome (e.g., size ranges 240-290â bp, 330-380â bp, and 420-470â bp). However, less significant effects of Dnase1 and Dffb deletions were observed regarding different sizes of DNA fragments. Interestingly, the aberration in plasma DNA jagged ends related to multinucleosomes was observed in human subjects with familial systemic lupus erythematosus with Dnase1l3 deficiency and human subjects with sporadic systemic lupus erythematosus. CONCLUSIONS: Detailed understanding of the relationship between nuclease and plasma DNA jaggedness has opened up avenues for biomarker development.
Asunto(s)
Ácidos Nucleicos Libres de Células , Lupus Eritematoso Sistémico , Animales , Biomarcadores , Ácidos Nucleicos Libres de Células/genética , ADN/genética , Desoxirribonucleasas/genética , Endodesoxirribonucleasas/genética , Femenino , Humanos , Lupus Eritematoso Sistémico/genética , Ratones , Nucleosomas/genética , EmbarazoRESUMEN
Circulating DNA in plasma consists of short DNA fragments. The biological processes generating such fragments are not well understood. DNASE1L3 is a secreted DNASE1-like nuclease capable of digesting DNA in chromatin, and its absence causes anti-DNA responses and autoimmunity in humans and mice. We found that the deletion of Dnase1l3 in mice resulted in aberrations in the fragmentation of plasma DNA. Such aberrations included an increase in short DNA molecules below 120 bp, which was positively correlated with anti-DNA antibody levels. We also observed an increase in long, multinucleosomal DNA molecules and decreased frequencies of the most common end motifs found in plasma DNA. These aberrations were independent of anti-DNA response, suggesting that they represented a primary effect of DNASE1L3 loss. Pregnant Dnase1l3-/- mice carrying Dnase1l3+/- fetuses showed a partial restoration of normal frequencies of plasma DNA end motifs, suggesting that DNASE1L3 from Dnase1l3-proficient fetuses could enter maternal systemic circulation and affect both fetal and maternal DNA fragmentation in a systemic as well as local manner. However, the observed shortening of circulating fetal DNA relative to maternal DNA was not affected by the deletion of Dnase1l3 Collectively, our findings demonstrate that DNASE1L3 plays a role in circulating plasma DNA homeostasis by enhancing fragmentation and influencing end-motif frequencies. These results support a distinct role of DNASE1L3 as a regulator of the physical form and availability of cell-free DNA and may have important implications for the mechanism whereby this enzyme prevents autoimmunity.
Asunto(s)
Ácidos Nucleicos Libres de Células/sangre , Fragmentación del ADN , ADN/sangre , Endodesoxirribonucleasas/metabolismo , Motivos de Nucleótidos , Animales , Ácidos Nucleicos Libres de Células/genética , ADN/genética , Endodesoxirribonucleasas/genética , Femenino , Feto/metabolismo , Eliminación de Gen , Ratones , Ratones Noqueados , EmbarazoRESUMEN
Comorbidities influence survival in patients with chronic lymphocytic leukaemia (CLL) treated with chemo-immunotherapy or ibrutinib. While idelalisib has been studied in patients with comorbidities, their impact has not been investigated. We analysed 481 patients treated with idelalisib on two randomised trials (NCT01659021 and NCT01539512). Comorbidities were assessed using the Cumulative Illness Risk Scale (CIRS). Patients received idelalisib + anti-CD20 (rituximab or ofatumumab; n = 284) or anti-CD20 alone (n = 197). The median age was 69 years. We found that comorbidities did not significantly affect outcomes of idelalisib therapy. The objective response rate (ORR) was 79·3% versus 85·8%, the median progression-free survival (PFS) was 16·3 versus 19·1 months, and the median overall survival (OS) was 39·8 versus 49·8 months in patients treated with idelalisib who had a CIRS score of >6 versus ≤6, correspondingly. Treatment with idelalisib + anti-CD20 was associated with superior PFS and ORR when compared to anti-CD20 monotherapy in patients who had high comorbidities (CIRS score of >6) or at least one severe comorbidity (median PFS 16·3 vs. 6·9 months and 16·6 vs. 6·5 months; odds ratio 20·1 and 33·2; P < 0·0001). Thus, comorbidities do not portend inferior outcomes in patients with CLL treated with idelalisib in combination with anti-CD20 therapy.
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Antineoplásicos/uso terapéutico , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Purinas/uso terapéutico , Quinazolinonas/uso terapéutico , Adulto , Anciano , Anciano de 80 o más Años , Anticuerpos Monoclonales Humanizados/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Comorbilidad , Supervivencia sin Enfermedad , Femenino , Humanos , Leucemia Linfocítica Crónica de Células B/epidemiología , Masculino , Persona de Mediana Edad , Supervivencia sin Progresión , Rituximab/uso terapéutico , Resultado del TratamientoRESUMEN
BACKGROUND: Plate fixation is frequently used to treat displaced midshaft clavicular fractures, however the ideal plate choice remains subject to discussion; reconstruction locking compression plates (RLCPs) are cheaper and can be easily contoured, whereas anatomically pre-contoured locking compression plates (ALCPs) are thought to provide better stability and therefore lower rates of mechanical failure. To compare the incidence of mechanical failures, functional and radiological outcomes in patients with midshaft clavicular fractures treated with ALCPs versus RLCPs. METHODS: A propensity score matched retrospective cohort study was conducted across two centers. One hundred and six consecutively recruited patients with displaced midshaft clavicular fractures, who were treated with plate fixation and had a minimum follow-up of 6 months, were matched on gender, age, fracture grading, energy of injury, and fracture location. The resulting groups included 53 ALCP-treated fractures and 53 matched controls treated with RLCPs. RESULTS: During a mean follow-up of 20.5 months, there were no implant deformities in the ALCP group whereas the RLCP group had 6 patients (11.3%, p = 0.012) with implant deformities (5 occurrences of plate bending with fracture union, and 1 plate breakage with nonunion). Despite the higher rate of plate deformities in the RLCP group, there were no statistically significant differences in number of patients recovering full shoulder range of motion (ALCP 90.6%, RLCP 88.7%, p = 0.751), incidence of rest pain (ALCP 13.2%, RLCP 9.4%, p = 0.542), or implant removals (ALCP 49.1%, RLCP 56.6%, p = 0.439). CONCLUSION: ALCPs may be superior to RLCPs in terms of implant stability but appear to produce similar clinical results.
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Placas Óseas , Clavícula/lesiones , Fracturas Óseas/cirugía , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Tornillos Óseos , Clavícula/cirugía , Remoción de Dispositivos , Femenino , Fijación Interna de Fracturas/instrumentación , Curación de Fractura , Fracturas Óseas/diagnóstico por imagen , Hong Kong , Humanos , Masculino , Persona de Mediana Edad , Puntaje de Propensión , Radiografía , Rango del Movimiento Articular , Estudios Retrospectivos , Resultado del Tratamiento , Adulto JovenRESUMEN
BACKGROUND: Resection of insular tumours utilising modern neurosurgical techniques has become commonplace since its safety and reduced morbidity was first established. Interest has grown in the cognitive consequences of insula neurosurgery and studies have largely shown postoperative stability or minor decline. Major or widespread improvements in cognitive functioning following resection of insular tumours have not previously been reported. CASE DESCRIPTION: A 34-year-old, left-handed man with a right insular low-grade glioma (LGG) presented with seizures, nausea, altered sensation, poor balance and extensive cognitive decline. Comprehensive neuropsychological assessment highlighted a striking left hemispatial neglect and impairments in attention, working memory, verbal learning and fluency. During an awake craniotomy with functional cortical mapping, he reported intraoperative improvements in hand function and processing speed. Resolution of the neglect and significant improvements in cognition, mood and functioning were observed at follow-up and sustained over several years. CONCLUSIONS: This case highlights that right insular LGGs can cause significant cognitive and functional deficits and that neurosurgery has the potential to alleviate these difficulties to an extent beyond those documented in the extant literature.
RESUMEN
OBJECTIVE: Growing evidence suggests that psychological treatments are effective for improving outcomes in both epilepsy and psychogenic nonepileptic seizures (PNES). However, the risk of in-session seizures may cause concerns about safety and about seizures disrupting treatment. This study explores the risk of in-session seizures in patients with epilepsy and those with PNES, the timings of seizures during psychological therapy, and the responses of therapists to seizures. METHODS: Consecutive patients with epilepsy or PNES attending 2 neurology centers in the United Kingdom for psychological treatment to help with their seizure disorders were studied. Information about seizures during outpatient psychological therapy sessions was gathered using a 12-item pro forma. RESULTS: Ninety-seven patients with epilepsy and 195 with PNES were evaluated. One in 32 patients with epilepsy and 1 in 8 with PNES had at least 1 in-session seizure. A seizure occurred in 1 in 136 treatment sessions provided to patients with epilepsy, and 1 in 36 sessions provided to those with PNES. The risk of in-session seizures was significantly greater in patients with PNES than in patients with epilepsy (odds ratio [OR] 4.4, 95% confidence interval [CI] 1.3-15.2). Seizures tended to occur in the first half of treatment programs and individual sessions and only disrupted sessions briefly. Only one patient with PNES required in-patient observation not involving overnight admission. SIGNIFICANCE: In-session seizures do occur and are much more common in patients with PNES than in those with epilepsy. Seizures rarely caused major disruption to psychological treatment, and could almost invariably be managed by the treating therapist without help from additional medical staff. Nonetheless, this research suggests that psychological therapy providers should anticipate the occurrence of in-session seizures and have safe management plans in place. The greater frequency of in-session seizures in PNES may add to our understanding of the mechanisms triggering these seizures.
Asunto(s)
Terapia Cognitivo-Conductual/métodos , Manejo de la Enfermedad , Trastornos Psicofisiológicos/diagnóstico , Trastornos Psicofisiológicos/terapia , Convulsiones/diagnóstico , Convulsiones/terapia , Terapia de Aceptación y Compromiso/métodos , Terapia de Aceptación y Compromiso/tendencias , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Terapia Cognitivo-Conductual/tendencias , Electroencefalografía/tendencias , Epilepsia/diagnóstico , Epilepsia/psicología , Epilepsia/terapia , Femenino , Humanos , Masculino , Persona de Mediana Edad , Trastornos Psicofisiológicos/psicología , Estudios Retrospectivos , Convulsiones/psicología , Resultado del Tratamiento , Adulto JovenRESUMEN
Plasma consists of DNA released from multiple tissues within the body. Using genome-wide bisulfite sequencing of plasma DNA and deconvolution of the sequencing data with reference to methylation profiles of different tissues, we developed a general approach for studying the major tissue contributors to the circulating DNA pool. We tested this method in pregnant women, patients with hepatocellular carcinoma, and subjects following bone marrow and liver transplantation. In most subjects, white blood cells were the predominant contributors to the circulating DNA pool. The placental contributions in the plasma of pregnant women correlated with the proportional contributions as revealed by fetal-specific genetic markers. The graft-derived contributions to the plasma in the transplant recipients correlated with those determined using donor-specific genetic markers. Patients with hepatocellular carcinoma showed elevated plasma DNA contributions from the liver, which correlated with measurements made using tumor-associated copy number aberrations. In hepatocellular carcinoma patients and in pregnant women exhibiting copy number aberrations in plasma, comparison of methylation deconvolution results using genomic regions with different copy number status pinpointed the tissue type responsible for the aberrations. In a pregnant woman diagnosed as having follicular lymphoma during pregnancy, methylation deconvolution indicated a grossly elevated contribution from B cells into the plasma DNA pool and localized B cells as the origin of the copy number aberrations observed in plasma. This method may serve as a powerful tool for assessing a wide range of physiological and pathological conditions based on the identification of perturbed proportional contributions of different tissues into plasma.
Asunto(s)
Carcinoma Hepatocelular/genética , Metilación de ADN , ADN/genética , Neoplasias Hepáticas/genética , Análisis de Secuencia de ADN/métodos , Trasplante de Tejidos , Adulto , Algoritmos , Linfocitos B/metabolismo , Trasplante de Médula Ósea , Carcinoma Hepatocelular/sangre , ADN/sangre , ADN/química , Variaciones en el Número de Copia de ADN/genética , Femenino , Feto/metabolismo , Humanos , Hígado/metabolismo , Hígado/patología , Neoplasias Hepáticas/sangre , Trasplante de Hígado , Persona de Mediana Edad , Neutrófilos/metabolismo , Placenta/metabolismo , Embarazo , Linfocitos T/metabolismoRESUMEN
BACKGROUND: Accumulating evidence suggests the origin of juvenile myelomonocytic leukemia (JMML) is closely associated with fetal development. Nevertheless, the contribution of embryonic progenitors to JMML pathogenesis remains unexplored. We hypothesized that expression of JMML-initiating PTPN11 mutations in HSC-independent yolk sac erythromyeloid progenitors (YS EMPs) would result in a mouse model of pediatric myeloproliferative neoplasm (MPN). RESULTS: E9.5 YS EMPs from VavCre+;PTPN11D61Y embryos demonstrated growth hypersensitivity to granulocyte-macrophage colony-stimulating factor (GM-CSF) and hyperactive RAS-ERK signaling. Mutant EMPs engrafted the spleens of neonatal recipients, but did not cause disease. To assess MPN development during unperturbed hematopoiesis we generated CSF1R-MCM+;PTPN11E76K ;ROSAYFP mice in which oncogene expression was restricted to EMPs. Yellow fluorescent protein-positive progeny of mutant EMPs persisted in tissues one year after birth and demonstrated hyperactive RAS-ERK signaling. Nevertheless, these mice had normal survival and did not demonstrate features of MPN. CONCLUSIONS: YS EMPs expressing mutant PTPN11 demonstrate functional and molecular features of JMML but do not cause disease following transplantation nor following unperturbed development. Developmental Dynamics 246:1001-1014, 2017. © 2017 Wiley Periodicals, Inc.