Production Of Dna Minicircles Less Than 250 Base Pairs Through A Novel Concentrated Dna Circularization Assay Enabling Minicircle Design With Nf-κb Inhibition Activity

Double-stranded DNA minicircles of less than 1000 bp in length have great interest in both fundamental research and therapeutic applications. Although minicircles have shown promising activity in gene therapy thanks to their good biostability and better intracellular trafficking, minicircles down to 250 bp in size have not yet been investigated from the test tube to the cell for lack of an efficient production method. Herein, we report a novel versatile plasmidfree method for the production of DNA minicircles comprising fewer than 250 bp. We designed a linear nicked DNA double-stranded oligonucleotide bluntended substrate for efficient minicircle production in a ligase-mediated and bending protein-assisted circularization reaction at high DNA concentration of 2M. This one pot multi-step reaction based-method yields hundreds of micrograms of minicircle with sequences of any base composition and position and containing or not a variety of site-specifically chemical modifications or physiological supercoiling. Biochemical and cellular studies were then conducted to design a 95 bp minicircle capable of binding in vitro two NF-κB transcription factors per minicircle and to efficiently inhibiting NF-κB-dependent transcriptional activity in human cells. Therefore, our production method could pave the way for the design of minicircles as new decoy nucleic acids. © The Author(s) 2016.45

Hypochloremia and hyponatremia as the initial presentation of cystic fibrosis in three adults

Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Most diagnoses of CF are made during infancy or childhood, and are based on respiratory or digestive involvement. Initial extracellular dehydration leading to the diagnosis of CF is usual in infants but has only exceptionally been reported in adults. We describe three new adult cases of CF initially presenting with depletive hyponatremia and hypochloremia following exposure to heat. At first consultation, these patients had no symptoms suggestive of CF. One patient presented with a seizure induced by hyponatremia. The two other patients were siblings carrying a novel c.4434insA mutation in exon 24 of CFTR. Acute dehydration is a very rare initial manifestation of CF but may be life-threatening. The possibility of CF should not be ignored in cases of depletive hyponatremia, hypochloremia or hypokalemic metabolic alkalosis, even in otherwise healthy patients

Exophiala dermatitidis Revealing Cystic Fibrosis in Adult Patients with Chronic Pulmonary Disease

Cystic fibrosis (CF) is a genetic inherited disease due to mutations in the gene cystic fibrosis transmembrane conductance regulator (CFTR). Because of the huge diversity of CFTR mutations, the CF phenotypes are highly heterogeneous, varying from typical to mild form of CF, also called atypical CF. These atypical features are more frequently diagnosed at adolescence or adulthood, and among clinical signs and symptoms leading to suspect a mild form of CF, colonization or infection of the respiratory tract due to well-known CF pathogens should be a warning signal. Exophiala dermatitidis is a melanized dimorphic fungus commonly detected in respiratory specimens from CF patients, but only very rarely from respiratory specimens from non-CF patients. We described here two cases of chronic colonization of the airways by E. dermatitidis, with recurrent pneumonia and hemoptysis in one patient, which led clinicians to diagnose mild forms of CF in these elderly patients who were 68- and 87-year-old. These cases of late CF diagnosis suggest that airway colonization or respiratory infections due to E. dermatitidis in patients with bronchiectasis should led to search for a mild form of CF, regardless of the age and associated symptoms. On a broader level, in patients with chronic respiratory disease and recurrent pulmonary infections, an allergic bronchopulmonary mycosis or an airway colonization by CF-related fungi like E. dermatitidis or some Aspergillus, Scedosporium or Rasamsonia species, should be considered as potential markers of atypical CF and should led clinicians to conduct investigations for CF diagnosis

Variants Within TSC2 Exons 25 and 31 Are Very Unlikely to Cause Clinically Diagnosable Tuberous Sclerosis

Inactivating mutations in TSC1 and TSC2 cause tuberous sclerosis complex (TSC). The 2012 international consensus meeting on TSC diagnosis and management agreed that the identification of a pathogenic TSC1 or TSC2 variant establishes a diagnosis of TSC, even in the absence of clinical signs. However, exons 25 and 31 of TSC2 are subject to alternative splicing. No variants causing clinically diagnosed TSC have been reported in these exons, raising the possibility that such variants would not cause TSC. We present truncating and in‐frame variants in exons 25 and 31 in three individuals unlikely to fulfil TSC diagnostic criteria and examine the importance of these exons in TSC using different approaches. Amino acid conservation analysis suggests significantly less conservation in these exons compared with the majority of TSC2 exons, and TSC2 expression data demonstrates that the majority of TSC2 transcripts lack exons 25 and/or 31 in many human adult tissues. In vitro assay of both exons shows that neither exon is essential for TSC complex function. Our evidence suggests that variants in TSC2 exons 25 or 31 are very unlikely to cause classical TSC, although a role for these exons in tissue/stage specific development cannot be excluded

Very Active Neutral P,O-Chelated Nickel Catalysts for Ethylene Polymerization

ABSTRACT: A series of highly active nickel-based neutral catalysts for ethylene polymerization is presented. These catalysts are obtained by direct complexation of simple fluorinated ketoylides onto bis-(1,5-cyclooctadiene)nickel. Catalyst formation readily occurs in the presence of an olefin, but due to the electron deficiency of the ligand, it hardly happens in the absence of an olefin or another Lewis base. Activities greater than 2 × 10 6 (gPE/gNi)/h and productivities higher than 15 × 10 6 gPE/molNi are typically observed. These catalysts are also active for the polymerization of R-olefins such as 1-hexene and 1-propene. Polymer characterization indicates that highly linear, low molecular weight PEHD is formed by these complexes

Characterization of mesenchymal stem cells in pre-B acute lymphoblastic leukemia

Components of the bone marrow microenvironment (BMM) have been shown to mediate the way in which leukemia develops, progresses and responds to treatment. Increasing evidence shows that leukemic cells hijack the BMM, altering its functioning and establishing leukemia-supportive interactions with stromal and immune cells. While previous work has highlighted functional defects in the mesenchymal stem cell (MSC) population from the BMM of acute leukemias, thorough characterization and molecular profiling of MSCs in pre-B cell acute lymphoblastic leukemia (B-ALL), the most common cancer in children, has not been conducted. Here, we investigated the cellular and transcriptome profiles of MSCs isolated from the BMM of an immunocompetent BCR-ABL1+ model of B-ALL. Leukemia-associated MSCs exhibited reduced self-renewal capacity in vitro and significant changes in numerous molecular signatures, including upregulation of inflammatory signaling pathways. Additionally, we found downregulation of genes involved in extracellular matrix organization and osteoblastogenesis in leukemia-associated MSCs. This study provides cellular and molecular insights into the role of MSCs during B-ALL progression

Building the Future Therapies for Down Syndrome: The Third International Conference of the T21 Research Society

Research focused on Down syndrome has increased in the last several years to advance understanding of the consequences of trisomy 21 (T21) on molecular and cellular processes and, ultimately, on individuals with Down syndrome. The Trisomy 21 Research Society (T21RS) is the premier scientific organization for researchers and clinicians studying Down syndrome. The Third International Conference of T21RS, held June 6–9, 2019, in Barcelona, Spain, brought together 429 scientists, families, and industry representatives to share the latest discoveries on underlying cellular and molecular mechanisms of T21, define cognitive and behavioral challenges and better understand comorbidities associated with Down syndrome, including Alzheimer’s disease and leukemia. Presentation of cutting-edge results in neuroscience, neurology, model systems, psychology, cancer, biomarkers and molecular and phar­ma­cological therapeutic approaches demonstrate the compelling interest and continuing advancement in all aspects of understanding and ameliorating conditions associated with T21