Rapid detection of copy number variations and point mutations in BRCA1/2 genes using a single workflow by ion semiconductor sequencing pipeline

Molecular analysis of BRCA1 (MIM# 604370) and BRCA2 (MIM #600185) genes is essential for familial breast and ovarian cancer prevention and treatment. An efficient, rapid, cost-effective accurate strategy for the detection of pathogenic variants is crucial. Mutations detection of BRCA1/2 genes includes screening for single nucleotide variants (SNVs), small insertions or deletions (indels), and Copy Number Variations (CNVs). Sanger sequencing is unable to identify CNVs and therefore Multiplex Ligation Probe amplification (MLPA) or Multiplex Amplicon Quantification (MAQ) is used to complete the BRCA1/2 genes analysis. The rapid evolution of Next Generation Sequencing (NGS) technologies allows the search for point mutations and CNVs with a single platform and workflow. In this study we test the possibilities of NGS technology to simultaneously detect point mutations and CNVs in BRCA1/2 genes, using the OncomineTM BRCA Research Assay on Personal Genome Machine (PGM) Platform with Ion Reporter Software for sequencing data analysis (Thermo Fisher Scientific). Comparison between the NGS-CNVs, MLPA and MAQ results shows how the NGS approach is the most complete and fast method for the simultaneous detection of all BRCA mutations, avoiding the usual time consuming multistep approach in the routine diagnostic testing of hereditary breast and ovarian cancers

Pretransplant recipient blood CD14+ preDC levels correlate with increased acute GVHD after allogeneic PBSC transplantation

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Complications and outcome of cats with congenital extrahepatic portosystemic shunts treated with thin film: Thirty-four cases (2008-2017)

Background: Congenital extrahepatic portosystemic shunts (CEHPSS) are rare in cats. Outcome after attenuation of CEHPSS with thin film has been described in a small number of cases. Objectives: To describe the clinical presentation, postoperative complications, and outcome of cats treated with thin film to attenuate CEHPSS. Animals: Thirty‐four cats with CEHPSS were identified from the database of 3 institutions over 9 years. Methods: Retrospective study. Medical records were reviewed to identify cats with a diagnosis of a CEHPSS that underwent surgical attenuation. Congenital extrahepatic portosystemic shunts were suspected from clinical signs, clinicopathologic findings, and diagnostic imaging, and confirmed at exploratory laparotomy. Cats treated with thin film band attenuation were included. Postoperative complications and follow‐up were recorded. Results: Complications were recorded in 11 of 34 cats. Deaths related to CEHPSS occurred in 6 of 34; 4 cats did not survive to discharge. Persistent seizures were the cause of death in 4 cats. Seizures were recorded in 8 of 34 cats after surgery; all these cats received preoperative antiepileptic drugs. Serum bile acid concentrations normalized in 25 of 28 of the cats for which data was available. Three cats had persistently increased serum bile acid concentrations and underwent a second exploratory laparotomy. One had a patent shunt, the other 2 had multiple acquired portosystemic shunts. Median follow‐up was 8 months (0.5‐84 months). Conclusions and Clinical Importance: Congenital extrahepatic portosystemic shunts attenuation using thin film in cats carries a good short‐ and mid‐term prognosis if they survive the postoperative period. Seizures were the most common cause of death

Molecular Monitoring of BCR-ABL Transcripts after Allogeneic Stem Cell Transplantation for Chronic Myeloid Leukemia

AbstractThe monitoring of minimal residual disease (MRD) through low sensitivity real-time (RT) polymerase chain reaction (PCR) analysis of BCR-ABL transcripts allows early detection of chronic myeloid leukemia (CML) relapse after allogeneic hematopoietic stem cell transplantation (HSCT). The introduction of more sensitive techniques, such as RT quantitative (Q)-PCR, may lead to an overestimation of the risk of CML relapse. In this study, we reviewed the results of peripheral blood RT Q-PCR in CML patients who underwent allogeneic HSCT from 1983 to 2007. In our laboratory, RT Q-PCR analysis was routinely performed since 2002. Eighty-seven of 189 patients had available RT Q-PCR data; 63 patients had at least 3 RT Q-PCR analyses assessable. Fifty-two of 63 patients (83%) had, at least once, detectable transcript levels, but with an BCR-ABL/ABL ratio <.1% defined as .1% confirmed by the finding of Ph+ cells in bone marrow. No patients with persistent undetectable transcripts relapsed (P = .19). Relapse did not correlate with the number of occurrences o

118: Elevated CD14+ Cell Dose in Marrow Graft Correlates with Increased Mortality after Allogeneic Transplantation

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Selective mitochondrial antioxidant MitoTEMPO reduces renal dysfunction and systemic inflammation in experimental sepsis in rats

BACKGROUND: Excess mitochondrial reactive oxygen species (mROS) in sepsis is associated with organ failure, in part by generating inflammation through the NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3) inflammasome. We determined the impact of a mitochondrial-targeted antioxidant (MitoTEMPO) on mitochondrial dysfunction in renal proximal tubular epithelial cells, peritoneal immune cell function ex vivo, and organ dysfunction in a rat model of sepsis. METHODS: The effects of MitoTEMPO were assessed ex vivo using adenosine triphosphate and lipopolysaccharide-stimulated rat peritoneal immune cells and fresh rat kidney slices exposed to serum from septic rats. We assessed mROS production and phagocytotic capacity (flow cytometry), mitochondrial functionality (multiphoton imaging, respirometry), and NLRP3 inflammasome activation in cell culture. The effect of MitoTEMPO on organ dysfunction was evaluated in a rat model of faecal peritonitis. RESULTS: MitoTEMPO decreased septic serum-induced mROS (P0.05). In vivo, compared with untreated septic animals, MitoTEMPO reduced systemic IL-1β (P=0.01), reduced renal oxidative stress as determined by urine isoprostane levels (P=0.04), and ameliorated renal dysfunction (reduced serum urea (P<0.001) and creatinine (P=0.05). CONCLUSIONS: Reduction of mROS by a mitochondria-targeted antioxidant reduced IL-1β, and protected mitochondrial, cellular, and organ functionality after septic insults

392: Delayed Recovery of Myeloid (mDC) and Plasmacytoid (pDC) Dendritic Cells at 3 Months after Allogeneic HSC Transplantation Correlates with an Increased Transplant-related and Overall Mortality Independently of GVHD

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