Molecular analysis of afibrinogenemic mutations caused by a homozygous FGA1238 bp deletion, and a compound heterozygous FGA1238 bp deletion and novel FGA c.54+3A > C substitution

We identified two afibrinogenemic girls in two Japanese families and performed molecular analysis to clarify the mechanisms of fibrinogen defects. Genetic analyses were performed by PCR amplification of the fibrinogen gene and DNA sequence analysis. To analyze the mechanisms of mature fibrinogen defects in plasma, we cloned minigenes from the proposita's PCR-amplified DNA, transfected them into CHO cells, and sequenced the cDNA amplified with the RT reaction followed by PCR. Sequence analyses indicated that one was caused by a homozygous 1238 bp deletion of the fibrinogen A alpha-chain gene (FGA Delta 1238) and the other was a compound heterozygous FGA Delta 1238 and novel FGA c.54+3A > C substitution. The minigene corresponding to FGA Delta 1238 generates two aberrant mRNAs, both of which may induce a frameshift and terminate prematurely. In contrast, the minigene corresponding to FGA c.54+3A > C generates two aberrant mRNAs, one of which may induce a frameshift and terminate prematurely, and the other uses a cryptic 5' splice site in exon 1, resulting in the deletion of six amino acids in signal peptides. Molecular analyses of both genetic variants suggest that the lack of a mature A alpha-chain, impaired assembly, and/or secretion of the fibrinogen molecule may lead to afibrinogenemia.ArticleINTERNATIONAL JOURNAL OF HEMATOLOGY. 96(1):39-46 (2012)journal articl

Molecular analysis of afibrinogenemic mutations caused by a homozygous FGA1238 bp deletion, and a compound heterozygous FGA1238 bp deletion and novel FGA c.54+3A>C substitution

We identified two afibrinogenemic girls in two Japanese families and performed molecular analysis to clarify the mechanisms of fibrinogen defects. Genetic analyses were performed by PCR amplification of the fibrinogen gene and DNA sequence analysis. To analyze the mechanisms of mature fibrinogen defects in plasma, we cloned minigenes from the proposita's PCR-amplified DNA, transfected them into CHO cells, and sequenced the cDNA amplified with the RT reaction followed by PCR. Sequence analyses indicated that one was caused by a homozygous 1238 bp deletion of the fibrinogen A alpha-chain gene (FGA Delta 1238) and the other was a compound heterozygous FGA Delta 1238 and novel FGA c.54+3A > C substitution. The minigene corresponding to FGA Delta 1238 generates two aberrant mRNAs, both of which may induce a frameshift and terminate prematurely. In contrast, the minigene corresponding to FGA c.54+3A > C generates two aberrant mRNAs, one of which may induce a frameshift and terminate prematurely, and the other uses a cryptic 5' splice site in exon 1, resulting in the deletion of six amino acids in signal peptides. Molecular analyses of both genetic variants suggest that the lack of a mature A alpha-chain, impaired assembly, and/or secretion of the fibrinogen molecule may lead to afibrinogenemia.ArticleINTERNATIONAL JOURNAL OF HEMATOLOGY. 96(1):39-46 (2012)journal articl

Abdominal and Thigh Muscle Attenuation Is Associated With Visceral Fat and Age in Children and Adolescents With Obesity

Objective . This study investigated relationships between adipose tissue deposition within skeletal muscle and morphological and biochemical variables in children with obesity. Methods . Fifty-one Japanese children (16 girls) aged 7 to 16 years were assigned to either mild (<20%), moderate (≥20% to <50%), or severe obesity groups (≥50%). Computed tomography images were taken to calculate the cross-sectional area (CSA) of muscle, subcutaneous and visceral adipose tissue (VAT, for abdomen only), and muscle signal intensities, as an index of intramuscular adipose tissue in the anterior, lateral, and posterior muscles of the abdomen and quadriceps, hamstring and adductor muscles in the thigh. Fasting blood samples were collected to measure plasma lipids, glutamate oxaloacetate transaminase, glutamate oxaloacetate transaminase, uric acid, glucose, and HbA1c. Results . Signal intensity in the severe obesity group was significantly lower than mild and moderate obesity groups in the abdomen and significantly lower than the moderate obesity group in the thigh. Stepwise regression analysis with signal intensity as dependent variable revealed that VAT CSA and age in abdominal muscles and VAT CSA, age, and triglycerides in thigh muscles are predictors of signal intensities. Conclusions . These results suggest that VAT CSA and age are predictors of intramuscular adipose tissue of the abdominal and thigh in children with obesity

β2 adrenergic agonist suppresses eosinophil-induced epithelial-to-mesenchymal transition of bronchial epithelial cells

application/pdf内容の要旨・審査結果の要旨 / 三重大学大学院医学系研究科 生命医科学専攻 臨床医学系講座 小児科学分

β2 adrenergic agonist suppresses eosinophil-induced epithelial-to-mesenchymal transition of bronchial epithelial cells

application/pdfBackground: Epithelial-mesenchymal transition is currently recognized as an important mechanism for the increased number of myofibroblasts in cancer and fibrotic diseases. We have already reported that epithelial-mesenchymal transition is involved in airway remodeling induced by eosinophils. Procaterol is a selective and full β2 adrenergic agonist that is used as a rescue of asthmatic attack inhaler form and orally as a controller. In this study, we evaluated whether procaterol can suppress epithelial-mesenchymal transition of airway epithelial cells induced by eosinophils.Methods: Epithelial-mesenchymal transition was assessed using a co-culture system of human bronchial epithelial cells and primary human eosinophils or an eosinophilic leukemia cell line.Results: Procaterol significantly inhibited co-culture associated morphological changes of bronchial epithelial cells, decreased the expression of vimentin, and increased the expression of E-cadherin compared to control. Butoxamine, a specific β2-adrenergic antagonist, significantly blocked changes induced by procaterol. In addition, procaterol inhibited the expression of adhesion molecules induced during the interaction between eosinophils and bronchial epithelial cells, suggesting the involvement of adhesion molecules in the process of epithelial-mesenchymal transition. Forskolin, a cyclic adenosine monophosphate-promoting agent, exhibits similar inhibitory activity of procaterol.Conclusions: Overall, these observations support the beneficial effect of procaterol on airway remodeling frequently associated with chronic obstructive pulmonary diseases.本文 / Allergy Center, Mie National Hospital. Department of Immunology, Mie University Graduate School of Medicine11

β2 adrenergic agonist suppresses eosinophil-induced epithelial-to-mesenchymal transition of bronchial epithelial cells

Abstract Background Epithelial-mesenchymal transition is currently recognized as an important mechanism for the increased number of myofibroblasts in cancer and fibrotic diseases. We have already reported that epithelial-mesenchymal transition is involved in airway remodeling induced by eosinophils. Procaterol is a selective and full β2 adrenergic agonist that is used as a rescue of asthmatic attack inhaler form and orally as a controller. In this study, we evaluated whether procaterol can suppress epithelial-mesenchymal transition of airway epithelial cells induced by eosinophils. Methods Epithelial-mesenchymal transition was assessed using a co-culture system of human bronchial epithelial cells and primary human eosinophils or an eosinophilic leukemia cell line. Results Procaterol significantly inhibited co-culture associated morphological changes of bronchial epithelial cells, decreased the expression of vimentin, and increased the expression of E-cadherin compared to control. Butoxamine, a specific β2-adrenergic antagonist, significantly blocked changes induced by procaterol. In addition, procaterol inhibited the expression of adhesion molecules induced during the interaction between eosinophils and bronchial epithelial cells, suggesting the involvement of adhesion molecules in the process of epithelial-mesenchymal transition. Forskolin, a cyclic adenosine monophosphate-promoting agent, exhibits similar inhibitory activity of procaterol. Conclusions Overall, these observations support the beneficial effect of procaterol on airway remodeling frequently associated with chronic obstructive pulmonary diseases