αA-crystallin R49Cneo mutation influences the architecture of lens fiber cell membranes and causes posterior and nuclear cataracts in mice

<p>Abstract</p> <p>Background</p> <p>αA-crystallin (CRYAA/HSPB4), a major component of all vertebrate eye lenses, is a small heat shock protein responsible for maintaining lens transparency. The R49C mutation in the αA-crystallin protein is linked with non-syndromic, hereditary human cataracts in a four-generation Caucasian family.</p> <p>Methods</p> <p>This study describes a mouse cataract model generated by insertion of a neomycin-resistant (neo^r) gene into an intron of the gene encoding mutant R49C αA-crystallin. Mice carrying the neo^rgene and wild-type <it>Cryaa </it>were also generated as controls. Heterozygous knock-in mice containing one wild type gene and one mutated gene for αA-crystallin (WT/R49C^neo) and homozygous knock-in mice containing two mutated genes (R49C^neo/R49C^neo) were compared.</p> <p>Results</p> <p>By 3 weeks, WT/R49C^neomice exhibited large vacuoles in the cortical region 100 μm from the lens surface, and by 3 months posterior and nuclear cataracts had developed. WT/R49C^neomice demonstrated severe posterior cataracts at 9 months of age, with considerable posterior nuclear migration evident in histological sections. R49C^neo/R49C^neomice demonstrated nearly complete lens opacities by 5 months of age. In contrast, R49C mice in which the neo^rgene was deleted by breeding with CreEIIa mice developed lens abnormalities at birth, suggesting that the neo^rgene may suppress expression of mutant R49C αA-crystallin protein.</p> <p>Conclusion</p> <p>It is apparent that modification of membrane and cell-cell interactions occurs in the presence of the αA-crystallin mutation and rapidly leads to lens cell pathology <it>in vivo</it>.</p

Determinants of the urinary and serum metabolome in children from six European populations

Background Environment and diet in early life can affect development and health throughout the life course. Metabolic phenotyping of urine and serum represents a complementary systems-wide approach to elucidate environment–health interactions. However, large-scale metabolome studies in children combining analyses of these biological fluids are lacking. Here, we sought to characterise the major determinants of the child metabolome and to define metabolite associations with age, sex, BMI and dietary habits in European children, by exploiting a unique biobank established as part of the Human Early-Life Exposome project (http://www.projecthelix.eu). Methods Metabolic phenotypes of matched urine and serum samples from 1192 children (aged 6–11) recruited from birth cohorts in six European countries were measured using high-throughput 1H nuclear magnetic resonance (NMR) spectroscopy and a targeted LC-MS/MS metabolomic assay (Biocrates AbsoluteIDQ p180 kit). Results We identified both urinary and serum creatinine to be positively associated with age. Metabolic associations to BMI z-score included a novel association with urinary 4-deoxyerythronic acid in addition to valine, serum carnitine, short-chain acylcarnitines (C3, C5), glutamate, BCAAs, lysophosphatidylcholines (lysoPC a C14:0, lysoPC a C16:1, lysoPC a C18:1, lysoPC a C18:2) and sphingolipids (SM C16:0, SM C16:1, SM C18:1). Dietary-metabolite associations included urinary creatine and serum phosphatidylcholines (4) with meat intake, serum phosphatidylcholines (12) with fish, urinary hippurate with vegetables, and urinary proline betaine and hippurate with fruit intake. Population-specific variance (age, sex, BMI, ethnicity, dietary and country of origin) was better captured in the serum than in the urine profile; these factors explained a median of 9.0% variance amongst serum metabolites versus a median of 5.1% amongst urinary metabolites. Metabolic pathway correlations were identified, and concentrations of corresponding metabolites were significantly correlated (r > 0.18) between urine and serum. Conclusions We have established a pan-European reference metabolome for urine and serum of healthy children and gathered critical resources not previously available for future investigations into the influence of the metabolome on child health. The six European cohort populations studied share common metabolic associations with age, sex, BMI z-score and main dietary habits. Furthermore, we have identified a novel metabolic association between threonine catabolism and BMI of children

Discovery of a novel anticancer agent with both anti-topoisomerase I and II activities in hepatocellular carcinoma SK-Hep-1 cells in vitro and in vivo: Cinnamomum verum component 2-methoxycinnamaldehyde

Daw-Shyong Perng,1 Yu-Hsin Tsai,2 Jonathan Cherng,3 Jeng-Shing Wang,4 Kuo-Shen Chou,5 Chia-Wen Shih,6 Jaw-Ming Cherng7 1Department of Gastroenterology, E-Da Hospital, I-Shou University, Kaohsiung, Taiwan, Republic of China; 2Sierra College, Rocklin, CA, USA; 3Faculty of Medicine, Medical University of Lublin, Lublin, Poland; 4Department of Internal Medicine, Antai Tian-Sheng Memorial Hospital, Pingtung, 5Department of Family Medicine, Saint Mary&rsquo;s Hospital Luodong, 6Department of Pathology, Lotung Poh-Ai Hospital, 7Department of Internal Medicine, Saint Mary&rsquo;s Hospital Luodong, Yilan, Taiwan, Republic of China Abstract: Cinnamomum verum is used to make the spice cinnamon and has been used as a traditional Chinese herbal medicine for various applications. We evaluated the anticancer effect of 2-methoxycinnamaldehyde (2-MCA), a constituent of the bark of the plant, and its underlying molecular biomarkers associated with carcinogenesis in human hepatocellular carcinoma SK-Hep-1 cell line. The results show that 2-MCA suppressed proliferation and induced apoptosis as indicated by mitochondrial membrane potential loss, activation of caspase-3 and caspase-9, increase in the DNA content in sub-G1, and morphological characteristics of apoptosis, including blebbing of plasma membrane, nuclear condensation, fragmentation, apoptotic body formation, and long comet tail. In addition, 2-MCA also induced lysosomal vacuolation with increased volume of acidic compartments, suppressions of nuclear transcription factors NF-&kappa;B, cyclooxygenase-2, prostaglandin E2 (PGE2), and both topoisomerase I and II activities in a dose-dependent manner. Further study reveals the growth-inhibitory effect of 2-MCA was also evident in a nude mice model. Taken together, the data suggest that the growth-inhibitory effect of 2-MCA against SK-Hep-1 cells is accompanied by downregulations of NF-&kappa;B-binding activity, inflammatory responses involving cyclooxygenase-2 and PGE2, and proliferative control involving apoptosis, both topoisomerase I and II activities, together with an upregulation of lysosomal vacuolation and volume of acidic compartments. Similar effects (including all of the above-mentioned effects) were found in other tested cell lines, including human hepatocellular carcinoma Hep 3B, lung adenocarcinoma A549, squamous cell carcinoma NCI-H520, colorectal adenocarcinoma COLO 205, and T-lymphoblastic MOLT-3 (results not shown). Our data &shy;suggest that 2-MCA could be a potential agent for anticancer therapy. Keywords: 2-methoxycinnamaldehyde, anticancer, SK-Hep-1 cells, topoisomerase I, topoisomerase I