Keratin 5-Cre-driven deletion of Ncstn in an acne inversa-like mouse model leads to a markedly increased IL-36a and Sprr2 expression / 医学前沿

Familial acne inversa (AI) is an autoinflammatory disorder that affects hair follicles and is caused by loss-of-function mutations in γ-secretase component genes. We and other researchers showed that nicastrin (NCSTN) is the most frequently mutated gene in familial AI. In this study, we generated a keratin 5-Cre-driven epidermis-specific Ncstn conditional knockout mutant in mice. We determined that this mutant recapitulated the major phenotypes of AI, including hyperkeratosis of hair follicles and inflammation. In Ncstn;K5-Cre mice, the IL-36a expression level markedly increased starting from postnatal day 0 (P0), and this increase occurred much earlier than those of TNF-α, IL-23A, IL-1β, and TLR4. RNA-Seq analysis indicated that Sprr2d, a member of the small proline-rich protein 2 family, in the skin tissues of the Ncstn;K5-Cre mice was also upregulated on P0. Quantitative reverse-transcription polymerase chain reaction showed that other Sprr2 genes had a similar expression pattern. Our findings suggested that IL-36a might be a key inflammatory cytokine in the pathophysiology of AI and involved in the malfunction of the skin barrier in the pathogenesis of AI.

Detection of pathogenic mutations in Marfan syndrome by targeted next-generation semiconductor sequencing / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To detect pathogenic mutations in Marfan syndrome (MFS) using an Ion Torrent Personal Genome Machine (PGM) and to validate the result of targeted next-generation semiconductor sequencing for the diagnosis of genetic disorders.</p><p><b>METHODS</b>Peripheral blood samples were collected from three MFS patients and a normal control with informed consent. Genomic DNA was isolated by standard method and then subjected to targeted sequencing using an Ion Ampliseq(TM) Inherited Disease Panel. Three multiplex PCR reactions were carried out to amplify the coding exons of 328 genes including FBN1, TGFBR1 and TGFBR2. DNA fragments from different samples were ligated with barcoded sequencing adaptors. Template preparation and emulsion PCR, and Ion Sphere Particles enrichment were carried out using an Ion One Touch system. The ion sphere particles were sequenced on a 318 chip using the PGM platform. Data from the PGM runs were processed using an Ion Torrent Suite 3.2 software to generate sequence reads. After sequence alignment and extraction of SNPs and indels, all the variants were filtered against dbSNP137. DNA sequences were visualized with an Integrated Genomics Viewer. The most likely disease-causing variants were analyzed by Sanger sequencing.</p><p><b>RESULTS</b>The PGM sequencing has yielded an output of 855.80 Mb, with a > 100 × median sequencing depth and a coverage of > 98% for the targeted regions in all the four samples. After data analysis and database filtering, one known missense mutation (p.E1811K) and two novel premature termination mutations (p.E2264X and p.L871FfsX23) in the FBN1 gene were identified in the three MFS patients. All mutations were verified by conventional Sanger sequencing.</p><p><b>CONCLUSION</b>Pathogenic FBN1 mutations have been identified in all patients with MFS, indicating that the targeted next-generation sequencing on the PGM sequencers can be applied for accurate and high-throughput testing of genetic disorders.</p>

Effects of PGF2? on the glucose-stimulated insulin secretion in NIT-1 beta cells / 中国药理学通报

Aim To investigate the effects of PGF_(2?) upon glucose-stimulated insulin secretion and the calcium response in NIT-1 beta cells.Methods Using the radioimmunoassay(RIA),the amount of PGF_(2?) augmentation of glucose-stimulated insulin secretion was determined in different conditions and the confocal laser scanning methods by Fluo-3AM as a fluorescent probe were used to analyze the NIT-1 beta cell intracellular calcium response in correlated various terms.Results In the presence of 16.5 mmol?L~(-1) glucose,PGF_(2?)(0.1,1,5 ?mol?L~(-1)) dose-dependently augmented glucose-induced insulin secretion in NIT-1 beta cells,especially at 5 ?mol?L~(-1)(P0.05).Meanwhile,Exposure of the NIT-1 cells to 5 ?mol?L~(-1) PGF_(2?) induced a rapid increase of intracellular calcium(P