Genetic heterogeneity of pseudoxanthoma elasticum: the Chinese signature profile of ABCC6 and ENPP1 mutations.

Pseudoxanthoma elasticum (PXE), an autosomal recessive disorder characterized by ectopic mineralization, is caused by mutations in the ABCC6 gene. We examined clinically 29 Chinese PXE patients from unrelated families, so far the largest cohort of Asian PXE patients. In a subset of 22 patients, we sequenced ABCC6 and another candidate gene, ENPP1, and conducted pathogenicity analyses for each variant. We identified a total of 17 distinct mutations in ABCC6, 15 of them being, to our knowledge, previously unreported, including 5 frameshift and 10 missense variants. In addition, a missense mutation in combination with a recurrent nonsense mutation in ENPP1 was discovered in a pediatric PXE case. No cases with p.R1141X or del23-29 mutations, common in Caucasian patient populations, were identified. The 10 missense mutations in ABCC6 were expressed in the mouse liver via hydrodynamic tail-vein injections. One mutant protein showed cytoplasmic accumulation indicating abnormal subcellular trafficking, while the other nine mutants showed correct plasma membrane location. These nine mutations were further investigated for their pathogenicity using a recently developed zebrafish mRNA rescue assay. Minimal rescue of the morpholino-induced phenotype was achieved with eight of the nine mutant human ABCC6 mRNAs tested, implying pathogenicity. This study demonstrates that the Chinese PXE population harbors unique ABCC6 mutations. These genetic data have implications for allele-specific therapy currently being developed for PXE

Investigation of Cytotoxicity of Phosphoryl Choline Modified Single-Walled Carbon Nanotubes under a Live Cell Station

Single-walled carbon nanotubes (SWCNTs) and various modified SWCNTs have drawn a lot of attention due to their potential applications in biomedical field. Before further moving on to real clinical applications, hydrophobicity and toxicity of SWCNTs should be investigated thoroughly. In this paper, 2-methacryloyloxy ethyl phosphorylcholine (MPC) was adopted to modify SWCNTs and phosphoryl choline was grafted onto SWCNTs as small molecule moieties and polymeric chains, which made SWCNTs dispersed stably both in water and in cell culture medium for a long time. Cytotoxicity of pristine and modified SWCNTs were assayed upon successful preparation of the designed modified SWCNT. Furthermore, the internalization of SWCNTs by three cells was investigated using a live cell station under normal culture temperature (37°C) and low temperature (4°C). The results showed that the internalization of modified SWCNTs was related to both the active transport and the passive transport. Although the modification with phosphoryl choline remarkably reduced the cytotoxicity of SWCNTs, the results were probably due to other reasons such as the decrease in the ratio of cells which internalized modified SWCNTs since the cells without SWCNTs occupation still exhibited normal states

Robust Genetic Diagnosis of Split Hand-Foot Malformation by Exome Sequencing

ABSTRACT: Purpose: The present study aimed to evaluate the genetic diagnostic yield and accuracy of exome sequencing in Chinese patients with split hand–foot malformation (SHFM), a severe heterogeneous congenital anomaly characterized by hypodevelopment of the central ray of the hands and feet. Methods: A cohort of seven families and five sporadic patients with SHFM was investigated. Genomic DNA was prepared from the peripheral blood of affected as well as unaffected individuals. Whole exome sequencing (WES) was performed to identify the pathogenic mutations. Array-based comparative genomic hybridization (aCGH), CytoScan, quantitative polymerase chain reaction (qPCR), and Sanger sequencing were performed to validate the findings of WES. WES data of an additional cohort of 24 patients with non-SHFM congenital hand anomalies were analyzed as the control. Results: Pathogenic variants of TP63, c.G956A p.R319H, and c.T602A:p.L201H, were identified in two families by WES. In the remaining patients, copy number analysis of the WES data by XHMM software identified pathogenic 10q 24 duplication in five individuals from three families, which was further validated via CytoScan and qPCR; however, WES could not detect duplication in 10q24 in an additional cohort of 24 individuals with non-SHFM congenital hand anomaly. Importantly, qPCR analysis of the 10q24 region copy number revealed a definite consistency with WES data in all individuals. Genotype–phenotype analysis did not present any unique feature that could differentiate between the families with TP63 mutation and 10q24 duplication. Conclusions: Our study demonstrated that WES is an accurate and sensitive method to detect the pathogenic 10q24 duplication. Collectively, with TP63 mutation, a single WES testing could yield a diagnosis rate of about 40% (5/12) for the SHFM patients, at least in our cohort. As the genotype–phenotype correlation remains unclear, WES could be used as a cost-effective method for the genetic diagnosis of SHFM

Microstructures of YBa1.85Eu0.15Cu3O7-delta superconducting films grown on SrTiO3 and YSZ substrates

A detailed atomic scale microstructure analysis of Eu-doped YBa1.85Eu0.15Cu3O7-delta (YEBCO) thin films with 100 nm in thickness has been carried out by a combination of scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Interesting regular-like arranged network of nanoscale undulations is observed on the surface of YEBCO film on (0 0 1) SrTiO3 substrate. TEM image clearly indicates that the film is always c-axis oriented, but lots of natural precipitates of Y2O3 are involved both at the interface and deep in the film. Desirable size and number density of Y2O3 are thought to be important for acting as efficient flux pinning centers. In the case of YEBCO film on (0 0 1) yttrium stabilized ZrO2 (YSZ) substrate, few cracks and outgrowths appear due to much larger lattice mismatch and dissimilar crystal structure between the film and substrate, but surface quality is still much better compared to the parent YBaCu3O7-delta film. Besides, highly textured BaZrO3 layer at the interface and alpha-axis grains with small dimensions in the film are formed. Interface stability of two kinds of films studied, namely YEBCO/STO and YEBCO/YSZ, is also assessed comprehensively by first principle calculations. (C) 2010 Elsevier B.V. All rights reserved

A potential biomarker hsa-miR-200a-5p distinguishing between benign thyroid tumors with papillary hyperplasia and papillary thyroid carcinoma.

Papillary thyroid carcinoma (PTC) is the most common endocrine cancer with a significantly increase of the incidence recently. Several cytokines, such as thyroid peroxidase (TPO), cluster of differentiation 56 (CD56), Galectin-3, mesothelial cell (MC), cytokeratin 19 (CK19) and BRAF (B-raf) were recommended to be tested by immunohistochemistry (IHC) for a definitive diagnosis, but were still limited in clinical use because of their relative lower sensitivity and specificity. MicroRNA (miRNA), as a new molecular biomarkers, however, has not been reported yet so far. To address this, hsa-miR-200a-5p, a miRNA, was selected and detected in PTC patients by in situ hybrization with benign thyroid tumor with papillary hyperplasia as a control, and the differential expression of hsa-miR-200a-5p between fresh PTC tissues and control was detected by qRT-PCR. Expressive levels of cytokines of TPO, CD56, Galectin-3, MC, CK19 and B-raf were also detected by immunohistochemistry. The correlation was analyzed by SPSS software using Spearman methods. As expected, the hsa-miR-200a-5p expressive level was significantly increased in PTC patients, compared to that of control, and was consistent with that of TPO, CD56, Galectin-3, MC, CK19 and B-raf. In addition, expression of hsa-miR-200a-5p showed negative correlation to that of TPO (rs = - 0.734; **: P < 0.01) and CD56 (rs = - 0.570; **: P < 0.01), but positive correlation to that of Galectin-3 (rs = 0.601; **: P < 0.01), MC (rs = 0.508; **: P < 0.01), CK19 (rs = 0.712; **: P < 0.01) and B-raf (rs = 0.378; **: P < 0.01). PTC and papillary benign thyroid papillary hyperplasia are difficult to distinguish in morphology, so requiring immunohistochemistry to further differentiate the diagnosis, however, for the existing clinical common diagnostic marker for immunohistochemistry, the sensitivity and accuracy are low, it is easy to miss diagnosis. Therefore, there is an urgent need for a rapid and sensitive molecular marker. So miR-200a-5p can be used to assist in the diagnosis of PTC at the molecular level, and as a biomarker, can be effectively used to distinguish between PTC and benign thyroid tumor with papillary hyperplasia

miR-17-92 ameliorates renal ischemia reperfusion injury

There is limited information on the role of miR-17-92 in renal tubular pathophysiology. Therefore, the present study was performed to determine whether miR-17-92 plays a role in ischemia-reperfusion injury (IRI)-induced acute kidney injury. We originally demonstrated that miR-17-92 is up-regulated following IRI in vivo. To explore the roles of miR-17-92 in the IRI process, we first generated a renal proximal tubule-specific miR-17-92 deletion (PT-miR-17-92−/−) knockout mouse model with Cre driven by the Kap promoter. We found that PT-deficient miR-17-92 mice had more severe renal dysfunction and renal structures than their littermates. Compared with sham-operated mice, both wide-type (WT) mice and PT-miR-17-92−/− mice showed increased serum levels of creatinine and urea. However, the levels of serum urea and creatinine in PT-miR-17-92−/− mice after the IRI operation were significantly higher than the levels in WT mice. In addition, PT-miR-17-92−/− mice showed higher levels of serum potassium and phosphonium after the IRI operation. Histological analysis revealed that PT-miR-17-92−/− mice had substantial histopathologic changes, such as tubular dilation and tubular necrosis. Overexpression of miR-17-92 could partially reverse the side-effects of IRI on the proximal tubules in vivo. Furthermore, we employed a quantitative proteomic strategy and identified 16 proteins as potential targets of miR-17-92. Taken together, our findings suggested that miR-17-92 may ameliorates IRI-induced acute kidney injury. Our results indicate that pharmacologic modulation of these miRNAs may have therapeutic potential for acute kidney injury. Keywords: Acute kidney injury, Ischemia reperfusion injury, miR-17-9

In Situ Identification of Unknown Crystals in Acute Kidney Injury Using Raman Spectroscopy

Raman spectroscopy is a well-established and powerful tool for in situ biomolecular evaluation. Type 2 crystal nephropathies are characterized by the deposition of crystalline materials in the tubular lumen, resulting in rapid onset of acute kidney injury without specific symptoms. Timely crystal identification is essential for its diagnosis, mechanism exploration and therapy, but remains challenging. This study aims to develop a Raman spectroscopy-based method to assist pathological diagnosis of type 2 crystal nephropathies. Unknown crystals in renal tissue slides from a victim suffered extensive burn injury were detected by Raman spectroscopy, and the inclusion of crystals was determined by comparing Raman data with established database. Multiple crystals were scanned to verify the reproducibility of crystal in situ. Raman data of 20 random crystals were obtained, and the distribution and uniformity of substances in crystals were investigated by Raman imaging. A mouse model was established to mimic the crystal nephropathy to verify the availability of Raman spectroscopy in frozen biopsy. All crystals on the human slides were identified to be calcium oxalate dihydrate, and the distribution and content of calcium oxalate dihydrate on a single crystal were uneven. Raman spectroscopy was further validated to be available in identification of calcium oxalate dihydrate crystals in the biopsy specimens. Here, a Raman spectroscopy-based method for in situ identification of unknown crystals in both paraffin-embedded tissues and biopsy specimens was established, providing an effective and promising method to analyze unknown crystals in tissues and assist the precise pathological diagnosis in both clinical and forensic medicine

Alteration of resting brain function by genetic variation in angiotensin converting enzyme in amnestic-type mild cognitive impairment of Chinese Han

Using a cross-sectional case–control study of amnestic-mild cognitive impairment (aMCI), we characterised the relationships among cognitive function, serum levels of angiotensin converting enzyme (ACE), brain activity, and ACE insertion or deletion (I/D) polymorphism. Forty-eight patients with aMCI and 36 well-matched normal controls were assessed by a comprehensive battery of standardized neuropsychological tests. In addition, regional homogeneity (ReHo) approaches were used to analyze blood oxygen level-dependent functional magnetic resonance imaging data on the resting state in all subjects, and genotyping of the serum ACE was measured in aMCI patients. The D carriers with aMCI patients were found to have markedly higher serum ACE levels than I homozygote carriers. Importantly, compared with the carried I homozygote group of patients with aMCI, the D carriers of aMCI patients were significantly impaired in the AVLT-delayed recall and had decreased ReHo over the bilateral precuneus, left middle occipital gyrus, right inferior parietal lobe, and right angular gyrus, whilst increased ReHo was found mainly in the left medial frontal gyrus, right paracentral lobe, and right anterior cingulate cortex. The findings indicated that ACE genotype was associated with episodic memory, serum levels of ACE, and resting-state brain activity in aMCI patients, and the findings of cognitive function and brain activity further suggests that the ACE D allele may have a specific role in semantic memory dysfunction and brain activity in aMCI