Here we present the first diploid genome sequence of an Asian individual. The genome was sequenced to 36-fold average coverage using massively parallel sequencing technology. We aligned the short reads onto the NCBI human reference genome to 99.97% coverage, and guided by the reference genome, we used uniquely mapped reads to assemble a high-quality consensus sequence for 92% of the Asian individual's genome. We identified approximately 3 million single-nucleotide polymorphisms (SNPs) inside this region, of which 13.6% were not in the dbSNP database. Genotyping analysis showed that SNP identification had high accuracy and consistency, indicating the high sequence quality of this assembly. We also carried out heterozygote phasing and haplotype prediction against HapMap CHB and JPT haplotypes (Chinese and Japanese, respectively), sequence comparison with the two available individual genomes (J. D. Watson and J. C. Venter), and structural variation identification. These variations were considered for their potential biological impact. Our sequence data and analyses demonstrate the potential usefulness of next-generation sequencing technologies for personal genomics.

BackgroundEndoscopic transsphenoidal surgery has gradually come to be regarded as a preferred option in the treatment of pituitary adenomas because of its advantages of improved visualization and its minimal invasiveness. The aim of this study was to compare and evaluate the outcomes and complications of endoscopic and microscopic transsphenoidal surgery in the treatment of pituitary adenomas.MethodsWe performed a systematic literature search of MEDLINE, EMBASE, the Cochrane Library and the Web of Science between January 1992 and May 2013. Studies with consecutive patients that explicitly and fully compared endoscopic and microscopic approaches in the treatment of pituitary adenomas were included.ResultsA total of 15 studies (n = 1,014 patients) met the inclusion criteria among 487 studies that involved endoscopic surgery and 527 studies that dealt with microscopic surgery. The rate of gross tumor removal was higher in the endoscopic group than in the microscopic group. The post-operative rates of septal perforation were less frequent in patients who underwent endoscopic surgery. There was no significant difference between the two techniques in the incidence rates of meningitis, diabetes insipidus, cerebrospinal fluid leak, epistaxis or hypopituitarism. The post-operative hospital stay was significantly shorter for the endoscopic surgery group compared with the microscopic surgery group (P < 0.05). There was no significant difference in the length of the operation (P > 0.05).ConclusionsThe present study indicates that the endoscopic transsphenoidal approach is safer and more effective than microscopic surgery in the treatment of pituitary adenomas.

Sperm malformation is a direct factor for male infertility. Multiple morphological abnormalities of the flagella (MMAF), a severe form of asthenoteratozoospermia, are characterized by immotile spermatozoa with malformed and/or absent flagella in the ejaculate. Previous studies indicated genetic heterogeneity in MMAF. To further define genetic factors underlying MMAF, we performed whole-exome sequencing in a cohort of 90 Chinese MMAF-affected men. Two cases (2.2%) were identified as carrying bi-allelic missense DNAH8 variants, variants which were either absent or rare in the control human population and were predicted to be deleterious by multiple bioinformatic tools. Re-analysis of exome data from a second cohort of 167 MMAF-affected men from France, Iran, and North Africa permitted the identification of an additional male carrying a DNAH8 homozygous frameshift variant. DNAH8 encodes a dynein axonemal heavy-chain component that is expressed preferentially in the testis. Hematoxylin-eosin staining and electron microscopy analyses of the spermatozoa from men harboring bi-allelic DNAH8 variants showed a highly aberrant morphology and ultrastructure of the sperm flagella. Immunofluorescence assays performed on the spermatozoa from men harboring bi-allelic DNAH8 variants revealed the absent or markedly reduced staining of DNAH8 and its associated protein DNAH17. Dnah8-knockout male mice also presented typical MMAF phenotypes and sterility. Interestingly, intracytoplasmic sperm injections using the spermatozoa from Dnah8-knockout male mice resulted in good pregnancy outcomes. Collectively, our experimental observations from humans and mice demonstrate that DNAH8 is essential for sperm flagellar formation and that bi-allelic deleterious DNAH8 variants lead to male infertility with MMAF.

BackgroundCirculating tumor cells (CTCs) hold great potential in both clinical application and basic research for the managements of cancer. However, it remains to be an enormous challenge to obtain efficient detection of pancreatic CTCs. New detection platforms for the detection of pancreatic CTCs are urgently required.MethodsIn the present study, we applied a newly-developed platform integrated subtraction enrichment and immunostaining-fluorescence in situ hybridization (SE-iFISH) to analyze clinical significance of pancreatic CTCs. Immunostaining of CK, CD45, DAPI and FISH with the centromere of chromosome 8 (CEP8) were utilized to identify CTCs. Cells with features of CK+/CD45-/DAPI+/CEP8 = 2, CK+/CD45-/DAPI+/CEP8 > 2, CK-/CD45-/DAPI+/CEP8 > 2 were defined as pancreatic CTCs. The Kaplan-Meier method and Cox proportional hazards model were used to analyze the relationship of CTC level and other clinicopathological factors with pancreatic cancer clinical outcomes.ResultsCTC count in pancreatic cancer was higher than healthy individuals (median, 3 vs 0 per 7.5 ml; P < 0.001). SE-iFISH platform yielded a sensitivity of 88 % and specificity of 90 % in pancreatic cancer at the cutoff value of 2 cells/7.5 ml. Pancreatic cancer patients with lower CTC count (<3/7.5 ml) had substantially better overall survival (OS) compared with these with higher CTC count (≥3/7.5 ml) (15.2 vs 10.2 months, P = 0.023). Multivariate analysis indicated that higher CTC count was a strong indicator for worse OS (HR = 4.547, P = 0.016).ConclusionOur current data showed that CTCs could be detected in pancreatic cancer patients in various stages, whether localized, locally advanced and metastatic. Besides, CTCs have shown the potential implication in predicting prognosis of pancreatic cancer.

Summary Objective Mutations in SCN8A, a voltage‐gated sodium‐channel type VIII alpha subunit gene, have recently been recognized as one of the pathogenic mechanisms leading to epilepsy and intellectual/developmental disabilities (IDDs). The aim of this study was to detect SCN8A mutations in Chinese patients with epilepsy of unknown etiology and ID/DD. Methods We used targeted next‐generation sequencing to identify SCN8A mutations in Chinese patients with epilepsy of unknown etiology and IDDs. A filter process was performed to prioritize rare variants of potential functional significance. Sanger sequencing confirmed the variants and determined the parental origin. We followed all patients with SCN8A mutations in our cohort and analyzed their clinical data. Results Five de novo SCN8A mutations were identified, including four novel mutations (p.Ala890Thr, p.Leu407Phe, p.Arg850Gln, and p.Ser1596Cys) and one reported (p.Arg1617Gln). Polyphen2 and SIFT software predicted that all five mutations probably damaged Nav1.6 protein function; Mutation Taster indicated that all mutations were disease‐causing. Three of these five patients were controlled well by sodium channel blockers (SCBs). Two of these three patients remained seizure free for 6 and 1.5 months, respectively. One patient had sudden unexpected death in epilepsy (SUDEP) at the age of 1 year and 4 months. Significance Five SCN8A mutations were first reported in Chinese patients with epilepsy and ID/DD, expanding the phenotype and mutation spectrum of SCN8A mutations. Although three of these patients were controlled well by SCBs in our study, the effectiveness of SCBs should be validated in more patients with epilepsy caused by SCN8A mutations in the future. One of our five patients had sudden unexpected death in epilepsy SUDEP, suggesting that we should pay more attention to SUDEP in epileptic patients with SCN8A mutations.

ObjectiveEpilepsy and intellectual/developmental disabilities (ID/DD) have a high rate of co-occurrence. Here, we investigated gene mutations in Chinese children with unexplained epilepsy and ID/DD.MethodsWe used targeted next-generation sequencing to detect mutations within 300 genes related to epilepsy and ID/DD in 253 Chinese children with unexplained epilepsy and ID/DD. A series of filtering criteria was used to find the possible pathogenic variations. Validation and parental origin analyses were performed by Sanger sequencing. We reviewed the phenotypes of patients with each mutated gene.ResultsWe identified 32 novel and 16 reported mutations within 24 genes in 46 patients. The detection rate was 18% (46/253) in the whole group and 26% (17/65) in the early-onset (before three months after birth) epilepsy group. To our knowledge, we are the first to report KCNAB1 is a disease-causing gene of epilepsy by identifying a novel de novo mutation (c.1062dupCA p.Leu355HisfsTer5) within this gene in one patient with early infantile epileptic encephalopathy (EIEE). Patients with an SCN1A mutation accounted for the largest proportion, 17% (8/46). A total of 38% (9/24) of the mutated genes re-occurred at least 2 times and 63% (15/24) occurred only one time. Ion channel genes are the most common (8/24) and genes related to synapse are the next most common to occur (5/24).SignificanceWe have established genetic diagnosis for 46 patients of our cohort. Early-onset epilepsy had the highest detection rate. KCNAB1 mutation was first identified in EIEE patient. We expanded the phenotype and mutation spectrum of the genes we identified. The mutated genes in this cohort are mostly isolated. This suggests that epilepsy and ID/DD phenotypes occur as a consequence of brain dysfunction caused by a highly diverse population of mutated genes. Ion channel genes and genes related to synapse were more common mutated in this patient cohort.

Accumulating evidence indicates the absence of paternally derived miRNAs, piwiRNAs, and proteins may be one important factor contributing to developmental failure in somatic cell cloned embryos. In the present study, we found microRNA-449b (miR-449b) was highly expressed in sperm. Target gene predictions and experimental verification indicate that several embryonic development-related genes, including CDK6, c-MYC, HDAC1 and BCL-2, are targets of miR-449b. We therefore investigated the role of miR-449b using somatic cell nuclear transfer (SCNT) embryo model. Bovine fetal fibroblasts, expressing miR-449b through a doxycycline (dox) induced expression system were used as nuclear donor cells for SCNT. The results showed that miR-449b expression in SCNT embryos significantly enhanced the cleavage rate at 48 h after activation and the levels of H3K9 acetylation at the 2-cell to 8-cell stages, meanwhile, significantly decreased the apoptosis index of blastocysts. In addition, we verified miR-449b could regulate the expression levels of CDK6, c-MYC, HDAC1 and BCL-2. In conclusion, the present study shows that miR-449b expression improves the first cleavage division, epigenetic reprogramming and apoptotic status of bovine preimplantation cloned embryos.

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