Quantitative proteomics and phosphoproteomics reveal insights into mechanisms of ocnus function in Drosophila testis development.

BACKGROUND: Testis is the only organ supporting sperm production and with the largest number of proteins and tissue-specific proteins in animals. In our previous studies, we have found that knockdown of ocnus (ocn), a testis-specific gene, resulted in much smaller testis with no germ cells in Drosophila melanogaster. However, the molecular consequences of ocn knockdown in fly testes are unknown. RESULTS: In this study, through iTRAQ quantitative proteomics sequencing, 606 proteins were identified from fly abdomens as having a significant and at least a 1.5-fold change in expression after ocn knockdown in fly testes, of which 85 were up-regulated and 521 were down-regulated. Among the differential expressed proteins (DEPs), apart from those proteins involved in spermatogenesis, the others extensively affected biological processes of generation of precursor metabolites and energy, metabolic process, and mitochondrial transport. Protein-protein interaction (PPI) analyses of DEPs showed that several kinases and/or phosphatases interacted with Ocn. Re-analyses of the transcriptome revealed 150 differential expressed genes (DEGs) appeared in the DEPs, and their changing trends in expressions after ocn knockdown were consistent. Many common down-regulated DEGs and DEPs were testis-specific or highly expressed in the testis of D. melanogaster. Quantitative RT-PCR (qRT-PCR) confirmed 12 genes appeared in both DEGs and DEPs were significantly down-regulated after ocn knockdown in fly testes. Furthermore, 153 differentially expressed phosphoproteins (DEPPs), including 72 up-regulated and 94 down-regulated phosphorylated proteins were also identified (13 phosphoproteins appeared in both up- and down-regulated groups due to having multiple phosphorylation sites). In addition to those DEPPs associated with spermatogenesis, the other DEPPs were enriched in actin filament-based process, protein folding, and mesoderm development. Some DEPs and DEPPs were involved in Notch, JAK/STAT, and cell death pathways. CONCLUSIONS: Given the drastic effect of the ocn knockdown on tissue development and testis cells composition, the differences in protein abundance in the ocn knockdown flies might not necessarily be the direct result of differential gene regulation due to the inactivation of ocn. Nevertheless, our results suggest that the expression of ocn is essential for Drosophila testis development and that its down-regulation disturbs key signaling pathways related to cell survival and differentiation. These DEPs and DEPPs identified may provide significant candidate set for future studies on the mechanism of male reproduction of animals, including humans.

Protective effect of vitamin B6 against doxorubicin-induced cardiotoxicity by modulating NHE1 expression.

Doxorubicin (DOX) has been used to treat various types of cancer, but its application is limited due to its heart toxicity as well as other drawbacks. Chronic inhibition of Na+ /H+ exchanger (NHE1) reduces heart failure and reduces the production of reactive oxygen species (ROS); vitamin B6 (VitB6 ) has been demonstrated to have a crucial role in antioxidant mechanism. So, this study was designed to explore the effect of VitB6 supplement on the DOX-induced cardiotoxicity and to imply whether NHE1 is involved. Ultrasonic cardiogram analysis revealed that VitB6 supplement could alleviate DOX-induced cardiotoxicity; hematoxylin and eosin (HE) and Masson's staining further confirmed this effect. Furthermore, VitB6 supplement exhibited significant antioxidative stress and antiapoptosis effect, which was evidenced by decreased serum malondialdehyde (MDA) content and increased serum superoxide dismutase (SOD) content, and decreased Bcl-2-associated X protein/B-cell lymphoma-2 ratio, respectively. Collectively, VitB6 supplement may exert antioxidative and antiapoptosis effects to improve cardiac function by decreasing NHE1 expression and improve DOX-induced cardiotoxicity.

GATA factor-regulated solute carrier ensemble reveals a nucleoside transporter-dependent differentiation mechanism.

Developmental-regulatory networks often include large gene families encoding mechanistically-related proteins like G-protein-coupled receptors, zinc finger transcription factors and solute carrier (SLC) transporters. In principle, a common mechanism may confer expression of multiple members integral to a developmental process, or diverse mechanisms may be deployed. Using genetic complementation and enhancer-mutant systems, we analyzed the 456 member SLC family that establishes the small molecule constitution of cells. This analysis identified SLC gene cohorts regulated by GATA1 and/or GATA2 during erythroid differentiation. As >50 SLC genes shared GATA factor regulation, a common mechanism established multiple members of this family. These genes included Slc29a1 encoding an equilibrative nucleoside transporter (Slc29a1/ENT1) that utilizes adenosine as a preferred substrate. Slc29a1 promoted erythroblast survival and differentiation ex vivo. Targeted ablation of murine Slc29a1 in erythroblasts attenuated erythropoiesis and erythrocyte regeneration in response to acute anemia. Our results reveal a GATA factor-regulated SLC ensemble, with a nucleoside transporter component that promotes erythropoiesis and prevents anemia, and establish a mechanistic link between GATA factor and adenosine mechanisms. We propose that integration of the GATA factor-adenosine circuit with other components of the GATA factor-regulated SLC ensemble establishes the small molecule repertoire required for progenitor cells to efficiently generate erythrocytes.

Risk-adapted stereotactic body radiotherapy for patients with cervical spinal metastases.

Owing to the complex anatomical structure and biomechanics, the current standard palliative treatments for cervical spinal metastases are associated with a high risk of recurrence and complications. Stereotactic body radiotherapy (SBRT) can provide radical dose to tumors while protecting normal organs to the maximum extent. However, the efficacy and safety of SBRT for cervical spinal metastases is not well characterized. Data from 71 patients with cervical spine metastases who were treated with SBRT using CyberKnife between 2006 and 2021 were obtained from our prospectively maintained database. Primary endpoint was pain response at 12 weeks following SBRT completion; secondary endpoints included local control (LC), overall survival (OS), and adverse events. Standard-risk patients were planned to receive 30 Gy (range 21-36) with median fractions of 3 (range 1-3) and high-risk patients 35 Gy (range 24-50) with median fractions of 5 (range 4-5) according to the spinal cord and esophagus dose constraints. The median follow-up time was 17.07 months (range 3.1-118.9). After 12 weeks of SBRT completion, 54 (98.2%) of 55 patients with baseline pain achieved pain response and 46 (83.6%) achieved complete pain response. LC rates were 93.1% and 90% at 1 year and 2 year, respectively. The 1-year and 2-year OS rates were 66.2% and 37.4%, respectively. Eight patients experienced grades 1-4 adverse events (six vertebral compression fracture [VCF], five of them had VCF before SBRT; and two hemiparesis). No grade 5 adverse events were observed. Therefore, risk-adapted SBRT for cervical spine metastases achieved high pain control and LC rates with acceptable adverse events.

Use of an in silico knowledge discovery approach to determine mechanistic studies of silver nanoparticles-induced toxicity from in vitro to in vivo.

BACKGROUND: Silver nanoparticles (AgNPs) are considered a double-edged sword that demonstrates beneficial and harmful effects depending on their dimensions and surface coating types. However, mechanistic understanding of the size- and coating-dependent effects of AgNPs in vitro and in vivo remains elusive. We adopted an in silico decision tree-based knowledge-discovery-in-databases process to prioritize the factors affecting the toxic potential of AgNPs, which included exposure dose, cell type and AgNP type (i.e., size and surface coating), and exposure time. This approach also contributed to effective knowledge integration between cell-based phenomenological observations and in vitro/in vivo mechanistic explorations. RESULTS: The consolidated cell viability assessment results were used to create a tree model for generalizing cytotoxic behavior of the four AgNP types: SCS, LCS, SAS, and LAS. The model ranked the toxicity-related parameters in the following order of importance: exposure dose > cell type > particle size > exposure time ≥ surface coating. Mechanistically, larger AgNPs appeared to provoke greater levels of autophagy in vitro, which occurred during the earlier phase of both subcytotoxic and cytotoxic exposures. Furthermore, apoptosis rather than necrosis majorly accounted for compromised cell survival over the above dosage range. Intriguingly, exposure to non-cytotoxic doses of AgNPs induced G2/M cell cycle arrest and senescence instead. At the organismal level, SCS following a single intraperitoneal injection was found more toxic to BALB/c mice as compared to SAS. Both particles could be deposited in various target organs (e.g., spleen, liver, and kidneys). Morphological observation, along with serum biochemical and histological analyses, indicated that AgNPs could produce pancreatic toxicity, apart from leading to hepatic inflammation. CONCLUSIONS: Our integrated in vitro, in silico, and in vivo study revealed that AgNPs exerted toxicity in dose-, cell/organ type- and particle type-dependent manners. More importantly, a single injection of lethal-dose AgNPs (i.e., SCS and SAS) could incur severe damage to pancreas and raise blood glucose levels at the early phase of exposure.

Comparative Ubiquitome Analysis Reveals Deubiquitinating Effects Induced by Wolbachia Infection in Drosophila melanogaster.

The endosymbiotic Wolbachia bacteria frequently cause cytoplasmic incompatibility (CI) in their insect hosts, where Wolbachia-infected males cross with uninfected females, leading to no or fewer progenies, indicating a paternal modification by Wolbachia. Recent studies have identified a Wolbachia protein, CidB, containing a DUB (deubiquitylating enzyme) domain, which can be loaded into host sperm nuclei and involved in CI, though the DUB activity is not necessary for CI in Drosophila melanogaster. To investigate whether and how Wolbachia affect protein ubiquitination in testes of male hosts and are thus involved in male fertility, we compared the protein and ubiquitinated protein expressions in D. melanogaster testes with and without Wolbachia. A total of 643 differentially expressed proteins (DEPs) and 309 differentially expressed ubiquitinated proteins (DEUPs) were identified to have at least a 1.5-fold change with a p-value of <0.05. Many DEPs were enriched in metabolic pathway, ribosome, RNA transport, and post-translational protein modification pathways. Many DEUPs were involved in metabolism, ribosome, and proteasome pathways. Notably, 98.1% DEUPs were downregulated in the presence of Wolbachia. Four genes coding for DEUPs in ubiquitin proteasome pathways were knocked down, respectively, in Wolbachia-free fly testes. Among them, Rpn6 and Rpn7 knockdown caused male sterility, with no mature sperm in seminal vesicles. These results reveal deubiquitylating effects induced by Wolbachia infection, suggesting that Wolbachia can widely deubiquitinate proteins that have crucial functions in male fertility of their hosts, but are not involved in CI. Our data provide new insights into the regulatory mechanisms of endosymbiont/host interactions and male fertility.

Survival of Tricuspid Valve Replacement in Patients With Previous Tricuspid Valve Surgery.

OBJECTIVES: This study was performed to investigate the short-term and long-term survival of patients who underwent reoperative tricuspid valve replacement (TVR). METHODS: A retrospective analysis was performed of 273 patients who underwent TVRs while hospitalised in Beijing Anzhen Hospital from November 1993 to August 2018. Fifty-six (56) of them underwent reoperative TVR: 36 had previous tricuspid valve repair and 20 had previous TVR. Follow-up was 100% complete, with a mean follow-up of 8 years (range, 1-15 years). RESULTS: The overall in-hospital mortality was 17.9% (n=10). In the univariate analysis, the overall in-hospital mortality and renal failure rate in the replacement group were lower than those in the repair group (5.0% vs 25%; p=0.046 and 27.8% vs 5%; p=0.040). However, in-hospital mortality was no longer statistically significant after multivariate adjustment (adjusted OR 0.318; 95% CI 0.030-3.338; p=0.340). There was no significant difference in survival between the patients with previous repair and those with previous replacement (log-rank test, p=0.839). Factors that correlated with long-term mortality on multivariate analysis were age >60 years (adjusted HR 11.753; 95% CI 1.686-81.915; p=0.013); cardiopulmonary bypass time (adjusted HR 1.019; 95% CI 1.005-1.034; p=0.009); intensive care unit time (adjusted HR 1.024; 95% CI 1.006-1.042; p=0.009); and ventilation time (adjusted HR 0.982; 95% CI 0.965-0.998; p=0.030). CONCLUSIONS: Reoperative TVR was associated with high in-hospital mortality and morbidity. Overall in-hospital mortality was similar between the previous replacement group and the previous repair group. Previous tricuspid valve repair and replacement had similar long-term survival.

[Genetic testing and prenatal diagnosis for thirteen Chinese pedigrees affected with oculocutaneous albinism].

OBJECTIVE: To identify the causative variants in 13 Chinese pedigrees affected with oculocutaneous albinism (OCA) so as to provide genetic counseling and prenatal diagnosis to them. METHODS: Thirteen unrelated pedigrees with clinically diagnosed OCA were collected and classified based on the manifestation of skin and eyes. With informed consent obtained from the participants, peripheral blood samples were collected from the probands and their family members for the extraction of genomic DNA. Candidate variants were screened by targeted capture and next generation sequencing, and the results were validated by Sanger sequencing. Prenatal diagnosis was provided to the families upon their subsequent pregnancies. RESULTS: Causative variants were detected in all probands, including 10 with compound heterozygotes or homozygotes for TYR gene variants and 3 with compound heterozygotes for OCA2 gene variants. Among these, two variants [TYR: c.650G>C (p.Arg217Pro) and OCA2: c.516-2A>T] were unreported previously. The pathogenicity of the novel TYR: c.650G>C (p.Arg217Pro) variant was verified through bioinformatic analysis and prediction of three dimensional structure of the protein. Prenatal diagnosis was provided to 6 fetuses with a high risk for OCA. Four fetuses were found to be carriers, one did not carry the variants of the proband, and one was affected with OCA. CONCLUSION: Identification of the pathogenic variants in the 13 probands, including 2 novel ones, has expanded the mutational spectrum of OCA and enabled genetic counseling and prenatal diagnosis for the families.

Chromosomal analysis for embryos from balanced chromosomal rearrangement carriers using next generation sequencing.

We aimed to use next generation sequencing (NGS) to investigate chromosomal abnormalities in blastocyst trophectoderm (TE) samples, and reproductive outcomes with the different types of chromosomal rearrangements (CR) and for each sex of CR carrier. A total of 1189 blastocyst TE samples were evaluated using NGS to detect chromosomal unbalanced translocations as well as aneuploidy, including blastocytes from 637 blastocysts from carriers of balanced CR and 552 blastocysts from carriers of normal chromosomes. The optimal embryos had lower chromosomal abnormality rates compared to the poor-quality embryos. The experimental group had significantly reduced rates of normal embryos and euploidy, and higher rates of total abnormalities, aneuploidy and unbalanced chromosomal aberrations. Carriers of reciprocal translocations had a reduced rate of normal embryos and an increased percentage of embryos with total abnormalities and unbalanced chromosomal aberrations compared with carriers of Robertsonian translocations. Couples with female carriers of chromosomal abnormalities had significantly reduced rates of normal embryos and euploidy, and a higher percentage of embryos with total abnormalities, aneuploidy, and unbalanced chromosomal aberrations compared with couples of male carriers. Our preimplantation genetic testing (PGT) study identified higher rates of chromosomal abnormalities, including chromosomal unbalanced translocations and aneuploidy, in blastocysts from CR carriers, especially from the female carriers, in a Chinese population. The PGT cycles successfully improved clinical outcomes by increasing the fertilization rate and reducing the early spontaneous abortion rate compared with the in vitro fertilization and intracytoplasmic sperm injection cycles, especially for CR carriers.

[Short-term and long-term outcomes of tricuspid valve replacement in patients with left ventricular dysfunction].

Objective: To examine the short-term and long-term outcomes of tricuspid valve replacement (TVR) in patients with left ventricular dysfunction. Methods: The clinical data of 24 patients with left ventricular dysfunction who received TVR at Department of Cardiovascular Surgery, Beijing Anzhen Hospital, Capital Medical University from November 1993 to August 2018 were consecutively enrolled. There were 14 males and 10 females,aged (41.9±13.2) years old (range: 19 to 66 years old). The preoperative ejection fraction was (42.9±6.4)% (range: 21% to 49%), while less than 35% in 3 patients. The scores of Charlson comorbidity index were as follows: 5 patients for 0, 6 patients for 1, 7 patients for 2, 1 patient for 3 and 5 patients for 4. The European system for cardiac operative risk evaluation (EuroSCORE) Ⅰ was 3.6±2.1 (range: 1 to 9). The EuroSCORE Ⅱ was (4.91±2.40)% (range: 1.58% to 11.60%). The model for end-staged liver disease score was 1.8±1.2 (range: 0.2 to 7.1). The simplified model for end-staged liver disease score was 5.6±2.5 (range: 1.5 to 13.4). Follow-up was conducted by clinic. The long-term survival rate was calculated by Kaplan-Meier survival curve. Results: In-hospital mortality was 16.7% (4/24), including 1 patient for multiple organ failure and 3 patients for low cardiac output syndrome (LCOS). One patient needed continuous renal replacement therapy and 6 patients suffered from LCOS. The follow-up time was 1 to 19 years, with a median of 8 years. During the follow-up period, 4 patients died, including 2 deaths for cardiogenic cause, 1 death for anticoagulant complications, and 1 death for lung cancer. The 1, 5 and 10-year survival rates were 76.2%, 71.4% and 64.9%, respectively. Conclusion: The short-term and long-term clinical outcomes of TVR in patients with left ventricular dysfunction are acceptable, but the mortality and morbidity are still high.

The Effectiveness of Next-Generation Sequencing-Based Preimplantation Genetic Testing for Balanced Translocation Couples.

The purpose of this study was to evaluate the effectiveness of next-generation sequencing (NGS)-based preimplantation genetic testing (PGT) for balanced translocation carriers to identify normal/balanced blastocysts and to measure pregnancy outcomes following euploid embryo transfer. We enrolled 75 couples with a balanced translocation who underwent 83 PGT cycles (58 cycles for carriers with reciprocal translocations and 25 cycles for carriers with Robertsonian translocations) and 388 blastocysts were diagnosed. Moreover, we transferred single euploid blastocysts through frozen embryo transfer and calculated the biochemical pregnancy, clinical pregnancy, miscarriage, and ongoing pregnancy rates per embryo transfer cycle. Despite a mean maternal age of 29.8 years and mean of 4.34 embryos biopsied, there was a 32.8% chance of recording no chromosomally normal/balanced embryos for reciprocal translocation carriers. The proportion of normal/balanced embryos was significantly higher (44.1 vs. 27.8%) in Robertsonian translocation carriers than in reciprocal translocation carriers. Female carriers had a significantly lower (23.3 vs. 42.4%, 34.7 vs. 54.7%, respectively) percentage of normal/balanced embryos than male carriers, regardless of the translocation. After transfering single blastocysts, we obtained a 64.4% clinical pregnancy rate per transfer, and the clinical miscarriage rate was 5.7%. Amniocentesis results showed that all karyotypes of the fetuses were consistent with PGT results. The clinical outcomes are probably not influenced by the type of translocation, maternal age, and blastocyst morphology following the transfer of euploid blastocysts. Therefore, we conclude that NGS-based PGT is an efficient method for analyzing balanced translocation carriers, and aneuploidy screening had good clinical outcomes.

Genotypic and phenotypic characterization of Chinese patients with osteogenesis imperfecta.

Osteogenesis imperfecta (OI) is a rare hereditary skeletal dysplasia, characterized by recurrent fractures and bone deformity. This study presents a clinical characterization and mutation analysis of 668 patients, aiming to establish the mutation spectrum and to elucidate genotype-phenotype correlations in Chinese OI patients. We identified 274 sequence variants (230 in type I collagen encoding genes and 44 in noncollagen genes), including 102 novel variants, in 340 probands with a detection rate of 90%. Compared with 47 loss-of-function variants detected in COL1A1, neither nonsense nor frameshift variants were found in COL1A2 (p < 0.0001). The major cause of autosomal recessive OI was biallelic variants in WNT1 (56%, 20/36). It is noteworthy that three genomic rearrangements, including one gross deletion and one gross duplication in COL1A1 as well as one gross deletion in FKBP10, were detected in this study. Of ten individuals with glycine substitutions that lie towards the N-terminal end of the triple-helical region of the α1(I) chain, none exhibited hearing loss, suggesting a potential genotype-phenotype correlation. The findings in this study expanded the mutation spectrum and identified novel correlations between genotype and phenotype in Chinese OI patients.

How do Wolbachia modify the Drosophila ovary? New evidences support the "titration-restitution" model for the mechanisms of Wolbachia-induced CI.

BACKGROUND: Cytoplasmic incompatibility (CI) is the most common phenotype induced by endosymbiont Wolbachia and results in embryonic lethality when Wolbachia-modified sperm fertilize eggs without Wolbachia. However, eggs carrying the same strain of Wolbachia can rescue this embryonic death, thus producing viable Wolbachia-infected offspring. Hence Wolbachia can be transmitted mainly by hosts' eggs. One of the models explaining CI is "titration-restitution", which hypothesized that Wolbachia titrated-out some factors from the sperm and the Wolbachia in the egg would restitute the factors after fertilization. However, how infected eggs rescue CI and how hosts' eggs ensure the proliferation and transmission of Wolbachia are not well understood. RESULTS: By RNA-seq analyses, we first compared the transcription profiles of Drosophila melanogaster adult ovaries with and without the wMel Wolbachia and identified 149 differentially expressed genes (DEGs), of which 116 genes were upregulated and 33 were downregulated by Wolbachia infection. To confirm the results obtained from RNA-seq and to screen genes potentially associated with reproduction, 15 DEGs were selected for quantitative RT-PCR (qRT-PCR). Thirteen genes showed the same changing trend as RNA-seq analyses. To test whether these genes are associated with CI, we also detected their expression levels in testes. Nine of them exhibited different changing trends in testes from those in ovaries. To investigate how these DEGs were regulated, sRNA sequencing was performed and identified seven microRNAs (miRNAs) that were all upregulated in fly ovaries by Wolbachia infection. Matching of miRNA and mRNA data showed that these seven miRNAs regulated 15 DEGs. Wolbachia-responsive genes in fly ovaries were involved in biological processes including metabolism, transportation, oxidation-reduction, immunity, and development. CONCLUSIONS: Comparisons of mRNA and miRNA data from fly ovaries revealed 149 mRNAs and seven miRNAs that exhibit significant changes in expression due to Wolbachia infection. Notably, most of the DEGs showed variation in opposite directions in ovaries versus testes in the presence of Wolbachia, which generally supports the "titration-restitution" model for CI. Furthermore, genes related to metabolism were upregulated, which may benefit maximum proliferation and transmission of Wolbachia. This provides new insights into the molecular mechanisms of Wolbachia-induced CI and Wolbachia dependence on host ovaries.

Risk-adapted stereotactic body radiation therapy for central and ultra-central early-stage inoperable non-small cell lung cancer.

To determine the therapeutic efficacy and safety of risk-adapted stereotactic body radiation therapy (SBRT) schedules for patients with early-stage central and ultra-central inoperable non-small cell lung cancer. From 2006 to 2015, 80 inoperable T1-2N0M0 NSCLC patients were treated with two median dose levels: 60 Gy in six fractions (range, 48-60 Gy in 4-8 fractions) prescribed to the 74% isodose line (range, 58%-79%) for central lesions (ie within 2 cm of, but not abutting, the proximal bronchial tree; n = 43), and 56 Gy in seven fractions (range, 48-60 Gy in 5-10 fractions) prescribed to the 74% isodose line (range, 60%-80%) for ultra-central lesions (ie abutting the proximal bronchial tree; n = 37) on consecutive days. Primary endpoint was overall survival (OS); secondary endpoints included progression-free survival (PFS), tumor local control rate (LC), and toxicity. Median OS and PFS were 64.47 and 32.10 months (respectively) for ultra-central patients, and not reached for central patients. Median time to local failure, regional failure, and any distant failures for central versus ultra-central lesions were: 27.37 versus 26.07 months, 20.90 versus 12.53 months, and 20.85 versus 15.53 months, respectively, all P < .05. Multivariate analyses showed that tumor categorization (ultra-central) and planning target volume ≥52.76 mL were poor prognostic factors of OS, PFS, and LC, respectively (all P < .05). There was one grade 5 toxicity; all other toxicities were grade 1-2. Our results showed that ultra-central tumors have a poor OS, PFS, and LC compared with central patients because of the use of risk-adapted SBRT schedules that allow for equal and favorable toxicity profiles.

Catalytic Asymmetric Synthesis of Butenolides and Butyrolactones.

γ-Butenolides, γ-butyrolactones, and derivatives, especially in enantiomerically pure form, constitute the structural core of numerous natural products which display an impressive range of biological activities which are important for the development of novel physiological and therapeutic agents. Furthermore, optically active γ-butenolides and γ-butyrolactones serve also as a prominent class of chiral building blocks for the synthesis of diverse biological active compounds and complex molecules. Taking into account the varying biological activity profiles and wide-ranging structural diversity of the optically active γ-butenolide or γ-butyrolactone structure, the development of asymmetric synthetic strategies for assembling such challenging scaffolds has attracted major attention from synthetic chemists in the past decade. This review offers an overview of the different enantioselective synthesis of γ-butenolides and γ-butyrolactones which employ catalytic amounts of metal complexes or organocatalysts, with emphasis focused on the mechanistic issues that account for the observed stereocontrol of the representative reactions, as well as practical applications and synthetic potentials.

Aspirin plus clopidogrel versus aspirin mono-therapy for ischemic stroke: a meta-analysis.

Objectives. Stroke is a common condition after a transient ischemic attack (TIA) or minor ischemic stroke (IS). Adding clopidogrel to aspirin may yield more beneficial outcomes than aspirin mono-therapy; meanwhile, the risk of bleeding in the acute phase remains poorly understood. Therefore, there is increasing emphasis on the risks and benefits of clopidogrel with aspirin compared with aspirin mono-therapy in an effort to treat TIA/IS. Design. We searched several electronic databases, including PubMed, Cochrane, and Embase, to identify eligible randomized controlled trials (RCTs) based on the index words comparing dual-antiplatelet therapy to aspirin mono-therapy for secondary stroke prevention updated to December, 2018. Results. A total of 11 RCTs met our inclusion criteria. The pooled analysis showed that clopidogrel plus aspirin was associated with a trend toward a reduction in recurrent IS (RR = 0.72, 95%CI = 0.65-0.81, p < .001), but not the recurrent stroke rate (RR = 0.81, 95% CI = 0.63-1.03, p = .09) than aspirin mono-therapy. There were differences in bleeding episodes (RR = 1.81, 95%CI = 1.65-1.99, p < .001), moderate-severe major bleeding (RR = 1.64, 95% CI = 1.24-2.16, p = .0005), or mild bleeding (RR = 2.25, 95%CI = 1.54-3.31, p < .001) between the study groups. Meanwhile, no benefit of reducing the risk of intracranial hemorrhage with dual-antiplatelet therapy was found in TIA/IS patients (RR = 1.44, 95% CI = 0.95-2.19, p = .09). Conclusions. The addition of clopidogrel to aspirin for patients with TIA or IS appeared to significantly reduce the risk of IS recurrence with a possible increase in the risk of bleeding compared with aspirin alone.

Novel NTRK1 mutations in Chinese patients with congenital insensitivity to pain with anhidrosis.

Congenital insensitivity to pain with anhidrosis (CIPA) is a rare autosomal recessive disorder, characterized by loss of algesthesis and inability to sweat. CIPA is known to be caused by mutations in the neurotrophic tyrosine kinase receptor type 1 gene ( NTRK1). However, the details of NTRK1 mutations in Chinese CIPA patients remain unclear. In the present study, we recruited 36 CIPA patients from 34 unrelated families in mainland China. Blood samples from these patients and their available familial members were collected and subjected to genetic analysis. We identified 27 mutations in NTRK1 from this cohort, including 15 novel mutations. Interestingly, we discovered two forms of novel recurrent mutations: the first was a large intragenic deletion c.429-374_717 + 485del mediated by recombination between Alu elements, and the second was a deep intronic substitutions c.[851-798C > T;851-794C > G]. All probands were homozygotes or compound heterozygotes of these mutations. Current findings expand our knowledge about the mutation spectrum of NTRK1 in Chinese CIPA patients and provide more evidence for precise diagnosis of the clinically suspected patients with CIPA.

Novel mutations in TPM2 and PIEZO2 are responsible for distal arthrogryposis (DA) 2B and mild DA in two Chinese families.

BACKGROUND: Distal arthrogryposis (DA) is a group of clinically and genetically heterogeneous disorders that involve multiple congenital limb contractures and comprise at least 10 clinical subtypes. Here, we describe our findings in two Chinese families: Family 1 with DA2B (MIM 601680) and Family 2 with mild DA. METHODS: To map the disease locus, two-point linkage analysis was performed with microsatellite markers closed to TPM2, TNNI2/TNNT3 and TNNC2. In Family 1, a positive LOD (logarithm of odds) score was only obtained at the microsatellite marker close to TPM2 and mutation screening was performed using direct sequencing of TPM2 in the proband. In Family 2, for the LOD score that did not favor linkage to any markers, whole-exome sequencing (WES) was performed on the proband. PCR-restriction fragment length polymorphism (RFLP) and bioinformatics analysis were then applied to identify the pathogenic mutations in two families. In order to correlate genotype with phenotype in DA, retrospective analyses of phenotypic features according to the TPM2 and PIEZO2 mutation spectrums were carried out. RESULTS: A heterozygous missense mutation c.308A > G (p.Q103R) in TPM2 in Family 1, and a novel variation c.8153G > A (p.R2718Q) in PIEZO2 in Family 2 were identified. Each of the two novel variants was co-segregated with the DA manifestations in the corresponding family. Bioinformatics analysis from several tools supported the pathogenicity of the mutations. Furthermore, our study suggests that there is no relation between the types or locations of TPM2 mutations and the clinical characteristics, and that different inheritance modes and mutation types concerning PIEZO2 cause distinct clinical manifestations. CONCLUSIONS: We report two novel mutations within TPM2 and PIEZO2 responsible for DA2B and mild DA in two Chinese families, respectively. Our study expands the spectrum of causal mutations in the TPM2 and PIEZO2 genes.

Clinical characteristics and spectrum of NF1 mutations in 12 unrelated Chinese families with neurofibromatosis type 1.

BACKGROUND: Neurofibromatosis type 1 (NF1) is a common autosomal dominant disorder caused by a heterozygous germline mutation in the tumor suppressor gene NF1. Because of the existence of highly homologous pseudogenes, the large size of the gene, and the heterogeneity of mutation types and positions, the detection of variations in NF1 is more difficult than that for an ordinary gene. METHODS: In this study, we collected samples from 23 patients among 46 study participants from 12 unrelated Chinese families with NF1. We used a combination of Sanger sequencing, targeted next-generation sequencing, and multiplex ligation-dependent probe amplification to identify potential mutations of different types. RESULTS: Seven recurrent mutations and four novel mutations were identified with the aforementioned methods, which were subsequently confirmed by either restriction fragment length polymorphism analysis or Sanger sequencing. Truncating mutations accounted for 73% (8/11) of all mutations identified. We also exhaustively investigated the clinical manifestations of NF1 in patients via acquired pathography, photographs and follow-up. However, no clear genotype-phenotype correlation has been found to date. CONCLUSION: In conclusion, the novel mutations identified broaden the spectrum of NF1 mutations in Chinese; however, obvious correlations between genotype and phenotype were not observed in this study.

Identification and characterization of two novel C-type lectins from the larvae of housefly, Musca domestica L.

Lectins and antimicrobial peptides (AMPs) are widely distributed in various insects and play crucial roles in primary host defense against pathogenic microorganisms. Two AMPs (cecropin and attacin) have been identified and characterized in the larvae of housefly. In this study, two novel C-type lectins (CTLs) were obtained from Musca domestica, while their agglutinating and antiviral properties were evaluated. Real-time PCR analysis showed that the mRNA levels of four immune genes (MdCTL1, MdCTL2, Cecropin, and Attacin) from M. domestica were significantly upregulated after injection with killed Gram-negative Escherichia coli. Moreover, purified MdCTL1-2 proteins can agglutinate E. coli and Staphylococcus aureus in the presence of calcium ions, suggesting their immune function is Ca2+ dependent. Sequence analysis indicated that typical WND and QPD motifs were found in the Ca2+ -binding site 2 of carbohydrate recognition domain from MdCTL1-2, which was consistent with their agglutinating activities. Subsequently, antiviral experiments indicated that MdCTL1-2 proteins could significantly reduce the infection rate of Spodoptera frugiperda 9 cells by the baculovirus Autographa californica multicapsid nucleopolyhedrovirus, indicating they might play important roles in insect innate immunity against microbial pathogens. In addition, MdCTL1-2 proteins could effectively inhibit the replication of influenza H1 N1 virus, which was similar to the effect of ribavirin. These results suggested that two novel CTLs could be considered a promising drug candidate for the treatment of influenza. Moreover, it is believed that the discovery of the CTLs with antiviral effects in M. domestica will improve our understanding of the molecular mechanism of insect immune response against viruses.