Research on Pneumatic Control of a Pressurized Self-Elevating Mat for an Offshore Wind Power Installation Platform.

Efficient deep-water offshore wind power installation platforms with a pressurized self-elevating mat are a new type of equipment used for installing offshore wind turbines. However, the unstable internal pressure of the pressurized self-elevating mat can cause serious harm to the platform. This paper studies the pneumatic control system of the self-elevating mat to improve the precision of its pressure control. According to the pneumatic control system structure of the self-elevating mat, the pneumatic model of the self-elevating mat is established, and a conventional PID controller and fuzzy PID controller are designed and established. It can be seen via Simulink simulation that the fuzzy PID controller has a smaller adjustment time and overshoot, but its anti-interference ability is relatively weak. The membership degree and fuzzy rules of the fuzzy PID controller are optimized using a neural network algorithm, and a fuzzy neural network PID controller based on BP neural network optimization is proposed. The simulation results show that the overshoot of the optimized controller is reduced by 9.71% and the stability time is reduced by 68.9% compared with the fuzzy PID. Finally, the experiment verifies that the fuzzy neural network PID controller has a faster response speed and smaller overshoot, which improves the pressure control accuracy and robustness of the self-elevating mat and provides a scientific basis for the engineering applications of the self-elevating mat.

Prenatal diagnosis and early childhood outcome of fetuses with extremely large nuchal translucency.

OBJECTIVE: To evaluate the prenatal and perinatal outcome of fetuses with extremely large nuchal translucency (eNT) thickness (≥ 6.5 mm). METHODS: 193 (0.61%) singleton fetuses with eNT were retrospectively included. Anomaly scan, echocardiography, and chromosomal and genetic test were included in our antenatal investigation. Postnatal follow-up was offered to all newborns. RESULTS: Major congenital anomalies included congenital heart defect (32.6%, 63/193), hydrops fetalis (13.5%, 26/193), omphalocele (9.3%, 18/193), and skeletal dysplasia (7.8%, 15/193) et al. Abnormal karyotype was identified in 81/115 (70.4%) cases including Turner syndrome (n = 47), Trisomy 18 (n = 17), Trisomy 21 (n = 9), and Trisomy 13 (n = 3). Chromosomal microarray analysis provided informative results with 3.6% (1/28) incremental diagnostic yield over conventional karyotyping. The diagnostic yield of exome sequencing is 10.0% (2/20). There was no significant increase [Odds Ratio (OR) = 1.974; 95% confidence interval 0.863-4.516; P = 0.104] in the incidence of chromosomal defects despite the presence of other structural anomalies. Only 13 fetuses were successfully followed up and survived at term, no one was found with developmental delay or mental retardation. CONCLUSIONS: Extremely large NT has a high risk of chromosomal abnormality. CMA and ES improve chromosomal genomic and genetic diagnosis of fetal increased NT. When cytogenetic analysis and morphology assessment are both normal, the outcome is good.

Generation of NANOS3-mCherry reporter human embryonic stem cell line SYSUe-009-A using CRISPR/Cas9.

NANOS3 is a zinc-finger containing RNA-binding protein that has been demonstrated to be highly expressed in human primordial germ cell(hPGC), thus NANOS3 can serve as a marker for hPGC development. Due to the ethical and technical restrictions, it is difficult to get primary human germline cells. Human primordial germ cell-like cells (hPGCLCs) generated from pluripotent stem cells is an excellent alternatives in human germ cell-related studies. This hESC line with an mCherry knock-in at the site before NANOS3's stop codon serves as a useful tool to learn human PGC specification.

Conflicting Roles of ZFP36L1 in Regulating the Progression of Muscle Invasive Bladder Cancer.

As the most common carcinoma of the human urinary system, bladder cancer (BC) is characterized by high recurrence, and poor prognosis after metastasis. In the past decade, genome-wide expression and sequencing studies had identified key genes and pathways related to BC, and pictured the comprehensive molecular features of the disease. Our previous study indicated that the coding gene of zinc finger protein 36 like 1 (ZFP36L1) mutated frequently in bladder tumor tissues and may be a potential suppressor for BC. The present study aimed to further investigate the role of ZFP36L1 in BC, and the survival analysis based on TCGA dataset revealed that high expressing level of ZFP36L1 associated with poorer prognosis of the patients with muscle invasive bladder cancer (MIBC). The associations of ZFP36L1 expression to the clinicopathological and molecular biological features also implicated the high level of ZFP36L1 may related to worse outcomes of patients. Also, GSEA indicated that high expression of ZFP36L1 significantly associated with enhanced activity of cancer metastasis related pathways. Functions of ZFP36L1 in MIBC were investigated further, and it was found that while ZFP36L1 suppressed the self-renewal of bladder cancer cells, it promoted the invasiveness of the cells markedly. Taken together, these results led to the conflicting roles of ZFP36L1 in regulating the progression of MIBC, and revealed further researches are needed to clarify the functions of the gene in tumor initiation and recurrence.

Collagen-targeted tumor-specific transepithelial penetration enhancer mediated intravesical chemoimmunotherapy for non-muscle-invasive bladder cancer.

Intravesical instillation of chemotherapeutics or immune-stimulating agents could reduce the recurrence rate of non-muscle-invasive bladder cancer (NMIBC) after transurethral resection of the bladder tumors. Its efficacy, however, remains to be improved due to the bladder epithelial barrier. Although certain transmucosal delivery carriers are able to enhance the transepithelial penetration of intravesical agents, they could hardly differentiate carcinoma and adjacent normal tissues of the bladder wall. Here, we reported polyethylene glycol (PEG) & glutaraldehyde co-modified fluorinated chitosan (PGFCS) as a collagen-targeted transepithelial penetration enhancer, which could create a tumor-targeted adhesive interface by the aldehyde-selective reaction with collagen amines enriched in the tumor, thus opening the transepithelial-delivery barrier at the tumor site though the fluorinated-chitosan-mediated tight junction regulation. Interestingly, with the help of PGFCS pre-treatment, intravesical instillation of chemotherapeutics pirabucin (THP) combined with immune stimulating agent interleukin-12 could trigger potent antitumor chemoimmunotherapeutic responses in destructing orthotopic bladder tumors and inhibiting cancer recurrence. Our work presents a unique type of tumor-specific transepithelial penetration enhancer, which shows great potential for safe and effective intravesical instillation of NMIBC.

The oncogenic role of HIF-1α/miR-182-5p/ZFP36L1 signaling pathway in nasopharyngeal carcinoma.

BACKGROUND: Accumulating evidence indicates that dysregulation of miR-182-5p can serve as diagnostic and prognostic biomarkers for some cancers, whereas the role of miR-182-5p has not been explored in nasopharyngeal carcinoma (NPC). Our study aims to elucidate the biological function of miR-182-5p in NPC and the potential molecular mechanism involved. METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to determine miR-182-5p expression in NPC primary tissues and cell lines. Immunohistochemistry (IHC) for ZFP36L1 was conducted in NPC samples. Western blot was used to evaluate protein expression in cell lines. A series of functional assays were carried out to evaluate the roles of miR-182-5p and ZFP36L1 in tumor development and progression of NPC. Bioinformatics tools and luciferase reporter assays were utilized to identify the potential mechanisms of action. Moreover, rescue experiments were applied to explore whether ZFP36L1 mediated the effects of miR-182-5p in NPC. RESULTS: Up-regulation of miR-182-5p was significantly associated with tumor development and poor prognosis in patients with NPC. Functional study demonstrated that miR-182-5p overexpression enhanced, whereas suppression of miR-182-5p impeded NPC cell proliferation, migration, tumorigenesis and metastasis. Mechanistically, miR-182-5p interacted with ZFP36L1 at two sites in its 3' un-translated region (UTR) and repressed ZFP36L1 expression in NPC. Consistently, an inverse correlation was observed between the expression levels of miR-182-5p and ZFP36L1 using clinical NPC tissues, and down-regulation of ZFP36L1 in NPC predicts poor survival. Furthermore, overexpression of miR-182-5p in NPC was partly attributable to the transcriptional activation effect induced by hypoxia-inducible factor 1α (HIF-1α). CONCLUSIONS: Our data suggests that miR-182-5p facilitates cell proliferation and migration in NPC through its ability to down-regulate ZFP36L1 expression, and that the HIF-1α/miR-182-5p/ZFP36L1 axis may serve as a novel therapeutic target in the management of NPC.

Characterization of the Genitourinary Microbiome of 1,165 Middle-Aged and Elderly Healthy Individuals.

Accumulated evidence shows that complex microbial communities resides in the healthy human urinary tract and can change in urological disorders. However, there lacks a comprehensive profiling of the genitourinary microbiota in healthy cohort. Here, we performed 16S rRNA gene sequencing of midstream urine specimens from 1,172 middle-aged and elderly healthy individuals. The core microbiota included 6 dominant genera (mean relative abundance >5%), including Prevotella, Streptococcus, Lactobacillus, Gardnerella, Escherichia-Shigella, and Veillonella, and 131 low-abundance genera (0.01-5%), displaying a distinct microbiome profiles to that of host-matched gut microbiota. The composition and diversity of genitourinary microbiome (GM) were distinct between genders and may fluctuate with ages. Several urotypes were identified by the stratification of microbiome profiles, which were mainly dominated by the six most predominant genera. The prevalence of urotypes was disparate between genders, and the male sample additionally harbored other urotypes dominated by Acinetobacter, Corynebacterium, Staphylococcus, or Sphingomonas. Peptoniphilus, Ezakiella, and Porphyromonas were co-occurred and co-abundant, and they may play crucial roles as keystone genera and be associated with increased microbial diversity. Our results delineated the microbial structure and diversity landscape of the GM in healthy middle-aged and elderly adults and provided insights into the influence of gender and age to it.

Generation of GPAM knockout human embryonic stem cell line SYSUe-008-A using CRISPR/Cas9.

GPAM (glycerol-3-phosphateacyltransferase1) is a mitochondrial enzyme that catalyze an essential step in glycerolphospholipids and triacylglycerol biosynthesis process. Loss-of-function mutation of GPAM has been shown to lead to hypomyelination of corticospinal tract in cerebral palsy patient. To model this rare disease with human brain organoid, we generated a GPAM knockout human embryonic stem cell line SYSUe-008-A by CRISPR/cas9. The GPAM knockout cell line maintains a normal karyotype and shows comparable level of pluripotent stem cell marker expression and differentiation potential as wild-type human embryonic stem cells.

Generation of an induced pluripotent stem cell line SYSUi-004-A from a child of microcephaly with TYW1 mutations.

We generated an induced human pluripotent stem cell line from a child with microcephaly carrying compound heterozygous mutations of TYW1 inherited from healthy parents. This iPS cell line showed typical embryonic stem cell-like morphology, expressed pluripotent markers that comparable to human embryonic stem cells. Moreover, these cells have the ability to differentiate into three germ layers and maintain a normal karyotype.

Cancer stem cell-specific expression profiles reveal emerging bladder cancer biomarkers and identify circRNA_103809 as an important regulator in bladder cancer.

Bladder cancer stem cells (BCSCs), exhibiting self-renewal and differentiation capacities, may contribute to the tumor initiation, metastasis, recurrence and drug resistance of bladder cancer. However, the underlying functional mechanisms of BCSCs remain to be clarified. In this study, we describe the differentially-expressed mRNAs, lncRNAs, and circRNAs in BCSCs compared with that in bladder cancer non-stem cells (BCNSCs) through the transcriptome microarray data analysis using bladder cancer patients' specimens. CircRNA_103809, the top one among the highly expressed circRNA identified in BCSCs, promotes the self-renewal, migration and invasion capabilities of bladder cancer by acting as a miR-511 sponge. Additionally, GO and KEGG pathway analysis suggest the differentially expressed genes identified may be involved in the cellular metabolism, differentiation and metastasis regulation of the cancer cells. Co-expression networks of lncRNAs/mRNAs and circRNAs/mRNAs constructed by WGCNA give a picture of the non-coding/coding RNAs regulating patterns in BCSCs. Notably, as core genes in the networks, AHCY, C6orf136 and LRIG1 show high potential to be prognosticators for bladder cancer. Therefore, further studies of non-coding RNA functional mechanisms in BCSCs is valuable for detecting the pathogenic mechanisms and discovering novel biomarkers in bladder cancer.

Roles of an N-terminal coiled-coil-containing domain in the localization and function of Bem3, a Rho GTPase-activating protein in budding yeast.

GTPase-activating proteins (GAPs) play critical roles in the spatial and temporal control of small GTPases. The budding yeast Bem3 is a GAP for Cdc42, a Rho GTPase crucial for actin and septin organization. Bem3 localizes to the sites of polarized growth. However, the amino acid sequence determinants mediating recruitment of Bem3 to its physiological sites of action and those important for Bem3 function are not clear. Here, we show that Bem3's localization is guided by two distinct targeting regions-the PX-PH-domain-containing TD1 and the coiled-coil-containing TD2. TD2 localization is largely mediated by its interaction with the polarisome component Epo1 via heterotypic coiled-coil interaction. This finding reveals a novel role for the polarisome in linking Bem3 to its functional target, Cdc42. We also show that the coiled-coil domain of Bem3 interacts homotypically and this interaction is important for the regulation of Cdc42 by Bem3. Moreover, we show that overexpression of a longer version of the TD2 domain disrupts septin-ring assembly in a RhoGAP-independent manner, suggesting that TD2 may be capable of interacting with proteins implicated in septin-ring assembly. Furthermore, we show that the longer version of TD2 interacts with Kss1, a MAPK involved in filamentous growth. Kss1 is reported to localize mainly in the nucleus. We find that Kss1 also localizes to the sites of polarized growth and Bem3 interacts with Kss1 at the septin-ring assembly site. Our study provides new insights in Bem3's localization and function.

Kin2, the Budding Yeast Ortholog of Animal MARK/PAR-1 Kinases, Localizes to the Sites of Polarized Growth and May Regulate Septin Organization and the Cell Wall.

MARK/PAR-1 protein kinases play important roles in cell polarization in animals. Kin1 and Kin2 are a pair of MARK/PAR-1 orthologs in the budding yeast Saccharomyces cerevisiae. They participate in the regulation of secretion and ER stress response. However, neither the subcellular localization of these two kinases nor whether they may have other cellular functions is clear. Here, we show that Kin2 localizes to the sites of polarized growth in addition to localization on the plasma membrane. The localization to polarity sites is mediated by two targeting domains-TD1 and TD2. TD1 locates in the N-terminal region that spans the protein kinase domain whereas TD2 locates in the C-terminal end that covers the KA1 domain. We also show that an excess of Kin2 activity impaired growth, septin organization, and chitin deposition in the cell wall. Both TD1 and TD2 contribute to this function. Moreover, we find that the C-terminal region of Kin2 interacts with Cdc11, a septin subunit, and Pea2, a component of the polarisome that is known to play a role in septin organization. These findings suggest that Kin2 may play a role in the regulation of the septin cytoskeleton and the cell wall. Finally, we show that the C-terminal region of Kin2 interacts with Rho3, a Rho GTPase, whereas the N-terminal region of Kin2 interacts with Bmh1, a 14-3-3 protein. We speculate that Kin2 may be regulated by Bmh1, Rho3, or Pea2 in vivo. Our study provides new insight in the localization, function, and regulation of Kin2.

The GTPase-activating protein Rga1 interacts with Rho3 GTPase and may regulate its function in polarized growth in budding yeast.

In budding yeast, Rga1 negatively regulates the Rho GTPase Cdc42 by acting as a GTPase-activating protein (GAP) for Cdc42. To gain insight into the function and regulation of Rga1, we overexpressed Rga1 and an N-terminally truncated Rga1-C538 (a.a. 538-1007) segment. Overexpression of Rga1-C538 but not full-length Rga1 severely impaired growth and cell morphology in wild-type cells. We show that Rga1 is phosphorylated during the cell cycle. The lack of phenotype for full-length Rga1 upon overexpression may result from a negative regulation by G1-specific Pho85, a cyclin-dependent kinase (CDK). From a high-copy suppressor screen, we isolated RHO3, SEC9, SEC1, SSO1, SSO2, and SRO7, genes involved in exocytosis, as suppressors of the growth defect caused by Rga1-C538 overexpression. Moreover, we detected that Rga1 interacts with Rho3 in two-hybrid and bimolecular fluorescence complementation (BiFC) assays. Rga1 preferentially interacts with the GTP-bound form of Rho3 and the interaction requires the GAP domain and additional sequence upstream of the GAP domain. Our data suggest that the interaction of Rga1 with Rho3 may regulate Rho3's function in polarized bud growth.

[Application of fluorescence in situ hybridization to prenatal diagnosis of aneuploidy in 110 uncultured amniotic fluid samples].

OBJECTIVE: To optimize the prenatal diagnosis platform by using domestically made fluorescence in situ hybridization(FISH) kit and to explore the clinical application of FISH to rapid prenatal diagnosis of a wide range of chromosomal abnormalities. METHODS: Amniotic fluid samples from 110 pregnant women were studied with the rapid prenatal diagnosis method of FISH and the conventional cell culture method of karyotyping, the results from both methods were compared. RESULTS: Four cases of trisomy 21, 1 case of trisomy 18, 58 cases of 46, XX, and 47 cases of 46, XY were detected by FISH in the 110 amniotic fluid samples. It is concordant with the results from conventional karyotype analysis. The concordance rate is 100%. CONCLUSION: Domestically made FISH kit can be used to rapidly and accurately detect the most common chromosome aneuploidies by using less sample volume while the price is relatively low. FISH can be a reliable and rapid prenatal diagnostic tool as an adjunct to classical cytogenetic study. It can be used for rapid and accurate prenatal diagnosis of women with high risk of maternal serum screening.

[Application of array-based comparative genomic hybridization in precise diagnosis of unbalanced chromosome aberration].

OBJECTIVE: To evaluate the method of array-based comparative genomic hybridization (array-CGH) in identifying unbalanced chromosome aberrations. METHODS: Four cases that could not be diagnosed by conventional cytogenetic technique were selected to undergo array-CGH analysis. DNA samples were extracted and hybridized with the Affymetrix SNP 6.0 arrays using Human Mapping SNP6.0 assay kit following the manufacturer's standard protocol. The data were analyzed by two professional software packages, GCOS and Genotyping Console. RESULTS: By using array-CGH technique, all the four cases were diagnosed precisely through identifying two duplications and two complex derivative chromosomes. CONCLUSION: Array-CGH is an effective method for whole-genome identification of unbalanced chromosomal aberrations with high sensitivity and specificity. It has a great value to investigate the correlations between genotype and phenotype in clinical service, especially in prenatal diagnosis.

[Application of spectral karyotyping in diagnosis and prenatal diagnosis of the marker chromosome].

OBJECTIVE: To determine the value of spectral karyotyping(SKY) in identification of the marker chromosome. METHODS: Selected six cases that could not be identified in clinic were studied, using samples of peripheral blood from four cases, and samples of amonic fluid and fetal cord blood for prenatal diagnosis in two cases were investigated. All cases were analyzed with the routine SKY method, and the results with the SKY View software. The SKY results were identified by using fluorescence in situ hybridization (FISH). And C-banding technique was used to help diagnose the heterochromatin. RESULTS: SKY was successfully performed on all of 6 cases. The origin of all marker chromosomes was identified by SKY. Except case No. 4, the others were confirmed by FISH. It helped determine the pregnancy outcome in two cases of prenatal diagnosis: one case of genetic marker chromosome continued the pregnancy, and another case of de novo marker chromosome was terminated of the pregnancy. CONCLUSION: SKY may be a valuable tool to diagnose the marker chromosome with rapidness,direct-viewing and sensitiveness. It can be used to assess the prognosis and the pregnancy outcome.

[Application of spectral karyotyping in diagnosis of complex chromosome aberration].

OBJECTIVE: To determine the value of spectral karyotyping (SKY) to identify the complex chromosome aberration. METHODS: Four cases were selected that can not be identified by standard cytogenetic techniques. The chromosome specimens were detected by the routine SKY method, and the results were analyzed by the SKY View software. RESULTS: By using SKY a case of complex chromosome rearrangements and two cases of chromosome duplication were identified. However it could not identify the chromosome inversion and the breakpoint of chromosome aberration. CONCLUSION: SKY may be a valuable tool in identification of complex chromosome translocation, rearrangement, minute aberration and unknown derivative chromosomes. Though SKY can not replace the standard cytogenetic techniques, but it will be the benefit supplementary.

[HLA-B gene polymorphism detected by high-resolution sequence-based typing in Guangdong Han populations].

OBJECTIVE: To investigate the genetic polymorphism of HLA-B locus in Guangdong Han population and compare the characteristic of the allele frequency distribution with that in other populations. METHODS: A total of 562 cord blood samples from Guangzhou Cord Blood Bank were analyzed by sequence-based typing. Then the sequences encompassing exons 2, 3, and 4 for HLA-B gene were analyzed by direct sequencing of PCR products. The allele frequency distribution of HLA-B in this population was compared with that in other populations. RESULTS: A total of 59 different HLA-B alleles were detected, and among them were 6 HLA-B alleles with frequencies higher than 5%: HLA-B*4601 (14.5%), HLA-B*400101 (14.4%), HLA-B*1502 (11.5%), HLA-B*1301 (8.6%), HLA-B*5801 (8.1%) and HLA-B*380201 (6.4%); the total frequency of these six alleles was 63.5%. At the same time, there were 30 kinds of HLA-B allele with frequencies lower than 0.5%; the total frequency of these alleles was 4.9%. Maximum variation at HLA-B was seen in the HLA-B*15 allele family (nine alleles). Comparison of the HLA-B frequencies in different populations showed a close relationship of Guandong Han population with the Chinese populations from Hong Kong and Singapore, respectively. CONCLUSION: The results have shown the characteristic of HLA-B distribution and provided more accurate genotypic data that may serve as normal reference value for the Han population in Guangdong, China.