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Microdeletions and microduplications of the 16p11.2 chromosomal locus are associated with syndromic neurodevelopmental disorders and reciprocal physiological conditions such as macro/microcephaly and high/low body mass index. To facilitate cellular and molecular investigations into these phenotypes, 65 clones of human induced pluripotent stem cells (hiPSCs) were generated from 13 individuals with 16p11.2 copy number variations (CNVs). To ensure these cell lines were suitable for downstream mechanistic investigations, a customizable bioinformatic strategy for the detection of random integration and expression of reprogramming vectors was developed and leveraged towards identifying a subset of 'footprint'-free hiPSC clones. Transcriptomic profiling of cortical neural progenitor cells derived from these hiPSCs identified alterations in gene expression patterns which precede morphological abnormalities reported at later neurodevelopmental stages. Interpreting clinical information--available with the cell lines by request from the Simons Foundation Autism Research Initiative--with this transcriptional data revealed disruptions in gene programs related to both nervous system function and cellular metabolism. As demonstrated by these analyses, this publicly available resource has the potential to serve as a powerful medium for probing the etiology of developmental disorders associated with 16p11.2 CNVs. [ABSTRACT FROM AUTHOR]

Objective: To express CD14 protein and prepare anti-CD14 monoclonal antibody. Method: The CD14 encoding sequence was cloned from human peripheral blood lymphocytes, which was connected to the pRSETC vector, the expression plasmid pRSETC/CD14 was transfected into BL21 (DE3). Expression of recombinant protein was induced by using IPTG. SDS-PAGE and western-blot were used to detected the purified recombinant protein. Hybridoma cells stably secreting anti-CD14 monoclonal antibodies were prepared by using hybridoma technology, the activity of purified antibody was identified by Western blot. Results: The coding sequence of CD14 was obtained. SDS-PAGE showed that recombinant protein expressed in the forms of inclusion body. And moreover, SDS-PAGE showed the purification rate of the purified product was higher than 95%. The positive hybridoma clone which secreted the mAb against CD14 protein were obtained, high purity monoclonal antibody which had the binding activity to CD14 protein was prepared. Conclusion: D14 protein and anti-CD14 monoclonal antibody were successfully prepared, which provided the experimental basis for the development of CD14 detection technology. [ABSTRACT FROM AUTHOR]

Imitating manipulation skills through observing human demonstrations in everyday life is promising in allowing service robots to be programed quickly, as well as to perform human-like behaviors. Such a Learning by demonstration (LbD) problem is challenging because robots are expected to adapt their learned behaviors to the changes of task parameters and the environment, rather than simply cloning the human teacher's motion. In this paper, we propose a hierarchical and parameterized LbD framework that combines symbolic and trajectory learning of pick-and-place manipulation tasks. We have extended the two-step parameterized learning method with error compensation for learning Environment-adaptive Action Primitives (EaAPs), which is capable of adapting robot's reproduced trajectories to new task instances as well as environmental changes. To arrive at refined plans in situations of under-specified human demonstrations, we propose to model the semantics of demonstrated activities with PDDL-based skill scripts. Therefore, latent motion primitives that are impossible to be learned directly from observing human demonstration in noisy video data can be inferred. The proposed method is implemented as a hierarchical LbD framework and has been evaluated on real robot hardware to illustrate the effectiveness of the proposed approach. [ABSTRACT FROM AUTHOR]

Kazuo Ishiguro’s 2005 novel Never Let Me Go presents an alternative history in which artificial reproduction is viable and human clones are mass produced. Even though they differ from other human beings only in their inception, the clones are perceived as artificial human beings, born and raised to be professional nonhumans at the service of natural humans. This article focuses on the novel’s depictions of the alterations that humans will undergo as a result of radical technological advancement. The analysis will concentrate on Ishiguro’s employment of the school as chronotope (in the sense of Bakhtin’s conceptualization). School as a concept is a quintessential humanist institution, the purpose of which is to develop the individual’s subjectivity to its full capacity. Hailsham School, however, represents the new discursive order of posthumanism that reveres technology but never considers the full extent of the ethical issues accompanying the new techniques. At Hailsham, the most advanced humanist discourse on human rights is ironically metamorphosed into a reactionary discourse of exploitation and exclusion. [ABSTRACT FROM AUTHOR]

Research in science, technology, engineering, and mathematics (STEM) fields has become much more complex in the twenty-first century. As a result, the students of our Graduate School, who are all Ph.D. candidates, need to be trained in essential skills and processes that are crucial for success in academia and beyond. Some research problems are inherently complex in that they raise deep moral dilemmas, such as antimicrobial resistance, sustainability, dual-use research of concern (defined as well-intentioned scientific research that may be misused for nefarious purposes), and human cloning. Dealing with moral dilemmas is one of several core competencies that twenty-first-century Ph.D. students must acquire. However, this might prove difficult for STEM Ph.D. students who have had limited exposure to moral philosophy. Since the task of dealing with moral dilemmas in STEM research requires input from both scientific and philosophical disciplines, it is argued with the help of the 4 examples above that this task be explicitly modelled as an interdisciplinary process. Furthermore, it is argued that a particular model from the interdisciplinary education literature could serve as a learning tool to support ethical decision-making in research ethics and integrity courses for doctoral students. [ABSTRACT FROM AUTHOR]

Copyright of Revista Iberoamericana de Ciencia, Tecnologia y Sociedad is the property of Centro de Estudios sobre Ciencia, Desarrollo y Educacion Superior and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Background: Universal gene targets are in persistent demand by real‐time quantitative polymerase chain reaction (RT‐qPCR)‐based methods in acute leukemia (AL) diagnosis and monitoring. Human Krüppel‐like factor 3 (hKLF3), a newly cloned human transcription factor, has proved to be a regulator of hematopoiesis. Methods: Sanger sequencing was performed in bone marrow (BM) samples from 17 AL patients for mutations in hKLF3 coding exons. hKLF3 expression in peripheral blood (PB) and BM samples from 45 AL patients was dynamically detected by RT‐qPCR. PB samples from 31 healthy donors were tested as normal controls. Results: No mutation was sequenced in hKLF3 coding exons. hKLF3 expression in PB of AL was significantly lower than that in healthy donors [0.30 (0.02‐1.07) vs 1.18 (0.62‐3.37), P <.0001]. Primary acute myeloid leukemia (AML) exhibited the least expression values compared with secondary AML and acute lymphoblastic leukemia. Receiver operating characteristic (ROC) analyses suggested that hKLF3 expression in PB was a good marker for AML diagnosis with an AUC of 0.99 (95% CI 0.98‐1.00) and an optimum cutoff value of 0.67 (sensitivity 93.94% and specificity 93.55%). hKLF3 expression was upregulated significantly when AML patients acquired morphological complete remission (CR), and the level of hKLF3 seemed to be higher in patients with deeper CR than in patients with minimal residual disease (MRD). Paired PB and BM samples showed highly consistent alteration in hKLF3 expression (r =.6533, P =.001). Besides, a significantly converse correlation between decreased hKLF3 expression in PB and markers for leukemic load was observed. Conclusions: hKLF3 expression in PB may act as a potential marker for AL diagnosis and monitoring. [ABSTRACT FROM AUTHOR]

Insulin has captured researchers' attention worldwide. There is a rapid global rise in the number of diabetic patients, which increases the demand for insulin. Current methods of insulin production are expensive and time-consuming. A PCR-based strategy was employed for the cloning and verification of human insulin. The human insulin protein was then overexpressed in E. coli on a laboratory scale. Thereafter, optimisation of human insulin expression was conducted. The yield of human insulin produced was approximately 520.92 (mg/L), located in the intracellular fraction. Human insulin was detected using the MALDI-TOF-MS and LC–MS methods. The crude biosynthesised protein sequence was verified using protein sequencing, which had a 100% similarity to the human insulin sequence. The biological activity of human insulin was tested in vitro using a MTT assay, which revealed that the crude biosynthesised human insulin displayed a similar degree of efficacy to the standard human insulin. This study eliminated the use of affinity tags since an untagged pET21b expression vector was employed. Tedious protein renaturation, inclusion body recovery steps, and the expensive enzymatic cleavage of the C-peptide of insulin were eliminated, thereby making this method of biosynthesising human insulin a novel and more efficient method. [ABSTRACT FROM AUTHOR]

Copyright of Al-Adab / Al-ādāb is the property of Republic of Iraq Ministry of Higher Education & Scientific Research (MOHESR) and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Background. In the field of periodontal guided tissue regeneration, microperforated membranes have recently proved to be very promising periodontal regenerative tissue engineering tools. Regenerative periodontal approaches, employing gingival mesenchymal stem/progenitor cells in combination with these novel membranes, would occur mostly in inflamed microenvironmental conditions intraorally. This in turn entails the investigation into how inflammation would affect the proliferation as well as the migration dynamics of gingival mesenchymal stem/progenitor cells. Materials and Methods. Clones of human gingival mesenchymal stem/progenitor cells (GMSCs) from inflamed gingival tissues were characterized for stem/progenitor cells' characteristics and compared to clones of healthy human GMSCs (n = 3), to be subsequently seeded on perforated collagen-coated poly-tetra-floro-ethylene (PTFE) membranes with a pore size 0.4 and 3 microns and polycarbonic acid membranes of 8 microns pore size in Transwell systems. The population doubling time and the MTT test of both populations were determined. Fetal bovine serum (FBS) was used as a chemoattractant in the culturing systems, and both groups were compared to their negative controls without FBS. Following 24 hours of incubation period, migrating cells were determined on the undersurface of microperforated membranes and the membrane-seeded cells were examined by scanning electron microscopy. Results. GMSCs demonstrated all predefined stem/progenitor cell characteristics. GMSCs from inflamed gingival tissues showed significantly shorter population doubling times. GMSCs of inflamed and healthy tissues did not show significant differences in their migration abilities towards the chemoattractant, with no cellular migration observed in the absence of FBS. GMSCs from healthy gingival tissue migrated significantly better through larger micropores (8 microns). Scanning electron microscopic images proved the migratory activity of the cells through the membrane pores. Conclusions. Inflammation appears to boost the proliferative abilities of GMSCs. In terms of migration through membrane pores, GMSCs from healthy as well as inflamed gingival tissues do not demonstrate a difference in their migration abilities through smaller pore sizes, whereas GMSCs from healthy gingival tissues appear to migrate significantly better through larger micropores. [ABSTRACT FROM AUTHOR]

Next-generation sequencing (NGS) enables the sequencing of thousands of genes, the exomes and even entire genomes in single experiments for $2000 or less. Genome sequencing reveals not only the exome but also the regulatory elements of transcription/translation. RNA sequencing determines which genes and spliced transcripts are expressed. We have been studying the blood group ABO genes, A and B glycosyltransferases (AT/BT), and A/B oligosaccharide antigens since we cloned human AT cDNAs in 1990. We have applied diverse scientific disciplines including genetics, biochemistry, enzymology and glycobiology to those studies and demonstrated the central dogma of ABO: A/B alleles at the ABO genetic locus encode AT/BT, which synthesize A/B antigens, respectively. We identified 4 amino acid substitutions between AT and BT and inactivating mutations in O alleles, elucidating the allelic basis of ABO. We became the first who succeeded in genotyping the ABO locus, discriminating AA/AO genotypes, as well as BB/BO, which was impossible by the immunological approach. We have also demonstrated mutations in several subgroup alleles and cis-AB and B(A) alleles. Taking advantage of big data recently generated on genome sequences and associated information deposited in public databases, we expanded our research from humans to other organisms, as well as from ATs/BTs and ABO genes to evolutionarily related α1,3-Gal(NAc) transferases and their genes. Combining phylogenetic analyses with functional assays we developed, we have successfully scaled up our research to the genomic level, deciphering the molecular mechanisms responsible for important biological phenomena associated with ABO. [ABSTRACT FROM AUTHOR]

Background: In Southeast Asia, people are often coinfected with different species of malaria (Plasmodium falciparum [Pf] and Plasmodium vivax [Pv]) as well as with multiple clones of the same species. Whether particular species or clones within mixed infections are more readily transmitted to mosquitoes remains unknown.Methods: Laboratory-reared Anopheles dirus were fed on blood from 119 Pf-infected Cambodian adults, with 5950 dissected to evaluate for transmitted infection. Among 12 persons who infected mosquitoes, polymerase chain reaction and amplicon deep sequencing were used to track species and clone-specific transmission to mosquitoes.Results: Seven of 12 persons that infected mosquitoes harbored mixed Pf/Pv infection. Among these 7 persons, all transmitted Pv with 2 transmitting both Pf and Pv, leading to Pf/Pv coinfection in 21% of infected mosquitoes. Up to 4 clones of each species were detected within persons. Shifts in clone frequency were detected during transmission. However, in general, all parasite clones in humans were transmitted to mosquitoes, with individual mosquitoes frequently carrying multiple transmitted clones.Conclusions: Malaria diversity in human hosts was maintained in the parasite populations recovered from mosquitoes fed on their blood. However, in persons with mixed Pf/Pv malaria, Pv appears to be transmitted more readily, in association with more prevalent patent gametocytemia. [ABSTRACT FROM AUTHOR]

• Easy-to-use approaches are lacking for site-directed modification of adenovector (Ad). • Construct an intermediate plasmid for site-directed mutagenesis by DNA assembly. • Restore modified intermediate plasmid to adenoviral plasmid by restriction-ligation. • Combined DNA assembly and restriction-ligation can be used for Ad modification. Commonly used and well accepted approaches are lacking for site-directed modification of adenoviral vectors. Here, we attempt to introduce an easy-to-implement strategy for such purpose with an example of establishing a replication competent adenoviral vector system from pKAd5 plasmid, an infectious clone of human adenovirus 5 (HAdV-5). PCR products of GFP expression cassette and plasmid backbone were fused with the EcoRI/NdeI-digested fragment of pKAd5 to generate a modified intermediate plasmid pMDXE3GA by DNA assembly. NdeI-digested fragment of pMDXE3GA was brought back to pKAd5 to form the adenoviral plasmid pKAd5XE3GA by restriction-ligation cloning. Recombinant adenovirus HAdV5-XE3GA was rescued, amplified and purified. The expression of GFP and the propagation of virus in adherent HEp-2 and suspension K562 cells were investigated. Expression of target gene was significantly enhanced in both cell lines infected with HAdV5-XE3GA due to virus replication. However, propagation of virus could not sustain in culture of K562 cells. Shuttle plasmid pSh5RC-GFP was constructed to facilitate exchange of transgene. In summary, the strategy of combined DNA assembly and restriction-ligation cloning is functional, cost-effective and suitable for genetic modification of adenovirus. [ABSTRACT FROM AUTHOR]

Objectives: Increasing numbers of linezolid-resistant Enterococcus carrying optrA are being reported across different niches worldwide. We aimed to characterize the first optrA-carrying Enterococcus faecalis obtained from food-producing animals and retail meat samples in Tunisia.Methods: Seven optrA-carrying E. faecalis obtained from chicken faeces (n=3, August 2017) and retail chicken meat (n=4, August 2017) in Tunisia were analysed. Antimicrobial susceptibility was determined by disc diffusion, broth microdilution and Etest against 13 antibiotics, linezolid and tedizolid, respectively (EUCAST/CLSI). optrA stability (∼600 bacterial generations), transfer (filter mating) and location (S1-PFGE/hybridization) were characterized. WGS (Illumina-HiSeq) was done for four representatives that were analysed through in silico and genomic mapping tools.Results: Four MDR clones carrying different virulence genes were identified in chicken faeces (ST476) and retail meat (the same ST476 clone plus ST21 and ST859) samples. MICs of linezolid and tedizolid were stably maintained at 8 and 1-2 mg/L, respectively. optrA was located in the same transferable chromosomal Tn6674-like element in ST476 and ST21 clones, similar to isolates from pigs in Malaysia and humans in China. ST859 carried a non-conjugative plasmid of ∼40 kb with an impB-fexA-optrA segment, similar to plasmids from pigs and humans in China.Conclusions: The same chromosomal and transferable Tn6674-like element was identified in different E. faecalis clones from humans and animals. The finding of retail meat contaminated with the same linezolid-resistant E. faecalis strain obtained from a food-producing animal highlights the potential role of the food chain in the worrisome dissemination of optrA that can be stably maintained without selective pressure over generations. [ABSTRACT FROM AUTHOR]

With the development of new technologies, more and more issues have arisen the legal framework and ethics must respond to. From an ethical perspective, codes of ethics have been formulated that prohibit the use of new technologies in order to create hybrid individuals or to create human clones of people who exist or have existed. In terms of international law, we will highlight a few countries that have laws and articles of law that are of interest for this work, but we will also highlight the importance of observing the human rights. [ABSTRACT FROM AUTHOR]

Extended-spectrum β-lactamase (ESBL)-producing Enterobacterales are a major public health problem, and wastewater from municipal wastewater treatment plants (WWTPs) is a potential means of spreading them into the environment and community. Our objective was to isolate ESBL-producing E. coli and other Enterobacterales from wastewater after treatment at Croatia's largest WWTP and to characterize these isolates by phenotypic and genotypic testing. Of the 200 bacterial isolates, 140 were confirmed as Enterobacterales by MALDI-TOF MS, with Escherichia coli and Klebsiella spp. predominating (69% and 7%, respectively). All 140 enterobacterial isolates were multidrug-resistant (MDR) and produced ESBLs. The most prevalent ESBL genes among the isolates tested were bla CTX-M-15 (60%), bla TEM-116 (44%), and bla CTX-M-3 (13%). Most isolates (94%) carried more than one ESBL gene in addition to bla CTX-M. Genes encoding plasmid-mediated AmpC, most notably bla EBC , were detected in 22% of isolates, whereas genes encoding carbapenemases (bla OXA-48 , bla NDM-1 , bla VIM-1) were less represented (10%). In E. coli , 9 different sequence types (ST) were found, with the emerging high-risk clones ST361 (serotype A-O9:H30) and pandemic ST131 (serotype B2–O25:H4) predominating (32% and 15%, respectively). Other high-risk E. coli clones included ST405 (3%), ST410 (3%), CC10 (3%), ST10 (3%), and ST38 (2%), and emerging clones included ST1193 (2%) and ST635 (2%). Whole-genome sequencing of three representative E. coli from two dominant clone groups (ST361 and ST131) and one extensively drug-resistant K. oxytoca revealed the presence of multiple plasmids and resistance genes to several other antibiotic classes, as well as association of the bla CTX-M-15 gene with transposons and insertion sequences. Our findings indicate that treated municipal wastewater contributes to the spread of emerging and pandemic MDR E. coli clones and other enterobacterial strains of clinical importance into the aquatic environment, with the risk of reintroduction into humans. [Display omitted] • Treated wastewater harbored MDR Enterobacterales isolates of clinical significance. • Most E. coli isolates belonged to high-risk clones related to human infections. • Bla CTX-M-15 was the most frequently found ESBL gene, followed by bla TEM-116. • Some ESBL-producing Klebsiella spp. harbored OXA-48 or NDM-1 carbapenemase genes. • Some ESBL-producing E. cloacae cplx. harbored GES-5 and VIM-1 carbapenemase genes. [ABSTRACT FROM AUTHOR]

[Display omitted] • Brca1 is required for development. • Mouse models help inform protein functions. • Mouse models can be used to discern therapeutic vulnerabilities. • DNA repair pathways are re-wired in a Brca1 mutation-dependent manner. Brca1 mouse models were first reported in the mid-1990′s shortly after cloning the human gene. Since then, many mouse models with a range of mutations have been generated, some mimic patient mutations, others are designed to probe specific protein domains and functions. In this review, we discuss early and recent studies using engineered Brca1 mouse alleles, and their implications for understanding Brca1 protein function in the context of DNA repair, tumorigenesis, and anti-cancer therapeutics. [ABSTRACT FROM AUTHOR]

Human adipose stem cells can differentiate into various mesodermic lineages, including adipogenic, osteogenic, chondrogenic, myogenic and endothelial pathways. In addition, these cells types possess immunomodulatory properties, potentially useful for autoimmune and autoinflammatory diseases. However, single-cell expanded clones have shown that the cells can present a variety of differentiation potential, which may be partly due to epigenetic differences among them. The objective of this study was to assess if DNA methylation plays a role in the differentiation potential observed between different cell clones obtained from the same donor. To this end, the methylation profile of five clonal cell lines of human adipose stem cells obtained by liposuction from two donors was analyzed. Previous reports demonstrated that cell lines 1.7 and 1.22 from Donor 1 and 3.5 from Donor 3 were adipogenic-osteogenic, but not cell lines 1.10 and 3.10. The genes analyzed were neuronal, endothelial, myogenic, osteogenic, adipogenic, extracellular matrix, cell cycle, cytoskeleton and metabolic enzymes. All clones analyzed in this study displayed a similar pattern of methylation in most of the gene families: 85.5% were hypomethylated genes and 14.5% hypermethylated. In conclusion, the methylation pattern of the 1113 genes studied in this report was not a consistent tool to identify the differentiation potential of human adipose stem cells. [ABSTRACT FROM AUTHOR]

• Porcine IL-17B (poIL-17B) and porcine IL-17E (poIL-17E) were cloned and characterized. • The significant enhanced expression of PR-39 and pBD-1 was observed when poIL-17B/E were overexpressed in IPEC-J2. • poIL-17B/E could inhibit effectively pathogenic microorganism, while inhibitory capability of poIL-17B was stronger than that of poIL-17E. The interleukin-17 (IL-17) family plays a critical role in host defense, allergic reactions, and even tumorigenesis on different mucous membranes. IL-17 family has been cloned in human and mouse, as well IL-17A, IL-17 F in swine. So far, current knowledge on the cloning and biological functions of porcine IL-17B (poIL-17B) and porcine IL-17E (poIL-17E) is limited. In this study, poIL-17B and poIL-17E, mainly expressed in intestine, were cloned and characterized. Expression of poIL-17B and poIL-17E was upregulated after pathogenic microorganism infection. Moreover, the significant enhanced expression of antibacterial peptides PR-39 and pBD-1 was observed when poIL-17B and poIL-17E were over-expressed in the small intestinal epithelial cell line IPEC-J2. This demonstrated that poIL-17B and poIL-17E might have anti-infective capability. Pathogens infection data showed that pathogens could up-regulate poIL-17B/E expression levels. After stimulating the cells with the pathogen, continued with probiotics, the expression of poIL-17B/E was down-regulated. Meanwhile, the induced expression of poIL-17E was greater than that of poIL-17B. Invasion data indicated that poIL-17B and poIL-17E both could inhibit effectively pathogenic microorganism, while inhibitory capability of poIL-17B was stronger than that of poIL-17E. Therefore, poIL-17B and poIL-17E both could be important members against intestinal infection in the porcine IL-17 family. This study provided a theoretical basis for the prevention of intestinal diseases in pigs and thus achieved healthy farming. [ABSTRACT FROM AUTHOR]

This paper gives an introduction to the interdisciplinary special section. Against the historical and ethical background of reproductive technologies, it explores future scenarios of human reproduction and analyzes ways of mutual engagement between fictional and academic endeavors. The underlying idea is that we can make use of human reproduction scenarios in at least two ways: we can use them to critique technologies by imagining terrible consequences for humanity but also to defend positions that favor scientific and technological development. [ABSTRACT FROM AUTHOR]

This essay examines the fantasy of life extension enabled through the transfer of one's consciousness to new, cloned bodies in the event of disease, accident, or old age. This vision has recently been dramatized in both fiction and film, bearing witness to the power of this imaginary scenario. This eventuality would raise wide-ranging ethical issues, which speculative bioethics should begin to contemplate. Interestingly, it is young adult fiction that has recently provided an extensive and consistent cluster of novels dealing not only with this topic but also with the interrelated notion of clones purposely grown to replace a loved one, with a concomitant array of further ethical concerns. These and related topics will be examined here through the lens of fiction and recent theoretical work on future biotechnological scenarios. [ABSTRACT FROM AUTHOR]

Background : The unique phenomenon of endothelial antibody capture (endocapt) leads to site-specific accumulation of antibodies on the endothelium after its locoregional injection. The potential of this phenomenon has already been demonstrated in animal models. In the present study, the translational potential of several human endothelium-specific antibodies for their use in the endocapt-based approach was analysed. Methods : The binding of different endothelium-surface specific monoclonal antibody clones was analysed in human tissue and in endothelial cells using image-based immunofluorescence and the determination of half-maximal effective concentration (EC 50). The potential of endocapt-based locoregional application of antibodies or antibody-coated liposomes was analysed ex vivo using isolated mouse liver perfusion and in vivo using superselective injection in tumour models. Results: Eight out of ten antibody clones were assigned to the group of "fast binding antibodies". Different antibody clones showed various binding kinetics to the same endothelial marker whereas the binding kinetics of single antibody clones was independent from the tissue type. Anti-CD49e, anti-CD31, anti-CD34 and anti-CD102 antibodies showed the lowest EC 50 of antibody binding concentration and constant results in EC 50 determination of antibody binding to cells and human tissue. Experimental studies using anti-mouse CD49e antibody and coated immunoliposomes confirmed their effective capture by endothelial cells in vitro and in vivo by which fluorescent liver segment labelling was achieved. Conclusions: Our findings identify the high potential of several human antibody clones, especially anti-CD49e, -CD31, -CD34 and -CD102, for endocapt technology. We also propose important translational implications of these antibodies for image-guided liver surgery and for use of nanoparticles/immunoliposomes. Toxicological studies are indispensable for further translational development of new antibodies for endocapt. The phenomenon of endothelial antibody capture (endocapt) leads to site-specific accumulation of antibodies on the endothelium after its locoregional injection. This phenomenon broadly prevents systemic circulation of the antibody or antibody-drug conjugates. In the present study, our findings identify several human antibody clones promising for endothelial capture technology. This study provided the experimental demonstration of liver segment labelling ex vivo using isolated mouse liver perfusion and in vivo using superselective injection in tumor models. In addition, this study proposed the important translational implications of selected antibodies for image-guided liver surgery and for use of nanoparticles/immunoliposomes. [ABSTRACT FROM AUTHOR]

A method of stance detection in text is proposed. This method is based on the machine learning of ensembles of classifiers. It is known that ensembles have advantages over individual classifiers, which often improves the quality of classification. An important issue is determining the classifiers that should be included in such an ensemble. The method of constructing ensembles proposed in this paper, which is based on a cross-validation procedure, makes it possible to optimize the parameters of the base classifiers, evaluate the effectiveness of each combination of classifiers included in the set, and select the optimal combination. For testing the proposed method, corpora of Russian language messages in Internet forums and the social network VKontakte have been formed. These messages concern three socially significant issues—vaccination of children, Unified State Exam, and human cloning. The experimental study shows the advantage of the proposed method over other classifiers. [ABSTRACT FROM AUTHOR]

Objectives: Antimicrobial resistance (AMR) to critically important antimicrobials (CIAs) amongst Gram-negative bacteria can feasibly be transferred amongst wildlife, humans and domestic animals. This study investigated the ecology, epidemiology and origins of CIA-resistant Escherichia coli carried by Australian silver gulls (Chroicocephalus novaehollandiae), a gregarious avian wildlife species that is a common inhabitant of coastal areas with high levels of human contact.Methods: Sampling locations were widely dispersed around the perimeter of the Australian continent, with sites separated by up to 3500 km. WGS was used to study the diversity and molecular characteristics of resistant isolates to ascertain their epidemiological origin.Results: Investigation of 562 faecal samples revealed widespread occurrence of extended-spectrum cephalosporin-resistant (21.7%) and fluoroquinolone-resistant (23.8%) E. coli. Genome sequencing revealed that CIA-resistant E. coli isolates (n = 284) from gulls predominantly belonged to human-associated extra-intestinal pathogenic E. coli (ExPEC) clones, including ST131 (17%), ST10 (8%), ST1193 (6%), ST69 (5%) and ST38 (4%). Genomic analysis revealed that gulls carry pandemic ExPEC-ST131 clades (O25:H4 H30-R and H30-Rx) and globally emerging fluoroquinolone-resistant ST1193 identified among humans worldwide. Comparative analysis revealed that ST131 and ST1193 isolates from gulls overlapped extensively with human clinical isolates from Australia and overseas. The present study also detected single isolates of carbapenem-resistant E. coli (ST410-blaOXA-48) and colistin-resistant E. coli (ST345-mcr-1).Conclusions: The carriage of diverse CIA-resistant E. coli clones that strongly resemble pathogenic clones from humans suggests that gulls can act as ecological sponges indiscriminately accumulating and disseminating CIA-resistant bacteria over vast distances. [ABSTRACT FROM AUTHOR]

• Genetic heterogeneity in human T-ALL cell lines contributes to large differences in cell death responses, and could explain failure of induction therapy. • Single cell T-ALL clones do not revert to parental phenotype, or resemble the parental phenotype of a population of genetically-diverse T-ALL cells. Genetic modification of human leukemic cell lines using CRISPR-Cas9 has become a staple of gene-function studies. Single-cell cloning of modified cells is frequently used to facilitate studies of gene function. Inherent in this approach is an assumption that the genetic drift, amplified in some cell lines by mutations in DNA replication and repair machinery, as well as non-genetic factors will not introduce significant levels of experimental cellular heterogeneity in clones derived from parental populations. In this study, we characterize the variation in cell death of fifty clonal cell lines generated from human Jurkat and MOLT-4 T-cells edited by CRISPR-Cas9. We demonstrate a wide distribution of sensitivity to chemotherapeutics between non-edited clonal human leukemia T-cell lines, and also following CRISPR-Cas9 editing at the NLRP1 locus, or following transfection with non-targeting sgRNA controls. The cell death sensitivity profile of clonal cell lines was consistent across experiments and failed to revert to the non-clonal parental phenotype. Whole genome sequencing of two clonal cell lines edited by CRISPR-Cas9 revealed unique and shared genetic variants, which had minimal read support in the non-clonal parental population and were not suspected CRISPR-Cas9 off-target effects. These variants included genes related to cell death and drug metabolism. The variation in cell death phenotype of clonal populations of human T-cell lines may be a consequence of T-cell line genetic instability, and to a lesser extent clonal heterogeneity in the parental population or CRISPR-Cas9 off-target effects not predicted by current models. This work highlights the importance of genetic variation between clonal T-cell lines in the design, conduct, and analysis of experiments to investigate gene function after single-cell cloning. [ABSTRACT FROM AUTHOR]

Purpose: Although MAGEC2 was first cloned from a human hepatocellular carcinoma (HCC) cDNA library by serum screening, the detailed attributes of MAGEC2 in HCC have rarely been elucidated. Patients and methods: In this study, The Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO) databases were consulted to analyse the expression of MAGEC2 mRNA in liver cancer. Immunohistochemistry (IHC) analysis was performed to detect MAGEC2 expression in HCC, and the relationship between MAGEC2 expression and the clinicopathological characteristics of HCC patients was evaluated. Then, we employed the short hairpin (sh)RNA-mediated knockdown of MAGEC2 in HCC cell lines to explore the function of MAGEC2 in HCC development. Finally, the expression of epithelial-mesenchymal transition (EMT) markers in HCC xenografts and clinical samples was investigated. Results: The results showed a remarkably higher level of MAGEC2 expression in HCC tissues than in noncancerous tissues, and MAGEC2 expression could be used as an independent prognostic factor for overall survival in HCC. Moreover, sh-MAGEC2 inhibited a series of HCC malignant behaviours both in vitro and in vivo. Finally, decreased MAGEC2 expression and low levels of EMT markers were detected in sh-MAGEC2 xenografts, while increased MAGEC2 expression and high levels of EMT markers were observed in invasive and metastatic HCC samples. Conclusion: Taken together, our data imply that MAGEC2 is a novel prognostic marker for HCC and that MAGEC2 significantly promotes HCC tumourigenesis by inducing EMT. Targeting MAGEC2 may provide a promising therapeutic strategy for HCC treatment. [ABSTRACT FROM AUTHOR]

Highlights • Three mutants with evidently improved kcat/Km were selected by directed evolution. • Revealed several 'hotspots' related with kcat/Km and thermostability of Bman26. • Structural-functional analysis gave insights into the molecular mechanism of key residues. • New findings will help promote the development of industrially useful β-mannanase. Abstract A β-mannanase gene was cloned from Bacillus sp. MK-2 and expressed in Bacillus subtilis WB800. The ORF of the β-mannanase gene was 1104 bp in length, encoding 367 aa. The deduced amino acid sequence shared high sequence identity with the β-mannanase from Bacillus subtilis , and belongs to glycosyl hydrolase family 26. The purified recombinant enzyme had a specific activity of 2802 U/mg and displayed optimum activity at pH 6.0 and 55 °C. To obtain an enzyme with high specific activity and facilitate its industrial applications, molecular engineering of Bman26 was undertaken using random mutagenesis in Bacillus subtilis WB800. Three positive mutants with substantially improved specific activities were selected and studied. The best performing mutant was K291E, for which the single amino acid substitution led to a 3.5-fold increase in k cat /K m. Mutants Q112R and L211I also exhibited an apparently increased k cat /K m towards konjac glucomannan, approximately 200% and 80% improvement, respectively. Structural-functional analysis indicated that a slight conformational change could dramatically affect certain enzyme characteristics. In addition, three amino acid sites (Gly88-Leu212-Lys288) in Bman26 were found to have close relationships with the enzyme's thermal stability. These new findings will help promote the development of industrially useful β-mannanase, with both good thermal stability and high specific activity. [ABSTRACT FROM AUTHOR]

Abstract Testicular 3β-hydroxysteroid dehydrogenase (HSD3B) is a steroidogenic enzyme, catalyzing the conversion of 3β-hydroxysteroids into 3-keto-steroids. Two distinct isoforms in the human are cloned, HSD3B1 and HSD3B2, and HSD3B2 is located in the testis. HSD3B2 is a two-substrate enzyme, which binds to cofactor NAD+ and a 3β-steroid. Many endocrine disruptors, including industrial compounds (phthalates, bisphenols, and perfluoroalkyl substances), insecticides and biocides (organochlorine insecticides and organotins), food additives (butylated hydroxyanisole, resveratrol, gossypol, flavones, and isoflavones), and drugs (etomidate, troglitazone, medroxyprogesterone acetate, and ketoconazole) inhibit testicular HSD3B, possibly interfering with androgen synthesis. In this review, we discuss the distinct testicular isoform of HSD3B, its gene, chemistry, subcellular location, and the endocrine disruptors that directly inhibit testicular HSD3B and their inhibitory modes. Highlights • HSD3B catalyzes 3β-dehydrogenase of 3β-hydroxysteroids. • Testicular HSD3B is critical for testosterone synthesis. • There is a SAR difference for phthalate-mediated inhibition of HSD3B. • Some flavones and isoflavones are very potent HSD3B inhibitors. [ABSTRACT FROM AUTHOR]

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Science performances, or science shows, combine scientific content with theatrical techniques for the purpose of engaging audiences with science. Despite the fragmented nature of the evidence on their effectiveness, science shows are commonly used in informal science learning, particularly in science centres, schools and museums. We have collated the literature into a comprehensive review of the history, typology and evidence base for science performance, drawing on a variety of literature including research discussing classroom demonstrations, science shows used in schools and museums, museum theatre and informal science learning. We found there is good qualitative and quantitative evidence to support the use of science shows both in informal science learning and as part of the school curriculum. Science shows that use demonstrations linked by a common theme have successfully produced both cognitive and affective outcomes, and theatrical science shows are frequently used to create awareness and stimulate discussion about complex and controversial science-related issues, such as HIV-AIDS and human cloning. The key features of structure, emotion and audience involvement that are present in good science shows make them a highly valuable and effective tool for both formal and informal science learning. Further research is needed to investigate the long-term impact of science performance. [ABSTRACT FROM AUTHOR]

Stem cell and cloning studies bear promising potential in the field of regenerative medicine with their exciting and innovative features. The wide range of applications of these cellular treatments and their permanent repairing qualities foretell that a completely different scientific paradigm is emerging for the future. However, satisfactory insight has not yet been developed with respect to the moral status and legal aspects of these studies. Furthermore, their potentially untoward side effects are not understood well enough to compare with the benefits they provide for human life. In general, attempts are made to place the subject in a pattern composed of the sanctity of life, the unique characteristics of being an individual, the boundaries drawn by beliefs, and the basic principles of law. This article will discuss the benefits of stem cell and cloning studies as well as their undeveloped medical, ethical, religious, and legal aspects. [ABSTRACT FROM AUTHOR]

Emerging evidence suggests the potential use of rapamycin in treatment of several neurological disorders. The drug readily crosses the blood brain barrier and may exert direct immunomodulatory effects within the brain. Microglia are the main innate immune cells of the brain, thus critically involved in the initiation and development of inflammatory processes at this level. However, there are conflicting data from rodent studies about the pharmacological effects of rapamycin on microglial inflammatory responses. Considering that rodent microglia display relevant biochemical and pharmacological differences compared to human microglia, in the present study we studied the effects of rapamycin in an experimental model of human microglia, the human microglial clone 3 (HMC3) cell line. Rapamycin was tested in the nM range both under basal conditions and in cells activated with a pro-inflammatory cytokine cocktail, consisting in a mixture of interferon-γ and interleukin-1β (II). The drug significantly increased II stimulatory effect on interleukin-6 (IL-6) expression and release in the HMC3 cells, while reducing the production of free oxygen radicals (ROS) both under basal conditions and in cells activated with II. Consistently with its known molecular mechanism of action, rapamycin reduced the extent of activation of the so-called 'mechanistic' target of rapamycin complex 1 (mTORC1) kinase and the total amount of intracellular proteins. In contrast to rodent cells, rapamycin did not alter human microglial cell viability nor inhibited cell proliferation. Moreover, rapamycin did not exert any significant effect on the morphology of the HMC3 cells. All together these data suggest that the inhibition of mTORC1 in human microglia by rapamycin results in complex immunomodulatory effects, including a significant increase in the expression and release of the pro-inflammatory IL-6. [ABSTRACT FROM AUTHOR]

Cementum is the mineralized tissue covering the tooth root that functions in tooth attachment and post-eruptive adjustment of tooth position. It has been reported to be highly similar to bone in several respects but remains poorly understood in terms of development and regeneration. Here, we investigate whether cementocytes, the residing cells in cellular cementum, have the potential to be protagonist in cementum homeostasis, responding to endocrine signals and directing local cementum metabolism. Cells from healthy erupted human teeth were isolated using sequential collagenase/EDTA digestions, and maintained in standard cell culture conditions. A cementocyte-like cell line was cloned (HCY-23, for human cementocyte clone 23), which presented a cementocyte compatible gene expression signature, including the expression of dentin matrix protein 1 (DMP1), sclerostin (SOST), and E11/gp38/podoplanin (E11). In contrast, these cells did not express the odontoblast/dentin marker dentin sialoprotein (DSPP). HCY-23 cells produced mineral-like nodules in vitro under differentiation conditions, and were highly responsive to inorganic phosphate (Pi). Within the limits of the present study, it can be concluded that cementocytes are phosphate-responsive cells, and have the potential do play a key role in periodontal homeostasis and regeneration. [ABSTRACT FROM AUTHOR]

This paper explores the representation of cannibalism in David Mitchell’s novel, Cloud Atlas, and delineates two levels on which it appears; the first context portrays cannibalistic tribes, while the second one displays an industry of producing and recycling human clones as fast food. In addition, this analysis focuses on the confrontation of what it means to be civilized or savage, and on the emerging social issues revolving around consumer culture, as the juxtaposition of consumer society and the figure of the cannibal challenges the concept of consumer society by questioning if it is civilised at all or not. [ABSTRACT FROM AUTHOR]

Among bioethical themes at the centre of public debate, one of the most sensitive is cloning, which has been controversial since its official appearance on the scene in the 1990s. The debate on the practice and its possible future application to humans has never ceased since Dolly the sheep was first born in 1996. It gathered new momentum when in late January 2018 Chinese researchers announced that they had successfully cloned two macaque monkeys applying a perfected version of the technique originally used to create Dolly. A barrier had been broken, and inevitably a whole series of ethical concerns were raised. This article looks at the coverage of the 2018 monkey cloning success in the daily press worldwide, and at the discursive procedures used to illustrate its significance, including the popularisation strategies deployed. The focus of the analysis is on ethical implications, with special regard to animal testing ethics and to the possible extension of the technique to humans. [ABSTRACT FROM AUTHOR]