bluuebunny/crossref_metadata_2025_split huggingface dataset:
A molecular survey and phylogenetic analysis of porcine circovirus type 3 using oral fluid, faeces and serum
Authors: Plut Jan , Jamnikar-Ciglenecki Urska , Golinar-Oven Irena , Knific Tanja , Stukelj Marina 
Published: August 2020

Abstract
Background
Porcine circovirus type 3 is the most recently discovered porcine circovirus, and an emerging pathogen. In this study the status of its presence on some Slovenian farms is reported. The effectiveness of the vaccine against porcine circovirus type 2 was assessed against porcine circovirus type 3.
Group samples of oral fluid, faeces and individual serum samples were taken from six different pig categories and tested for presence of viral DNA, using both real time and conventional PCR. Positive samples were subjected to direct Sanger sequencing. Nucleotide sequences were analyzed and compared to GenBank PCV3 sequences.

Results
Positive samples were sent for genome sequencing, which confirmed the presence of virus in all different pig categories on five farms. A high to moderate correlation of strong statistical significance was found between individual serum samples, oral fluid and faeces. Slovenian PCV3 was found to be distributed in a way similar to that of other countries. Slovenian PCV3 nt sequences are highly related, sharing more than 99.5% nt identity. On one farm a commercially available vaccine against porcine circovirus type 2 was used on 3-week-old pigs. It did not affect the presence of porcine circovirus type 3 in oral fluid or sera of any of the seven age groups of pigs, each with two control groups.

Conclusions
The results constitute the first discovery of the virus in Slovenia. Genome sequencing has revealed a high degree of similarity between Slovenian and GenBank isolates.
Neural differentiation of canine mesenchymal stem cells/multipotent mesenchymal stromal cells
Authors: Prpar Mihevc Sonja , Kokondoska Grgich Vesna , Kopitar Andreja Nataša , Mohorič Luka , Majdič Gregor 
Published: August 2020

Abstract
Background
The ability of adipose tissue-derived multipotent mesenchymal stromal cells/mesenchymal stem cells (ASCs) to differentiate in neural lineages promises progress in the field of regenerative medicine, especially for replacing neuronal tissue damaged by different neurological disorders. Reprogramming of ASCs can be induced by the growth medium with neurogenic inductors and specific growth factors. We investigated the neural differentiation potential of canine ASCs using several growth media (KEM, NIMa, NIMb, NIMc) containing various combinations of neurogenic inductors: B27 supplement, valproic acid, forskolin, N2-supplement, and retinoic acid. Cells were first preconditioned in the pre-differentiation neural induction medium (mitogenically stimulated; STIM1), followed by the induction of neuronal differentiation.

Results
After 3, 6, and 9 days of neural induction, elongated neural-like cells with bipolar elongations were observed, and some oval cells with light nuclei appeared. The expression of neuronal markers tubulin beta III (TUBB3), neurofilament H (NF-H), microtubule-associated protein-2 (MAP2), and glial fibrillary acidic protein (GFAP) was observed using immunocytochemistry, which confirmed the differentiation into neurons and glial cells. Flow cytometry analysis showed high GFAP expression (between 70 and 90% of all cells) after cells had been growing three days in the neural induction medium a (NIMa). Around 25% of all cells also expressed adult neuronal markers NF-H and MAP2. After nine days of ASCs differentiation, the expression of all neural markers was reduced. There were no differences between the neural differentiation of ASCs isolated from female or male dogs.

Conclusions
The differentiation repertoire of canine ASCs extends beyond mesodermal lineages. Using a defined neural induction medium, the canine ASCs differentiated into neural lineages and expressed markers of neuronal and glial cells, and also displayed the typical neuronal morphology. Differentiated ASCs can thus be a source of neural cellular lineages for the regenerative therapy of nerve damage and could be useful in the future for therapy or the modelling of neurodegenerative diseases.
Cul o 2 specific IgG3/5 antibodies predicted Culicoides hypersensitivity in a group imported Icelandic horses
Authors: Raza Fahad , Ivanek Renata , Freer Heather , Reiche Dania , Rose Horst , Torsteinsdóttir Sigurbjörg , Svansson Vilhjálmur , Björnsdóttir Sigríður , Wagner Bettina 
Published: August 2020

Abstract
Background
Culicoides hypersensitivity (CH) is induced in horses by salivary allergens of Culicoides midges. In Iceland, the causal Culicoides species for CH are not present. Previous epidemiological data indicated that Icelandic horses are more susceptible to CH when they are exported from Iceland and first exposed to Culicoides at adult age. Horses born in countries where Culicoides is endemic, develop the disease less frequently. Here, we established a longitudinal allergy model to identify predictive and diagnostic serological biomarkers of CH.

Results
Sixteen adult Icelandic horses from Iceland were imported to the Northeastern United States (US) during the winter and were kept in the same environment with natural Culicoides exposure for the next two years. None of the horses showed clinical allergy during the first summer of Culicoides exposure. In the second summer, 9/16 horses (56%) developed CH. Allergen specific IgE and IgG isotype responses in serum samples were analysed using nine potential Culicoides allergens in a fluorescent bead-based multiplex assay. During the first summer of Culicoides exposure, while all horses were still clinically healthy, Cul o 2 specific IgG3/5 antibodies were higher in horses that developed the allergic disease in the second summer compared to those that did not become allergic (p = 0.043). The difference in Cul o 2 specific IgG3/5 antibodies between the two groups continued to be detectable through fall (p = 0.035) and winter of the first year. During the second summer, clinical signs first appeared and Cul o 3 specific IgG3/5 isotypes were elevated in allergic horses (p = 0.041). Cul o 2 specific IgG5 (p = 0.035), and Cul o 3 specific IgG3/5 (p = 0.043) were increased in late fall of year two when clinical signs started to improve again.

Conclusions
Our results identified IgG5 and IgG3/5 antibodies against Cul o 2 and Cul o 3, respectively, as markers for CH during and shortly after the allergy season in the Northeastern US. In addition, Cul o 2 specific IgG3/5 antibodies may be valuable as a predictive biomarker of CH in horses that have been exposed to Culicoides but did not yet develop clinical signs.
FGL1 regulates acquired resistance to Gefitinib by inhibiting apoptosis in non-small cell lung cancer
Authors: Sun Cuilan , Gao Weiwei , Liu Jiatao , Cheng Hao , Hao Jiqing 
Published: August 2020

AbstractBackgroundThis study investigated the role of fibrinogen-like protein 1 (FGL1) in regulating gefitinib resistance of PC9/GR non-small cell lung cancer (NSCLC).MethodsThe effect of different concentrations of gefitinib on cell proliferation were evaluated using the CCK-8 assay. FGL1 expression in the normal human bronchial epithelial cell line Beas-2B, as well as four lung tumor cell lines, H1975, A549, PC9, and PC9/GR, was investigated by using western blotting and qRT-PCR.FGL1was knocked down using small interfering RNA to evaluate the effects of FGL1 on PC9 and PC9/GR. The correlation between FGL1 expression and gefitinib resistance was determined in vitro via CCK-8 and colony formation assays, and flow cytometry and in vivo via flow cytometry and immunohistochemistry.ResultsFGL1 expression was significantly upregulated in non-small cell lung cancer cells with EGFR mutation and higher in the gefitinib-resistant NSCLC cell line PC9/GR than in the gefitinib-sensitive NSCLC cell line PC9. Further, FGL1 expression in PC9 and PC9/GR cells increased in response to gefitinib treatment in a dose-dependent manner. Knockdown ofFGL1suppressed cell viability, reduced the gefitinib IC50 value, and enhanced apoptosis in PC9 and PC9/GR cells upon gefitinib treatment. Mouse xenograft experiments showed thatFGL1knockdown in PC9/GR tumor cells enhanced the inhibitory and apoptosis-inducing actions of gefitinib. The potential mechanism of gefitinib in inducing apoptosis of PC9/GR cells involves inhibition of PARP1 and caspase 3 expression via suppression of FGL1.ConclusionsFGL1 confers gefitinib resistance in the NSCLC cell line PC9/GR by regulating the PARP1/caspase 3 pathway. Hence, FGL1 is a potential therapeutic target to improve the treatment response of NSCLC patients with acquired resistance to gefitinib.
The local transcriptional regulators SacR1 and SacR2 act as repressors of fructooligosaccharides metabolism in Lactobacillus plantarum
Authors: Chen Chen , Wang Linlin , Yu Haiyan , Tian Huaixiang 
Published: August 2020

Abstract
Background
In Lactobacillus plantarum, fructooligosaccharides (FOS) metabolism is controlled by both global and local regulatory mechanisms. Although catabolite control protein A has been identified as a global regulator of FOS metabolism, the functions of local regulators remain unclear. This study aimed to elucidate the roles of two local regulators, SacR1 and SacR2, in the regulation of FOS metabolism in L. plantarum both in vitro and in vivo.

Results
The inactivation of sacR1 and sacR2 affected the growth and production of metabolites for strains grown on FOS or glucose, respectively. A reverse transcription-quantitative PCR analysis of one wild-type and two mutant strains (ΔsacR1 and ΔsacR2) of L. plantarum identified SacR1 and SacR2 as repressors of genes relevant to FOS metabolism in the absence of FOS, and these genes could be induced or derepressed by the addition of FOS. The analysis predicted four potential transcription factor binding sites (TFBSs) in the putative promoter regions of two FOS-related clusters. The binding of SacR1 and SacR2 to these TFBSs both in vitro and in vivo was verified using electrophoretic mobility shift assays and chromatin immunoprecipitation, respectively. A consensus sequence of WNNNNNAACGNNTTNNNNNW was deduced for the TFBSs of SacR1 and SacR2.

Conclusion
Our results identified SacR1 and SacR2 as local repressors for FOS metabolism in L. plantarum. The regulation is achieved by the binding of SacR1 and SacR2 to TFBSs in the promoter regions of FOS-related clusters. The results provide new insights into the complex network regulating oligosaccharide metabolism by lactic acid bacteria.
A common approach for absolute quantification of short chain CoA thioesters in prokaryotic and eukaryotic microbes
Authors: Gläser Lars , Kuhl Martin , Jovanovic Sofija , Fritz Michel , Vögeli Bastian , Erb Tobias J. , Becker Judith , Wittmann Christoph 
Published: August 2020

AbstractBackgroundThioesters of coenzyme A participate in 5% of all enzymatic reactions. In microbial cell factories, they function as building blocks for products of recognized commercial value, including natural products such as polyketides, polyunsaturated fatty acids, biofuels, and biopolymers. A core spectrum of approximately 5–10 short chain thioesters is present in many microbes, as inferred from their genomic repertoire. The relevance of these metabolites explains the high interest to trace and quantify them in microbial cells.ResultsHere, we describe a common workflow for extraction and absolute quantification of short chain CoA thioesters in different gram-positive and gram-negative bacteria and eukaryotic yeast, i.e.Corynebacterium glutamicum,Streptomyces albus,Pseudomonas putida, andYarrowia lipolytica. The approach assessed intracellular CoA thioesters down to the picomolar level and exhibited high precision and reproducibility for all microbes, as shown by principal component analysis. Furthermore, it provided interesting insights into microbial CoA metabolism. A succinyl-CoA synthase defective mutant ofC. glutamicum exhibited an unaffected level of succinyl-CoA that indicated a complete compensation by thel-lysine pathway to bypass the disrupted TCA cycle. Methylmalonyl-CoA, an important building block of high-value polyketides, was identified as dominant CoA thioester in the actinomyceteS.albus. The microbe revealed a more than 10,000-fold difference in the abundance of intracellular CoA thioesters. A recombinant strain ofS. albus, which produced different derivatives of the antituberculosis polyketide pamamycin, revealed a significant depletion of CoA thioesters of the ethylmalonyl CoA pathway, influencing product level and spectrum.ConclusionsThe high relevance of short chain CoA thioesters to synthetize industrial products and the interesting insights gained from the examples shown in this work, suggest analyzing these metabolites in microbial cell factories more routinely than done so far. Due to its broad application range, the developed approach appears useful to be applied this purpose. Hereby, the possibility to use one single protocol promises to facilitate automatized efforts, which rely on standardized workflows.
Therapeutic inhibition of microRNA-21 (miR-21) using locked-nucleic acid (LNA)-anti-miR and its effects on the biological behaviors of melanoma cancer cells in preclinical studies
Authors: Javanmard Shaghayegh Haghjooy , Vaseghi Golnaz , Ghasemi Ahmad , Rafiee Laleh , Ferns Gordon A. , Esfahani Hajar Naji , Nedaeinia Reza 
Published: August 2020

AbstractBackgroundMelanoma is a cancer that has a high mortality rate in the absence of targeted therapy. Conventional therapies such as surgery, chemotherapy, and radiotherapy are associated with poor prognosis. The expression of miR-21 appears to be of clinical importance, and the regulation of its expression appears to be an opportunity for treatment.MethodsIn this current study, we aimed to evaluate the effects of miR-21 inhibition in- vitro and in-vivo. In-vitro studies have investigated LNA-anti-miR-21 in mouse melanoma cells (B16F10), and in-vivo studies have proposed a model of melanoma in male C57BL/6 mice. To evaluate the anticancer effects of LNA-anti-miR-21, a QRT-PCR analysis was performed using the 2−ΔΔCTmethod to determine the degree of inhibition of oncomiR-21. The MTT test, propidium iodide/AnnexinV in-vitro, and tumor volume measurement using the QRT-PCR test with the 2−ΔΔCTmethod were used to estimate the inhibition of miR-21 and the expression of downstream genes including:SNAI1, Nestin (Nes), Oct-4, andNF-kBfollowing miR-21 inhibition. Finally, immunohistochemistry was conducted for an in-vivo animal study.ResultsMiR-21 expression was inhibited by 80% after 24 h of B16F10 cell line transfection with LNA-anti-miR-21. The MTT test showed a significant reduction in the number of transfected cells with LNA-anti-miR-21. The transfected cells showed a significant increase in apoptosis in comparison with the control and scrambled LNA groups. According to our in vivo findings, anti-miR-21 could reduce tumor growth and volume in mice receiving intraperitoneal anti-miR after 9 days. The expression of theSNAI1gene was significantly reduced compared to the controls. Immunohistochemical analysis showed no change inCD133andNF-kBmarkers.ConclusionOurfindings suggest LNA-anti-miR-21 can be potentially used as an anticancer agent for the treatment of melanoma.
RACK1 promotes miR-302b/c/d-3p expression and inhibits CCNO expression to induce cell apoptosis in cervical squamous cell carcinoma
Authors: Wang Jing , Chen Shengcai 
Published: August 2020

Abstract
Background
Cervical squamous cell carcinoma (CSCC) is one of the main causes of cancer-related deaths in women worldwide. The present study was conducted with the main objective of determining the potential role of receptor for activated protein kinase C1 (RACK1) in CSCC through regulation of microRNA (miR)-302b/c/d-3p and Cyclin O (CCNO).

Methods
The expression of RACK1, miR-302b/c/d-3p and CCNO in CSCC tissues and cells was measured by RT-qPCR and Western blot analysis. The interaction among RACK1, miR-302b/c/d-3p, and CCNO was determined by dual luciferase reporter assay. Subsequently, effects of RACK1, miR-302b/c/d-3p and CCNO on CSCC cell cycle entry, proliferation and apoptosis were investigated with the use of flow cytometry, EdU, and TUNEL assays. Furthermore, mouse xenograft model of CSCC cells was established to verify the function of RACK1 in vivo.

Results
RACK1 and miR-302b/c/d-3p were down-regulated and CCNO was overexpressed in CSCC. CCNO was identified as the target of miR-302b/c/d-3p. Importantly, overexpressed miR-302b-3p, miR-302c-3p or miR-302d-3p or RACK1 enhanced the apoptosis and suppressed the proliferation of CSCC cells in vitro, while inhibiting tumor growth in vivo by targeting CCNO.

Conclusions
On all accounts, overexpressed RACK1 could dampen the progression of CSCC through miR-302b/c/d-3p-mediated CCNO inhibition.
The characterization of extracellular vesicles-derived microRNAs in Thai malaria patients
Authors: Ketprasit Nutpakal , Cheng Iris Simone , Deutsch Fiona , Tran Nham , Imwong Mallika , Combes Valery , Palasuwan Duangdao 
Published: August 2020

Abstract
Background
Extracellular vesicles (EVs) have been broadly studied in malaria for nearly a decade. These vesicles carry various functional biomolecules including RNA families such as microRNAs (miRNA). These EVs-derived microRNAs have numerous roles in host-parasite interactions and are considered promising biomarkers for disease severity. However, this field lacks clinical studies of malaria-infected samples. In this study, EV specific miRNAs were isolated from the plasma of patients from Thailand infected with Plasmodium vivax and Plasmodium falciparum. In addition, it is postulated that these miRNAs were differentially expressed in these groups of patients and had a role in disease onset through the regulation of specific target genes.

Methods
EVs were purified from the plasma of Thai P. vivax-infected patients (n = 19), P. falciparum-infected patients (n = 18) and uninfected individuals (n = 20). EV-derived miRNAs were then prepared and abundance of hsa-miR-15b-5p, hsa-miR-16-5p, hsa-let-7a-5p and hsa-miR-150-5p was assessed in these samples. Quantitative polymerase chain reaction was performed, and relative expression of each miRNA was calculated using hsa-miR-451a as endogenous control. Then, the targets of up-regulated miRNAs and relevant pathways were predicted by using bioinformatics. Receiver Operating Characteristic with Area under the Curve (AUC) was then calculated to assess their diagnostic potential.

Results
The relative expression of hsa-miR-150-5p and hsa-miR-15b-5p was higher in P. vivax-infected patients compared to uninfected individuals, but hsa-let-7a-5p was up-regulated in both P. vivax-infected patients and P. falciparum-infected patients. Bioinformatic analysis revealed that these miRNAs might regulate genes involved in the malaria pathway including the adherens junction and the transforming growth factor-β pathways. All up-regulated miRNAs could potentially be used as disease biomarkers as determined by AUC; however, the sensitivity and specificity require further investigation.

Conclusion
An upregulation of hsa-miR-150-5p and hsa-miR-15b-5p was observed in P. vivax-infected patients while hsa-let-7a-5p was up-regulated in both P. vivax-infected and P. falciparum-infected patients. These findings will require further validation in larger cohort groups of malaria patients to fully understand the contribution of these EVs miRNAs to malaria detection and biology.
Vulnerability and agency across treatment-seeking journeys for acutely ill children: how family members navigate complex healthcare before, during and after hospitalisation in a rural Kenyan setting
Authors: Zakayo Scholastica M. , Njeru Rita W. , Sanga Gladys , Kimani Mary N. , Charo Anderson , Muraya Kui , Sarma Haribondhu , Uddin Md. Fakhar , Berkley James A. , Walson Judd L. , Kelley Maureen , Marsh Vicki , Molyneux Sassy 
Published: August 2020

Abstract
Background
Child mortality rates during hospitalisation for acute illness and after discharge are unacceptably high in many under-resourced settings. Childhood vulnerability to recurrent illness, and death, is linked to their families’ situations and ability to make choices and act (their agency). We examined vulnerability and agency across treatment-seeking journeys for acutely ill children and considered the implications for policy and practice.

Method
A qualitative sub-study was embedded within the prospective CHAIN Network cohort study, which is investigating mechanisms of inpatient and post-hospital discharge mortality among acutely ill young children across a spectrum of nutritional status. Primary data were collected from household members of 20 purposively selected cohort children over 18 months through formal interviews (total n = 74), complemented by informal discussions and observations. Data were analysed using narrative and thematic approaches.

Results
Treatment-seeking pathways were often long and complex, particularly for children diagnosed as severely malnourished. Family members’ stories reveal that children’s carers, usually mothers, navigate diverse challenges related to intersecting vulnerabilities at individual, household and facility levels. Specific challenges include the costs of treatment-seeking, confusing and conflicting messaging on appropriate care and nutrition, and poor continuity of care. Strong power inequities were observed between family members and health staff, with many mothers feeling blamed for their child’s condition. Caregivers’ agency, as demonstrated in decision-making and actions, often drew on the social support of others but was significantly constrained by their situation and broader structural drivers.

Conclusion
To support children’s care and recovery, health systems must be more responsive to the needs of families facing multiple and interacting vulnerabilities. Reducing incurred treatment costs, improving interpersonal quality of care, and strengthening continuity of care across facilities is essential. Promising interventions need to be co-designed with community representatives and health providers and carefully tested for unintended negative consequences and potential for sustainable scale-up.