bluuebunny/crossref_metadata_2025 huggingface dataset:
Precision medicine phase II study evaluating the efficacy of a double immunotherapy by durvalumab and tremelimumab combined with olaparib in patients with solid cancers and carriers of homologous recombination repair genes mutation in response or stable after olaparib treatment
Authors: Fumet Jean-David , Limagne Emeric , Thibaudin Marion , Truntzer Caroline , Bertaut Aurélie , Rederstorff Emilie , Ghiringhelli Francois
Published: August 2020

Background

Tumors with deficient homologous repair are sensitive to PARP inhibitors such as olaparib which is known to have immunogenic properties. Durvalumab (D) is a human monoclonal antibody (mAb) which inhibits binding of programmed cell death ligand 1 (PD-L1) to its receptor. Tremelimumab (T) is a mAb directed against the cytotoxic T-lymphocyte-associated protein 4 (CTLA-4). This study is designed to evaluate the efficacy of combination of olaparib, durvalumab and tremelimumab in patients with a solid tumors with a mutation in homologous gene repair.

Methods

This phase II study will assess the efficacy and safety of olaparib/D/T association in patients (n = 213) with several types of solid cancers (breast cancer, ovarian cancer, pancreatic cancer, endometrial cancer, prostate cancer and others) with at least one mutation in homologous repair genes (BRCA1, BRCA2, PALB2, ATM, FANCA, FANCB, FANCC, FANCE, FANCF, CHEK2, RAD51, BARD1, MRE11, RAD50, NBS1, HDAC2), LKB1/STK11, INPP4B, STAG2, ERG, CHEK1, BLM, LIG4, ATR, ATRX, CDK12). Good performance status patients and corresponding to specific inclusion criteria of each cohort will be eligible. STEP1: Patients will receive olaparib 300 mg BID. In absence of progression after 6 weeks of olaparib, they will follow STEP 2 with olaparib and immunotherapy by durvalumab (1500 mg Q4W) + tremelimumab (75 mg IV Q4W) during 4 months and will further pursue durvalumab alone until disease progression, death, intolerable toxicity, or patient/investigator decision to stop (for a maximum duration of 24 months, and 36 months for ovarian cohort). Primary endpoint is safety and efficacy according to progression-free survival (PFS) of olaparib + immunotherapy (durvalumab + tremelimumab) during 4 months followed by durvalumab alone as maintenance in patients with solid cancers and in response or stable, after prior molecular target therapy by olaparib; secondary endpoints include overall survival (OS), disease control rate (DCR), response rate after 6 weeks of olaparib, safety of olaparib/durvalumab/tremelimumab association. Blood, plasma and tumor tissue will be collected for potential prognostic and predictive biomarkers.

Discussion

This study is the first trial to test the combination of olaparib and double immunotherapy based on molecular screening.

Trial registration

NCT04169841, date of registration November 20, 2019
Effect of Tropicamide on crystalline Lens rise in low-to-moderate myopic eyes
Authors: Chen Zhuoyi , Li Tao , Li Meiyan , Xu Ye , Zhou Xingtao
Published: August 2020

Background

Cycloplegics have been reported to induce changes in the lens thickness. However, the studies of correlation between cycloplegia and the lens position are limited. This study aims to investigate changes in crystalline lens rise (CLR) and other anterior segment parameters after inducing cycloplegia with tropicamide.

Methods

In this consecutive case study, 39 children (20 boys and 19 girls; mean age, 9.51 ± 1.75 years, mean spherical equivalence [SE], − 1.9 ± 1.5 D) with low-to moderate myopia were examined using CASIA 2 both before and after 30 min of administering 5-cycles (each 5 min apart) of 0.5% tropicamide. Measurements included CLR, crystalline lens thickness (CLT), mean radius of curvature of the anterior/posterior surface of the lens (Rf_ave/Rb_ave), anterior chamber depth (ACD), anterior chamber width (ACW), and central corneal thickness (CCT). Correlations of CLT and CLR with ACD, SE, and age were assessed respectively.

Results

CLT and CLR decreased significantly after cycloplegia (p < 0.001 andp < 0.001, respectively); whereas CCT, ACD, and Rf_ave increased (p = 0.008,p < 0.001,p < 0.001, respectively). A positive correlation was found between CLR and SE (r = 0.565,p < 0.001). However, a negative correlation between ACD and CLR was found before and after cycloplegia (r = − 0.430,p = 0.006;r = − 0.342,p = 0.035, respectively).

Conclusions

The crystalline lens appeared thinner and moved backward after cycloplegia. ACD increased mainly due to the backward movement of the crystalline lens. These results aid in elucidating the impact of crystalline lens changes during the process of accommodation.
A case series of children with adenovirus pneumonia: three-year experiences in a tertiary PICU
Authors: Shi Jingyi , Zhou Yiping , Wang Fei , Wang Chunxia , Miao Huijie , Sun Ting , Shan Yijun , Cui Yun , Zhang Yucai
Published: August 2020

Background

Describe the outcome of adenovirus pneumonia in a pediatric intensive care unit (PICU) over a 3-year period, to identify the risk factors that may be associated with worse outcome.

Methods

A retrospective observational study was performed in the PICU of children’s hospital in Shanghai from July 2016 to June 2019. Sixty-seven children over 29 days to 14 years old with adenovirus pneumonia who were admitted to PICU with acute hypoxemic respiratory failure were included in this study. The primary outcome was hospital mortality, and secondary outcomes were hospital and PICU length of stay (LOS), and risk factors of worse outcome.

Results

Of 67 children with severe adenovirus pneumonia, the hospital mortality was 16.42% (11/67) and 28-day mortality was 14.93% (10/67). Median Pediatric Risk of Mortality III (PRISM III) score at admission was 13 (interquartile range [IQR], 10–15). Median PICU LOS stay was 11 days (8-18d) and hospital LOS was 22 days (16-31d). Among children with extracorporeal membrane oxygenation (n = 9), 6 cases survived and 3 cases died. The patients who need renal replacement therapy, neuromuscular blockade, parenteral nutrition, and packed red blood cell perfusion had higher hospital mortality (p < 0.001, p = 0.041, p = < 0.001, p = 0.012, respectively). Multivariate logistic analysis indicated that liver dysfunction and nosocomial infection were associated with high risk of mortality.

Conclusions

The hospital mortality of adenovirus pneumonia in our PICU was 16.42%. Patients complicated liver dysfunction and co-infection & nosocomial infection were associated with poor outcome.
Intra- and inter-rater reliability of thoracic spine mobility and posture assessments in subjects with thoracic spine pain
Authors: Takatalo Jani , Ylinen Jari , Pienimäki Tuomo , Häkkinen Arja
Published: August 2020

Background

The thoracic spine (TS) has been neglected in the study of the spine despite its essential role in the stability and posture of the entire spinal complex. Therefore, there is an inevitable need to investigate the reproducibility of different thoracic spinal posture measures used in subjects with TS pain.

Methods

Thirty-two subjects (16 females and 16 males, mean age 39 years) were evaluated by two physiotherapists on the same day to gauge inter-rater reliability and on two consecutive days to gauge intra-rater reliability. TS posture was assessed by observation, and thoracic spine mobility was measured by manual assessment of segmental flexion and extension mobility in a seated position. Additionally, posterior-to-anterior accessory mobility in a prone position was assessed manually. Moreover, cervicothoracic flexion in a seated position, thoracic posture, and thoracic flexion and extension mobility in a standing position were assessed with a tape measure, and flexion and extension mobility in a seated position and TS posture in seated and standing positions were measured with an inclinometer. The intraclass correlation coefficient (ICC), standard error of measurement (SEM), mean difference (MD), Bland-Altman (B&A) plot features and coefficient of repeatability (CR) were calculated.

Results

The mean and standard deviation (SD) of the duration of TS pain was 22 (SD 45) months, with the intensity of pain being rated at 27 (SD 21) mm on a visual analogue scale (VAS). Intra-rater reliability was very strong (ICC ≥ 0.80) for the evaluation of seated and standing upper TS posture, standing whole TS posture and seated lower TS posture with an inclinometer. Moreover, TS posture evaluation with a measuring tape, posture inspection in a seated position, and manual assessment of segmental extension were found to have very strong intra-rater reliability. Inter-rater reliability was very strong for inclinometer measurements of standing and seated upper TS posture as well as standing whole TS posture.

Conclusion

Intra-rater reliability was higher than inter-rater reliability in most of the evaluated measurements. Overall, posture measurements with an inclinometer were more reliable than mobility measurements with the same instrument. The manual assessments can be used reliably when same evaluator performs the examination.

Trial registration

Clinical Trials, NCT01884818. Registered 24 June 2013,https://clinicaltrials.gov/ct2/show/NCT01884818?cond=thoracic+spine&cntry=FI&rank=1
Increased risk of knee osteoarthritis in patients using oral N-acetylcysteine: a nationwide cohort study
Authors: Yeh Ying-Ting , Liang Chung-Chao , Chang Chia-Ling , Hsu Chung-Y. , Li Pei-Chen
Published: August 2020

Background

Knee osteoarthritis (OA) is known to be a progressive degenerative disorder; however, recent evidence suggests that inflammatory mediators contribute to cartilage degradation. Studies have reported that N-acetylcysteine (NAC) had a promising effect on the reduction of the synthesis of proinflammatory and structural mediators by synovial cells. Given the lack of relevant clinical trials, we conducted this study to determine the relationship between NAC use and risk of knee OA.

Methods

We designed a retrospective cohort study from 2000 to 2013. Patients who received oral NAC over 28 days within 1 year after the first prescription were defined as the case group, whereas those without NAC use were considered as candidates of the control group. We adopted 1:4 propensity-score matching by age, sex, index year, and comorbidities to obtain the control group. The primary outcome was a new diagnosis of knee OA during the follow-up period.

Results

Our study sample comprised 12,928 people who used NAC and 51,715 NAC nonusers. NAC users had a significantly higher incidence of osteoarthritis (adjusted hazard ratio: 1.42, P < .001) than did NAC nonusers. Also, in analyses stratified by age group and sex, all subgroups exhibited a significantly higher incidence of knee osteoarthritis (P < .0001) among NAC users than among NAC nonusers. The use of oral NAC was associated with nearly four-fold increased the risk of knee OA in the young age group.

Conclusions

Long-term use of oral NAC is associated with a higher risk of knee OA.
Elevated non-high-density lipoprotein cholesterol corresponds to a high risk of nephrolithiasis in children
Authors: Chen Meiyuan , Xiao Jing , Du Yuan , Wang Miaomiao , Ruan Jimeng , Tian Ye
Published: August 2020

Background

Dyslipidemia contributes to the development of nephrolithiasis in adults; however its relationship to urolithiasis in children remains debatable, and will be clarified in the present work.

Methods

A case–control study was performed involving 58 pediatric patients diagnosed with upper urinary tract stones as well as 351 controls. Age, gender, body mass index (BMI), serum calcium, serum uric acid, blood glucose, blood lipids, and compositions of stones were compared.

Results

According to the univariate analysis, uric acid was higher (P < 0.01) but serum calcium lower in the stone group than the control (P < 0.05). As for the blood lipids, non-high-density lipoprotein cholesterol (non-HDL-c) was significantly higher in the stone group as compared to the control (P < 0.01), while total cholesterol, triglycerides, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol did not show statistical difference between the two groups. In the multivariate analysis, only non-HDL-c and serum uric acid were increased in the stone group (P = 0.003 and P = 0.008). In the stone compositions’ analysis, serum uric acid and non-HDL-c were associated with percentage of uric acid and pure calcium oxalate stones, respectively.

Conclusion

Non-high-density lipoprotein cholesterol may act as a lipid risk factor for urolithiasis in children.
Study of hospitalization and mortality in Korean diabetic patients using the diabetes complications severity index
Authors: Yoo Hyunju , Choo Eunjung , Lee Sukhyang
Published: August 2020

Background

The prevalence of type 2 diabetes mellitus (T2DM) is expected to increase from 7.7% in 2017 to 8.4% in 2045 worldwide. Diabetes complications contribute to morbidity and mortality. To evaluate whether the diabetes complications severity index (DCSI) was associated with increased risks of mortality and hospitalization.

Methods

A retrospective cohort study was conducted using the National Health Insurance Database (NHID) sample cohort of 1,102,047 patients (2002–2015) in Korea. Diabetes complications were evaluated at 2 years after the initial diagnosis and during the subsequent follow-up period (mean duration 6.56 ± 2.81 years). The type and severity of complications were evaluated on the basis of the International Classification of Disease Ninth (ICD-9) codes used in DCSI with 7 categories and 55 subcategories of complications. The Cox proportional hazard and Poisson regression models were used to evaluate the mortality and hospitalization rates. The incidence and relative risk of diabetes complications as well as the risk of mortality and hospitalization were the main outcome measures.

Results

A total of 27,871 patients were finally included and grouped by the number of complications present at 2 years. Four hundred ninety patients (5.37%) died without complications, 659 (7.31%) died with one complication and 1153 (11.85%) died with two or more complications. As DCSI at index date increased, the risk of additional new diabetes complications increased by 26% [relative risk (RR) 1.26, 95% CI 1.25–1.27]. The risks of mortality and hospitalization were linearly related to DCSI [hazard ratio 1.13 (95% CI 1.11–1.16), relative risk 1.04 (95% CI 1.03–1.06)].

Conclusions

Patients with higher incidence and severity of diabetes complications have increased risks of mortality and hospitalization.
Efficiency of tooth bleaching agent on staining and discoloration characteristics of nicotine stained dental enamel model
Authors: Lertsukprasert Nattha , Locharoenrat Kitsakorn
Published: August 2020

Background

Surface staining and deeper discoloration characteristics of peroxide-based bleaching agents in the nicotine stain in dental enamel model were evaluated in the present study.

Methods

Nicotine stained dental enamel fragments (n = 36) were prepared and were subjected to the bleaching ingredients for a fixed treatment time of 30 min. The bleaching agents were composed of limonene, coconut diethanolamide, and carbamide peroxide served as solvent, nonionic surfactant, and oxidizer, respectively. Optical analysis was carried out considering color stability via colorimeter and UV-Vis spectrometer.

Results

Degrees of color variations were significantly influenced by nicotine content and bleaching ingredient factors. They varied in the range of approximately 3.00 and 5.00 units for all tooth-bleaching agents. The most prominent degrees of color variation elevations were obtained in the tooth bleaching formulae set #2 (1.0% limonene + 20% coconut diethanolamide) in the stained tooth model in comparison to set #1 (0.5% limonene + 10% coconut diethanolamide) and set #3 (1.5% limonene + 30% coconut diethanolamide), partly due to the perceptible color changes. The lowest degree of color variation under a dose limitation was found in the tooth bleaching formulae set #2 + 10% carbamide peroxide formulation. Absorbance spectra were also evaluated after the interaction of bleaching treatment. They confirmed a relationship between nicotine content and discoloration characteristics of the tooth bleaching formulae set #2 + 10% carbamide peroxide.

Conclusions

Carbamide peroxide is considered as generator of free radicals. It converts the color of stains to clear by oxidizing the organic compounds in the stained dental enamel model, achieving whiteness enhancement.
The impact of a hands-on arthrocentesis workshop in undergraduate medical education
Authors: Ladurner Andreas , Nijman Thomas , Gill Tiffany K. , Smitham Peter J.
Published: August 2020

Background

To evaluate the impact of a training programme for arthrocentesis on procedural skills enhancement and self-confidence in medical students.

Methods

Participants were provided a structured workshop on injection models. A self-confidence questionnaire and medical knowledge assessment were performed. Retention of knowledge and skills were assessed at a later time point during a formal OSCE examination and compared to participants who had not attended a lecture and clinical attachments only.P-values, 95% confidence intervals about the mean, standard error of the mean, and standard deviations of the differences were calculated.

Results

All participants gained self-confidence, and improvement of their skills was significant. The mean self-confidence with performing an arthrocentesis procedure increased from 1.3 pre- to 5.9 points post-workshop (10-point Likert scale). The knee was the joint students felt most confident with (1.3 to 6.5 points). Knowledge on the selection of corticosteroid preparations (1.2 to 5.8 points) gained substantially, as well as confidence in providing post-injection advice (1.9 to 6.6 points). Upon the OSCE examination, attendance to the workshop resulted in a significant higher total score (16.2 vs 14.8 points,p < 0.05).

Conclusions

A workshop for arthrocentesis procedures, in conjunction with other learning activities, is well suited to increasing skills and self-confidence in fourth year medical students and allows for developing important baseline knowledge and practicing invasive techniques without risk to a patient.

Trial registration

This trial has been approved by the human research ethics committee of the University of Adelaide (Ethics approval No H-2019-134).
Evaluating stakeholder involvement in building a decision support tool for NHS health checks: co-producing the WorkHORSE study
Authors: Lloyd-Williams Ffion , Hyseni Lirije , Guzman-Castillo Maria , Kypridemos Chris , Collins Brendan , Capewell Simon , Schwaller Ellen , O’Flaherty Martin
Published: August 2020

Background

Stakeholder engagement is being increasingly recognised as an important way to achieving impact in public health. The WorkHORSE (Working Health Outcomes Research Simulation Environment) project was designed to continuously engage with stakeholders to inform the development of an open access modelling tool to enable commissioners to quantify the potential cost-effectiveness and equity of the NHS Health Check Programme.

An objective of the project was to evaluate the involvement of stakeholders in co-producing the WorkHORSE computer modelling tool and examine how they perceived their involvement in the model building process and ultimately contributed to the strengthening and relevance of the modelling tool.

Methods

We identified stakeholders using our extensive networks and snowballing techniques. Iterative development of the decision support modelling tool was informed through engaging with stakeholders during four workshops. We used detailed scripts facilitating open discussion and opportunities for stakeholders to provide additional feedback subsequently. At the end of each workshop, stakeholders and the research team completed questionnaires to explore their views and experiences throughout the process.

Results

30 stakeholders participated, of which 15 attended two or more workshops. They spanned local (NHS commissioners, GPs, local authorities and academics), third sector and national organisations including Public Health England.

Stakeholders felt valued, and commended the involvement of practitioners in the iterative process. Major reasons for attending included: being able to influence development, and having insight and understanding of what the tool could include, and how it would work in practice. Researchers saw the process as an opportunity for developing a common language and trust in the end product, and ensuring the support tool was transparent. The workshops acted as a reality check ensuring model scenarios and outputs were relevant and fit for purpose.

Conclusions

Computational modellers rarely consult with end users when developing tools to inform decision-making. The added value of co-production (continuing collaboration and iteration with stakeholders) enabled modellers to produce a “real-world” operational tool. Likewise, stakeholders had increased confidence in the decision support tool’s development and applicability in practice.
Patient-appraised beneficial moments during inpatient psychiatric treatment
Authors: Locher Cosima , Mansour Ramin , Koechlin Helen , Büchi Stefan
Published: August 2020

Background

Psychiatric inpatients receive a multidisciplinary treatment approach, covering psychiatry, nursing, occupational therapy, and psychology. Research findings reveal that the effectiveness of any treatment is associated with three types of factors: specific (e.g., treatment techniques), common (e.g., clinician-patient relationship, patients’ expectations) and extra-therapeutic. However, there is little published research on the factors and events which inpatients themselves consider to be beneficial (‘beneficial moments’).

Methods

Inpatients (N = 107) of a psychiatric clinic completed a questionnaire to elicit their appraisal of beneficial moments. A qualitative content analysis was applied. The coding procedure was conducted independently by two authors.

Results

Self-appraised beneficial moments were found in five areas: therapy-specific components (number of quotations,N = 204), positive relationships (N = 140), clinical setting and environment (N = 52), inpatients’ new insights (N = 36), and factors unrelated to either therapy or the clinic (N = 30). In total, 44% of the quotations were related to specific factors, 49% to common factors, and 7% to extra-therapeutic factors.

Conclusions

Inpatients judge both specific and common factors as crucial for the therapeutic benefit they gain during their stay at the clinic. Our results differ from meta-analytical findings, where the impact of specific factors on symptom improvement has shown to be much smaller (i.e., 17%) than appraised by patients in our study (i.e., 44%). Our study underlines the importance of a patient-centred care approach as well as shared decision making and patient-clinician communication. For clinical practice, knowledge of inpatients’ perspectives on beneficial moments is crucial in order to reinforce precisely these therapeutic components.
Impact of intermittent preventive treatment of malaria in pregnancy with dihydroartemisinin-piperaquine versus sulfadoxine-pyrimethamine on the incidence of malaria in infancy: a randomized controlled trial
Authors: Kakuru Abel , Jagannathan Prasanna , Kajubi Richard , Ochieng Teddy , Ochokoru Harriet , Nakalembe Miriam , Clark Tamara D. , Ruel Theodore , Staedke Sarah G. , Chandramohan Daniel , Havlir Diane V. , Kamya Moses R. , Dorsey Grant
Published: August 2020

Background

Intermittent preventive treatment of malaria during pregnancy (IPTp) with dihydroartemisinin-piperaquine (DP) significantly reduces the burden of malaria during pregnancy compared to sulfadoxine-pyrimethamine (SP), the current standard of care, but its impact on the incidence of malaria during infancy is unknown.

Methods

We conducted a double-blind randomized trial to compare the incidence of malaria during infancy among infants born to HIV-uninfected pregnant women who were randomized to monthly IPTp with either DP or SP. Infants were followed for all their medical care in a dedicated study clinic, and routine assessments were conducted every 4 weeks. At all visits, infants with fever and a positive thick blood smear were diagnosed and treated for malaria. The primary outcome was malaria incidence during the first 12 months of life. All analyses were done by modified intention to treat.

Results

Of the 782 women enrolled, 687 were followed through delivery from December 9, 2016, to December 5, 2017, resulting in 678 live births: 339 born to mothers randomized to SP and 339 born to those randomized to DP. Of these, 581 infants (85.7%) were followed up to 12 months of age. Overall, the incidence of malaria was lower among infants born to mothers randomized to DP compared to SP, but the difference was not statistically significant (1.71 vs 1.98 episodes per person-year, incidence rate ratio (IRR) 0.87, 95% confidence interval (CI) 0.73–1.03, p = 0.11). Stratifying by infant sex, IPTp with DP was associated with a lower incidence of malaria among male infants (IRR 0.75, 95% CI 0.58–0.98, p = 0.03) but not female infants (IRR 0.99, 95% CI 0.79–1.24, p = 0.93).

Conclusion

Despite the superiority of DP for IPTp, there was no evidence of a difference in malaria incidence during infancy in infants born to mothers who received DP compared to those born to mothers who received SP. Only male infants appeared to benefit from IPTp-DP suggesting that IPTp-DP may provide additional benefits beyond birth. Further research is needed to further explore the benefits of DP versus SP for IPTp on the health outcomes of infants.

Trial registration

ClinicalTrials.gov, NCT02793622. Registered on June 8, 2016.
Evaluation of nutraceutical properties of Leucaena leucocephala leaf pellets fed to goat kids infected with Haemonchus contortus
Authors: Marie-Magdeleine Carine , Ceriac Steve , Barde Dingamgoto Jesse , Minatchy Nathalie , Periacarpin Fred , Pommier Frederic , Calif Brigitte , Philibert Lucien , Bambou Jean-Christophe , Archimède Harry
Published: August 2020

Background

Leucaena leucocephala, as a shrub legume rich in condensed tannins, is a candidate for the integrated control of gastrointestinal parasitic pathogen nematodes. Here, we investigated the anthelmintic potential of the nutraceutical L. leucocephala, transformed into pellets, against Haemonchus contortus.

Results

Creole goat kids were fed an iso-nitrogenous diet of Dichantium hay with alfalfa pellets or Leucaena pellets or an alfalfa–leucaena mixture in varying ratios. The artificial infection of kids with H. contortus led to infection levels comparable to those observed in the farm environment on the basis of egg excretion. The anthelmintic activity of Leucaena, compared to alfalfa, was demonstrated by its potential to reduce egg excretions (1524 vs. 3651 eggs/g) and the larval development of excreted eggs (3.5 vs. 24%). This anthelmintic potential was reported when the ratio of Leucaena incorporation in the diet was high (50% DM intake). The voluntary dry matter intake (79.3 vs. 77.0 g Large Weigth0.75), the total tract dry matter digestion (0.585 vs. 0.620), and the growth (57.1 vs. 71.3 g/d) of kids fed Leucaena compared to alfalfa indicate that Leucaena has a high feed value. The Leucaena, even at the highest intakes in the diets, has not shown any signs of poisoning in kids.

Conclusions

Leucaena fulfilled the conditions to be a good nutraceutical, and pelleting is a good method for its use.
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

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

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

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

Background

This study investigated the role of fibrinogen-like protein 1 (FGL1) in regulating gefitinib resistance of PC9/GR non-small cell lung cancer (NSCLC).

Methods

The 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.

Results

FGL1 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.

Conclusions

FGL1 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

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

Background

Thioesters 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.

Results

Here, 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.

Conclusions

The 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

Background

Melanoma 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.

Methods

In 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.

Results

MiR-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.

Conclusion

Ourfindings 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

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

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.