bluuebunny/crossref_metadata_2025 huggingface dataset:
The Gnoseology of Catharsis
Authors:   Ufa State Institute of Arts named after Zagir Ismagilov, Verbitskaya Galina Ya.
Published: January 2021

A work of art induces the recipient towards cognition and creativity by means of experience of cathartic emotion and the pain of convulsion. Art is a guide for life creativity which helps people make a metaphysical effort for overcoming despair. As the result of the conducted research a model has been developed of the emergence, development and realization of cathartic emotion. All the elements of the model are inwardly interconnected and mutually connected and are in the condition of a dialogue between the polarities of manifestations, the intensity of emotion, in a cause-and-effect relationship, in the connection of the invariant with the variant parts. The content and realization of the levels of experiencing cathartic emotion are disclosed in the dialogue of polar opposite fundamental principles of existence: fear vs. joy, compassion vs. pleasure, death vs. revival. This is what comprises the content and semantic signification of catharsis. Art is not exhausted by cathartic impact in the sense of purification, the most important thing is in the emergence of the insurmountable desire for creative activity. In other words, it is the reception of a work of art, the effort and artistry, the disclosure of the best in oneself, the generation of the creative gift, the emergence of the necessity for creative activity. On the essential level catharsis presents the outcome of the conflict, the resolution of the contradiction by means of the author’s creative insight and the admission of the audience into the highest level of understanding and sensation of the conflicts of life. It is particularly an aesthetic attitude towards the world created by perception of art. The artistic experience creates the possibility of resolving the eternal contradictions-antinomies of life as an infinite process of search for solutions of insoluble problems and an alternate perception of life and its meaning.
Amateur Piano Music-Making and the Ufa Section of the Imperial Russian Musical Society
Authors:   Ufa State Institute of Arts named after Zagir Ismagilov, Garipova Ninel F.
Published: January 2021

The geographic position of Ufa, which in the early 19th century was a deep province, was not conducive to the development of musical culture. However, we must consider as an important element in its formation the active spread of household music-making and the wish of amateurs to participate in the city’s concert life. The “Society for Singing, Music and the Art of Drama” was founded in 1885 in Ufa following the wishes of the city residents. The twenty-year-long existence of the Society has left a considerable trace in the development of musical education and the exposure of the public to the academic genres of the art of piano performance; it played a significant role in the development of musical literacy and the musical hearing of the residents of Ufa. In virtue of a number of existing social reasons the Society was closed down, but following the request of the most educated part of the local nobility and intelligentsia the Ufa Section of the Imperial Russian Musical Society (IRMS). Having existed for only a few years, until the revolution of 1917, it was able to lead the art of music to a new, higher level. Professionals with a higher musical education were conducive to the further expansion of promotion of music with their concert performances and teaching lessons in the musical classes and enhanced the development of the art of professional music in Bashkiria.
The “Notebooks” of Leopold Mozart (“Die Notenbücher der Geschwister Mozart”) As a Specimen of Instructive Compositions
Authors:   Ufa State Institute of Arts named after Zagir Ismagilov, Alexeyeva Irina V. , Kirsanova Olga Yu.
Published: January 2021

One of the interesting realms of contemporary musicology is the issue of study and evaluation of the historical musical text of the early classical period, remote from us in terms of time. No less significant is the comprehension of its didactic potential, as well as the adaptation of scholarly results of research and their application in pedagogical practice. More relevant is the study of the techniques of interaction on the part of the performer with this musical text. The article acquaints the readers with the unique album for clavier, absent in Russian performing and instructive practice — “Die Notenbücher der Geschwister Mozart” in its original version. Its study presents the possibility to immerse into the specificity of the artistic content and pedagogical “secrets” of one of the opuses of the instructive direction reflecting the specificity of 18th century instrumental music-making. Analytical immersion into the musical text of the “Notebooks” (such is the version of the translation of the title of the analogous albums of J.S. Bach, according to Russian publishers) as a historical document of the epoch is aided by turning to its “intonational-lexical vocabulary” stipulated by culturological context. The album discloses the practical secrets of adaptation of the musical score form of notation in its transcription into two-lined form and demonstrated signs of the reduction of the musical text in correspondence with the peculiarities of keyboard instruments. For a present-day beginning performer, the creative potential of the pieces in the collection consists in the possibility of reverse unfolding of two-line keyboard music into an ensemble score the means of which have been fixated into the musical text.
The Musical Legacy of Sergei Rachmaninoff
Authors:   Center for Comprehensive Art Studies, Saratov State L.V. Sobinov Conservatory, Demchenko Alexander I.
Published: January 2021

“The musical legacy of Sergei Rachmaninoff” — this is the first lecture from the authorial cycle of Doctor of Arts, Professor Alexander Demchenko “The Classics of 20th Century Russian Music.” Its following sections will be dedicated to such composers as Igor Stravinsky, Sergei Prokofiev, Nikolai Myaskovsky, Dmitri Shostakovich, Aram Khachaturian, Georgiy Sviridov, odion Shchedrin and Alfred Schnittke. The lecture is supposed to include listening to a number of musical fragments chosen to give a general perception of the range of the composer’s artistic explorations. The preferential performance versions and durations of the corresponding musical fragments are given. The publication of the lecture is addressed to students and faculty members of conservatories, artistic institutions of higher education, as well as music colleges and high schools.
Biomechanical Study of a Tricompartmental Unloader Brace for Patellofemoral or Multicompartment Knee Osteoarthritis
Authors: McGibbon Chris A. , Brandon Scott , Bishop Emily L. , Cowper-Smith Chris , Biden Edmund N.
Published: January 2021

Objective: Off-loader knee braces have traditionally focused on redistributing loads away from either the medial or lateral tibiofemoral (TF) compartments. In this article, we study the potential of a novel “tricompartment unloader” (TCU) knee brace intended to simultaneously unload both the patellofemoral (PF) and TF joints during knee flexion. Three different models of the TCU brace are evaluated for their potential to unload the knee joint.

Methods: A sagittal plane model of the knee was used to compute PF and TF contact forces, patellar and quadriceps tendon forces, and forces in the anterior and posterior cruciate ligaments during a deep knee bend (DKB) test using motion analysis data from eight participants. Forces were computed for the observed (no brace) and simulated braced conditions. A sensitivity and validity analysis was conducted to determine the valid output range for the model, and Statistical Parameter Mapping was used to quantify the effectual region of the different TCU brace models.

Results: PF and TF joint force calculations were valid between ~0 and 100 degrees of flexion. All three simulated brace models significantly (p < 0.001) reduced predicted knee joint loads (by 30–50%) across all structures, at knee flexion angles >~30 degrees during DKB.

Conclusions: The TCU brace is predicted to reduce PF and TF knee joint contact loads during weight-bearing activity requiring knee flexion angles between 30 and 100 degrees; this effect may be clinically beneficial for pain reduction or rehabilitation from common knee injuries or joint disorders. Future work is needed to assess the range of possible clinical and prophylactic benefits of the TCU brace.
Measures of Interjoint Coordination Post-stroke Across Different Upper Limb Movement Tasks
Authors: Schwarz Anne , Veerbeek Janne M. , Held Jeremia P. O. , Buurke Jaap H. , Luft Andreas R.
Published: January 2021

Background:Deficits in interjoint coordination, such as the inability to move out of synergy, are frequent symptoms in stroke subjects with upper limb impairments that hinder them from regaining normal motor function. Kinematic measurements allow a fine-grained assessment of movement pathologies, thereby complementing clinical scales, like the Fugl–Meyer Motor Assessment of the Upper Extremity (FMMA-UE). The study goal was to investigate the effects of the performed task, the tested arm, the dominant affected hand, upper limb function, and age on spatiotemporal parameters of the elbow, shoulder, and trunk. The construct validity of the metrics was examined by relating them with each other, the FMMA-UE, and its arm section.

Methods:This is a cross-sectional observational study including chronic stroke patients with mild to moderate upper limb motor impairment. Kinematic measurements were taken using a wearable sensor suit while performing four movements with both upper limbs: (1) isolated shoulder flexion, (2) pointing, (3) reach-to-grasp a glass, and (4) key insertion. The kinematic parameters included the joint ranges of shoulder abduction/adduction, shoulder flexion/extension, and elbow flexion/extension; trunk displacement; shoulder–elbow correlation coefficient; median slope; and curve efficiency. The effects of the task and tested arm on the metrics were investigated using a mixed-model analysis. The validity of metrics compared to clinically measured interjoint coordination (FMMA-UE) was done by correlation analysis.

Results:Twenty-six subjects were included in the analysis. The movement task and tested arm showed significant effects (p< 0.05) on all kinematic parameters. Hand dominance resulted in significant effects on shoulder flexion/extension and curve efficiency. The level of upper limb function showed influences on curve efficiency and the factor age on median slope. Relations with the FMMA-UE revealed the strongest and significant correlation for curve efficiency (r= 0.75), followed by shoulder flexion/extension (r= 0.68), elbow flexion/extension (r= 0.53), and shoulder abduction/adduction (r= 0.49). Curve efficiency additionally correlated significantly with the arm subsection, focusing on synergistic control (r= 0.59).

Conclusion:The kinematic parameters of the upper limb after stroke were influenced largely by the task. These results underpin the necessity to assess different relevant functional movements close to real-world conditions rather than relying solely on clinical measures.

Study Registration:clinicaltrials.gov, identifier NCT03135093 and BASEC-ID 2016-02075.
ROS-Sensitive Nanoparticles Co-delivering Dexamethasone and CDMP-1 for the Treatment of Osteoarthritis Through Chondrogenic Differentiation Induction and Inflammation Inhibition
Authors: Wu Xiaodong , Li Pengpeng , Cheng Jian , Xu Qiang , Lu Beiji , Han Conghui , Huo Weiling
Published: January 2021

Objective: Osteoarthritis (OA) is a common subtype of arthritis. To date, treatment of OA focuses primarily on alleviating pain and improving joint function. The lack of a vascular system within synovial joints and the rapid removal of agents due to synovial exchange hinder continuous delivery of OA drugs. However, these obstacles are being addressed by promising nanoscale drugs.

Methods: We synthesize and assemble a hydrogen peroxide [H2O2, belongs to the category of active oxygen species (ROS)]-sensitive nanomicelle, which is loaded with the anti-inflammation drug dexamethasone and chondrogenic differentiation factor cartilage-derivedmor-phogeneticprotein-1. The micelle can induce bone marrow mesenchymal stem cells to repair cartilage while inhibiting joint inflammation.

Results: The prepared nanoparticles were of uniform size and displayed an obvious core-shell structure. Under H2O2 stimulation, the shell layer could be removed gradually. The drug-loaded micelle effectively inhibited proliferation of activated macrophages, induced macrophage apoptosis with an anti-inflammatory effect, and caused the BMSCs to differentiate into chondrocytes.

Conclusion: This work provides an experimental and theoretical basis for further development of a drug-loaded micelle in the healing of osteoarthritis.
Discovering the Potential of Dental Pulp Stem Cells for Corneal Endothelial Cell Production: A Proof of Concept
Authors: Bosch Begoña M. , Salero Enrique , Núñez-Toldrà Raquel , Sabater Alfonso L. , Gil F. J. , Perez Roman A.
Published: January 2021

Failure of corneal endothelium cell monolayer is the main cause leading to corneal transplantation. Autologous cell-based therapies are required to reconstruct in vitro the cell monolayer. Several strategies have been proposed using embryonic stem cells and induced pluripotent stem cells, although their use has ethical issues as well as limited clinical applications. For this purpose, we propose the use of dental pulp stem cells isolated from the third molars to form the corneal endothelium cell monolayer. We hypothesize that using dental pulp stem cells that share an embryological origin with corneal endothelial cells, as they both arise from the neural crest, may allow a direct differentiation process avoiding the use of reprogramming techniques, such as induced pluripotent stem cells. In this work, we report a two-step differentiation protocol, where dental pulp stem cells are derived into neural crest stem-like cells and, then, into corneal endothelial-like cells. Initially, for the first-step we used an adhesion culture and compared two initial cell sources: a direct formation from dental pulp stem cells with the differentiation from induced pluripotent stem cells. Results showed significantly higher levels of early stage marker AP2 for the dental pulp stem cells compared to induced pluripotent stem cells. In order to provide a better environment for neural crest stem cells generation, we performed a suspension method, which induced the formation of neurospheres. Results showed that neurosphere formation obtained the peak of neural crest stem cell markers expression after 4 days, showing overexpression of AP2, Nestin, and p75 markers, confirming the formation of neural crest stem-like cells. Furthermore, pluripotent markers Oct4, Nanog, and Sox2 were as well-upregulated in suspension culture. Neurospheres were then directly cultured in corneal endothelial conditioned medium for the second differentiation into corneal endothelial-like cells. Results showed the conversion of dental pulp stem cells into polygonal-like cells expressing higher levels of ZO-1, ATP1A1, COL4A2, and COL8A2 markers, providing a proof of the conversion into corneal endothelial-like cells. Therefore, our findings demonstrate that patient-derived dental pulp stem cells may represent an autologous cell source for corneal endothelial therapies that avoids actual transplantation limitations as well as reprogramming techniques.
On the Applicability of a Precursor Derived from Organic Waste Streams for Bacteria-Based Self-Healing Concrete
Authors: Rossi Emanuele , Vermeer Chris M. , Mors Renee , Kleerebezem Robbert , Copuroglu Oguzhan , Jonkers Henk M.
Published: January 2021

Bacteria-based self-healing concrete has the ability to heal cracks due to the bacterial conversion of incorporated organic compounds into calcium carbonate. Precipitates seal the cracks, theoretically increasing the service life of constructions. The aim of this paper is to propose a precursor for bacteria-based self-healing concrete derived from organic waste streams, produced is in line with the circular economy principle and ideally more affordable than other substrates. To verify the applicability of the proposed healing agent, some fundamental requirements of the proposed system are studied, such as its influence on functional properties, crack sealing capacity and evidence of bacterial activity in concrete.
Fas-Activated Serine/Threonine Kinase Governs Cardiac Mitochondrial Complex I Functional Integrity in Ischemia/Reperfusion Heart
Authors: Chen Xiyao , Hu Guangyu , Wang Yuanyuan , Li Congye , Zhang Fuyang
Published: January 2021

Cardiac energy homeostasis is strictly controlled by the mitochondrial complex-mediated respiration. In the heart, mitochondrial complex I is highly susceptible to functional and structural destroy after ischemia/reperfusion (I/R), thereby contributing to myocardial energy insufficiency and cardiomyocyte death. Fas-activated serine/threonine kinase (FASTK) is recently recognized as a key modulator of mitochondrial gene expression and respiration. However, the role of FASTK in cardiac I/R process is undetermined. Here, we show that FASTK expression was down-regulated in the post-I/R heart. The reactive oxygen species scavenger N-acetyl-L-cysteine reversed I/R-induced FASTK down-regulation. Genetic deletion of FASTK exacerbated I/R-induced cardiac dysfunction, enlarged myocardial infarct size, and increased cardiomyocyte apoptosis. Compared with the wild type control, the FASTK deficient heart exhibited a lower mRNA expression of NADH dehydrogenase subunit-6 (MTND6, a mitochondrial gene encoding a subunit of complex I) and was more vulnerable to I/R-associated complex I inactivation. Replenishment of FASTK expression via adenovirus-mediated gene delivery restored mitochondrial complex I activity and ameliorated cardiomyocyte death induced by I/R, whereas these beneficial effects were blocked by the co-treatment with rotenone, a specific complex I inhibitor. in vivo experiments further confirmed that cardiac overexpression of FASTK ameliorated I/R-related MTND6 down-regulation and mitochondrial complex I inactivation, thereby protecting the heart against I/R injury. Collectively, these data for the first time identify that the down-regulation of FASTK is a direct culprit behind the loss of mitochondrial complex I functional integrity and cardiac injury induced by I/R process. Targeting FASTK might be a promising and effective strategy for MI/R intervention.
Ubiquitination and the Proteasome as Drug Targets in Trypanosomatid Diseases
Authors: Bijlmakers Marie-José
Published: January 2021

The eukaryotic pathogens Trypanosoma brucei, Trypanosoma cruzi and Leishmania are responsible for debilitating diseases that affect millions of people worldwide. The numbers of drugs available to treat these diseases, Human African Trypanosomiasis, Chagas' disease and Leishmaniasis are very limited and existing treatments have substantial shortcomings in delivery method, efficacy and safety. The identification and validation of novel drug targets opens up new opportunities for the discovery of therapeutic drugs with better efficacy and safety profiles. Here, the potential of targeting the ubiquitin-proteasome system in these parasites is reviewed. Ubiquitination is the posttranslational attachment of one or more ubiquitin proteins to substrates, an essential eukaryotic mechanism that regulates a wide variety of cellular processes in many different ways. The best studied of these is the delivery of ubiquitinated substrates for degradation to the proteasome, the major cellular protease. However, ubiquitination can also regulate substrates in proteasome-independent ways, and proteasomes can degrade proteins to some extent in ubiquitin-independent ways. Because of these widespread roles, both ubiquitination and proteasomal degradation are essential for the viability of eukaryotes and the proteins that mediate these processes are therefore attractive drug targets in trypanosomatids. Here, the current understanding of these processes in trypanosomatids is reviewed. Furthermore, significant recent progress in the development of trypanosomatid-selective proteasome inhibitors that cure mouse models of trypanosomatid infections is presented. In addition, the targeting of the key enzyme in ubiquitination, the ubiquitin E1 UBA1, is discussed as an alternative strategy. Important differences between human and trypanosomatid UBA1s in susceptibility to inhibitors predicts that the selective targeting of these enzymes in trypanosomatids may also be feasible. Finally, it is proposed that activating enzymes of the ubiquitin-like proteins SUMO and NEDD8 may represent drug targets in these trypanosomatids as well.
The CRISPR-Cas Mechanism for Adaptive Immunity and Alternate Bacterial Functions Fuels Diverse Biotechnologies
Authors: Newsom Sydney , Parameshwaran Hari Priya , Martin Lindsie , Rajan Rakhi
Published: January 2021

Bacterial and archaeal CRISPR-Cas systems offer adaptive immune protection against foreign mobile genetic elements (MGEs). This function is regulated by sequence specific binding of CRISPR RNA (crRNA) to target DNA/RNA, with an additional requirement of a flanking DNA motif called the protospacer adjacent motif (PAM) in certain CRISPR systems. In this review, we discuss how the same fundamental mechanism of RNA-DNA and/or RNA-RNA complementarity is utilized by bacteria to regulate two distinct functions: to ward off intruding genetic materials and to modulate diverse physiological functions. The best documented examples of alternate functions are bacterial virulence, biofilm formation, adherence, programmed cell death, and quorum sensing. While extensive complementarity between the crRNA and the targeted DNA and/or RNA seems to constitute an efficient phage protection system, partial complementarity seems to be the key for several of the characterized alternate functions. Cas proteins are also involved in sequence-specific and non-specific RNA cleavage and control of transcriptional regulator expression, the mechanisms of which are still elusive. Over the past decade, the mechanisms of RNA-guided targeting and auxiliary functions of several Cas proteins have been transformed into powerful gene editing and biotechnological tools. We provide a synopsis of CRISPR technologies in this review. Even with the abundant mechanistic insights and biotechnology tools that are currently available, the discovery of new and diverse CRISPR types holds promise for future technological innovations, which will pave the way for precision genome medicine.
A Novel Marmoset (Callithrix jacchus) Model of Human Inhalational Q Fever
Authors: Nelson Michelle , Salguero Francisco J. , Hunter Laura , Atkins Timothy P.
Published: January 2021

Common marmosets (Callithrix jacchus)were shown to be susceptible to inhalational infection withCoxiella burnetii, in a dose-dependent manner, producing a disease similar to human Q fever, characterized by a resolving febrile response. Illness was also associated with weight loss, liver enzyme dysfunction, characteristic cellular activation, circulating INF-γand bacteraemia. ViableC. burnetiiwas recovered from various tissues during disease and from 75% of the animal’s lungs on 28 days post challenge, when there were no overt clinical features of disease but there was histological evidence of macrophage and lymphocyte infiltration into the lung resulting in granulomatous alveolitis. Taken together, these features of disease progression, physiology and bacterial spread appear to be consistent with human disease and therefore the common marmoset can be considered as a suitable model for studies on the pathogenesis or the development of medical counter measures of inhalational Q fever.
Genomic Analysis of Korean Patient With Microcephaly
Authors: Lee Jiwon , Park Jong Eun , Lee Chung , Kim Ah Reum , Kim Byung Joon , Park Woong-Yang , Ki Chang-Seok , Lee Jeehun
Published: January 2021

Microcephaly is a prevalent phenotype in patients with neurodevelopmental problems, often with genetic causes. We comprehensively investigated the clinical phenotypes and genetic background of microcephaly in 40 Korean patients. We analyzed their clinical phenotypes and radiologic images and conducted whole exome sequencing (WES) and analysis of copy number variation (CNV). Infantile hypotonia and developmental delay were present in all patients. Thirty-four patients (85%) showed primary microcephaly. The diagnostic yield from the WES and CNV analyses was 47.5%. With WES, we detected pathogenic or likely pathogenic variants that were previously associated with microcephaly in 12 patients (30%); nine of these were de novo variants with autosomal dominant inheritance. Two unrelated patients had mutations in the KMT2A gene. In 10 other patients, we found mutations in the GNB1, GNAO1, TCF4, ASXL1, SMC1A, VPS13B, ACTG1, EP300, and KMT2D genes. Seven patients (17.5%) were diagnosed with pathogenic CNVs. Korean patients with microcephaly show a genetic spectrum that is different from that of patients with microcephaly of other ethnicities. WES along with CNV analysis represents an effective approach for diagnosis of the underlying causes of microcephaly.
Allelic Diversity, Structural Analysis, and Genome-Wide Association Study (GWAS) for Yield and Related Traits Using Unexplored Common Bean (Phaseolus vulgaris L.) Germplasm From Western Himalayas
Authors: Mir Reyazul Rouf , Choudhary Neeraj , Bawa Vanya , Jan Sofora , Singh Bikram , Bhat Mohd Ashraf , Paliwal Rajneesh , Kumar Ajay , Chitikineni Annapurna , Thudi Mahendar , Varshney Rajeev Kumar
Published: January 2021

The north-western Indian Himalayas possesses vast diversity in common bean germplasm due to several years of natural adaptation and farmer’s selection. Systematic efforts have been made for the first time for the characterization and use of this huge diversity for the identification of genes/quantitative trait loci (QTLs) for yield and yield-contributing traits in common bean in India. A core set of 96 diverse common bean genotypes was characterized using 91 genome-wide genomic and genic simple sequence repeat (SSR) markers. The study of genetic diversity led to the identification of 691 alleles ranging from 2 to 21 with an average of 7.59 alleles/locus. The gene diversity (expected heterozygosity,He) varied from 0.31 to 0.93 with an average of 0.73. As expected, the genic SSR markers detected less allelic diversity than the random genomic SSR markers. The traditional clustering and Bayesian clustering (structural analysis) analyses led to a clear cut separation of a core set of 96 genotypes into two distinct groups based on their gene pools (Mesoamerican and Andean genotypes). Genome-wide association mapping for pods/plant, seeds/pod, seed weight, and yield/plant led to the identification of 39 significant marker–trait associations (MTAs) including 15 major, 15 stable, and 13 both major and stable MTAs. Out of 39 MTAs detected, 29 were new MTAs reported for the first time, whereas the remaining 10 MTAs were already identified in earlier studies and therefore declared as validation of earlier results. A set of seven markers was such, which were found to be associated with multiple (two to four) different traits. The important MTAs will be used for common bean molecular breeding programs worldwide for enhancing common bean yield.
A Seven-Long Non-coding RNA Signature Improves Prognosis Prediction of Lung Adenocarcinoma: An Integrated Competing Endogenous RNA Network Analysis
Authors: Li Rang , Han Kedong , Xu Dehua , Chen Xiaolin , Lan Shujin , Liao Yuanjun , Sun Shengnan , Rao Shaoqi
Published: January 2021

Early and precise prediction is an important way to reduce the poor prognosis of lung adenocarcinoma (LUAD) patients. Nevertheless, the widely used tumor, node, and metastasis (TNM) staging system based on anatomical information only often could not achieve adequate performance on foreseeing the prognosis of LUAD patients. This study thus aimed to examine whether the long non-coding RNAs (lncRNAs), known highly involved in the tumorigenesis of LUAD through the competing endogenous RNAs (ceRNAs) mechanism, could provide additional information to improve prognosis prediction of LUAD patients. To prove the hypothesis, a dataset consisting of both RNA sequencing data and clinical pathological data, obtained from The Cancer Genome Atlas (TCGA) database, was analyzed. Then, differentially expressed RNAs (DElncRNAs, DEmiRNAs, and DEmRNAs) were identified and a lncRNA–miRNA–mRNA ceRNA network was constructed based on those differentially expressed RNAs. Functional enrichment analysis revealed that this ceRNA network was highly enriched in some cancer-associated signaling pathways. Next, lasso-Cox model was run 1,000 times to recognize the potential survival-related combinations of the candidate lncRNAs in the ceRNA network, followed by the “best subset selection” to further optimize these lncRNA-based combinations, and a seven-lncRNA prognostic signature with the best performance was determined. Based on the median risk score, LUAD patients could be well distinguished into high-/low-risk subgroups. The Kaplan–Meier survival curve showed that LUAD patients in the high-risk group had significantly shorter overall survival than those in the low-risk group (log-rank test P = 4.52 × 10–9). The ROC curve indicated that the clinical genomic model including both the TNM staging system and the signature had a superior performance in predicting the patients’ overall survival compared to the clinical model with the TNM staging system only. Further stratification analysis suggested that the signature could work well in the different strata of the stage, gender, or age, rendering it to be a wide application. Finally, a ceRNA subnetwork related to the signature was extracted, demonstrating its high involvement in the tumorigenesis mechanism of LUAD. In conclusion, the present study established a lncRNA-based molecular signature, which can significantly improve prognosis prediction for LUAD patients.
Targeted Gene Editing in Porcine Spermatogonia
Authors: Webster Dennis , Bondareva Alla , Solin Staci , Goldsmith Taylor , Su Lin , Lara Nathalia de Lima e Martins , Carlson Daniel F. , Dobrinski Ina
Published: January 2021

To study the pathophysiology of human diseases, develop innovative treatments, and refine approaches for regenerative medicine require appropriate preclinical models. Pigs share physiologic and anatomic characteristics with humans and are genetically more similar to humans than are mice. Genetically modified pigs are essential where rodent models do not mimic the human disease phenotype. The male germline stem cell or spermatogonial stem cell (SSC) is unique; it is the only cell type in an adult male that divides and contributes genes to future generations, making it an ideal target for genetic modification. Here we report that CRISPR/Cas9 ribonucleoprotein (RNP)-mediated gene editing in porcine spermatogonia that include SSCs is significantly more efficient than previously reported editing with TALENs and allows precise gene editing by homology directed repair (HDR). We also established homology-mediated end joining (HMEJ) as a second approach to targeted gene editing to enable introduction of larger transgenes and/or humanizing parts of the pig genome for disease modeling or regenerative medicine. In summary, the approaches established in the current study result in efficient targeted genome editing in porcine germ cells for precise replication of human disease alleles.
Profiling of MicroRNAs and Their Targets in Roots and Shoots Reveals a Potential MiRNA-Mediated Interaction Network in Response to Phosphate Deficiency in the Forestry Tree Betula luminifera
Authors: Zhang Junhong , Lin Yan , Wu Fangmin , Zhang Yuting , Cheng Longjun , Huang Menghui , Tong Zaikang
Published: January 2021

Inorganic phosphate (Pi) is often lacking in natural and agro-climatic environments, which impedes the growth of economically important woody species. Plants have developed strategies to cope with low Pi (LP) availability. MicroRNAs (miRNAs) play important roles in responses to abiotic stresses, including nutrition stress, by regulating target gene expression. However, the miRNA-mediated regulation of these adaptive responses and their underlying coordinating signals are still poorly understood in forestry trees such as Betula luminifera. Transcriptomic libraries, small RNA (sRNA) libraries, and a mixed degradome cDNA library of B. luminifera roots and shoots treated under LP and normal conditions (CK) were constructed and sequenced using next-generation deep sequencing. A comprehensive B. luminifera transcriptome derived from its roots and shoots was constructed, and a total of 76,899 unigenes were generated. Analysis of the transcriptome identified 8,095 and 5,584 differentially expressed genes in roots and shoots, respectively, under LP conditions. sRNA sequencing analyses indicated that 66 and 60 miRNAs were differentially expressed in roots and shoots, respectively, under LP conditions. A total of 109 and 112 miRNA–target pairs were further validated in the roots and shoots, respectively, using degradome sequencing. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis of differential miRNA targets indicated that the “ascorbate and aldarate metabolism” pathway responded to LP, suggesting miRNA-target pairs might participating in the removing of reactive oxidative species under LP stress. Moreover, a putative network of miRNA–target interactions involved in responses to LP stress in B. luminifera is proposed. Taken together, these findings provide useful information to decipher miRNA functions and establish a framework for exploring P signaling networks regulated by miRNAs in B. luminifera and other woody plants. It may provide new insights into the genetic engineering of high use efficiency of Pi in forestry trees.
Reduced CD5 on CD8+ T Cells in Tumors but Not Lymphoid Organs Is Associated With Increased Activation and Effector Function
Authors: Alotaibi Faizah , Vincent Mark , Min Wei-Ping , Koropatnick James
Published: January 2021

CD5, a member of the scavenger receptor cysteine-rich superfamily, is a marker for T cells and a subset of B cells (B1a). CD5 associates with T-cell and B-cell receptors and increased CD5 is an indication of B cell activation. In tumor-infiltrating lymphocytes (TILs) isolated from lung cancer patients, CD5 levels were negatively correlated with anti-tumor activity and tumor‐mediated activation-induced T cell death, suggesting that CD5 could impair activation of anti-tumor T cells. We determined CD5 levels in T cell subsets in different organs in mice bearing syngeneic 4T1 breast tumor homografts and assessed the relationship between CD5 and increased T cell activation and effector function by flow cytometry. We report that T cell CD5 levels were higher in CD4+T cells than in CD8+T cells in 4T1 tumor-bearing mice, and that high CD5 levels on CD4+T cells were maintained in peripheral organs (spleen and lymph nodes). However, both CD4+and CD8+T cells recruited to tumors had reduced CD5 compared to CD4+and CD8+T cells in peripheral organs. In addition, CD5high/CD4+T cells and CD5high/CD8+T cells from peripheral organs exhibited higher levels of activation and associated effector function compared to CD5low/CD4+T cell and CD5low/CD8+T cell from the same organs. Interestingly, CD8+T cells among TILs and downregulated CD5 were activated to a higher level, with concomitantly increased effector function markers, than CD8+/CD5highTILs. Thus, differential CD5 levels among T cells in tumors and lymphoid organs can be associated with different levels of T cell activation and effector function, suggesting that CD5 may be a therapeutic target for immunotherapeutic activation in cancer therapy.
Key Components of Inflammasome and Pyroptosis Pathways Are Deficient in Canines and Felines, Possibly Affecting Their Response to SARS-CoV-2 Infection
Authors: Cui Haoran , Zhang Leiliang
Published: January 2021

SARS-CoV-2 causes the ongoing COVID-19 pandemic. Natural SARS-COV-2 infection has been detected in dogs, cats and tigers. However, the symptoms in canines and felines were mild. The underlying mechanisms are unknown. Excessive activation of inflammasome pathways can trigger cytokine storm and severe damage to host. In current study, we performed a comparative genomics study of key components of inflammasome and pyroptosis pathways in dogs, cats and tigers. Cats and tigers do not have AIM2 and NLRP1. Dogs do not contain AIM2, and encode a short form of NLRC4. The activation sites in GSDMB were absent in dogs, cats and tigers, while GSDME activation sites in cats and tigers were abolished. We propose that deficiencies of inflammasome and pyroptosis pathways might provide an evolutionary advantage against SARS-CoV-2 by reducing cytokine storm-induced host damage. Our findings will shed important lights on the mild symptoms in canines and felines infected with SARS-CoV-2.
IL-21–Deficient T Follicular Helper Cells Support B Cell Responses Through IL-27 in Patients With Chronic Hepatitis B
Authors: Khanam Arshi , Ayithan Natarajan , Tang Lydia , Poonia Bhawna , Kottilil Shyam
Published: January 2021

Chronic Hepatitis B (CHB) affects over 350 million people worldwide. Current treatment does result in reduced complications; however, a cure (development of antibodies to the S antigen) is not achieved, requiring life-long therapy. Humoral responses contribute to viral elimination by secreting neutralizing antibodies; though, effective induction of humoral immunity require CD4T cell differentiation into T follicular helper (TFH) cells that support B cell response through interleukin-21 (IL-21). In CHB, mechanism of TFH-B interactions is seldom described. During CHB, TFH cells are defective in producing IL-21 in response to hepatitis B surface antigen (HBsAg). However, regardless of low IL-21, TFH cells efficiently support B cell responses by producing interleukin-27 (IL-27), which directs the formation of plasmablasts and plasma cells from memory and naïve B cells by enhancing B lymphocyte-induced maturation protein-1. IL-27 not only improved total antibody production but HBsAg-specific IgG and IgM secretion that are essential for viral clearance. Importantly, IL-27+TFH cells were significantly associated with HBV DNA reduction. Therefore, these findings imply a novel mechanism of TFH mediated B cell help in CHB and suggest that IL-27 effectively compensate the function of IL-21 by supporting TFH-B cell function, required for protective antibody response and may contribute to viral clearance by providing potential target for achieving a functional cure.
A Novel Branched DNA-Based Flowcytometric Method for Single-Cell Characterization of Gene Therapy Products and Expression of Therapeutic Genes
Authors: Garcia-Perez Laura , van Eggermond Marja C.J.A. , Maietta Elisa , van der Hoorn Marie-Louise P. , Pike-Overzet Karin , Staal Frank J. T.
Published: January 2021

Many preclinical and clinical studies of hematopoietic stem cell-based gene therapy (GT) are based on the use of lentiviruses as the vector of choice. Assessment of the vector titer and transduction efficiency of the cell product is critical for these studies. Efficacy and safety of the modified cell product are commonly determined by assessing the vector copy number (VCN) using qPCR. However, this optimized and well-established method in the GT field is based on bulk population averages, which can lead to misinterpretation of the actual VCN per transduced cell. Therefore, we introduce here a single cell-based method that allows to unmask cellular heterogeneity in the GT product, even when antibodies are not available. We use Invitrogen’s flow cytometry-based PrimeFlow™ RNA Assay with customized probes to determine transduction efficiency of transgenes of interest, promoter strength, and the cellular heterogeneity of murine and human stem cells. The assay has good specificity and sensitivity to detect the transgenes, as shown by the high correlations between PrimeFlow™-positive cells and the VCN. Differences in promoter strengths can readily be detected by differences in percentages and fluorescence intensity. Hence, we show a customizable method that allows to determine the number of transduced cells and the actual VCN per transduced cell in a GT product. The assay is suitable for all therapeutic genes for which antibodies are not available or too cumbersome for routine flow cytometry. The method also allows co-staining of surface markers to analyze differential transduction efficiencies in subpopulations of target cells.
Targeting Indoleamine 2,3-Dioxygenase in Cancer Models Using the Novel Small Molecule Inhibitor NTRC 3883-0
Authors: Grobben Yvonne , de Man Jos , van Doornmalen Antoon M. , Muller Michelle , Willemsen-Seegers Nicole , Vu-Pham Diep , Mulder Winfried R. , Prinsen Martine B. W. , de Wit Joeri , Sterrenburg Jan Gerard , van Cauter Freek , den Ouden Judith E. , van Altena Anne M. , Massuger Leon F. , Uitdehaag Joost C. M. , Buijsman Rogier C. , Zaman Guido J. R.
Published: January 2021

Indoleamine 2,3-dioxygenase (IDO1) is a key regulator of immune suppression by catalyzing the oxidation of L-tryptophan. IDO1 expression has been related to poor prognosis in several cancers and to resistance to checkpoint immunotherapies. We describe the characterization of a novel small molecule IDO1 inhibitor, NTRC 3883-0, in a panel of biochemical and cell-based assays, and various cancer models. NTRC 3883-0 released the inhibitory effect of IDO1 on CD8-positive T cell proliferation in co-cultures of IDO1-overexpressing cells with healthy donor lymphocytes, demonstrating its immune modulatory activity. In a syngeneic mouse model using IDO1-overexpressing B16F10 melanoma cells, NTRC 3883-0 effectively counteracted the IDO1-induced modulation of L-tryptophan and L-kynurenine levels, demonstrating its in vivo target modulation. Finally, we studied the expression and activity of IDO1 in primary cell cultures established from the malignant ascites of ovarian cancer patients. In these cultures, IDO1 expression was induced upon stimulation with IFNγ, and its activity could be inhibited by NTRC 3883-0. Based on these results, we propose the use of ascites cell-based functional assays for future patient stratification. Our results are discussed in light of the recent discontinuation of clinical trials of more advanced IDO1 inhibitors and the reconsideration of IDO1 as a valid drug target.
Distinct Features and Functions of Systemic and Mucosal Humoral Immunity Among SARS-CoV-2 Convalescent Individuals
Authors: Butler Savannah E. , Crowley Andrew R. , Natarajan Harini , Xu Shiwei , Weiner Joshua A. , Bobak Carly A. , Mattox Daniel E. , Lee Jiwon , Wieland-Alter Wendy , Connor Ruth I. , Wright Peter F. , Ackerman Margaret E.
Published: January 2021

Understanding humoral immune responses to SARS-CoV-2 infection will play a critical role in the development of vaccines and antibody-based interventions. We report systemic and mucosal antibody responses in convalescent individuals who experienced varying severity of disease. Whereas assessment of neutralization and antibody-mediated effector functions revealed polyfunctional antibody responses in serum, only robust neutralization and phagocytosis were apparent in nasal wash samples. Serum neutralization and effector functions correlated with systemic SARS-CoV-2-specific IgG response magnitude, while mucosal neutralization was associated with nasal SARS-CoV-2-specific IgA. Antibody depletion experiments support the mechanistic relevance of these correlations. Associations between nasal IgA responses, virus neutralization at the mucosa, and less severe disease suggest the importance of assessing mucosal immunity in larger natural infection cohorts. Further characterization of antibody responses at the portal of entry may define their ability to contribute to protection from infection or reduced risk of hospitalization, informing public health assessment strategies and vaccine development efforts.
Prediction of Rubber Fiber Concrete Strength Using Extreme Learning Machine
Authors: Zhang Jingkui , Xu Juncai , Liu Changshun , Zheng Ji
Published: January 2021

The conventional design method of concrete mix ratio relies on a large number of tests for trial mixing and optimization, and the workload is massive. It is challenging to cope with today's diverse raw materials and the concrete's specific performance to fit modern concrete development. To innovate the design method of concrete mix ratio and effectively use the various complex novel raw materials, the traditional mix ratio test method can be replaced with the intelligent optimization algorithm, and the concrete performance prediction can be realized rapidly and accurately. The mixed ratio of the rubber fiber concrete was designed with its 28-day strength test. Then the range and variance analysis of the orthogonal test results were carried out to determine the optimal mix ratio and its influencing factors. A data set containing 114 sets of valid test data was collected by combining the rubber concrete mix test data published in recent years. Based on this data set, there are six influencing factors; rubber content, rubber particle size, and polypropylene fiber content are considered as the input variables, and the 28-day concrete compression, splitting tensile, and flexural strength are considered as the output variables. A strength prediction model of rubber fiber concrete is established based on the extreme learning machine (ELM). For verifying the ELM prediction model's performance, this article has conducted a comparison experiment between this model and other intelligent algorithm models. The results show that the model has the advantages of high accuracy and high generalization ability compared with other algorithm models such as conventional neural networks. It can be used as an effective method for predicting concrete performance. The method allows for the innovation and development of concrete mixing technology.
Cryopreservation and Resuscitation of Natural Aquatic Prokaryotic Communities
Authors: Rain-Franco Angel , de Moraes Guilherme Pavan , Beier Sara
Published: January 2021

Experimental reproducibility in aquatic microbial ecology is critical to predict the dynamics of microbial communities. However, controlling the initial composition of naturally occurring microbial communities that will be used as the inoculum in experimental setups is challenging, because a proper method for the preservation of those communities is lacking. To provide a feasible method for preservation and resuscitation of natural aquatic prokaryote assemblages, we developed a cryopreservation procedure applied to natural aquatic prokaryotic communities. We studied the impact of inoculum size, processing time, and storage time on the success of resuscitation. We further assessed the effect of different growth media supplemented with dissolved organic matter (DOM) prepared from naturally occurring microorganisms on the recovery of the initially cryopreserved communities obtained from two sites that have contrasting trophic status and environmental heterogeneity. Our results demonstrated that the variability of the resuscitation process among replicates decreased with increasing inoculum size. The degree of similarity between initial and resuscitated communities was influenced by both the growth medium and origin of the community. We further demonstrated that depending on the inoculum source, 45–72% of the abundant species in the initially natural microbial communities could be detected as viable cells after cryopreservation. Processing time and long-term storage up to 12 months did not significantly influence the community composition after resuscitation. However, based on our results, we recommend keeping handling time to a minimum and ensure identical incubation conditions for repeated resuscitations from cryo-preserved aliquots at different time points. Given our results, we recommend cryopreservation as a promising tool to advance experimental research in the field of microbial ecology.