bluuebunny/crossref_metadata_2025_split huggingface dataset:
Development of Scaffold-free 3D Cholangiocyte Organoids to Study the Progression of Primary Sclerosing Cholangitis
Authors: Zhang Wenjun , Kyritsi Konstantina , Isidan Abdulkadir , Park Yujin , Li Ping , Cross-Najafi Arthur A , Lopez Kevin , Kennedy Lindsey , Sato Keisaku , Glaser Shannon , Francis Heather , Alpini Gianfranco , Ekser Burcin 
Published: October 2022

ABSTRACTOrganoids are novelin vitromodels to study intercellular crosstalk between the different types of cells in the pathophysiology of disease. To better understand the underlying mechanisms driving the progression of primary sclerosing cholangitis (PSC), we developed scaffold-free multi-cellular 3D cholangiocyte organoids (3D-CHO) using ‘primary’ liver cell lines derived from normal and PSC patients. Human liver samples from healthy donors and late-stage PSC patients were used to isolate ‘primary’ cholangiocytes (EPCAM+/CK-19+), liver endothelial cells (LECs, CD31+), and hepatic stellate cells (HSCs, CD31−/CD68−/Desmin+/Vitamin A+). 3D-CHOs were formed using cholangiocytes:HSCs:LECs and kept viable for up to 1 month. Isolated primary cell lines and 3D-CHOs were further characterized by immunofluorescence (IF), qRT-PCR, and transmission electron microscopy. Gene expressions for cholangiocytes (SOX9, CFTR, EpCAM, AE, SCT, SCTR), fibrosis (ACTA2, COL1A1, DESMIN, TGFβ1), angiogenesis (PECAM, VEGF, CDH5, vWF), and inflammation (IL-6, TNF-α) confirmed PSC phenotypes of 3D-CHOs. Since cholangiocytes develop a neuroendocrine phenotype and express neuromodulators, confocal-IF demonstrated that neurokinin-1 receptor (NK-1R, expressed by cholangiocytes and upregulated in PSC), was localized within CK-19+cholangiocytes. Moreover, 3D-CHOs from PSC patients confirmed PSC phenotypes with upregulated NK-1R, tachykinin precursor 1, and downregulated membrane metalloendopeptidase. Our viable scaffold-free multiple-cell 3D-CHOs showed superiority as anin vitromodel in mimicking PSCin vivophenotypes compared to 2D cell culture, which can be used in PSC disease-related research.
Fever temperatures modulate intraprotein dynamics and enhance the binding affinity between monoclonal antibodies and the Spike protein from SARS-CoV-2
Authors: Kim Dong Gun , Kim Hak Sung , Choi Yoonjoo , Stan Razvan C. 
Published: October 2022

AbstractFever is a typical symptom of most infectious diseases. While prolonged fever may be clinically undesirable, mild reversible fever (< 39°C, 312K) can potentiate the immune responses against pathogens. Here, using molecular dynamics, we investigated the effect of febrile temperatures (38°C to 40°C, 311K to 313K) on the immune complexes formed by the SARS-CoV-2 spike protein with two neutralizing antibodies. We found that, at mild fever temperatures (311-312K), the binding affinities of the two antibodies improve when compared to the physiological body temperature (37°C, 310K). Furthermore, only at 312K, antibodies exert distinct mechanical effects on the receptor binding domains of the spike protein that may hinder SARS-CoV-2 infectivity. Enhanced antibody binding affinity may thus be obtained using appropriate temperature conditions.
Novel estimators for family-based genome-wide association studies increase power and robustness
Authors: Guan Junming , Nehzati Seyed Moeen , Benjamin Daniel J. , Young Alexander I. 
Published: October 2022

AbstractA goal of genome-wide association studies (GWASs) is to estimate the causal effects of alleles carried by an individual on that individual (‘direct genetic effects’). Typical GWAS designs, however, are susceptible to confounding due to gene-environment correlation and non-random mating (population stratification and assortative mating). Family-based GWAS, in contrast, is robust to such confounding since it uses random, within-family genetic variation. When both parents are genotyped, a regression controlling for parental genotype provides the most powerful approach. However, parental genotypes are often missing. We have previously shown that imputing the genotypes of missing parent(s) can increase power for estimation of direct genetic effects over using genetic differences between siblings. We extend the imputation method, which previously only applied to samples with at least one genotyped sibling or parent, to ‘singletons’ (individuals without any genotyped relatives). By including singletons, the effective sample size for estimation of direct effects can be increased by up to 50%. We apply this method to 408,254 ‘White British’ individuals from the UK Biobank, obtaining an effective sample size increase of between 25% and 43% (depending upon phenotype) by including 368,629 singletons. While this approach maximizes power, it can be biased when there is strong population structure. We therefore introduce an imputation based estimator that is robust to population structure and more powerful than other robust estimators. We implement our estimators in the software package snipar using an efficient linear-mixed model (LMM) specified by a sparse genetic relatedness matrix. We examine the bias and variance of different family-based and standard GWAS estimators theoretically and in simulations with differing levels of population structure, enabling researchers to choose the appropriate approach depending on their research goals.
Capturing differences in the regulation of LRRK2 dynamics and conformational states by small molecule kinase inhibitors
Authors: Weng Jui-Hung , Ma Wen , Wu Jian , Silletti Steve , McCammon J. Andrew , Taylor Susan 
Published: October 2022

AbstractMutations in the human leucine rich repeat protein kinase-2 (LRRK2) create risk factors for Parkinson’s Disease, and pathological functions of LRRK2 are often correlated with aberrant kinase activity. Past research has focused on developing selective LRRK2 kinase inhibitors. We showed previously that in addition to influencing intrinsic kinase activity, the global conformation of the LRRK2 protein plays a vital role in regulating LRRK2 signaling pathways. Deciphering the allosteric regulation in LRRK2 provides novel strategies for drug discovery. In this study, we combined enhanced sampling simulations with HDX-MS to analyze the inhibitor-induced dynamic changes and the allosteric communications in the C-terminal half of LRRK2, LRRK2RCKW. We find that a type I inhibitor (MLi-2) locks the kinase into a closed, active-like configuration, whereas a type II inhibitor (Rebastinib) shifts the kinase to an open, inactive configuration. While both type I and type II inhibitors reduce the kinase activity effectively, they have distinct effects on the LRRK2 conformational dynamics. Specifically, binding of MLi-2 stabilizes the kinase domain in a closed conformation and reduces the global dynamics of LRRK2RCKW, leading to a more compact LRRK2RCKWstructure. In contrast, binding of Rebastinib stabilizes an open conformation where communication between the N- and C-lobe is severed, which promotes a more extended LRRK2RCKWstructure. Rebastinib, based on HDX-MS, creates a more dynamic kinase domain especially at domain interfaces associated with the C-lobe. Our results also reveal the importance of the Dk-helix, which plays a crucial role in propagating communication between the kinase domain and the GTPase domain.
YAP condensates are highly organized hubs for YAP/TEAD transcription
Authors: Hao Siyuan , Fuehrer Hannah , Flores Eduardo , Demmerle Justin , Lippincott-Schwartz Jennifer , Liu Zhe , Sukenik Shahar , Cai Danfeng 
Published: October 2022

SummaryYAP/TEAD signaling is essential for organismal development, cell proliferation, and cancer progression. As a transcriptional coactivator, how YAP activates its downstream target genes is incompletely understood. YAP forms biomolecular condensates in response to hyperosmotic stress, concentrating transcription-related factors to activate downstream target genes. However, whether YAP forms condensates under other signals, how YAP condensates organize and function, and how YAP condensates activate transcription in general are unknown. Here, we report that endogenous YAP forms sub-micron scale condensates in response to Hippo pathway regulation and actin cytoskeletal tension. The transcription factor TEAD1 actively stabilizes YAP condensates, which also recruit BRD4, a coactivator that is enriched at active enhancers. Using single molecule tracking, we found that YAP condensates slowed YAP diffusion within condensate boundaries, a possible mechanism for promoting YAP target search. These results reveal that YAP condensate formation is a highly regulated process that is critical for YAP/TEAD target gene expression.
A HUMANIZED FCRN TRANSGENIC MOUSE FOR PRECLINICAL PHARMACOKINETICS STUDIES
Authors: Conner Christopher M. , van Fossan Don , Read Kristen , Cowley Dale O. , Alvarez Oscar , Xu Shannon (Xiang-Ru) , Webb David R. , Jarnagin Kurt 
Published: October 2022

ABSTRACTMonoclonal antibodies (mAbs) are one of the fastest-growing classes of drugs and have been approved to treat several diseases, including cancers and autoimmune disorders. Preclinical pharmacokinetics studies are performed to determine the therapeutically meaningful dosages and efficacy of candidate drugs. These studies are typically performed in non-human primates; however, using primates is costly and raises ethical considerations. As a result, rodent models that better mimic human-like pharmacokinetics have been generated and remain an area of active investigation.Pharmacokinetic characteristics of a candidate drug, such as half-life, are partly controlled by antibody binding to the human neonatal receptor hFCRN. Due to the abnormally high binding of human antibodies to mouse FCRN, traditional laboratory rodents do not accurately model the pharmacokinetics of human mAbs. In response, humanized rodents expressing hFCRNhave been generated. However, these models generally use large inserts randomly integrated into the mouse genome. Here, we report the production and characterization of a CRISPR/Cas9-mediated hFCRNtransgenic mouse termed SYNB-hFCRN. Using CRISPR/Cas9-assisted gene targeting, we prepared a strain with a simultaneous knockout of mFcrnand insertion of a hFCRNmini-gene under the control of the endogenous mouse promoter. These mice are healthy and express hFCRN in the appropriate tissues and immune cell subtypes. Pharmacokinetic evaluation of human IgG and adalimumab (Humira®) demonstrate hFCRN-mediated protection. These newly generated SYNB-hFCRNmice provide another valuable animal model for use in preclinical pharmacokinetics studies during early drug development.Materials availabilityCryopreserved germline cells from the strains are available from the MMRRC. Dr. Jarnagin will provide sequencing and other evidence supporting strain identity and the plasmids used for constructions.
Dissecting the genetic architecture of sunflower head diameter using genome-wide association study
Authors: Delen Yavuz , Mural Ravi V. , Xu Gen , Delen Semra Palali , Schnable James C. , Yang Jinliang , Dweikat Ismail 
Published: October 2022

ABSTRACTSunflower (Helianthus annuusL.) plays an essential role in meeting the edible oil demand worldwide. Sunflower seed yield can be decomposed into several yield component traits, one of which is the head diameter. In 2019, 2020, and 2022, we evaluated the head diameter phenotypic variation on a set of diverse sunflower accessions (N=342) in replicated field trials. By combining three years of field data, the broad sense heritability (H2) of the head diameter trait was estimated to be 0.88. Then, a subset of N=274 accessions was genotyped by using the tunable genotyping-by-sequencing (tGBS) method, resulting in 226,779 high-quality SNPs. With these SNPs and the head diameter phenotype, the genome-wide association study (GWAS) was conducted using two statistical approaches: 1) the mixed linear model (MLM) and 2) the fixed and random model circulating probability unification (farmCPU). The MLM and farmCPU GWAS approaches identified 106 and 8 significant SNPs that were placed close to 53 and 21 genes, respectively. Two significant peaks were identified in MLM, with a strong signal on chromosome 10 and a less strong signal on chromosome 16. The farmCPU method detected the same signals on chromosomes 10 and 16 and several additional significant signals on other chromosomes. The head diameter associated genetic loci and the underlying candidate genes can be leveraged for further functional validation and serve as a basis for sunflower oil yield improvement.
Spatiotemporal gene expression atlas of the extremophyteSchrenkiella parvula
Authors: Wijesinghege Chathura , Wang Guannan , Pantha Pramod , Tran Kieu-Nga , Dassanayake Maheshi 
Published: October 2022

SUMMARYExtremophytes are naturally selected to survive environmental stresses, but scarcity of genetic resources for them developed with spatiotemporal resolution limit their use in stress biology.Schrenkiella parvulais one of the leading extremophyte models with initial molecular genomic resources developed to study its tolerance mechanisms to high salinity. Here we present a transcriptome atlas forS. parvulawith subsequent analyses to highlight its diverse gene expression networks associated with salt responses. We included spatiotemporal expression profiles, expression specificity of each gene, and co-expression and functional gene networks representing 115 transcriptomes sequenced from 35 tissue and developmental stages examining their responses before and after 27 salt treatments in our current study. The highest number of tissue-preferentially expressed genes were found in seeds and siliques while genes in seedlings showed the broadest expression profiles among developmental stages. Seedlings had the highest magnitude of overall transcriptomic responses to salinity compared to mature tissues and developmental stages. Differentially expressed genes in response to salt were largely mutually exclusive but shared common stress response pathways spanning across tissues and developmental stages. Our foundational dataset created forS. parvularepresenting a stress-adapted wild plant lays the groundwork for future functional, comparative, and evolutionary studies using extremophytes aiming to uncover novel stress tolerant mechanisms.SIGNIFICANCE STATEMENTConcerted transcriptomic responses coordinated across developmental stages and tissues are required to complete a plant lifecycle under salt stress. Transcriptomic resources created with spatiotemporal resolution for plants are rare and for stress-adapted plants rarer. We present a transcriptome atlas enabling discovery of genes and networks evolved as adaptations to salt stress in a model extremophyte,Schrenkiella parvula. The spatiotemporally resolved gene expression networks are largely non-overlapping but functionally connected through synergistic stress responsive pathways.
Hsp47 Promotes Biogenesis of Multi-subunit Neuroreceptors in the Endoplasmic Reticulum
Authors: Wang Ya-Juan , Di Xiao-Jing , Han Dong-Yun , Nashmi Raad , Henderson Brandon J. , Moss Fraser J. , Mu Ting-Wei 
Published: October 2022

ABSTRACTProtein homeostasis (proteostasis) deficiency is an important contributing factor to neurodegenerative, neurological, and metabolic diseases. However, how the proteostasis network orchestrates the folding and assembly of multi-subunit membrane proteins is not well understood. Previous proteomics studies identified Hsp47 (Gene:SERPINH1), a heat shock protein in the endoplasmic reticulum lumen, as the most enriched interacting chaperone for gamma-aminobutyric type A (GABAA) receptors. Here, we show that Hsp47 enhances neuronal GABAAreceptor functional surface expression, acting after Binding immunoglobulin Protein (BiP) to preferentially bind the folded conformation of GABAAreceptors. Therefore, Hsp47 promotes the subunit-subunit interaction, the receptor assembly process, and the anterograde trafficking of GABAAreceptors. These Hsp47 properties are also extended to other Cys-loop receptors, including nicotinic acetylcholine receptors. Therefore, in addition to its known function as a collagen chaperone, this work establishes that Hsp47 also plays a critical and general role in the maturation of multi-subunit neuroreceptors.HighlightsHsp47 positively regulates the functional surface expression of endogenous GABAAreceptors.Hsp47 acts after BiP and preferentially binds the folded conformation of GABAAreceptors.Hsp47 promotes the subunit-subunit assembly of GABAAreceptors.Hsp47 plays a critical and general role in the maturation of multi-subunit neuroreceptors.
Identification of aPrunusMAX1 Homolog as a Unique Strigol Synthase from Carlactone Bypassing 5-Deoxystrigol
Authors: Wu Sheng , Zhou Anqi , Hiugano Kozue , Yoda Akiyoshi , Xie Xiaonan , Yamane Kenji , Miura Kenji , Nomura Takahito , Li Yanran 
Published: October 2022

AbstractStrigol was the first strigolactone (SL) to be discovered, but the biosynthetic pathway remains elusive. Here, through rapid gene screening using a microbial SL-producing platform, we functionally identified a strigol synthase (PpMAX1c, a cytochrome P450 711A enzyme) inPrunusthat synthesizes strigol directly from the SL precursor carlactone through catalyzing multi-step oxidations and C-ring cyclization, bypassing the synthesis of 5-deoxystrigol. The function of PpMAX1c was validated through reconstructing the biosynthesis of strigol inNicotiana benthamiana. Additional genomic analysis and functional verification confirm that peach also encodes an orobanchol synthase (PpCYP722C, a cytochrome P450 722C enzyme), which hints at the presence of both strigol-type and orobanchol-type SLs in peach and was confirmed through metabolic analysis of peach seedlings. This work highlights the catalytic diversity of the largely unexplored family of CYP711A homologs and sets the foundation to characterize the roles of different types of SLs in the economically importantPrunus.
Mitoxantrone Targets Both Host and Bacteria to Overcome Vancomycin Resistance inEnterococcus faecalis
Authors: da Silva Ronni A. G. , Wong Jun Jie , Antypas Haris , Choo Pei Yi , Goh Karlyn , Jolly Shreya , Liang Cui , Sing Leona Tay Kwan , Veleba Mark , Hu Guangan , Chen Jianzhu , Kline Kimberly A. 
Published: October 2022

AbstractAmong Enterococci, intrinsic and acquired resistance to antibiotics such as β-lactams and vancomycin critically limit treatment options for infection with these opportunistic pathogens. Antimicrobials that enhance the host immune response are emerging as alternative approaches, with the potential to overcome bacterial resistance. Here, we investigate the antibiotic and immunological activity of the anticancer agent mitoxantrone (MTX)in vitroandin vivoagainst vancomycin resistantEnterococcus faecalis(VRE). We show that,in vitro, MTX is a potent antibiotic against Gram-positive bacteria with a minimal inhibitory concentration (MIC) of ~1 μg/ml through induction of reactive oxygen species and DNA damage. MTX synergises with vancomycin and lowers the vancomycin concentration required to kill VRE by over 140-fold. This synergy is specific to vancomycin-resistant, but not susceptible strains because vancomycin rendered the resistant strains more permeable to MTX and thus MTX-mediated DNA damage. In a murine wound infection model, MTX treatment effectively reduced VRE bacterial numbers by 120-fold and with further reduction when combined with vancomycin. Wounds treated with MTX had significantly higher numbers of macrophages and higher pro-inflammatory cytokines compared to untreated wounds. In addition, MTX augmented intracellular bacterial killing by both murine and human macrophages by upregulating the expression of lysosomal hydrolases cathepsins D and H, and β-Hexosaminidase. These results show that MTX is a potent antibiotic against Gram-positive bacteria, synergizes with vancomycin, enhances macrophage recruitment and intracellular bactericidal activity, and represents a promising dual bacterium- and host-targeted therapeutic for overcoming vancomycin resistance.One sentence summaryMitoxantrone synergizes with vancomycin against vancomycin resistant bacterial strains via direct antibiotic activity and by augmenting both host macrophage recruitment to the site of infection and macrophage bactericidal activity.
Blended care in patients with knee and/or hip osteoarthritis in physical therapy: a Delphi study on needs and preconditions
Authors: Weber F. , Kloek C. , Arntz A. , Grüneberg C. , Veenhof C. 
Published: October 2022

AbstractIntroductionOsteoarthritis is a major public health concern. Despite existing evidence-based treatment options, the health care situation remains unsatisfactory. Digital care options, especially combined with in-person sessions seems to be promising. Therefore, the aim of this study was to investigate the needs, preconditions, barriers and facilitators of blended physical therapy.MethodsThis Delphi study consisted of interviews, an online survey and focus groups. Participants were physical therapists, patients with hip or knee osteoarthritis with or without experience in digital care and stakeholders of the health care system. In the first phase, interviews were conducted with patients and physical therapists. The interview guide was based on the “Consolidated Framework For Implementation Research”. The interviews focused on experiences with digital and blended care. Furthermore, needs, facilitators and barriers were discussed. In the second phase, an online survey and focus groups served the process to confirm the needs and collect preconditions.ResultsNine physical therapists, seven patients and six stakeholders confirm that an increase of acceptance of digital care by physical therapists and patients is crucial. One of the most frequently mentioned facilitator was conducting regular in-person sessions. Physical therapists and patients conclude that blended physical therapy needs to be tailored to the patients’ needs. The reimbursement of blended physical therapy needs to be clarified.DiscussionMost importantly, it is necessary to strengthen the acceptance of patients and physical therapists towards digital care. Overall, for development and implementation purposes, it is crucial to take the needs and preconditions into account.