Biology subjects

Hou, Y.

Publications and source records attributed to Hou, Y..

At least 145 records · Page 8Linked to original sources

bioRxiv2022

Single-cell transcriptional profiling reveals cellular and molecular divergence in human maternal-fetal interface

Placenta play essential role in successful pregnancy, as the most important organ connecting and interplaying between mother and fetus. However, the cellular and molecular characteristics of fetal origin and maternal origin cell populations within the fetomaternal interface still is poorly understood. Here, we profiled the transcriptomes of single cells with well-defined maternal-fetal origin that consecutively localized from fetal section (FS), middle section (Mid_S) to maternal section (Mat_S) within the human full-term placenta. Then, we initially identified the cellular and molecular heterogeneity of cytotrophoblast cell (CTB) and stromal cell (STR) with the spatial location and fetal/maternal origin, also highlighted STR cells from fetal origins showed greater proliferation ability in Mat_S compared to cells from FS or Mid_S. Further, by integrating analysis with the first-trimester placental single cell transcriptome data, we revealed that a subpopulation of trophoblast progenitor-like cells (TPLCs) existed in the full-term placenta and mainly distributed in Mid_S, with high expression of pool of putative cell surface makers and unique molecular features. Moreover, through the extravillous cytotrophoblast (EVT) subsets differentiation trajectory and regulation network analysis, we proposed a putative key transcription factor PRDM6 that promoted the differentiation of endovascular extravillous trophoblast cells (enEVT). Finally, based on the integrated analyses of single cell transcriptional profiling of preeclampsia (PE) and match-trimester normal placenta, we highlighted the defective EVT subgroup composition and down-regulation of PRDM6 may lead to an abnormal enEVT differentiation process in PE. Together, our study offers important resources for better understanding of human placenta, stem cell-based therapy as well as PE, and provides new insights on the study of tissue heterogeneity, the clinical prevention and control of PE as well as the maternal-fetal interface.

developmental biology↗

bioRxiv2021

Long-term labeling and imaging of synaptically-connected neuronal networks in vivo using nontoxic, double-deletion-mutant rabies viruses

Monosynaptic tracing is a widely-used technique for mapping neural circuitry, but its cytotoxicity has confined it primarily to anatomical applications. Here we present a second-generation system for labeling direct inputs to targeted neuronal populations with minimal toxicity, using double-deletion-mutant rabies viruses. Spread of the viruses requires expression of both deleted viral genes in trans in postsynaptic source cells; suppressing this expression with doxycycline following an initial period of viral replication reduces toxicity to postsynaptic cells. Longitudinal two-photon imaging in vivo indicated that over 90% of both presynaptic and source cells survived for the full twelve-week course of imaging. Ex vivo whole-cell recordings at 5 weeks postinfection showed that the second-generation system perturbs input and source cells much less than does the first-generation system. Finally, two-photon calcium imaging of labeled networks of visual cortex neurons showed that their visual response properties appeared normal for 10 weeks, the longest we followed them.

neuroscience↗

bioRxiv2021

Screening and selection of 21 novel microhaplotype markers for ancestry inference in ten Chinese Subpopulations

Genetic findings suggested that ethnolinguistically diverse populations in China harbor differentiated genetic structure and complex evolutionary admixture history, which provide the genetic basis and theoretical foundation for forensic biogeographical ancestry inference (BGAI). Forensic assays for BGAI among intracontinental eastern Eurasians were previously conducted mainly based on the SNPs or InDels. Microhaplotypes, as a set of closely linked SNPs within 200 base pairs, possess the advantages of both STR and SNP and have great potential in forensic ancestry inference. However, the developed forensic assay based on the ancestry informative microhaplotypes in the BGAI remained to be comprehensively explored, especially in China with enriching genetic diversity. Here, we described a new BGAI panel based on 21 novel identified ancestry informative microhaplotypes that focused on dissected finer-scale ancestry composition of Chinese populations. We initially screened all possible microhaplotypes with high Fst values among five East Asian populations and finally employed 21 candidate microhaplotypes in two multiplex SNaPshot assays. Forensic amplification efficiency and statistically/physically phased haplotypes of the 21 microhaplotypes were validated using both SNaPshot and massively parallel sequencing (MPS) platforms. Followingly, we validated the efficiency of these microhaplotypes for BGAI in 764 individuals from ten Chinese populations. Fine-scale ancestry source and ancestry proportion estimated by the principal component analysis (PCA), multidimensional scaling (MDS), phylogenetic tree and model-based STRUCTURE among worldwide populations and East Asians showed that our customized panel could provide a higher discrimination resolution in both continental population stratification and East Asian regional substructure. East Asian populations could be further classified into linguistically/geographically different intracontinental subpopulations (Tibeto-Burman, Tai-Kadai and others). Finally, we obtained a higher estimated accuracy using training and tested datasets in the microhaplotype-based panel than traditional SNP-based panels. Generally, the above results demonstrated that this microhaplotype panel was robust and suitable for forensic BGAI in Chinese populations, which not only provided a high discriminatory power for continental populations but also discriminated East Asians into linguistically restricted subpopulations.

genetics↗

bioRxiv2021

A Network-based Deep Learning Framework Catalyzes GWAS and Multi-Omics Findings to Biology and Drug Repurposing for Alzheimer's Disease

Human genome sequencing studies have identified numerous loci associated with complex diseases, including Alzheimers disease (AD). Translating human genetic findings (i.e., genome-wide association studies [GWAS]) to pathobiology and therapeutic discovery, however, remains a major challenge. To address this critical problem, we present a network topology-based deep learning framework to identify disease-associated genes (NETTAG). NETTAG is capable of integrating multigenomics data along with the protein-protein interactome to infer putative risk genes and drug targets impacted by GWAS loci. Specifically, we leverage non-coding GWAS loci effects on expression quantitative trait loci (eQTLs), histone-QTLs, and transcription factor binding-QTLs, enhancers and CpG islands, promoter regions, open chromatin, and promoter flanking regions. The key premises of NETTAG are that the disease risk genes exhibit distinct functional characteristics compared to non-risk genes and therefore can be distinguished by their aggregated genomic features under the human protein interactome. Applying NETTAG to the latest AD GWAS data, we identified 156 putative AD-risk genes (i.e., APOE, BIN1, GSK3B, MARK4, and PICALM). We showed that predicted risk genes are: 1) significantly enriched in AD-related pathobiological pathways, 2) more likely to be differentially expressed regarding transcriptome and proteome of AD brains, and 3) enriched in druggable targets with approved medicines (i.e., choline and ibudilast). In summary, our findings suggest that understanding of human pathobiology and therapeutic development could benefit from a network-based deep learning methodology that utilizes GWAS findings under the multimodal genomic analyses.

bioinformatics↗

bioRxiv2021

Profiling of Nascent Lariat Intermediates Reveals Key Genetic Determinants of the Timing of Human Co-transcriptional Splicing

As splicing is intimately coupled with transcription, understanding splicing mechanisms requires an understanding of splicing timing, which is currently limited. Here, we developed CoLa-seq (co-transcriptional lariat sequencing), a genomic assay that reports splicing timing relative to transcription through analysis of nascent lariat intermediates. In human cells, we mapped 165,282 branch points and characterized splicing timing for over 70,000 introns. Splicing timing varies dramatically across introns, with regulated introns splicing later than constitutive introns. Machine learning-based modeling revealed genetic elements predictive of splicing timing, notably the polypyrimidine tract, intron length, and regional GC content, which illustrate the significance of the broader genomic context of an intron and the impact of co-transcriptional splicing. The importance of the splicing factor U2AF in early splicing rationalizes surprising observations that most introns can splice independent of exon definition. Together, these findings establish a critical framework for investigating the mechanisms and regulation of co-transcriptional splicing. HighlightsO_LICoLa-seq enables cell-type specific, genome-wide branch point annotation with unprecedented efficiency. C_LIO_LICoLa-seq captures co-transcriptional splicing for tens of thousands of introns and reveals splicing timing varies dramatically across introns. C_LIO_LIModeling uncovers key genetic determinants of splicing timing, most notably regional GC content, intron length, and the polypyrimidine tract, the binding site for U2AF2. C_LIO_LIEarly splicing precedes transcription of a downstream 5 SS and in some cases accessibility of the upstream 3 SS, precluding exon definition. C_LI

molecular biology↗

bioRxiv2021

Screening and Identification of Lujo Virus Entry Inhibitors from an FDA-Approved Drugs Library

The Lujo virus (LUJV) belongs to the Old World (OW) genus Mammarenavirus (family Arenaviridae); it is categorized as a biosafety level (BSL) 4 agent. Currently, there are no U.S. Food and Drug Administration (FDA)-approved drugs or vaccines specifically for LUJV or other pathogenic OW mammarenaviruses. Here, a high-throughput screening of an FDA-approved drug library was conducted using pseudotype viruses bearing LUJV envelope glycoprotein (GPC) to identify inhibitors of LUJV entry. Three hit compounds, trametinib, manidipine, and lercanidipine, were identified as LUJV entry inhibitors in the micromolar range. Mechanistic studies revealed that trametinib inhibited LUJV GPC-mediated membrane fusion by targeting C410 (located in the transmembrane (TM) domain), while manidipine and lercanidipine inhibited LUJV entry by acting as calcium channel blockers. Meanwhile, all three hits extended their antiviral spectra to the entry of other pathogenic mammarenaviruses. Furthermore, all three could inhibit the authentic prototype mammarenavirus, lymphocytic choriomeningitis virus (LCMV), and could prevent infection at the micromolar level. This study shows that trametinib, manidipine, and lercanidipine are candidates for LUJV therapy, and highlights the critical role of calcium in LUJV infection. The presented findings reinforce the notion that the key residue(s) located in the TM domain of GPC provide an entry-targeted platform for designing mammarenavirus inhibitors. IMPORTANCETo date, only one LUJV outbreak has been recorded; it occurred in 2008 and resulted in a fatality rate of 80% (4/5 cases). Pathogenesis studies and therapeutic strategies are therefore urgently needed. Repurposing approved drugs can accelerate the development of drug design and facilitate the understanding of infectious mechanisms. Here, three compounds, trametinib, manidipine, and lercanidipine, were identified as entry inhibitors against LUJV. Studying the underling mechanisms revealed that a key residue (C410) in LUJV GPC modulates its sensitivity/resistance to trametinib and demonstrated the critical role of calcium in LUJV infection.

microbiology↗

bioRxiv2021

Comparative proteomic analysis of vancomycin-sensitive and vancomycin-intermediate resistant S. aureus

The extensive use of vancomycin has led to the development of MRSA (methicillin-resistant Staphylococcus aureus) strains with varying degrees of resistance to vancomycin. Accumulation of surplus cell wall material, decreased cross-linking of peptidoglycan, and/ or other cell wall alterations has been put forward to explain the VISA phenotype. To our knowledge, the protein profiles of hVISA and VISA strains are rarely analyzed via quantitative comparative proteomics. In this study, we subjected subcellular fractions isolated from two isogenic S. aureus strains (vancomycin-intermediate resistant S. aureus) to proteomic analysis, using an integrated quantitative proteomic approach assisted by bioinformatic analysis. In total,128 up-regulated proteins (mainly AhpC, Adh, ArgF et al.) were identified,21 down-regulated proteins (mainly LtaS, SdrD, MsrR, MsrB, OatA et al.) were obtained. The largest group of differentially expressed proteins is composed of enzymic proteins associated with metabolic and catalytic activity, which accounts for 50% and 51% of the total proteins, respectively.Some proteins which take an indispensable part in the regulatory networks of S. aureus with vancomycin treatment are related to Cell wall metabolism ( MurA ), Cell adhesion ( SdrC, SdrD, ClfA, ClfB ), Proteolysis ( Atl, LytM, SceD ) and Pressure response ( MsrA, MsrB, AhpC ) process.In conclusion, our proteomic study revealed regulatory proteins associated with vancomycin resistance in S. aureus, and some of these proteins are involved in the regulation of cell metabolism and function, providing protential targets for further development of drug resistance strategies.

microbiology↗

bioRxiv2021

Identification of TAZ as the essential molecular switch in orchestrating SCLC phenotypic transition and metastasis

Small cell lung cancer (SCLC) is a recalcitrant cancer featured with high metastasis. However, the exact cell type contributing to metastasis remains elusive. Using Rb1L/L/Trp53L/L mouse model, we identify the NCAMhiCD44lo/- subpopulation as SCLC metastasizing cell (SMC), which is progressively transitioned from non-metastasizing NCAMloCD44hi cell (Non-SMC). Integrative chromatin accessibility and gene expression profiling studies reveal an important role of SWI/SNF complex, and knockout of its central component, Brg1, significantly inhibits such phenotypic transition and metastasis. Mechanistically, TAZ is silenced by SWI/SNF complex during SCLC malignant progression, and its knockdown promotes SMC transition and metastasis. Importantly, ectopic TAZ expression reversely drives SMC-to-Non-SMC transition and alleviates metastasis. Single-cell RNA-sequencing analyses identify SMC as the dominant subpopulation in human SCLC metastasis, and immunostaining data show a positive correlation between TAZ and patient prognosis. These data uncover high SCLC plasticity and identify TAZ as key molecular switch in orchestrating SCLC phenotypic transition and metastasis.

cancer biology↗

bioRxiv2021

Orfamide-A-mediated bacterial-algal interactions involve specific Ca2+ signalling pathways

The antagonistic bacterium Pseudomonas protegens secretes the cyclic lipopeptide orfamide A, which triggers a Ca2+ signal, causing the deflagellation of the green microalga Chlamydomonas reinhardtii. By investigating targeted synthetic orfamide A variants and inhibitors, we found that at least two Ca2+-signalling pathways and TRP channels are involved in this response.

microbiology↗

bioRxiv2021

Genomic insights into the differentiated population admixture structure and demographic history of North East Asians

North China and South Siberia, mainly populated by Altaic-speaking populations, possess extensive ethnolinguistic diversity and serve as the crossroad for the initial peopling of America and western-eastern trans-continental communication. Yet, the complex scenarios of genetic origin, population structure, and admixture history of North-East Asia remain to be fully characterized, especially for Mongolic people in China with a genome-wide perspective. Thus, we genotyped genome-wide SNPs for 510 individuals from 38 Chinese Mongolic, Tungusic, and Sinitic populations to explore the sharing alleles and haplotypes within the studied groups and following merged it with 3508 modern and ancient Eurasian individuals to reconstruct the deep evolutionary and natural selection history of northern East Asians. We identified significant substructures within Altaic-speaking populations with the primary common ancestry linked to the Neolithic northern East Asians: Western Turkic people harbored more western Eurasian ancestry; Northern Mongolic people in Siberia and eastern Tungusic people in Amur River Basin (ARB) possessed dominant Neolithic Mongolian Plateau (MP) or ARB ancestry; Southern Mongolic people in China owned obvious genetic impact from Neolithic Yellow River Basin (YRB) farmers. Additionally, we found the differentiated admixture history between western and eastern Mongolians and geographically close Northeast Hans: the former received a genetic impact from western Eurasians and the latter retained the dominant YRB and ARB Neolithic ancestry. Moreover, we demonstrated that Kalmyk people from the northern Caucasus Mountain possessed a strong genetic affinity with Neolithic MP people, supporting the hypothesis of their eastern Eurasian origin and long-distance migration history. We also illuminated that historic pastoral empires in the MP contributed considerably to the gene pool of northern Mongolic people but rarely to southern ones. We finally found natural signatures in Mongolians associated with alcohol metabolism. Generally, our results not only illuminated that complex population migration and admixture of Neolithic ancestral sources from the MP or ARB played an important role in the spread of Altaic-speaking populations and Proto-Altaic language, which partly supported the Northeast Asia-origin hypothesis, but also demonstrated that the observed multi-sources of genetic diversity contributed significantly to the modern existing extensive ethnolinguistic diversity in North-East Asia.

genomics↗

bioRxiv2021

Novel Roles of G-quadruplexes on Enhancers in human chromatin

G-quadruplexes (G4), stable four-stranded non-canonical DNA structures, are highly related to function of promoters and initiation of gene transcription. We found that G4 structures were also enriched in the enhancers across different cell lines. However, the relationship between G4 structures and enhancer activity remains unknown. Here, we proved that G4 structures on enhancers lead to the re-positioning of nucleosomes create nucleosome depleted regions (NDRs). Moreover, stable NDRs and special secondary structures of G4 help enhancers to recruit abundant TFs to co-bind, especially for architectural proteins including CTCF, RAD21, and SMC3. These architectural proteins, which play critical roles in the formation of higher-order chromatin organization, further influenced the chromatin interactions of G4 enhancers. Additionally, we revealed that G4 enhancers harbored significantly higher enrichment of eQTLs than typical enhancers, suggesting G4 enhancers displayed more enhancer regulatory activity. We found that most super enhancers (SEs) contain G4 structures. Even though the enrichment of chromatin accessibility and histone modifications around G4-containing SEs are not significantly higher than those around other SEs, G4-containing SEs still possess much more TFs across different cell lines. According to these results, we proposed a model in which the formation of G4 structures on enhancer exclude nucleosome occupancy and recruit abundant TFs which lead to the stable chromatin interaction between G4 enhancers and their target genes. Because of the relevance between G4 structures and enhancers, we hypothesized that G4 structures may be a potential markers indicating enhancer regulatory activity.

bioinformatics↗

bioRxiv2021

Tracking neural markers of template formation and implementation in attentional inhibition under different distractor consistency

Performing visual search tasks requires optimal attention deployment to promote targets and inhibit distractors. Rejection templates based on the distractors feature can be built to constrain the search process. We measured electroencephalography (EEG) of human participants of both sexes when they performed a visual search task in conditions where the distractor cues were constant within a block (fixed-cueing) or changed on a trial-by-trial basis (varied-cueing). In the fixed-cueing condition, sustained decoding of the cued colors could be achieved during the retention interval and the participants with higher decoding accuracy showed larger suppression benefit of the distractor cueing in the search period. In the varied-cueing condition, the cued color could only be transiently decoded after its onset and the higher decoding accuracy was observed from the participants who demonstrated lower suppression benefit. The differential neural representations of the to-be-ignored color in the two cueing conditions as well as their reverse associations with behavioral performance implied that rejection templates were formed in the fixed-cueing condition but not in the varied-cueing condition. Additionally, we observed stronger posterior alpha lateralization and mid-frontal theta/beta power during the retention interval of the varied-cueing condition, indicating the cognitive costs in template formation caused by the trialwise change of distractor colors. Taken together, our findings revealed the neural markers associated with the critical roles of distractor consistency in linking template formation to successful inhibition.

neuroscience↗

bioRxiv2021

Metagenome-genome-wide association studies reveal human genetic impact on the oral microbiome

The oral microbiota contains billions of microbial cells, which could contribute to diseases in a number of body sites. Challenged by eating, drinking and dental hygiene on a daily basis, the oral microbiota is regarded as highly dynamic. Here, we report significant human genomic associations with the oral metagenome from more than 1,915 individuals, for both the tongue dorsum and saliva. We identified five genetic loci associated with oral microbiota at study-wide significance (p < 3.16 x 10-11). Four of the five associations were well replicated in an independent cohort of 1,439 individuals: rs1196764 at APPL2 with Prevotella jejuni, Oribacterium uSGB 3339 and Solobacterium uSGB 315; rs3775944 at the serum uric acid transporter SLC2A9 with Oribacterium uSGB 1215, Oribacterium uSGB 489 and Lachnoanaerobaculum umeaense; rs4911713 near OR11H1 with species F0422 uSGB 392; and rs36186689 at LOC105371703 with Eggerthia. Further analyses confirmed 84% (386/455 for tongue dorsum) and 85% (391/466 for saliva) of genetic-microbiota associations including 6 genome-wide significant associations mutually validated between the two niches. Human genetics accounted for at least 10% of oral microbiome compositions between individuals. Machine learning models showed that polygenetic risk score dominated over oral microbiome in predicting predisposing risk of dental diseases such as dental calculus and gingival bleeding. These findings indicate that human genetic differences are one explanation for a stable or recurrent oral microbiome in each individual.

genetics↗

bioRxiv2021

Network medicine links SARS-CoV-2/COVID-19 infection to brain microvascular injury and neuroinflammation in dementia-like cognitive impairment

BackgroundDementia-like cognitive impairment is an increasingly reported complication of SARS-CoV-2 infection. However, the underlying mechanisms responsible for this complication remain unclear. A better understanding of causative processes by which COVID-19 may lead to cognitive impairment is essential for developing preventive interventions. MethodsIn this study, we conducted a network-based, multimodal genomics comparison of COVID-19 and neurologic complications. We constructed the SARS-CoV-2 virus-host interactome from protein-protein interaction assay and CRISPR-Cas9 based genetic assay results, and compared network-based relationships therein with those of known neurological manifestations using network proximity measures. We also investigated the transcriptomic profiles (including single-cell/nuclei RNA-sequencing) of Alzheimers disease (AD) marker genes from patients infected with COVID-19, as well as the prevalence of SARS-CoV-2 entry factors in the brains of AD patients not infected with SARS-CoV-2. ResultsWe found significant network-based relationships between COVID-19 and neuroinflammation and brain microvascular injury pathways and processes which are implicated in AD. We also detected aberrant expression of AD biomarkers in the cerebrospinal fluid and blood of patients with COVID-19. While transcriptomic analyses showed relatively low expression of SARS-CoV-2 entry factors in human brain, neuroinflammatory changes were pronounced. In addition, single-nucleus transcriptomic analyses showed that expression of SARS-CoV-2 host factors (BSG and FURIN) and antiviral defense genes (LY6E, IFITM2, IFITM3, and IFNAR1) was significantly elevated in brain endothelial cells of AD patients and healthy controls relative to neurons and other cell types, suggesting a possible role for brain microvascular injury in COVID-19-mediated cognitive impairment. Notably, individuals with the AD risk allele APOE E4/E4 displayed reduced levels of antiviral defense genes compared to APOE E3/E3 individuals. ConclusionOur results suggest significant mechanistic overlap between AD and COVID-19, strongly centered on neuroinflammation and microvascular injury. These results help improve our understanding of COVID-19-associated neurological manifestations and provide guidance for future development of preventive or treatment interventions.

systems biology↗

bioRxiv2021

Instructor-learner neural synchronization during elaborated feedback predicts learning transfer

The provision of feedback with complex information beyond the correct answer, i.e., elaborated feedback, can powerfully shape learning outcomes such as transfer, i.e., the ability to extend what has been learned in one context to new contexts. However, an understanding of neurocognitive processes of elaborated feedback during instructor-learner interactions remains elusive. Here, a two-person interactive design is used during simultaneous recording of functional near-infrared spectroscopy (fNIRS) signals from adult instructor-learner dyads. Instructors either provided elaborated feedback (i.e., correct answer and an example) or simple feedback (i.e., correct answer only) to learners during a concept learning task. Our results showed that elaborated feedback produced comparable levels of retention to simple feedback, however, transfer was significantly enhanced by elaboration. We also noted significant instructor-learner neural synchronization in frontoparietal regions during the provision of elaborated feedback, especially when examples were provided. Further, interpersonal neural synchronization in the parietal cortex successfully predicted transfer of knowledge to novel contexts. This prediction was retained for both learner-delayed and learner-preceding neural synchronization. These findings point toward transfer effects of elaborated feedback provided in a social context can be predictable through interpersonal neural synchronization, which may hold important implications for real-world learning and pedagogical efficacy. Educational Impact and Implications StatementFeedback provides learners with crucial information regarding the gap between what has currently been achieved and what remains to be achieved, and thus plays a critical role in any learning process. In real-world settings, feedback is typically provided and received through social interaction, and high-quality "elaborated feedback" contains complex information that goes beyond the correct answer. This study aims to elucidate the neurocognitive processes underpinning elaborated feedback during instructor-learner interactions. We detected significant instructor-learner neural synchronization in mutual frontoparietal brain regions during elaborated feedback, particularly during the provision of specific elaborated information (i.e., concrete examples). Moreover, this synchronization (including learner-delayed and learner-preceded synchronization) in the parietal region predicted whether the learners transferred learning to novel examples of learned psychology concepts. This study advances current understanding on the neural mechanisms for elaborated feedback and the role of social interaction in feedback effects. These results may have important implications for successful real-world learning and communication, and related pedagogical applications in educational settings.

scientific communication and education↗

bioRxiv2021

Single-cell sequencing characterizes intratumoral profile of immune cells in old mice

It has long been thought that aging is a major risk factor for cancer incidence. However, accumulating evidence indicates increased resistance of old animals to tumor growth. A systematic understanding of how old individuals defend against tumor invasion is currently lacking. Here we investigated the differences of age-associated alterations in tumor-infiltrating immune cells between young and old mice using single-cell RNA analysis. Our results showed that a higher proportion of cytotoxic CD8+ T cells, nTregs, cDC, and M1-type macrophages, while a higher percentage of exhausted CD8+ T cells, iTregs, pDC, and M2-type macrophages were found in young mice. Importantly, TCR diversity analysis showed top 10 TCR clones consisted primarily of exhausted CD8+ T cells in young mice whereas tip clones were predominantly cytotoxic CD8+ T cells in old mice. Consistently, trajectory inference demonstrated that CD8+ T cells preferentially differentiated into cytotoxic cells in old mice in contrast to exhausted cells in young mice. Meanwhile, we confirmed the main distinctions between young and old mice by flow cytometry. Collectively, our data revealed that a significantly higher proportion of effector immune cells in old mice defend against tumor progression, providing a framework for the immunotherapy of elderly patients with tumors.

immunology↗

bioRxiv2021

Novel EGFRvIII-CAR transgenic mice for rigorous preclinical studies in syngeneic mice

BackgroundRigorous preclinical studies of chimeric antigen receptor (CAR) immunotherapy will require large quantities of consistent and high-quality CAR-transduced T (CART)-cells that can be used in syngeneic mouse glioblastoma (GBM) models. To this end, we developed a novel transgenic (Tg) mouse strain with a fully murinized CAR targeting epidermal growth factor receptor variant III (EGFRvIII). MethodsWe first established the murinized version of EGFRvIII-CAR and validated its function using a retroviral vector (RV) in C57BL/6J mice bearing syngeneic SB28 GBM expressing EGFRvIII. Next, we created C57BL/6J-background Tg mice carrying the anti-EGFRvIII-CAR downstream of a Lox-Stop-Lox cassette in the Rosa26 locus. We bred these mice with CD4-Cre Tg mice to allow CAR expression on T-cells and evaluated the function of the CART-cells both in vitro and in vivo. In order to inhibit immunosuppressive myeloid cells within SB28 GBM, we also evaluated a combination approach of CART and an anti-EP4 compound (ONO-AE3-208). ResultsBoth RV- and Tg-CART-cells demonstrated specific cytotoxic activities against SB28-EGFRvIII cells. A single intravenous infusion of EGFRvIII-CART-cells prolonged the survival of glioma-bearing mice when preceded by a lymphodepletion regimen with recurrent tumors displaying profound EGFRvIII loss. The addition of ONO-AE3-208 resulted in long-term survival in a fraction of CART-treated mice and those survivors demonstrated delayed growth of subcutaneously re-challenged both EGFRvIII+ and parental EGFRvIII- SB28. ConclusionOur new syngeneic CAR Tg mouse model can serve as a useful tool to address clinically relevant questions and develop future immunotherapeutic strategies. Importance of studyThe majority of preclinical studies evaluating CART therapy for GBM have utilized xenografts implanted into immunocompromised mice. Because the successful development of these strategies will depend on the understanding of critical interactions between therapeutic cells and the endogenous immune environment, it is essential to develop a novel immunocompetent system which allows us to study these interactions in a robust and reproducible manner. To this end, we created a Tg mouse strain in which all T-cells express a murinized EGFRvIII-CAR. T-cells derived from these mice demonstrated consistent CAR expression and EGFRvIII-specific cytotoxicity while traditional transduction with a CAR vector showed batch-to-batch variability. The syngeneic system also gave us the opportunity to evaluate a combination regimen with blockade of myeloid-derived suppressor cells. The Tg-CART mice represent a novel system for robust, and reproducible preclinical investigations.

immunology↗

bioRxiv2021

YHP: Y-chromosome Haplogroup Predictor for predicting male lineages based on Y-STRs

Human Y chromosome reflects the evolutionary process of males. Male lineage tracing by Y chromosome is of great use in evolutionary, forensic, and anthropological studies when male samples exist or especially when the biological sample is a mixture of male and female individuals. Identifying the male lineage based on the specific distribution of Y haplogroups narrows down the investigation scope. Integrating previously published datasets with genotypes of Y chromosome short tandem repeats (Y-STRs) and high-resolution haplogroups (122 haplogroups in total), we developed YHP (Y Haplogroup Predictor), an open-access and userfriendly software package to predict haplogroups, compare the similarity, and conduct mismatch analysis of samples with Y-STR profiles. The software is available at Github (https://github.com/cissy123/YHP-Y-Haplogroup-Predictor-). Author SummaryFamilial searching has been used in forensic, anthropologic, and personalized scenarios. Software packages have been developed to assist in male familial searching, such as predicting Y-SNP haplogroups by Y-STRs. However, these software packages, in general, achieve this goal with a rough resolution. In this study, we developed a software package to conduct high-resolution haplogroup inference to help familial searching and at the same time reduce the cost, since it does not require tiresome Y-SNP sequencing.

genetics↗