Organoid single-cell genomic atlas uncovers human-specific features of brain development.

The human brain has undergone substantial change since humans diverged from chimpanzees and the other great apes1,2. However, the genetic and developmental programs that underlie this divergence are not fully understood. Here we have analysed stem cell-derived cerebral organoids using single-cell transcriptomics and accessible chromatin profiling to investigate gene-regulatory changes that are specific to humans. We first analysed cell composition and reconstructed differentiation trajectories over the entire course of human cerebral organoid development from pluripotency, through neuroectoderm and neuroepithelial stages, followed by divergence into neuronal fates within the dorsal and ventral forebrain, midbrain and hindbrain regions. Brain-region composition varied in organoids from different iPSC lines, but regional gene-expression patterns remained largely reproducible across individuals. We analysed chimpanzee and macaque cerebral organoids and found that human neuronal development occurs at a slower pace relative to the other two primates. Using pseudotemporal alignment of differentiation paths, we found that human-specific gene expression resolved to distinct cell states along progenitor-to-neuron lineages in the cortex. Chromatin accessibility was dynamic during cortex development, and we identified divergence in accessibility between human and chimpanzee that correlated with human-specific gene expression and genetic change. Finally, we mapped human-specific expression in adult prefrontal cortex using single-nucleus RNA sequencing analysis and identified developmental differences that persist into adulthood, as well as cell-state-specific changes that occur exclusively in the adult brain. Our data provide a temporal cell atlas of great ape forebrain development, and illuminate dynamic gene-regulatory features that are unique to humans.

Integrative analysis of single-cell embryo data reveals transcriptome signatures for the human pre-implantation inner cell mass.

As the source of embryonic stem cells (ESCs), inner cell mass (ICM) can form all tissues of the embryo proper, however, its role in early human lineage specification remains controversial. Although a stepwise differentiation model has been proposed suggesting the existence of ICM as a distinct developmental stage, the underlying molecular mechanism remains unclear. In the present study, we perform an integrated analysis on the public human preimplantation embryonic single-cell transcriptomic data and apply a trajectory inference algorithm to measure the cell plasticity. In our results, ICM population can be clearly discriminated on the dimension-reduced graph and confirmed by compelling evidences, thus validating the two-step hypothesis of lineage commitment. According to the branch probabilities and differentiation potential, we determine the precise time points for two lineage segregations. Further analysis on gene expression dynamics and regulatory network indicates that transcription factors including GSC, PRDM1, and SPIC may underlie the decisions of ICM fate. In addition, new human ICM marker genes, such as EPHA4 and CCR8 are discovered and validated by immunofluorescence. Given the potential clinical applications of ESCs, our analysis provides a further understanding of human ICM cells and facilitates the exploration of more unique characteristics in early human development.

Hypomorphic and hypermorphic mouse models of Fsip2 indicate its dosage-dependent roles in sperm tail and acrosome formation.

Loss-of-function mutations in multiple morphological abnormalities of the sperm flagella (MMAF)-associated genes lead to decreased sperm motility and impaired male fertility. As an MMAF gene, the function of fibrous sheath-interacting protein 2 (FSIP2) remains largely unknown. In this work, we identified a homozygous truncating mutation of FSIP2 in an infertile patient. Accordingly, we constructed a knock-in (KI) mouse model with this mutation. In parallel, we established an Fsip2 overexpression (OE) mouse model. Remarkably, KI mice presented with the typical MMAF phenotype, whereas OE mice showed no gross anomaly except for sperm tails with increased length. Single-cell RNA sequencing of the testes uncovered altered expression of genes related to sperm flagellum, acrosomal vesicle and spermatid development. We confirmed the expression of Fsip2 at the acrosome and the physical interaction of this gene with Acrv1, an acrosomal marker. Proteomic analysis of the testes revealed changes in proteins sited at the fibrous sheath, mitochondrial sheath and acrosomal vesicle. We also pinpointed the crucial motifs of Fsip2 that are evolutionarily conserved in species with internal fertilization. Thus, this work reveals the dosage-dependent roles of Fsip2 in sperm tail and acrosome formation.

Technical strategy for monozygotic twin discrimination by single-nucleotide variants.

Monozygotic (MZ) twins are theoretically genetically identical. Although they are revealed to accumulate mutations after the zygote splits, discriminating between twin genomes remains a formidable challenge in the field of forensic genetics. Single-nucleotide variants (SNVs) are responsible for a substantial portion of genetic variation, thus potentially serving as promising biomarkers for the identification of MZ twins. In this study, we sequenced the whole genome of a pair of female MZ twins when they were 27 and 33 years old to approximately 30 × coverage using peripheral blood on an Illumina NovaSeq 6000 Sequencing System. Potentially discordant SNVs supported by whole-genome sequencing were validated extensively by amplicon-based targeted deep sequencing and Sanger sequencing. In total, we found nine bona fide post-twinning SNVs, all of which were identified in the younger genomes and found in the older genomes. None of the SNVs occurred within coding exons, three of which were observed in introns, supported by whole-exome sequencing results. A double-blind test was employed, and the reliability of MZ twin discrimination by discordant SNVs was endorsed. All SNVs were successfully detected when input DNA amounts decreased to 0.25 ng, and reliable detection was limited to seven SNVs below 0.075 ng input. This comprehensive analysis confirms that SNVs could serve as cost-effective biomarkers for MZ twin discrimination.

Epigenetic regulation of human-specific gene expression in the prefrontal cortex.

BACKGROUND: Changes in gene expression levels during brain development are thought to have played an important role in the evolution of human cognition. With the advent of high-throughput sequencing technologies, changes in brain developmental expression patterns, as well as human-specific brain gene expression, have been characterized. However, interpreting the origin of evolutionarily advanced cognition in human brains requires a deeper understanding of the regulation of gene expression, including the epigenomic context, along the primate genome. Here, we used chromatin immunoprecipitation sequencing (ChIP-seq) to measure the genome-wide profiles of histone H3 lysine 4 trimethylation (H3K4me3) and histone H3 lysine 27 acetylation (H3K27ac), both of which are associated with transcriptional activation in the prefrontal cortex of humans, chimpanzees, and rhesus macaques. RESULTS: We found a discrete functional association, in which H3K4me3HP gain was significantly associated with myelination assembly and signaling transmission, while H3K4me3HP loss played a vital role in synaptic activity. Moreover, H3K27acHP gain was enriched in interneuron and oligodendrocyte markers, and H3K27acHP loss was enriched in CA1 pyramidal neuron markers. Using strand-specific RNA sequencing (ssRNA-seq), we first demonstrated that approximately 7 and 2% of human-specific expressed genes were epigenetically marked by H3K4me3HP and H3K27acHP, respectively, providing robust support for causal involvement of histones in gene expression. We also revealed the co-activation role of epigenetic modification and transcription factors in human-specific transcriptome evolution. Mechanistically, histone-modifying enzymes at least partially contribute to an epigenetic disturbance among primates, especially for the H3K27ac epigenomic marker. In line with this, peaks enriched in the macaque lineage were found to be driven by upregulated acetyl enzymes. CONCLUSIONS: Our results comprehensively elucidated a causal species-specific gene-histone-enzyme landscape in the prefrontal cortex and highlighted the regulatory interaction that drove transcriptional activation.

A Multi-Information Spreading Model for One-Time Retweet Information in Complex Networks.

In the realm of online social networks, the spreading of information is influenced by a complex interplay of factors. To explore the dynamics of one-time retweet information spreading, we propose a Susceptible-Infected-Completed (SIC) multi-information spreading model. This model captures how multiple pieces of information interact in online social networks by introducing inhibiting and enhancement factors. The SIC model considers the completed state, where nodes cease to spread a particular piece of information after transmitting it. It also takes into account the impact of past and present information received from neighboring nodes, dynamically calculating the probability of nodes spreading each piece of information at any given moment. To analyze the dynamics of multiple information pieces in various scenarios, such as mutual enhancement, partial competition, complete competition, and coexistence of competition and enhancement, we conduct experiments on BA scale-free networks and the Twitter network. Our findings reveal that competing information decreases the likelihood of its spread while cooperating information amplifies the spreading of mutually beneficial content. Furthermore, the strength of the enhancement factor between different information pieces determines their spread when competition and cooperation coexist. These insights offer a fresh perspective for understanding the patterns of information propagation in multiple contexts.

Single-cell-resolution transcriptome map of human, chimpanzee, bonobo, and macaque brains.

Identification of gene expression traits unique to the human brain sheds light on the molecular mechanisms underlying human evolution. Here, we searched for uniquely human gene expression traits by analyzing 422 brain samples from humans, chimpanzees, bonobos, and macaques representing 33 anatomical regions, as well as 88,047 cell nuclei composing three of these regions. Among 33 regions, cerebral cortex areas, hypothalamus, and cerebellar gray and white matter evolved rapidly in humans. At the cellular level, astrocytes and oligodendrocyte progenitors displayed more differences in the human evolutionary lineage than the neurons. Comparison of the bulk tissue and single-nuclei sequencing revealed that conventional RNA sequencing did not detect up to two-thirds of cell-type-specific evolutionary differences.

In-depth analysis reveals complex molecular aetiology in a cohort of idiopathic cerebral palsy.

Cerebral palsy is the most prevalent physical disability in children; however, its inherent molecular mechanisms remain unclear. In the present study, we performed in-depth clinical and molecular analysis on 120 idiopathic cerebral palsy families, and identified underlying detrimental genetic variants in 45% of these patients. In addition to germline variants, we found disease-related postzygotic mutations in ∼6.7% of cerebral palsy patients. We found that patients with more severe motor impairments or a comorbidity of intellectual disability had a significantly higher chance of harbouring disease-related variants. By a compilation of 114 known cerebral-palsy-related genes, we identified characteristic features in terms of inheritance and function, from which we proposed a dichotomous classification system according to the expression patterns of these genes and associated cognitive impairments. In two patients with both cerebral palsy and intellectual disability, we revealed that the defective TYW1, a tRNA hypermodification enzyme, caused primary microcephaly and problems in motion and cognition by hindering neuronal proliferation and migration. Furthermore, we developed an algorithm and demonstrated in mouse brains that this malfunctioning hypermodification specifically perturbed the translation of a subset of proteins involved in cell cycling. This finding provided a novel and interesting mechanism for congenital microcephaly. In another cerebral palsy patient with normal intelligence, we identified a mitochondrial enzyme GPAM, the hypomorphic form of which led to hypomyelination of the corticospinal tract in both human and mouse models. In addition, we confirmed that the aberrant Gpam in mice perturbed the lipid metabolism in astrocytes, resulting in suppressed astrocytic proliferation and a shortage of lipid contents supplied for oligodendrocytic myelination. Taken together, our findings elucidate novel aspects of the aetiology of cerebral palsy and provide insights for future therapeutic strategies.

Recursive traffic percolation on urban transportation systems.

This paper proposes a recursive traffic percolation framework to capture the dynamics of cascading failures and analyze potential overloaded bottlenecks. In particular, compared to current work, the influence of external flow is considered, providing a new perspective for the study of regional commuting. Finally, we present an empirical study to verify the accuracy and effectiveness of our framework. Further analysis indicates that external flows from different regions affect the network. Our work requires only primary data and verifies the improvement of the functional network.

Recent Advances in Bacterial Persistence Mechanisms.

The recurrence of bacterial infectious diseases is closely associated with bacterial persisters. This subpopulation of bacteria can escape antibiotic treatment by entering a metabolic status of low activity through various mechanisms, for example, biofilm, toxin-antitoxin modules, the stringent response, and the SOS response. Correspondingly, multiple new treatments are being developed. However, due to their spontaneous low abundance in populations and the lack of research on in vivo interactions between persisters and the host's immune system, microfluidics, high-throughput sequencing, and microscopy techniques are combined innovatively to explore the mechanisms of persister formation and maintenance at the single-cell level. Here, we outline the main mechanisms of persister formation, and describe the cutting-edge technology for further research. Despite the significant progress regarding study techniques, some challenges remain to be tackled.

Transport and lymphatic uptake of monoclonal antibodies after subcutaneous injection.

The subcutaneous injection has emerged to become a feasible self-administration practice for biotherapeutics due to the patient comfort and cost-effectiveness. However, the available knowledge about transport and absorption of these agents after subcutaneous injection is limited. Here, a mathematical framework to study the subcutaneous drug delivery of mAbs from injection to lymphatic uptake is presented. A three-dimensional poroelastic model is exploited to find the biomechanical response of the tissue by taking into account tissue deformation during the injection. The results show that including tissue deformability noticeably changes tissue poromechanical response due to the significant dependence of interstitial pressure on the tissue deformation. Moreover, the importance of the amount of lymph fluid at the injection site and the injection rate on the drug uptake to lymphatic capillaries is highlighted. Finally, variability of lymphatic uptake due to uncertainty in parameters including tissue poromechanical and lymphatic absorption parameters is evaluated. It is found that interstitial pressure due to injection is the major contributing factor in short-term lymphatic absorption, while the amount of lymph fluid at the site of injection determines the long-term absorption of the drug. Finally, it is shown that the lymphatic uptake results are consistent with experimental data available in the literature.

Variation of microRNA expression in the human placenta driven by population identity and sex of the newborn.

BACKGROUND: Analysis of lymphocyte cell lines revealed substantial differences in the expression of mRNA and microRNA (miRNA) among human populations. The extent of such population-associated differences in actual human tissues remains largely unexplored. The placenta is one of the few solid human tissues that can be collected in substantial numbers in a controlled manner, enabling quantitative analysis of transient biomolecules such as RNA transcripts. Here, we analyzed microRNA (miRNA) expression in human placental samples derived from 36 individuals representing four genetically distinct human populations: African Americans, European Americans, South Asians, and East Asians. All samples were collected at the same hospital following a unified protocol, thus minimizing potential biases that might influence the results. RESULTS: Sequence analysis of the miRNA fraction yielded 938 annotated and 70 novel miRNA transcripts expressed in the placenta. Of them, 82 (9%) of annotated and 11 (16%) of novel miRNAs displayed quantitative expression differences among populations, generally reflecting reported genetic and mRNA-expression-based distances. Several co-expressed miRNA clusters stood out from the rest of the population-associated differences in terms of miRNA evolutionary age, tissue-specificity, and disease-association characteristics. Among three non-environmental influenced demographic parameters, the second largest contributor to miRNA expression variation after population was the sex of the newborn, with 32 miRNAs (3% of detected) exhibiting significant expression differences depending on whether the newborn was male or female. Male-associated miRNAs were evolutionarily younger and correlated inversely with the expression of target mRNA involved in neuron-related functions. In contrast, both male and female-associated miRNAs appeared to mediate different types of hormonal responses. Demographic factors further affected reported imprinted expression of 66 placental miRNAs: the imprinting strength correlated with the mother's weight, but not height. CONCLUSIONS: Our results showed that among 12 assessed demographic variables, population affiliation and fetal sex had a substantial influence on miRNA expression variation among human placental samples. The effect of newborn-sex-associated miRNA differences further led to expression inhibition of the target genes clustering in specific functional pathways. By contrast, population-driven miRNA differences might mainly represent neutral changes with minimal functional impacts.

Upregulation of Nogo-B by hypoxia inducible factor-1 and activator protein-1 in hepatocellular carcinoma.

Nogo-B is an important regulator of tumor angiogenesis. Expression of Nogo-B is remarkably upregulated in multiple tumor types, especially hepatocellular carcinoma (HCC). Here, we show the transcriptional regulation mechanisms of Nogo-B in liver cancer. In response to hypoxia, expression of Nogo-B significantly increased in HCC tissues and cells. The distal hypoxia-responsive element in the promoter was essential for transcriptional activation of Nogo-B under hypoxic conditions, which is the specific site for hypoxia inducible factor-1α (HIF-1α) binding. In addition, Nogo-B expression was associated with c-Fos expression in HCC tissues. Nogo-B expression was induced by c-Fos, yet inhibited by a dominant negative mutant A-Fos. Deletion and mutation analysis of the predicted activator protein-1 binding sites revealed that functional element mediated the induction of Nogo-B promoter activity, which was confirmed by ChIP. These results indicate that HIF-1α and c-Fos induce the expression of Nogo-B depending on tumor microenvironments, such as hypoxia and low levels of nutrients, and play a role in upregulation of Nogo-B in tumor angiogenesis.

The Interface Motion and Hydrodynamic Shear of the Liquid Slosh in Syringes.

PURPOSE: Interface motion and hydrodynamic shear of the liquid slosh during the insertion of syringes upon autoinjector activation may damage the protein drug molecules. Experimentally validated computational fluid dynamics simulations are used in this study to investigate the interfacial motion and hydrodynamic shear due to acceleration and deceleration of syringes. The goal is to explore the role of fluid viscosity, air gap size, syringe acceleration, syringe tilt angle, liquid-wall contact angle, surface tension and fill volume on the interface dynamics caused by autoinjector activation. METHODS: A simplified autoinjector platform submerged in water is built to record the syringe and liquid motion without obstruction of view. The syringe kinematics is imported to the simulations based on OpenFOAM InterIsoFoam solver, which is used to study the effects of various physical parameters. RESULTS: The simulations agree with experiments on the air-liquid interface profile and interface area. The interfacial area and the volume of fluid subject to high strain rate decrease with the solution viscosity, increase with the air gap height, syringe velocity, tilt angle and syringe wall hydrophobicity, and hardly change with the surface tension and liquid column height. The hydrodynamic shear mainly occurs near the syringe wall and entrained bubbles. CONCLUSION: For a given dose of drug solution, the syringe with smaller radius and larger length will generate less liquid slosh. Reducing the air volume and syringe wall hydrophobicity are also helpful to reduce interface area and effective shear. The interface motion is reduced when the syringe axis is aligned with the gravitational direction.

An Evaluation of Morphological Changes and Deformability of Suspended Red Blood Cells Prepared Using Whole Blood with Different Hemoglobin Levels of Tibetans.

BACKGROUND: The hypoxic environment stimulates the human body to increase the levels of hemoglobin (HGB) and hematocrit and the number of red blood cells. Such enhancements have individual differences, leading to a wide range of HGB in Tibetans' whole blood (WB). STUDY DESIGN: WB of male Tibetans was divided into 3 groups according to different HGB (i.e., A: >120 but ≤185 g/L, B: >185 but ≤210 g/L, and C: >210 g/L). Suspended red blood cells (SRBC) processed by collected WB and stored in standard conditions were examined aseptically on days 1, 14, 21, and 35 after storage. The routine biochemical indexes, deformability, cell morphology, and membrane proteins were tested. RESULTS: Mean corpuscular volume, adenosine triphosphate, pH, and deformability were not different in group A vs. those in storage (p > 0.05). The increased rate of irreversible morphology of red blood cells was different among the 3 groups, but there was no difference in the percentage of red blood cells with an irreversible morphology after 35 days of storage. Group C performed better in terms of osmotic fragility and showed a lower rigid index than group A. Furthermore, SDS-PAGE revealed similar cross-linking degrees of cell membrane protein but the band 3 protein of group C seemed to experience weaker clustering than that of group A as detected by Western Blot analysis after 35 days of storage. CONCLUSIONS: There was no difference in deformability or morphological changes in the 3 groups over the 35 days of storage. High HGB levels of plateau SRBC did not accelerate the RBC change from a biconcave disc into a spherical shape and it did not cause a reduction in deformability during 35 days of preservation in bank conditions.

Conservation, evolution, and regulation of splicing during prefrontal cortex development in humans, chimpanzees, and macaques.

Changes in splicing are known to affect the function and regulation of genes. We analyzed splicing events that take place during the postnatal development of the prefrontal cortex in humans, chimpanzees, and rhesus macaques based on data obtained from 168 individuals. Our study revealed that among the 38,822 quantified alternative exons, 15% are differentially spliced among species, and more than 6% splice differently at different ages. Mutations in splicing acceptor and/or donor sites might explain more than 14% of all splicing differences among species and up to 64% of high-amplitude differences. A reconstructed trans-regulatory network containing 21 RNA-binding proteins explains a further 4% of splicing variations within species. While most age-dependent splicing patterns are conserved among the three species, developmental changes in intron retention are substantially more pronounced in humans.

Disruption of an Evolutionarily Novel Synaptic Expression Pattern in Autism.

Cognitive defects in autism spectrum disorder (ASD) include socialization and communication: key behavioral capacities that separate humans from other species. Here, we analyze gene expression in the prefrontal cortex of 63 autism patients and control individuals, as well as 62 chimpanzees and macaques, from natal to adult age. We show that among all aberrant expression changes seen in ASD brains, a single aberrant expression pattern overrepresented in genes involved synaptic-related pathways is enriched in nucleotide variants linked to autism. Furthermore, only this pattern contains an excess of developmental expression features unique to humans, thus resulting in the disruption of human-specific developmental programs in autism. Several members of the early growth response (EGR) transcription factor family can be implicated in regulation of this aberrant developmental change. Our study draws a connection between the genetic risk architecture of autism and molecular features of cortical development unique to humans.

Self-awareness control effect of cooperative epidemics on complex networks.

Coinfection mechanism is a common interacting mode between multiple diseases in real spreading processes, where the diseases mutually increase their susceptibility, and has aroused widespread studies in network science. We use the bond percolation theory to characterize the coinfection model under two self-awareness control strategies, including immunization strategy and quarantine strategy, and to study the impacts of the synergy effect and control strategies on cooperative epidemics. We find that strengthening the synergy effect can reduce the epidemic threshold and enhance the outbreak size of coinfected networks. On Erdos-Rényi networks, the synergy effect will induce a crossover phenomenon of phase transition, i.e., make the type of phase transition from being continuous to discontinuous. Self-awareness control strategies play a non-negligible role in suppressing cooperative epidemics. In particular, increasing immunization or the quarantine rate can enhance the epidemic threshold and reduce the outbreak size of cooperative epidemics, and lead to a crossover phenomenon of transition from being discontinuous to continuous. The impact of quarantine strategy on cooperative epidemics is more significant than the immunization strategy, which is verified on scale-free networks.

Conserved expression of lincRNA during human and macaque prefrontal cortex development and maturation.

The current annotation of the human genome includes more than 12,000 long intergenic noncoding RNAs (lincRNA). While a handful of lincRNA have been shown to play important regulatory roles, the functionality of most remains unclear. Here, we examined the expression conservation and putative functionality of lincRNA in human and macaque prefrontal cortex (PFC) development and maturation. We analyzed transcriptome sequence (RNA-seq) data from 38 human and 40 macaque individuals covering the entire postnatal development interval. Using the human data set, we detected the expression of 5835 lincRNA annotated in GENCODE and further identified 1888 novel lincRNA. Most of these lincRNA show low DNA sequence conservation, as well as low expression levels. Remarkably, developmental expression patterns of these lincRNA were as conserved between humans and macaques as those of protein-coding genes. Transfection of development-associated lincRNA into human SH-SY5Y cells affected gene expression, indicating their regulatory potential. In brain, expression of these putative target genes correlated with the expression of the corresponding lincRNA during human and macaque PFC development. These results support the potential functionality of lincRNA in primate PFC development.

TCP10L acts as a tumor suppressor by inhibiting cell proliferation in hepatocellular carcinoma.

TCP10L (T-complex 10 (mouse)-like) has been identified as a liver and testis-specific gene. Although a potential transcriptional suppression function of TCP10L has been reported previously, biological function of this gene still remains largely elusive. In this study, we reported for the first time that TCP10L was significantly down-regulated in clinical hepatocellular carcinoma (HCC) samples when compared to the corresponding non-tumorous liver tissues. Furthermore, TCP10L expression was highly correlated with advanced cases exceeding the Milan criteria. Overexpression of TCP10L in HCC cells suppressed colony formation, inhibited cell cycle progression through G0/G1 phase, and attenuated cell growth in vivo. Consistently, silencing of TCP10L promoted cell cycle progression and cell growth. Therefore, our study has revealed a novel suppressor role of TCP10L in HCC, by inhibiting proliferation of HCC cells, which may facilitate the diagnosis and molecular therapy in HCC.