Mapping the Mouse Cell Atlas by Microwell-Seq.

Single-cell RNA sequencing (scRNA-seq) technologies are poised to reshape the current cell-type classification system. However, a transcriptome-based single-cell atlas has not been achieved for complex mammalian systems. Here, we developed Microwell-seq, a high-throughput and low-cost scRNA-seq platform using simple, inexpensive devices. Using Microwell-seq, we analyzed more than 400,000 single cells covering all of the major mouse organs and constructed a basic scheme for a mouse cell atlas (MCA). We reveal a single-cell hierarchy for many tissues that have not been well characterized previously. We built a web-based "single-cell MCA analysis" pipeline that accurately defines cell types based on single-cell digital expression. Our study demonstrates the wide applicability of the Microwell-seq technology and MCA resource.

Construction of a human cell landscape at single-cell level.

Single-cell analysis is a valuable tool for dissecting cellular heterogeneity in complex systems1. However, a comprehensive single-cell atlas has not been achieved for humans. Here we use single-cell mRNA sequencing to determine the cell-type composition of all major human organs and construct a scheme for the human cell landscape (HCL). We have uncovered a single-cell hierarchy for many tissues that have not been well characterized. We established a 'single-cell HCL analysis' pipeline that helps to define human cell identity. Finally, we performed a single-cell comparative analysis of landscapes from human and mouse to identify conserved genetic networks. We found that stem and progenitor cells exhibit strong transcriptomic stochasticity, whereas differentiated cells are more distinct. Our results provide a useful resource for the study of human biology.

Construction of a cross-species cell landscape at single-cell level.

Individual cells are basic units of life. Despite extensive efforts to characterize the cellular heterogeneity of different organisms, cross-species comparisons of landscape dynamics have not been achieved. Here, we applied single-cell RNA sequencing (scRNA-seq) to map organism-level cell landscapes at multiple life stages for mice, zebrafish and Drosophila. By integrating the comprehensive dataset of > 2.6 million single cells, we constructed a cross-species cell landscape and identified signatures and common pathways that changed throughout the life span. We identified structural inflammation and mitochondrial dysfunction as the most common hallmarks of organism aging, and found that pharmacological activation of mitochondrial metabolism alleviated aging phenotypes in mice. The cross-species cell landscape with other published datasets were stored in an integrated online portal-Cell Landscape. Our work provides a valuable resource for studying lineage development, maturation and aging.

How many cell types are there in nature? How do they change during the life cycle? These are two fundamental questions that researchers have been trying to understand in the area of biology. In this study, single-cell mRNA sequencing data were used to profile over 2.6 million individual cells from mice, zebrafish and Drosophila at different life stages, 1.3 million of which were newly collected. The comprehensive datasets allow investigators to construct a cross-species cell landscape that helps to reveal the conservation and diversity of cell taxonomies at genetic and regulatory levels. The resources in this study are assembled into a publicly available website at http://bis.zju.edu.cn/cellatlas/.

Large Room Temperature Magnetization Enhancement in a Copper-Based Photoactive Metal-Organic Framework.

The tris(pyridin-4-yl)amine ligand was found to exhibit a radical-actuated coloration phenomenon, and a novel copper-based color-changeable metal-organic framework (MOF) was synthesized via this photoactive ligand. After light irradiation, the photogenerated stable radicals in this framework induced increasing amplitude of magnetization (32%) at room temperature, being the largest enhancement among radical-based photochromic systems.

An Efficient Agrobacterium rhizogenes-Mediated Hairy Root Transformation Method in a Soybean Root Biology Study.

The stable genetic transformation of soybean is time-consuming and inefficient. As a simple and practical alternative method, hairy root transformation mediated by Agrobacterium rhizogenes is widely applied in studying root-specific processes, nodulation, biochemical and molecular functions of genes of interest, gene editing efficiency of CRISPR/Cas9, and biological reactors and producers. Therefore, many laboratories have developed unique protocols to obtain hairy roots in composite plants composed of transgenic roots and wild-type shoots. However, these protocols still suffer from the shortcomings of low efficiency and time, space, and cost consumption. To address this issue, we developed a new protocol efficient regeneration and transformation of hairy roots (eR&T) in soybean, by integrating and optimizing the main current methods to achieve high efficiency in both hairy root regeneration and transformation within a shorter period and using less space. By this eR&T method, we obtained 100% regeneration of hairy roots for all explants, with an average 63.7% of transformation frequency, which promoted the simultaneous and comparative analysis of the function of several genes. The eR&T was experimentally verified Promoter:GUS reporters, protein subcellular localization, and CRISPR/Cas9 gene editing experiments. Employing this approach, we identified several novel potential regulators of nodulation, and nucleoporins of the Nup107-160 sub-complex, which showed development-dependent and tissue-dependent expression patterns, indicating their important roles in nodulation in soybean. Thus, the new eR&T method is an efficient and economical approach for investigating not only root and nodule biology, but also gene function.

Application of fractional calculus methods to viscoelastic behaviours of solid propellants.

A three-branch viscoelastic model based on fractional derivatives is proposed for the viscoelastic behaviours of solid propellants. The simulation results show a satisfactory agreement with the stress relaxation modulus and complex modulus of solid propellants. As a comparison, the static modulus is also characterized by traditional viscoelastic model with integer-order derivatives. Results show that the application of the fractional derivatives to the viscoelastic constitutive model can effectively reduce the number of the required parameters while giving an accurate prediction of viscoelastic behaviours of solid propellants. Moreover, a simple and effective direct search method based on simulated annealing and Powell's mothed is proposed for the data fitting. This article is part of the theme issue 'Advanced materials modelling via fractional calculus: challenges and perspectives'.

Association of TCF4 polymorphisms and Fuchs' endothelial dystrophy: a meta-analysis.

BACKGROUND: Studies investigating the associations between transcription factor 4 (TCF4) genetic polymorphisms and Fuchs' endothelial dystrophy (FED) have reported controversial results. Therefore, this meta-analysis aims to clarify the effects of TCF4 polymorphisms on FED risk. METHODS: A meta-analysis was conducted to assess the association between four single nucleotide polymorphisms (SNPs) inTCF4 and the risk of FED. Relevant studies were selected through an extensive search of PubMed, EMBASE, and the Web of Science databases. Pooled odds ratio (OR) and 95 % confidence interval (CI) were calculated using the random-effects model. RESULTS: Thirteen studies were included in this systematic review and meta-analysis. The pooled results showed that there was a strong positive association between the TCF4 rs613872 polymorphism and FED risk in all the genetic models tested (G allele vs. T allele: OR = 4.19, 95 % CI = 3.53-4.97; GG vs. GT/TT: OR = 4.27, 95 % CI = 2.54-7.19; GG/GT vs. TT: OR = 6.29, 95 % CI = 4.23-8.93; GG VS. TT: OR = 10.64, 95 % CI = 5.28-21.41; GT VS. TT: OR = 6.08, 95 % CI = 4.28-8.64). Statistic evidence was also detected for a significant association between three other SNPs and the risk of FED. CONCLUSIONS: This meta-analysis suggested a genetic association between four TCF4 polymorphisms (rs613872, rs2286812, rs17595731, and rs9954153) and the risk of FED.

[Ophthalmic manifestations in acquired immune deficiency syndrome patients with cryptococcal meningitis].

OBJECTIVE: Analysis ophthalmic manifestations in acquired immune deficiency syndrome (AIDS) patients with cryptococcal meningitis. METHODS: Retrospective study. Forty AIDS patients (including 32 men and 8 women) with cryptococcal meningitis were studied. The age of the patients varied from 11 to 67 years, average on (38 ± 12) years (mean ± standard deviation). All the patients underwent detailed ocular examinations including corrected visual acuity, eye movement, intraocular pressure, slit lamp microscope, indirect ophthalmoscope and color fundus photography. The CD4(+) T cell count was evaluated and Lumbar puncture was done to examine the cerebral spinal fluid (CSF). Eighteen patients were followed up for a period between 2 weeks to 4 years. Four patients died during hospitalization. The remaining 18 patients failed to be followed up due to moving away. RESULTS: Binocular diplopia were present in 3 patients. It showed Binocular or monocular abducens nerve palsy. Papilledema were found in 37 eyes of 19 cases. One person had unilateral papilledema. Among 19 cases, 24 eyes of 12 cases were accompanied with peripapillary hemorrhages and 7 cases of 13 eyes without peripapillary hemorrhages. Optic atrophy occurs in 6 cases of 12 eyes and multifocal choroiditis in 2 cases of 4 eyes. Twenty-seven eyes of 14 patients had normal fundus or cytomegalovirus retinitis (CMVR) only. CONCLUSIONS: AIDS patients with cryptococcal meningitis merger may be accompanied by a variety of eye complications. Papilledema are the most common manifestations.

Reversible acetylation regulates salt-inducible kinase (SIK2) and its function in autophagy.

Salt-inducible kinase 2 (SIK2) is a serine/threonine protein kinase belonging to the AMP-activated protein kinase (AMPK) family. SIK2 has been shown to function in the insulin-signaling pathway during adipocyte differentiation and to modulate CREB-mediated gene expression in response to hormones and nutrients. However, molecular mechanisms underlying the regulation of SIK2 kinase activity remains largely elusive. Here we report a dynamic, post-translational regulation of its kinase activity that is coordinated by an acetylation-deacetylation switch, p300/CBP-mediated Lys-53 acetylation inhibits SIK2 kinase activity, whereas HDAC6-mediated deacetylation restores the activity. Interestingly, overexpression of acetylation-mimetic mutant of SIK2 (SIK2-K53Q), but not the nonacetylatable K53R variant, resulted in accumulation of autophagosomes. Further consistent with a role in autophagy, knockdown of SIK2 abrogated autophagosome and lysosome fusion. Consequently, SIK2 and its kinase activity are indispensable for the removal of TDP-43Δ inclusion bodies. Our findings uncover SIK2 as a critical determinant in autophagy progression and further suggest a mechanism in which the interplay among kinase and deacetylase activities contributes to cellular protein pool homeostasis.

Homocysteine, methylenetetrahydrofolate reductase C677T polymorphism, and risk of retinal vein occlusion: an updated meta-analysis.

BACKGROUND: To evaluate the role of plasma total homocysteine (tHcy) and homozygosity for the thermolabile variant of the methylenetetrahydrofolate reductase (MTHFR) C677T genotype in the risk of retinal vein occlusion (RVO). METHODS: Relevant studies were selected through an extensive search of PubMed, EMBASE, and the Web of Science databases. Summary weighted mean differences (WMDs) or odds ratios (ORs) and 95% confidence intervals (CI) were calculated with a random-effects model. RESULTS: Forty-two studies with 6445 participants were included in this updated systematic review and meta-analysis. The mean plasma tHcy level in the RVO patients was significantly higher than in the controls (WMD =2.13 µmol/L; 95% CI: 1.29 to 2.98, P < 0.001), but there was evidence of between-study heterogeneity (P < 0.001). No significant association between MTHFR C677T genotype and RVO was found under all genetic models. CONCLUSION: There was some evidence that plasma tHcy is associated with an increased risk of RVO. There was no evidence to suggest an association between homozygosity for the MTHFR C677T genotype and RVO.

Camellia oil exhibits anti-fatigue property by modulating antioxidant capacity, muscle fiber, and gut microbial composition in mice.

Camellia seed oil (CO) has high nutritional value and multiple bioactivities. However, the specific anti-fatigue characteristics and the implied mechanism of CO have not yet been fully elucidated. Throughout this investigation, male C57BL/6J mice, aged 8 weeks, underwent exhaustive exercise with or without CO pretreatment (2, 4, and 6 mL/kg BW) for 28 days. CO could extend the rota-rod and running time, reduce blood urea nitrogen levels and serum lactic acid, and increase muscle and hepatic glycogen, adenosine triphosphate, and anti-oxidative indicators. Additionally, CO could upregulate the mRNA and Nrf2 protein expression levels, as well as enhance the levels of its downstream antioxidant enzymes and induce the myofiber-type transformation from fast to slow and attenuate the gut mechanical barrier. Moreover, CO could ameliorate gut dysbiosis by reducing Firmicutes to Bacteroidetes ratio at the phylum level, increasing the percentage of Alistipes, Alloprevotella, Lactobacillus, and Muribaculaceae, and decreasing the proportion of Dubosiella at the genus level. In addition, specific bacterial taxa, which were altered by CO, showed a significant correlation with partial fatigue-related parameters. These findings suggest that CO may alleviate fatigue by regulating antioxidant capacity, muscle fiber transformation, gut mechanical barrier, and gut microbial composition in mice. PRACTICAL APPLICATION: Our study revealed that camellia seed oil (CO) could ameliorate exercise-induced fatigue in mice by modulating antioxidant capacity, muscle fiber, and gut microbial composition in mice. Our results promote the application of CO as an anti-fatigue functional food that targets oxidative stress, myofiber-type transformation, and microbial community.

Single-cell landscape analysis reveals systematic senescence in mammalian Down syndrome.

BACKGROUND: Down syndrome (DS), which is characterized by various malfunctions, is the most common chromosomal disorder. As the DS population continues to grow and most of those with DS live beyond puberty, early-onset health problems have become apparent. However, the cellular landscape and molecular alterations have not been thoroughly studied. METHODS: This study utilized single-cell resolution techniques to examine DS in humans and mice, spanning seven distinct organs. A total of 71 934 mouse and 98 207 human cells were analyzed to uncover the molecular alterations occurring in different cell types and organs related to DS, specifically starting from the fetal stage. Additionally, SA-ß-Gal staining, western blot, and histological study were employed to verify the alterations. RESULTS: In this study, we firstly established the transcriptomic profile of the mammalian DS, deciphering the cellular map and molecular mechanism. Our analysis indicated that DS cells across various types and organs experienced senescence stresses from as early as the fetal stage. This was marked by elevated SA-ß-Gal activity, overexpression of cell cycle inhibitors, augmented inflammatory responses, and a loss of cellular identity. Furthermore, we found evidence of mitochondrial disturbance, an increase in ribosomal protein transcription, and heightened apoptosis in fetal DS cells. This investigation also unearthed a regulatory network driven by an HSA21 gene, which leads to genome-wide expression changes. CONCLUSION: The findings from this study offer significant insights into the molecular alterations that occur in DS, shedding light on the pathological processes underlying this disorder. These results can potentially guide future research and treatment development for DS.

Polysaccharides from Enteromorpha prolifera alleviate hypercholesterolemia via modulating the gut microbiota and bile acid metabolism.

Polysaccharides from Enteromorpha prolifera (EP) possess multiple biological activities, while the role of EP in hypercholesterolemia and its relationship with the gut microbiota have not been elucidated. To address this issue, fifty male C57BL/6J mice were randomly subjected to a basal diet and a high-fat and high-cholesterol diet, and 3 treatment groups were fed an HFHC diet supplemented with different dosages of EP (100, 200 and 300 mg kg-1 day-1) for 12 weeks. Here we show that EP intervention lowered serum concentrations of total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) and inhibited hepatic cholesterol deposition. EP intervention also upregulated the gene expression related to the hepatic cholesterol uptake and bile acid synthetic pathway. Apart from that, EP altered the gut microbiota, pre-dominantly increasing microbes associated with bile acid metabolism, such as norank_f_ Muribaculaceae. Moreover, bile acid profile analysis revealed that EP could alter the fecal bile acid profile and reduce fecal conjugated bile acids. Further correlation analysis indicated the negative correlation of Bacteroides, norank_f_ Muribaculaceae and Ileibacterium abundance with the levels of fecal conjugated bile acids and serum TC and LDL-C, while the abundance of Proteobacteria and Lachnoclosteridium showed a positive association with conjugated bile acids and serum TC. To sum up, the above findings revealed that EP may alleviate hypercholesterolemia and regulate cholesterol metabolism in ways that promote a favorable fecal microbiota composition and modulate bile acid metabolism.

Systematic identification of cell-fate regulatory programs using a single-cell atlas of mouse development.

Waddington's epigenetic landscape is a metaphor frequently used to illustrate cell differentiation. Recent advances in single-cell genomics are altering our understanding of the Waddington landscape, yet the molecular mechanisms of cell-fate decisions remain poorly understood. We constructed a cell landscape of mouse lineage differentiation during development at the single-cell level and described both lineage-common and lineage-specific regulatory programs during cell-type maturation. We also found lineage-common regulatory programs that are broadly active during the development of invertebrates and vertebrates. In particular, we identified Xbp1 as an evolutionarily conserved regulator of cell-fate determinations across different species. We demonstrated that Xbp1 transcriptional regulation is important for the stabilization of the gene-regulatory networks for a wide range of mouse cell types. Our results offer genetic and molecular insights into cellular gene-regulatory programs and will serve as a basis for further advancing the understanding of cell-fate decisions.

Cell landscape of larval and adult Xenopus laevis at single-cell resolution.

The rapid development of high-throughput single-cell RNA sequencing technology offers a good opportunity to dissect cell heterogeneity of animals. A large number of organism-wide single-cell atlases have been constructed for vertebrates such as Homo sapiens, Macaca fascicularis, Mus musculus and Danio rerio. However, an intermediate taxon that links mammals to vertebrates of more ancient origin is still lacking. Here, we construct the first Xenopus cell landscape to date, including larval and adult organs. Common cell lineage-specific transcription factors have been identified in vertebrates, including fish, amphibians and mammals. The comparison of larval and adult erythrocytes identifies stage-specific hemoglobin subtypes, as well as a common type of cluster containing both larval and adult hemoglobin, mainly at NF59. In addition, cell lineages originating from all three layers exhibits both antigen processing and presentation during metamorphosis, indicating a common regulatory mechanism during metamorphosis. Overall, our study provides a large-scale resource for research on Xenopus metamorphosis and adult organs.

Tracing cell-type evolution by cross-species comparison of cell atlases.

Cell types are the basic building units of multicellular life, with extensive diversities. The evolution of cell types is a crucial layer of comparative cell biology but is thus far not comprehensively studied. We define a compendium of cell atlases using single-cell RNA-seq (scRNA-seq) data from seven animal species and construct a cross-species cell-type evolutionary hierarchy. We present a roadmap for the origin and diversity of major cell categories and find that muscle and neuron cells are conserved cell types. Furthermore, we identify a cross-species transcription factor (TF) repertoire that specifies major cell categories. Overall, our study reveals conservation and divergence of cell types during animal evolution, which will further expand the landscape of comparative genomics.

Polymerase chain reaction analysis of aqueous humor specimens in the diagnosis of cytomegalovirus retinitis in AIDS patients.

PURPOSE: To determine the value of the polymerase chain reaction analysis of aqueous humor specimens as a tool to diagnose cytomegalovirus retinitis in AIDS patients. METHODS: In all, 63 AIDS patients were evaluated in this study. They were sorted into two diagnostic categories: eyes with active cytomegalovirus retinitis and eyes without active cytomegalovirus retinitis. The aqueous humor and blood samples were collected and analyzed by polymerase chain reaction. RESULTS: A total of 49 patients had active cytomegalovirus retinitis (77.8%) and 14 patients had inactive cytomegalovirus retinitis or normal fundus (22.2%). The mean average of patients was 39 years (range: 22-59). The majority of patients were male (90.5%). Cytomegalovirus DNA was detected in 46 and 7 of 49 aqueous and blood samples, respectively, from AIDS patients with active cytomegalovirus retinitis. We did not detect cytomegalovirus DNA in any of the eyes without active cytomegalovirus retinitis. The sensitivity of polymerase chain reaction in the detection of cytomegalovirus in aqueous humor and blood samples was 93.5% and 14.3%, respectively. CONCLUSIONS: The polymerase chain reaction analysis is a safe, highly specific, and sensitive method to diagnose cytomegalovirus retinitis.

A single-cell survey of cellular hierarchy in acute myeloid leukemia.

BACKGROUND: Acute myeloid leukemia (AML) is a fatal hematopoietic malignancy and has a prognosis that varies with its genetic complexity. However, there has been no appropriate integrative analysis on the hierarchy of different AML subtypes. METHODS: Using Microwell-seq, a high-throughput single-cell mRNA sequencing platform, we analyzed the cellular hierarchy of bone marrow samples from 40 patients and 3 healthy donors. We also used single-cell single-molecule real-time (SMRT) sequencing to investigate the clonal heterogeneity of AML cells. RESULTS: From the integrative analysis of 191727 AML cells, we established a single-cell AML landscape and identified an AML progenitor cell cluster with novel AML markers. Patients with ribosomal protein high progenitor cells had a low remission rate. We deduced two types of AML with diverse clinical outcomes. We traced mitochondrial mutations in the AML landscape by combining Microwell-seq with SMRT sequencing. We propose the existence of a phenotypic "cancer attractor" that might help to define a common phenotype for AML progenitor cells. Finally, we explored the potential drug targets by making comparisons between the AML landscape and the Human Cell Landscape. CONCLUSIONS: We identified a key AML progenitor cell cluster. A high ribosomal protein gene level indicates the poor prognosis. We deduced two types of AML and explored the potential drug targets. Our results suggest the existence of a cancer attractor.

scMCA: A Tool to Define Mouse Cell Types Based on Single-Cell Digital Expression.

For decades, people have been trying to define cell type with the combination of expressed genes. The choice of the limited number of genes for the classification limits the precision of this system. Here, we build a "single-cell Mouse Cell Atlas (scMCA) analysis" pipeline based on scRNA-seq datasets covering all mouse cell types. We build the scMCA reference and then use the tool "scMCA" to match single-cell digital expression to its closest cell type.