Chromatin and gene-regulatory dynamics of the developing human cerebral cortex at single-cell resolution

1. Download : Download high-res image (270KB)
2. Download : Download full-size image

     

Stem cell-based modeling and single-cell multiomics reveal gene-regulatory mechanisms underlying human skeletal development

1. Download : Download high-res image (343KB)
2. Download : Download full-size image

     

Random forest for gene selection and microarray data classification

A random forest method has been selected to perform both gene selection and classification of the microarray data. In this embedded method, the selection of smallest possible sets of genes with lowest error rates is the key factor in achieving highest classification accuracy. Hence, improved gene selection method using random forest has been proposed to obtain the smallest subset of genes as well as biggest subset of genes prior to classification. The option for biggest subset selection is done to assist researchers who intend to use the informative genes for further research. Enhanced random forest gene selection has performed better in terms of selecting the smallest subset as well as biggest subset of informative genes with lowest out of bag error rates through gene selection. Furthermore, the classification performed on the selected subset of genes using random forest has lead to lower prediction error rates compared to existing method and other similar available methods.     

Failure to obtain hybridomas between human macrophages and human tumoral U-937 cells is probably due to parental macrophages

Summary Despite more than 50 attempts and the use of various methods, it has been impossible to establish homologous hybridomas between human mature macrophages and 8-azaguanine-resistant U-937 clones prepared in the laboratory. To rule out the possibility that these clones were unsuitable for the selection of hybrids, a study of their properties was done. It was shown that U-937 wild type cells were able to produce HPRT, whereas 8-azaguanine (8-aza)-resistant clones did not. Curiously, exonic and intronic HPRT sequences were undetectable both in wild type and in 8-aza-resistant cell genomes, under conditions where they were detected in control cells. Chromosome analysis of the clone UM9, one of the most frequently used in fusion experiments, revealed many qualitative and quantitative differences with the U-937 wild type cells. 8-aza-resistant U-937 cells were capable of fusion with human macrophages and gave rise to heterokaryons and probably to synkaryons, which survived for weeks without dividing in hypoxanthine-aminopterin-thymidine medium. The results could be interpreted in terms of the existence of a transacting negative regulatory mechanism of the macrophage genome on the proliferative capacity of homospecific hybridomas.     

Proteomics and metabolomics analyses reveal the cucurbit sieve tube system as a complex metabolic space

The plant vascular system, and specifically the phloem, plays a pivotal role in allocation of fixed carbon to developing sink organs. Although the processes involved in loading and unloading of sugars and amino acids are well characterized, little information is available regarding the nature of other metabolites in the sieve tube system (STS) at specific sites along the pathway. Here, we elucidate spatial features of metabolite composition mapped with phloem enzymes along the cucurbit STS. Phloem sap (PS) was collected from the loading (source), unloading (apical sink region) and shoot–root junction regions of cucumber, watermelon and pumpkin. Our PS analyses revealed significant differences in the metabolic and proteomic profiles both along the source–sink pathway and between the STSs of these three cucurbits. In addition, metabolite profiles established for PS and vascular tissue indicated the presence of distinct compositions, consistent with the operation of the STS as a unique symplasmic domain. In this regard, at various locations along the STS we could map metabolites and their related enzymes to specific metabolic pathways. These findings are discussed with regard to the function of the STS as a unique and highly complex metabolic space within the plant vascular system.     

Global metabolic profiling to model biological processes of aging in twins

Aging is intimately linked to system‐wide metabolic changes that can be captured in blood. Understanding biological processes of aging in humans could help maintain a healthy aging trajectory and promote longevity. We performed untargeted plasma metabolomics quantifying 770 metabolites on a cross‐sectional cohort of 268 healthy individuals including 125 twin pairs covering human lifespan (from 6 months to 82 years). Unsupervised clustering of metabolic profiles revealed 6 main aging trajectories throughout life that were associated with key metabolic pathways such as progestin steroids, xanthine metabolism, and long‐chain fatty acids. A random forest (RF) model was successful to predict age in adult subjects (≥16 years) using 52 metabolites (R² = .97). Another RF model selected 54 metabolites to classify pediatric and adult participants (out‐of‐bag error = 8.58%). These RF models in combination with correlation network analysis were used to explore biological processes of healthy aging. The models highlighted established metabolites, like steroids, amino acids, and free fatty acids as well as novel metabolites and pathways. Finally, we show that metabolic profiles of twins become more dissimilar with age which provides insights into nongenetic age‐related variability in metabolic profiles in response to environmental exposure.     

Ionic channels and membrane hyperpolarization in human macrophages

Summary Microelectrode impalement of human macrophages evokes a transient hyperpolarizing response (HR) of the membrane potential. This HR was found to be dependent on the extracellular concentration of K⁺ but not on that of Na⁺ or Cl^–. It was not influenced by low temperature (12°C) or by 0.2mm ouabain, but was blocked by 0.2mm quinine or 0.2mm Mg²⁺-EGTA. These findings indicate that the HR in human macrophages is caused by the activation of a K⁺ (Ca²⁺) conductance. Two types of ionic channels were identified in intact cells by use of the patch-clamp technique in the cell-attached-patch configuration, low and high-conductance voltage-dependent K⁺ channels. The low-conductance channels had a mean conductance of 38 pS with Na⁺-saline and 32 pS with K⁺-saline in the pipette. The high-coductance channels had a conductance of 101 and 114 pS with Na⁺- and K⁺-saline in the pipette, respectively. Cell-attached patch measurements made during evocation of an HR by microelectrode penetration showed enhanced channel activity associated with the development of the HR. These channels were also high-conductance channels (171 pS with Na⁺- and 165 pS K⁺-saline in the pipette) and were voltage dependent. They were, however, active at less positive potentials than the high-conductance K⁺ channels seen prior to the microelectrode-evoked HR. It is concluded that the high-conductance voltage-dependent ionic channels active during the HR in human macrophages contribute to the development of the HR.     

随机森林模型在分类与回归分析中的应用

随机森林(random forest)模型是由Breiman和Cutler在2001年提出的一种基于分类树的算法。它通过对大量分类树的汇总提高了模型的预测精度,是取代神经网络等传统机器学习方法的新的模型。随机森林的运算速度很快,在处理大数据时表现优异。随机森林不需要顾虑一般回归分析面临的多元共线性的问题,不用做变量选择。现有的随机森林软件包给出了所有变量的重要性。另外,随机森林便于计算变量的非线性作用,而且可以体现变量间的交互作用(interaction)。它对离群值也不敏感。本文通过3个案例,分别介绍了随机森林在昆虫种类的判别分析、有无数据的分析(取代逻辑斯蒂回归)和回归分析上的应用。案例的数据格式和R语言代码可为研究随机森林在分类与回归分析中的应用提供参考。     

The single-cell and spatial transcriptional landscape of human gastrulation and early brain development

1. Download : Download high-res image (214KB)
2. Download : Download full-size image

     

Phenotype correlations reveal the relationships of physiological systems underlying human ageing

Ageing is characterized by degeneration and loss of function across multiple physiological systems. To study the mechanisms and consequences of ageing, several metrics have been proposed in a hierarchical model, including biological, phenotypic and functional ageing. In particular, phenotypic ageing and interconnected changes in multiple physiological systems occur in all ageing individuals over time. Recently, phenotypic age, a new ageing measure, was proposed to capture morbidity and mortality risk across diverse subpopulations in US cohort studies. Although phenotypic age has been widely used, it may overlook the complex relationships among phenotypic biomarkers. Considering the correlation structure of these phenotypic biomarkers, we proposed a composite phenotype analysis (CPA) strategy to analyse 71 biomarkers from 2074 individuals in the Rugao Longitudinal Ageing Study. CPA grouped these biomarkers into 18 composite phenotypes according to their internal correlation, and these composite phenotypes were mostly consistent with prior findings. In addition, compared with prior findings, this strategy exhibited some different yet important implications. For example, the indicators of kidney and cardiovascular functions were tightly connected, implying internal interactions. The composite phenotypes were further verified through associations with functional metrics of ageing, including disability, depression, cognitive function and frailty. Compared to age alone, these composite phenotypes had better predictive performances for functional metrics of ageing. In summary, CPA could reveal the hidden relationships of physiological systems and identify the links between physiological systems and functional ageing metrics, thereby providing novel insights into potential mechanisms underlying human ageing.     

Understanding heterogeneity of human bone marrow plasma cell maturation and survival pathways by single-cell analyses

1. Download : Download high-res image (273KB)
2. Download : Download full-size image

     

森林结构复杂度对单木分割精度的影响——以田横岛为例

单木分割对于森林资源调查具有重要的意义,不同结构复杂度的森林单木分割算法的选择以及分割参数的选取对分割精度有着很大的影响。以山东田横岛为研究区,基于无人机正射影像与激光雷达数据,首先提取海岛森林典型植被二维与三维特征,然后利用随机森林算法对不同树种的树木进行分类,最后基于分类后的点云数据,选取不同结构复杂度的森林样地,对比分析聚类算法、层堆叠算法、分水岭算法在不同复杂度林区的适用性。结果表明:(1)随机森林算法结合单木二维、三维特征可有效对混交林树木进行分类,模型总体的精度为94.51%,Kappa系数为0.9038;(2)聚类算法对结构简单的林区具有更高的分割精度(F=96.41),但依赖于分割参数的选取;面对复杂单木集群,分水岭算法总体得分波动最小(ΔF=14.56),表现出较强的稳定性;(3)混交林预先进行树种分类可有效改善单木分割环境,相比于直接进行单木分割,聚类算法、层堆叠算法、分水岭算法的分割精度均得到不同程度的提升(ΔF₁=10.06,ΔF₂=9.51,ΔF₃=12.6)。     

基于随机森林模型的茹河流域水沙变化及其影响因素研究

水沙关系不协调已成为制约黄河高质量发展的关键问题,揭示径流量与输沙量变化的主要影响因素,对适应生态经济发展条件下的水土流失治理工作具有指导意义。以茹河流域为研究对象,采用M-K检验和Pettitt检验分析茹河流域1989—2019年水沙变化趋势,运用双累积曲线法定量研究了降雨和人类活动对茹河流域径流量、输沙量变化的贡献率,通过随机森林回归分析探索了影响水沙变化的主控因子。结果表明：(1)茹河流域径流量和输沙量峰值对应情况良好,但在2012年之后径流量和输沙量呈现出明显分异,径流量逐渐增加,输沙量平稳减小。(2)茹河流域径流量突变点为1996年和2003年,输沙量突变点为1996年和2004年,影响期的两个阶段中,人类活动对径流量的贡献率分别为95.95%和77.52%,对输沙量的贡献率分别为78.31%和77.91%。(3)显著影响径流量的因子为农作物播种面积、耕地面积、草地面积和人口密度,显著影响输沙量的因子为植被覆盖度、水利环境投资占比、人口密度和工业总产值。(4)农作物播种面积和耕地面积的变化对径流量的影响均呈显著正相关,草地面积和人口密度对径流量响应关系为显著负相关;植被覆盖度...     

Effect of cytokines and lipopolysaccharide on CD14 antigen expression in human monocytes and macrophages.

The 52 kD myeloid membrane glycoprotein CD14 represents the receptor for complexes of lipopolysaccharide (LPS) and LPS binding protein (LBP); it is involved in LPS induced tumor necrosis factor-alpha production. Expression of CD14 increases in monocytes differentiating into macrophages, and it is reduced by rIFNg in monocytes in vitro. In the present study CD14 membrane antigen expression was investigated in cultures of human mononuclear leucocytes (PBL), in elutriated, purified monocytes, and in blood monocyte derived Teflon cultured macrophages. Cells were incubated for 15 or 45 h with rIL-1, rIL-2, rIL-3, rIL-5, rIL-6, rTNFa, rGM-CSF, rM-CSF, rTGFb1, rIFNa, lipopolysaccharide (LPS), and, as a control, rIFNg. The monoclonal antibodies Leu-M3 and MEM 18 were used for labelling of CD14 antigen by indirect immunofluorescence and FACS analysis of scatter gated monocytes or macrophages. IFNg concentrations were determined in PBL culture supernatants by ELISA. rIFNa and rIL-2 reduced CD14 in 15 and 45 h PBL cultures, an effect mediated by endogenous IFNg, since it was abolished by simultaneous addition of an anti-IFNg antibody. rIFNa and rIL-2 were ineffective in purified monocytes or macrophages. rIL-4 strongly reduced CD14 in PBL and purified monocytes after 45 h, whereas in macrophages the decrease was weak, although measurable after 15 h. The other cytokines investigated did not change CD14 antigen expression. Cycloheximide alone reduced CD14, but when added in combination with rIFNg the effect on CD14 downregulation was more pronounced. The effect of rIFNg on CD14 in PBL cultures was dose-dependently inhibited by rIL-4 and this inhibition is probably due to an IL-4 mediated blockade of IFNg secretion. LPS at a low dose increased CD14, at a high dose it produced a variable decrease of CD14 in PBL, which was probably due to LPS induced IFNg secretion. LPS strongly enhanced CD14 in 45 h cultures of purified monocytes. The results, showing that CD14 antigen expression is upregulated by LPS and downregulated by rIFNg and rIL-4, suggest that the LPS-LBP receptor is involved in the feedback response of IFNg and IL-4 to LPS stimulation.     

On the mechanism of binding of human lymphocyte products to guinea pig macrophages

When lymphocytes from a majority of patients with cancer are incubated with encephalitogenic factor, a lymphocyte product is released that reduces the anodic electrophoretic mobilities of guinea pig macrophages and fixed, tanned sheep erythrocytes. Although these reactions are not specific for cancer, it is distinctly possible that in patients with cancer, products from stimulated lymphocytes are capable of altering the surfaces of the patients' own macrophages, thereby modifying the course of their disease. In this paper, we attempt to elucidate some mechanisms for the binding of lymphocyte products to macrophages, such as occurs in the macrophage electrophoretic mobility (MEM) test, since this may be of general interest. Binding of lymphocyte product to macrophages has been monitored by measurements of their electrophoretic mobilities and by electron microscopic determination of the density of binding of electron-dense, cationic colloidal iron hydroxide particles to their surfaces. The results show that the lymphocyte products reduce the net surface negativity of the macrophages by (coulombic) binding of this net positively charged material to sialic acids at the macrophage surface. Product-binding can be prevented by prior treatment of the macrophages with neuraminidase. It appears that only a minority of sialic acids are involved in the binding process, which occurs without demonstrable blocking of adjacent sialic acids or redistribution of such sites over the macrophage surface. Parallel experiments with fixed tanned erythrocytes also suggest that binding of lymphocyte products is not solely determined by surface sialic acids, although it cannot occur without them.     

Prolactin induces chitotriosidase expression in human macrophages through PTK,PI3‐K,and MAPK pathways

We previously reported that prolactin (PRL) induces chitotriosidase (CHIT‐1) mRNA expression in human macrophages. In this investigation we determined the signaling pathways involved in CHIT‐1 induction in response to PRL. The CHIT‐1 induction PRL‐mediated was reduced by wortmannin and LY‐294002, inhibitors of phosphatidylinositol 3‐kinase (PI3‐K) and by genistein an inhibitor of protein tyrosine kinase (PTK). Pre‐treatment of macrophages with SB203580, a specific inhibitor of the mitogen‐activated kinases (MAPK) p38, or with U0126, an inhibitor of MAPK p44/42, prevented both basal and exogenous PRL‐mediated CHIT‐1 expression. No significant effects on CHIT‐1 induction PRL‐mediated were observed with a protein kinase C inhibitor (PKC), rottlerin, or with an Src inhibitor, PP2, or with JAK2 inhibitor, AG490. In addition, PRL induced a phosphorylation of AKT that was prevented both by the two MAPK inhibitors SB203580 and U0126 and by the PI3‐K inhibitors wortmannin and LY‐294002. In conclusion, our results indicate that PRL up‐regulated CHIT‐1 expression via PTK, PI3‐K, MAPK, and signaling transduction components. J. Cell. Biochem. 107: 881–889, 2009. © 2009 Wiley‐Liss, Inc.     

Serum metabolomics reveals many novel metabolic markers of heart failure, including pseudouridine and 2-oxoglutarate

There is intense interest in the identification of novel biomarkers which improve the diagnosis of heart failure. Serum samples from 52 patients with systolic heart failure (EF < 40% plus signs and symptoms of failure) and 57 controls were analyzed by gas chromatography – time of flight – mass spectrometry and the raw data reduced to 272 statistically robust metabolite peaks. 38 peaks showed a significant difference between case and control (p < 5 × 10⁻⁵). Two such metabolites were pseudouridine, a modified nucleotide present in t- and rRNA and a marker of cell turnover, as well as the tricarboxylic acid cycle intermediate 2-oxoglutarate. Furthermore, 3 further new compounds were also excellent discriminators between patients and controls: 2-hydroxy, 2-methylpropanoic acid, erythritol and 2,4,6-trihydroxypyrimidine. Although renal disease may be associated with heart failure, and metabolites associated with renal disease and other markers were also elevated (e.g. urea, creatinine and uric acid), there was no correlation within the patient group between these metabolites and our heart failure biomarkers, indicating that these were indeed biomarkers of heart failure and not renal disease per se. These findings demonstrate the power of data-driven metabolomics approaches to identify such markers of disease. Warwick B. Dunn, David I. Broadhurst and Sasalu M. Deepak contributed equally to this work.     

Developmental landscape of human forebrain at a single-cell level identifies early waves of oligodendrogenesis

1. Download : Download high-res image (184KB)
2. Download : Download full-size image

     

Mycobacterial growth in human macrophages: variation according to donor, inoculum and bacterial strain

The microbicidal capacity of the macrophage is frequently evaded by mycobacteria, leading to tuberculosis (TB). We investigated a number of parameters affecting the rate of growth of mycobacteria in human monocyte-derived macrophages (MDM). The results show a great deal of variation in the growth of both Mycobacterium bovis BCG and M. tuberculosis H37Rv, using a large number of human macrophage donors, (132 and 40, respectively), but no correlation was seen with the TB status of the MDM donor. Clumping of the mycobacteria resulted in more vigorous growth in MDM, suggesting that inoculum size could affect disease progression. The growth rates of 17 clinical isolates of M. tuberculosis were measured in macrophages derived from three donors and no consistent or marked differences between isolates were observed over the 5-day period of growth measurement. However, all 17 clinical strains grew consistently faster than H37Rv in the same experiments.