CR1 variants contribute to FSGS susceptibility across multiple populations.

Focal segmental glomerulosclerosis (FSGS) is a common cause of nephrotic syndrome with an annual incidence in the United States in African-Americans compared to European-Americans of 24 cases and 5 cases per million, respectively. Among glomerular diseases in Europe and Latin-America, FSGS was the second most frequent diagnosis, and in Asia the fifth. We expand previous efforts in understanding genetics of FSGS by performing a case-control study involving ethnically-diverse groups FSGS cases (726) and a pool of controls (13,994), using panel sequencing of approximately 2,500 podocyte-expressed genes. Through rare variant association tests, we replicated known risk genes - KANK1, COL4A4, and APOL1. A novel significant association was observed for the gene encoding complement receptor 1 (CR1). High-risk rare variants in CR1 in the European-American cohort were commonly observed in Latin- and African-Americans. Therefore, a combined rare and common variant analysis was used to replicate the CR1 association in non-European populations. The CR1 risk variant, rs17047661, gives rise to the Sl1/Sl2 (R1601G) allele that was previously associated with protection against cerebral malaria. Pleiotropic effects of rs17047661 may explain the difference in allele frequencies across continental ancestries and suggest a possible role for genetically-driven alterations of adaptive immunity in the pathogenesis of FSGS.

Systems Immunology Approaches to Metabolism.

Over the last decade, immunometabolism has emerged as a novel interdisciplinary field of research and yielded significant fundamental insights into the regulation of immune responses. Multiple classical approaches to interrogate immunometabolism, including bulk metabolic profiling and analysis of metabolic regulator expression, paved the way to appreciating the physiological complexity of immunometabolic regulation in vivo. Studying immunometabolism at the systems level raised the need to transition towards the next-generation technology for metabolic profiling and analysis. Spatially resolved metabolic imaging and computational algorithms for multi-modal data integration are new approaches to connecting metabolism and immunity. In this review, we discuss recent studies that highlight the complex physiological interplay between immune responses and metabolism and give an overview of technological developments that bear the promise of capturing this complexity most directly and comprehensively.

Gene, cell type, and drug prioritization analysis suggest genetic basis for the utility of diuretics in treating Alzheimer disease.

We introduce a user-friendly tool for risk gene, cell type, and drug prioritization for complex traits: GCDPipe. It uses gene-level GWAS-derived data and gene expression data to train a model for the identification of disease risk genes and relevant cell types. Gene prioritization information is then coupled with known drug target data to search for applicable drug agents based on their estimated functional effects on the identified risk genes. We illustrate the utility of our approach in different settings: identification of the cell types, implicated in disease pathogenesis, was tested in inflammatory bowel disease (IBD) and Alzheimer disease (AD); gene target and drug prioritization was tested in IBD and schizophrenia. The analysis of phenotypes with known disease-affected cell types and/or existing drug candidates shows that GCDPipe is an effective tool to unify genetic risk factors with cellular context and known drug targets. Next, analysis of the AD data with GCDPipe suggested that gene targets of diuretics, as an Anatomical Therapeutic Chemical drug subgroup, are significantly enriched among the genes prioritized by GCDPipe, indicating their possible effect on the course of the disease.

Tumor-associated macrophages respond to chemotherapy by detrimental transcriptional reprogramming and suppressing stabilin-1 mediated clearance of EGF.

Introduction: Tumor resistance to chemotherapy and metastatic relapse account for more than 90% of cancer specific mortality. Tumor-associated macrophages (TAMs) can process chemotherapeutic agents and impair their action. Little is known about the direct effects of chemotherapy on TAMs. Methods: The effect of chemotherapeutic platinum agent cisplatin was assessed in the model system of human ex vivo TAMs. Whole-transcriptome sequencing for paired TAMs stimulated and not stimulated by cisplatin was analysed by NGS. Endocytic uptake of EGF was quantified by flow cytometry. Confocal microscopy was used to visualize stabilin-1-mediated internalization and endocytic trafficking of EGF in CHO cells expressing ectopically recombinant stabilin-1 and in stabilin-1+ TAMs. In cohort of patients with breast cancer, the effect of platinum therapy on the transcriptome of TAMs was validated, and differential expression of regulators of endocytosis was identified. Results: Here we show that chemotherapeutic agent cisplatin can initiate detrimental transcriptional and functional programs in TAMs, without significant impairment of their viability. We focused on the clearance function of TAMs that controls composition of tumor microenvironment. For the first time we demonstrated that TAMs' scavenger receptor stabilin-1 is responsible for the clearance of epidermal growth factor (EGF), a potent stimulator of tumor growth. Cisplatin suppressed both overall and EGF-specific endocytosis in TAMs by bidirectional mode: suppression of positive regulators and stimulation of negative regulators of endocytosis, with strongest effect on synaptotagmin-11 (SYT11), confirmed in patients with breast cancer. Conclusion: Our data demonstrate that synergistic action of cytostatic agents and innovative immunomodulators is required to overcome cancer therapy resistance.

Network analysis of large-scale ImmGen and Tabula Muris datasets highlights metabolic diversity of tissue mononuclear phagocytes.

The diversity of mononuclear phagocyte (MNP) subpopulations across tissues is one of the key physiological characteristics of the immune system. Here, we focus on understanding the metabolic variability of MNPs through metabolic network analysis applied to three large-scale transcriptional datasets: we introduce (1) an ImmGen MNP open-source dataset of 337 samples across 26 tissues; (2) a myeloid subset of ImmGen Phase I dataset (202 MNP samples); and (3) a myeloid mouse single-cell RNA sequencing (scRNA-seq) dataset (51,364 cells) assembled based on Tabula Muris Senis. To analyze such large-scale datasets, we develop a network-based computational approach, genes and metabolites (GAM) clustering, for unbiased identification of the key metabolic subnetworks based on transcriptional profiles. We define 9 metabolic subnetworks that encapsulate the metabolic differences within MNP from 38 different tissues. Obtained modules reveal that cholesterol synthesis appears particularly active within the migratory dendritic cells, while glutathione synthesis is essential for cysteinyl leukotriene production by peritoneal and lung macrophages.

Integrated multi-omics analysis of adverse cardiac remodeling and metabolic inflexibility upon ErbB2 and ERRα deficiency.

Functional oncogenic links between ErbB2 and ERRα in HER2+ breast cancer patients support a therapeutic benefit of co-targeted therapies. However, ErbB2 and ERRα also play key roles in heart physiology, and this approach could pose a potential liability to cardiovascular health. Herein, using integrated phosphoproteomic, transcriptomic and metabolic profiling, we uncovered molecular mechanisms associated with the adverse remodeling of cardiac functions in mice with combined attenuation of ErbB2 and ERRα activity. Genetic disruption of both effectors results in profound effects on cardiomyocyte architecture, inflammatory response and metabolism, the latter leading to a decrease in fatty acyl-carnitine species further increasing the reliance on glucose as a metabolic fuel, a hallmark of failing hearts. Furthermore, integrated omics signatures of ERRα loss-of-function and doxorubicin treatment exhibit common features of chemotherapeutic cardiotoxicity. These findings thus reveal potential cardiovascular risks in discrete combination therapies in the treatment of breast and other cancers.

Genotype imputation and polygenic score estimation in northwestern Russian population.

Numerous studies demonstrated the lack of transferability of polygenic score (PGS) models across populations and the problem arising from unequal presentation of ancestries across genetic studies. However, even within European ancestry there are ethnic groups that are rarely presented in genetic studies. For instance, Russians, being one of the largest, diverse, and yet understudied group in Europe. In this study, we evaluated the reliability of genotype imputation for the Russian cohort by testing several commonly used imputation reference panels (e.g. HRC, 1000G, HGDP). HRC, in comparison with two other panels, showed the most accurate results based on both imputation accuracy and allele frequency concordance between masked and imputed genotypes. We built polygenic score models based on GWAS results from the UK biobank, measured the explained phenotypic variance in the Russian cohort attributed to polygenic scores for 11 phenotypes, collected in the clinic for each participant, and finally explored the role of allele frequency discordance between the UK biobank and the study cohort in the resulting PGS performance.

Case Report: Supernormal Vascular Aging in Leningrad Siege Survivors.

Age-related changes in the vascular system play an important role in the biological age and lifespan of a person and maybe affected from an early age onward. One of the indicators of changes in the vascular system is arterial wall stiffness and its main measure, i.e., carotid-femoral pulse wave velocity (cfPWV). We examined arterial wall stiffness in a sample of 305 Leningrad Siege survivors to assess how hunger and stressful conditions during fetal development and early childhood affected the state of the cardiovascular system at a later age and what factors may neutralize the negative impact sustained in early childhood. Here, we presented an evaluation of two unique patients with supernormal vascular aging (SUPERNOVA) phenotype from this cohort and described the details of congruence between hereditary resistance and practiced lifestyle yielding slower biological aging rate.

Shiny GATOM: omics-based identification of regulated metabolic modules in atom transition networks.

Multiple high-throughput omics techniques provide different angles on systematically quantifying and studying metabolic regulation of cellular processes. However, an unbiased analysis of such data and, in particular, integration of multiple types of data remains a challenge. Previously, for this purpose we developed GAM web-service for integrative metabolic network analysis. Here we describe an updated pipeline GATOM and the corresponding web-service Shiny GATOM, which takes as input transcriptional and/or metabolomic data and finds a metabolic subnetwork most regulated between the two conditions of interest. GATOM features a new metabolic network topology based on atom transition, which significantly improves interpretability of the analysis results. To address computational challenges arising with the new network topology, we introduce a new variant of the maximum weight connected subgraph problem and provide a corresponding exact solver. To make the used networks up-to-date we upgraded the KEGG-based network construction pipeline and developed one based on the Rhea database, which allows analysis of lipidomics data. Finally, we simplified local installation, providing R package mwcsr for solving relevant graph optimization problems and R package gatom, which implements the GATOM pipeline. The web-service is available at https://ctlab.itmo.ru/shiny/gatom and https://artyomovlab.wustl.edu/shiny/gatom.

GWAS of depression in 4,520 individuals from the Russian population highlights the role of MAGI2 (S-SCAM) in the gut-brain axis.

We present the results of the depression Genome-wide association studies study performed on a cohort of Russian-descent individuals, which identified a novel association at chromosome 7q21 locus. Gene prioritization analysis based on already known depression risk genes indicated MAGI2 (S-SCAM) as the most probable gene from the locus and potential susceptibility gene for the disease. Brain and gut expression patterns were the main features highlighting functional relatedness of MAGI2 to the previously known depression risk genes. Local genetic covariance analysis, analysis of gene expression, provided initial suggestive evidence of hospital anxiety and depression scale and diagnostic and statistical manual of mental disorders scales having a different relationship with gut-brain axis disturbance. It should be noted, that while several independent methods successfully in silico validate the role of MAGI2, we were unable to replicate genetic association for the leading variant in the MAGI2 locus, therefore the role of rs521851 in depression should be interpreted with caution.

Non-canonical glutamine transamination sustains efferocytosis by coupling redox buffering to oxidative phosphorylation.

Macrophages rely on tightly integrated metabolic rewiring to clear dying neighboring cells by efferocytosis during homeostasis and disease. Here we reveal that glutaminase-1-mediated glutaminolysis is critical to promote apoptotic cell clearance by macrophages during homeostasis in mice. In addition, impaired macrophage glutaminolysis exacerbates atherosclerosis, a condition during which, efficient apoptotic cell debris clearance is critical to limit disease progression. Glutaminase-1 expression strongly correlates with atherosclerotic plaque necrosis in patients with cardiovascular diseases. High-throughput transcriptional and metabolic profiling reveals that macrophage efferocytic capacity relies on a non-canonical transaminase pathway, independent from the traditional requirement of glutamate dehydrogenase to fuel ɑ-ketoglutarate-dependent immunometabolism. This pathway is necessary to meet the unique requirements of efferocytosis for cellular detoxification and high-energy cytoskeletal rearrangements. Thus, we uncover a role for non-canonical glutamine metabolism for efficient clearance of dying cells and maintenance of tissue homeostasis during health and disease in mouse and humans.

Loss of Mir146b with aging contributes to inflammation and mitochondrial dysfunction in thioglycollate-elicited peritoneal macrophages.

Macrophages undergo programmatic changes with age, leading to altered cytokine polarization and immune dysfunction, shifting these critical immune cells from protective sentinels to disease promoters. The molecular mechanisms underlying macrophage inflammaging are poorly understood. Using an unbiased RNA sequencing (RNA-seq) approach, we identified Mir146b as a microRNA whose expression progressively and unidirectionally declined with age in thioglycollate-elicited murine macrophages. Mir146b deficiency led to altered macrophage cytokine expression and reduced mitochondrial metabolic activity, two hallmarks of cellular aging. Single-cell RNA-seq identified patterns of altered inflammation and interferon gamma signaling in Mir146b-deficient macrophages. Identification of Mir146b as a potential regulator of macrophage aging provides novel insights into immune dysfunction associated with aging.

Markov chain Monte Carlo for active module identification problem.

BACKGROUND: Integrative network methods are commonly used for interpretation of high-throughput experimental biological data: transcriptomics, proteomics, metabolomics and others. One of the common approaches is finding a connected subnetwork of a global interaction network that best encompasses significant individual changes in the data and represents a so-called active module. Usually methods implementing this approach find a single subnetwork and thus solve a hard classification problem for vertices. This subnetwork inherently contains erroneous vertices, while no instrument is provided to estimate the confidence level of any particular vertex inclusion. To address this issue, in the current study we consider the active module problem as a soft classification problem. RESULTS: We propose a method to estimate probabilities of each vertex to belong to the active module based on Markov chain Monte Carlo (MCMC) subnetwork sampling. As an example of the performance of our method on real data, we run it on two gene expression datasets. For the first many-replicate expression dataset we show that the proposed approach is consistent with an existing resampling-based method. On the second dataset the jackknife resampling method is inapplicable due to the small number of biological replicates, but the MCMC method can be run and shows high classification performance. CONCLUSIONS: The proposed method allows to estimate the probability that an individual vertex belongs to the active module as well as the false discovery rate (FDR) for a given set of vertices. Given the estimated probabilities, it becomes possible to provide a connected subgraph in a consistent manner for any given FDR level: no vertex can disappear when the FDR level is relaxed. We show, on both simulated and real datasets, that the proposed method has good computational performance and high classification accuracy.

A 300-kb microduplication of 7q36.3 in a patient with triphalangeal thumb-polysyndactyly syndrome combined with congenital heart disease and optic disc coloboma: a case report.

BACKGROUND: Triphalangeal thumb-polysyndactyly syndrome (TPT-PS) is a rare well-defined autosomal dominant disorder characterized by long thumbs with three phalanges combined with pre- and postaxial polydactyly/syndactyly of limbs. By now, the syndrome has been reported in several large families from different ethnic backgrounds, with a high degree of inter- and intrafamilial variability. The genome locus responsible for TPT-PS has been mapped to the 7q36.3 region harboring a long-range sonic hedgehog (SHH) regulatory sequence (ZRS). Both single-nucleotide variants and complete duplications of ZRS were shown to cause TPT-PS and similar limb phenotypes. TPT-PS usually forms as isolated limb pathology not associated with additional malformations, in particular, with cardiovascular abnormalities. CASE PRESENTATION: Here we report on a rare Russian neonatal case of TPT-PS combined with severe congenital heart disease, namely double outlet right ventricle, and microphthalmia with optic disc coloboma. Pedigree analysis revealed TPT-PS of various expressivity in 10 family members throughout five generations, while the cardiac defect and the eye pathology were detected only in the proband. To extend the knowledge on genotype-phenotype spectrum of TPT-PS, the careful clinical and genomic analysis of the family was performed. High-resolution array-based comparative genomic hybridization (array-CGH) revealed a ~ 300 kb microduplication of 7q36.3 locus (arr[GRCh37] 7q36.3(156385810_156684811) × 3) that co-segregated with TPT-PS in the proband and her mother. The duplication encompassed three genes including LMBR1, the intron 5 of which is known to harbor ZRS. Based on whole-exome sequencing data, no additional pathogenic mutations or variants of uncertain clinical significance were found in morbid cardiac genes or genes associated with a microphthalmia/anophthalmia/coloboma spectrum of ocular malformations. CONCLUSIONS: The results support the previous data, indicating that complete ZRS duplication underlies TPT-PS, and suggest a broader phenotypic impact of the 7q36.3 microduplication. Potential involvement of the 7q36.3 microduplication in the patient's cardiac and eye malformations is discussed. However, the contribution of some additional genetic/epigenetic factors to the complex patient`s phenotype cannot be excluded entirely. Further comprehensive functional studies are needed to prove the possible involvement of the 7q36.3 locus in congenital heart disease and eye pathology.

LMNA Mutations G232E and R482L Cause Dysregulation of Skeletal Muscle Differentiation, Bioenergetics, and Metabolic Gene Expression Profile.

Laminopathies are a family of monogenic multi-system diseases resulting from mutations in the LMNA gene which include a wide range of neuromuscular disorders. Although lamins are expressed in most types of differentiated cells, LMNA mutations selectively affect only specific tissues by mechanisms that remain largely unknown. We have employed the combination of functional in vitro experiments and transcriptome analysis in order to determine how two LMNA mutations associated with different phenotypes affect skeletal muscle development and metabolism. We used a muscle differentiation model based on C2C12 mouse myoblasts genetically modified with lentivirus constructs bearing wild-type human LMNA (WT-LMNA) or R482L-LMNA/G232E-LMNA mutations, linked to familial partial lipodystrophy of the Dunnigan type and muscular dystrophy phenotype accordingly. We have shown that both G232E/R482L-LMNA mutations cause dysregulation in coordination of pathways that control cell cycle dynamics and muscle differentiation. We have also found that R482/G232E-LMNA mutations induce mitochondrial uncoupling and a decrease in glycolytic activity in differentiated myotubes. Both types of alterations may contribute to mutation-induced muscle tissue pathology.

Tonic TCR Signaling Inversely Regulates the Basal Metabolism of CD4+ T Cells.

The contribution of self-peptide-MHC signaling in CD4+ T cells to metabolic programming has not been definitively established. In this study, we employed LLO118 and LLO56, two TCRtg CD4+ T cells that recognize the same Listeria epitope. We previously have shown that LLO56 T cells are highly self-reactive and respond poorly in a primary infection, whereas LLO118 cells, which are less self-reactive, respond well during primary infection. We performed metabolic profiling and found that naive LLO118 had a dramatically higher basal respiration rate, a higher maximal respiration rate, and a higher glycolytic rate relative to LLO56. The LLO118 cells also exhibited a greater uptake of 2-NBD-glucose, in vitro and in vivo. We extended the correlation of low self-reactivity (CD5lo) with high basal metabolism using two other CD4+ TCRtg cells with known differences in self-reactivity, AND and Marilyn. We hypothesized that the decreased metabolism resulting from a strong interaction with self was mediated through TCR signaling. We then used an inducible knock-in mouse expressing the Scn5a voltage-gated sodium channel. This channel, when expressed in peripheral T cells, enhanced basal TCR-mediated signaling, resulting in decreased respiration and glycolysis, supporting our hypothesis. Genes and metabolites analysis of LLO118 and LLO56 T cells revealed significant differences in their metabolic pathways, including the glycerol phosphate shuttle. Inhibition of this pathway reverts the metabolic state of the LLO118 cells to be more LLO56 like. Overall, these studies highlight the critical relationship between peripheral TCR-self-pMHC interaction, metabolism, and the immune response to infection.

Generation of two iPSC lines (FAMRCi007-A and FAMRCi007-B) from patient with Emery-Dreifuss muscular dystrophy and heart rhythm abnormalities carrying genetic variant LMNA p.Arg249Gln.

Human iPSC lines were generated from peripheral blood mononuclear cells of patient carrying LMNA mutation associated with Emery-Dreifuss muscular dystrophy accompanied by atrioventricular block and paroxysmal atrial fibrillation. Reprogramming factors OCT4, KLF4, SOX2, CMYC were delivered using Sendai virus transduction. iPSCs were characterized in order to prove the pluripotency markers expression, normal karyotype, ability to differentiate into three embryonic germ layers. Generated iPSC lines would be useful model to investigate disease development associated with genetic variants in LMNA gene.

Generation of two iPSC lines (FAMRCi006-A and FAMRCi006-B) from patient with dilated cardiomyopathy and Emery-Dreifuss muscular dystrophy associated with genetic variant LMNAp.Arg527Pro.

Mutations in LMNA gene are known to cause a broad range of diseases called laminopathies. We have generated two induced pluripotent stem cell lines FAMRCi006-A and FAMRCi006-B from a patient carrying LMNA p. p.Arg527Pro mutation associated with Emery-Dreifuss muscular dystrophy and dilated cardiomyopathy. Patient-specific peripheral blood mononuclear cells were reprogrammed to iPSCs using Sendai virus reprogramming system. Characterization of iPSCs had revealed pluripotency marker expression, normal karyotype, ability to differentiate into three embryonic germ layers. The reported iPSC lines could be a useful tool for in vitro modeling of laminopathies associated with LMNA genetic variants.

Transcriptomic profiling of experimental arterial injury reveals new mechanisms and temporal dynamics in vascular healing response.

OBJECTIVE: Endovascular interventions cause arterial injury and induce a healing response to restore vessel wall homeostasis. Complications of defective or excessive healing are common and result in increased morbidity and repeated interventions. Experimental models of intimal hyperplasia are vital for understanding the vascular healing mechanisms and resolving the clinical problems of restenosis, vein graft stenosis, and dialysis access failure. Our aim was to systematically investigate the transcriptional, histologic, and systemic reaction to vascular injury during a prolonged time. METHODS: Balloon injury of the left common carotid artery was performed in male rats. Animals (n = 69) were euthanized before or after injury, either directly or after 2 hours, 20 hours, 2 days, 5 days, 2 weeks, 6 weeks, and 12 weeks. Both injured and contralateral arteries were subjected to microarray profiling, followed by bioinformatic exploration, histologic characterization of the biopsy specimens, and plasma lipid analyses. RESULTS: Immune activation and coagulation were key mechanisms in the early response, followed by cytokine release, tissue remodeling, and smooth muscle cell modulation several days after injury, with reacquisition of contractile features in later phases. Novel pathways related to clonal expansion, inflammatory transformation, and chondro-osteogenic differentiation were identified and immunolocalized to neointimal smooth muscle cells. Analysis of uninjured arteries revealed a systemic component of the reaction after local injury, underlined by altered endothelial signaling, changes in overall tissue bioenergy metabolism, and plasma high-density lipoprotein levels. CONCLUSIONS: We demonstrate that vascular injury induces dynamic transcriptional landscape and metabolic changes identifiable as early, intermediate, and late response phases, reaching homeostasis after several weeks. This study provides a temporal "roadmap" of vascular healing as a publicly available resource for the research community.

Clinical Response to Personalized Exercise Therapy in Heart Failure Patients with Reduced Ejection Fraction is Accompanied by Skeletal Muscle Histological Alterations.

Abstract: Heart failure (HF) is associated with skeletal muscle wasting and exercise intolerance. This study aimed to evaluate the exercise-induced clinical response and histological alterations. One hundred and forty-four HF patients were enrolled. The individual training program was determined as a workload at or close to the lactate threshold (LT1); clinical data were collected before and after 12 weeks/6 months of training. The muscle biopsies from eight patients were taken before and after 12 weeks of training: histology analysis was used to evaluate muscle morphology. Most of the patients demonstrated a positive response after 12 weeks of the physical rehabilitation program in one or several parameters tested, and 30% of those showed improvement in all four of the following parameters: oxygen uptake (VO2) peak, left ventricular ejection fraction (LVEF), exercise tolerance (ET), and quality of life (QOL); the walking speed at LT1 after six months of training showed a significant rise. Along with clinical response, the histological analysis detected a small but significant decrease in both fiber and endomysium thickness after the exercise training course indicating the stabilization of muscle mechanotransduction system. Together, our data show that the beneficial effect of personalized exercise therapy in HF patients depends, at least in part, on the improvement in skeletal muscle physiological and biochemical performance.