Biology subjects

Adam, M.

Publications and source records attributed to Adam, M..

36 records · Page 2Linked to original sources

bioRxiv2022

Vascular ultrasound for in vivo assessment of arterial pathologies in a murine model of atherosclerosis and aortic aneurysm

BackgroundVascular diseases like atherosclerosis or aortic aneurysms are common pathologies in the western world, promoting various, potentially fatal conditions. Hence, a plethora of animal models have been developed to investigate underlying mechanisms and potential therapeutics. Here we evaluate high resolution (HR) ultrasound in mouse models of atherosclerosis and abdominal aortic aneurysm (AAA) for noninvasive monitoring of morphological and functional vascular changes in vivo. MethodsEight-week-old ApoE-/- mice were used for disease models. For induction of atherosclerosis, mice were fed a western diet over 12 weeks. To trigger AAA development, osmotic minipumps were implanted, permanently releasing Angiotensin II continuously for 28 days. All animals were on C57Bl6/J background. HR vascular ultrasound of the carotid artery or the abdominal aorta was performed, respectively. Images obtained were analyzed by a speckle tracking algorithm (VevoVasc software) and were correlated with histological analyses by Picro Sirius Red staining and automated collagen quantification. ResultsArterial wall distensibility and global radial strain (GRS) as measures of arterial wall elasticity were reduced in the carotids of atherosclerotic mice as well as in the aortas of AAA mice. Pulse wave velocity (PWV) was elevated in both disease models. Intima-media thickness (IMT) was significantly increased in the atherosclerosis model. Matching those findings, area of the tunica media was enlarged in ApoE-/- mice fed a western diet, and in Angiotensin II treated mice as measured by automated image analysis, depicting higher collagen depositions in diseased arteries. Simple regression analysis revealed a strong correlation of media collagen content and area in AAA with IMT and GRS, respectively. In atherosclerosis, media collagen content significantly correlated with PWV and GRS, whereas wall distensibility was associated with the size of media area. ConclusionVascular imaging using latest generation HR ultrasound devices is suitable to trace changes of arterial wall properties in murine models of atherosclerosis and AAA. Obtained results not only correlate with histological findings but deliver information on functional parameters which may be used as early disease and risk markers in a longitudinal experimental approach.

physiology↗

bioRxiv2022

PI 3-kinase isoform p110alpha controls smooth muscle cell functionality and protects against aortic aneurysm formation

BackgroundCatalytic class IA PI 3-kinase isoform p110 is a crucial regulator of cellular proliferation and survival in numerous cell types. While p110 is critically involved in pathogenic vascular remodeling, its physiological role for vascular integrity under stress conditions has not been studied. We report a protective function of smooth muscle p110 against abdominal aortic aneurysm (AAA) formation. Methods & ResultsIn mice lacking p110 in smooth muscle cells (sm-p110-/-), perfusion of the infrarenal aorta with porcine pancreatic elastase (PPE) yielded substantially enhanced AAA formation compared to wild type controls. This disease phenotype is partly attributable to a subtle preexisting vascular phenotype under basal conditions, as sm-p110-/- mice displayed a smaller media area, deranged aortic wall structure (detached smooth muscle cells, increased apoptotic cell death), and a diminished functional responsiveness of aortic rings to vasodilators. Furthermore, p110 is also implicated in regenerative processes during AAA development: Whereas wild type mice showed increased media hypertrophy, neointima formation and proliferation upon PPE intervention, these vascular remodeling processes were diminished in sm-p110-/- mice. Concomitantly, increased numbers of elastic fiber breaks and ECM degradation were detected in sm-p110-/- aorta. Mechanistically, we found that lack of p110 expression impaired smooth muscle cell proliferation, expression of contractile marker genes and production of elastin fibers. This phenotype largely depended on reduced phosphorylation and inactivation of FOXO1, as specific FOXO1 inhibition fully rescued proliferation of p110-/- smooth muscle cells, and knockdown of FOXO1 increased expression of calponin and elastin. ConclusionsSmooth muscle p110 protects against AAA disease by maintaining aortic wall homoeostasis and promoting SMC proliferation to compensate for cell loss during AAA development. Our findings have potential implications for current approaches aimed at p110 inhibition for cancer therapy and suggest new pharmacological strategies to activate p110 signaling in AAA disease.

cell biology↗

bioRxiv2022

Inhibition of myeloperoxidase prevents thoracic aortic aneurysm formation in Marfan mice

Marfan syndrome (MFS) is the most prevalent inherited connective tissue disorder, still remains uncurable, and is characterized by high mortality at early age driven by dissection and rupture of thoracic aortic aneurysms. MFS is caused by mutations in the fibrillin-1 gene and aberrant TGF{beta} signaling. Here we addressed whether myeloperoxidase (MPO), a leukocyte derived enzyme with potent matrix modulating properties also influences the aortic phenotype in MFS. MFS patients displayed increased circulating MPO levels compared to controls as well as marked aortic MPO deposition. In an MFS mouse model, MPO induced inflammatory endothelial activation and endothelial to mesenchymal transition which triggered aortic leukocyte recruitment. Moreover, MPO directly contributed to adverse extracellular matrix remodeling by promoting oxidative stress and nitration of proteins within the vascular wall. Genetic MPO deficiency and pharmacological MPO inhibition attenuated MFS-related aneurysm formation. We herein identify MPO as a critical mediator of MFS-related thoracic aortic aneurysm formation and - in the absence of any pharmacological treatment so far in this disease - a first anti-inflammatory target to modulate disease progression.

immunology↗

bioRxiv2022

Inhibition of MLKL impairs abdominal aortic aneurysm development by attenuating smooth muscle cell necroptosis

BackgroundReceptor-interacting serine/threonine-protein kinase 1 and 3 (RIPK1 and RIPK3) dependent cell death has been identified as a crucial mediator of abdominal aortic aneurysm (AAA) development. RIPK3 mediates phosphorylation of Mixed lineage kinase domain like pseudokinase (MLKL) thereby inducing its oligomerization and translocation to the cell membrane. Given the dual role of RIPKs being involved in necroptosis as well as in apoptosis induction, the specific role of MLKL-induced necroptotic cell death in AAA remains unclear. MethodsWe monitored elastase-perfusion (PPE) induced progression of AAA in C57BL/6N (WT), RIPK1 kinase-inactive (Ripk1D138N/D138N), MLKL knockout (Mlkl-/-) and MLKL phospho-deficient (MlklAA) mice by ultrasound measurements, histological analyses and bulk mRNA sequencing to assess structural and molecular aortic changes. Bone marrow transplantations in WT and MlklAA mice were utilized to dissect the role of MLKL in smooth muscle cells (SMCs) and myeloid cells in AAA development. MLKL expressing human SMCs were generated to investigate necroptosis-induced proinflammatory cytokine secretion and subsequent polymorphonuclear neutrophil (PMN) migration and activation in vitro. ResultsUltrasound analysis showed that ~70% of the WT animals developed PPE induced-AAA with significant aortic structural alterations and enhanced myeloid cell infiltration. In contrast, Ripk1D138N/D138N, MlklAA, and Mlkl-/- mice were protected from AAA. This protection was associated with reduced adverse extracellular matrix (ECM) remodeling and leukocyte infiltration. MLKL deficiency was associated with a significant downregulation of genes involved in fibrinolysis, anti-inflammatory response, immune response and complement activation in aortic tissue in AAA. Bone marrow transplantation studies showed the lack of MLKL in SMCs to be the main driver of AAA protection. Proinflammatory cytokine secretion was elevated in necroptosis induced SMCs and resulted in a significant accumulation and activation of PMN. ConclusionsOverall, these findings indicate that MLKL-induced necroptotic SMC death and subsequent proinflammatory leukocyte activation play a causative role in AAA development and suggest that pharmacological inhibition of MLKL may represent a promising treatment strategy for AAA disease.

cell biology↗

bioRxiv2022

Single-cell transcriptomic profiling of kidney fibrosis identifies a novel specific fibroblast marker and putative disease target

BackgroundPersistent kidney fibroblast activation and tubular epithelial cell (TEC) injury are key contributors to CKD. However, transcriptional and cellular identities of advanced kidney disease, along with renal fibroblast specific markers and molecular targets contributing to persistent tubular injury, remain elusive. MethodsWe performed single-cell RNA sequencing with two clinically relevant murine kidney fibrosis models. Day 28 post-injury was chosen to ensure advanced fibrotic disease. Identified gene expression signatures were validated using multiple quantitative molecular analyses. ResultsWe revealed comprehensive single cell transcriptomic profiles of two independent kidney fibrosis models compared to normal control. Both models exhibited key CKD characteristics including renal blood flow decline, inflammatory expansion and proximal tubular loss. We identified novel populations including "secretory", "migratory" and "contractile" activated fibroblasts, specifically labelled by newly identified fibroblast-specific Gucy1a3 expression. Fibrotic kidneys elicited elevated embryonic and pro-fibrotic signaling, including separate "Embryonic" and "Pro-fibrotic" TEC clusters. Also, fibrosis caused enhanced cell-to-cell crosstalk, particularly between activated fibroblasts and pro-fibrotic TECs. Analysis of factors mediating mesenchymal phenotype in the injured epithelium identified persistent elevation of Ahnak, previously reported in AKI, in both CKD models. AHNAK knockdown in primary human renal proximal tubular epithelial cells induced a pro-fibrotic phenotype and exacerbated TGF{beta} response via p38, p42/44, pAKT, BMP and MMP signaling. ConclusionsOur study comprehensively examined kidney fibrosis in two independent models at the singe-cell resolution, providing a valuable resource for the field. Moreover, we newly identified Gucy1a3 as a kidney activated fibroblast specific marker and validated AHNAK as a putative disease target. Significance StatementMechanistic understanding of kidney fibrosis is principal for mechanistic understanding and developing targeted strategies against CKD. However, specific markers and molecular targets of key effector cells - activated kidney fibroblasts and injured tubular epithelial cells - remain elusive. Here, we created comprehensive single cell transcriptomic profiles of two clinically relevant kidney fibrosis models. We revealed "secretory", "contractile" and "migratory" fibroblasts and identified Gucy1a3 as a novel marker selectively labelling all three populations. We revealed that kidney fibrosis elicited remarkable epithelial-to-stromal crosstalk and pro-fibrotic signaling in the tubular cells. Moreover, we mechanistically validated AHNAK as a putative novel kidney injury target in a primary human in vitro model of epithelial-to-mesenchymal transition. Our findings advance understanding of and targeted intervention in fibrotic kidney disease.

molecular biology↗

bioRxiv2022

Single nuclei RNAseq analysis of HD mouse models and human brain reveals impaired oligodendrocyte maturation and potential role for thiamine metabolism

The complexity of affected brain regions and cell types is a challenge for Huntingtons disease (HD) treatment. Here we used single nucleus RNA sequencing (snRNAseq) to investigate mechanism of pathology in the cortex and striatum from R6/2 mice at 8 and 12w and in three regions of human HD post-mortem tissue. We identified cell type-specific and cell agnostic signatures and found changes suggesting oligodendrocytes (OLs) and oligodendrocyte precursors (OPCs) were arrested in intermediate maturation states. OL-lineage regulators OLIG1 and OLIG2 were negatively correlated with CAG length in human OPCs, and ATACseq analysis of HD mouse NeuN-negative cells showed decreased accessibility of sites regulated by OL maturation genes. Glucose and lipid metabolism were implicated in abnormal cell maturation and PRKCE and Thiamine Pyrophosphokinase 1 were identified as central genes. High dose thiamine/biotin treatment of R6/1 HD mice to target thiamine metabolism not only restored OL maturation, but also rescued pathology in neurons. These findings reveal insights into HD OL pathology that spans multiple brain regions and link OL maturation deficits to abnormal thiamine metabolism.

neuroscience↗

bioRxiv2022

Interstitial Notch signaling regulates nephron development via the Gata3-Renin axis in the mouse kidney

Notch signaling in the renal interstitium is known to be required for the formation of mesangial cells and Ren1 (Renin)-expressing cells. However, little is known about how interstitial Notch signaling affects nephron development. We found that blocking Notch signaling in the renal interstitium in mice caused developmental arrest of proximal tubules accompanied by defective formation of mesangial cells. We examined the interstitial Pdgfrb mutant kidney which exhibits a similar mesangial cell defect and found that the Pdgfrb mutant kidney showed normal proximal tubule development, suggesting that the absence of mesangial cells was not the cause of defective proximal tubule development. Our single cell RNA-seq analysis of the interstitial Rbpj mutant kidney showed that a subset of proximal tubule genes were downregulated in the mutant kidney and that Gata3 was downregulated in the mutant interstitium during the development of Ren1-expressing cells. We found that deleting Gata3 in the interstitium caused the loss of Renin and the developmental arrest of proximal tubules, phenocopying the interstitial Notch/Rbpj mutants. Our results suggest that interstitial Notch signaling regulates the development of proximal tubules via the Gata3-Renin axis in the mouse kidney.

developmental biology↗

bioRxiv2022

Uncovering molecular iron compounds in the living human brain.

Strict iron regulation is essential for normal brain function. The iron homeostasis, determined by the milieu of available iron compounds, is impaired in aging, neurodegenerative diseases and cancer. However, non-invasive assessment of different molecular iron environments implicating brain tissues iron homeostasis remains a challenge. We present a novel magnetic resonance imaging (MRI) technology sensitive to the iron homeostasis of the living brain (the r1-r2* relaxivity). In vitro, our MRI approach reveals the distinct paramagnetic properties of ferritin, transferrin and ferrous iron. In the in vivo human brain, we validate our approach against ex vivo iron compounds quantification and gene expression. Our approach varies with the iron mobilization capacity across brain regions and in aging. It reveals brain tumors iron homeostasis, and enhances the distinction between tumor tissue and non-pathological tissue without contrast agents. Therefore, our approach may allow for non-invasive research and diagnosis of iron homeostasis in living human brains. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=133 SRC="FIGDIR/small/490254v2_ufig1.gif" ALT="Figure 1"> View larger version (50K): org.highwire.dtl.DTLVardef@17e2722org.highwire.dtl.DTLVardef@13b646aorg.highwire.dtl.DTLVardef@3d9bfeorg.highwire.dtl.DTLVardef@cc5b7d_HPS_FORMAT_FIGEXP M_FIG Non-invasive assessment of normal and impaired iron homeostasis in living human brains. C_FIG

neuroscience↗

bioRxiv2022

Accelerated cognitive decline in obese mouse model of Alzheimer's disease is linked to sialic acid-driven immune deregulation

Systemic immunity supports healthy brain homeostasis. Accordingly, conditions causing systemic immune deregulation may accelerate onset of neurodegeneration in predisposed individuals. Here we show that, in the 5xFAD mouse model of Alzheimers disease (AD), high-fat diet-induced obesity accelerated cognitive decline, which was associated with immune deviations comprising increased splenic frequencies of exhausted CD4+ T effector memory cells and CD4+FOXP3+ regulatory T cells (Tregs). Non-targeted plasma metabolomics identified N-acetylneuraminic acid (NANA), the predominant sialic acid, as the major obesity-induced metabolite in 5xFAD mice, the levels of which directly correlated with Tregs abundance and inversely correlated with cognitive performance. Visceral adipose tissue macrophages were identified by sNuc-Seq as one potential source of NANA. Exposure to NANA led to immune deregulation in middle-aged wild-type mice, and ex vivo in human T cells. Our study identified diet-induced immune deregulation, potentially via sialic acid, as a previously unrecognized link between obesity and AD.

neuroscience↗

bioRxiv2021

The landscape of extrachromosomal circular DNA in medulloblastoma

Extrachromosomal circular DNA (ecDNA) is an important driver of aggressive tumor growth, promoting high oncogene copy number, intratumoral heterogeneity, accelerated evolution of drug resistance, enhancer rewiring, and poor outcome. ecDNA has been reported in medulloblastoma (MB), the most common malignant pediatric brain tumor, but the ecDNA landscape and its association with specific MB subgroups, its impact on enhancer rewiring, and its potential clinical implications, are not known. We assembled a retrospective cohort of 468 MB patient samples with available whole genome sequencing (WGS) data covering the four major MB subgroups WNT, SHH, Group 3 and Group 4. Using computational methods for the detection and reconstruction of ecDNA1, we find ecDNA in 82 patients (18%) and observe that ecDNA+ MB patients are more than twice as likely to relapse and three times as likely to die of disease. In addition, we find that individual medulloblastoma tumors often harbor multiple ecDNAs, each containing different amplified oncogenes along with co-amplified non-coding regulatory enhancers. ecDNA was substantially more prevalent among 31 analyzed patient-derived xenograft (PDX) models and cell lines than in our patient cohort. By mapping the accessible chromatin and 3D conformation landscapes of MB tumors that harbor ecDNA, we observe frequent candidate "enhancer rewiring" events that spatially link oncogenes with co-amplified enhancers. Our study reveals the frequency and diversity of ecDNA in a subset of highly aggressive tumors and suggests enhancer rewiring as a frequent oncogenic mechanism of ecDNAs in MB. Further, these results demonstrate that ecDNA is a frequent and potent driver of poor outcome in MB patients.

cancer biology↗

bioRxiv2021

A LINCS microenvironment perturbation resource for integrative assessment of ligand-mediated molecular and phenotypic responses

The phenotype of a cell and its underlying molecular state is strongly influenced by extracellular signals, including growth factors, hormones, and extracellular matrix. While these signals are normally tightly controlled, their dysregulation leads to phenotypic and molecular states associated with diverse diseases. To develop a detailed understanding of the linkage between molecular and phenotypic changes, we generated a comprehensive dataset that catalogs the transcriptional, proteomic, epigenomic and phenotypic responses of MCF10A mammary epithelial cells after exposure to the ligands EGF, HGF, OSM, IFNG, TGFB and BMP2. Systematic assessment of the molecular and cellular phenotypes induced by these ligands comprise the LINCS Microenvironment (ME) perturbation dataset, which has been curated and made publicly available for community-wide analysis and development of novel computational methods (synapse.org/LINCS_MCF10A). In illustrative analyses, we demonstrate how this dataset can be used to discover functionally related molecular features linked to specific cellular phenotypes.

systems biology↗

bioRxiv2020

An integrated multi-omic analysis of iPSC-derived motor neurons from C9ORF72 ALS patients

Neurodegenerative diseases present a challenge for systems biology, due to the lack of reliable animal models and the difficulties in obtaining samples from patients at early stages of disease, when interventions might be most effective. Studying induced pluripotent stem cell (iPSC)-derived neurons could overcome these challenges and dramatically accelerate and broaden therapeutic strategies. Here we undertook a network-based multi-omic characterization of iPSC-derived motor neurons from ALS patients carrying genetically dominant hexanucleotide expansions in C9orf72 to gain a deeper understanding of the relationship between DNA, RNA, epigenetics and protein in the same pool of tissue. ALS motor neurons showed the expected C9orf72-related alterations to specific nucleoporins and production of dipeptide repeats. RNA-seq, ATAC-seq and data-independent acquisition mass-spectrometry (DIA-MS) proteomics were then performed on the same motor neuron cultures. Using integrative computational methods that combined all of the omics, we discovered a number of novel dysregulated pathways including biological adhesion and extracellular matrix organization and disruption in other expected pathways such as RNA splicing and nuclear transport. We tested the relevance of these pathways in vivo in a C9orf72 Drosophila model, analyzing the data to determine which pathways were causing disease phenotypes and which were compensatory. We also confirmed that some pathways are altered in late-stage neurodegeneration by analyzing human postmortem C9 cervical spine data. To validate that these key pathways were integral to the C9 signature, we prepared a separate set of C9orf72 and control motor neuron cultures using a different differentiation protocol and applied the same methods. As expected, there were major overall differences between the differentiation protocols, especially at the level of in individual omics data. However, a number of the core dysregulated pathways remained significant using the integrated multiomic analysis. This new method of analyzing patient specific neural cultures allows the generation of disease-related hypotheses with a small number of patient lines which can be tested in larger cohorts of patients.

neuroscience↗

bioRxiv2020

Combinatorial transcriptional profiling of mouse and human enteric neurons identifies shared and disparate subtypes in situ.

AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSBACKGROUND & AIMSC_ST_ABSThe enteric nervous system (ENS) coordinates essential intestinal functions through the concerted action of diverse enteric neurons (EN). However, integrated molecular knowledge of EN subtypes is lacking. To compare human and mouse ENs, we transcriptionally profiled healthy ENS from adult humans and mice. We aimed to identify transcripts marking discrete neuron subtypes and visualize conserved EN subtypes for humans and mice in multiple bowel regions. METHODSHuman myenteric ganglia and adjacent smooth muscle were isolated by laser-capture microdissection for RNA-Seq. Ganglia-specific transcriptional profiles were identified by computationally subtracting muscle gene signatures. Nuclei from mouse myenteric neurons were isolated and subjected to single-nucleus RNA-Seq (snRNA-Seq), totaling over four billion reads and 25,208 neurons. Neuronal subtypes were defined using mouse snRNA-Seq data. Comparative informatics between human and mouse datasets identified shared EN subtype markers, which were visualized in situ using hybridization chain reaction (HCR). RESULTSSeveral EN subtypes in the duodenum, ileum, and colon are conserved between humans and mice based on orthologous gene expression. However, some EN subtype-specific genes from mice are expressed in completely distinct morphologically defined subtypes in humans. In mice, we identified several neuronal subtypes that stably express gene modules across all intestinal segments, with graded, regional expression of one or more marker genes. CONCLUSIONSOur combined transcriptional profiling of human myenteric ganglia and mouse EN provides a rich foundation for developing novel intestinal therapeutics. There is congruency among some EN subtypes, but we note multiple species differences that should be carefully considered when relating findings from mouse ENS research to human GI studies. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=123 SRC="FIGDIR/small/187211v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@1b41277org.highwire.dtl.DTLVardef@12856feorg.highwire.dtl.DTLVardef@1674dadorg.highwire.dtl.DTLVardef@11ca1c8_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG

neuroscience↗

bioRxiv2019

Single Cell Profiling of Acute Kidney Injury Reveals Novel Transcriptional Signatures, Mixed Identities and Epithelial-to-Stromal Crosstalk

Acute kidney injury (AKI) is a rapid decline of renal function, with an incidence of up to 67% of intensive care unit patients. Current treatments are merely supportive, emphasizing the need for deeper understanding that could lead to improved therapies. We used single cell RNA sequencing, in situ hybridization and protein expression analyses to create comprehensive renal cell specific transcriptional profiles of multiple AKI stages. We revealed that AKI induces marked dedifferentiation, renal developmental gene activation and mixed identities in injured renal tubules. Moreover, we identified potential pathologic crosstalk between epithelial and stromal cells, and several novel genes involved in AKI. We also demonstrated the definitive effects of age on AKI outcome, and showed that renal developmental genes hold a potential as novel AKI markers. Moreover, our study provides the resource power which will aid in unraveling the molecular genetics of AKI.

developmental biology↗

bioRxiv2019

Transcriptional regulation of MGE progenitor proliferation by PRDM16 controls cortical GABAergic interneuron production

The mammalian cortex is populated by neurons derived from neural progenitors located throughout the embryonic telencephalon. Excitatory neurons are derived from progenitors located in the dorsal telencephalon, while inhibitory interneurons are generated by ventral telencephalic progenitors. The transcriptional regulator PRDM16 is expressed by radial glia, neural progenitors present in both regions; however, its mechanisms of action are still not fully understood. It is unclear if PRDM16 functions plays a role in neurogenesis in both dorsal and ventral progenitor lineages, and if so, whether it does so by regulating common or unique networks of genes. Here, we show that Prdm16 expression in MGE progenitors is required for maintaining their proliferative capacity and for the production of proper numbers of pallial GABAergic interneurons. PRDM16 binds to cis-regulatory elements and represses the expression of region-specific neuronal differentiation genes, thereby controlling the timing of neuronal maturation. Our results highlight the importance of PRDM16 for the development of both excitatory and inhibitory cortical circuits. We demonstrate the existence of convergent developmental gene expression programs regulated by PRDM16, which utilize both common and region-specific sets of genes in the cortex and the MGE to control the proliferative capacity of neural progenitors, ensuring the generation of correct numbers of cortical neurons.

neuroscience↗

bioRxiv2019

Identification of the lymphangioleiomyomatosis cell and its uterine origin

Lymphangioleiomyomatosis (LAM) is a metastasizing neoplasm of reproductive age women that causes cystic lung remodeling and progressive respiratory failure. The source of LAM cells that invade the lung and the reasons that LAM targets women have remained elusive. We employed single cell and single nuclei RNA sequencing on LAM lesions within explanted LAM lungs, known to contain smooth muscle like cells bearing mTOR activating mutations in TSC1 or TSC2, and identified a unique population of cells that were readily distinguished from those of endogenous lung cells. LAMCORE cells shared closest transcriptomic similarity to normal uterus and neural crest. Immunofluorescence microscopy demonstrated the expression of LAMCORE cell signature genes within LAM lesions in both lung and uterus. Serum aptamer proteomics and ELISA identified biomarkers predicted to be secreted by LAMCORE cells. Single cell transcriptomics strongly supports a uterine neural crest origin of LAMCORE cells; providing insights into disease pathogenesis and informing future treatment strategies for LAM.\n\nSIGNIFICANCEPresent study identified a novel population of LAMCORE cells, which is likely originated from uterine neural crest; identified novel LAM cell-specific secretome proteins that hold promise as potential biomarkers and therapeutic targets. Advancing the understanding of LAM pathogenesis and metastasis model may yield broader insights into the biology of cancer.

cancer biology↗

bioRxiv2018

Single cell RNA-seq study of wild type and Hox9,10,11 mutant developing uterus

The uterus is a remarkable organ that must guard against infections while maintaining the ability to support growth of a fetus without rejection. The Hoxa10 and Hoxa11 genes have previously been shown to play essential roles in uterus development and function. In this report we show that the Hoxc9,10,11 genes play a redundant role in the formation of uterine glands. In addition, we use single cell RNA-seq to create a high resolution gene expression atlas of the developing wild type mouse uterus. Cell types and subtypes are defined, for example dividing endothelial cells into arterial, venous, capillary, and lymphatic, while epithelial cells separate into luminal and glandular subtypes. Further, a surprising heterogeneity of stromal and myocyte cell types are identified. Transcription factor codes and ligand/receptor interactions are characterized. We also used single cell RNA-seq to globally define the altered gene expression patterns in all developing uterus cell types for two Hox mutants, with 8 or 9 mutant Hox genes. The mutants show a striking disruption of Wnt signaling as well as the Cxcl12/Cxcr4 ligand/receptor axis.\n\nSummary statementA single cell RNA-seq study of the developing mouse uterus defines cellular heterogeneities, lineage specific gene expression programs and perturbed pathways in Hox9,10,11 mutants.

developmental biology↗

bioRxiv2018

Ehmt1 forebrain haploinsufficiency leads to impaired memory, sensory gating and information processing

Regulators of chromatin dynamics and transcription are increasingly implicated in the aetiology of neurodevelopmental disorders (NDDs). Haploinsufficiency of EHMT1, encoding a histone methyl-transferase, is associated with several NDDs, including Kleefstra syndrome, developmental delay and autism spectrum disorder. Using a mouse model of Ehmt1 haploinsufficiency (Ehmt1D6Cre/+), we examined a number of brain and behavioural endophenotypes of relevance to NDDs. Specifically, we show that Ehmt1D6Cre/+ mice have deficits in information processing, evidenced by abnormal sensory-motor gating, a complete absence of object recognition memory and a reduced magnitude of auditory evoked potentials in both paired-pulse inhibition and mismatch negativity (MMN). The electrophysiological experiments show that differences in magnitude response to auditory stimulus were associated with marked reductions in total and evoked beta- and gamma-band oscillatory activity, as well as significant reductions in phase synchronisation. The pattern of electrophysiological deficits in Ehmt1D6Cre/+ matches those seen in control mice following administration of the selective NMDA-R antagonist, ketamine. This, coupled with reduction of Grin1 mRNA expression in Ehmt1D6Cre/+ hippocampus, suggests that Ehmt1 haploinsufficiency may lead to disruption in NMDA-R. Taken together, these data indicate that reduced Ehmt1 dosage during forebrain development leads to abnormal circuitry formation, which in turn results in profound information processing deficits. Such information processing deficits are likely paramount to our understanding of the cognitive and neurological dysfunctions shared across the NDDs associated with EHMT1 haploinsufficiency.

neuroscience↗