Oligodendrocyte Cell Research Articles

Adolescent binge ethanol impacts H3K9me3-occupancy at synaptic genes and the regulation of oligodendrocyte development.

Binge drinking in adolescence can disrupt myelination and cause brain structural changes that persist into adulthood. Alcohol consumption at a younger age increases the susceptibility of these changes. Animal models to understand ethanol's actions on myelin and white matter show that adolescent binge ethanol can alter the developmental trajectory of oligodendrocytes, myelin structure, and myelin fiber density. Oligodendrocyte differentiation is epigenetically regulated by H3K9 trimethylation (H3K9me3). Prior studies have shown that adolescent binge ethanol dysregulates H3K9 methylation and decreases H3K9-related gene expression in the PFC. Here, we assessed ethanol-induced changes to H3K9me3 occupancy at genomic loci in the developing adolescent PFC. We further assessed ethanol-induced changes at the transcription level with qPCR time course approaches in oligodendrocyte-enriched cells to assess changes in oligodendrocyte progenitor and oligodendrocytes specifically. Adolescent binge ethanol altered H3K9me3 regulation of synaptic-related genes and genes specific for glutamate and potassium channels in a sex-specific manner. In PFC tissue, we found an early change in gene expression in transcription factors associated with oligodendrocyte differentiation that may lead to the later significant decrease in myelin-related gene expression. This effect appeared stronger in males. Further exploration in oligodendrocyte cell enrichment time course and dose response studies could suggest lasting dysregulation of oligodendrocyte maturation at the transcriptional level. Overall, these studies suggest that binge ethanol may impede oligodendrocyte differentiation required for ongoing myelin development in the PFC by altering H3K9me3 occupancy at synaptic-related genes. We identify potential genes that may be contributing to adolescent binge ethanol-related myelin loss.

Understanding the Role of CDH4 in Multiple System Atrophy Brain.

Multiple system atrophy (MSA) is a rapidly progressive neurodegenerative disease clinically characterized by parkinsonism, cerebellar ataxia, and autonomic dysfunction. A major pathological feature of MSA is the presence of α-synuclein aggregates in oligodendrocytes, the myelinating cells of the central nervous system. A genome-wide association study revealed that the CDH4 gene is associated with MSA. However, virtually nothing is known about the role of CDH4 in the context of MSA. Our aim was to compare the expression of CDH4 between MSA and control brains, and to investigate its relationship with α-synuclein in oligodendrocytes. RNA and protein were prepared from putamen, motor cortex white matter, cerebellum, and superior occipital cortex tissues collected from MSA (N = 11) and control (N = 13) brains. The expression of CDH4 was measured at mRNA and protein levels by qPCR and western blotting. Oligodendrocyte cells were cultured on plates and transfected with CDH4 cDNA and its impact on α-synuclein was analyzed. Firstly, we found that CDH4 in MSA brain was significantly elevated in the disease-affected motor cortex white matter in MSA (N = 11) compared to controls (N = 13) and unaltered in the disease-unaffected superior occipital cortex. Secondly, we determined that increases in CDH4 expression caused changes in the cellular levels of α-synuclein in oligodendrocytes. When put together, these results provide evidence that support the GWAS association of CDH4 with MSA.

APOE4 impairs myelination via cholesterol dysregulation in oligodendrocytes

AbstractBackgroundAPOE4 is the strongest genetic risk factor for late‐onset Alzheimer’s disease (AD), yet the cellular and molecular mechanisms by which APOE4 contributes to AD risk remain largely unknownMethodWe performed single‐nuclear RNAsequencing from the prefrontal cortex (Brodmann’s Area 10) of thirty‐two individuals, 12 APOE3/3 and 12 APOE3/4 carriers each, and one subgroup with 8 APOE4/4 carriers, and validated hypotheses using mouse models and patient‐derived iPSC lines.ResultDifferential expression and pathway analysis revealed that APOE4 is strongly associated with dysregulation of cholesterol homeostasis across multiple cell types, with oligodendrocytes being the most impacted. Strikingly, in human and mouse brains, APOE4 oligodendrocytes contained aberrant intracellular accumulations of cholesterol, whereas in non‐carriers, cholesterol preferentially localized to the oligodendrocyte cell membrane along myelinated neuronal tracts. Mass‐spectrometry‐based lipidomic analysis further revealed that APOE4 oligodendrocytes and APOE4 human post‐mortem brain tissue had elevated levels of cholesteryl esters, associated with intracellular storage of cholesterol as lipid droplets. Cholesterol transport and bioavailability are rate‐limiting steps in axonal myelination. Consistent with this, we discovered both APOE4 human and mouse brains were hypomyelinated relative to non‐carriers. We found that a small molecule known to facilitate cholesterol transport reduced cholesterol accumulation in APOE4 oligodendrocytes, increased axonal myelination, and improved memory in APOE4 mice.ConclusionThese results provide a transcriptional atlas of cell‐type specific APOE4 effects in the human brain. Our results demonstrate that cholesterol homeostasis in oligodendrocytes is highly dysregulated in APOE4 oligodendrocytes and identify hypomyelination as a key APOE4‐associated phenotype. Collectively, these data establish a functional link between APOE4, cholesterol homeostasis, and myelination that presents new therapeutic opportunities for Alzheimer’s disease.

Single-cell RNA sequencing unveils Lrg1's role in cerebral ischemia‒reperfusion injury by modulating various cells

Background and purposeCerebral ischemia‒reperfusion injury causes significant harm to human health and is a major contributor to stroke-related deaths worldwide. Current treatments are limited, and new, more effective prevention and treatment strategies that target multiple cell components are urgently needed. Leucine-rich alpha-2 glycoprotein 1 (Lrg1) appears to be associated with the progression of cerebral ischemia‒reperfusion injury, but the exact mechanism of it is unknown.MethodsWild-type (WT) and Lrg1 knockout (Lrg1−/−) mice were used to investigate the role of Lrg1 after cerebral ischemia‒reperfusion injury. The effects of Lrg1 knockout on brain infarct volume, blood‒brain barrier permeability, and neurological score (based on 2,3,5-triphenyl tetrazolium chloride, evans blue dye, hematoxylin, and eosin staining) were assessed. Single-cell RNA sequencing (scRNA-seq), immunofluorescence, and microvascular albumin leakage tests were utilized to investigate alterations in various cell components in brain tissue after Lrg1 knockout.ResultsLrg1 expression was increased in various cell types of brain tissue after cerebral ischemia‒reperfusion injury. Lrg1 knockout reduced cerebral edema and infarct size and improved neurological function after cerebral ischemia‒reperfusion injury. Single-cell RNA sequencing analysis of WT and Lrg1−/− mouse brain tissues after cerebral ischemia‒reperfusion injury revealed that Lrg1 knockout enhances blood‒brain barrier (BBB) by upregulating claudin 11, integrin β5, protocadherin 9, and annexin A2. Lrg1 knockout also promoted an anti-inflammatory and tissue-repairing phenotype in microglia and macrophages while reducing neuron and oligodendrocyte cell death.ConclusionsOur results has shown that Lrg1 mediates numerous pathological processes involved in cerebral ischemia‒reperfusion injury by altering the functional states of various cell types, thereby rendering it a promising therapeutic target for cerebral ischemia‒reperfusion injury.

Compartmentalized ciliation changes of oligodendrocytes in aged mouse optic nerve.

Primary cilia are microtubule-based sensory organelles that project from the apical surface of most mammalian cells, including oligodendrocytes, which are myelinating cells of the central nervous system (CNS) that support critical axonal function. Dysfunction of CNS glia is associated with aging-related white matter diseases and neurodegeneration, and ciliopathies are known to affect CNS white matter. To investigate age-related changes in ciliary profile, we examined ciliary length and frequency in the retinogeniculate pathway, a white matter tract commonly affected by diseases of aging but in which expression of cilia has not been characterized. We found expression of Arl13b, a marker of primary cilia, in a small group of Olig2-positive oligodendrocytes in the optic nerve, optic chiasm, and optic tract in young and aged C57BL/6 wild-type mice. While the ciliary length and ciliated oligodendrocyte cells were constant in young mice in the retinogeniculate pathway, there was a significant increase in ciliary length in the anterior optic nerve as compared to the aged animals. Morphometric analysis confirmed a specific increase in the ciliation rate of CC1+ /Olig2+ oligodendrocytes in aged mice compared with young mice. Thus, the prevalence of primary cilia in oligodendrocytes in the visual pathway and the age-related changes in ciliation suggest that they may play important roles in white matter and age-associated optic neuropathies.

Antioxidant, Anti-Inflammatory and Pro-Differentiative Effects of Chlorogenic Acid on M03-13 Human Oligodendrocyte-like Cells

Chlorogenic acid (CGA), a polyphenol found mainly in coffee and tea, exerts antioxidant, anti-inflammatory and anti-apoptotic effects at the gastrointestinal level. However, although CGA is known to cross the blood–brain barrier (BBB), its effects on the CNS are still unknown. Oligodendrocytes (OLs), the myelin-forming cells in the CNS, are the main target in demyelinating neuroinflammatory diseases such as multiple sclerosis (MS). We evaluated the antioxidant, anti-inflammatory and anti-apoptotic roles of CGA in M03-13, an immortalized human OL cell line. We found that CGA reduces intracellular superoxide ions, mitochondrial reactive oxygen species (ROS) and NADPH oxidases (NOXs) /dual oxidase 2 (DUOX2) protein levels. The stimulation of M03-13 cells with TNFα activates the nuclear factor kappa-light-chain-enhancer of activated B cell (NF-kB) pathway, leading to an increase in superoxide ion, NOXs/DUOX2 and phosphorylated extracellular regulated protein kinase (pERK) levels. In addition, tumor necrosis factor alpha (TNF-α) stimulation induces caspase 8 activation and the cleavage of poly-ADP-ribose polymerase (PARP). All these TNFα-induced effects are reversed by CGA. Furthermore, CGA induces a blockade of proliferation, driving cells to differentiation, resulting in increased mRNA levels of myelin basic protein (MBP) and proteolipid protein (PLP), which are major markers of mature OLs. Overall, these data suggest that dietary supplementation with this polyphenol could play an important beneficial role in autoimmune neuroinflammatory diseases such as MS.

Unraveling the transcriptomic signatures of Parkinson's disease and major depression using single-cell and bulk data.

Motor symptoms are well-characterized in Parkinson's disease (PD). However, non-motor symptoms, such as depression, are commonly observed and can appear up to 10 years before motor features, resulting in one-third of individuals being misdiagnosed with a neuropsychiatric disorder. Thus, identifying diagnostic biomarkers is crucial for accurate PD diagnosis during its prodromal or early stages. We employed an integrative approach, combining single nucleus RNA and bulk mRNA transcriptomics to perform comparative molecular signatures analysis between PD and major depressive disorder (MDD). We examined 39,834 nuclei from PD (GSE202210) and 32,707 nuclei from MDD (GSE144136) in the dorsolateral prefrontal cortex (dlPFC) of Brodmann area 9. Additionally, we analyzed bulk mRNA peripheral blood samples from PD compared to controls (GSE49126, GSE72267), as well as MDD compared to controls (GSE39653). Our findings show a higher proportion of astrocytes, and oligodendrocyte cells in the dlPFC of individuals with PD vs. MDD. The excitatory to inhibitory neurons (E/I) ratio analysis indicates that MDD has a ratio close to normal 80/20, while PD has a ratio of 62/38, indicating increased inhibition in the dlPFC. Microglia displayed the most pronounced differences in gene expression profiles between the two conditions. In PD, microglia display a pro-inflammatory phenotype, while in MDD, they regulate synaptic transmission through oligodendrocyte-microglia crosstalk. Analysis of bulk mRNA blood samples revealed that the COL5A, MID1, ZNF148, and CD22 genes were highly expressed in PD, whereas the DENR and RNU1G2 genes were highly expressed in MDD. CD22 is involved in B-cell activation and the negative regulation of B-cell receptor signaling. Additionally, CD86, which provides co-stimulatory signals for T-cell activation and survival, was found to be a commonly differentially expressed gene in both conditions. Pathway analysis revealed several immune-related pathways common in both conditions, including the complement and coagulation cascade, and B-cell receptor signaling. This study demonstrates that bulk peripheral immune cells play a role in both conditions, but neuroinflammation in the dlPFC specifically manifests in PD as evidenced by the analysis of single nucleus dlPFC datasets. Integrating these two omics levels offers a better understanding of the shared and distinct molecular pathophysiology of PD and MDD in both the periphery and the brain. These findings could lead to potential diagnostic biomarkers, improving accuracy and guiding pharmacological treatments.

Pathogenic myelin-specific antibodies in multiple sclerosis target conformational proteolipid protein 1-anchored membrane domains.

B cell clonal expansion and cerebrospinal fluid (CSF) oligoclonal IgG bands are established features of the immune response in multiple sclerosis (MS). Clone-specific recombinant monoclonal IgG1 Abs (rAbs) derived from MS patient CSF plasmablasts bound to conformational proteolipid protein 1 (PLP1) membrane complexes and, when injected into mouse brain with human complement, recapitulated histologic features of MS pathology: oligodendrocyte cell loss, complement deposition, and CD68+ phagocyte infiltration. Conformational PLP1 membrane epitopes were complex and governed by the local cholesterol and glycolipid microenvironment. Abs against conformational PLP1 membrane complexes targeted multiple surface epitopes, were enriched within the CSF compartment, and were detected in most MS patients, but not in inflammatory and noninflammatory neurologic controls. CSF PLP1 complex Abs provide a pathogenic autoantibody biomarker specific for MS.

Attenuation of Immunogenicity in MOG-Induced Oligodendrocytes by the Probiotic Bacterium Lactococcus Sp. PO3.

Background and Objectives: Milk is healthy and includes several vital nutrients and microbiomes. Probiotics in milk and their derivatives modulate the immune system, fight inflammation, and protect against numerous diseases. The present study aimed to isolate novel bacterial species with probiotic potential for neuroinflammation. Materials and Methods: Six milk samples were collected from lactating dairy cows. Bacterial isolates were obtained using standard methods and were evaluated based on probiotic characteristics such as the catalase test, hemolysis, acid/bile tolerance, cell adhesion, and hydrophobicity, as well as in vitro screening. Results: Nine morphologically diverse bacterial isolates were found in six different types of cow's milk. Among the isolates, PO3 displayed probiotic characteristics. PO3 was a Gram-positive rod cell that grew in an acidic (pH-2) salty medium containing bile salt and salinity (8% NaCl). PO3 also exhibited substantial hydrophobicity and cell adhesion. The sequencing comparison of the 16S rRNA genes revealed that PO3 was Lactococcus raffinolactis with a similarity score of 99.3%. Furthermore, PO3 was assessed for its neuroanti-inflammatory activity on human oligodendrocyte (HOG) cell lines using four different neuroimmune markers: signal transducer and activator of transcription (STAT-3), myelin basic protein (MBP), glial fibrillary acidic protein (GFAP), and GLAC in HOG cell lines induced by MOG. Unlike the rest of the evaluated neuroimmune markers, STAT-3 levels were elevated in the MOG-treated HOG cell lines compared to the untreated ones. The expression level of STAT-3 was attenuated in both PO3-MOG-treated and only PO3-treated cell lines. On the contrary, in PO3-treated cell lines, MBP, GFAP, and GLAC were significantly expressed at higher levels when compared with the MOG-treated cell lines. Conclusions: The findings reported in this article are to be used as a foundation for further in vivo research in order to pave the way for the possible use of probiotics in the treatment of neuroinflammatory diseases, including multiple sclerosis.

Does DNA methylation in the fetal brain leave an epigenetic memory in the blood?

Epigenetic memory is an emerging concept that refers to the process in which epigenetic changes occurring early-in life can lead to long-term programs of gene regulation in time and space. By leveraging neural network regression modeling of DNA methylation data in pigs, we show that specific methylations in the adult blood can reliably predict methylation changes that occurred in the fetal brain. Genes associated with these methylations represented known markers of specific cell types of blood including bone marrow hematopoietic progenitor cells, and ependymal and oligodendrocyte cells of brain. This suggested that methylation changes that occurred in the developing brain were maintained as an epigenetic memory in the blood through the adult life.

Differential expression of the circadian clock network correlates with tumour progression in gliomas

BackgroundGliomas are tumours arising mostly from astrocytic or oligodendrocytic precursor cells. These tumours are classified according to the updated WHO classification from 2021 in 4 grades depending on molecular and histopathological criteria. Despite novel multimodal therapeutic approaches, the vast majority of gliomas (WHO grade III and IV) are not curable. The circadian clock is an important regulator of numerous cellular processes and its dysregulation had been found during the progression of many cancers, including gliomas.ResultsIn this study, we explore expression patterns of clock-controlled genes in low-grade glioma (LGG) and glioblastoma multiforme (GBM) and show that a set of 45 clock-controlled genes can be used to distinguish GBM from normal tissue. Subsequent analysis identified 17 clock-controlled genes with a significant association with survival. The results point to a loss of correlation strength within elements of the circadian clock network in GBM compared to LGG. We further explored the progression patterns of mutations in LGG and GBM, and showed that tumour suppressor APC is lost late both in LGG and GBM. Moreover, HIF1A, involved in cellular response to hypoxia, exhibits subclonal losses in LGG, and TERT, involved in the formation of telomerase, is lost late in the GBM progression. By examining multi-sample LGG data, we find that the clock-controlled driver genes APC, HIF1A, TERT and TP53 experience frequent subclonal gains and losses.ConclusionsOur results show a higher level of disrgulation at the gene expression level in GBM compared to LGG, and indicate an association between the differentially expressed clock-regulated genes and patient survival in both LGG and GBM. By reconstructing the patterns of progression in LGG and GBM, our data reveals the relatively late gains and losses of clock-regulated glioma drivers. Our analysis emphasizes the role of clock-regulated genes in glioma development and progression. Yet, further research is needed to asses their value in the development of new treatments.

ATRT-06. ETMR TUMORIGENESIS MIRRORS RADIAL GLIAL DEVELOPMENT

Abstract Embryonal tumors with multilayered rosettes (ETMRs) are rare pediatric brain tumors defined primarily by overexpression of the C19MC microRNA cluster. These tumors affect mostly children under the age of 3. However, little is known about the origin and developmental contexts of ETMRs. We hypothesized that the early onset of these tumors suggests an embryonic tumorigenic event. To elucidate the embryonic contexts of which ETMRs arise, we curated three mouse embryogenesis datasets representing the full spectrum of murine embryonic development. We generated timepoint structure signatures of this atlas and projected them onto a single nuclei RNA sequencing dataset of 8 primary ETMRs. Consistent with existing literature, we discovered that ETMRs resemble a spectrum of radial glial, neuronal, and oligodendrocytic cell types. We also recapitulated potential ETMR driver mechanisms such as overexpression of WNT and SHH signaling by gene set enrichment analysis. We validated the enrichment of lineage and driver markers through GSEA and marker gene enrichment in a separate cohort of ETMRs characterized by bulk RNA sequencing. Taken together, the embryonic context of ETMR development may play crucial roles in the early identification and treatments of this lethal disease.

Comprehensive analyses of brain cell communications based on multiple scRNA-seq and snRNA-seq datasets for revealing novel mechanism in neurodegenerative diseases.

Complex cellular communications between glial cells and neurons are critical for brain normal function and disorders, and single-cell level RNA-sequencing datasets display more advantages for analyzing cell communications. Therefore, it is necessary to systematically explore brain cell communications when considering factors such as sex and brain region. We extracted a total of 1,039,459 cells derived from 28 brain single-cell RNA-sequencing (scRNA-seq) or single-nucleus RNA-sequencing (snRNA-seq) datasets from the GEO database, including 12 human and 16 mouse datasets. These datasets were further divided into 71 new sub-datasets when considering disease, sex, and region conditions. In the meanwhile, we integrated four methods to evaluate ligand-receptor interaction score among six major brain cell types (microglia, neuron, astrocyte, oligodendrocyte, OPC, and endothelial cell). For Alzheimer's disease (AD), disease-specific ligand-receptor pairs when compared with normal sub-datasets, such as SEMA4A-NRP1, were identified. Furthermore, we explored the sex- and region-specific cell communications and identified that WNT5A-ROR1 among microglia cells displayed close communications in male, and SPP1-ITGAV displayed close communications in the meninges region from microglia to neurons. Furthermore, based on the AD-specific cell communications, we constructed a model for AD early prediction and confirmed the predictive performance using multiple independent datasets. Finally, we developed an online platform for researchers to explore brain condition-specific cell communications. This research provided a comprehensive study to explore brain cell communications, which could reveal novel biological mechanisms involved in normal brain function and neurodegenerative diseases such as AD.

Effects of acute low-moderate dose ionizing radiation to human brain organoids.

Human exposure to low-to-moderate dose ionizing radiation (LMD-IR) is increasing via environmental, medical, occupational sources. Acute exposure to LMD-IR can cause subclinical damage to cells, resulting in altered gene expression and cellular function within the human brain. It has been difficult to identify diagnostic and predictive biomarkers of exposure using traditional research models due to factors including lack of 3D structure in monolayer cell cultures, limited ability of animal models to accurately predict human responses, and technical limitations of studying functional human brain tissue. To address this gap, we generated brain/cerebral organoids from human induced pluripotent stem cells to study the radiosensitivity of human brain cells, including neurons, astrocytes, and oligodendrocytes. While organoids have become popular models for studying brain physiology and pathology, there is little evidence to confirm that exposing brain organoids to LMD-IR will recapitulate previous in vitro and in vivo observations. We hypothesized that exposing brain organoids to proton radiation would (1) cause a time- and dose-dependent increase in DNA damage, (2) induce cell type-specific differences in radiosensitivity, and (3) increase expression of oxidative stress and DNA damage response genes. Organoids were exposed to 0.5 or 2 Gy of 250 MeV protons and samples were collected at 30 minute, 24 hour, and 48 hour timepoints. Using immunofluorescence and RNA sequencing, we found time- and dose-dependent increases in DNA damage in irradiated organoids; no changes in cell populations for neurons, oligodendrocytes, and astrocytes by 24 hours; decreased expression of genes related to oligodendrocyte lineage, astrocyte lineage, mitochondrial function, and cell cycle progression by 48 hours; increased expression of genes related to neuron lineage, oxidative stress, and DNA damage checkpoint regulation by 48 hours. Our findings demonstrate the possibility of using organoids to characterize cell-specific radiosensitivity and early radiation-induced gene expression changes within the human brain, providing new avenues for further study of the mechanisms underlying acute neural cell responses to IR exposure at low-to-moderate doses.

Myelin Oligodendrocyte Glycoprotein (MOG) Associated Diseases: Updates in Pediatric Practice

Myelin oligodendrocyte glycoprotein (MOG) is a membrane bound protein found on the surface of oligodendrocyte cells and the outermost surface of myelin sheaths. MOG is posited to play a role as a cell surface receptor or cell adhesion molecule, though there is no definitive answer to its exact function at this time. In the last few decades, there has been a recognition of anti-MOG-antibodies (MOG-Abs) in association with a variety of neurologic conditions, though primarily demyelinating and white matter disorders. In addition, MOG associated disease (MOGAD) appears to have a predilection for pediatric populations and in some patients may have a relapsing course. There has been considerable debate as to whether MOG-Abs are truly directly pathogenic or a disease biomarker associated with neuorinflammatory disease. In this manuscript we will review the current literature surrounding MOGAD, review new clinical phenotypes, discuss treatment and prognosis, and provide insight into potential future directions that studies may focus on.

Free fatty acids support oligodendrocyte survival in a mouse model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the white matter degeneration. Although changes in blood lipids are involved in the pathogenesis of neurological diseases, the pathological role of blood lipids in ALS remains unclear. We performed lipidome analysis on the plasma of ALS model mice, mutant superoxide dismutase 1 (SOD1G93A) mice, and found that the concentration of free fatty acids (FFAs), including oleic acid (OA) and linoleic acid (LA), decreased prior to disease onset. An in vitro study revealed that OA and LA directly inhibited glutamate-induced oligodendrocytes cell death via free fatty acid receptor 1 (FFAR1). A cocktail containing OA/LA suppressed oligodendrocyte cell death in the spinal cord of SOD1G93A mice. These results suggested that the reduction of FFAs in the plasma is a pathogenic biomarker for ALS in the early stages, and supplying a deficiency in FFAs is a potential therapeutic approach for ALS by preventing oligodendrocyte cell death.

Abstract 338: Novel Role Of Endothelial Cd47 In The Regulation Of Pathogenesis Of Atherosclerosis

Background: The anti-phagocytic molecule CD47 is upregulated in atherosclerotic plaques of patients with cerebrovascular disease. However, the role of endothelial cell-specific CD47 in the pathogenesis of atherosclerosis has not yet been defined. Kojima et.al . described atheroprotective effect of anti-CD47 antibody by enhancing efferocytosis in atherosclerosis. In this study, we determined whether endothelialCD47 regulates efferocytosis in atherosclerosis. Methods: Susceptibility to atherosclerosis was evaluated in cross sections of the aortic sinus in atherosclerotic mice. Using single-cell RNA sequencing and efferocytosis assays, we investigated the role of CD47 in regulating efferocytosis in atherosclerotic mouse model. Results: Endothelial cell-specific deletion of CD47 led to significant plaque reduction in aortic sinus of athero-CD47iECKO mice. Cluster analysis and gene expression profiles of 20,000 aortic endothelial cells from atherosclerotic mice identified 10 major cell types: endothelial cells, VSMCs, fibroblasts, B cells, T cells, macrophage, erythrocytes, oligodendrocytes cells, Schwann cells, and pericytes. In athero-CD47iECKO group, subpopulation of endothelial cells as well as fibroblasts, T-cells, B-cells, macrophages, erythrocytes, oligodendrocytes, and pericytes were increased, while vascular smooth muscle cell numbers decreased. Using efferocytosis assay on isolated primary endothelial cells from control and CD47iECKO mice, we observed CD47 deletion induced engulfment of apoptotic Jurkat cells, which had been exposed to NF-κB inhibitor, BAY11-7082 to stimulate apoptotic attributes. Quantitative PCR analyses showed that CD47 deletion increases the expression of efferocytosis receptor FasL, CX3CL1, Gpr132, MerTK, Scarb1, Sirpa, TSP1, and HMGB1. Conclusion: Our study indicates that genetic deletion of CD47 in endothelial cells attenuates atheroma progression by enhancing efferocytosis. This study also reveals phenotypic heterogenicity of endothelial cells at the single-cell level during atherogenesis and provides deeper understanding of endothelial cell biology in atherosclerosis that may facilitate the development of novel therapeutic interventions for cardiovascular disease.

Cover Image, Volume 140, Issue 20

The cover image by Senay Ustunel features a co-culturing system wherein neuroblastoma (SH-SY5Y) and oligodendrocyte (MO3.13) cell lines grow within a 3D liquid crystal elastomer (LCE) scaffold. LCE scaffolds can effectively impact cellular function, differentiation, and myelination of co-cultured cell lines, allowing the mimicry of a more endogenous environment and providing support for long term studies to manipulate cellular crosstalk, like in this case to study myelination. DOI: 10.1002/app.53883

Neuroprotective Effect of Wharton's Jelly-Derived Mesenchymal Stem Cell-Conditioned Medium (WJMSC-CM) on Diabetes-Associated Cognitive Impairment by Improving Oxidative Stress, Neuroinflammation, and Apoptosis.

According to strong evidence, diabetes mellitus increases the risk of cognitive impairment. Mesenchymal stem cells have been shown to be potential therapeutic agents for neurological disorders. In the current study, we aimed to examine the effects of Wharton's jelly-derived mesenchymal stem cell-conditioned medium (WJMSC-CM) on learning and memory, oxidative stress, apoptosis, and histological changes in the hippocampus of diabetic rats. Randomly, 35 male Sprague Dawley rats weighing 260-300 g were allocated into five groups: control, diabetes, and three diabetic groups treated with insulin, WJMSC-CM, and DMEM. The injections of insulin (3 U/day, S.C.) and WJMSC-CM (10 mg/week, I.P.) were done for 60 days. The Morris water maze and open field were used to measure cognition and anxiety-like behaviors. Colorimetric assays were used to determine hippocampus glutathione (GSH), malondialdehyde (MDA) levels, and antioxidant enzyme activity. The histopathological evaluation of the hippocampus was performed by Nissl staining. The expression levels of Bax, Bcl-2, BDNF, and TNF-α were detected by real-time polymerase chain reaction (RT-PCR). According to our findings, WJMSC-CM significantly reduced and increased blood glucose and insulin levels, respectively. Enhanced cognition and improved anxiety-like behavior were also found in WJMSC-CM-treated diabetic rats. In addition, WJMSC-CM treatment reduced oxidative stress by lowering MDA and elevating GSH and antioxidant enzyme activity. Reduced TNF-α and enhanced Bcl-2 gene expression levels and elevated neuronal and nonneuronal (astrocytes and oligodendrocytes) cells were detected in the hippocampus of WJMSC-CM-treated diabetic rats. In conclusion, WJMSC-CM alleviated diabetes-related cognitive impairment by reducing oxidative stress, neuroinflammation, and apoptosis in diabetic rats.

BMPRII+ neural precursor cells isolated and characterized from organotypic neurospheres: an in vitro model of human fetal spinal cord development.

Roof plate secretion of bone morphogenetic proteins (BMPs) directs the cellular fate of sensory neurons during spinal cord development, including the formation of the ascending sensory columns, though their biology is not well understood. Type-II BMP receptor (BMPRII), the cognate receptor, is expressed by neural precursor cells during embryogenesis; however, an in vitro method of enriching BMPRII+ human neural precursor cells (hNPCs) from the fetal spinal cord is absent. Immunofluorescence was undertaken on intact second-trimester human fetal spinal cord using antibodies to BMPRII and leukemia inhibitory factor (LIF). Regions of highest BMPRII+ immunofluorescence localized to sensory columns. Parenchymal and meningeal-associated BMPRII+ vascular cells were identified in both intact fetal spinal cord and cortex by co-positivity with vascular lineage markers, CD34/CD39. LIF immunostaining identified a population of somas concentrated in dorsal and ventral horn interneurons, mirroring the expression of LIF receptor/CD118. A combination of LIF supplementation and high-density culture maintained culture growth beyond 10 passages, while synergistically increasing the proportion of neurospheres with a stratified, cytoarchitecture. These neurospheres were characterized by BMPRII+/MAP2ab+/-/βIII-tubulin+/nestin-/vimentin-/GFAP-/NeuN- surface hNPCs surrounding a heterogeneous core of βIII-tubulin+/nestin+/vimentin+/GFAP+/MAP2ab-/NeuN- multipotent precursors. Dissociated cultures from tripotential neurospheres contained neuronal (βIII-tubulin+), astrocytic (GFAP+), and oligodendrocytic (O4+) lineage cells. Fluorescence-activated cell sorting-sorted BMPRII+ hNPCs were MAP2ab+/-/βIII-tubulin+/GFAP-/O4- in culture. This is the first isolation of BMPRII+ hNPCs identified and characterized in human fetal spinal cords. Our data show that LIF combines synergistically with high-density reaggregate cultures to support the organotypic reorganization of neurospheres, characterized by surface BMPRII+ hNPCs. Our study has provided a new methodology for an in vitro model capable of amplifying human fetal spinal cord cell numbers for > 10 passages. Investigations of the role BMPRII plays in spinal cord development have primarily relied upon mouse and rat models, with interpolations to human development being derived through inference. Because of significant species differences between murine biology and human, including anatomical dissimilarities in central nervous system (CNS) structure, the findings made in murine models cannot be presumed to apply to human spinal cord development. For these reasons, our human in vitro model offers a novel tool to better understand neurodevelopmental pathways, including BMP signaling, as well as spinal cord injury research and testing drug therapies.