Cell Types Research Articles

Reactive Astrocytes Release GDNF to Promote Brain Recovery and Neuronal Survival Following Ischemic Stroke

Astrocytes are important glia cell type in the central nervous system. These cells can undergo transformation to a reactive state upon injury such as focal ischemic stroke (FIS). Reactive astrocytes are distinct from normal or homeostatic astrocytes in morphology, protein profiles and metabolic functions. Glial cell-derived neurotrophic factor (GDNF) was discovered as a potent survival neurotrophic factor for multiple subtypes of neurons and can be released from reactive astrocytes. In our previous study, we found that GDNF expression was upregulated in reactive astrocytes following ischemic stroke. Specific knock out of GDNF in reactive astrocytes exacerbated brain damage and motor deficits after ischemic stroke. Here, using in vitro and in vivo ischemia models, we investigated the effects of GDNF overexpression in astrocytes on neuronal survival and brain recovery after ischemia. We observed that astrocyte specific GDNF overexpression by viral transduction could decrease brain infarction and promote motor function recovery after photothrombosis (PT)-induced FIS. In addition, GDNF overexpression in astrocytes could increase the proliferation of reactive astrocytes and reduce oxidative stress after PT. Using the oxygen-glucose deprivation (OGD) model of cultured astrocytes, we confirmed that this ischemic insult could upregulate GDNF expression and increase its release to extracellular space. Transfection of GDNF DNA plasmid could further increase GDNF release after OGD. To further study the effects of reactive astrocytes-derived extracellular GDNF on neuronal survival after ischemia, cultured neurons subjected to OGD were exposed to astrocyte conditioned medium (ACM). The ACM collected from OGD subjected astrocyte culture could significantly reduce neuronal death, while neutralizing antibodies against GDNF and its receptors including GFRα1, RET and p-RET could suppress this beneficial effect. We also found that reactive astrocytes-derived GDNF could trigger the activation of RET receptors in cultured neurons and suppress neuronal mitochondrial fission and caspase-dependent cell apoptosis after OGD. Overall, our results indicate that reactive astrocytes-derived GDNF could play an important role in neuronal survival and functional recovery and underscore the non-cell autonomy underlying astrocyte-neuron interactions in brain repair after ischemic stroke.

Macrophage-derived SPP1 exacerbate myocardial injury by interacting with fibroblasts in viral myocarditis

BackgroundViral myocarditis (VMC) is an inflammatory myocardial condition triggered by viral infections which involves pathogenic-related damage and immune-mediated damage. However, the precise immunopathogenic mechanisms underlying VMC remain elusive.MethodsWe performed single-cell RNA sequencing on mouse hearts during the acute phase of CVB3-induced VMC. After manually annotating cell types, functional analyses of macrophage were performed by cell ratio changes, customized gene set module scoring and CellPhoneDB. Utilizing indirect co-culture experiments in vitro, the effects of macrophage-derived SPP1 on cardiac fibroblasts were investigated. Depletion of macrophages and inhibition of SPP1 expression in mice were carried out to study the effects of macrophage-derived SPP1 on cardiac function, inflammation levels, and myocardial injury in mice with VMC.ResultsOur data revealed that macrophages are the major immune cells which infiltrate the heart during the acute phase of VMC, particularly a macrophage subpopulation which highly expresses Spp1 (Spp1+ macrophages) and exhibited characteristics of peripheral blood monocytes. Spp1+ macrophages communicate extensively with fibroblasts during VMC, and that SPP1 promotes fibroblast conversion to an inflammatory phenotype with high Ccl2/Ccl7 expression. This in turn increases monocyte chemotaxis to the heart. Besides, a partial depletion of macrophages in the early stages of VMC attenuated myocardial inflammation and myocardial injury in mice. Inhibition of SPP1 reduced cardiac macrophage infiltration, attenuated myocardial inflammation, and improved cardiac function in VMC mice.ConclusionOur findings suggested that Spp1+ macrophages could self-recruit, and macrophage-derived SPP1 exacerbated myocardial immune injury by promoting high Ccl2/Ccl7 expression in fibroblasts. Our study advances understandings of VMC pathogenesis, and provides novel insight into potential immunotherapies for VMC.

Distinct landscapes of fibroblast subtypes in arteries of patients with giant cell arteritis.

Giant cell arteritis (GCA) is a systemic vasculitis of large- and medium-sized arteries characterized by granulomatous inflammation and vascular remodelling. Although fibroblasts are the predominant cell type in the adventitia, their role in GCA pathogenesis is largely unknown. This study aimed to investigate the distribution of fibroblast subtypes in relation to vascular remodeling in GCA. Temporal artery biopsies (TAB) from patients with GCA(n = 9) and controls(n = 15) and aorta tissues from GCA(n = 9)- and atherosclerosis(n = 11)-related aneurysms were examined. Immunohistochemical and immunofluorescence stainings for fibroblast subtype markers (CD90, PDGFRA, FAP, podoplanin, CD248, α-SMA), cellular proliferation (Ki67) and remodelling-related growth factors (TGF-β, FGF21, PDGFB) were performed to evaluate the distribution of fibroblast subtypes in relation to active remodelling pathways. To evaluate the role of FAP in TGF-β-induced fibroblast proliferation, human aortic adventitial fibroblasts (HAoAF) were stimulated in vitro with TGF-β and transfected with small interference RNA targeting FAP. In GCA-TAB, CD90+FAP+activated fibroblasts and CD90+podoplanin+immunofibroblasts were predominantly located in the adventitia. CD90+α-SMA+myofibroblasts were observed mainly in the intima, and CD90+CD248+fibroblasts in the adventitia-media border and intima. High FGF21 and PDGFB expression in the intima was associated with intimal hyperplasia in GCA-positive TAB. GCA-affected aortas showed a different landscape of fibroblast subtypes: CD90+FAP+activated fibroblasts, CD90+podoplanin+immunofibroblasts and CD90+CD248+fibroblasts accumulated especially in structurally disrupted media. ∼80% of proliferative fibroblasts in TAB and aorta were FAP positive. FAP knockdown suppressed TGFβ-induced proliferation of HAoAF in vitro. This study documents a distinct spatial distribution pattern of fibroblast subtypes in GCA-affected arteries. The data suggest different roles for fibroblasts in remodelling of specific arterial vascular beds in GCA.

Dual role of FOXG1 in regulating gliogenesis in the developing neocortex via the FGF signalling pathway.

In the developing vertebrate central nervous system, neurons and glia typically arise sequentially from common progenitors. Here, we report that the transcription factor Forkhead Box G1 (Foxg1) regulates gliogenesis in the mouse neocortex via distinct cell-autonomous roles in progenitors and postmitotic neurons that regulate different aspects of the gliogenic FGF signalling pathway. We demonstrate that loss of Foxg1 in cortical progenitors at neurogenic stages causes premature astrogliogenesis. We identify a novel FOXG1 target, the pro-gliogenic FGF pathway component Fgfr3, which is suppressed by FOXG1 cell-autonomously to maintain neurogenesis. Furthermore, FOXG1 can also suppress premature astrogliogenesis triggered by the augmentation of FGF signalling. We identify a second novel function of FOXG1 in regulating the expression of gliogenic cues in newborn neocortical upper-layer neurons. Loss of FOXG1 in postmitotic neurons non-autonomously enhances gliogenesis in the progenitors via FGF signalling. These results fit well with the model that newborn neurons secrete cues that trigger progenitors to produce the next wave of cell types, astrocytes. If FGF signalling is attenuated in Foxg1 null progenitors, they progress to oligodendrocyte production. Therefore, loss of FOXG1 transitions the progenitor to a gliogenic state, producing either astrocytes or oligodendrocytes depending on FGF signalling levels. Our results uncover how FOXG1 integrates extrinsic signalling via the FGF pathway to regulate the sequential generation of neurons, astrocytes, and oligodendrocytes in the cerebral cortex.

A Mathematical Exploration of SDH-b Loss in Chromaffin Cells

The succinate dehydrogenase (SDH) is a four-subunit enzyme complex (SDH-a, SDH-b, SDH-c, and SDH-d) central to cell carbon metabolism. The SDH bridges the tricarboxylic acid cycle to the electron transport chain. A pathological loss of the SDH-b subunit leads to a cell-wide signalling cascade that shifts the cell’s metabolism into a pseudo-hypoxic state akin to the so-called Warburg effect (or aerobic glycolysis). This trait is a hallmark of phaeochromocytomas, a rare tumour arising from chromaffin cells; a type of cell that lies in the medulla of the adrenal gland. In this study, we leverage the insights from a mathematical model constructed to underpin the metabolic implications of SDH-b dysfunction in phaeochromocytomas. We specifically investigate why chromaffin cells seemingly have the ability to maintain electron transport chain’s Complex I function when confronted with the loss of the SDH-b subunit while other cells do not. Our simulations indicate that retention of Complex I is associated with cofactor oxidation, which enables cells to manage mitochondrial swelling and limit the reversal of the adenosine triphosphate synthase, supporting cell fitness, without undergoing lysis. These results support previous hypotheses that point to mitochondrial proton leaks as a critical factor of future research. Moreover, the model asserts that control of the proton gradient across the mitochondrial inner membrane is rate-limiting upon fitness management of SDH-b deficient cells.

Interleukin-10 resistance in type 2 diabetes is associated with defective STAT3 signaling in human blood leukocytes.

Chronic inflammation is strongly implicated in the pathophysiology of type 2 diabetes (T2D), highlighting the need to better understand inflammatory processes in people living with T2D. Hyperglycemia blunts the anti-inflammatory actions of interleukin-10 (IL-10) - the most potent anti-inflammatory cytokine - but the mechanistic basis remains unclear. To test the hypothesis that signaling defects underpin this hyporesponsiveness to IL-10 action, fasted blood samples were obtained from individuals living with T2D (n=17, age: 64±9 yrs, HbA1c: 7.2±1.1%) and their age-matched counterparts without diabetes (n=19, 65±8 yrs, 5.5±0.3%). Blood leukocytes were analyzed for IL-10-mediated signaling, gene expression, and cytokine secretion using flow cytometry, qPCR, and whole-blood cultures, respectively. Despite no overt elevations in circulating pro- and anti-inflammatory cytokine concentrations, blood leukocytes from individuals with T2D exhibited exaggerated cytokine secretion when exposed to lipopolysaccharide (LPS) (p<0.05). IL-10's ability to activate its canonical transcription factor signal transducer and activator of transcription 3 (STAT3) was blunted in CD14 monocytes and CD4 lymphocytes from people with T2D (p<0.01) - a defect associated with lower IL-10 receptor expression on both cell types (p<0.05). This upstream signaling defect was accompanied by attenuated suppressor of cytokine signaling 3 (SOCS3) mRNA levels in IL-10 treated mononuclear cells (p=0.059) and higher lipopolysaccharide (LPS)-stimulated cytokine secretion from blood leukocytes exposed to IL-10 (p<0.01). Our findings identify defective IL-10-mediated signaling and gene expression as a potential mechanism underpinning IL-10 resistance in T2D, highlighting need for further investigation into therapeutic approaches targeting IL-10.

Simultaneous isolation and culture of endothelial colony-forming cells, endothelial cells and vascular smooth muscle cells from human umbilical cords

BackgroundRegulation of the human umbilical circulation under physiological and pathological conditions remains poorly understood. We previously demonstrated that intrauterine growth restriction (IUGR) is associated with sex-specific alterations in the human umbilical circulation. Our data strongly suggest a differential contribution of subcellular compartmentation depending on fetal sex, vessel type and the presence of IUGR. We therefore developed a protocol to isolate and culture umbilical vascular cells to further investigate the relative contribution of each cell type and subcellular compartmentation to the human umbilical circulation regulation.Methods and resultsHuman umbilical cords and cord blood were collected just after delivery. Mononuclear cells were recovered from cord blood using a Ficoll gradient and cultured to obtain endothelial colony-forming cells (ECFCs). Endothelial cells (ECs) were isolated from human umbilical vein (HUV) and arteries (HUAs) by collagenase/dispase digestion, and vascular smooth muscle cells (SMCs) by migration from vascular explants. All cell types were characterized by visualization, and by analysis of biomarkers using immunocytofluorescence and Western blot. ECFCs were also submitted to polychromatic flow cytometry analysis.ConclusionsThis protocol enables simultaneous isolation and culture of ECFCs, HUVECs, HUAECs, HUVSMCs and HUASMCs from the same umbilical cord. It is simpler, faster and more cost-effective than other previously published methods, with good success rates. This will be helpful to further investigate the regulatory mechanisms implicated in the human umbilical circulation under physiological and pathological conditions and to study the influence of fetal sex.

Nucleus pulposus cell-mimicking nanoparticles for cell-specific HIF1A editing to modulate SASP-mediated disc inflammation via autophagy activation.

Nucleus pulposus cell-mimicking nanoparticles for cell-specific HIF1A editing to modulate SASP-mediated disc inflammation via autophagy activation.

V-domain Ig Suppressor of T cell Activation Expression During Hemorrhage or Sepsis-Induced Acute Respiratory Distress Syndrome: Insights From a Mouse Model.

V-domain Ig Suppressor of T cell Activation Expression During Hemorrhage or Sepsis-Induced Acute Respiratory Distress Syndrome: Insights From a Mouse Model.

Nuclear Argonaute protein NRDE-3 switches small RNA partners during embryogenesis to mediate temporal-specific gene regulatory activity.

RNA interference (RNAi) is a conserved pathway that utilizes Argonaute proteins and their associated small RNAs to exert gene regulatory function on complementary transcripts. While the majority of germline-expressed RNAi proteins reside in perinuclear germ granules, it is unknown whether and how RNAi pathways are spatially organized in other cell types. Here, we find that the small RNA biogenesis machinery is spatially and temporally organized during Caenorhabditis elegans embryogenesis. Specifically, the RNAi factor, SIMR-1, forms visible concentrates during mid-embryogenesis that contain an RNA-dependent RNA polymerase, a poly-UG polymerase, and the unloaded nuclear Argonaute protein, NRDE-3. Curiously, coincident with the appearance of the SIMR granules, the small RNAs bound to NRDE-3 switch from predominantly CSR-class 22G-RNAs to ERGO-dependent 22G-RNAs. NRDE-3 binds ERGO-dependent 22G-RNAs in the somatic cells of larvae and adults to silence ERGO-target genes; here we further demonstrate that NRDE-3-bound, CSR-class 22G-RNAs repress transcription in oocytes. Thus, our study defines two separable roles for NRDE-3, targeting germline-expressed genes during oogenesis to promote global transcriptional repression, and switching during embryogenesis to repress recently duplicated genes and retrotransposons in somatic cells, highlighting the plasticity of Argonaute proteins and the need for more precise temporal characterization of Argonaute-small RNA interactions.

Developing a gene expression classifier for breast cancer diagnosis.

Breast cancer (BC) is the most common type of cancer in women worldwide. Solid tumors are complex structures composed of many cell types and extracellular matrix components. Understanding solid tumors is crucial for developing effective treatments. This study aimed to develop a gene expression classifier to predict BC with high accuracy. The study first identified the most important genes for cancer through differential expression analysis (DEA) between breast cancer and adjacent normal breast samples. The R package STRINGdb was then used to create a protein-protein interaction network (PPI) to examine upregulated genes and find clusters. Enrichment analyses were performed to identify overrepresented biological functions and pathways. A logistic regression prediction model was developed using a breast cancer dataset from TCGA and evaluated using discrimination and calibration measures. BUB1 expression in breast cancer was also investigated using quantitative analysis. Two significant clusters were identified, with cell cycle checkpoints and M phase key pathways in one cluster and extracellular matrix organization in the other. A prediction model using the hub gene set (COMP, FN1, SDC1, BUB1, TTK, and NUSAP1) showed high sensitivity (97.2%) and specificity (96.1%), and an AUC of 0.994. Three hub genes (COMP, FN1, and SDC1) were identified through the PPI network, strongly linked to extracellular matrix organization (BUB1, TTK, and NUSAP1) as hub genes involved in M phase and cell cycle checkpoints. Overall, the study identified hub pathways and genes that accurately distinguish between cancer and normal samples, presenting promising new possibilities for early cancer detection and improved BC therapy.

Human BioMolecular Atlas Program (HuBMAP): 3D Human Reference Atlas construction and usage.

The Human BioMolecular Atlas Program (HuBMAP) aims to construct a 3D Human Reference Atlas (HRA) of the healthy adult body. Experts from 20+ consortia collaborate to develop a Common Coordinate Framework (CCF), knowledge graphs and tools that describe the multiscale structure of the human body (from organs and tissues down to cells, genes and biomarkers) and to use the HRA to characterize changes that occur with aging, disease and other perturbations. HRA v.2.0 covers 4,499 unique anatomical structures, 1,195 cell types and 2,089 biomarkers (such as genes, proteins and lipids) from 33 ASCT+B tables and 65 3D Reference Objects linked to ontologies. New experimental data can be mapped into the HRA using (1) cell type annotation tools (for example, Azimuth), (2) validated antibody panels or (3) by registering tissue data spatially. This paper describes HRA user stories, terminology, data formats, ontology validation, unified analysis workflows, user interfaces, instructional materials, application programming interfaces, flexible hybrid cloud infrastructure and previews atlas usage applications.

Integration of single-nuclei transcriptome and bulk RNA-seq to unravel the role of AhWRKY70 in regulating stem cell development in Arachis hypogaea L.

Peanut stem is a vital organ to provide mechanical support and energy for aerial tissue development. However, the transcriptional regulatory mechanisms underlying stem development at a single-cell resolution remain unclear. Herein, single-nuclei isolation coupled with fluorescent-activated cell sorting was employed to construct a cell atlas of peanut seedling stems using microdroplets-based single-nuclei RNA-sequencing. This approach yielded 29 308 cells with 53 349 expressed genes underlying the identification of five cell types characterized by known marker genes. Additionally, 2053 differentially expressed genes (DEGs) were identified across different cell types. Furthermore, 3306 core-DEGs involved in cell development trajectories were used to construct a transcription factor (TF) interaction network, providing insights into specific biological pathways and transcriptional regulation dynamics underlying cell-type differentiation. Additionally, 1446 DEGs associated with different cell-cycle profile were identified, revealing that peanut stem elongation and cell expansion are closely linked to auxin-responsive pathway. This was supported by the examination of endogenous phytohormones and the identification of 10 hormone-responsive DEGs. Moreover, AhWRKY70 was localized in the nucleus and is highly enriched in stem cortex and xylem cells and exhibits a tissue-specific expression pattern that regulates stem growth. Overexpression of AhWRKY70 in Arabidopsis led to accelerated stem growth by modulating the phytohormone signalling pathway, influencing the expression of sixteen auxin and ethylene-responsive genes as demonstrated by transcriptome sequencing. In conclusion, the single-cell atlas provides a foundational dataset for understanding gene expression heterogeneity in peanut seedling stems. The elucidation of AhWRKY70 function expands our understanding of the roles of WRKY family members in peanut.

Isolation of bovine milk-derived extracellular vesicles via a capillary-channeled polymer (C-CP) fiber stationary phase.

Extracellular vesicles (EVs) are released by all cell types into the extracellular environment. A subset of EVs, known as exosomes, range in size from 30 to 200nm and are of biochemical interest due to their function as vehicles of intercellular communication. Their ability to transport proteinaceous species and genetic material at the cellular level makes them prime candidates as vectors in gene therapies. Focusing on biotherapeutics, bovine milk-derived extracellular vesicles (MDEVs) hold particular promise as an alternative to other exosome sources for therapeutics delivery. Bovine milk poses unique challenges due to the complex colloidal matrix, composed predominantly of fats and proteins like casein, which form micelles that exhibit exosome-like characteristics, specifically size and density. When faced with complex matrices like milk, conventional size/density-based isolation methods including ultracentrifugation and size exclusion chromatography struggle to provide high purity yields on practical time and cost scales. When paired with a stepwise hydrophobic interaction chromatography (HIC) gradient, polyester (PET) capillary-channeled polymer (C-CP) fibers in column and spin-down tips formats have been used effectively to isolate exosomes from highly diverse sources. Here, PET C-CP fiber columns are demonstrated to isolate MDEVs from pre-treated raw milk, yielding concentrations of 1.5 × 1010 particles mL⁻1 with purities of ~2 × 1010 EVs µg-1 protein in less than 20min. The efficacy of the isolation process is verified by a suite of characterization methods. Implementing PET C-CP fiber columns for MDEV isolation addresses the challenges associated with conventional isolation methods, holding promise for scale-up towards therapeutic applications.

A skin organoid-based infection platform identifies an inhibitor specific for HFMD

The EV-A71 poses a serious threat to the health and lives of children. The EV-A71 can be transmitted by direct and indirect skin contact. Therefore, there is an urgent need to create novel skin models using human-derived cells to study the biology and pathogenesis of the virus and facilitate drug screening. Here, we use human induced pluripotent stem cells-derived skin organoids (hiPSC-SOs) as a model for EV-A71 infection and find that multiple cell types within the skin organoids, including epidermal cells, hair follicle cells, fibroblasts, and nerve cells, express EV-A71 receptors and are susceptible to EV-A71 infection. We elucidate the specific response of different cell types to EV-A71 and reveal that EV-A71 infection can degrade extracellular collagen and affect fibroblasts. We find that EV-A71 can mediate epidermal cell damage through autophagy and Integrin/Hippo-YAP/TAZ signaling pathways, thereby promoting hyperproliferation of progenitor cells. Based on this finding, we identify an autophagy-associated protein as a drug target of EV-A71 and discover an EV-A71 replication inhibitor. Altogether, these data suggest that hiPSC-SOs can be used as an infectious disease model to study skin infectious diseases, providing a valuable resource for drug screening to identify candidate virus therapeutics.

Enhanced Performance of Dye-Sensitized Solar Cells by Lanthanum Doped-TiO2 Photoelectrodes

Dye-sensitized solar cells (DSSCs) are a type of thin-film solar cell in which metal oxide nanomaterials are used as photoanodes. With the aim to improving the efficiency of DSSC, pure and La3+ doped-TiO2 nanoparticles were synthesized by sol-gel method and the quality of synthesized materials was investigated through various characterizations like structural, elemental, morphological and optical studies. The XRD pattern revealed the presence of anatase-TiO2 phase. The FESEM results evidenced the spherical nanoparticles with less agglomeration. The UV-Vis spectra showed enhanced light absorption with the addition of La. With the addition of La, the band gap energy of the TiO2 nanoparticles reduced from 3.06 to 2.72 eV. These nanoparticles were integrated as photoanodes in DSSC and the photocurrent density-voltage (J-V) measurements were carried out. The highest energy conversion efficiency (η) of 4.75 for pure TiO2 device was achieved while 5.8% was obtained for the 5% La-doped TiO2 device. This enhancement of η confirmed that 5% La-doped TiO2 is highly suitable for DSSC device fabrication. It was also observed that the high efficiency (∼5.8%) with the highest current density of 11.11 mA/cm2 - was retained even under low-light conditions.

Development of NIR photocleavable nanoparticles with BDNF for vestibular neuron regeneration

Among nanoparticle platforms, light or photoresponsive nanoparticles have emerged as a promising drug delivery strategy with spatiotemporal control while minimizing off-target effects. The characteristic absorption spectrum of the photoresponsive moiety dictates the wavelength of light needed to activate bond cleavage. However, the low tissue penetration depth limit and short-wavelength ultraviolet (UV) cellular toxicity are considered disadvantageous. This study developed a vestibular ganglion neuron organoid as a model for vestibulopathy. UV and near-infrared (NIR) radiation targeted the inner ear and neural cells, followed by toxicity evaluation. A significantly smaller toxicity of NIR light was confirmed. The photocleavage release of brain-derived neurotrophic factor (BDNF) was used by applying NIR wavelength. The results indicate that polyethylene glycol octamethylene diamine derivative conjugated with leucomethylene blue with an ethanolamine linker nanoparticle can be effectively disassembled and release BDNF when using the 808 nm laser as a trigger. The findings of the cytotoxicity assay suggest that photocleavable nanoparticles (PCNs) and laser irradiation are safe and biocompatible for human-derived and neural progenitor types of cells. Phototriggered BDNF release by NIR laser supported the growth and differentiation of human neural progenitor cells in culture. In addition, the vestibulopathy organoid exhibited a significant regenerative effect. This study harnesses the full potential of NIR laser PCNs to treat vestibular neuropathies.Graphical abstract

Temporal Profiling of Male Cortical Astrocyte Transcription Predicts Molecular Shifts From Early Development to Aging.

Astrocytes are the most abundant glial cell type in the central nervous system (CNS). Astrocytes are born during the early postnatal period in the rodent brain and mature alongside neurons, demonstrating remarkable morphological structural complexity, which is attained in the second postnatal month. Throughout this period of development and across the remainder of the lifespan, astrocytes participate in CNS homeostasis, support neuronal partners, and contribute to nearly all aspects of CNS function. In the present study, we analyzed astrocyte gene expression in the cortex of wild-type male rodents throughout their lifespan (postnatal 7 days to 18 months). A pairwise timepoint comparison of differential gene expression during early development and CNS maturation (7-60 days) revealed four unique astrocyte gene clusters, each with hundreds of genes, which demonstrate unique temporal profiles. These clusters are distinctively related to cell division, cell morphology, cellular communication, and vascular structure and regulation. A similar analysis across adulthood and in the aging brain (3 to 18 months) identified similar patterns of grouped gene expression related to cell metabolism and cell structure. Additionally, our analysis identified that during the aging process astrocytes demonstrate a bias toward shorter transcripts, with loss of longer genes related to synapse development and a significant increase in shorter transcripts related to immune regulation and the response to DNA damage. Our study highlights the critical role that astrocytes play in maintaining CNS function throughout life and reveals molecular shifts that occur during development and aging in the cortex of male mice.

Human CSPG4-targeting CAR-macrophages inhibit melanoma growth.

Approximately half of melanoma patients relapse or fail to respond to current standards of care, highlighting the need for new treatment options. Engineering T-cells with chimeric antigen receptors (CARs) has revolutionized the treatment of hematological malignancies but has been clinically less effective in solid tumors. We therefore sought to engineer alternative immune cell types to inhibit melanoma progression. Engineering macrophages with CARs has emerged as a promising approach to overcome some of the challenges faced by CAR-T cells; however, whether these engineered macrophages can effectively inhibit melanoma growth is unknown. To determine whether CAR-macrophages (CAR-Ms) specifically target and kill melanoma cells, we engineered CAR-Ms targeting chondroitin sulfate proteoglycan 4 (CSPG4), an antigen expressed in melanoma. CSPG4-targeting CAR-Ms exhibited specific phagocytosis of CSPG4-expressing melanoma cells. We developed 3D approaches to show that CSPG4-targeting CAR-Ms efficiently infiltrated melanoma spheroids. Furthermore, combining CSPG4-targeting CAR-Ms with strategies inhibiting CD47/SIRPα "don't eat me" signaling synergistically enhanced CAR-M-mediated phagocytosis and robustly inhibited melanoma spheroid growth in 3D. Importantly, CSPG4-targeting CAR-Ms inhibited melanoma tumor growth in mouse models. These results suggest engineering macrophages against melanoma antigens is a promising solid tumor immunotherapy approach for treating melanoma.

Identification and validation of biomarkers related to ferroptosis in idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a kind of interstitial lung disease (ILD). It has a high incidence rate and mortality. Its pathogenesis remains unclear. So far, no effective methods have been found for the early diagnosis of IPF. Ferroptosis has been reported to be critical in the initiation and progression of IPF. Therefore, our aim was to identify the hub gene related to ferroptosis co-expressed in the peripheral blood and pulmonary tissue of patients with IPF. Sequencing data were obtained from the Gene Expression Omnibus database. A comprehensive analysis was conducted on the differentially expressed genes (DEGs) to extract ferroptosis-related differentially expressed genes (FRDEGs). The results showed that ferroptosis-related signal paths were highly enriched in IPF, and 10 FRDEGs were identified.The hub gene was predicted through protein-protein interactions (PPI) and Cytoscape. The diagnostic utility of the hub gene was proven by enzyme-linked immunosorbent assay (ELISA) in serum and by immunohistochemistry (IHC) in pulmonary tissues. The results of ELISA indicated that the levels of ATM in the serum of patients with IPF were significantly lower than the normal levels. In contrast, the results of IHC showed that the expression of ATM in the pulmonary tissues of IPF patients exhibited a notably elevated trend. The immune status was assessed by the CIBERSORT method and so was the relevance between ATM and immune cells. These findings unveiled significant differences in various immune cell types in peripheral blood and pulmonary tissue between the IPF group and the control group. Furthermore, ATM was associated with various immune cells. This study suggests that as a ferroptosis-related gene, ATM assumes a pivotal role in the diagnosis and treatment of IPF. This discovery presents a novel approach for the clinical diagnosis and therapy of IPF.