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Science Advances Nov 2023infection is a major risk factor for the development of gastric cancer. The bacteria reside in close proximity to gastric surface mucous as well as stem and progenitor...
infection is a major risk factor for the development of gastric cancer. The bacteria reside in close proximity to gastric surface mucous as well as stem and progenitor cells. Here, we take advantage of wild-type and genetically engineered murine gastric organoids and organoid-derived monolayers to study the cellular targets of -induced DNA damage and replication stress and to explore possible interactions with preexisting gastric cancer driver mutations. We find using alkaline comet assay, single-molecule DNA fiber assays, and immunofluorescence microscopy of DNA repair foci that induces transcription-dependent DNA damage in actively replicating, Leucine-rich-repeat containing G-Protein-Coupled Receptor 5 (Lgr5)-positive antral stem and progenitor cells and their Troy-positive corpus counterparts, but not in other gastric epithelial lineages. Infection-dependent DNA damage is aggravated by inactivation, but not by or loss, or overexpression. Our data suggest that induces DNA damage in stem and progenitor cells, especially in settings of hyperproliferation due to constitutively active Wnt signaling.
Topics: Animals; Humans; Mice; DNA Damage; Genes, Tumor Suppressor; Helicobacter Infections; Helicobacter pylori; Receptors, G-Protein-Coupled; Stem Cells; Stomach Neoplasms
PubMed: 37967175
DOI: 10.1126/sciadv.adh0322 -
Cell Host & Microbe Feb 2024T cells are critical in mediating the early control of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) breakthrough infection. However, it remains unknown...
T cells are critical in mediating the early control of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) breakthrough infection. However, it remains unknown whether memory T cells can effectively cross-recognize new SARS-CoV-2 variants with a broad array of mutations, such as the emergent hypermutated BA.2.86 variant. Here, we report in two separate cohorts, including healthy controls and individuals with chronic lymphocytic leukemia, that SARS-CoV-2 spike-specific CD4 and CD8 T cells induced by prior infection or vaccination demonstrate resilient immune recognition of BA.2.86. In both cohorts, we found largely preserved SARS-CoV-2 spike-specific CD4 and CD8 T cell magnitudes against mutated spike epitopes of BA.2.86. Functional analysis confirmed that both cytokine expression and proliferative capacity of SARS-CoV-2 spike-specific T cells to BA.2.86-mutated spike epitopes are similarly sustained. In summary, our findings indicate that memory CD4 and CD8 T cells continue to provide cell-mediated immune recognition to highly mutated emerging variants such as BA.2.86.
Topics: Humans; Memory T Cells; CD8-Positive T-Lymphocytes; COVID-19; SARS-CoV-2; Epitopes; Spike Glycoprotein, Coronavirus; Antibodies, Viral
PubMed: 38211584
DOI: 10.1016/j.chom.2023.12.010 -
Neural Regeneration Research Jun 2024Inflammation plays a crucial role in the regeneration of fish and avian retinas. However, how inflammation regulates Müller glia (MG) reprogramming remains unclear....
Inflammation plays a crucial role in the regeneration of fish and avian retinas. However, how inflammation regulates Müller glia (MG) reprogramming remains unclear. Here, we used single-cell RNA sequencing to investigate the cell heterogeneity and interactions of MG and immune cells in the regenerating zebrafish retina. We first showed that two types of quiescent MG (resting MG1 and MG2) reside in the uninjured retina. Following retinal injury, resting MG1 transitioned into an activated state expressing known reprogramming genes, while resting MG2 gave rise to rod progenitors. We further showed that retinal microglia can be categorized into three subtypes (microglia-1, microglia-2, and proliferative) and pseudotime analysis demonstrated dynamic changes in microglial status following retinal injury. Analysis of cell-cell interactions indicated extensive crosstalk between immune cells and MG, with many interactions shared among different immune cell types. Finally, we showed that inflammation activated Jak1-Stat3 signaling in MG, promoting their transition from a resting to an activated state. Our study reveals the cell heterogeneity and crosstalk of immune cells and MG in zebrafish retinal repair, and may provide valuable insights into future mammalian retina regeneration.
PubMed: 38934409
DOI: 10.4103/NRR.NRR-D-23-02083 -
Nature Communications Dec 2023Although patients with rheumatoid arthritis (RA) typically exhibit symmetrical joint involvement, some patients develop alternative disease patterns in response to...
Although patients with rheumatoid arthritis (RA) typically exhibit symmetrical joint involvement, some patients develop alternative disease patterns in response to treatment, suggesting that different molecular mechanism may underlie disease progression depending on joint location. Here, we identify joint-specific changes in RA synovium and synovial fibroblasts (SF) between knee and hand joints. We show that the long non-coding RNA HOTAIR, which is only expressed in knee SF, regulates more than 50% of this site-specific gene expression in SF. HOTAIR is downregulated after stimulation with pro-inflammatory cytokines and is expressed at lower levels in knee samples from patients with RA, compared with osteoarthritis. Knockdown of HOTAIR in knee SF increases PI-Akt signalling and IL-6 production, but reduces Wnt signalling. Silencing HOTAIR inhibits the migratory function of SF, decreases SF-mediated osteoclastogenesis, and increases the recruitment of B cells by SF. We propose that HOTAIR is an important epigenetic factor in joint-specific gene expression in RA.
Topics: Humans; Arthritis, Rheumatoid; Fibroblasts; Gene Expression; Osteoarthritis; RNA, Long Noncoding; Synovial Fluid; Synovial Membrane
PubMed: 38071204
DOI: 10.1038/s41467-023-44053-w -
Redox Biology Nov 2023Excessive light exposure can damage photoreceptors and lead to blindness. Oxidative stress serves a key role in photo-induced retinal damage. Free radical scavengers...
Excessive light exposure can damage photoreceptors and lead to blindness. Oxidative stress serves a key role in photo-induced retinal damage. Free radical scavengers have been proven to protect against photo-damaged retinal degeneration. Fullerol, a potent antioxidant, has the potential to protect against ultraviolet-B (UVB)-induced cornea injury by activating the endogenous stem cells. However, its effects on cell fate determination of Müller glia (MG) between gliosis and de-differentiation remain unclear. Therefore, we established a MG lineage-tracing mouse model of light-induced retinal damage to examine the therapeutic effects of fullerol. Fullerol exhibited superior protection against light-induced retinal injury compared to glutathione (GSH) and reduced oxidative stress levels, inhibited gliosis by suppressing the TGF-β pathway, and enhanced the de-differentiation of MG cells. RNA sequencing revealed that transcription candidate pathways, including Nrf2 and Wnt10a pathways, were involved in fullerol-induced neuroprotection. Fullerol-mediated transcriptional changes were validated by qPCR, Western blotting, and immunostaining using mouse retinas and human-derived Müller cell lines MIO-M1 cells, confirming that fullerol possibly modulated the Nrf2, Wnt10a, and TGF-β pathways in MG, which suppressed gliosis and promoted the de-differentiation of MG in light-induced retinal degeneration, indicating its potential in treating retinal diseases.
Topics: Animals; Mice; Humans; Ependymoglial Cells; Retinal Degeneration; Gliosis; NF-E2-Related Factor 2; Retina; Neuroglia; Transforming Growth Factor beta
PubMed: 37816275
DOI: 10.1016/j.redox.2023.102911 -
Aging and Disease May 2024Age-related macular degeneration (AMD) is a progressive neurodegeneration disease that causes photoreceptor demise and vision impairments. In AMD pathogenesis, the... (Review)
Review
Age-related macular degeneration (AMD) is a progressive neurodegeneration disease that causes photoreceptor demise and vision impairments. In AMD pathogenesis, the primary death of retinal neurons always leads to the activation of resident microglia. The migration of activated microglia to the ongoing retinal lesion and their morphological transformation from branching to ameboid-like are recognized as hallmarks of AMD pathogenesis. Activated microglia send signals to Müller cells and promote them to react correspondingly to damaging stimulus. Müller cells are a type of neuroglia cells that maintain the normal function of retinal neurons, modulating innate inflammatory responses, and stabilize retinal structure. Activated Müller cells can accelerate the progression of AMD by damaging neurons and blood vessels. Therefore, the crosstalk between microglia and Müller cells plays a homeostatic role in maintaining the retinal environment, and this interaction is complicatedly modulated. In particular, the mechanism of mutual regulation between the two glia populations is complex under pathological conditions. This paper reviews recent findings on the crosstalk between microglia and Müller glia during AMD pathology process, with special emphasis on its therapeutic potentials.
Topics: Humans; Macular Degeneration; Ependymoglial Cells; Microglia; Neuroinflammatory Diseases; Animals
PubMed: 37728589
DOI: 10.14336/AD.2023.0823-3 -
Neural Regeneration Research Dec 2023Cynops orientalis (C. orientalis) has a pronounced ability to regenerate its spinal cord after injury. Thus, exploring the molecular mechanism of this process could...
Cynops orientalis (C. orientalis) has a pronounced ability to regenerate its spinal cord after injury. Thus, exploring the molecular mechanism of this process could provide new approaches for promoting mammalian spinal cord regeneration. In this study, we established a model of spinal cord thoracic transection injury in C. orientalis, which is an endemic species in China. We performed RNA sequencing of the contused axolotl spinal cord at two early time points after spinal cord injury - during the very acute stage (4 days) and the subacute stage (7 days) - and identified differentially expressed genes; additionally, we performed Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses, at each time point. Transcriptome sequencing showed that 13,059 genes were differentially expressed during C. orientalis spinal cord regeneration compared with uninjured animals, among which 4273 were continuously down-regulated and 1564 were continuously up-regulated. Down-regulated genes were most enriched in the Gene Ontology term "multicellular organismal process" and in the ribosome pathway at 10 days following spinal cord injury. We found that multiple genes associated with energy metabolism were down-regulated and multiple genes associated with the lysosome were up-regulated after spinal cord injury, indicating the importance of low metabolic activity during wound healing. Immune response-associated pathways were activated during the early acute phase (4 days), while the expression of extracellular matrix proteins such as glycosaminoglycan and collagen, as well as tight junction proteins, was lower at 10 days post-spinal cord injury than 4 days post-spinal cord injury. However, compared with 4 days post-injury, at 10 days post-injury neuroactive ligand-receptor interactions were no longer down-regulated, up-regulated differentially expressed genes were enriched in pathways associated with cancer and the cell cycle, and SHH, VIM, and Sox2 were prominently up-regulated. Immunofluorescence staining showed that glial fibrillary acidic protein was up-regulated in axolotl ependymoglial cells after injury, similar to what is observed in mammalian astrocytes after spinal cord injury, even though axolotls do not form a glial scar during regeneration. We suggest that low intracellular energy production could slow the rapid amplification of ependymoglial cells, thereby inhibiting reactive gliosis, at early stages after spinal cord injury. Extracellular matrix degradation slows cellular responses, represses the expression of neurogenic genes, and reactivates a transcriptional program similar to that of embryonic neuroepithelial cells. These ependymoglial cells act as neural stem cells: they migrate and proliferate to repair the lesion and then differentiate to replace lost glial cells and neurons. This provides the regenerative microenvironment that allows axon growth after injury.
PubMed: 37449639
DOI: 10.4103/1673-5374.373717 -
Acta Biomaterialia Sep 2023Papillary and reticular dermis show distinct extracellular matrix (ECM) and vascularization corresponding to their specific functions. These characteristics are...
Papillary and reticular dermis show distinct extracellular matrix (ECM) and vascularization corresponding to their specific functions. These characteristics are associated with gene expression patterns of fibroblasts freshly isolated from their native microenvironment. In order to assess the relevance of these fibroblast subpopulations in a tissue engineering context, we investigated their contribution to matrix production and vascularization using cell sheet culture conditions. We first performed RNA-seq differential expression analysis to determine whether several rounds of cell amplification and high-density culture affected their gene expression profile. Bioinformatics analysis revealed that expression of angiogenesis-related and matrisome gene signatures were maintained, resulting in papillary and reticular ECMs that differ in composition and structure. The impact of secreted or ECM-associated factors was then assessed using two independent 3D angiogenesis assays: -1/ a fibrin hydrogel-based assay allowing investigation of diffusible secreted factors, -2/ a scaffold-free cell-sheet based assay for investigation of fibroblast-produced microenvironment. These analyses revealed that papillary fibroblasts secrete highly angiogenic factors and produce a microenvironment characterised by ECM remodelling capacity and dense and branched microvascular network, whereas reticular fibroblasts produced more structural core components of the ECM associated with less branched and larger vessels. These features mimick the characteristics of both the ECM and the vasculature of dermis subcompartments. In addition to showing that skin fibroblast populations differentially regulate angiogenesis via both secreted and ECM factors, our work emphasizes the importance of papillary and reticular fibroblasts for engineering and modelling dermis microenvironment and vascularization. STATEMENT OF SIGNIFICANCE: Recent advances have brought to the forefront the central role of microenvironment and vascularization in tissue engineering for regenerative medicine and microtissue modelling. We have investigated the role of papillary and reticular fibroblast subpopulations using scaffold-free cell sheet culture. This approach provides differentiated cells conditions allowing the production of their own microenvironment. Analysis of gene expression profiles and characterisation of the matrix produced revealed strong and specific angiogenic properties that we functionally characterized using 3D angiogenesis models targeting the respective role of either secreted or matrix-bound factors. This study demonstrates the importance of cell-generated extracellular matrix and questions the importance of cell source and the relevance of hydrogels for developing physio-pathologically relevant tissue engineered substitutes.
Topics: Humans; Dermis; Cell Culture Techniques; Tissue Engineering; Epidermis; Neovascularization, Pathologic; Fibroblasts; Extracellular Matrix
PubMed: 37406716
DOI: 10.1016/j.actbio.2023.06.040 -
Frontiers in Cellular Neuroscience 2023The possible applications for human retinal organoids (HROs) derived from human induced pluripotent stem cells (hiPSC) rely on the robustness and transferability of the...
The possible applications for human retinal organoids (HROs) derived from human induced pluripotent stem cells (hiPSC) rely on the robustness and transferability of the methodology for their generation. Standardized strategies and parameters to effectively assess, compare, and optimize organoid protocols are starting to be established, but are not yet complete. To advance this, we explored the efficiency and reliability of a differentiation method, called CYST protocol, that facilitates retina generation by forming neuroepithelial cysts from hiPSC clusters. Here, we tested seven different hiPSC lines which reproducibly generated HROs. Histological and ultrastructural analyses indicate that HRO differentiation and maturation are regulated. The different hiPSC lines appeared to be a larger source of variance than experimental rounds. Although previous reports have shown that HROs in several other protocols contain a rather low number of cones, HROs from the CYST protocol are consistently richer in cones and with a comparable ratio of cones, rods, and Müller glia. To provide further insight into HRO cell composition, we studied single cell RNA sequencing data and applied CaSTLe, a transfer learning approach. Additionally, we devised a potential strategy to systematically evaluate different organoid protocols side-by-side through parallel differentiation from the same hiPSC batches: In an explorative study, the CYST protocol was compared to a conceptually different protocol based on the formation of cell aggregates from single hiPSCs. Comparing four hiPSC lines showed that both protocols reproduced key characteristics of retinal epithelial structure and cell composition, but the CYST protocol provided a higher HRO yield. So far, our data suggest that CYST-derived HROs remained stable up to at least day 200, while single hiPSC-derived HROs showed spontaneous pathologic changes by day 200. Overall, our data provide insights into the efficiency, reproducibility, and stability of the CYST protocol for generating HROs, which will be useful for further optimizing organoid systems, as well as for basic and translational research applications.
PubMed: 37868194
DOI: 10.3389/fncel.2023.1166641 -
Frontiers in Molecular Neuroscience 2023Loss of neurons in the neural retina is a leading cause of vision loss. While humans do not possess the capacity for retinal regeneration, zebrafish can achieve this...
INTRODUCTION
Loss of neurons in the neural retina is a leading cause of vision loss. While humans do not possess the capacity for retinal regeneration, zebrafish can achieve this through activation of resident Müller glia. Remarkably, despite the presence of Müller glia in humans and other mammalian vertebrates, these cells lack an intrinsic ability to contribute to regeneration. Upon activation, zebrafish Müller glia can adopt a stem cell-like state, undergo proliferation and generate new neurons. However, the underlying molecular mechanisms of this activation subsequent retinal regeneration remains unclear.
METHODS/RESULTS
To address this, we performed single-cell RNA sequencing (scRNA-seq) and report remarkable heterogeneity in gene expression within quiescent Müller glia across distinct dorsal, central and ventral retina pools of such cells. Next, we utilized a genetically driven, chemically inducible nitroreductase approach to study Müller glia activation following selective ablation of three distinct photoreceptor subtypes: long wavelength sensitive cones, short wavelength sensitive cones, and rods. There, our data revealed that a region-specific bias in activation of Müller glia exists in the zebrafish retina, and this is independent of the distribution of the ablated cell type across retinal regions. Notably, gene ontology analysis revealed that injury-responsive dorsal and central Müller glia express genes related to dorsal/ventral pattern formation, growth factor activity, and regulation of developmental process. Through scRNA-seq analysis, we identify a shared genetic program underlying initial Müller glia activation and cell cycle entry, followed by differences that drive the fate of regenerating neurons. We observed an initial expression of AP-1 and injury-responsive transcription factors, followed by genes involved in Notch signaling, ribosome biogenesis and gliogenesis, and finally expression of cell cycle, chromatin remodeling and microtubule-associated genes.
DISCUSSION
Taken together, our findings document the regional specificity of gene expression within quiescent Müller glia and demonstrate unique Müller glia activation and regeneration features following neural ablation. These findings will improve our understanding of the molecular pathways relevant to neural regeneration in the retina.
PubMed: 37575968
DOI: 10.3389/fnmol.2023.1087136