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1.
Dev Cell ; 2024 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-38848717

RESUMO

The histone H3 lysine 9 methyltransferase SETDB1 controls transcriptional repression to direct stem cell fate. Here, we show that Setdb1 expression by adult muscle stem cells (MuSCs) is required for skeletal muscle regeneration. We find that SETDB1 represses the expression of endogenous retroviruses (ERVs) in MuSCs. ERV de-repression in Setdb1-null MuSCs prevents their amplification following exit from quiescence and promotes cell death. Multi-omics profiling shows that chromatin decompaction at ERV loci activates the DNA-sensing cGAS-STING pathway, entailing cytokine expression by Setdb1-null MuSCs. This is followed by aberrant infiltration of inflammatory cells, including pathological macrophages. The ensuing histiocytosis is accompanied by myofiber necrosis, which, in addition to progressive MuSCs depletion, completely abolishes tissue repair. In contrast, loss of Setdb1 in fibro-adipogenic progenitors (FAPs) does not impact immune cells. In conclusion, genome maintenance by SETDB1 in an adult somatic stem cell is necessary for both its regenerative potential and adequate reparative inflammation.

2.
Curr Top Dev Biol ; 158: 151-177, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38670704

RESUMO

The process of skeletal muscle regeneration involves a coordinated interplay of specific cellular and molecular interactions within the injury site. This review provides an overview of the cellular and molecular components in regenerating skeletal muscle, focusing on how these cells or molecules in the niche regulate muscle stem cell functions. Dysfunctions of muscle stem cell-to-niche cell communications during aging and disease will also be discussed. A better understanding of how niche cells coordinate with muscle stem cells for muscle repair will greatly aid the development of therapeutic strategies for treating muscle-related disorders.


Assuntos
Homeostase , Músculo Esquelético , Regeneração , Nicho de Células-Tronco , Regeneração/fisiologia , Humanos , Músculo Esquelético/fisiologia , Músculo Esquelético/citologia , Animais , Nicho de Células-Tronco/fisiologia , Células-Tronco/citologia , Células-Tronco/fisiologia , Células-Tronco/metabolismo
3.
Cell Stem Cell ; 31(1): 89-105.e6, 2024 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-38141612

RESUMO

Stem cells are known for their resilience and enhanced activity post-stress. The mammary gland undergoes frequent remodeling and is subjected to recurring stress during the estrus cycle, but it remains unclear how mammary stem cells (MaSCs) respond to the stress and contribute to regeneration. We discovered that cytotoxic stress-induced activation of CD11c+ ductal macrophages aids stem cell survival and prevents differentiation. These macrophages boost Procr+ MaSC activity through IL1ß-IL1R1-NF-κB signaling during the estrus cycle in an oscillating manner. Deleting IL1R1 in MaSCs results in stem cell loss and skewed luminal differentiation. Moreover, under cytotoxic stress from the chemotherapy agent paclitaxel, ductal macrophages secrete higher IL1ß levels, promoting MaSC survival and preventing differentiation. Inhibiting IL1R1 sensitizes MaSCs to paclitaxel. Our findings reveal a recurring inflammatory process that regulates regeneration, providing insights into stress-induced inflammation and its impact on stem cell survival, potentially affecting cancer therapy efficacy.


Assuntos
Glândulas Mamárias Animais , Células-Tronco , Feminino , Animais , Diferenciação Celular/fisiologia , Transdução de Sinais , Paclitaxel/farmacologia , Paclitaxel/metabolismo
4.
STAR Protoc ; 4(4): 102750, 2023 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-38041820

RESUMO

Studying skeletal muscle stem cells (MuSCs) quiescence is challenging as they quickly activate within hours of isolation from muscle. Here, we present a protocol to disassociate and characterize fixed peptides from quiescent MuSCs using trapped ion-mobility time-of-flight mass spectrometry (MS). We describe steps for mouse perfusion, fluorescence-activated cell sorting preparation and sorting, protein extraction, digestion, and liquid chromatography MS analysis. This protocol can be applied to other less-abundant somatic stem cell types using mouse lines with a reporter. For complete details on the use and execution of this protocol, please refer to Zeng et al. (2022, 2023).1,2.


Assuntos
Células-Tronco Adultas , Proteômica , Animais , Camundongos , Fibras Musculares Esqueléticas , Divisão Celular , Movimento Celular
5.
Dev Cell ; 58(15): 1383-1398.e6, 2023 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-37321216

RESUMO

Age-associated impairments in adult stem cell functions correlate with a decline in somatic tissue regeneration capacity. However, the mechanisms underlying the molecular regulation of adult stem cell aging remain elusive. Here, we provide a proteomic analysis of physiologically aged murine muscle stem cells (MuSCs), illustrating a pre-senescent proteomic signature. During aging, the mitochondrial proteome and activity are impaired in MuSCs. In addition, the inhibition of mitochondrial function results in cellular senescence. We identified an RNA-binding protein, CPEB4, downregulated in various aged tissues, which is required for MuSC functions. CPEB4 regulates the mitochondrial proteome and activity through mitochondrial translational control. MuSCs devoid of CPEB4 induced cellular senescence. Importantly, restoring CPEB4 expression rescued impaired mitochondrial metabolism, improved geriatric MuSC functions, and prevented cellular senescence in various human cell lines. Our findings provide the basis for the possibility that CPEB4 regulates mitochondrial metabolism to govern cellular senescence, with an implication of therapeutic intervention for age-related senescence.


Assuntos
Proteoma , Proteômica , Idoso , Animais , Humanos , Camundongos , Envelhecimento/fisiologia , Senescência Celular , Músculo Esquelético/fisiologia , Músculos , Proteínas de Ligação a RNA
6.
STAR Protoc ; 4(3): 102376, 2023 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-37352103

RESUMO

Chromatin accessibility is critical for cell identity. Conventional ATAC-seq can examine chromatin accessibility on freshly prepared muscle stem cells or satellite cells (SCs); however, isolating SCs in mice remains challenging. Here, we present a protocol to preserve the in vivo chromatin profile of SCs by applying paraformaldehyde (PFA) perfusion throughout the mouse before SC isolation. We describe steps for PFA perfusion and FACS sorting of SCs. We then detail library preparation for ATAC-seq. For complete details on the use and execution of this protocol, please refer to Dong et al.1.


Assuntos
Cromatina , Células Satélites de Músculo Esquelético , Animais , Camundongos , Cromatina/genética , Sequenciamento de Cromatina por Imunoprecipitação , Análise de Sequência de DNA/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Músculos
7.
Dev Cell ; 58(12): 1052-1070.e10, 2023 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-37105173

RESUMO

Organismal homeostasis and regeneration are predicated on committed stem cells that can reside for long periods in a mitotically dormant but reversible cell-cycle arrest state defined as quiescence. Premature escape from quiescence is detrimental, as it results in stem cell depletion, with consequent defective tissue homeostasis and regeneration. Here, we report that Polycomb Ezh1 confers quiescence to murine muscle stem cells (MuSCs) through a non-canonical function. In the absence of Ezh1, MuSCs spontaneously exit quiescence. Following repeated injuries, the MuSC pool is progressively depleted, resulting in failure to sustain proper muscle regeneration. Rather than regulating repressive histone H3K27 methylation, Ezh1 maintains gene expression of the Notch signaling pathway in MuSCs. Selective genetic reconstitution of the Notch signaling corrects stem cell number and re-establishes quiescence of Ezh1-/- MuSCs.


Assuntos
Transdução de Sinais , Células-Tronco , Camundongos , Animais , Divisão Celular , Pontos de Checagem do Ciclo Celular , Músculos
8.
Methods Mol Biol ; 2640: 369-395, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36995608

RESUMO

Skeletal muscle possesses a remarkable regenerative capacity, mainly relying on a population of undifferentiated and unipotent muscle progenitors, called muscle stem cells (MuSCs) or satellite cells, and their interplay with various cell types within the niche. Investigating the cellular composition of skeletal muscle tissues and the heterogeneity among various cell populations is crucial to the unbiased understanding of how cellular networks work in harmony at the population level in the context of skeletal muscle homeostasis, regeneration, aging, and diseases. As opposed to probing the average profile in a cell population, single-cell RNA-seq has unlocked access to the transcriptomic landscape characterization of individual cells in a highly parallel manner. This chapter describes the workflow for single-cell transcriptomic analysis of mononuclear cells in skeletal muscle by taking advantage of the droplet-based single-cell RNA-seq platform, Chromium Single Cell 3' solution from 10x Genomics®. Using this protocol, we can reveal insights into muscle-resident cell-type identities, which can be exploited to study the muscle stem cell niche further.


Assuntos
Células Satélites de Músculo Esquelético , Transcriptoma , Músculo Esquelético/metabolismo , Células-Tronco , Genômica , Análise de Célula Única
9.
Methods Mol Biol ; 2640: 445-452, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36995612

RESUMO

Uncovering the transcriptomic signatures of quiescent muscle stem cells elicits the regulatory networks on stem cell quiescence. However, the spatial clues of the transcripts are missing in the commonly used quantitative analysis such as qPCR and RNA-seq. Visualization of RNA transcripts using single-molecule in situ hybridization provides additional subcellular localization clues to understanding gene expression signatures. Here, we provide an optimized protocol of smFISH analysis on Fluorescence-Activated Cell Sorting isolated muscle stem cells to visualize low-abundance transcripts.


Assuntos
Perfilação da Expressão Gênica , Células-Tronco , Hibridização in Situ Fluorescente/métodos , RNA/genética , Músculos
10.
Cell Rep ; 41(8): 111694, 2022 11 22.
Artigo em Inglês | MEDLINE | ID: mdl-36417861

RESUMO

The establishment of a functional vasculature requires endothelial cells to enter quiescence during the completion of development, otherwise pathological overgrowth occurs. How such a transition is regulated remains unclear. Here, we uncover a role of Zeb1 in defining vascular quiescence entry. During quiescence acquisition, Zeb1 increases along with the progressive decline of endothelial progenitors' activities, with Zeb1 loss resulting in endothelial overgrowth and vascular deformities. RNA sequencing (RNA-seq) and assay for transposase-accessible chromatin sequencing (ATAC-seq) analyses reveal that Zeb1 represses Wif1, thereby activating Wnt/ß-catenin signaling. Knockdown of Wif1 rescues the overgrowth induced by Zeb1 deletion. Importantly, local administration of surrogate Wnt molecules in the retina ameliorates the overgrowth defects of Zeb1 mutants. These findings show a mechanism by which Zeb1 induces quiescence of endothelial progenitors during the establishing of vascular homeostasis, providing molecular insight into the inherited neovascular pathologies associated with human ZEB1 mutations, suggesting pharmacological activation of Wnt/ß-catenin signaling as a potential therapeutical approach.


Assuntos
Células Endoteliais , beta Catenina , Humanos , beta Catenina/metabolismo , Células Endoteliais/metabolismo , Via de Sinalização Wnt/genética , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética
11.
iScience ; 25(9): 104954, 2022 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-36093058

RESUMO

Regulation of chromatin accessibility is critical for cell fate decisions. Chromatin structure responds to extrinsic environments rapidly. The traditional adult stem cell isolation approach requires tissue dissociation, which triggers stem cell activation and leads to alterations in chromatin structure. To preserve the in vivo chromatin states, we utilized the PFA-perfusion-based isolation approach and characterized the DNA regulatory landscapes during muscle stem cell quiescence exit and aging. We showed that aged SCs display a chronically activated chromatin signature. Detailed analysis of the chromatin accessibility profiles identified key enhancer elements for SC quiescence. Constant activation of the enhancer elements promotes stemness and prevents SCs from differentiation, whereas genetic deletion causes cell-cycle arrest and leads to defects in activation. Our comprehensive characterization of the chromatin accessibility and transcriptomic landscapes in SC quiescence and aging broadens our understanding of these processes and identifies key distal regulatory elements for SC function.

12.
Nat Commun ; 13(1): 947, 2022 02 17.
Artigo em Inglês | MEDLINE | ID: mdl-35177647

RESUMO

Skeletal muscle stem cells, also called Satellite Cells (SCs), are actively maintained in quiescence but can activate quickly upon extrinsic stimuli. However, the mechanisms of how quiescent SCs (QSCs) activate swiftly remain elusive. Here, using a whole mouse perfusion fixation approach to obtain bona fide QSCs, we identify massive proteomic changes during the quiescence-to-activation transition in pathways such as chromatin maintenance, metabolism, transcription, and translation. Discordant correlation of transcriptomic and proteomic changes reveals potential translational regulation upon SC activation. Importantly, we show Cytoplasmic Polyadenylation Element Binding protein 1 (CPEB1), post-transcriptionally affects protein translation during SC activation by binding to the 3' UTRs of different transcripts. We demonstrate phosphorylation-dependent CPEB1 promoted Myod1 protein synthesis by binding to the cytoplasmic polyadenylation elements (CPEs) within its 3' UTRs to regulate SC activation and muscle regeneration. Our study characterizes CPEB1 as a key regulator to reprogram the translational landscape directing SC activation and subsequent proliferation.


Assuntos
Músculo Esquelético/lesões , Biossíntese de Proteínas/genética , Regeneração/genética , Células Satélites de Músculo Esquelético/fisiologia , Fatores de Transcrição/metabolismo , Fatores de Poliadenilação e Clivagem de mRNA/metabolismo , Regiões 3' não Traduzidas/genética , Animais , Linhagem Celular , Células Cultivadas , Modelos Animais de Doenças , Regulação da Expressão Gênica , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Músculo Esquelético/citologia , Proteína MyoD/biossíntese , Proteômica , RNA-Seq
13.
Curr Opin Cell Biol ; 73: 124-132, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34534837

RESUMO

The chromatin landscape represents a critical regulatory layer for precise transcriptional control. Chromosome architecture restrains the physical access to the DNA elements and is one of the determinants that specifies cell identity. Adult stem cells possess the unique ability to differentiate into a specific lineage. One of the underexplored areas in skeletal muscle biology is the molecular mechanism guiding the chromatin organization changes in muscle stem cell specification, myogenic determination, and differentiation. In this review, we focus on the regulatory network guiding the progression of muscle stem cells to differentiated progeny. We summarize recent findings regarding the mechanisms directing myogenic cell fate decision and differentiation, with a particular focus on three-dimensional chromosome architecture and long noncoding RNA-associated chromatin accessibility changes.


Assuntos
Cromatina , Desenvolvimento Muscular , Diferenciação Celular , Músculo Esquelético , Células-Tronco
14.
PLoS Genet ; 17(7): e1009635, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34237064

RESUMO

The intracellular level of fatty aldehydes is tightly regulated by aldehyde dehydrogenases to minimize the formation of toxic lipid and protein adducts. Importantly, the dysregulation of aldehyde dehydrogenases has been implicated in neurologic disorder and cancer in humans. However, cellular responses to unresolved, elevated fatty aldehyde levels are poorly understood. Here, we report that ALH-4 is a C. elegans aldehyde dehydrogenase that specifically associates with the endoplasmic reticulum, mitochondria and peroxisomes. Based on lipidomic and imaging analysis, we show that the loss of ALH-4 increases fatty aldehyde levels and reduces fat storage. ALH-4 deficiency in the intestine, cell-nonautonomously induces NHR-49/NHR-79-dependent hypodermal peroxisome proliferation. This is accompanied by the upregulation of catalases and fatty acid catabolic enzymes, as indicated by RNA sequencing. Such a response is required to counteract ALH-4 deficiency since alh-4; nhr-49 double mutant animals are sterile. Our work reveals unexpected inter-tissue communication of fatty aldehyde levels and suggests pharmacological modulation of peroxisome proliferation as a therapeutic strategy to tackle pathology related to excess fatty aldehydes.


Assuntos
Aldeído Desidrogenase/genética , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Peroxissomos/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Aldeído Desidrogenase/química , Aldeído Desidrogenase/metabolismo , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans/citologia , Caenorhabditis elegans/genética , Caenorhabditis elegans/crescimento & desenvolvimento , Proteínas de Caenorhabditis elegans/genética , Regulação da Expressão Gênica , Lipase/genética , Lipase/metabolismo , Gotículas Lipídicas/metabolismo , Lipólise/genética , Mutação , Peroxissomos/genética , Receptores Citoplasmáticos e Nucleares/genética
15.
Development ; 148(3)2021 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-33558315

RESUMO

Quiescence is a cellular state in which a cell remains out of the cell cycle but retains the capacity to divide. The unique ability of adult stem cells to maintain quiescence is crucial for life-long tissue homeostasis and regenerative capacity. Quiescence has long been viewed as an inactive state but recent studies have shown that it is in fact an actively regulated process and that adult stem cells are highly reactive to extrinsic stimuli. This has fuelled hopes of boosting the reactivation potential of adult stem cells to improve tissue function during ageing. In this Review, we provide a perspective of the quiescent state and discuss how quiescent adult stem cells transition into the cell cycle. We also discuss current challenges in the field, highlighting recent technical advances that could help overcome some of these challenges.


Assuntos
Ciclo Celular/fisiologia , Divisão Celular/fisiologia , Células-Tronco/citologia , Células-Tronco/fisiologia , Células-Tronco Adultas/citologia , Células-Tronco Adultas/fisiologia , Diferenciação Celular , Proliferação de Células , Epigenômica , Homeostase/fisiologia , Humanos
16.
Proc Natl Acad Sci U S A ; 117(51): 32464-32475, 2020 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-33293420

RESUMO

Epigenetics regulation plays a critical role in determining cell identity by controlling the accessibility of lineage-specific regulatory regions. In muscle stem cells, epigenetic mechanisms of how chromatin accessibility is modulated during cell fate determination are not fully understood. Here, we identified a long noncoding RNA, LncMyoD, that functions as a chromatin modulator for myogenic lineage determination and progression. The depletion of LncMyoD in muscle stem cells led to the down-regulation of myogenic genes and defects in myogenic differentiation. LncMyoD exclusively binds with MyoD and not with other myogenic regulatory factors and promotes transactivation of target genes. The mechanistic study revealed that loss of LncMyoD prevents the establishment of a permissive chromatin environment at myogenic E-box-containing regions, therefore restricting the binding of MyoD. Furthermore, the depletion of LncMyoD strongly impairs the reprogramming of fibroblasts into the myogenic lineage. Taken together, our study shows that LncMyoD associates with MyoD and promotes myogenic gene expression through modulating MyoD accessibility to chromatin, thereby regulating myogenic lineage determination and progression.


Assuntos
Cromatina/genética , RNA Longo não Codificante/genética , Células Satélites de Músculo Esquelético/fisiologia , Animais , Diferenciação Celular/genética , Linhagem da Célula , Transdiferenciação Celular , Cromatina/metabolismo , Feminino , Fibroblastos/citologia , Fibroblastos/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos Endogâmicos C57BL , Desenvolvimento Muscular/fisiologia , Proteína MyoD/genética , Mioblastos/citologia , Mioblastos/fisiologia , Células Satélites de Músculo Esquelético/citologia
17.
STAR Protoc ; 1(3): 100128, 2020 12 18.
Artigo em Inglês | MEDLINE | ID: mdl-33377022

RESUMO

Quiescent muscle stem cells, also called satellite cells (SCs), are essential for muscle regeneration. However, quiescent SCs are quickly activated during fluorescence-activated cell sorting (FACS) isolation. Here, we present an optimized protocol to isolate quiescent muscle stem cells from fixative-perfused mice and generate high-quality cDNA libraries for RNA-sequencing analysis. Fixation preserves the signatures of quiescent muscle stem cells in vivo. Isolated SCs can be used for downstream analysis such as immunofluorescence, RNA sequencing, and mass spectrometry. For complete information on the use and execution of this protocol, please refer to Yue et al. (2020).


Assuntos
Separação Celular/métodos , Perfusão/métodos , Células Satélites de Músculo Esquelético/citologia , Animais , Fixadores/química , Biblioteca Gênica , Camundongos , Músculo Esquelético/citologia , Regeneração/fisiologia , Análise de Sequência de RNA/métodos , Fixação de Tecidos/métodos
18.
Dev Cell ; 53(6): 661-676.e6, 2020 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-32502396

RESUMO

Adult stem cells are essential for tissue regeneration. However, the mechanisms underlying the activation of quiescent adult stem cells remain elusive. Using skeletal muscle stem cells, also called satellite cells (SCs), we demonstrate prevalent intron retention (IR) in the transcriptome of quiescent SCs (QSCs). Intron-retained transcripts found in QSCs are essential for fundamental functions including RNA splicing, protein translation, cell-cycle entry, and lineage specification. Further analysis reveals that phosphorylated Dek protein modulates IR during SC quiescence exit. While Dek protein is absent in QSCs, Dek overexpression in vivo results in a global decrease of IR, quiescence dysregulation, premature differentiation of QSCs, and undermined muscle regeneration. Moreover, IR analysis on hundreds of public RNA-seq data show that IR is conserved among quiescent adult stem cells. Altogether, we illustrate IR as a conserved post-transcriptional regulation mechanism that plays an important role during stem cell quiescence exit.


Assuntos
Diferenciação Celular , Proteínas de Ligação a DNA/metabolismo , Íntrons , Proteínas Oncogênicas/metabolismo , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , Processamento Pós-Transcricional do RNA , Células Satélites de Músculo Esquelético/metabolismo , Animais , Células Cultivadas , Proteínas de Ligação a DNA/genética , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Oncogênicas/genética , Proteínas de Ligação a Poli-ADP-Ribose/genética , Células Satélites de Músculo Esquelético/citologia , Transcriptoma
19.
Cell Rep ; 31(3): 107530, 2020 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-32320664

RESUMO

Impairment of microglial clearance activity contributes to beta-amyloid (Aß) pathology in Alzheimer's disease (AD). While the transcriptome profile of microglia directs microglial functions, how the microglial transcriptome can be regulated to alleviate AD pathology is largely unknown. Here, we show that injection of interleukin (IL)-33 in an AD transgenic mouse model ameliorates Aß pathology by reprogramming microglial epigenetic and transcriptomic profiles to induce a microglial subpopulation with enhanced phagocytic activity. These IL-33-responsive microglia (IL-33RMs) express a distinct transcriptome signature that is highlighted by increased major histocompatibility complex class II genes and restored homeostatic signature genes. IL-33-induced remodeling of chromatin accessibility and PU.1 transcription factor binding at the signature genes of IL-33RM control their transcriptome reprogramming. Specifically, disrupting PU.1-DNA interaction abolishes the microglial state transition and Aß clearance that is induced by IL-33. Thus, we define a PU.1-dependent transcriptional pathway that drives the IL-33-induced functional state transition of microglia, resulting in enhanced Aß clearance.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/genética , Interleucina-33/farmacologia , Microglia/efeitos dos fármacos , Microglia/metabolismo , Proteínas Proto-Oncogênicas/genética , Transativadores/genética , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/genética , Peptídeos beta-Amiloides/metabolismo , Animais , Cromatina/genética , Cromatina/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Interleucina-33/genética , Masculino , Camundongos , Camundongos Transgênicos , Microglia/patologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Recombinantes/farmacologia , Transativadores/metabolismo , Transcriptoma/efeitos dos fármacos
20.
Light Sci Appl ; 9: 25, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32133128

RESUMO

Coherent Raman scattering (CRS) microscopy is widely recognized as a powerful tool for tackling biomedical problems based on its chemically specific label-free contrast, high spatial and spectral resolution, and high sensitivity. However, the clinical translation of CRS imaging technologies has long been hindered by traditional solid-state lasers with environmentally sensitive operations and large footprints. Ultrafast fibre lasers can potentially overcome these shortcomings but have not yet been fully exploited for CRS imaging, as previous implementations have suffered from high intensity noise, a narrow tuning range and low power, resulting in low image qualities and slow imaging speeds. Here, we present a novel high-power self-synchronized two-colour pulsed fibre laser that achieves excellent performance in terms of intensity stability (improved by 50 dB), timing jitter (24.3 fs), average power fluctuation (<0.5%), modulation depth (>20 dB) and pulse width variation (<1.8%) over an extended wavenumber range (2700-3550 cm-1). The versatility of the laser source enables, for the first time, high-contrast, fast CRS imaging without complicated noise reduction via balanced detection schemes. These capabilities are demonstrated in this work by imaging a wide range of species such as living human cells and mouse arterial tissues and performing multimodal nonlinear imaging of mouse tail, kidney and brain tissue sections by utilizing second-harmonic generation and two-photon excited fluorescence, which provides multiple optical contrast mechanisms simultaneously and maximizes the gathered information content for biological visualization and medical diagnosis. This work also establishes a general scenario for remodelling existing lasers into synchronized two-colour lasers and thus promotes a wider popularization and application of CRS imaging technologies.

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