Cell Regeneration最新文献

Intestinal homeostasis is sustained by self-renewal of intestinal stem cells (ISCs), which continuously divide and produce proliferative transit-amplifying (TA) and then progenitor cells. Eukaryotic translation initiation factor 5A (eIF5A), a conserved translation factor, involves in a variety of cellular processes, yet its role in intestinal homeostasis remains unclear. Here, we demonstrate that eIF5A is indispensable for maintaining intestinal epithelial homeostasis. Conditional knockout of Eif5a in the adult mouse intestinal epithelium leads to stem cell loss, suppressed cell proliferation, and increased apoptosis within the crypts, concurrent with shortened gut length, reduced mouse body weight and rapid animal mortality. Consistently, Eif5a deletion in intestinal organoids also exhibits resembling cellular phenotypes. Mass spectrometry analysis reveals a significant downregulation of mitochondrial proteins, particularly those involved in mitochondrial translation, upon eIF5A depletion. Analysis of a published single-cell RNA sequencing dataset shows that mitochondrial translation-related genes, including Dars2, are highly expressed in ISC, TA and progenitor cells. Furthermore, eIF5A-deficient organoids exhibit impaired mitochondrial function, characterized by reduced ATP levels and increased reactive oxygen species (ROS). These findings highlight a critical role for eIF5A in sustaining intestinal epithelial homeostasis by regulating mitochondrial translation, providing a new insight into the molecular mechanism underlying intestinal stem cell renewal and tissue maintenance.

Myocardial infarction (MI) is a leading cause of death and disability worldwide. The promotion of myocardial regeneration is a promising therapeutic strategy for acute MI. Using a zebrafish ventricular ablation system, we found that the Mongolian traditional medicine Eerdun-Wurile (EW) promotes myocardial regeneration in zebrafish. EW treatment significantly accelerated proliferation of myocardial cells and improved cardiac function. Transcriptome sequencing revealed a significant decrease in mevalonate diphosphate decarboxylase a (mvda) expression in the metronidazole-induced ventricular ablation group, whereas mvda expression was restored in the EW group. mvda knockdown using morpholino oligonucleotides reversed the EW-mediated myocardial regeneration, whereas mvda overexpression enhanced the regenerative ability. In conclusion, EW may promote zebrafish myocardial regeneration, accelerate myocardial cell proliferation, and improve cardiac function by upregulating mvda expression. Our data partially revealed the molecular mechanism by which EW promotes myocardial regeneration and repair, and provides experimental data and novel insights for advancing MI treatment.

Mechanical properties of cells have been proposed as potential biophysical markers for cell phenotypes and functions since they are vital for maintaining biological activities. However, current approaches used to measure single-cell mechanics suffer from low throughput, high technical complexity, and stringent equipment requirements, which cannot satisfy the demand for large-scale cell sample testing. In this study, we proposed to evaluate cell stiffness at the single-cell level using deep learning. The image-based deep learning models could non-invasively predict the stiffness ranges of mesenchymal stem cells (MSCs) and macrophages in situ with high throughput and high sensitivity. We further applied the models to evaluate MSC functions including senescence, stemness, and immunomodulatory capacity as well as macrophage diversity in phenotypes and functions. Our image-based deep learning models provide potential techniques and perspectives for cell-based mechanobiology research and clinical translation.

Alternative splicing is a key regulatory mechanism that generates transcriptomic diversity by selectively splicing pre-RNA molecules in different ways, leading to the production of multiple RNA isoforms from a single gene. This process is crucial for the fine-tuning of gene expression and is tightly regulated during various biological processes. Recent studies have highlighted how alternative splicing contributes to stem cells self-renewal and differentiation, as well as how dysregulation of splicing factors can impact stem cells behavior and lead to developmental abnormalities or diseases. This review summarizes the current understanding of alternative splicing in stem cells and development, focusing on the molecular mechanisms that govern alternative splicing regulation, the role of splicing factors, and the impact of splicing isoforms on stem cell fate determination and developmental processes. We also discuss emerging technologies, such as CRISPR/Cas-based tools, single-cell long-read RNA sequencing, imaging technologies and 3D culture systems, which are advancing our ability to study alternative splicing in vitro and in vivo. Overall, this field is rapidly evolving, revealing new insights into how alternative splicing shapes the molecular landscape and functions of stem cells and developmental processes.

N6-methyladenosine (m⁶A) plays crucial roles in development and cellular reprogramming. During embryonic development, pluripotency transitions from a naïve to a primed state, and modeling the reverse primed-to-naïve transition (PNT) provides a valuable framework for investigating pluripotency regulation. Here, we show that inhibiting METTL3 significantly promotes PNT in an m⁶A-dependent manner. Mechanistically, we found that suppressing METTL3 and YTHDF2 prolongs the lifetimes of pluripotency-associated mRNAs, such as Nanog and Sox2, during PNT. In addition, Gstp1 was identified as a downstream target of METTL3 inhibition and YTHDF2 knockout. Gstp1 overexpression enhances PNT, whereas its inhibition impedes the transition. Overall, our findings suggest that YTHDF2 facilitates the removal of pluripotency gene transcripts and Gstp1, thereby promoting PNT reprogramming through m⁶A-mediated posttranscriptional control.

The efficient differentiation of adult gastric stem cells into specific epithelial cell types is crucial for gastric homeostasis. Although it is well appreciated that the niche plays a critical role in gastric epithelium cell differentiation, the relevant molecular factors and the underlying regulatory mechanisms remain poorly understood. In this study, by combining the knowledge of the niche cells obtained from single-cell RNA sequencing and manipulation of signaling pathways, we achieved effective differentiation of various gastric epithelial cell types in mouse and human gastric organoids. These in vitro differentiated cells showed a similar gene expression profile to those in gastric tissues. Specifically, BMP4 signaling stimulates pit cell and parietal cell differentiation. Furthermore, BMP4 and EGF signaling cooperate to enhance pit cell differentiation, whereas inhibition of TGF-β and BMP4 signaling promotes chief cell differentiation. We demonstrated that Zbtb7b is a novel regulator controlling pit cell differentiation. In addition, BMP4, together with the small molecule Isoxazole 9, promotes parietal and enteroendocrine cell differentiation. Our data also revealed the different requirements of parietal and chief cell differentiation between mouse and human. Together, our findings provide a mechanistic insight into gastric epithelial cell differentiation and uncover its similarities and differences between mouse and human, laying a foundation for future investigation and potential clinical use of gastric organoids.

Spinal motoneurons control muscle fibers contraction and drive all motor behaviors in vertebrates. Although spinal motoneurons share the fundamental role of innervating muscle fibers, they exhibit remarkable diversity that reflects their specific identities. Defining the morphological changes during postnatal development is critical for elucidating this diversity. However, our understanding of the three-dimensional (3D) morphology of spinal motoneurons at these stages remains limited, largely due to the lack of high-throughput imaging tools. Using tiling light sheet microscopy combined with tissue clearing methods, we imaged motoneurons of the lateral and median motor column in the cervical and lumbar cord during postnatal development. By analyzing their soma size, we found that motoneurons innervating the upper limbs differentiate into two subpopulations with distinct soma size by postnatal day 14 (P14), while differentiation of motoneurons innervating the lower limbs is delayed. Furthermore, coupling adenovirus labeling with 3D volumetric reconstruction, we traced and measured the number and lengths of dendrites of flexor and extensor motoneurons in the lumbar cord, finding that the number of dendrites initially increases and subsequently declines as dendritic order rises. Together, these findings provide a quantitative analysis of the 3D morphological changes underlying spinal motoneuron diversity.

Gastric cancer peritoneal metastasis (GCPM) typically indicates a poor clinical prognosis and is frequently observed in diffuse gastric cancer (GC) patients with CDH1 loss of function. GCPM characterized for its aggressiveness and resistance to chemotherapy, most notably paclitaxel (PTX), poses significant treatment challenges. Previously, no mouse gastric adenocarcinoma (MGA) cell lines with Trp53 (encoding mouse p53) and Cdh1 (encoding mouse E-cadherin) mutations and a high potential for peritoneal metastasis in mice have been established. Here, we derived a mouse GC cell line, called MTC, from subcutaneously transplanted mouse Trp53^-/-Cdh1^-/- GC organoids. Through matching the short tandem repeat profile of MTC with those in current cell banks, we verified the uniqueness of MTC. Furtherly, we confirmed the features of MTC by detecting the expression of p53, E-cadherin, and pan-CK. After long-term exposure of the original MTC line to PTX, we developed a more aggressive, PTX-resistant cell line, termed MTC-R. Compared with MTC, MTC-R demonstrated enhanced tumorigenicity and high potential for peritoneal metastasis in subcutaneous and intraperitoneal tumour models both in BALB/c nude mice and C57BL/6 J mice. Transcriptome analysis revealed the ECM‒receptor interaction pathway activation during the development of PTX resistance, and dasatinib (DASA) was identified as a potential drug targeting this pathway. DASA showed promise in ameliorating disease progression and improving overall survival in MTC-R GCPM model in C57BL/6 J mice. Overall, we established a novel MGA cell line with Trp53 and Cdh1 mutations and its PTX-resistant variant and demonstrated the efficacy of DASA in treating PTX-resistant GCPM.

The fetal liver is the primary site for the expansion of hematopoietic stem and progenitor cells (HSPCs) during fetal hematopoiesis. However, the spatial organization of different hematopoietic progenitor populations within the fetal liver remains poorly characterized. In this study, we utilized SeekSpace, a high-resolution single-nucleus spatial transcriptomics platform, to map the spatial distribution of hematopoietic stem cells and multipotent progenitor cells (HSC/MPPs) and downstream restricted progenitors (RPs) in relation to other hematopoietic and stromal cell populations in the fetal liver at embryonic day 13.5. Using SeekSpace, we constructed a detailed single-cell spatial transcriptomic atlas of fetal liver hematopoiesis, revealing that both HSC/MPPs and many RPs undergo active expansion in the fetal liver, a process distinct from their behavior in adult bone marrow. Proximity analysis and in situ imaging demonstrated that HSC/MPPs expansion occurs in close association with macrophages and endothelial cells throughout the fetal liver, supported by signaling pathways involving IGF and collagen. In contrast, RPs exhibited no specific spatial proximity to other cell populations during their expansion. Collectively, this study provides a comprehensive resource for understanding the spatial and molecular mechanisms underlying HSC/MPPs and RP expansion during fetal liver hematopoiesis.

Autophagy is a crucial cellular process that facilitates the degradation of damaged organelles and protein aggregates, and the recycling of cellular components for the energy production and macromolecule synthesis. It plays an indispensable role in maintaining cellular homeostasis. Over recent decades, research has increasingly focused on the role of autophagy in regulating adult stem cells (SCs). Studies suggest that autophagy modulates various cellular processes and states of adult SCs, including quiescence, proliferation, self-renewal, and differentiation. The primary role of autophagy in these contexts is to sustain homeostasis, withstand stressors, and supply energy. Notably, the dysfunction of adult SCs during aging is correlated with a decline in autophagic activity, suggesting that autophagy is also involved in SC- and aging-associated disorders. Given the diverse cellular processes mediated by autophagy and the intricate mechanisms governing adult SCs, further research is essential to elucidate both universal and cell type-specific regulatory pathways of autophagy. This review discusses the role of autophagy in regulating adult SCs during quiescence, proliferation, self-renewal, and differentiation. Additionally, it summarizes the relationship between SC aging and autophagy, providing therapeutical insights into treating and ameliorating aging-associated diseases and cancers, and ultimately promoting longevity.