Cell and Tissue Research最新文献

Recent large-scale genome-wide association and single-cell RNA sequencing (scRNA-seq) studies have uncovered disease-associated genetic risk factors and cell type-specific genetic alterations. However, our understanding of how these genetic variants cause diseases and the underlying mechanisms remains largely unknown. Functional genomics screens using CRISPR-based technologies offer an effective tool for studying genes relevant to disease phenotypes. Here, we summarize recent CRISPR-based functional genomics screen approaches applied to human pluripotent stem cell (hPSC)-derived neurons and brain organoids. These screens have identified genes crucial for neurogenesis, neuronal survival, morphological development, and migration. Combining CRISPR-based genetic screens with scRNA-seq, researchers have revealed downstream genes and cellular pathways impacted by these genetic variants in human neural cells, providing new insights into the pathogenesis of neurodevelopmental disorders, such as microcephaly and autism spectrum disorders. Finally, we discuss current challenges and future directions for using CRISPR-based screens in furthering our understanding of neurological diseases and developing potential therapeutic strategies. Despite challenges, CRISPR-based screens have enormous potential for advancing the therapeutic development of many diseases.

Interstitial cells of Cajal (ICC) and PDGFRα⁺ cells regulate smooth muscle motility in the gastrointestinal (GI) tract, yet their function in the esophagus remains unknown. The mouse esophagus has been described as primarily skeletal muscle; however, ICC  have been identified in this region. This study characterizes the distribution of skeletal and smooth muscle cells (SMCs) and their spatial relationship to ICC, PDGFRα⁺ cells, and intramuscular motor neurons in the mouse esophagus. SMCs occupied approximately 30% of the distal esophagus, but their density declined in more proximal regions. Similarly, ANO1⁺ intramuscular ICC (ICC-IM) were distributed along the esophagus, with density decreasing proximally. While ICC-IM were closely associated with SMCs, they were also present in regions of skeletal muscle. Intramuscular, submucosal, and myenteric PDGFRα⁺ cells were densely distributed throughout the esophagus, yet only intramuscular PDGFRα⁺ cells in the lower esophageal sphincter (LES) and distal esophagus expressed SK3. ICC-IM and PDGFRα⁺ cells were closely associated with intramuscular nNOS⁺, VIP⁺, VAChT⁺, and TH⁺ neurons and GFAP⁺ cells resembling intramuscular enteric glia. These findings suggest that ICC-IM and PDGFRα⁺ cells may have roles in regulating esophageal motility due to their close proximity to each other and to skeletal muscle and SMCs, although further functional studies are needed to explore their role in this region. The mixed muscular composition and presence of interstitial cells in the mouse distal esophagus is anatomically similar to the transitional zone found in the human esophagus, and therefore, motility studies in the mouse may be translatable to humans.

α-Smooth muscle actin (α-SMA) is an actin isoform commonly found within vascular smooth muscle cells. Moreover, α-SMA-positive cells are localized in the dental follicle (DF). DF is derived from alveolar bone (AB), cementum, and periodontal ligament (PDL). Therefore, α-SMA-positive cells in the periodontal tissue are speculated to be a marker for mesenchymal stem cells during tooth development. In particular, the mechanism of osteoblast differentiation is not clear. This study demonstrated the fate of α-SMA-positive cells around the tooth germ immunohistochemically. First, α-SMA- and Runx2-positive localization at embryonic days (E) 13, E14, postnatal days (P) 9, and P15 was demonstrated. α-SMA- and Runx2-positive cells were detected in the upper part of the DF at P1. At P9 and P15, α-SMA-positive cells in the PDL were detected in the upper and lower parts. The positive reaction of Runx2 was also localized in the PDL. Then, the distribution of α-SMA-positive cell progeny at P9 and P15 were clarified using α-SMA-CreERT2/ROSA26-loxP-stop-loxP-tdTomato (α-SMA/tomato) mice. It has known that Runx2-positive cells differentiate into osteoblasts. In this study, some Runx2 and α-SMA-positive cells were localized in the DF and PDL. The lineage-tracing analysis demonstrated that the α-SMA/tomato-positive cells expressing Runx2 or Osterix were detected on the AB surface at P15. α-SMA/tomato-positive cells expressing type I collagen were found in the AB matrix. These results indicate that the progeny of the α-SMA-positive cells in the DF could differentiate into osteogenic cells. In conclusion, α-SMA could be a potential marker of progenitor cells that differentiate into osteoblasts.

The metabolic syndrome, or MetS, is currently a global health concern. The anti-inflammatory, anti-proliferative, and antioxidant properties of luteolin are some of its advantageous pharmacological characteristics. This research was designed to establish a MetS rat model and investigate the possible protective effect of luteolin on cardiovascular, hepatic, and metabolic changes in diet-induced metabolic syndrome in rats. Forty adult male albino rats were split into four groups: a negative control group, a group treated with luteolin, a group induced MetS (fed 20% fructose), and a group treated with luteolin (fed 20% fructose and given luteolin). Following the experiment after 8 weeks, biochemical, histological (light and electron), and immunohistochemistry analyses were performed on liver and heart tissues. Serum levels of cTnI, CK-MB, and LDH were significantly elevated in response to the cardiovascular effect of MetS. Furthermore, compared to the negative control group, the MetS group showed a marked increase in lipid peroxidation in the cardiac and hepatic tissues, as evidenced by elevated levels of MDA and a decline in the antioxidant defense system, as demonstrated by lower activities of GSH and SOD. The fatty liver-induced group exhibited histological alterations, including disrupted hepatic architecture, dilated and congested central veins, blood sinusoids, and portal veins. In addition to nuclear structural alterations, most hepatocytes displayed varying degrees of cytoplasmic vacuolation, mitochondrial alterations, and endoplasmic reticulum dilatation. These alterations were linked to inflammatory cellular infiltrations, collagen fiber deposition, active hepatic stellate cells, and scattered hypertrophied Kupffer cells, as demonstrated by electron microscopy and validated by immunohistochemical analysis. It is interesting to note that eosinophils were seen between the liver cells and in dilated blood sinusoids. Moreover, the biochemical (hepatic and cardiac) and histological (liver) changes were significantly less severe in luteolin-treated rat on a high-fructose diet. These results suggested that luteolin protects against a type of metabolic syndrome that is produced experimentally.

Decidual mesenchymal stromal cells (DMSC) were the source of extracellular vesicles (DMSC_EV). The xCELLigence real-time cell growth assay revealed increasing concentrations of EVs decreased DMSC attachment in the early growth phase but stimulated DMSC proliferation at day 7 when grown on tissue culture plastic (TCP). DMSC attachment and proliferation varied depending on the growth surface and DMSC_EV supplementation. DMSC attachment increased on decellularized and solubilized amniotic (s-dAM) whether or not EVs were added. Only Matrigel substrate increased DMSC attachment with added EVs. The addition of EVs increased DMSC proliferation only on the s-dAM substrate. DMSCs were more motile on s-dAM and decellularized and solubilized chorionic (s-dCM) membranes following EV addition. The osteogenic potential of DMSCs was improved on s-dAM substrates when supplanted with EVs. Finally, the levels of reactive oxygen species in DMSCs varied depending on the substrate but not on added EVs. We show that the addition of in vitro EVs isolated from the source being expanded (i.e., DMSCs) and the presence of ECM improve DMSC behaviours during ex vivo expansion. The inclusion of two key components of the MSC niche, EVs and ECM, benefitted the ex vivo expansion of MSCs. Added in vitro EVs increased the proliferation of DMSCs when grown on s-dAM but not on s-dCM, whereas they improved DMSC mobility on both surfaces. Testing different ECMs could be used to promote specific desired characteristics of DMSCs, and different combinations of EVs and ECM may enhance desirable MSC characteristics for specific therapeutic settings.

Blue light (BL)-induced retinal injury has become a very common problem due to over exposure to blue light-emitting sources. This study aimed to investigate the possible ameliorating impact of stromal vascular fraction cells (SVFCs) on BL-induced retinal injury. Forty male albino rats were randomly allocated into four groups. The control group rats were kept in 12-h light/12-h dark. Rats of SVFC-control as the control group, but rats were intravenously injected once by SVFCs. Rats of both the BL-group and BL-SVFC group were exposed to BL for 2 weeks; then rats of the BL-SVFC group were intravenously injected once by SVFCs. Following the BL exposure, rats were kept for 8 weeks. Physical and physiological studies were performed; then retinal tissues were collected for biochemical and histological studies. The BL-group showed physical and physiological changes indicating affection of the visual function. Biochemical marker assessment showed a significant increase in MDA, TLR4 and MYD88 tissue levels with a significant decrease in TAC levels. Histological and ultrastructural assessment showed disruption of the normal histological architecture with retinal pigment epithelium, photoreceptors, and ganglion cell deterioration. A significant increase in NF-κB, caspase-3, and GFAP immunoreactivity was also detected. BL-SVFC group showed a significant improvement in physical, physiological, and biochemical parameters. Retinal tissues revealed amelioration of retinal structural and ultrastructural deterioration and a significant decrease in NF-κB and caspase-3 immunoreactivity with a significant increase in GFAP immunoreaction. This study concluded that SVFCs could ameliorate the BL-induced retinal injury through TLR-4/MYD-88/NF-κB signaling inhibition, regenerative, anti-oxidative, and anti-apoptotic effects.

In this study, we aimed to promote the maturation of cardiomyocytes-like cells by mechanical stimulation, and evaluate their therapeutic potential against myocardial infarction. The cyclic tensile strain was used to induce the maturation of cardsiomyocyte-like cells from P19 cells in vitro. Western blot and qPCR assays were performed to examine protein and gene expression, respectively. High-resolution respirometry was used to assay cell function. The induced cells were then evaluated for their therapeutic effect. In vitro, we observed cyclic tensile strain induced P19 cell differentiation into cardiomyocyte-like cells, as indicated by the increased expression of cardiomyocyte maturation-related genes such as Myh6, Myl2, and Gja1. Furthermore, cyclic tensile strain increased the antioxidant capacity of cardiomyocytes by upregulating the expression Sirt1, a gene important for P19 maturation into cardiomyocyte-like cells. High-resolution respirometry analysis of P19 cells following cyclic tensile strain showed enhanced metabolic function. In vivo, stimulated P19 cells enhanced cardiac function in a mouse model of myocardial infarction, and these mice showed decreased infarction-related biomarkers. The current study demonstrates a simple yet effective mean to induce the maturation of P19 cells into cardiomyocyte-like cells, with a promising therapeutic potential for the treatment of myocardial infarction.

The umbilical cord epithelium (UCE) is the surface tissue that covers the umbilical cord (UC). It is widely considered a single-layered epithelium composed of squamous or cuboidal cells, which are in constant contact with amniotic fluid. The objective of this study was to elucidate the distinctive structural characteristics and abundance of specific proteins in this unique epithelium, many of which have not been previously demonstrated. Samples of the UC were obtained from term pregnancies (n = 12) and processed for examination using stereo, light, electron, and 3D high-resolution confocal microscopy. Sections displayed a range of stratification, ranging from a single squamous layer to 4-5 layers of round/cuboid cells, challenging the notion of considering it as a single-layered structure. Cells are located on a well-developed basement membrane (BM), as evidenced by the expression of BM-specific proteins and PAS staining. The cells possess distinctive cytoplasmic domains that are tightly bound to each other by desmosomes and interdigitating anchoring surfaces. Desquamations and limited organelles suggest that the cells have reached the final stages of differentiation and are no longer actively synthesizing proteins, despite maintaining stratification-specific expression levels of cytoskeletal, junctional, receptor, and stem cell proteins. Although definitive keratinization was not observed, the distribution of proteins and the distinctive structural organization of the single/multi-layered cells suggest that they exhibit plasticity, likely due to adaptive mechanisms in response to chemical and/or mechanical stimuli during fetal development. These structural alterations may facilitate the active transportation of soluble ingredients between the amniotic fluid and cord blood through an intercellular route.

Smell and taste are extensively studied in fish species as essential for finding food and selecting mates while avoiding toxic substances and predators. Depending on the evolutionary position and adaptation, a discrete variation in the morphology of these sense organs has been reported in numerous teleost species. Here, for the first time, we approach the phenotypic characterization of the olfactory epithelium and taste buds in the African turquoise killifish (Nothobranchius furzeri), a model organism known for its short lifespan and use in ageing research. Our observations indicate that the olfactory epithelium of N. furzeri is organized as a simple patch, lacking the complex folding into a rosette, with an average size of approximately 600 µm in length, 300 µm in width, and 70 µm in thickness. Three main cytotypes, including olfactory receptor neurons (CalbindinD28K), supporting cells (β-tubulin IV), and basal cells (Ki67), were identified across the epithelium. Further, we determined the taste buds' distribution and quantification between anterior (skin, lips, oral cavity) and posterior (gills, pharynx, oesophagus) systems. We identified the key cytotypes by using immunohistochemical markers, i.e. CalbindinD28K, doublecortin, and neuropeptide Y (NPY) for gustatory receptor cells, glial fibrillary acidic protein (GFAP) for supporting cells, and Ki67, a marker of cellular proliferation for basal cells. Altogether, these results indicate that N. furzeri is a microsmatic species with unique taste and olfactory features and possesses a well-developed posterior taste system compared to the anterior. This study provides fundamental insights into the chemosensory biology of N. furzeri, facilitating future investigations into nutrient-sensing mechanisms and their roles in development, survival, and ageing.

Hematopoietic stem cells (HSCs) drive cellular turnover in the hematopoietic system by balancing self-renewal and differentiation. In the adult bone marrow (BM), these cells are regulated by a complex cellular microenvironment known as "niche," which involves dynamic interactions between diverse cellular and non-cellular elements. During blood cell maturation, lineage branching is guided by clusters of genes that interact or counteract each other, forming complex networks of lineage-specific transcription factors. Disruptions in these networks can lead to obstacles in differentiation, lineage reprogramming, and ultimately malignant transformation, including acute myeloid leukemia (AML). Zinc Finger Protein 521 (Znf521/Zfp521), a conserved transcription factor enriched in HSCs in both human and murine hematopoiesis, plays a pivotal role in regulating HSC self-renewal and differentiation. Its enforced expression preserves progenitor cell activity, while inhibition promotes differentiation toward the lymphoid and myeloid lineages. Transcriptomic analysis of human AML patient samples has revealed upregulation of ZNF521 in AMLs with the t(9;11) fusion gene MLL-AF9. In vitro studies have shown that ZNF521 collaborates with MLL-AF9 to enhance the growth of transformed leukemic cells, increase colony formation, and activate MLL target genes. Conversely, inhibition of ZNF521 using short-hairpin RNA (shRNA) results in decreased leukemia proliferation, reduced colony formation, and induction of cell cycle arrest in MLL-rearranged AML cell lines. In vivo experiments have demonstrated that mZFP521-deficient mice transduced with MLL-AF9 experience a delay in leukemia development. This review provides an overview of the regulatory network involving ZNF521, which plays a crucial role in controlling both HSC self-renewal and differentiation pathways. Furthermore, we examine the impact of ZNF521 on the leukemic phenotype and consider it a potential marker for MLL-AF9⁺ AML.