Cell and Tissue Research最新文献

The integrin-associated proteins (IAPs) function in a tightly regulated and coordinated manner to maintain the complex cytoarchitecture at the myotendinous junctions (MTJs) of Drosophila indirect flight muscles (IFMs). Parvin, a conserved but less explored IAP, forms a ternary complex with ILK and PINCH (the IPP complex). Although the IPP complex is functionally conserved, playing a central role in integrin-mediated adhesion, its individual components may also exert independent roles. The present study investigates parvin as a critical regulator of IFM function and sarcomeric integrity. Downregulation of parvin in IFM leads to altered sarcomere organisation and lowered accumulation of essential thin filament genes, including Act88F, wupA, up, TpnC4, and TM2 transcripts. Furthermore, Z-disc-associated proteins such as Zasp52 and its binding partner α-actinin, which are vital for myofibril stability, showed markedly reduced expression in parvin-deficient muscles. Notably, Zasp52 failed to localise to the Z-discs in IFMs, despite being detectable in leg muscles, suggesting tissue-specific mislocalisation. The expression of sallimus, a titin orthologue contributing to muscle elasticity, remained unchanged. Our findings underscore parvin's essential role in preserving IFM ultrastructure and function. We also emphasise the importance of maintaining the stoichiometric balance within the IPP complex-including its extended member Ras suppressor 1 (RSU1)-for proper muscle performance. Additionally, perturbation of parvin expression in a tissue-specific manner revealed its broader role in fly viability and muscle-driven behaviours, including larval locomotion and leg muscle function. Collectively, this study positions parvin as a pivotal component in maintaining muscle integrity across multiple muscle types in Drosophila.

In the larvae of the mosquito Aedes aegypti, the three most important endocrine glands, the corpora allata (CA), the corpora cardiaca (CC), and the prothoracic gland (PG), together form the glandular complexes (GC). Using confocal laser scanning microscopy in combination with immunohistochemistry, in situ hybridization, ultrastructural expansion microscopy, and apoptosis studies, we were able to identify the different cell types of the GC and follow their fate during metamorphosis. Our studies revealed that the CC is not a well-defined organ but consists of individual cells randomly distributed within the GC and CA-CC complexes. Furthermore, imaging and in situ hybridization show that the CA is a compact organ composed of a single cell type. We observed that CA and CC survive during the larval-to-adult transition, while PG undergoes apoptosis and disappears within the first 24 h of adult life. This study lays the foundation for a more detailed understanding of the structure and changes in the major endocrine organs of mosquitoes, which are vectors of several important infectious diseases.

Angiogenesis, the formation of new blood vessels from pre-existing vasculature, is a crucial process in both physiological and pathological contexts, including cancer. Phospholipases A₂ (PLA₂s), enzymes found in snake venoms, have attracted attention due to their potential antiangiogenic properties. In this study, we explored the antiangiogenic effects of PLA₂ isoforms isolated from Bothrops diporus venom using a combination of in vivo and ex vivo models. The chorioallantoic membrane (CAM) assay revealed a significant reduction in vascular density and branching following PLA₂s treatment, with histological analysis confirming vascular regression, including vessel wall thinning and luminal collapse. Moreover, PLA₂s induced endothelial cell apoptosis, as shown by TUNEL staining, and reduced VEGF expression. The filter paper disc variant of the CAM assay further supported these findings, demonstrating inhibited neovascularization while preserving mature vessels. Additionally, the CAM explant assay showed a marked decrease in vascular complexity and branching. These results demonstrate the antiangiogenic effect of PLA₂ isoforms from B. diporus and suggest that these enzymes may modulate key angiogenic pathways. Based on our previous findings, this modulation may involve interference with integrin-mediated signaling, which could underlie the vascular effects observed. Thus, this work provides compelling evidence for the potential role of snake venom-derived PLA₂s in modulating angiogenesis and highlights the need for further research into their mechanisms and possible biomedical applications.

The settlement levels of indigenous bacteria show circadian rhythms in various regions of the rat alimentary tract. Numerous bacteria colonize between the mucosal folds of the ascending colon in rodents; however, the rhythm of bacteria colonizing the ascending colon remains to be clarified. Therefore, we first aimed to examine the diurnal changes in bacteria colonizing in the rat ascending colon. The settlement levels of indigenous bacteria were significantly higher at zeitgeber time (ZT) 18 (dark phase) than at ZT6 (light phase) in the region encompassing the aggregated lymphoid tissue in the ascending colon (ALT-AC). The bacterial composition around the ALT-AC was dominated by the phylum Firmicutes and the family Lachnospiraceae, displaying notable distinctions from the compositions found in cecal contents and feces. The relative abundance of some bacterial species around the ALT-AC, such as Mucispirillum schaedleri, changed significantly between ZT6 and ZT18. Furthermore, we explored the effect of bacterial expansion on gene expression in the ALT-AC at ZT18 by administrating antibiotics for 1 day to inhibit bacterial growth. The antibiotic-treated group exhibited significant downregulation of multiple genes, including those associated with cell proliferation (Plk3), differentiation into goblet cells (Spdef, Atoh1, Bhlha15), and Golgi organization (Gorasp2). These results suggested that indigenous bacteria around the rat ALT-AC undergo diurnal changes in both settlement levels, peaking at the dark phase, and bacterial composition. In addition, bacterial expansion during the dark phase can induce changes in the expression of diverse genes, including genes associated with goblet cell differentiation.

Podocytes are a type of epithelial cells that form the kidney nephron and are essential for ultrafiltration in the glomerulus. The majority of nephron constituent cells have primary cilia, which play an important role in the maintenance of normal tubular architecture of nephron. However, whether podocytes have primary cilia was only partially understood. In general, immunohistochemistry with an anti-acetylated α-tubulin antibody is often used to visualize primary cilia. α-Tubulin is highly acetylated throughout podocytes, and this antibody is not suitable to determine the presence or absence of primary cilia in podocytes, which is one reason why the presence of primary cilia in podocytes has remained unclear. In the present study, we determined the presence or absence of primary cilia in mature podocytes of six vertebrate species using a recently reported array tomography workflow optimized for whole glomerulus analysis. The proportion of podocytes with primary cilia tended to decrease with evolution as follows: 89.6% in river lampreys (Agnatha), 42.1% in zebrafish (Osteichthyes), 43.3% in African clawed frogs (Amphibia), 17.3% in Reeves' turtles (Reptilia), 10.4% in common quails (Aves), and 0.0% in Wistar rats (Mammalia). Our previous study has reported that, in rats, primary cilia are present in podocytes during development and disappear in mature podocytes. In other words, primary cilia disappear from podocytes during both phylogeny and ontogeny in vertebrates. We discuss the triggers and significance of primary cilium disappearance from vertebrate podocytes.

The compartments and lymphocyte subsets of the human spleen differ from the spleen of rodents. The red pulp removes old red cells or malformed erythrocytes found in spherocytosis. The high blood flow is also necessary to filter bacteria like pneumococci from the blood. Without a spleen or after splenectomy, there is the risk of a fatal postsplenectomy sepsis. Therefore, these patients have to be vaccinated. Splenic particles can regenerate. The spleen is very important in lymphocyte recirculation and lymphocyte production. The marginal zone B lymphocytes are unique and important for B memory in man.

The corpus luteum (CL) is an ovarian structure that secretes progesterone (P4) following ovulation, playing a crucial role in regulating the estrous cycle and maintaining pregnancy. Luteolysis, the structural and functional degradation of the CL, occurs through apoptosis and autophagy. Recent studies suggest that the circadian clock (CC) system, particularly the gene NR1D1, is involved in these processes. This study investigated the role of NR1D1 in bovine CL regression using an ex vivo model treated with prostaglandin F_2α (PGF_2α), the NR1D1 agonist GSK4112, and the antagonist SR8278. CL samples were classified into four estrous cycle stages based on ovarian morphology and analyzed for P4 secretion, as well as gene and protein expression related to steroid synthesis, the CC system, autophagy, and apoptosis. P4 levels, steroid synthesis-related genes, and CC system-related genes, including NR1D1, were highly expressed in the mid and late stages of the CL, whereas autophagy- and apoptosis-related genes peaked during regression. Western blotting and immunofluorescence revealed increased expression of NR1D1 and BMAL1 in the mid and late stages, while LC3 and CTSB were most prominent during regression. PGF_2α treatment reduced NR1D1 and BMAL1 expression, along with decreased P4 levels and increased apoptosis markers. GSK4112 suppressed steroid synthesis while upregulating autophagy- and apoptosis-related genes. Conversely, SR8278 reversed PGF_2α-induced luteal regression, restoring P4 and steroidogenic gene expression while suppressing CTSB. These findings suggest that NR1D1 interacts with PGF_2α to regulate CL regression, highlighting the CC system as a potential target for improving reproductive efficiency in cattle.

Reperfusion following myocardial infarction salvages the ischemic heart but paradoxically exacerbates injury. Yet, efficient treatment for cardiac ischemia/reperfusion injury is still missing in clinics. ATP release through Pannexin1 (PANX1) channels facilitates recruitment of leukocytes to the injured myocardium. Thus, PANX1 channel inhibition might confer cardioprotection. Currently available PANX1 channel blockers lack specificity or in vivo stability. Nanobodies offer a new therapeutic modality given their high target affinity, small size, and deep tissue penetration. Nanobodies targeting Panx1 were recently introduced. Here, their target specificity and selective PANX1 channel inhibition for cardiovascular purposes were validated in vitro. The two most promising candidates were further examined in the context of cardiac ischemia/reperfusion injury. Nanobody-1 (Nb1) and Nb9 reduced neutrophil adhesion to an endothelial monolayer. Nb1 did not affect left ventricular function ex vivo; however, Nb9 tended to diminish the performance of isolated hearts. Finally, in vivo application of Nb1, but not of Nb9 or a control Nb, at the onset of reperfusion increased the survival rate of mice. However, the infarct size observed after treatment with Nb1 was similar than the one found after treatment with the control Nb. In conclusion, Nb1 efficiently and specifically inhibits ATP release from endothelial cells thereby limiting leukocyte adhesion and improving the outcome of cardiac ischemia/reperfusion in mice. This warrants further studies to unveil the detailed molecular mechanism underlying the beneficial effects of Nb1.

Mechanisms governing somatic cell interactions in the testis are not well defined. The platelet-derived growth factor (PDGF) pathway mediates epithelial-mesenchymal interactions and is involved in testicular morphogenesis in rodents. However, its roles in the testis of higher mammals remain largely unknown. Here, we investigated how PDGF signaling inhibition affects immature (1-week-old) porcine testicular tubular somatic cells (TTSCs), including cell-cell communication and morphogenesis. From scRNA-seq data, we established the PDGF pathway signatures in crosstalk between testicular cells, identifying Sertoli cells as the primary source and peritubular myoid cells as the main recipients of PDGF. Further, we demonstrated that PDGF inhibition by CP673451 affects TTSC functions, proliferation and cytoskeleton, with reduced cell area, focal adhesion size and fibronectin production, and decreased expression of peritubular myoid cell-specific genes. PDGF inhibition did not impair testicular organoid formation and tubule morphogenesis in vitro, but it correlated with ablation of cytoplasmic extensions from the tubule surface, potentially related to interactions between TTSCs and the extracellular matrix. PDGF signaling can be transduced by primary cilia, sensory organelles that respond to environmental stimuli, and PDGF inhibition increased the percentage of ciliated cells and ciliary length in TTSCs. Comprehensive morphological characterization of primary cilia in the porcine testis indicated that these remain submerged in the cytoplasm. In conclusion, the PDGF signaling pathway is active in the immature pig testis and may influence testis morphogenesis by affecting cell-extracellular matrix interaction, cytoskeleton and primary cilia. However, the role of primary cilia-modulated PDGF signaling in the testis remains to be determined.

Previous studies revealed the presence of several tight junction (TJ) proteins in the centrosome and their interaction with various centriolar proteins, prompting us to analyze whether this also applies to the TJ protein ZO-2. Here, we found that ZO-2 colocalizes with CEP164 in the distal appendage of the mother centriole and is also present in the pericentriolar region, mitotic spindle poles, the basal body of primary cilia, and the tail of spermatozoa. The absence of ZO-2 altered the cellular content of centriolar proteins CEP164, centriolin, and CEP135, but did not change the morphology of centrioles. ZO-2 depletion inhibits the development of astral and mitotic spindle microtubules expressing EB1. At the spindle poles, ZO-2 depletion increases the accumulation of NuMA while reducing the levels of kinesin KIF14 and the TPX2 scaffold, and the accumulation of the kinase p-Aurora, leading to a decrease in mitotic spindle length, microtubule instability, and abnormal chromosome congression. KIF14, NuMA, and p-Aurora co-immunoprecipitate with ZO-2, and NuMA and Aurora-A bind to different segments of ZO-2. At the ciliary basal body, ZO-2 depletion reduces the content of CEP164, KIF14, and IFT-B protein IFT57, while increasing the expression of p-Aurora and pAKT. These changes block primary cilium development and the response to Sonic Hedgehog signaling pathway stimulation. These results suggest that, rather than being a centrosomal architectural component, ZO-2 enhances microtubule stability and serves as a scaffold that facilitates the adequate accumulation of spindle pole and centriole proteins, allowing proper poleward spindle microtubule flux and cilia development.