Cell and Tissue Research最新文献

Conservative treatment of the anterior cruciate ligament (ACL) is important for restoring functional activity and preventing secondary degeneration. However, the molecular mechanisms underlying ligament immobilization and its precise role in the healing process remain poorly understood. In this study, we investigated the effect of immobilization on the strength of the healed ACL during acute management. We performed surgery to heal the ACL of rats and immobilized the knees using Kirschner wires. The group in which only the surgery to promote ACL healing was performed was designated as the controlled anterior tibial instability group, whereas the group that underwent both surgery and immobilization was designated as the immobilization (IMM) group. After 1-2 weeks of immobilization, histological analyses using hematoxylin-eosin staining and immunohistochemical evaluation of collagen types I and III expression were performed. A comprehensive genetic analysis in the acute phase was performed via RNA sequencing. Furthermore, fibroblasts derived from rat ACL were used to recapitulate inflammation with interleukin-1β, and its effect on elongation stress (110%) was investigated using polymerase chain reaction. Joint immobilization for 2 weeks postoperatively increased the mechanical strength of the conservatively connected ligaments. Stretch stimulation of fibroblasts with interleukin-1β also decreased the expression of the extracellular matrix. Furthermore, bioinformatics analyses identified differentially expressed genes associated with the healing process in fixed versus unfixed ligaments. The results demonstrate that acute-phase immobilization, defined as fixation for 2 weeks following injury, enhances ligament strength by promoting extracellular matrix synthesis and organized regeneration, providing novel insights into optimizing conservative ACL therapy.

The purpose of this study was to investigate the role of hsa-miR-483-3p on the regulation of extracellular matrix (ECM) in cultured human trabecular meshwork (HTM) cells with known steroid response. Primary cultures of HTM cells with known steroid responsiveness [GC-responder (GC-R) and GC-Non-responder (GC-NR) cells] were grown on coverslip in 12-well plate until 80% confluence and treated with 100 nM dexamethasone (DEX) for 24 h and transfected with different concentrations of synthetic miRNA 483-3p mimic or inhibitor. After 24 h or 72 h post transfection, the cells were harvested for the following experiments: (i) percentage transfection efficiency, (ii) RNA isolation for qPCR analysis, (iii) immunofluorescence staining, and (iv) protein isolation for Western blotting, respectively. All experiments were performed in triplicate with three biological samples (n = 3). GC-R HTM cells showed significantly higher expression of SMAD4 as compared to GC-NR HTM cells. Similarly, DEX treatment up-regulated the SMAD4-dependent ECM proteins. The presence of a miR-483-3p mimic down-regulated SMAD4 expression and SMAD4-dependent ECM production in a dose-dependent manner by negatively down-regulating SMAD4/TGF-β2 signaling. The inhibition of SMAD4-dependent ECM production by miR-483-3p was more pronounced in GC-R HTM cells as compared to GC-NR cells. The down-regulation in ECM production by miR-483-3p is SMAD4-dependent and may play a protective role in mitigating steroid response in GC-R HTM cells. Using miR-483-3p mimics demonstrates therapeutic potential for the management of steroid-induced ocular hypertension and glaucoma.

Macromolecular crowding (MMC) is a biophysical phenomenon that has proven effective in enhancing extracellular matrix (ECM) deposition in vitro. However, MMCs potential in neuroglial cell cultures remains underexplored. This study investigates the effects of three distinct MMC agents [carrageenan (CR), dextran sulphate (DxS) and FicollⓇ cocktail (FC)] on ECM deposition and cell behaviour of Neu7 and primary astrocytes. While the viability and metabolic activity of Neu7 astrocytes were unaffected by any of the crowding agents, primary astrocytes exhibited a significant decrease in viability and metabolic activity in the presence of CR and DxS. The addition of CR, DxS, and FC resulted in a significant increase in deposition of fibronectin, collagen IV, collagen I, GFAP and CS56 in Neu7 astrocytes. In primary astrocytes, FC significantly enhanced the expression of astrocytic markers and increased the deposition of ECM proteins, including fibronectin and collagen IV. This study highlights the advantages of using FC as a MMC agent for enhancing ECM deposition in astrocytes. The method demonstrates potential for developing fast and more physiologically relevant in vitro models and improving drug screening processes for future studies. The observed benefits underscore the utility of FC in creating advanced cellular models that better mimic the native neural environment.

Neurons expressing Calb1 that encodes calbindin D-28K (Calb), a calcium-binding protein, are significant components of the sexually dimorphic nucleus (SDN) of the preoptic area in mice. Calb is therefore used as a marker to study the SDN. The number of Calb neurons in the SDN is greater in males than in females, and this sex difference emerges before puberty. However, the timing of emergence of this difference and the role of Calb1 in the SDN remain unclear. In this study, we investigated when the sex difference in Calb neurons appears and whether Calb1 is required for organizing the SDN. Profiling the temporal changes in Calb neurons revealed that these neurons dramatically increased in the SDN during postnatal days 15 to 20 in males but not in females, resulting in a significant difference between sexes. However, the Calb1 mRNA in the SDN remained unchanged during the late postnatal period and did not differ between sexes. Calb1 knockdown (KD) was performed by injecting an adeno-associated virus vector into the preoptic area of neonatal mice. The analysis of postnatal Calb1-KD mice revealed that Calb1 KD reduced not only Calb1 expression but also the number of neurons comprising the SDN. These findings suggest that compared with the female SDN, more Calb neurons migrate to and are incorporated into the male SDN, thereby yielding the sex difference in the number of Calb neurons at the late postnatal period, and that Calb1 plays a significant role in clustering neurons in the SDN.

Coronavirus-disease 2019 (COVID-19) affects the respiratory system with high morbidity in elderly and comorbid patients. Acute COVID-19 infection (CI) primarily leads to respiratory failure, long-term effects on respiratory skeletal muscle however remain vague. Thus, histopathological marker expression of oxidative stress, inflammation, satellite cell activity, myosin fiber composition, and cellular senescence were analyzed in intercostal muscle and diaphragm to compare respiratory muscle degeneration (RMD) in deceased CI-positive and control patients. Beside CI, the impact of BMI, age, sex, ventilation status, and duration of hospitalization on RMD were evaluated. CI-positive patients exhibited higher numbers of regenerative stem cells, but no association between CI status and RMD was observed. Interestingly, ventilation support and lung-associated comorbidities had no effect on expression of RMD markers (p > 0.05). However, intercostal muscle showed BMI-dependent changes in expression of RMD markers, regardless of the CI status, with increased cytokine expression (p = 0.04), reduced antioxidative capacity (p = 0.05), and low stem cell prevalence (p = 0.02) in patients with high BMI. Moreover, elderly patients demonstrated increased oxidative stress (p = 0.001) and cell senescence (p = 0.03) independent of CI status. Notably, immobility drives muscle fiber transformation to Myosin ST (p = 0.03), since prolonged hospitalization correlated with muscle fiber type shift. Limitations included incomplete retrospective data collection and absence of adequate samples for molecular analyses. Together, RMD is influenced by BMI, age and immobility rather than the CI status alone. Future studies including larger cohorts, molecular analyses, and evaluation of patient data in addition to CI status alone, will further support meaningful analyses and interpretation of RMD and its impact on post CI recovery.

Liquid-liquid phase separation (LLPS) is an emerging research field in cellular biology. LLPS-driven biomolecular condensates act as reaction chambers and regulatory hubs for critical processes, including chromatin architecture, gene expression, and metabolism. The dysregulation of these processes frequently impedes the proper execution of physiological functions. Current research indicates that abnormal phase separation plays a significant role in the pathogenesis of diseases and aging. This review briefly overviews the fundamental concepts and research methods related to phase separation. We also summarize studies concerning its physiological functions, particularly emphasizing its role in hematopoiesis. We further discuss how abnormal phase separation can lead to hematological disorders, specifically summarizing its involvement in the pathogenesis of leukemia. Despite recent advancements, elucidating LLPS mechanisms in hematopoiesis remains challenging due to the intricate interplay between biomolecular condensates and cellular function. Future research efforts aiming to reveal the role of LLPS in hematological diseases hold promise for novel therapeutic interventions and a deeper understanding of hematopoietic processes.

Our current understanding of brain organization in malacostracan crustaceans is strongly biased towards representatives of the Decapoda ("ten legged" crustaceans) such as crayfish, crabs, clawed lobsters and spiny lobsters. However, to understand aspects of brain evolution in crustaceans, a broader taxonomic sampling is essential. The peracarid crustaceans are a species-rich group that embraces representatives of, e.g. the Isopoda, Amphipoda and Mysida ("opossum shrimps"), taxa whose neuroanatomy has not been carefully examined. The current study sets out to analyze brain morphology of the mysid Neomysis integer (Leach, 1814; Peracarida, Mysida) using immunohistochemistry against the presynaptic protein synapsin and the neuropeptides RFamide, SIFamide and allatostatin combined with three-dimensional reconstruction of elements of the central olfactory pathway. Furthermore, we studied the inventory of sensilla on the first pair of antennae using cuticular autofluorescence. Anterograde filling with neuronal tracers allowed visualisation the central projections of the sensilla on the first pair of antennae. This species is known to display a sexual dimorphism in both the peripheral and central olfactory pathway. We focussed our analysis on this aspect because in contrast to Hexapoda, reports on a sexual dimorphism of the olfactory system are extremely rare in malacostracan crustaceans. We provide a detailed description of the sensilla associated with a male-specific structure on the pair of first antenna the "lobus masculinus". Furthermore, we analyzed the projection patterns of theses sensilla into the "male-specific neuropil" in the deutocerebrum and critically discuss our results in comparison to examples of sexual dimorphism in the chemosensory pathways in other malacostracan crustaceans and hexapods.

Mesenchymal stromal cells (MSC) are multipotent cells that can modulate immune cells, affecting macrophages, monocytes, and lymphocytes. Neutrophils are circulating leucocytes responsible for the first line of defense and can assume different phenotypes depending on their environment: N0, the naïve form, N1 (inflammatory), N2 (anti-inflammatory). This study explores the potentially protective roles of chorionic membrane MSCs and their products-conditioned medium and pre-conditioned cMSC-derived membrane microparticles (MP-cMSC)-on neutrophils. Conditioned medium treatment reduced the rate of apoptosis and enhanced the immunosuppressive potential consistent with an anti-inflammatory profile. MP-cMSC are a noteworthy cell-free therapy, consisting of artificially generated circular lipid bilayer structures with no cargo and approximately 200 nm in size. When added to neutrophil culture, MPs increased neutral red uptake, suggesting an enhanced phagocytic activity. In the MSC co-culture group, a reduced rate of apoptosis, increased neutral red uptake, and elevated programed death-ligand 1 (PD-L1) expression were observed. These findings suggest that the distinct effects elicited by conditioned media, microparticles, and co-culture are likely influenced by the specific nature of the interactions involved-whether purely paracrine, mediated through direct cell-to-cell contact, or a combination of both.