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Chinese Herbal Medicines Jan 2024This study is designed to investigate the mode of action of the synergistic effect of 5-fluorouracil (5-FU) and magnolol against cervical cancer.
OBJECTIVE
This study is designed to investigate the mode of action of the synergistic effect of 5-fluorouracil (5-FU) and magnolol against cervical cancer.
METHODS
Network pharmacological approach was applied to predict the molecular mechanism of 5-FU combined with magnolol against cervical cancer. CCK-8 assay, colony formation assay, immunofluorescence staining, adhesion assay, wound healing mobility assay, cell migration and invasion assay and Western blot analysis were conducted to validate the results of study.
RESULTS
Phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway was identified as the key pathway study. The experimental results showed that 5-FU combined with magnolol strongly inhibited cervical cancer cell proliferation, induced the morphological change of HeLa cells by down-regulating the expression of -actinin, tensin-2 and vinculin. Moreover, magnolol enhanced inhibitory effect of 5-FU on the cell adhesion, migration and invasion. The phosphorylation of AKT and PI3K and the expression of mTOR were strongly inhibited by the combination of 5-FU and magnolol. Moreover, the expression of E-cadherin and -catenin was upregulated and the expression of Snail, Slug and vimentin was down-regulated by the 5-FU together with magnolol.
CONCLUSION
Taken together, this study suggests that 5-FU combined with magnolol exerts a synergistic anti-cervical cancer effect by regulating the PI3K/AKT/mTOR and epithelial-mesenchymal transition (EMT) signaling pathways.
PubMed: 38375055
DOI: 10.1016/j.chmed.2023.01.004 -
ACS Applied Materials & Interfaces Feb 2024The complex interplay between cells and materials is a key focus of this research, aiming to develop optimal scaffolds for regenerative medicine. The need for tissue...
The complex interplay between cells and materials is a key focus of this research, aiming to develop optimal scaffolds for regenerative medicine. The need for tissue regeneration underscores understanding cellular behavior on scaffolds, especially cell adhesion to polymer fibers forming focal adhesions. Key proteins, paxillin and vinculin, regulate cell signaling, migration, and mechanotransduction in response to the extracellular environment. This study utilizes advanced microscopy, specifically the AiryScan technique, along with advanced image analysis employing the Density-Based Spatial Clustering of Applications with Noise (DBSCAN) cluster algorithm, to investigate protein distribution during osteoblast cell adhesion to polymer fibers and glass substrates. During cell attachment to both glass and polymer fibers, a noticeable shift in the local maxima of paxillin and vinculin signals is observed at the adhesion sites. The focal adhesion sites on polymer fibers are smaller and elliptical but exhibit higher protein density than on the typical glass surface. The characteristics of focal adhesions, influenced by paxillin and vinculin, such as size and density, can potentially reflect the strength and stability of cell adhesion. Efficient adhesion correlates with well-organized, larger focal adhesions characterized by increased accumulation of paxillin and vinculin. These findings offer promising implications for enhancing scaffold design, evaluating adhesion to various substrates, and refining cellular interactions in biomedical applications.
Topics: Paxillin; Vinculin; Focal Adhesions; Mechanotransduction, Cellular; Cell Adhesion; Polymers; Phosphoproteins; Focal Adhesion Protein-Tyrosine Kinases
PubMed: 38354103
DOI: 10.1021/acsami.3c19035 -
Biophysical Journal Dec 2023Transmission of cell-generated (i.e., endogenous) tension at cell-cell contacts is crucial for tissue shape changes during morphogenesis and adult tissue repair in...
Transmission of cell-generated (i.e., endogenous) tension at cell-cell contacts is crucial for tissue shape changes during morphogenesis and adult tissue repair in tissues such as epithelia. E-cadherin-based adhesions at cell-cell contacts are the primary means by which endogenous tension is transmitted between cells. The E-cadherin-β-catenin-α-catenin complex mechanically couples to the actin cytoskeleton (and thereby the cell's contractile machinery) both directly and indirectly. However, the key adhesion constituents required for substantial endogenous force transmission at these adhesions in cell-cell contacts are unclear. Due to the role of α-catenin as a mechanotransducer that recruits vinculin at cell-cell contacts, we expected α-catenin to be essential for sustaining normal levels of force transmission. Instead, using the traction force imbalance method to determine the inter-cellular force at a single cell-cell contact between cell pairs, we found that it is vinculin that is essential for sustaining normal levels of endogenous force transmission, with absence of vinculin decreasing the inter-cellular tension by over 50%. Our results constrain the potential mechanical pathways of force transmission at cell-cell contacts and suggest that vinculin can transmit forces at E-cadherin adhesions independent of α-catenin, possibly through β-catenin. Furthermore, we tested the ability of lateral cell-cell contacts to withstand external stretch and found that both vinculin and α-catenin are essential to maintain cell-cell contact stability under external forces.
Topics: alpha Catenin; Vinculin; beta Catenin; Cadherins; Cell Adhesion; Actins
PubMed: 38350000
DOI: 10.1016/j.bpj.2023.10.029 -
Materials Today. Bio Apr 2024Critical-size defects (CSDs) of the craniofacial bones cause aesthetic and functional complications that seriously impact the quality of life. The transplantation of...
Critical-size defects (CSDs) of the craniofacial bones cause aesthetic and functional complications that seriously impact the quality of life. The transplantation of human dental pulp stem cells (hDPSCs) is a promising strategy for bone tissue engineering. Chirality is commonly observed in natural biomolecules, yet its effect on stem cell differentiation is seldom studied, and little is known about the underlying mechanism. In this study, supramolecular chiral hydrogels were constructed using -phenylalanine (L/D-Phe) derivatives. The results of alkaline phosphatase expression analysis, alizarin red S assay, as well as quantitative real-time polymerase chain reaction and western blot analyses suggest that right-handed D-Phe hydrogel fibers significantly promoted osteogenic differentiation of hDPSCs. A rat model of calvarial defects was created to investigate the regulation of chiral nanofibers on the osteogenic differentiation of hDPSCs . The results of the animal experiment demonstrated that the D-Phe group exhibited greater and faster bone formation on hDPSCs. The results of RNA sequencing, vinculin immunofluorescence staining, a calcium fluorescence probe assay, and western blot analysis indicated that L-Phe significantly promoted adhesion of hDPSCs, while D-Phe nanofibers enhanced osteogenic differentiation of hDPSCs by facilitating calcium entry into cells and activate the MAPK pathway. These results of chirality-dependent osteogenic differentiation offer a novel therapeutic strategy for the treatment of CSDs by optimising the differentiation of hDPSCs into chiral nanofibers.
PubMed: 38347936
DOI: 10.1016/j.mtbio.2024.100971 -
Stem Cell Research & Therapy Feb 2024Pericytes (PCs) are multipotent contractile cells that wrap around the endothelial cells (ECs) to maintain the blood vessel's functionality and integrity. The...
BACKGROUND
Pericytes (PCs) are multipotent contractile cells that wrap around the endothelial cells (ECs) to maintain the blood vessel's functionality and integrity. The hyperglycemia associated with Type 2 diabetes mellitus (T2DM) was shown to impair the function of PCs and increase the risk of diabetes complications. In this study, we aimed to investigate the deleterious effect of the diabetic microenvironment on the regenerative capacities of human PCs.
METHODS
PCs isolated from human adipose tissue were cultured in the presence or absence of serum collected from diabetic patients. The functionality of PCs was analyzed after 6, 14, and 30 days.
RESULTS
Microscopic examination of PCs cultured in DS (DS-PCs) showed increased aggregate formation and altered surface topography with hyperbolic invaginations. Compared to PCs cultured in normal serum (NS-PCs), DS-PCs showed more fragmented mitochondria and thicker nuclear membrane. DS caused impaired angiogenic differentiation of PCs as confirmed by tube formation, decreased VEGF-A and IGF-1 gene expression, upregulated TSP1, PF4, actin-related protein 2/3 complex, and downregulated COL21A1 protein expression. These cells suffered more pronounced apoptosis and showed higher expression of Clic4, apoptosis facilitator BCl-2-like protein, serine/threonine protein phosphatase, and caspase-7 proteins. DS-PCs showed dysregulated DNA repair genes CDKN1A, SIRT1, XRCC5 TERF2, and upregulation of the pro-inflammatory genes ICAM1, IL-6, and TNF-α. Further, DS-treated cells also showed disruption in the expression of the focal adhesion and binding proteins TSP1, TGF-β, fibronectin, and PCDH7. Interestingly, DS-PCs showed resistance mechanisms upon exposure to diabetic microenvironment by maintaining the intracellular reactive oxygen species (ROS) level and upregulation of extracellular matrix (ECM) organizing proteins as vinculin, IQGAP1, and tubulin beta chain.
CONCLUSION
These data showed that the diabetic microenvironment exert a deleterious effect on the regenerative capacities of human adipose tissue-derived PCs, and may thus have possible implications on the vascular complications of T2DM. Nevertheless, PCs have shown remarkable protective mechanisms when initially exposed to DS and thus they could provide a promising cellular therapy for T2DM.
Topics: Humans; Diabetes Mellitus, Type 2; Pericytes; Endothelial Cells; Adipose Tissue; Apoptosis; Cells, Cultured
PubMed: 38331889
DOI: 10.1186/s13287-024-03643-1 -
Redox Biology Apr 2024Vascular diseases, a leading cause of death in human, are strongly associated with pathological damage to blood vessels. The selenoprotein (Sel) have been reported to...
Vascular diseases, a leading cause of death in human, are strongly associated with pathological damage to blood vessels. The selenoprotein (Sel) have been reported to play important roles in vascular disease. However, the role of SelO in vascular disease has not been conclusively investigated. The present experiment was to investigate the regulatory mechanism of the effect of SelO on the permeability of vascular endothelial. The H.E staining, FITC-Dextran staining, Dil-AC-LDL staining and FITC-WGA staining showed that vascular structure was damaged, and intercellular junctions were disrupted with selenium (Se)-deficient. Immunohistochemistry, qPCR and Western blot revealed decreased expression of the adhesion plaque proteins vinculin, talin and paxillin, decreased expression of the vascular connectivity effector molecules connexin, claudin-1 and E-cadherin and increased expression of JAM-A and N-cadherin, as well as decreased expression of the ZO-1 signaling pathways ZO-1, Rock, rhoGEF, cingulin and MLC-2. In a screening of 24 Sel present in mice, SelO showed the most pronounced changes in vascular tissues, and a possible association between SelO and vascular intercellular junction effectors was determined using IBM SPSS Statistics 25. Silencing of SelO, vascular endothelial intercellular junction adverse effects present. The regulatory relationship between SelO and vascular endothelial intercellular junctions was determined. The results showed that Se deficiency lead to increased vascular endothelial permeability and vascular tissue damage by decreasing SelO expression, suggesting a possible role for SelO in regulating vascular endothelial permeability.
Topics: Humans; Animals; Mice; Endothelial Cells; Selenium; Vascular Diseases; Permeability; Selenoproteins
PubMed: 38316067
DOI: 10.1016/j.redox.2024.103063 -
Experimental Cell Research Feb 2024The existing knowledge of the involvement of vinculin (VCL) in the control of ovarian cell functions is insufficient. To understand the role of VCL in the control of...
The existing knowledge of the involvement of vinculin (VCL) in the control of ovarian cell functions is insufficient. To understand the role of VCL in the control of basic porcine ovarian granulosa cell functions, we decreased VCL activity by small interfering RNA (VCL siRNA). The expression of VCL, accumulation of VCL protein, cell viability, proliferation (accumulation of PCNA and cyclin B1), proportion of proliferative active cells, apoptosis (accumulation of bax, caspase 3, p53, antiapoptotic marker bcl2, and bax/bcl-2 ratio), DNA fragmentation, and release of steroid hormones and IGF-I were analyzed by RT‒qPCR, Trypan blue exclusion test, quantitative immunocytochemistry, XTT assay, TUNEL assay, and ELISA. The suppression of VCL activity inhibited cell viability, the accumulation of the proliferation-related proteins PCNA and cyclin B1, the antiapoptotic protein bcl2, and the proportion of proliferative active cells. Moreover, VCL siRNA inhibited the release of progesterone, estradiol, and IGF-1. VCL siRNA increased the proportion of the proapoptotic proteins bax, caspase 3, p53, the proportion of DNA fragmented cells, and stimulated testosterone release. Taken together, the present study is the first evidence that inhibition of VCL suppresses porcine granulosa cell functions. Moreover, the results suggest that VCL can be a potent physiological stimulator of ovarian functions.
Topics: Female; Swine; Animals; Cyclin B1; Caspase 3; Proliferating Cell Nuclear Antigen; Tumor Suppressor Protein p53; bcl-2-Associated X Protein; Vinculin; Progesterone; Apoptosis; Proto-Oncogene Proteins c-bcl-2; Cell Proliferation; RNA, Small Interfering; Cells, Cultured; Insulin-Like Growth Factor I
PubMed: 38309674
DOI: 10.1016/j.yexcr.2024.113950 -
ELife Feb 2024Barrier functions of proliferative epithelia are constantly challenged by mechanical and chemical constraints. How epithelia respond to and cope with disturbances of...
Barrier functions of proliferative epithelia are constantly challenged by mechanical and chemical constraints. How epithelia respond to and cope with disturbances of barrier functions to allow tissue integrity maintenance is poorly characterised. Cellular junctions play an important role in this process and intracellular traffic contribute to their homeostasis. Here, we reveal that, in pupal , alteration of the bi- or tricellular septate junctions (SJs) triggers a mechanism with two prominent outcomes. On one hand, there is an increase in the levels of E-cadherin, F-actin, and non-muscle myosin II in the plane of adherens junctions. On the other hand, β-integrin/Vinculin-positive cell contacts are reinforced along the lateral and basal membranes. We found that the weakening of SJ integrity, caused by the depletion of bi- or tricellular SJ components, alters ESCRT-III/Vps32/Shrub distribution, reduces degradation and instead favours recycling of SJ components, an effect that extends to other recycled transmembrane protein cargoes including Crumbs, its effector β-Heavy Spectrin Karst, and β-integrin. We propose a mechanism by which epithelial cells, upon sensing alterations of the SJ, reroute the function of Shrub to adjust the balance of degradation/recycling of junctional cargoes and thereby compensate for barrier junction defects to maintain epithelial integrity.
Topics: Animals; Drosophila; Drosophila melanogaster; Drosophila Proteins; Epithelial Cells; Intercellular Junctions; Integrins
PubMed: 38305711
DOI: 10.7554/eLife.91246 -
BMC Medical Genomics Jan 2024Atherosclerosis (AS) is a pathology factor for cardiovascular diseases and instability of atherosclerotic plaques contributes to acute coronary events. This study...
BACKGROUND
Atherosclerosis (AS) is a pathology factor for cardiovascular diseases and instability of atherosclerotic plaques contributes to acute coronary events. This study identified a hub gene VCL for atherosclerotic plaques and discovered its potential therapeutic targets for atherosclerotic plaques.
METHODS
Differential expressed genes (DEGs) were screened between unstable and stable plaques from GSE120521 dataset and then used for construction of a protein-protein interactions (PPI) network. Through topological analysis, hub genes were identified within this PPI network, followed by construction of a diagnostic model. GSE41571 dataset was utilized to validate the diagnostic model. A key hub gene was identified and its association with immune characteristics and pathways were further investigated. Molecular docking and molecular dynamics (MD) simulation were employed to discover potential therapeutic targets.
RESULTS
According to the PPI network, 3 tightly connected protein clusters were found. Topological analysis identified the top 5 hub genes, Vinculin (VCL), Dystrophin (DMD), Actin alpha 2 (ACTA2), Filamin A (FLNA), and transgelin (TAGLN). Among these hub genes, VCL had the highest diagnostic value. VCL was selected for further analysis and we found that VCL was negatively correlated with immune score and AS-related inflammatory pathways. Next, we identified 408 genes that were highly correlated with VCL and determined potential drug candidates. The results from molecular docking and MD simulation showed compound DB07117 combined with VCL protein stably, the binding energy is -7.7 kcal/mol, indicating that compound DB07117 was a potential inhibitor of VCL protein.
CONCLUSION
This study identified VCL as a key gene for atherosclerotic plaques and provides a potential therapeutic target of VCL for the treatment of atherosclerotic plaques.
Topics: Humans; Molecular Docking Simulation; Plaque, Atherosclerotic; Vinculin; Protein Interaction Maps; Cardiovascular Diseases
PubMed: 38287421
DOI: 10.1186/s12920-024-01815-9 -
BioRxiv : the Preprint Server For... Jan 2024The ability of cells to sense and respond to mechanical forces is critical in many physiological and pathological processes. However, the mechanisms by which forces...
UNLABELLED
The ability of cells to sense and respond to mechanical forces is critical in many physiological and pathological processes. However, the mechanisms by which forces affect protein function inside cells remain unclear. Motivated by in vitro demonstrations of fluorescent proteins (FPs) undergoing reversible mechanical switching of fluorescence, we investigated if force-sensitive changes in FP function could be visualized in cells. Guided by a computational model of FP mechanical switching, we develop a formalism for its detection in Förster resonance energy transfer (FRET)-based biosensors and demonstrate its occurrence in a synthetic actin-crosslinker and the mechanical linker protein vinculin. We find that mechanical switching is reversible and altered by manipulation of cellular force generation as well as force-sensitive bond dynamics of the biosensor. Together, this work describes a new framework for assessing FP mechanical stability and provides a means of probing force-sensitive protein function inside cells.
MOTIVATION
The ability of cells to sense mechanical forces is critical in developmental, physiological, and pathological processes. Cells sense mechanical cues via force-induced alterations in protein structure and function, but elucidation of the molecular mechanisms is hindered by the lack of approaches to directly probe the effect of forces on protein structure and function inside cells. Motivated by in vitro observations of reversible fluorescent protein mechanical switching, we developed an approach for detecting fluorescent protein mechanical switching . This enables the visualization of force-sensitive protein function inside living cells.
PubMed: 38260589
DOI: 10.1101/2024.01.10.575065