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Frontiers in Cell and Developmental... 2022The tumor microenvironment plays an important role in cancer development and the use of 3D systems that decouple different elements of this microenvironment is critical...
The tumor microenvironment plays an important role in cancer development and the use of 3D systems that decouple different elements of this microenvironment is critical for the study of cancer progression. In neuroblastoma (NB), vitronectin (VN), an extracellular matrix protein, has been linked to poor prognosis and appears as a promising therapeutic target. Here, we developed hydrogels that incorporate VN into 3D polyethylene glycol (PEG) hydrogel networks to recapitulate the native NB microenvironment. The stiffness of the VN/PEG hydrogels was modulated to be comparable to the values reported for NB tissue samples. We used SK-N-BE (2) NB cells to demonstrate that PEGylated VN promotes cell adhesion as the native protein does. Furthermore, the PEGylation of VN allows its crosslinking into the hydrogel network, providing VN retention within the hydrogels that support viable cells in 3D. Confocal imaging and ELISA assays indicate that cells secrete VN also in the hydrogels and continue to reorganize their 3D environment. Overall, the 3D VN-based PEG hydrogels recapitulate the complexity of the native tumor extracellular matrix, showing that VN-cell interaction plays a key role in NB aggressiveness, and that VN could potentially be targeted in preclinical drug studies performed on the presented hydrogels.
PubMed: 36425532
DOI: 10.3389/fcell.2022.988699 -
Bioengineering (Basel, Switzerland) Nov 2022Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) can be differentiated into cardiomyocytes (hESC-CMs and iPSC-CMs, respectively), which hold... (Review)
Review
Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) can be differentiated into cardiomyocytes (hESC-CMs and iPSC-CMs, respectively), which hold great promise for cardiac regenerative medicine and disease modeling efforts. However, the most widely employed differentiation protocols require undefined substrates that are derived from xenogeneic (animal) products, contaminating resultant hESC- and iPSC-CM cultures with xenogeneic proteins and limiting their clinical applicability. Additionally, typical hESC- and iPSC-CM protocols produce CMs that are significantly contaminated by non-CMs and that are immature, requiring lengthy maturation procedures. In this review, we will summarize recent studies that have investigated the ability of purified extracellular matrix (ECM) proteins to support hESC- and iPSC-CM differentiation, with a focus on commercially available ECM proteins and coatings to make such protocols widely available to researchers. The most promising of the substrates reviewed here include laminin-521 with laminin-221 together or Synthemax (a synthetic vitronectin-based peptide coating), which both resulted in highly pure CM cultures. Future efforts are needed to determine whether combinations of specific purified ECM proteins or derived peptides could further improve CM maturation and culture times, and significantly improve hESC- and iPSC-CM differentiation protocols.
PubMed: 36550926
DOI: 10.3390/bioengineering9120720 -
EJNMMI Research Jul 2020The urokinase plasminogen activator receptor (uPAR) plays a multifaceted role in almost any process where migration of cells and tissue-remodeling is involved such as... (Review)
Review
The urokinase plasminogen activator receptor (uPAR) plays a multifaceted role in almost any process where migration of cells and tissue-remodeling is involved such as inflammation, but also in diseases as arthritis and cancer. Normally, uPAR is absent in healthy tissues. By its carefully orchestrated interaction with the protease urokinase plasminogen activator and its inhibitor (plasminogen activator inhibitor-1), uPAR localizes a cascade of proteolytic activities, enabling (patho)physiologic cell migration. Moreover, via the interaction with a broad range of cell membrane proteins, like vitronectin and various integrins, uPAR plays a significant, but not yet completely understood, role in differentiation and proliferation of cells, affecting also disease progression. The implications of these processes, either for diagnostics or therapeutics, have received much attention in oncology, but only limited beyond. Nonetheless, the role of uPAR in different diseases provides ample opportunity to exploit new applications for targeting. Especially in the fields of oncology, cardiology, rheumatology, neurology, and infectious diseases, uPAR-targeted molecular imaging could offer insights for new directions in diagnosis, surveillance, or treatment options.
PubMed: 32725278
DOI: 10.1186/s13550-020-00673-7 -
Molecular and Cellular Biochemistry Mar 2021Integrins are a group of transmembrane glycoprotein receptors that are responsible for platelet activation through bidirectional signalling. These receptors have left... (Review)
Review
Integrins are a group of transmembrane glycoprotein receptors that are responsible for platelet activation through bidirectional signalling. These receptors have left their footprints in various cellular events and have intrigued many groups of scientists that have led to some significant discoveries. A lot of the recent understanding of haemostasis has been possible due to the integrins filling the gaps in between several cellular mechanism. Apart from this, other important functions carried out by integrins are growth and maturation of cardiomyocytes, mechano-transduction, and interaction with actin cytoskeleton. The signalling cascade for integrin activation involves certain intracellular interacting proteins, which initiates the step-by-step activation procedure through 'inside-out' signalling. The signalling cascade gets activated through 'outside-in' signalling with the involvement of agonists such as ADP, Fibronectin, Vitronectin, and so on. This is a crucial step for the downstream processes of platelet spreading, followed by aggregation, clot progression and finally thrombus formation. Researchers throughout the world have shown direct relation of integrins with CVD and cardiac remodelling. The present review aims to summarize the information available so far on the involvement of integrins in thrombosis and its relationship to DVT. This information could be a bedrock of hidden answers to several questions on pathogenesis of deep vein thrombosis.
Topics: Actin Cytoskeleton; Animals; Blood Platelets; Cytoskeletal Proteins; Cytoskeleton; Disease Models, Animal; Heart; Hemostasis; Integrins; Ligands; Mice; Mice, Transgenic; Myocardium; Platelet Aggregation; Platelet Glycoprotein GPIIb-IIIa Complex; Signal Transduction; Talin; Thrombosis; Venous Thrombosis; rap GTP-Binding Proteins
PubMed: 33398665
DOI: 10.1007/s11010-020-03961-x -
Frontiers in Cell and Developmental... 2022Urokinase-type plasminogen activator receptor (uPAR or CD87) is a glycosyl-phosphatidyl-inositol anchored (GPI) membrane protein. The uPAR primary ligand is the serine... (Review)
Review
Urokinase-type plasminogen activator receptor (uPAR or CD87) is a glycosyl-phosphatidyl-inositol anchored (GPI) membrane protein. The uPAR primary ligand is the serine protease urokinase (uPA), converting plasminogen into plasmin, a broad spectrum protease, active on most extracellular matrix components. Besides uPA, the uPAR binds specifically also to the matrix protein vitronectin and, therefore, is regarded also as an adhesion receptor. Complex formation of the uPAR with diverse transmembrane proteins, including integrins, formyl peptide receptors, G protein-coupled receptors and epidermal growth factor receptor results in intracellular signalling. Thus, the uPAR is a multifunctional receptor coordinating surface-associated pericellular proteolysis and signal transduction, thereby affecting physiological and pathological mechanisms. The uPAR-initiated signalling leads to remarkable cellular effects, that include increased cell migration, adhesion, survival, proliferation and invasion. Although this is beyond the scope of this review, the uPA/uPAR system is of great interest to cancer research, as it is associated to aggressive cancers and poor patient survival. Increasing evidence links the uPA/uPAR axis to epithelial to mesenchymal transition, a highly dynamic process, by which epithelial cells can convert into a mesenchymal phenotype. Furthermore, many reports indicate that the uPAR is involved in the maintenance of the stem-like phenotype and in the differentiation process of different cell types. Moreover, the levels of anchor-less, soluble form of uPAR, respond to a variety of inflammatory stimuli, including tumorigenesis and viral infections. Finally, the role of uPAR in virus infection has received increasing attention, in view of the Covid-19 pandemics and new information is becoming available. In this review, we provide a mechanistic perspective, via the detailed examination of consolidated and recent studies on the cellular responses to the multiple uPAR activities.
PubMed: 35493073
DOI: 10.3389/fcell.2022.818616 -
Drugs & Aging Feb 2022Over recent years, great attention has been paid to the role of the complement system in the pathogenesis of age-related macular degeneration (AMD). In particular,... (Review)
Review
Over recent years, great attention has been paid to the role of the complement system in the pathogenesis of age-related macular degeneration (AMD). In particular, several studies have highlighted a link between AMD development and complement dysregulation, which can probably be explained as a complement cascade hyperactivation resulting from the presence of a series of risk factors such as aging; smoking; obesity; alcohol consumption; exposure to pesticides, industrial chemicals, or pollution; and other causes of oxidative stress. This hypothesis has been mainly supported by the presence of complement mediators as constituents of drusen, representing one of the earliest and most characteristic signs of retinal damage in AMD. Additionally, activated complement mediators and some complement regulators, such as vitronectin, have been found not only in the drusen and adjacent retinal areas but also in the peripheral blood of patients with AMD. Therefore, we aim to provide a review of recently studied complement factors to highlight their role in the pathogenesis of AMD and to evaluate new potential therapeutic strategies.
Topics: Aging; Complement Activation; Complement System Proteins; Humans; Macular Degeneration; Oxidative Stress
PubMed: 35050489
DOI: 10.1007/s40266-021-00914-x -
Cells Dec 2021Epithelial to mesenchymal transition (EMT) occurs during the pathological process associated with tumor progression and is considered to influence and promote the... (Review)
Review
Epithelial to mesenchymal transition (EMT) occurs during the pathological process associated with tumor progression and is considered to influence and promote the metastatic cascade. Characterized by loss of cell adhesion and apex base polarity, EMT enhances cell motility and metastasis. The key markers of the epithelial to mesenchymal transition are proteins characteristic of the epithelial phenotype, e.g., E-cadherin, cytokeratins, occludin, or desmoplakin, the concentration and activity of which are reduced during this process. On the other hand, as a result of acquiring the characteristics of mesenchymal cells, an increased amount of N-cadherin, vimentin, fibronectin, or vitronectin is observed. Importantly, epithelial cells undergo partial EMT where some of the cells show both epithelial and mesenchymal characteristics. The significant influence of epigenetic regulatory mechanisms is observed in the gene expression involved in EMT. Among the epigenetic modifications accompanying incorrect genetic reprogramming in cancer are changes in the level of DNA methylation within the CpG islands and posttranslational covalent changes of histone proteins. All observed modifications, which are stable but reversible changes, affect the level of gene expression leading to the development and progression of the disease, and consequently affect the uncontrolled growth of the population of cancer cells.
Topics: CpG Islands; DNA Methylation; Epigenesis, Genetic; Epithelial-Mesenchymal Transition; Gene Expression Regulation, Neoplastic; Humans; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms
PubMed: 34943943
DOI: 10.3390/cells10123435 -
Journal of Stem Cells & Regenerative... 2022Coronary heart disease (CHD) is a leading cause of death globally, while its current management is limited to reducing the myocardial infarction area without actually...
Coronary heart disease (CHD) is a leading cause of death globally, while its current management is limited to reducing the myocardial infarction area without actually replacing dead cardiomyocytes. Direct cell reprogramming is a method of cellular cardiomyoplasty which aims for myocardial tissue regeneration, and CD34 cells are one of the potential sources due to their shared embryonic origin with cardiomyocytes. However, the isolation and culture of non-adherent CD34 cells is crucial to obtain adequate cells for high-efficiency genetic modification. This study aimed to investigate the optimal method for isolation and culture of CD34 peripheral blood cells using certain culture media. A peripheral blood sample was obtained from a healthy subject and underwent pre-enrichment, isolation, and expansion. The culture was subsequently observed for their viability, adherence, and confluence. Day 0 observation of the culture showed a healthy CD34 cell with a round cell shape, without any adherent cells present yet. Day 4 of observation showed that CD34 cells within the blood plasma medium became adherent, indicated by their transformations into spindle or oval morphologies. Meanwhile, CD34 cells in vitronectin and fibronectin media showed no adherent cells and many of them died. Day 7 observation revealed more adherent CD34 cells in blood plasma medium, and which had 75% of confluence. In conclusion, the CD34 cells that were isolated using a combination of density and magnetic methods may be viable and adequately adhere in culture using blood plasma medium, but not in cultures using fibronectin and vitronectin.
PubMed: 36003658
DOI: 10.46582/jsrm.1801004 -
FEBS Letters Aug 2020Pathogens that colonize deep tissues and spread systemically encounter the innate host resistance mechanism of complement-mediated lysis and complement opsonization... (Review)
Review
Pathogens that colonize deep tissues and spread systemically encounter the innate host resistance mechanism of complement-mediated lysis and complement opsonization leading to engulfment and degradation by phagocytic cells. Yersinia and Salmonella species have developed numerous strategies to block the antimicrobial effects of complement. These include recruitment of complement regulatory proteins factor H, C4BP, and vitronectin (Vn) as well as interference in late maturation events such as assembly of C9 into the membrane attack complex that leads to bacterial lysis. This review will discuss the contributions of various surface structures (proteins, lipopolysaccharide, and capsules) to evasion of complement-mediated immune clearance of the systemic pathogens Yersiniae and Salmonellae. Bacterial proteins required for recruitment of complement regulatory proteins will be described, including the details of their interaction with host regulatory proteins, where known. The potential role of the surface proteases Pla (Yersinia pestis) and PgtE (Salmonella species) on the activity of complement regulatory proteins will also be addressed. Finally, the implications of complement inactivation on host cell interactions and host cell targeting for type 3 secretion will be discussed.
Topics: Animals; Bacterial Proteins; Complement System Proteins; Humans; Immune Evasion; Plasminogen Activators; Salmonella; Type III Secretion Systems; Yersinia pestis
PubMed: 32170725
DOI: 10.1002/1873-3468.13771 -
Cells May 2022The pathogenesis of age-related macular degeneration (AMD), a frequent disorder of the central retina, is incompletely understood. Genome-wide association studies (GWAS)...
The pathogenesis of age-related macular degeneration (AMD), a frequent disorder of the central retina, is incompletely understood. Genome-wide association studies (GWAS) suggest a strong contribution of genomic variation in AMD susceptibility. Nevertheless, little is known about biological mechanisms of the disease. We reported previously that the AMD-associated polymorphism rs704C > T in the vitronectin (VTN) gene influences protein expression and functional aspects of encoded vitronectin, a human blood and extracellular matrix (ECM) protein. Here, we refined the association of rs704 with AMD in 16,144 cases and 17,832 controls and noted that rs704 is carried exclusively by the neovascular AMD subtype. Interaction studies demonstrate that rs704 affects the ability of vitronectin to bind the angiogenic regulator plasminogen activator inhibitor 1 (PAI-1) but has no influence on stabilizing its active state. Western blot analysis and confocal imaging reveal a strong enrichment of PAI-1 in the ECM of cultured endothelial cells and RPE cell line ARPE-19 exposed to vitronectin. Large-scale gene expression of VTN and PAI-1 showed positive correlations and a statistically significant increase in human retinal and blood tissues aged 60 years and older. Our results suggest a mechanism by which the AMD-associated rs704 variant in combination with ageing may contribute to the vascular complications in AMD.
Topics: Aged; Angiogenesis Inhibitors; Endothelial Cells; Genome-Wide Association Study; Humans; Macular Degeneration; Middle Aged; Plasminogen Activator Inhibitor 1; Vascular Endothelial Growth Factor A; Visual Acuity; Vitronectin
PubMed: 35681461
DOI: 10.3390/cells11111766