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International Journal of Molecular... Feb 2023Estrogen-related receptor alpha (ERRα) plays an important role in endometrial cancer (EC) progression. However, the biological roles of ERRα in EC invasion and...
Estrogen-related receptor alpha (ERRα) plays an important role in endometrial cancer (EC) progression. However, the biological roles of ERRα in EC invasion and metastasis are not clear. This study aimed to investigate the role of ERRα and 3-hydroxy-3-methylglutaryl-CoA synthase 1 (HMGCS1) in regulating intracellular cholesterol metabolism to promote EC progression. ERRα and HMGCS1 interactions were detected by co-immunoprecipitation, and the effects of ERRα/HMGCS1 on the metastasis of EC were investigated by wound-healing and transwell chamber invasion assays. Cellular cholesterol content was measured to verify the relationship between ERRα and cellular cholesterol metabolism. Additionally, immunohistochemistry was performed to confirm that ERRα and HMGCS1 were related to EC progression. Furthermore, the mechanism was investigated using loss-of-function and gain-of-function assays or treatment with simvastatin. High expression levels of ERRα and HMGCS1 promoted intracellular cholesterol metabolism for invadopodia formation. Moreover, inhibiting ERRα and HMGCS1 expression significantly weakened the malignant progression of EC in vitro and in vivo. Our functional analysis showed that ERRα promoted EC invasion and metastasis through the HMGCS1-mediated intracellular cholesterol metabolism pathway, which was dependent on the epithelial-mesenchymal transition pathway. Our findings suggest that ERRα and HMGCS1 are potential targets to suppress EC progression.
Topics: Female; Humans; Cell Line, Tumor; Endometrial Neoplasms; Hydroxymethylglutaryl-CoA Synthase; Podosomes; Receptors, Estrogen; Neoplasm Invasiveness; Neoplasm Metastasis; ERRalpha Estrogen-Related Receptor
PubMed: 36835419
DOI: 10.3390/ijms24044010 -
Biomolecules Feb 2023The interface between the cellular actin network and diverse forms of integrin-mediated cell adhesions displays a unique capacity to serve as accurate chemical and... (Review)
Review
The interface between the cellular actin network and diverse forms of integrin-mediated cell adhesions displays a unique capacity to serve as accurate chemical and mechanical sensors of the cell's microenvironment. Focal adhesion-like structures of diverse cell types, podosomes in osteoclasts, and invadopodia of invading cancer cells display distinct morphologies and apparent functions. Yet, all three share a similar composition and mode of coupling between a protrusive structure (the lamellipodium, the core actin bundle of the podosome, and the invadopodia protrusion, respectively), and a nearby adhesion site. Cytoskeletal or external forces, applied to the adhesion sites, trigger a cascade of unfolding and activation of key adhesome components (e.g., talin, vinculin, integrin), which in turn, trigger the assembly of adhesion sites and generation of adhesion-mediated signals that affect cell behavior and fate. The structural and molecular mechanisms underlying the dynamic crosstalk between the actin cytoskeleton and the adhesome network are discussed.
Topics: Actins; Integrins; Cytoskeleton; Cell Adhesion; Actin Cytoskeleton
PubMed: 36830665
DOI: 10.3390/biom13020294 -
The Journal of Investigative Dermatology Aug 2023Signaling through the HGF receptor/Met in skin-resident Langerhans cells (LCs) and dermal dendritic cells (DCs) is essential for their emigration toward draining lymph...
Signaling through the HGF receptor/Met in skin-resident Langerhans cells (LCs) and dermal dendritic cells (DCs) is essential for their emigration toward draining lymph nodes upon inflammation-induced activation. In this study, we addressed the role of Met signaling in distinct steps of LC/dermal DC emigration from the skin by employing a conditionally Met-deficient mouse model (Met). We found that Met deficiency severely impaired podosome formation in DCs and concomitantly decreased the proteolytic degradation of gelatin. Accordingly, Met-deficient LCs failed to efficiently cross the extracellular matrix-rich basement membrane between the epidermis and the dermis. We further observed that HGF-dependent Met activation reduced the adhesion of bone marrow-derived LCs to various extracellular matrix factors and enhanced the motility of DCs in three-dimensional collagen matrices, which was not the case for Met-deficient LCs/DCs. We found no impact of Met signaling on the integrin-independent amoeboid migration of DCs in response to the CCR7 ligand CCL19. Collectively, our data show that the Met-signaling pathway regulates the migratory properties of DC in HGF-dependent and HGF-independent manners.
Topics: Mice; Animals; Podosomes; Cell Movement; Skin; Langerhans Cells; Signal Transduction; Dendritic Cells; Lymph Nodes
PubMed: 36813160
DOI: 10.1016/j.jid.2022.12.025 -
Free Radical Biology & Medicine Feb 2023Osteoclast differentiation and function are critical targets for anti-osteoporosis treatment. Oxidative stress also plays an important regulatory role in the...
Osteoclast differentiation and function are critical targets for anti-osteoporosis treatment. Oxidative stress also plays an important regulatory role in the differentiation of osteoclasts. Corylifol A (CA) is a flavonoid extracted from the Psoralea fruit. It has anti-inflammatory and antioxidant properties despite its unknown effect on osteoporosis. This study found that CA prevented estrogen-deficiency-induced bone loss and suppressed osteoclastogenesis in ovariectomized (OVX) mice by inhibiting intracellular reactive oxygen species (ROS) levels. In vivo, CA effectively prevented trabecular bone loss and reduced osteoclasts' number on the bone surface in OVX mice, as demonstrated in micro-CT, osteometry, and immunohistochemical data. However, CA did not affect cortical bone. In vitro, CA inhibited RANKL-induced podosome belt formation, osteoclastogenesis, and bone resorption functions. CA suppressed RANKL-induced ROS by boosting antioxidant enzymes (Catalase and NQO1) and NFATc1 signaling pathway related protein expression, including integrin αvβ3, NFATc1 and CTSK. Moreover, CA inhibited osteoclast-specific genes, including Ctsk, Acp5, and Mmp9. CA also attenuated the MAPK/ERK pathway, but did not affect the NF-κB signaling pathway. In terms of osteogenesis, CA did not inhibit or promote osteogenic differentiation and mineralization in vitro. These results reveal that CA could be a new replacement therapy for treating estrogen-deficiency osteoporosis via suppressing osteoclastogenesis and intracellular ROS.
Topics: Animals; Mice; Osteogenesis; Reactive Oxygen Species; Antioxidants; Osteoclasts; Bone Resorption; Osteoporosis; NF-kappa B; Estrogens; RANK Ligand; NFATC Transcription Factors; Cell Differentiation; Mice, Inbred C57BL
PubMed: 36649902
DOI: 10.1016/j.freeradbiomed.2023.01.017 -
The Journal of Biological Chemistry Feb 2023Osteoporosis is a chronic skeletal condition characterized by low bone mass and deteriorated microarchitecture of bone tissue and puts tens of millions of people at high...
Osteoporosis is a chronic skeletal condition characterized by low bone mass and deteriorated microarchitecture of bone tissue and puts tens of millions of people at high risk of fractures. New therapeutic agents like i-bodies, a class of next-generation single-domain antibodies, are needed to overcome some limitations of conventional treatments. An i-body is a human immunoglobulin scaffold with two long binding loops that mimic the shape and position of those found in shark antibodies, the variable new antigen receptors of sharks. Its small size (∼12 kDa) and long binding loops provide access to drug targets, which are considered undruggable by traditional monoclonal antibodies. Here, we have successfully identified a human receptor activator of nuclear factor-κB ligand (RANKL) i-body, ADR3, which demonstrates a high binding affinity to human RANKL (hRANKL) with no adverse effect on the survival or proliferation of bone marrow-derived macrophages. Differential scanning fluorimetry suggested that ADR3 is stable and able to tolerate a wide range of physical environments (including both temperature and pH). In addition, in vitro studies showed a dose-dependent inhibitory effect of ADR3 on osteoclast differentiation, podosome belt formation, and bone resorption activity. Further investigation on the mechanism of action of ADR3 revealed that it can inhibit hRANKL-mediated signaling pathways, supporting the in vitro functional observations. These clues collectively indicate that hRANKL antagonist ADR3 attenuates osteoclast differentiation and bone resorption, with the potential to serve as a novel therapeutic to protect against bone loss.
Topics: Humans; Bone Resorption; Cell Differentiation; Macrophages; Osteoclasts; RANK Ligand; Signal Transduction; Single-Domain Antibodies
PubMed: 36634847
DOI: 10.1016/j.jbc.2023.102889 -
International Journal of Molecular... Dec 2022Idiopathic pulmonary fibrosis (IPF) is characterized by abnormal fibroblast accumulation in the lung leading to extracellular matrix deposition and remodeling that...
Idiopathic pulmonary fibrosis (IPF) is characterized by abnormal fibroblast accumulation in the lung leading to extracellular matrix deposition and remodeling that compromise lung function. However, the mechanisms of interstitial invasion and remodeling by lung fibroblasts remain poorly understood. The invadosomes, initially described in cancer cells, consist of actin-based adhesive structures that coordinate with numerous other proteins to form a membrane protrusion capable of degrading the extracellular matrix to promote their invasive phenotype. In this regard, we hypothesized that invadosome formation may be increased in lung fibroblasts from patients with IPF. Public RNAseq datasets from control and IPF lung tissues were used to identify differentially expressed genes associated with invadosomes. Lung fibroblasts isolated from bleomycin-exposed mice and IPF patients were seeded with and without the two approved drugs for treating IPF, nintedanib or pirfenidone on fluorescent gelatin-coated coverslips for invadosome assays. Several matrix and invadosome-associated genes were increased in IPF tissues and in IPF fibroblastic foci. Invadosome formation was significantly increased in lung fibroblasts isolated from bleomycin-exposed mice and IPF patients. The degree of lung fibrosis found in IPF tissues correlated strongly with invadosome production by neighboring cells. Nintedanib suppressed IPF and PDGF-activated lung fibroblast invadosome formation, an event associated with inhibition of the PDGFR/PI3K/Akt pathway and TKS5 expression. Fibroblasts derived from IPF lung tissues express a pro-invadosomal phenotype, which correlates with the severity of fibrosis and is responsive to antifibrotic treatment.
Topics: Mice; Animals; Podosomes; Phosphatidylinositol 3-Kinases; Lung; Idiopathic Pulmonary Fibrosis; Fibroblasts; Fibrosis; Bleomycin
PubMed: 36613948
DOI: 10.3390/ijms24010499 -
The Journal of Cell Biology Feb 2023Invadopodia formation is regulated by Rho GTPases. However, the molecular mechanisms that control Rho GTPase signaling at invadopodia remain poorly understood. Here, we...
Invadopodia formation is regulated by Rho GTPases. However, the molecular mechanisms that control Rho GTPase signaling at invadopodia remain poorly understood. Here, we have identified ARHGAP17, a Cdc42-specific RhoGAP, as a key regulator of invadopodia in breast cancer cells and characterized a novel ARHGAP17-mediated signaling pathway that controls the spatiotemporal activity of Cdc42 during invadopodia turnover. Our results show that during invadopodia assembly, ARHGAP17 localizes to the invadopodia ring and restricts the activity of Cdc42 to the invadopodia core, where it promotes invadopodia growth. Invadopodia disassembly starts when ARHGAP17 translocates from the invadopodia ring to the core, in a process that is mediated by its interaction with the Cdc42 effector CIP4. Once at the core, ARHGAP17 inactivates Cdc42 to promote invadopodia disassembly. Our results in invadopodia provide new insights into the coordinated transition between the activation and inactivation of Rho GTPases.
Topics: cdc42 GTP-Binding Protein; Podosomes; rho GTP-Binding Proteins; Signal Transduction; Cell Line, Tumor; Humans; Breast Neoplasms; GTPase-Activating Proteins
PubMed: 36571786
DOI: 10.1083/jcb.202207020 -
Cells Dec 2022Tissue engineering strategies aim at characterizing and at optimizing the cellular component that is combined with biomaterials, for improved tissue regeneration. Here,...
Tissue engineering strategies aim at characterizing and at optimizing the cellular component that is combined with biomaterials, for improved tissue regeneration. Here, we present the immunoMap of apical papilla, the native tissue from which SCAPs are derived. We characterized stem cell niches that correspond to a minority population of cells expressing Mesenchymal stromal/Stem Cell (CD90, CD105, CD146) and stemness (SSEA4 and CD49f) markers as well as endothelial cell markers (VWF, CD31). Based on the colocalization of TKS5 and cortactin markers, we detected migration-associated organelles, podosomes-like structures, in specific regions and, for the first time, in association with stem cell niches in normal tissue. From six healthy teenager volunteers, each with two teeth, we derived twelve cell banks, isolated and amplified under 21 or 3% O. We confirmed a proliferative advantage of all banks when cultured under 3% versus 21% O. Interestingly, telomerase activity was similar to that of the highly proliferative hiPSC cell line, but unrelated to O concentration. Finally, SCAPs embedded in a thixotropic hydrogel and implanted subcutaneously in immunodeficient mice were protected from cell death with a slightly greater advantage for cells preconditioned at 3% O.
Topics: Animals; Mice; Cells, Cultured; Cell Differentiation; Stem Cells; Mesenchymal Stem Cells; Oxygen
PubMed: 36552862
DOI: 10.3390/cells11244098 -
International Journal of Molecular... Nov 2022TKS5 promotes invasion and migration through the formation of invadopodia in some tumour cells, and it also has an important physiological function in cell migration...
TKS5 promotes invasion and migration through the formation of invadopodia in some tumour cells, and it also has an important physiological function in cell migration through podosome formation in various nontumour cells. To date, the role of TKS5 in urothelial cells, and its potential role in BC initiation and progression, has not yet been addressed. Moreover, the contribution of TKS5 to ploidy control and chromosome stability has not been reported in previous studies. Therefore, in the present study, we wished to address the following questions: (i) Is TKS5 involved in the ploidy control of urothelial cells? (ii) What is the mechanism that leads to aneuploidy in response to TKS5 knockdown? (iii) Is TKS5 an oncogene or tumour-suppressor gene in the context of BC? (iv) Does TKS5 affect the proliferation, migration and invasion of BC cells? We assessed the gene and protein expressions via qPCR and Western blot analyses in a set of nontumour cell strains (Y235T, HBLAK and UROtsa) and a set of BC cell lines (RT4, T24, UMUC3 and J82). Following the shRNA knockdown in the TKS5-proficient cells and the ectopic TKS5 expression in the cell lines with low/absent TKS5 expression, we performed functional experiments, such as metaphase, invadopodia and gelatine degradation assays. Moreover, we determined the invasion and migration abilities of these genetically modified cells by using the Boyden chamber and wound-healing assays. The TKS5 expression was lower in the bladder cancer cell lines with higher invasive capacities (T24, UMUC3 and J82) compared to the nontumour cell lines from human ureter (Y235T, HBLAK and UROtsa) and the noninvasive BC cell line RT4. The reduced TKS5 expression in the Y235T cells resulted in augmented aneuploidy and impaired cell division. According to the Boyden chamber and wound-healing assays, TKS5 promotes the invasion and migration of bladder cancer cells. According to the present study, TKS5 regulates the migration and invasion processes of bladder cancer (BC) cell lines and plays an important role in genome stability.
Topics: Humans; Urinary Bladder Neoplasms; Urinary Bladder; Aneuploidy; Chromosomal Instability; Adaptor Proteins, Vesicular Transport
PubMed: 36430759
DOI: 10.3390/ijms232214283 -
IScience Dec 2022Podosomes, an important actin-based adhesive architecture, play critical roles in cell migration and matrix invasiveness. Here, we elucidate the ultrastructural...
Podosomes, an important actin-based adhesive architecture, play critical roles in cell migration and matrix invasiveness. Here, we elucidate the ultrastructural organization and regulation of podosome clusters in primary macrophages. With three-dimensional stochastic optical reconstruction microscopy (3D-STORM), we achieve ∼20/50 nm (lateral/axial) spatial resolution to resolve the mutual localization of podosome core and ring components, and further show that microtubules pass through podosomes at the layer of myosin IIA. The microtubule disruption-caused podosome dissolution is previously ascribed to Rho/ROCK-myosin signaling, yet inhibiting this pathway with Y27632 or blebbistatin only partially recovers podosome assembly, thus suggesting the contribution of the physical supporting of microtubules in stabilizing podosome structures. Through improved substrate-coating technique, we further corroborate that the matrix-degrading capability of macrophages depends on the formation of podosome clusters. Together, 3D-STORM super-resolution microscopy reveals the nanoscale spatial arrangement and the microtubule-dependent regulation of the matrix-degrading podosome clusters in macrophages.
PubMed: 36425766
DOI: 10.1016/j.isci.2022.105514