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1.
Cells ; 13(9)2024 Apr 26.
Article in English | MEDLINE | ID: mdl-38727288

ABSTRACT

Glioblastoma (GBM) is a devastating brain cancer for which new effective therapies are urgently needed. GBM, after an initial response to current treatment regimens, develops therapeutic resistance, leading to rapid patient demise. Cancer cells exhibit an inherent elevation of endoplasmic reticulum (ER) stress due to uncontrolled growth and an unfavorable microenvironment, including hypoxia and nutrient deprivation. Cancer cells utilize the unfolded protein response (UPR) to maintain ER homeostasis, and failure of this response promotes cell death. In this study, as integrins are upregulated in cancer, we have evaluated the therapeutic potential of individually targeting all αß1 integrin subunits using RNA interference. We found that GBM cells are uniquely susceptible to silencing of integrin α3. Knockdown of α3-induced proapoptotic markers such as PARP cleavage and caspase 3 and 8 activation. Remarkably, we discovered a non-canonical function for α3 in mediating the maturation of integrin ß1. In its absence, generation of full length ß1 was reduced, immature ß1 accumulated, and the cells underwent elevated ER stress with upregulation of death receptor 5 (DR5) expression. Targeting α3 sensitized TRAIL-resistant GBM cancer cells to TRAIL-mediated apoptosis and led to growth inhibition. Our findings offer key new insights into integrin α3's role in GBM survival via the regulation of ER homeostasis and its value as a therapeutic target.


Subject(s)
Apoptosis , Endoplasmic Reticulum Stress , Glioblastoma , Integrin alpha3 , Integrin beta1 , TNF-Related Apoptosis-Inducing Ligand , Humans , Glioblastoma/pathology , Glioblastoma/metabolism , Glioblastoma/genetics , Apoptosis/genetics , Cell Line, Tumor , Integrin beta1/metabolism , Integrin beta1/genetics , TNF-Related Apoptosis-Inducing Ligand/metabolism , TNF-Related Apoptosis-Inducing Ligand/pharmacology , Integrin alpha3/metabolism , Integrin alpha3/genetics , Brain Neoplasms/pathology , Brain Neoplasms/metabolism , Brain Neoplasms/genetics , Receptors, TNF-Related Apoptosis-Inducing Ligand/metabolism , Receptors, TNF-Related Apoptosis-Inducing Ligand/genetics
2.
JCI Insight ; 9(10)2024 May 22.
Article in English | MEDLINE | ID: mdl-38775153

ABSTRACT

Small cell lung cancer (SCLC) is the most aggressive lung cancer entity with an extremely limited therapeutic outcome. Most patients are diagnosed at an extensive stage. However, the molecular mechanisms driving SCLC invasion and metastasis remain largely elusive. We used an autochthonous SCLC mouse model and matched samples from patients with primary and metastatic SCLC to investigate the molecular characteristics of tumor metastasis. We demonstrate that tumor cell invasion and liver metastasis in SCLC are triggered by an Angiopoietin-2 (ANG-2)/Integrin ß-1-dependent pathway in tumor cells, mediated by focal adhesion kinase/Src kinase signaling. Strikingly, CRISPR-Cas9 KO of Integrin ß-1 or blocking Integrin ß-1 signaling by an anti-ANG-2 treatment abrogates liver metastasis formation in vivo. Interestingly, analysis of a unique collection of matched samples from patients with primary and metastatic SCLC confirmed a strong increase of Integrin ß-1 in liver metastasis in comparison with the primary tumor. We further show that ANG-2 blockade combined with PD-1-targeted by anti-PD-1 treatment displays synergistic treatment effects in SCLC. Together, our data demonstrate a fundamental role of ANG-2/Integrin ß-1 signaling in SCLC cells for tumor cell invasion and liver metastasis and provide a potentially new effective treatment strategy for patients with SCLC.


Subject(s)
Angiopoietin-2 , Integrin beta1 , Liver Neoplasms , Lung Neoplasms , Signal Transduction , Small Cell Lung Carcinoma , Animals , Angiopoietin-2/metabolism , Angiopoietin-2/genetics , Small Cell Lung Carcinoma/pathology , Small Cell Lung Carcinoma/metabolism , Small Cell Lung Carcinoma/genetics , Small Cell Lung Carcinoma/drug therapy , Mice , Lung Neoplasms/secondary , Lung Neoplasms/pathology , Lung Neoplasms/metabolism , Lung Neoplasms/genetics , Lung Neoplasms/drug therapy , Humans , Integrin beta1/metabolism , Integrin beta1/genetics , Liver Neoplasms/secondary , Liver Neoplasms/metabolism , Liver Neoplasms/pathology , Liver Neoplasms/genetics , Liver Neoplasms/drug therapy , Cell Line, Tumor , Neoplasm Invasiveness , Neoplasm Metastasis
3.
Cell Biol Toxicol ; 40(1): 32, 2024 May 20.
Article in English | MEDLINE | ID: mdl-38767703

ABSTRACT

BACKGROUND: Recent studies have emphasized the critical role of Telocytes (TCs)-derived exosomes in organ tissue injury and repair. Our previous research showed a significant increase in ITGB1 within TCs. Pulmonary Arterial Hypertension (PAH) is marked by a loss of microvessel regeneration and progressive vascular remodeling. This study aims to investigate whether exosomes derived from ITGB1-modified TCs (ITGB1-Exo) could mitigate PAH. METHODS: We analyzed differentially expressed microRNAs (DEmiRs) in TCs using Affymetrix Genechip miRNA 4.0 arrays. Exosomes isolated from TC culture supernatants were verified through transmission electron microscopy and Nanoparticle Tracking Analysis. The impact of miR-429-3p-enriched exosomes (Exo-ITGB1) on hypoxia-induced pulmonary arterial smooth muscle cells (PASMCs) was evaluated using CCK-8, transwell assay, and inflammatory factor analysis. A four-week hypoxia-induced mouse model of PAH was constructed, and H&E staining, along with Immunofluorescence staining, were employed to assess PAH progression. RESULTS: Forty-five miRNAs exhibited significant differential expression in TCs following ITGB1 knockdown. Mus-miR-429-3p, significantly upregulated in ITGB1-overexpressing TCs and in ITGB1-modified TC-derived exosomes, was selected for further investigation. Exo-ITGB1 notably inhibited the migration, proliferation, and inflammation of PASMCs by targeting Rac1. Overexpressing Rac1 partly counteracted Exo-ITGB1's effects. In vivo administration of Exo-ITGB1 effectively reduced pulmonary vascular remodeling and inflammation. CONCLUSIONS: Our findings reveal that ITGB1-modified TC-derived exosomes exert anti-inflammatory effects and reverse vascular remodeling through the miR-429-3p/Rac1 axis. This provides potential therapeutic strategies for PAH treatment.


Subject(s)
Exosomes , Integrin beta1 , MicroRNAs , Telocytes , rac1 GTP-Binding Protein , MicroRNAs/genetics , MicroRNAs/metabolism , Animals , Exosomes/metabolism , Exosomes/genetics , rac1 GTP-Binding Protein/metabolism , rac1 GTP-Binding Protein/genetics , Integrin beta1/metabolism , Integrin beta1/genetics , Mice , Telocytes/metabolism , Male , Myocytes, Smooth Muscle/metabolism , Myocytes, Smooth Muscle/pathology , Mice, Inbred C57BL , Pulmonary Arterial Hypertension/metabolism , Pulmonary Arterial Hypertension/genetics , Pulmonary Arterial Hypertension/pathology , Hypoxia/metabolism , Hypoxia/genetics , Hypoxia/complications , Cell Proliferation/genetics , Cell Movement/genetics , Humans , Vascular Remodeling/genetics , Neuropeptides
4.
J Cell Sci ; 137(10)2024 May 15.
Article in English | MEDLINE | ID: mdl-38813860

ABSTRACT

WW domain-containing transcription regulator 1 (WWTR1, referred to here as TAZ) and Yes-associated protein (YAP, also known as YAP1) are transcriptional co-activators traditionally studied together as a part of the Hippo pathway, and are best known for their roles in stem cell proliferation and differentiation. Despite their similarities, TAZ and YAP can exert divergent cellular effects by differentially interacting with other signaling pathways that regulate stem cell maintenance or differentiation. In this study, we show in mouse neural stem and progenitor cells (NPCs) that TAZ regulates astrocytic differentiation and maturation, and that TAZ mediates some, but not all, of the effects of bone morphogenetic protein (BMP) signaling on astrocytic development. By contrast, both TAZ and YAP mediate the effects on NPC fate of ß1-integrin (ITGB1) and integrin-linked kinase signaling, and these effects are dependent on extracellular matrix cues. These findings demonstrate that TAZ and YAP perform divergent functions in the regulation of astrocyte differentiation, where YAP regulates cell cycle states of astrocytic progenitors and TAZ regulates differentiation and maturation from astrocytic progenitors into astrocytes.


Subject(s)
Adaptor Proteins, Signal Transducing , Astrocytes , Cell Differentiation , Cell Proliferation , Neural Stem Cells , Signal Transduction , Trans-Activators , Transcriptional Coactivator with PDZ-Binding Motif Proteins , YAP-Signaling Proteins , Animals , Astrocytes/metabolism , Astrocytes/cytology , YAP-Signaling Proteins/metabolism , Adaptor Proteins, Signal Transducing/metabolism , Adaptor Proteins, Signal Transducing/genetics , Mice , Neural Stem Cells/metabolism , Neural Stem Cells/cytology , Transcriptional Coactivator with PDZ-Binding Motif Proteins/metabolism , Trans-Activators/metabolism , Trans-Activators/genetics , Phosphoproteins/metabolism , Phosphoproteins/genetics , Cell Cycle Proteins/metabolism , Cell Cycle Proteins/genetics , Integrin beta1/metabolism , Integrin beta1/genetics , Transcription Factors/metabolism , Transcription Factors/genetics , Bone Morphogenetic Proteins/metabolism , Cerebral Cortex/cytology , Cerebral Cortex/metabolism , Protein Serine-Threonine Kinases
5.
Int J Mol Sci ; 25(8)2024 Apr 12.
Article in English | MEDLINE | ID: mdl-38673859

ABSTRACT

The dynamic relationship between heart failure and cancer poses a dual challenge. While cardiac remodeling can promote cancer growth and metastasis, tumor development can ameliorate cardiac dysfunction and suppress fibrosis. However, the precise mechanism through which cancer influences the heart and fibrosis is yet to be uncovered. To further explore the interaction between heart failure and cancer, we used the MDX mouse model, which suffers from cardiac fibrosis and cardiac dysfunction. A previous study from our lab demonstrated that tumor growth improves cardiac dysfunction and dampens fibrosis in the heart and diaphragm muscles of MDX mice. We used breast Polyoma middle T (PyMT) and Lewis lung carcinoma (LLC) cancer cell lines that developed into large tumors. To explore whether the aggressiveness of the cancer cell line is crucial for the beneficial phenotype, we employed a PyMT breast cancer cell line lacking integrin ß1, representing a less aggressive cell line compared to the original PyMT cells. In addition, we examined immortalized and primary MEF cells. The injection of integrin ß1 KO PyMT cancer cells and Mouse Embryo Fibroblasts cells (MEF) resulted in the improvement of cardiac function and decreased fibrosis in the heart, diaphragm, and skeletal muscles of MDX mice. Collectively, our data demonstrate that the cancer line aggressiveness as well as primary MEF cells are sufficient to impose the beneficial phenotype. These discoveries present potential novel clinical therapeutic approaches with beneficial outcome for patients with fibrotic diseases and cardiac dysfunction that do not require tumor growth.


Subject(s)
Disease Models, Animal , Fibrosis , Mice, Inbred mdx , Muscular Dystrophy, Duchenne , Animals , Muscular Dystrophy, Duchenne/pathology , Muscular Dystrophy, Duchenne/metabolism , Mice , Cell Line, Tumor , Mice, Inbred C57BL , Female , Myocardium/pathology , Myocardium/metabolism , Integrin beta1/metabolism , Integrin beta1/genetics , Fibroblasts/metabolism , Fibroblasts/pathology , Humans
6.
Cancer Res ; 84(9): 1517-1533, 2024 May 02.
Article in English | MEDLINE | ID: mdl-38587552

ABSTRACT

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy characterized by an immunosuppressive tumor microenvironment enriched with cancer-associated fibroblasts (CAF). This study used a convergence approach to identify tumor cell and CAF interactions through the integration of single-cell data from human tumors with human organoid coculture experiments. Analysis of a comprehensive atlas of PDAC single-cell RNA sequencing data indicated that CAF density is associated with increased inflammation and epithelial-mesenchymal transition (EMT) in epithelial cells. Transfer learning using transcriptional data from patient-derived organoid and CAF cocultures provided in silico validation of CAF induction of inflammatory and EMT epithelial cell states. Further experimental validation in cocultures demonstrated integrin beta 1 (ITGB1) and vascular endothelial factor A (VEGFA) interactions with neuropilin-1 mediating CAF-epithelial cell cross-talk. Together, this study introduces transfer learning from human single-cell data to organoid coculture analyses for experimental validation of discoveries of cell-cell cross-talk and identifies fibroblast-mediated regulation of EMT and inflammation. SIGNIFICANCE: Adaptation of transfer learning to relate human single-cell RNA sequencing data to organoid-CAF cocultures facilitates discovery of human pancreatic cancer intercellular interactions and uncovers cross-talk between CAFs and tumor cells through VEGFA and ITGB1.


Subject(s)
Cancer-Associated Fibroblasts , Carcinoma, Pancreatic Ductal , Coculture Techniques , Epithelial-Mesenchymal Transition , Inflammation , Integrin beta1 , Pancreatic Neoplasms , Single-Cell Analysis , Tumor Microenvironment , Humans , Carcinoma, Pancreatic Ductal/pathology , Carcinoma, Pancreatic Ductal/metabolism , Carcinoma, Pancreatic Ductal/genetics , Cancer-Associated Fibroblasts/metabolism , Cancer-Associated Fibroblasts/pathology , Pancreatic Neoplasms/pathology , Pancreatic Neoplasms/metabolism , Pancreatic Neoplasms/genetics , Inflammation/pathology , Inflammation/metabolism , Integrin beta1/metabolism , Integrin beta1/genetics , Organoids/pathology , Organoids/metabolism , Vascular Endothelial Growth Factor A/metabolism , Vascular Endothelial Growth Factor A/genetics , Neuropilin-1/metabolism , Neuropilin-1/genetics , Gene Expression Regulation, Neoplastic , Cell Line, Tumor , Cell Communication
7.
Oncogene ; 43(21): 1620-1630, 2024 May.
Article in English | MEDLINE | ID: mdl-38570692

ABSTRACT

The role of the focal adhesion protein kindlin-3 as a tumor suppressor and its interaction mechanisms with extracellular matrix constitute a major field of investigation to better decipher tumor progression. Besides the well-described role of kindlin-3 in integrin activation, evidence regarding modulatory functions between melanoma cells and tumor microenvironment are lacking and data are needed to understand mechanisms driven by kindlin-3 inactivation. Here, we show that kindlin-3 inactivation through knockdown or somatic mutations increases BRAFV600mut melanoma cells oncogenic properties via collagen-related signaling by decreasing cell adhesion and enhancing proliferation and migration in vitro, and by promoting tumor growth in mice. Mechanistic analysis reveals that kindlin-3 interacts with the collagen-activated tyrosine kinase receptor DDR1 (Discoidin domain receptor 1) modulating its expression and its interaction with ß1-integrin. Kindlin-3 knockdown or mutational inactivation disrupt DDR1/ß1-integrin complex in vitro and in vivo and its loss improves the anti-proliferative effect of DDR1 inhibition. In agreement, kindlin-3 downregulation is associated with DDR1 over-expression in situ and linked to worse melanoma prognosis. Our study reveals a unique mechanism of action of kindlin-3 in the regulation of tumorigenesis mediated by the collagen-activated tyrosine kinase receptor DDR1 thus paving the way for innovative therapeutic targeting approaches in melanoma.


Subject(s)
Cell Proliferation , Discoidin Domain Receptor 1 , Melanoma , Membrane Proteins , Neoplasm Proteins , Humans , Discoidin Domain Receptor 1/genetics , Discoidin Domain Receptor 1/metabolism , Animals , Melanoma/pathology , Melanoma/genetics , Melanoma/metabolism , Mice , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Cell Proliferation/genetics , Membrane Proteins/genetics , Membrane Proteins/metabolism , Cell Line, Tumor , Integrin beta1/metabolism , Integrin beta1/genetics , Cell Movement/genetics , Cell Adhesion/genetics , Collagen/metabolism , Signal Transduction/genetics , Gene Expression Regulation, Neoplastic
8.
J Cell Sci ; 137(10)2024 May 15.
Article in English | MEDLINE | ID: mdl-38639242

ABSTRACT

WW domain-containing transcription regulator 1 (WWTR1, referred to here as TAZ) and Yes-associated protein (YAP, also known as YAP1) are transcriptional co-activators traditionally studied together as a part of the Hippo pathway, and are best known for their roles in stem cell proliferation and differentiation. Despite their similarities, TAZ and YAP can exert divergent cellular effects by differentially interacting with other signaling pathways that regulate stem cell maintenance or differentiation. In this study, we show in mouse neural stem and progenitor cells (NPCs) that TAZ regulates astrocytic differentiation and maturation, and that TAZ mediates some, but not all, of the effects of bone morphogenetic protein (BMP) signaling on astrocytic development. By contrast, both TAZ and YAP mediate the effects on NPC fate of ß1-integrin (ITGB1) and integrin-linked kinase signaling, and these effects are dependent on extracellular matrix cues. These findings demonstrate that TAZ and YAP perform divergent functions in the regulation of astrocyte differentiation, where YAP regulates cell cycle states of astrocytic progenitors and TAZ regulates differentiation and maturation from astrocytic progenitors into astrocytes.


Subject(s)
Adaptor Proteins, Signal Transducing , Astrocytes , Cell Differentiation , Cell Proliferation , Neural Stem Cells , Signal Transduction , Trans-Activators , Transcriptional Coactivator with PDZ-Binding Motif Proteins , YAP-Signaling Proteins , Animals , Astrocytes/metabolism , Astrocytes/cytology , YAP-Signaling Proteins/metabolism , Adaptor Proteins, Signal Transducing/metabolism , Adaptor Proteins, Signal Transducing/genetics , Mice , Neural Stem Cells/metabolism , Neural Stem Cells/cytology , Transcriptional Coactivator with PDZ-Binding Motif Proteins/metabolism , Trans-Activators/metabolism , Trans-Activators/genetics , Phosphoproteins/metabolism , Phosphoproteins/genetics , Cell Cycle Proteins/metabolism , Cell Cycle Proteins/genetics , Integrin beta1/metabolism , Integrin beta1/genetics , Transcription Factors/metabolism , Transcription Factors/genetics , Bone Morphogenetic Proteins/metabolism , Cerebral Cortex/cytology , Cerebral Cortex/metabolism , Protein Serine-Threonine Kinases
9.
J Biochem Mol Toxicol ; 38(4): e23688, 2024 Apr.
Article in English | MEDLINE | ID: mdl-38511888

ABSTRACT

In women, breast cancer (BC) accounts for 7%-10% of all cancer cases and is one of the most common cancers. To identify a new method for treating BC, the role of CD93 and its underlying mechanism were explored. MDA-MB-231 cells were used in this study and transfected with si-CD93, si-MMRN2, oe-CD93, si-integrin ß1, or oe-SP2 lentivirus. After MDA-MB-231 cells were transfected with si-NC or si-CD93, they were injected into nude mice by subcutaneous injection at a dose of 5 × 106/mouse to construct a BC animal model. The expression of genes and proteins and cell migration, invasion and vasculogenic mimicry were detected by RT‒qPCR, western blot, immunohistochemistry, immunofluorescence, Transwell, and angiogenesis assays. In pathological samples and BC cell lines, CD93 was highly expressed. Functionally, CD93 promoted the proliferation, migration, and vasculogenic mimicry of MDA-MB-231 cells. Moreover, CD93 interacts with MMRN2 and integrin ß1. Knockdown of CD93 and MMRN2 can inhibit the activation of integrin ß1, thereby inhibiting the PI3K/AKT/SP2 signaling pathway and inhibiting BC growth and vasculogenic mimicry. In conclusion, the binding of CD93 to MMRN2 can activate integrin ß1, thereby activating the PI3K/AKT/SP2 signaling pathway and subsequently promoting BC growth and vasculogenic mimicry.


Subject(s)
Breast Neoplasms , Integrin beta1 , Membrane Glycoproteins , Receptors, Complement , Animals , Female , Humans , Mice , Breast Neoplasms/pathology , Cell Line, Tumor , Cell Proliferation , Integrin beta1/genetics , Integrin beta1/metabolism , Mice, Nude , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction , Receptors, Complement/metabolism , Membrane Glycoproteins/metabolism
10.
Cell Rep ; 43(4): 113989, 2024 Apr 23.
Article in English | MEDLINE | ID: mdl-38536816

ABSTRACT

Attachment of circulating tumor cells to the endothelial cells (ECs) lining blood vessels is a critical step in cancer metastatic colonization, which leads to metastatic outgrowth. Breast and prostate cancers are common malignancies in women and men, respectively. Here, we observe that ß1-integrin is required for human prostate and breast cancer cell adhesion to ECs under shear-stress conditions in vitro and to lung blood vessel ECs in vivo. We identify IQGAP1 and neural Wiskott-Aldrich syndrome protein (NWASP) as regulators of ß1-integrin transcription and protein expression in prostate and breast cancer cells. IQGAP1 and NWASP depletion in cancer cells decreases adhesion to ECs in vitro and retention in the lung vasculature and metastatic lung nodule formation in vivo. Mechanistically, NWASP and IQGAP1 act downstream of Cdc42 to increase ß1-integrin expression both via extracellular signal-regulated kinase (ERK)/focal adhesion kinase signaling at the protein level and by myocardin-related transcription factor/serum response factor (SRF) transcriptionally. Our results identify IQGAP1 and NWASP as potential therapeutic targets to reduce early metastatic dissemination.


Subject(s)
Integrin beta1 , Neoplasm Metastasis , Serum Response Factor , ras GTPase-Activating Proteins , Humans , Integrin beta1/metabolism , Integrin beta1/genetics , ras GTPase-Activating Proteins/metabolism , ras GTPase-Activating Proteins/genetics , Cell Line, Tumor , Serum Response Factor/metabolism , Male , Female , Prostatic Neoplasms/pathology , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/genetics , Animals , Trans-Activators/metabolism , Cell Adhesion , Wiskott-Aldrich Syndrome Protein, Neuronal/metabolism , Wiskott-Aldrich Syndrome Protein, Neuronal/genetics , Breast Neoplasms/pathology , Breast Neoplasms/metabolism , Breast Neoplasms/genetics , Mice , Focal Adhesion Kinase 1/metabolism , Focal Adhesion Kinase 1/genetics , Gene Expression Regulation, Neoplastic , cdc42 GTP-Binding Protein/metabolism
11.
Nat Commun ; 15(1): 1622, 2024 Mar 04.
Article in English | MEDLINE | ID: mdl-38438343

ABSTRACT

Alveologenesis is a spatially coordinated morphogenetic event, during which alveolar myofibroblasts surround the terminal sacs constructed by epithelial cells and endothelial cells (ECs), then contract to form secondary septa to generate alveoli in the lungs. Recent studies have demonstrated the important role of alveolar ECs in this morphogenetic event. However, the mechanisms underlying EC-mediated alveologenesis remain unknown. Herein, we show that ECs regulate alveologenesis by constructing basement membranes (BMs) acting as a scaffold for myofibroblasts to induce septa formation through activating mechanical signaling. Rap1, a small GTPase of the Ras superfamily, is known to stimulate integrin-mediated cell adhesions. EC-specific Rap1-deficient (Rap1iECKO) mice exhibit impaired septa formation and hypo-alveolarization due to the decreased mechanical signaling in myofibroblasts. In Rap1iECKO mice, ECs fail to stimulate integrin ß1 to recruit Collagen type IV (Col-4) into BMs required for myofibroblast-mediated septa formation. Consistently, EC-specific integrin ß1-deficient mice show hypo-alveolarization, defective mechanical signaling in myofibroblasts, and disorganized BMs. These data demonstrate that alveolar ECs promote integrin ß1-mediated Col-4 recruitment in a Rap1-dependent manner, thereby constructing BMs acting as a scaffold for myofibroblasts to induce mechanical signal-mediated alveologenesis. Thus, this study unveils a mechanism of organ morphogenesis mediated by ECs through intrinsic functions.


Subject(s)
Endothelial Cells , Myofibroblasts , Animals , Mice , Basement Membrane , Integrin beta1/genetics , Morphogenesis
12.
Exp Cell Res ; 436(2): 113980, 2024 Mar 15.
Article in English | MEDLINE | ID: mdl-38401686

ABSTRACT

BACKGROUND: Hypospadias is a common congenital abnormality of the penile. Abnormal regulation of critical genes involved in urethral development leads to hypospadias. We used the Rab25-/- mice and foreskin fibroblasts transfected with lentivirus in vitro and in vivo to investigate the role of Rab25 in hypospadias. METHODS: The expression levels of various molecules in tissue samples and foreskin fibroblasts were confirmed using molecular biology methods (western blotting, PCR, immunohistochemistry, etc.). A scanning electron microscope (SEM) was used to visualize the external morphology of genital tubercles (GTs) of gestation day (GD) 18.5 male wild-type (WT) and Rab25-/- mice. RESULTS: An expanded distal cleft and V-shaped urethral opening were observed in GD 18.5 Rab25-/- mice. We demonstrated that Rab25 mediated hypospadias through the ß1 integrin/EGFR pathway. In addition, silencing Rab25 inhibited cell proliferation and migration and promoted apoptosis in the foreskin fibroblasts; Ki-67- and TUNEL-positive cells were mainly concentrated near the urethral seam. CONCLUSION: These findings suggest that Rab25 plays an essential role in hypospadias by activation of ß1 integrin/EGFR pathway, and Rab25 is a critical mediator of urethral seam formation in GD18.5 male fetal mice.


Subject(s)
Hypospadias , Humans , Male , Mice , Animals , Hypospadias/genetics , Hypospadias/metabolism , Integrin beta1/genetics , Integrin beta1/metabolism , Urethra/metabolism , Penis/metabolism , ErbB Receptors/metabolism , rab GTP-Binding Proteins/genetics
13.
J Transl Med ; 22(1): 154, 2024 Feb 14.
Article in English | MEDLINE | ID: mdl-38355577

ABSTRACT

BACKGROUND: Tumor cells of diffuse-type gastric cancer (DGC) are discohesive and infiltrate into the stroma as single cells or small subgroups, so the stroma significantly impacts DGC progression. Cancer-associated fibroblasts (CAFs) are major components of the tumor stroma. Here, we identified CAF-specific secreted molecules and investigated the mechanism underlying CAF-induced DGC progression. METHODS: We conducted transcriptome analysis for paired normal fibroblast (NF)-CAF isolated from DGC patient tissues and proteomics for conditioned media (CM) of fibroblasts. The effects of fibroblasts on cancer cells were examined by transwell migration and soft agar assays, western blotting, and in vivo. We confirmed the effect of blocking tubulointerstitial nephritis antigen-like 1 (TINAGL1) in CAFs using siRNA or shRNA. We evaluated the expression of TINAGL1 protein in frozen tissues of DGC and paired normal stomach and mRNA in formalin-fixed, paraffin-embedded (FFPE) tissue using RNA in-situ hybridization (RNA-ISH). RESULTS: CAFs more highly expressed TINAGL1 than NFs. The co-culture of CAFs increased migration and tumorigenesis of DGC. Moreover, CAFs enhanced the phosphorylation of focal adhesion kinase (FAK) and mesenchymal marker expression in DGC cells. In an animal study, DGC tumors co-injected with CAFs showed aggressive phenotypes, including lymph node metastasis. However, increased phosphorylation of FAK and migration were reduced by blocking TINAGL1 in CAFs. In the tissues of DGC patients, TINAGL1 was higher in cancer than paired normal tissues and detected with collagen type I alpha 1 chain (COL1A1) in the same spot. Furthermore, high TINAGL1 expression was significantly correlated with poor prognosis in several public databases and our patient cohort diagnosed with DGC. CONCLUSIONS: These results indicate that TINAGL1 secreted by CAFs induces phosphorylation of FAK in DGC cells and promotes tumor progression. Thus, targeting TINAGL1 in CAFs can be a novel therapeutic strategy for DGC.


Subject(s)
Cancer-Associated Fibroblasts , Nephritis, Interstitial , Stomach Neoplasms , Animals , Humans , Cancer-Associated Fibroblasts/metabolism , Cell Line, Tumor , Cell Movement/genetics , Fibroblasts/metabolism , Integrin beta1/genetics , Integrin beta1/metabolism , Nephritis, Interstitial/metabolism , Nephritis, Interstitial/pathology , RNA, Small Interfering/metabolism , Stomach Neoplasms/pathology , Tumor Microenvironment
14.
J Cell Mol Med ; 28(4): e18113, 2024 Feb.
Article in English | MEDLINE | ID: mdl-38332530

ABSTRACT

The resistance to anoikis plays a critical role in the metastatic progression of various types of malignancies, including gastric cancer (GC). Nevertheless, the precise mechanism behind anoikis resistance is not fully understood. Here, our primary focus was to examine the function and underlying molecular mechanism of Integrin beta-like 1 (ITGBL1) in the modulation of anoikis resistance and metastasis in GC. The findings of our investigation have demonstrated that the overexpression of ITGBL1 significantly augmented the resistance of GC cells to anoikis and promoted their metastatic potential, while knockdown of ITGBL1 had a suppressive effect on both cellular processes in vitro and in vivo. Mechanistically, we proved that ITGBL1 has a role in enhancing the resistance of GC cells to anoikis and promoting metastasis through the AKT/Fibulin-2 (FBLN2) axis. The inhibition of AKT/FBLN2 signalling was able to reverse the impact of ITGBL1 on the resistance of GC cells to anoikis and their metastatic capability. Moreover, the expression levels of ITGBL1 were found to be significantly elevated in the cancerous tissues of patients diagnosed with GC, and there was a strong correlation observed between high expression levels of ITGBL1 and worse prognosis among individuals diagnosed with GC. Significantly, it was revealed that within our cohort of GC patients, individuals exhibiting elevated ITGBL1 expression and diminished FBLN2 expression experienced the worst prognosis. In conclusion, the findings of our study indicate that ITGBL1 may serve as a possible modulator of resistance to anoikis and the metastatic process in GC.


Subject(s)
Anoikis , Calcium-Binding Proteins , Stomach Neoplasms , Humans , Anoikis/genetics , Stomach Neoplasms/pathology , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , Extracellular Matrix Proteins , Cell Line, Tumor , Neoplasm Metastasis , Integrin beta1/genetics
15.
Mol Cancer ; 23(1): 39, 2024 02 21.
Article in English | MEDLINE | ID: mdl-38378518

ABSTRACT

BACKGROUND: Focal adhesion signaling involving receptor tyrosine kinases (RTK) and integrins co-controls cancer cell survival and therapy resistance. However, co-dependencies between these receptors and therapeutically exploitable vulnerabilities remain largely elusive in HPV-negative head and neck squamous cell carcinoma (HNSCC). METHODS: The cytotoxic and radiochemosensitizing potential of targeting 10 RTK and ß1 integrin was determined in up to 20 3D matrix-grown HNSCC cell models followed by drug screening and patient-derived organoid validation. RNA sequencing and protein-based biochemical assays were performed for molecular characterization. Bioinformatically identified transcriptomic signatures were applied to patient cohorts. RESULTS: Fibroblast growth factor receptor (FGFR 1-4) targeting exhibited the strongest cytotoxic and radiosensitizing effects as monotherapy and combined with ß1 integrin inhibition, exceeding the efficacy of the other RTK studied. Pharmacological pan-FGFR inhibition elicited responses ranging from cytotoxicity/radiochemosensitization to resistance/radiation protection. RNA sequence analysis revealed a mesenchymal-to-epithelial transition (MET) in sensitive cell models, whereas resistant cell models exhibited a partial epithelial-to-mesenchymal transition (EMT). Accordingly, inhibition of EMT-associated kinases such as EGFR caused reduced adaptive resistance and enhanced (radio)sensitization to FGFR inhibition cell model- and organoid-dependently. Transferring the EMT-associated transcriptomic profiles to HNSCC patient cohorts not only demonstrated their prognostic value but also provided a conclusive validation of the presence of EGFR-related vulnerabilities that can be strategically exploited for therapeutic interventions. CONCLUSIONS: This study demonstrates that pan-FGFR inhibition elicits a beneficial radiochemosensitizing and a detrimental radioprotective potential in HNSCC cell models. Adaptive EMT-associated resistance appears to be of clinical importance, and we provide effective molecular approaches to exploit this therapeutically.


Subject(s)
Antineoplastic Agents , Carcinoma, Squamous Cell , Head and Neck Neoplasms , Humans , Squamous Cell Carcinoma of Head and Neck/drug therapy , Squamous Cell Carcinoma of Head and Neck/genetics , Carcinoma, Squamous Cell/drug therapy , Carcinoma, Squamous Cell/genetics , Carcinoma, Squamous Cell/pathology , Head and Neck Neoplasms/drug therapy , Head and Neck Neoplasms/genetics , Integrin beta1/genetics , Cell Line, Tumor , Receptor Protein-Tyrosine Kinases/genetics , Antineoplastic Agents/therapeutic use , ErbB Receptors/metabolism , Phenotype , Epithelial-Mesenchymal Transition/genetics
16.
Cell Rep ; 43(2): 113796, 2024 Feb 27.
Article in English | MEDLINE | ID: mdl-38367240

ABSTRACT

The acidic metabolic byproducts within the tumor microenvironment (TME) hinder T cell effector functions. However, their effects on T cell infiltration remain largely unexplored. Leveraging the comprehensive The Cancer Genome Atlas dataset, we pinpoint 16 genes that correlate with extracellular acidification and establish a metric known as the "tumor acidity (TuAci) score" for individual patients. We consistently observe a negative association between the TuAci score and T lymphocyte score (T score) across various human cancer types. Mechanistically, extracellular acidification significantly impedes T cell motility by suppressing podosome formation. This phenomenon can be attributed to the reduced expression of methyltransferase-like 3 (METTL3) and the modification of RNA N6-methyladenosine (m6A), resulting in a subsequent decrease in the expression of integrin ß1 (ITGB1). Importantly, enforced ITGB1 expression leads to enhanced T cell infiltration and improved antitumor activity. Our study suggests that modulating METTL3 activity or boosting ITGB1 expression could augment T cell infiltration within the acidic TME, thereby improving the efficacy of cell therapy.


Subject(s)
Integrin beta1 , Neoplasms , Humans , Cell- and Tissue-Based Therapy , Hydrogen-Ion Concentration , Integrin beta1/genetics , Methyltransferases/genetics , T-Lymphocytes , Tumor Microenvironment
17.
Exp Dermatol ; 33(1): e15006, 2024 Jan.
Article in English | MEDLINE | ID: mdl-38284200

ABSTRACT

Excessive ultraviolet B ray (UVB) exposure to sunlight results in skin photoageing. Our previous research showed that a Q-switched 1064 nm Nd: YAG laser can alleviate skin barrier damage through miR-24-3p. However, the role of autophagy in the laser treatment of skin photoageing is still unclear. This study aims to investigate whether autophagy is involved in the mechanism of Q-switched 1064 nm Nd: YAG in the treatment of skin ageing. In vitro, primary human dermal fibroblast (HDF) cells were irradiated with different doses of UVB to establish a cell model of skin photoageing. In vivo, SKH-1 hairless mice were irradiated with UVB to establish a skin photoageing mouse model and irradiated with laser. The oxidative stress and autophagy levels were detected by western blot, immunofluorescence and flow cytometer. String was used to predict the interaction protein of TGF-ß1, and CO-IP and GST-pull down were used to detect the binding relationship between TGFß1 and ITGB1. In vitro, UVB irradiation reduced HDF cell viability, arrested cell cycle, induced cell senescence and oxidative stress compared with the control group. Laser treatment reversed cell viability, senescence and oxidative stress induced by UVB irradiation and activated autophagy. Autophagy agonists or inhibitors can enhance or attenuate the changes induced by laser treatment, respectively. In vivo, UVB irradiation caused hyperkeratosis, dermis destruction, collagen fibres reduction, increased cellular senescence and activation of oxidative stress in hairless mice. Laser treatment thinned the stratum corneum of skin tissue, increased collagen synthesis and autophagy in the dermis, and decreased the level of oxidative stress. Autophagy agonist rapamycin and autophagy inhibitor 3-methyladenine (3-MA) can enhance or attenuate the effects of laser treatment on the skin, respectively. Also, we identified a direct interaction between TGFB1 and ITGB1 and participated in laser irradiation-activated autophagy, thereby inhibiting UVB-mediated oxidative stress further reducing skin ageing. Q-switched 1064 nm Nd: YAG laser treatment inhibited UVB-induced oxidative stress and restored skin photoageing by activating autophagy, and TGFß1 and ITGB1 directly incorporated and participated in this process.


Subject(s)
Integrin beta1 , Lasers, Solid-State , Skin Aging , Transforming Growth Factor beta1 , Animals , Humans , Mice , Autophagy , Collagen , Lasers, Solid-State/therapeutic use , Mice, Hairless , Transforming Growth Factor beta1/genetics , Integrin beta1/genetics
18.
BMC Cancer ; 24(1): 113, 2024 Jan 22.
Article in English | MEDLINE | ID: mdl-38254031

ABSTRACT

BACKGROUND: Extracellular vesicles (EVs) have been revealed to facilitate the development of oral squamous cavity cell carcinoma (OCSCC), while its supporting role in lymph node metastases is under continuous investigation. This study aimed to examine the function of cancer-associated fibroblasts (CAF)-derived EVs (CAF-EVs) during lymph node metastasis in OCSCC and the mechanisms. METHODS: CAF were isolated from OCSCC tissues of patients, and CAF-EVs were extracted and identified. EdU, colony formation, wound healing, and Transwell assays were performed. The OCSCC cells before and after CAF-EVs treatment were injected into mice to probe the effects of CAF-EVs on tumor growth and lymph node metastasis, respectively. The effect of CAF-EVs treatment on transcriptome changes in OCSCC cells was analyzed. Clinical data of patients with OCSCC were analyzed to determine the prognostic significance of the selected genes. Finally, loss-of-function assays were conducted to corroborate the involvement of polycomb complex protein BMI-1 (BMI1) and integrin beta1 (ITGB1). RESULTS: CAF-EVs promoted the malignant behavior of OCSCC cells and accelerated tumor growth and lymph node metastasis in mice. CAF-EVs significantly increased the expression of BMI1 and ITGB1, and the expression of BMI1 and ITGB1 was negatively correlated with the overall survival and relapse-free survival of OCSCC patients. Knockdown of BMI1 or ITGB1 in OCSCC cells abated the promoting effects of CAF-EVs in vitro and in vivo. CONCLUSION: CAF-EVs elicited the metastasis-promoting properties in OCSCC by elevating BMI1 and ITGB1, suggesting that BMI1 and ITGB1 could be potential biomarkers and therapeutic targets for OCSCC.


Subject(s)
Cancer-Associated Fibroblasts , Carcinoma, Squamous Cell , Extracellular Vesicles , Head and Neck Neoplasms , Mouth Neoplasms , Animals , Humans , Mice , Head and Neck Neoplasms/metabolism , Integrin beta1/genetics , Lymphatic Metastasis/genetics , Mouth Neoplasms/metabolism , Neoplasm Recurrence, Local , Polycomb Repressive Complex 1/genetics , Squamous Cell Carcinoma of Head and Neck/metabolism
19.
J Ethnopharmacol ; 323: 117699, 2024 Apr 06.
Article in English | MEDLINE | ID: mdl-38185262

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: Withania somnifera (L.) Dunal, known as Ashwagandha, has long been used in traditional medicine in Ayurveda, India, a representative adaptogen. The main active constituents of W. somnifera are withanolides, and the root is often used as a medicine with a wide range of pharmacological activities, which can be used to treat insomnia, neurasthenia, diabetes mellitus and skin cancer. AIM OF THE STUDY: Whole-component qualitative and quantitative analyses were performed on W. somnifera. We explored the ameliorative effect of the adaptogen representative plant W. somnifera on the senescence events of MGO-injured fibroblasts and its action mechanism and verified the hypotheses that WS can inhibit the accumulation of AGEs and regulate the dynamic balance among the components of the ECM by modulating the expression of integrin ß1 receptor; as a result, WS maintains cellular behavioural and biological functions in a normal range and retards the aging of skin from the cellular level. MATERIALS AND METHODS: In this study, the components of WS were first qualitatively and quantitatively analysed by HPLC fingerprinting and LC-MS detection. Second, a model of MGO-induced injury of CML-overexpressing fibroblasts was established. ELISA was used to detect CML expression and the synthesis of key extracellular matrix ECM protein components COL1, FN1, LM5 and TNC synthesis; CCK-8 was used to detect cell viability; EDU was used to detect cell proliferation capacity; fluorescence was used to detect cell adhesion capacity; and migration assay were used to detect cell migration capacity; qRT-PCR was used to detect the regulatory pathway TGF-ß1 and MMP-2, MMP-9 in ECMs; immunofluorescence was used to detect the expression of ITGB1; and WB was used to detect the expression of COL1, FN1, LM5, Tnc, TGF-ß1, MMP-2, MMP-9 and ITGB1. RESULTS: In total, 27 active ingredients were analysed from WS, which mainly consisted of withanolide components, such as withaferin A and withanolide A. Based on the model of MGO-induced fibroblast senescence injury, WS significantly inhibited CML synthesis. By up-regulating the expression of integrin ß1, it upregulated the expression of the TGF-ß1 gene, which is closely related to the generation of ECMs, downregulated the expression of the MMP-2 and MMP-9 genes, which are closely related to the degradation of ECMs, maintained the dynamic balance of the four types of ECMs, and improved cell viability as well as proliferation, migration and adhesion abilities. CONCLUSIONS: WS can prevent cellular behavioural dysfunction and delay skin ageing by reducing the accumulation of CML, upregulating the expression of the ITGB1 receptor, maintaining the normal function of ECM-integrin receptor interaction and preventing an imbalance between the production and degradation of protein components of ECMs. The findings reported in this study suggest that WS as a CML inhibitor can modulate ECM-integrin homeostasis and has great potential in the field of aging retardation.


Subject(s)
Withania , Withanolides , Transforming Growth Factor beta1/metabolism , Withania/metabolism , Integrin beta1/genetics , Integrin beta1/metabolism , Magnesium Oxide/metabolism , Matrix Metalloproteinase 2/metabolism , Matrix Metalloproteinase 9/metabolism , Integrins/metabolism , Withanolides/pharmacology , Withanolides/metabolism , Plant Extracts/pharmacology , Plant Extracts/metabolism , Fibroblasts/metabolism , Extracellular Matrix/metabolism , Plant Roots/chemistry
20.
J Exp Clin Cancer Res ; 43(1): 27, 2024 Jan 23.
Article in English | MEDLINE | ID: mdl-38254102

ABSTRACT

BACKGROUND: Peritoneal metastasis, which accounts for 85% of all epithelial ovarian carcinoma (EOC) metastases, is a multistep process that requires the establishment of adhesive interactions between cancer cells and the peritoneal membrane. Interrelations between EOC and the mesothelial stroma are critical to facilitate the metastatic process. No data is available so far on the impact of histone acetylation/deacetylation, a potentially relevant mechanism governing EOC metastasis, on mesothelial cells (MCs)-mediated adhesion. METHODS: Static adhesion and peritoneal clearance experiments were performed pretreating mesenchymal-like MCs and platinum-sensitive/resistant EOC cell lines with MS-275-a Histone deacetylase (HDAC)1-3 pharmacological inhibitor currently used in combination trials. Results were acquired by confocal microscopy and were analyzed with an automated Opera software. The role of HDAC1/2 was validated by genetic silencing. The role of α4-, α5-α1 Integrins and Fibronectin-1 was validated using specific monoclonal antibodies. Quantitative proteomic analysis was performed on primary MCs pretreated with MS-275. Decellularized matrices were generated from either MS-275-exposed or untreated cells to study Fibronectin-1 extracellular secretion. The effect of MS-275 on ß1 integrin activity was assessed using specific monoclonal antibodies. The role of Talin-1 in MCs/EOC adhesion was analyzed by genetic silencing. Talin-1 ectopic expression was validated as a rescue tool from MS-275-induced phenotype. The in vivo effect of MS-275-induced MC remodeling was validated in a mouse model of peritoneal EOC dissemination. RESULTS: Treatment of MCs with non-cytotoxic concentrations of MS-275 caused a consistent reduction of EOC adhesion. Proteomic analysis revealed several pathways altered upon MC treatment with MS-275, including ECM deposition/remodeling, adhesion receptors and actin cytoskeleton regulators. HDAC1/2 inhibition hampered actin cytoskeleton polymerization by downregulating actin regulators including Talin-1, impairing ß1 integrin activation, and leading to abnormal extracellular secretion and distribution of Fibronectin-1. Talin-1 ectopic expression rescued EOC adhesion to MS-275-treated MCs. In an experimental mouse model of metastatic EOC, MS-275 limited tumor invasion, Fibronectin-1 secretion and the sub-mesothelial accumulation of MC-derived carcinoma-associated fibroblasts. CONCLUSION: Our study unveils a direct impact of HDAC-1/2 in the regulation of MC/EOC adhesion and highlights the regulation of MC plasticity by epigenetic inhibition as a potential target for therapeutic intervention in EOC peritoneal metastasis.


Subject(s)
Benzamides , Carcinoma, Ovarian Epithelial , Cell Adhesion , Histone Deacetylase 1 , Histone Deacetylase 2 , Ovarian Neoplasms , Peritoneal Neoplasms , Animals , Female , Humans , Mice , Actin Cytoskeleton/metabolism , Antibodies, Monoclonal , Carcinoma, Ovarian Epithelial/metabolism , Epithelium , Extracellular Matrix Proteins/metabolism , Fibronectins , Histone Deacetylase 1/metabolism , Integrin alpha5 , Integrin beta1/genetics , Ovarian Neoplasms/genetics , Ovarian Neoplasms/metabolism , Peritoneal Neoplasms/genetics , Peritoneal Neoplasms/metabolism , Proteomics , Pyridines , Talin/genetics , Talin/metabolism , Histone Deacetylase 2/metabolism , Cell Adhesion/genetics
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