CD81 Controls Beige Fat Progenitor Cell Growth and Energy Balance via FAK Signaling.

Adipose tissues dynamically remodel their cellular composition in response to external cues by stimulating beige adipocyte biogenesis; however, the developmental origin and pathways regulating this process remain insufficiently understood owing to adipose tissue heterogeneity. Here, we employed single-cell RNA-seq and identified a unique subset of adipocyte progenitor cells (APCs) that possessed the cell-intrinsic plasticity to give rise to beige fat. This beige APC population is proliferative and marked by cell-surface proteins, including PDGFRα, Sca1, and CD81. Notably, CD81 is not only a beige APC marker but also required for de novo beige fat biogenesis following cold exposure. CD81 forms a complex with αV/ß1 and αV/ß5 integrins and mediates the activation of integrin-FAK signaling in response to irisin. Importantly, CD81 loss causes diet-induced obesity, insulin resistance, and adipose tissue inflammation. These results suggest that CD81 functions as a key sensor of external inputs and controls beige APC proliferation and whole-body energy homeostasis.

Fever Promotes T Lymphocyte Trafficking via a Thermal Sensory Pathway Involving Heat Shock Protein 90 and α4 Integrins.

Fever is an evolutionarily conserved response that confers survival benefits during infection. However, the underlying mechanism remains obscure. Here, we report that fever promoted T lymphocyte trafficking through heat shock protein 90 (Hsp90)-induced α4 integrin activation and signaling in T cells. By inducing selective binding of Hsp90 to α4 integrins, but not ß2 integrins, fever increased α4-integrin-mediated T cell adhesion and transmigration. Mechanistically, Hsp90 bound to the α4 tail and activated α4 integrins via inside-out signaling. Moreover, the N and C termini of one Hsp90 molecule simultaneously bound to two α4 tails, leading to dimerization and clustering of α4 integrins on the cell membrane and subsequent activation of the FAK-RhoA pathway. Abolishment of Hsp90-α4 interaction inhibited fever-induced T cell trafficking to draining lymph nodes and impaired the clearance of bacterial infection. Our findings identify the Hsp90-α4-integrin axis as a thermal sensory pathway that promotes T lymphocyte trafficking and enhances immune surveillance during infection.

Cell-matrix interface regulates dormancy in human colon cancer stem cells.

Cancer relapse after chemotherapy remains a main cause of cancer-related death. Although the relapse is thought to result from the propagation of resident cancer stem cells1, a lack of experimental platforms that enable the prospective analysis of cancer stem cell dynamics with sufficient spatiotemporal resolution has hindered the testing of this hypothesis. Here we develop a live genetic lineage-tracing system that allows the longitudinal tracking of individual cells in xenotransplanted human colorectal cancer organoids, and identify LGR5+ cancer stem cells that exhibit a dormant behaviour in a chemo-naive state. Dormant LGR5+ cells are marked by the expression of p27, and intravital imaging provides direct evidence of the persistence of LGR5+p27+ cells during chemotherapy, followed by clonal expansion. Transcriptome analysis reveals that COL17A1-a cell-adhesion molecule that strengthens hemidesmosomes-is upregulated in dormant LGR5+p27+ cells. Organoids in which COL17A1 is knocked out lose the dormant LGR5+p27+ subpopulation and become sensitive to chemotherapy, which suggests that the cell-matrix interface has a role in the maintenance of dormancy. Chemotherapy disrupts COL17A1 and breaks the dormancy in LGR5+p27+ cells through FAK-YAP activation. Abrogation of YAP signalling prevents chemoresistant cells from exiting dormancy and delays the regrowth of tumours, highlighting the therapeutic potential of YAP inhibition in preventing cancer relapse. These results offer a viable therapeutic approach to overcome the refractoriness of human colorectal cancer to conventional chemotherapy.

AMBRA1 levels predict resistance to MAPK inhibitors in melanoma.

Intrinsic and acquired resistance to mitogen-activated protein kinase inhibitors (MAPKi) in melanoma remains a major therapeutic challenge. Here, we show that the clinical development of resistance to MAPKi is associated with reduced tumor expression of the melanoma suppressor Autophagy and Beclin 1 Regulator 1 (AMBRA1) and that lower expression levels of AMBRA1 predict a poor response to MAPKi treatment. Functional analyses show that loss of AMBRA1 induces phenotype switching and orchestrates an extracellular signal-regulated kinase (ERK)-independent resistance mechanism by activating focal adhesion kinase 1 (FAK1). In both in vitro and in vivo settings, melanomas with low AMBRA1 expression exhibit intrinsic resistance to MAPKi therapy but higher sensitivity to FAK1 inhibition. Finally, we show that the rapid development of resistance in initially MAPKi-sensitive melanomas can be attributed to preexisting subclones characterized by low AMBRA1 expression and that cotreatment with MAPKi and FAK1 inhibitors (FAKi) effectively prevents the development of resistance in these tumors. In summary, our findings underscore the value of AMBRA1 expression for predicting melanoma response to MAPKi and supporting the therapeutic efficacy of FAKi to overcome MAPKi-induced resistance.

ORP2 couples LDL-cholesterol transport to FAK activation by endosomal cholesterol/PI(4,5)P2 exchange.

Low-density lipoprotein (LDL)-cholesterol delivery from late endosomes to the plasma membrane regulates focal adhesion dynamics and cell migration, but the mechanisms controlling it are poorly characterized. Here, we employed auxin-inducible rapid degradation of oxysterol-binding protein-related protein 2 (ORP2/OSBPL2) to show that endogenous ORP2 mediates the transfer of LDL-derived cholesterol from late endosomes to focal adhesion kinase (FAK)-/integrin-positive recycling endosomes in human cells. In vitro, cholesterol enhances membrane association of FAK to PI(4,5)P2 -containing lipid bilayers. In cells, ORP2 stimulates FAK activation and PI(4,5)P2 generation in endomembranes, enhancing cell adhesion. Moreover, ORP2 increases PI(4,5)P2 in NPC1-containing late endosomes in a FAK-dependent manner, controlling their tubulovesicular trafficking. Together, these results provide evidence that ORP2 controls FAK activation and LDL-cholesterol plasma membrane delivery by promoting bidirectional cholesterol/PI(4,5)P2 exchange between late and recycling endosomes.

Polarized focal adhesion kinase activity within a focal adhesion during cell migration.

Focal adhesion kinase (FAK) relays integrin signaling from outside to inside cells and contributes to cell adhesion and motility. However, the spatiotemporal dynamics of FAK activity in single FAs is unclear due to the lack of a robust FAK reporter, which limits our understanding of these essential biological processes. Here we have engineered a genetically encoded FAK activity sensor, dubbed FAK-separation of phases-based activity reporter of kinase (SPARK), which visualizes endogenous FAK activity in living cells and vertebrates. Our work reveals temporal dynamics of FAK activity during FA turnover. Most importantly, our study unveils polarized FAK activity at the distal tip of newly formed single FAs in the leading edge of a migrating cell. By combining FAK-SPARK with DNA tension probes, we show that tensions applied to FAs precede FAK activation and that FAK activity is proportional to the strength of tension. These results suggest tension-induced polarized FAK activity in single FAs, advancing the mechanistic understanding of cell migration.

Focal adhesion kinase regulates tendon cell mechanoresponse and physiological tendon development.

Tendons enable locomotion by transmitting high tensile mechanical forces between muscle and bone via their dense extracellular matrix (ECM). The application of extrinsic mechanical stimuli via muscle contraction is necessary to regulate healthy tendon function. Specifically, applied physiological levels of mechanical loading elicit an anabolic tendon cell response, while decreased mechanical loading evokes a degradative tendon state. Although the tendon response to mechanical stimuli has implications in disease pathogenesis and clinical treatment strategies, the cell signaling mechanisms by which tendon cells sense and respond to mechanical stimuli within the native tendon ECM remain largely unknown. Therefore, we explored the role of cell-ECM adhesions in regulating tendon cell mechanotransduction by perturbing the genetic expression and signaling activity of focal adhesion kinase (FAK) through both in vitro and in vivo approaches. We determined that FAK regulates tendon cell spreading behavior and focal adhesion morphology, nuclear deformation in response to applied mechanical strain, and mechanosensitive gene expression. In addition, our data reveal that FAK signaling plays an essential role in in vivo tendon development and postnatal growth, as FAK-knockout mouse tendons demonstrated reduced tendon size, altered mechanical properties, differences in cellular composition, and reduced maturity of the deposited ECM. These data provide a foundational understanding of the role of FAK signaling as a critical regulator of in situ tendon cell mechanotransduction. Importantly, an increased understanding of tendon cell mechanotransductive mechanisms may inform clinical practice as well as lead to the discovery of diagnostic and/or therapeutic molecular targets.

FOXS1 acts as an oncogene and induces EMT through FAK/PI3K/AKT pathway by upregulating HILPDA in prostate cancer.

Prostate cancer (PCa) is a widespread global health concern characterized by elevated rates of occurrence, and there is a need for novel therapeutic targets to enhance patient outcomes. FOXS1 is closely linked to different cancers, but its function in PCa is still unknown. The expression of FOXS1, its prognostic role, clinical significance in PCa, and the potential mechanism by which FOXS1 affects PCa progression were investigated through bioinformatics analysis utilizing public data. The levels of FOXS1 and HILPDA were evaluated in clinical PCa samples using various methods, such as western blotting, immunohistochemistry, and qRT-PCR. To examine the function and molecular mechanisms of FOXS1 in PCa, a combination of experimental techniques including CCK-8 assay, flow cytometry, wound-healing assay, Transwell assay, and Co-IP assay were employed. The FOXS1 expression levels were significantly raised in PCa, correlating strongly with tumor aggressiveness and an unfavorable prognosis. Regulating FOXS1 expression, whether upregulating or downregulating it, correspondingly enhanced or inhibited the growth, migration, and invasion capabilities of PCa cells. Mechanistically, we detected a direct interaction between FOXS1 and HILPDA, resulting in the pathway activation of FAK/PI3K/AKT and facilitation EMT in PCa cells. FOXS1 collaborates with HILPDA to initiate EMT, thereby facilitating the PCa progression through the FAK/PI3K/AKT pathway activation.

Focal Adhesion- and IGF1R-Dependent Survival and Migratory Pathways Mediate Tumor Resistance to mTORC1/2 Inhibition.

Aberrant signaling by the mammalian target of rapamycin (mTOR) contributes to the devastating features of cancer cells. Thus, mTOR is a critical therapeutic target and catalytic inhibitors are being investigated as anti-cancer drugs. Although mTOR inhibitors initially block cell proliferation, cell viability and migration in some cancer cells are quickly restored. Despite sustained inhibition of mTORC1/2 signaling, Akt, a kinase regulating cell survival and migration, regains phosphorylation at its regulatory sites. Mechanistically, mTORC1/2 inhibition promotes reorganization of integrin/focal adhesion kinase-mediated adhesomes, induction of IGFR/IR-dependent PI3K activation, and Akt phosphorylation via an integrin/FAK/IGFR-dependent process. This resistance mechanism contributes to xenograft tumor cell growth, which is prevented with mTOR plus IGFR inhibitors, supporting this combination as a therapeutic approach for cancers.

DCAF13 promotes ovarian cancer progression by activating FRAS1-mediated FAK signaling pathway.

Cullin-RING ubiquitin ligase 4 (CRL4) is closely correlated with the incidence and progression of ovarian cancer. DDB1- and CUL4-associated factor 13 (DCAF13), a substrate-recognition protein in the CRL4 E3 ubiquitin ligase complex, is involved in the occurrence and development of ovarian cancer. However, its precise function and the underlying molecular mechanism in this disease remain unclear. In this study, we confirmed that DCAF13 is highly expressed in human ovarian cancer and its expression is negatively correlated with the overall survival rate of patients with ovarian cancer. We then used CRISPR/Cas9 to knockout DCAF13 and found that its deletion significantly inhibited the proliferation, colony formation, and migration of human ovarian cancer cells. In addition, DCAF13 deficiency inhibited tumor proliferation in nude mice. Mechanistically, CRL4-DCAF13 targeted Fraser extracellular matrix complex subunit 1 (FRAS1) for polyubiquitination and proteasomal degradation. FRAS1 influenced the proliferation and migration of ovarian cancer cell through induction of the focal adhesion kinase (FAK) signaling pathway. These findings collectively show that DCAF13 is an important oncogene that promotes tumorigenesis in ovarian cancer cells by mediating FRAS1/FAK signaling. Our findings provide a foundation for the development of targeted therapeutics for ovarian cancer.

Focal-adhesion kinase regulates the sialylation of N-glycans via the PI4KIIα-PI4P pathway.

Sialylation is a terminal glycosylated modification of glycoproteins that regulates critical biological events such as cell adhesion and immune response. Our previous study showed that integrin α3ß1 plays a crucial role in regulating the sialylation of N-glycans. However, the underlying mechanism for the regulation remains unclear. This study investigated how sialylation is affected by focal adhesion kinase (FAK), which is a critical downstream signal molecule of integrin ß1. We established a stable FAK knockout (KO) cell line using the CRISPR/Cas9 system in HeLa cells. The results obtained from lectin blot, flow cytometric analysis, and MS showed that the sialylation levels were significantly decreased in the KO cells compared with that in wild-type (WT) cells. Moreover, phosphatidylinositol 4-phosphate (PI4P) expression levels were also reduced in the KO cells due to a decrease in the stability of phosphatidylinositol 4-kinase-IIα (PI4KIIα). Notably, the decreased levels of sialylation, PI4P, and the complex formation between GOLPH3 and ST3GAL4 or ST6GAL1, which are the main sialyltransferases for modification of N-glycans, were significantly restored by the re-expression of FAK. Furthermore, the decreased sialylation and phosphorylation of Akt and cell migration caused by FAK deficiency all were restored by overexpressing PI4KIIα, which suggests that PI4KIIα is one of the downstream molecules of FAK. These findings indicate that FAK regulates sialylation via the PI4P synthesis pathway and a novel mechanism is suggested for the integrin-FAK-PI4KIIα-GOLPH3-ST axis modulation of sialylation in N-glycans.

Plant phenolics inhibit focal adhesion kinase and suppress host cell invasion by uropathogenic Escherichia coli.

Traditional folk treatments for the prevention and management of urinary tract infections (UTIs) and other infectious diseases often include plants and plant extracts that are rich in phenolic compounds. These have been ascribed a variety of activities, including inhibition of bacterial interactions with host cells. Here, we tested a panel of four well-studied phenolic compounds-caffeic acid phenethyl ester (CAPE), resveratrol, catechin, and epigallocatechin gallate-for the effects on host cell adherence and invasion by uropathogenic Escherichia coli (UPEC). These bacteria, which are the leading cause of UTIs, can bind and subsequently invade bladder epithelial cells via an actin-dependent process. Intracellular UPEC reservoirs within the bladder are often protected from antibiotics and host defenses and likely contribute to the development of chronic and recurrent infections. In cell culture-based assays, only resveratrol had a notable negative effect on UPEC adherence to bladder cells. However, both CAPE and resveratrol significantly inhibited UPEC entry into the host cells, coordinate with attenuated phosphorylation of the host actin regulator Focal Adhesion Kinase (FAK or PTK2) and marked increases in the numbers of focal adhesion structures. We further show that the intravesical delivery of resveratrol inhibits UPEC infiltration of the bladder mucosa in a murine UTI model and that resveratrol and CAPE can disrupt the ability of other invasive pathogens to enter host cells. Together, these results highlight the therapeutic potential of molecules like CAPE and resveratrol, which could be used to augment antibiotic treatments by restricting pathogen access to protective intracellular niches.IMPORTANCEUrinary tract infections (UTIs) are exceptionally common and increasingly difficult to treat due to the ongoing rise and spread of antibiotic-resistant pathogens. Furthermore, the primary cause of UTIs, uropathogenic Escherichia coli (UPEC), can avoid antibiotic exposure and many host defenses by invading the epithelial cells that line the bladder surface. Here, we identified two plant-derived phenolic compounds that disrupt activation of the host machinery needed for UPEC entry into bladder cells. One of these compounds, resveratrol, effectively inhibited UPEC invasion of the bladder mucosa in a mouse UTI model, and both phenolic compounds significantly reduced host cell entry by other invasive pathogens. These findings suggest that select phenolic compounds could be used to supplement existing antibacterial therapeutics by denying uropathogens shelter within host cells and tissues and help explain some of the benefits attributed to traditional plant-based medicines.

Epithelial-mesenchymal transition status is a remarkable biomarker for the combination treatment with avutometinib and defactinib in KRAS-mutated non-small cell lung cancer.

BACKGROUND: Recent therapeutic strategies for KRAS-mutated cancers that inhibit the MAPK pathway have attracted considerable attention. The RAF/MEK clamp avutometinib (VS-6766/CH5126766/RO5126766/CKI27) is promising for patients with KRAS-mutated cancers. Although avutometinib monotherapy has shown clinical activity in patients with KRAS-mutated cancers, effective combination strategies will be important to develop. METHODS: Using a phosphorylation kinase array kit, we explored the feedback mechanism of avutometinib in KRAS-mutated NSCLC cells, and investigated the efficacy of combining avutometinib with inhibitors of the feedback signal using in vitro and in vivo experiments. Moreover, we searched for a biomarker for the efficacy of combination therapy through an in vitro study and analysis using the The Cancer Genome Atlas Programme dataset. RESULTS: Focal adhesion kinase (FAK) phosphorylation/activation was increased after avutometinib treatment and synergy between avutometinib and FAK inhibitor, defactinib, was observed in KRAS-mutated NSCLC cells with an epithelial rather than mesenchymal phenotype. Combination therapy with avutometinib and defactinib induced apoptosis with upregulation of Bim in cancer cells with an epithelial phenotype in an in vitro and in vivo study. CONCLUSIONS: These results demonstrate that the epithelial-mesenchymal transition status may be a promising biomarker for the efficacy of combination therapy with avutometinib and defactinib in KRAS-mutated NSCLC.

IGFBP2 induces podocyte apoptosis promoted by mitochondrial damage via integrin α5/FAK in diabetic kidney disease.

Podocyte apoptosis or loss is the pivotal pathological characteristic of diabetic kidney disease (DKD). Insulin-like growth factor-binding protein 2 (IGFBP2) have a proinflammatory and proapoptotic effect on diseases. Previous studies have shown that serum IGFBP2 level significantly increased in DKD patients, but the precise mechanisms remain unclear. Here, we found that IGFBP2 levels obviously increased under a diabetic state and high glucose stimuli. Deficiency of IGFBP2 attenuated the urine protein, renal pathological injury and glomeruli hypertrophy of DKD mice induced by STZ, and knockdown or deletion of IGFBP2 alleviated podocytes apoptosis induced by high concentration of glucose or in DKD mouse. Furthermore, IGFBP2 facilitated apoptosis, which was characterized by increase in inflammation and oxidative stress, by binding with integrin α5 (ITGA5) of podocytes, and then activating the phosphorylation of focal adhesion kinase (FAK)-mediated mitochondrial injury, including membrane potential decreasing, ROS production increasing. Moreover, ITGA5 knockdown or FAK inhibition attenuated the podocyte apoptosis caused by high glucose or IGFBP2 overexpression. Taken together, these findings unveiled the insight mechanism that IGFBP2 increased podocyte apoptosis by mitochondrial injury via ITGA5/FAK phosphorylation pathway in DKD progression, and provided the potential therapeutic strategies for diabetic kidney disease.

New insights into FAK structure and function in focal adhesions.

Focal adhesion kinase (FAK; also known as PTK2) was discovered three decades ago and is now recognised as a key player in the regulation of cell-matrix adhesion and mesenchymal cell migration. Although it is essential during development, FAK also drives invasive cancer progression and metastasis. On a structural level, the basic building blocks of FAK have been described for some time. However, a picture of how FAK integrates into larger assemblies in various cellular environments, including one of its main cellular locations, the focal adhesion (FA) complex, is only beginning to emerge. Nano-resolution data from cellular studies, as well as atomic structures from reconstituted systems, have provided first insights, but also point to challenges that remain for obtaining a full structural understanding of how FAK is integrated in the FA complex and the structural changes occurring at different stages of FA maturation. In this Review, we discuss the known structural features of FAK, the interactions with its partners within the FA environment on the cell membrane and propose how its initial assembly in nascent FAs might change during FA maturation under force.

Phosphorylated paxillin and phosphorylated FAK constitute subregions within focal adhesions.

Integrin-mediated adhesions are convergence points for multiple signaling pathways. Their inner structure and diverse functions can be studied with super-resolution microscopy. Here, we examined the spatial organization within focal adhesions by analyzing several adhesion proteins with structured illumination microscopy (SIM). Paxillin (Pax) serves as a scaffold protein and signaling hub in focal adhesions, and focal adhesion kinase (FAK, also known as PTK2) regulates the dynamics of adhesions. We found that their phosphorylated forms, pPax and pFAK, form spot-like, spatially defined clusters within adhesions in several cell lines and confirmed these findings with additional super-resolution techniques. These clusters showed a more regular separation from each other compared with more randomly distributed signals for FAK or paxillin. Mutational analysis indicated that the active (open) FAK conformation is a prerequisite for the pattern formation of pFAK. Live-cell super-resolution imaging revealed that organization in clusters is preserved over time for FAK constructs; however, distance between clusters is dynamic for FAK, while paxillin is more stable. Combined, these data introduce spatial clusters of pPax and pFAK as substructures in adhesions and highlight the relevance of paxillin-FAK binding for establishing a regular substructure in focal adhesions.

Plexin D1 negatively regulates macrophage-derived foam cell migration via the focal adhesion kinase/Paxillin pathway.

BACKGROUND: Macrophage-derived foam cell formation is a hallmark of atherosclerosis and is retained during plaque formation. Strategies to inhibit the accumulation of these cells hold promise as viable options for treating atherosclerosis. Plexin D1 (PLXND1), a member of the Plexin family, has elevated expression in atherosclerotic plaques and correlates with cell migration; however, its role in macrophages remains unclear. We hypothesize that the guidance receptor PLXND1 negatively regulating macrophage mobility to promote the progression of atherosclerosis. METHODS: We utilized a mouse model of atherosclerosis based on a high-fat diet and an ox-LDL- induced foam cell model to assess PLXND1 levels and their impact on cell migration. Through western blotting, Transwell assays, and immunofluorescence staining, we explored the potential mechanism by which PLXND1 mediates foam cell motility in atherosclerosis. RESULTS: Our study identifies a critical role for PLXND1 in atherosclerosis plaques and in a low-migration capacity foam cell model induced by ox-LDL. In the aortic sinus plaques of ApoE-/- mice, immunofluorescence staining revealed significant upregulation of PLXND1 and Sema3E, with colocalization in macrophages. In macrophages treated with ox-LDL, increased expression of PLXND1 led to reduced pseudopodia formation and decreased migratory capacity. PLXND1 is involved in regulating macrophage migration by modulating the phosphorylation levels of FAK/Paxillin and downstream CDC42/PAK. Additionally, FAK inhibitors counteract the ox-LDL-induced migration suppression by modulating the phosphorylation states of FAK, Paxillin and their downstream effectors CDC42 and PAK. CONCLUSION: Our findings indicate that PLXND1 plays a role in regulating macrophage migration by modulating the phosphorylation levels of FAK/Paxillin and downstream CDC42/PAK to promoting atherosclerosis.

Recycling machinery of integrin coupled with focal adhesion turnover via RAB11-UNC13D-FAK axis for migration of pancreatic cancer cells.

BACKGROUND: Recycling of integrin via endosomal vesicles is critical for the migration of cancer cells, which leads to the metastasis of pancreatic cancer and devastating cancer-related death. So, new diagnostic and therapeutic molecules which target the recycling of endosomal vesicles need to be developed. METHODS: Public databases including TCGA, ICGC, GSE21501, GSE28735, and GENT are analyzed to derive diagnostic and therapeutic targets. To reveal biological roles and underlying mechanisms of molecular targets, various molecular biological experiments were conducted. RESULTS: First, we identified UNC13D's overexpression in patients with pancreatic cancer (n = 824) and its prognostic significance and high hazard ratio (HR) in four independent pancreatic cancer cohorts (TCGA, n = 178, p = 0.014, HR = 3.629; ICGC, n = 91, p = 0.000, HR = 4.362; GSE21501, n = 102, p = 0.002, HR = 2.339; GSE28735, n = 45, p = 0.022, HR = 2.681). Additionally, its expression is associated with the clinicopathological progression of pancreatic cancer. Further biological studies have shown that UNC13D regulates the migration of pancreatic cancer cells by coupling the exocytosis of recycling endosomes with focal adhesion turnover via the regulation of FAK phosphorylation. Immunoprecipitation and immunocytochemistry showed the formation of the RAB11-UNC13D-FAK axis in endosomes during integrin recycling. We observed that UNC13D directly interacted with the FERM domain of FAK and regulated FAK phosphorylation in a calcium-dependent manner. Finally, we found co-expression of UNC13D and FAK showed the poorest survival (TCGA, p = 0.000; ICGC, p = 0.036; GSE28735, p = 0.006). CONCLUSIONS: We highlight that UNC13D, a novel prognostic factor, promotes pancreatic cancer progression by coupling integrin recycling with focal adhesion turnover via the RAB11-UNC13D-FAK axis for the migration of pancreatic cancer cells.

The suppression of cell motility through the reduction of FAK activity and expression of cell adhesion proteins by hAMSCs secretome in MDA-MB-231 breast cancer cells.

Breast cancer is a leading cause of death in women worldwide. Cancer therapy based on stem cells is considered as a novel and promising platform. In the present study, we explore the therapeutic effects of human amniotic mesenchymal stromal cells (hAMSCs) through the reduction of focal adhesion kinase (FAK) activity, SHP-2, and cell adhesion proteins such as Paxillin, Vinculin, Fibronectin, Talin, and integrin αvß3 expression in MDA-MB-231 breast cancer cells. For this purpose, we employed a co-culture system using 6-well plate transwell. After 72 h, hAMSCs-treated MDA-MB-231 breast cancer cells, the activity of focal adhesion kinase (FAK) and the expression of SHP-2 and cell adhesion proteins such as Paxillin, Vinculin, Fibronectin, Talin, and integrin αvß3 expression were analyzed using western blot. The shape and migration of cells were also analyzed. Based on our results, a significant reduction in tumor cell motility through downregulation of the tyrosine phosphorylation level of FAK (at Y397 and Y576/577 sites) and cell adhesion expression in MDA-MB-231 breast cancer cells was demonstrated. Our findings indicate that hAMSCS secretome has therapeutic effects on cancer cell migration through downregulation of FAK activity and expression of cell adhesion proteins.

Effect of extracellular matrix stiffness on efficacy of Dapagliflozin for diabetic cardiomyopathy.

BACKGROUND: Extracellular matrix (ECM) stiffness is closely related to the progress of diabetic cardiomyopathy (DCM) and the response of treatment of DCM to anti-diabetic drugs. Dapagliflozin (Dapa) has been proven to have cardio-protective efficacy for diabetes and listed as the first-line drug to treat heart failure. But the regulatory relationship between ECM stiffness and treatment efficacy of Dapa remains elusive. MATERIALS AND METHODS: This work investigated the effect of ECM stiffness on DCM progression and Dapa efficacy using both in vivo DCM rat model and in vitro myocardial cell model with high glucose injury. First, through DCM rat models with various levels of myocardial injury and administration with Dapa treatment for four weeks, the levels of myocardial injury, myocardial oxidative stress, expressions of AT1R (a mechanical signal protein) and the stiffness of myocardial tissues were obtained. Then for mimicking the stiffness of myocardial tissues at early and late stages of DCM, we constructed cell models through culturing H9c2 myocardial cells on the polyacrylamide gels with two stiffness and exposed to a high glucose level and without/with Dapa intervention. The cell viability, reactive oxygen species (ROS) levels and expressions of mechanical signal sensitive proteins were obtained. RESULTS: The DCM progression is accompanied by the increased myocardial tissue stiffness, which can synergistically exacerbate myocardial cell injury with high glucose. Dapa can improve the ECM stiffness-induced DCM progression and its efficacy on DCM is more pronounced on the soft ECM, which is related to the regulation pathway of AT1R-FAK-NOX2. Besides, Dapa can inhibit the expression of the ECM-induced integrin ß1, but without significant impact on piezo 1. CONCLUSIONS: Our study found the regulation and effect of biomechanics in the DCM progression and on the Dapa efficacy on DCM, providing the new insights for the DCM treatment. Additionally, our work showed the better clinical prognosis of DCM under early Dapa intervention.