Β1 Integrin Research Articles

Salmonella YrbD protein mediates invasion into the host by interacting with β2 integrin1

Salmonella enterica serovar Typhimurium, the causative agent of gastroenteritis, is one of the most successful intracellular pathogens. Although certain host factors for Salmonella infection have been unveiled, the factors mediating Salmonella entry, particularly the invasion process, remain obscure. Here, we have unearthed β2 integrin, a crucial member of the integrin family, as an important host factor facilitating Salmonella invasion. It demonstrated that overexpression of β2 integrin promotes Salmonella invasion, while the knockdown of β2 integrin significantly diminishes the extent of invasion. Moreover, Salmonella exhibits specific binding affinity towards β2 integrin, and the block of β2 integrin on cell surface substantially reduces the infection of cells in vitro. The ectodomain soluble protein of β2 integrin neutralized Salmonella infection both in cells (in vitro) and in mice (in vivo). Additionally, Salmonella protein YrbD directly interacts with β2 integrin to facilitate its invasion. To our knowledge, this study showed for the first time that the protein YrbD mediates Salmonella adhesion and internalization into host cells by interacting with β2 integrin. These findings not only broaden our understanding of the mechanisms underlying Salmonella entry, but also identify a prospective target for therapeutic control.

The association between integrin β4 overexpression and lymphovascular invasion in papillary thyroid cancer.

Lymphovascular invasion is an independent prognostic marker in papillary thyroid carcinomas. In addition, integrin β4 is associated with advanced progression and metastasis in many malignancies. We aimed to investigate the relationship between integrin β4 and lymphovascular invasion in papillary thyroid carcinoma. 73 patients with papillary thyroid cancer (48 patients with lymphovascular invasion and 25 patients without) were included in our study. The immunohistochemical staining score for integrin b4 was evaluated according to the percentage and intensity of staining. The staining intensity was scored as 0 (no staining), 1 (weak staining - light yellow), 2 (medium staining - yellow-brown), and 3 (strong staining - brown). The staining was scored by multiplying the percentage and intensity of staining. The mean percentage of integrin b4 staining was 63.54 ± 22.26% in the group with lymphovascular invasion and 10.2 ± 22.48% in the group without lymphovascular invasion (p < 0.001). When evaluated in terms of staining score, it was found to be 107.08 ± 45.29 in the group with lymphovascular invasion and 16.2 ± 40.03 in the group without lymphovascular invasion (p < 0.001). There was a linear relationship between the percentage of integrin β4 and the staining scores (r² = 0.881; p < 0.001). In the by receiver-operating characteristic (ROC) curve analysis for the cut-off value of the percentage of integrin b4 staining, the area under the curve was found to be 0.916. The cut-off value for the percentage of integrin b4 was found to be 35 (sensitivity 91.7% and specificity 88%) (odds 80.66%). A significant relationship was found between integrin b4 expression and lymphovascular invasion in papillary thyroid carcinomas. Integrin b4 expression level can be used as a marker to predict the presence of lymphovascular invasion in papillary thyroid carcinomas, especially in large tumours where it may not be possible to sample the entire tumour.

Photoactivatable substrates show diverse phenotypes of leader cells in collective migration when moving along different extracellular matrix proteins.

In cancer metastasis, collectively migrating clusters are discriminated into leader and follower cells that move through extracellular matrices (ECMs) with different characteristics. The impact of changes in ECM protein types on leader cells and migrating clusters is unknown. To address this, we investigated the response of leader cells and migrating clusters upon moving from one ECM protein to another using a photoactivatable substrate bearing photocleavable PEG (PCP), whose surface changes from protein-repellent to protein-adhesive in response to light. We chose laminin and collagen I for our study since they are abundant in two distinct regions in living tissues, namely basement membrane and connective tissue. Using the photoactivatable substrates, the precise deposition of the first ECM protein in the irradiated areas was achieved, followed by creating well-defined cellular confinements. Secondary irradiation enabled the deposition of the second ECM protein in the new irradiated regions, resulting in region-selective heterogeneous and homogenous ECM protein-coated surfaces. Different tendencies in leader cell formation from laminin into laminin compared to those migrating from laminin into collagen were observed. The formation of focal adhesion and actin structures for cells within the same cluster in the ECM proteins responded according to the underlying ECM protein type. Finally, integrin β1 was crucial for the appearance of leader cells for clusters migrating from laminin into collagen. However, when it came to laminin into laminin, integrin β1 was not responsible. This highlights the correlation between leader cells in collective migration and the biochemical signals that arise from underlying extracellular matrix proteins.

EXOSC5 maintains cancer stem cell activity in endometrial cancer by regulating the NTN4/integrin β1 signalling axis.

Endometrial carcinoma (EC) is a common type of uterine cancer in developed countries, originating from the uterine epithelium. The incidence rate of EC in Taiwan has doubled from 2005. Cancer stem cells (CSCs) are a subpopulation of cancer cells that have high tumorigenicity and play a crucial role in the malignant processes of cancer. Targeting molecules associated with CSCs is essential for effective cancer treatments. This study delves into the role of Exosome component 5 (EXOSC5) in EC. Data from The Cancer Genome Atlas suggests a correlation between high EXOSC5 mRNA expression and unfavorable EC prognosis. EXOSC5 knockdown diminished EC-CSC self-renewal and reduced expression of key cancer stemness proteins, including c-MYC and SOX2. Intriguingly, this knockdown significantly curtailed tumorigenicity and CSC frequency in EC tumor spheres. A mechanistic examination revealed a reduction in netrin4 (NTN4) levels in EXOSC5-depleted EC cells. Moreover, NTN4 treatment amplified EC cell CSC activity and, when secreted, NTN4 partnered with integrin β1, subsequently triggering the FAK/SRC axis to elevate c-MYC activity. A clear positive relation between EXOSC5 and NTN4 was evident in 93 EC tissues. In conclusion, EXOSC5 augments NTN4 expression, activating c-MYC via the integrin β1/FAK/SRC pathway, offering potential avenues for EC diagnosis and treatment.

Regulatory effects of stress release from decellularized periosteum on proliferation, migration, and osteogenic differentiation of periosteum-derived cells.

Bone injury is often associated with tears in the periosteum and changes in the internal stress microenvironment of the periosteum. In this study, we investigated the biological effects of periosteal prestress release on periosteum-derived cells (PDCs) and the potential mechanisms of endogenous stem cell recruitment. Decellularized periosteum with natural extracellular matrix (ECM) components was obtained by a combination of physical, chemical, and enzymatic decellularization. The decellularized periosteum removed immunogenicity while retaining the natural network structure and composition of the ECM. The Young's modulus has no significant difference between the periosteum before and after decellularization. The extracted PDCs were further composited with the decellularized periosteum and subjected to 20% stress release. It was found that the proliferative capacity of PDCs seeded on decellularized periosteum was significantly enhanced 6 h after stress release of the periosteum. The cell culture supernatant obtained after periosteal prestress release was able to significantly promote the migration ability of PDCs within 24 h. Enzyme-linked immunosorbnent assay (ELISA) experiments showed that the expression of stroma-derived factor-1α (SDF-1α) and vascular endothelial growth factor (VEGF) in the supernatant increased significantly after 3 h and 12 h of stress release, respectively. Furthermore, periosteal stress release promoted the high expression of osteogenic markers osteocalcin (OCN), osteopontin (OPN), and collagen type I of PDCs. The change in stress environment caused by the release of periosteal prestress was sensed by integrin β1, a mechanoreceptor on the membrane of PDCs, which further stimulated the expression of YAP in the nucleus. These investigations provided a novel method to evaluate the importance of mechanical stimulation in periosteum, which is also of great significance for the design and fabrication of artificial periosteum with mechanical regulation function.

Hypoxia-driven TNS4 fosters HNSCC tumorigenesis by stabilizing integrin α5β1 complex and triggering FAK-mediated Akt and TGFβ signaling pathways.

Head and neck squamous cell carcinoma (HNSCC) remains a formidable clinical challenge due to its high recurrence rate and limited targeted therapeutic options. This study aims to elucidate the role of tensin 4 (TNS4) in the pathogenesis of HNSCC across clinical, cellular, and animal levels. We found a significant upregulation of TNS4 expression in HNSCC tissues compared to normal controls. Elevated levels of TNS4 were associated with adverse clinical outcomes, including diminished overall survival. Functional assays revealed that TNS4 knockdown attenuated, and its overexpression augmented, the oncogenic capabilities of HNSCC cells both in vitro and in vivo. Mechanistic studies revealed that TNS4 overexpression promotes the interaction between integrin α5 and integrin β1, thereby activating focal adhesion kinase (FAK). This TNS4-mediated FAK activation simultaneously enhanced the PI3K/Akt signaling pathway and facilitated the interaction between TGFβRI and TGFβRII, leading to the activation of the TGFβ signaling pathway. Both of these activated pathways contributed to HNSCC tumorigenesis. Additionally, we found that hypoxia-inducible factor 1α (HIF-1α) transcriptionally regulated TNS4 expression. In conclusion, our findings provide the basis for innovative TNS4-targeted therapeutic strategies, which could potentially improve prognosis and survival rates for patients with HNSCC.

Myosin light chain 6 (Myl6) interacts with kindlin-3 and is required to support integrin αIIbβ3 activation in platelets in mice

BackgroundKindlin-3 in platelets plays an essential role in supporting integrin αIIbβ3 activation, platelet spreading, aggregation, and clot retraction by binding to the integrin β3 cytoplasmic tail. However, the mechanism by which kindlin-3 mediates the crosstalk between integrin αIIbβ3 and myosin in platelets remains unknown. ObjectivesTo examine the role of myosin light chain 6 (Myl6) in supporting integrin αIIbβ3 activation in platelets. MethodsMyl6fl/flPF4-Cre mice with a deficiency of Myl6 in the megakaryocyte lineage were generated, and integrin αIIbβ3 activation in Myl6-deficient platelets was analyzed. ResultsWe identified a novel kindlin-3 binding protein, Myl6, an essential light chain of myosin in platelets. Myl6fl/flPF4-Cre mice exhibited significant macrothrombocytopenia resulting from defective proplatelet formation. In the absence of Myl6, integrin αIIbβ3 activation in platelets was significantly suppressed, and platelet aggregation was substantially impaired. Interestingly, the deficiency of Myl6 in platelets preferentially affected the binding of a multivalent ligand compared to a monovalent ligand to integrin αIIbβ3 upon activation, indicating that Myl6 may contribute to the avidity modulation of integrin αIIbβ3 by binding to kindlin-3. Furthermore, blood coagulation ability was impaired in Myl6fl/flPF4-Cre mice, and consistently, these mice exhibited defects in both hemostatic and thrombotic functions. ConclusionIn summary, these results suggest that Myl6, as a novel kindlin-3 binding partner, is required to support integrin αIIbβ3 activation in platelets, which plays an important role in both hemostasis and thrombosis.

CYR61 Is Overexpressed in Human Melanoma Tissue Ex Vivo and Promotes Melanoma Cell Survival and Proliferation Through Its Binding Ligand Integrin β3 In Vitro.

CYR61 Is Overexpressed in Human Melanoma Tissue Ex Vivo and Promotes Melanoma Cell Survival and Proliferation Through Its Binding Ligand Integrin β3 In Vitro.

Multifunctional and bioinspired titanium surface with multilayer nanofilms for novel dental implant applications.

Introduction: Smart multifunctional surfaces targeting intricate biological events or versatile therapeutic strategies are imminent to achieve long-term transmucosal implant success. Methods: This study used dopamine (DA), graphene oxide (GO), and type IV collagen (COL-IV) to construct multilayer nanofilms (DGCn) based on their universal adhesive and biomimetic properties to design a versatile and bioactive titanium implant. The characterization of DGCn on different titanium surfaces was performed, and its loading capacity, release profile, in situ gene delivery, and in vitro biological properties were preliminarily evaluated. Results: Our results demonstrate that hydrogenated TiO2 nanotubes (H) provide a better platform for the DGCn coating than machined Ti and air-TiO2 nanotubes. The H-DGC10 displayed the most stable surface with excellent loading capacity, sustained-release profile, and in situ gene transfection efficiency; this could be due to the high specific surface area of H and GO, as well as the functional groups in H, DA, and GO. Moreover, the H-DGC10 exhibited good biocompatibility for human oral epithelial cells and promoted the expression of integrin β4 and laminin 332, both being hemidesmosome-related proteins. Discussion: Our findings suggest that H-DGCn can be designed as a smart multifunctional interface for titanium implants to achieve long-term transmucosal implant success and aid in versatile therapeutic strategies.

Oleuropein Stimulates Migration of Human Trophoblast Cells and Expression of Invasion-Associated Markers.

Successful pregnancy establishment requires highly synchronized cross talk between the invasive trophoblast cells and the receptive maternal endometrium. Any disturbances in this tightly regulated process may lead to pregnancy complications. Local factors such as nutrients, hormones, cytokines and reactive oxygen species modulate the invasion of extravillous trophoblasts through critical signaling cascades. Epidemiological studies strongly indicate that a Mediterranean diet can significantly impact molecular pathways during placentation. Therefore, the aim of the current study was to examine whether oleuropein (OLE), one of the main compounds of the Mediterranean diet, may influence trophoblast cell adhesion and migration, as well as the expression of invasion-associated molecular markers and inflammatory pathways fostering these processes. HTR-8/SVneo cells were incubated with OLE at selected concentrations of 10 and 100 µM for 24 h. Results showed that OLE did not affect trophoblast cell viability, proliferation and adhesion after 24 h in in vitro treatment. The mRNA expression of integrin subunits α1, α5 and β1, as well as matrix-degrading enzymes MMP-2 and -9, was significantly increased after treatment with 10 µM OLE. Furthermore, OLE at a concentration of 10 µM significantly increased the protein expression of integrin subunits α1 and β1. Also, OLE inhibited the activation of JNK and reduced the protein expression of COX-2. Finally, a lower concentration of OLE 10 µM significantly stimulated migration of HTR-8/SVneo cells. In conclusion, the obtained results demonstrate the effects of OLE on the function of trophoblast cells by promoting cell migration and stimulating the expression of invasion markers. As suggested from results, these effects may be mediated via inhibition of the JNK signaling pathway.

USP10 promotes migration and cisplatin resistance in esophageal squamous cell carcinoma cells.

Cisplatin-based chemotherapy is the main treatment option for advanced or metastatic esophageal squamous cell carcinoma (ESCC). However, most ESCC patients develop drug resistance within 2years after receiving cisplatin chemotherapy. Ubiquitin-specific protease 10 (USP10) is abnormally expressed in a variety of cancers, but the mechanistic roles of USP10 in ESCC are still obscure. Here, the effects of USP10 on the migration and cisplatin resistance of ESCC in vivo and in vitro and the underlying mechanisms have been investigated by bioinformatics analysis, RT-PCR, western blotting, immunoprecipitation, immunohistochemistry, cell migration and MTS cell proliferation assays, deubiquitination assay, and mouse tail vein injection model. USP10 was significantly up-regulated in ESCC tissues compared with adjacent normal tissues in both public databases and clinical samples and was closely associated with overall survival. Subsequent results revealed that USP10 contributed to the migration and cisplatin resistance of ESCC cells, while knocking down USP10 in cisplatin-resistant cells exhibited opposite effects in vitro and in vivo. Further Co-IP experiments showed that integrin β1 and YAP might be targets for USP10 deubiquitination. Moreover, deficiency of USP10 significantly inhibited the migrative and chemo-resistant abilities of ESCC cells, which could be majorly reversed by integrin β1 or YAP reconstitution. Altogether,USP10 was required for migration and cisplatin resistance in ESCC through deubiquinating and stabilizing integrin β1/YAP, highlighting that inhibition of USP10 may be a potential therapeutic strategy for ESCC.

A synergistic regulation works in matrix stiffness-driven invadopodia formation in HCC

Growing evidence has suggested that increased matrix stiffness can significantly strengthen the malignant characteristics of hepatocellular carcinoma (HCC) cells. However, whether and how increased matrix stiffness regulates the formation of invadopodia in HCC cells remain largely unknown. In the study, we developed different experimental systems in vitro and in vivo to explore the effects of matrix stiffness on the formation of invadopodia and its relevant molecular mechanism. Our results demonstrated that increased matrix stiffness remarkably augmented the migration and invasion abilities of HCC cells, upregulated the expressions of invadopodia-associated genes and enhanced the number of invadopodia. Two regulatory pathways contribute to matrix stiffness-driven invadopodia formation together in HCC cells, including direct triggering invadopodia formation through activating integrin β1 or Piezo1/ FAK/Src/Arg/cortactin pathway, and indirect stimulating invadopodia formation through improving EGF production to activate EGFR/Src/Arg/cortactin pathway. Src was identified as the common hub molecule of two synergistic regulatory pathways. Simultaneously, activation of integrin β1/RhoA/ROCK1/MLC2 and Piezo1/Ca2+/MLCK/MLC2 pathways mediate matrix stiffness-reinforced cell migration. This study uncovers a new mechanism by which mechanosensory pathway and biochemical signal pathway synergistically regulate the formation of invadopodia in HCC cells.

Rab25 suppresses colon cancer cell invasion through upregulating claudin‑7 expression.

Ras‑related protein25 (Rab25) is a member of small GTPase and is implicated in cancer cell progression of various types of cancer. Growing evidence suggests the context‑dependent role of Rab25 in cancer invasiveness. Claudin‑7 is a tight junction protein and has been known to suppress cancer cell invasion. Although Rab25 was reported to repress cancer aggressiveness through recycling β1 integrin to the plasma membrane, the detailed underlying mechanism remains to be elucidated. The present study identified the critical role of claudin‑7 in Rab25‑induced suppression of colon cancer invasion. 3D Matrigel system and modified Boyden chamber analysis showed that enforced expression of Rab25 attenuated colon cancer cell invasion. In addition, Rab25 inactivated epidermal growth factor receptor (EGFR) and increased E‑cadherin expression. Unexpectedly, it was observed that Rab25 induces claudin‑7 expression through protein stabilization. In addition, ectopic claudin‑7 expression reduced EGFR activity and Snail expression as well as colon cancer cell invasion. However, silencing of claudin‑7 expression reversed the tumor suppressive role of Rab25, thereby increasing colon cancer cell invasiveness. Collectively, the present data indicated that Rab25 inactivates EGFR and colon cancer cell invasion by upregulating claudin‑7 expression.

The role of EDIL3 in maintaining cartilage extracellular matrix and inhibiting osteoarthritis development

AimsTherapeutic agents that prevent chondrocyte loss, extracellular matrix (ECM) degradation, and osteoarthritis (OA) progression are required. The expression level of epidermal growth factor (EGF)-like repeats and discoidin I-like domains-containing protein 3 (EDIL3) in damaged human cartilage is significantly higher than in undamaged cartilage. However, the effect of EDIL3 on cartilage is still unknown.MethodsWe used human cartilage plugs (ex vivo) and mice with spontaneous OA (in vivo) to explore whether EDIL3 has a chondroprotective effect by altering OA-related indicators.ResultsEDIL3 protein prevented chondrocyte clustering and maintained chondrocyte number and SOX9 expression in the human cartilage plug. Administration of EDIL3 protein prevented OA progression in STR/ort mice by maintaining the number of chondrocytes in the hyaline cartilage and the number of matrix-producing chondrocytes (MPCs). It reduced the degradation of aggrecan, the expression of matrix metalloproteinase (MMP)-13, the Osteoarthritis Research Society International (OARSI) score, and bone remodelling. It increased the porosity of the subchondral bone plate. Administration of an EDIL3 antibody increased the number of matrix-non-producing chondrocytes (MNCs) in cartilage and exacerbated the serum concentrations of OA-related pro-inflammatory cytokines, including monocyte chemotactic protein-3 (MCP-3), RANTES, interleukin (IL)-17A, IL-22, and GROα. Administration of β1 and β3 integrin agonists (CD98 protein) increased the expression of SOX9 in OA mice. Hence, EDIL3 might activate β1 and β3 integrins for chondroprotection. EDIL3 may also protect cartilage by attenuating the expression of IL-1β-enhanced phosphokinase proteins in chondrocytes, especially glycogen synthase kinase 3 alpha/beta (GSK-3α/β) and phospholipase C gamma 1 (PLC-γ1).ConclusionEDIL3 has a role in maintaining the cartilage ECM and inhibiting the development of OA, making it a potential therapeutic drug for OA.Cite this article: Bone Joint Res 2023;12(12):734–746.

Heart Failure Promotes Cancer Progression in an Integrin β1-Dependent Manner.

Heart failure and cancer are currently the deadliest diseases in the Western world, posing the most pressing clinical challenges that remain unmet today. Both conditions share similar risk factors, including age, genetics, lifestyle, chronic inflammation, stress, and more. Furthermore, medications that are being used to counteract cancer frequently result in cardiotoxicity and the spontaneous emergence of heart failure. Thus, heart failure and cancer display an intimate connection and share similarities. Recent studies show that cardiac remodeling and heart failure promote cancer progression and metastasis. Using three different mouse models for heart failure revealed that the communication between the remodeled heart and the tumor is facilitated through multiple secreted factors. Among these factors, Periostin was consistently found to be elevated in all models and was shown to be required in vitro. Yet, whether Periostin is necessary for tumor promotion in vivo is unknown. Towards this end, we examined tumor promotion in mice lacking Periostin following transverse aortic constriction (TAC). Despite the loss of Periostin, tumor growth was promoted in the TAC-operated mice. This likely occurred due to increased levels of various cytokines and growth factors in Periostin KO mice. Many of these factors are potential ligands of Integrin receptors. Therefore, we next studied the role of Integrin receptors in the tumor-promotion phenotype following heart failure. We generated cancer cells with an Integrin β1 loss of function mutation and examined tumor growth in the presence and absence of heart failure. Integrin β1 KO cancer cells fail to display cardiac-remodeling-dependent tumor-promotion. Interestingly, a previous study showed that renal cell carcinoma cells (Renca) fail to be promoted following a myocardial infarction. Consistently, we show that Renca cells do not respond to secreted factors derived from the failing heart both in vitro and in vivo. Interestingly, Renca cells display low basal mRNA levels of Integrin β1 which may explain the inability of heart failure to promote their growth. The findings may have significant clinical relevance to cardio-oncology patients who suffer from cancers with high levels of Integrin β1. Chemotherapy leading to cardiotoxicity in these patients may generate a vicious cycle with poor prognosis.

Reversible SAHH inhibitor ameliorates MIA-induced osteoarthritis of rats through suppressing MEK/ERK pathway

Osteoarthritis (OA) is characterized by gradual articular cartilage degradation, accompanied by persistent low-grade joint inflammation, correlating with radiographic and pain-related progression. The latent therapeutic potential of DZ2002, a reversible inhibitor of S-adenosyl-L-homocysteine hydrolase (SAHH), holds promise for OA intervention. This study endeavored to examine the therapeutic efficacy of DZ2002 within the milieu of OA. The cytotoxicity of DZ2002 was evaluated using the MTT assay on bone marrow-derived macrophages. The inhibitory impact of DZ2002 during the process of osteoclastogenesis was assessed using TRAP staining, analysis of bone resorption pits, and F-actin ring formation. Mechanistic insights were derived from qPCR and Western blot analyses. Through the intra-articular injection of monosodium iodoacetate (MIA), an experimental rat model of OA was successfully instituted. This was subsequently accompanied by a series of assessments including Von Frey filament testing, analysis of weight-bearing behaviors, and micro-CT imaging, all aimed at assessing the effectiveness of DZ2002. The findings emphasized the effectiveness of DZ2002 in mitigating osteoclastogenesis induced by M-CSF/RANKL, evident through a reduction in TRAP-positive OCs and bone resorption. Moreover, DZ2002 modulated bone resorption-associated gene and protein expression (CTSK, CTR, Integrin β3) via the MEK/ERK pathway. Encouragingly, DZ2002 also alleviates MIA-induced pain, cartilage degradation, and bone loss. In conclusion, DZ2002 emerges as a potential therapeutic contender for OA, as evidenced by its capacity to hinder in vitro M-CSF/RANKL-induced osteoclastogenesis and mitigate in vivo osteoarthritis progression. This newfound perspective provides substantial support for considering DZ2002 as a compelling agent for osteoarthritis intervention.

MiRNA-ome plasma analysis unveils changes in blood–brain barrier integrity associated with acute liver failure in rats

BackgroundHepatic encephalopathy (HE) symptoms associated with liver insufficiency are linked to the neurotoxic effects of ammonia and other toxic metabolites reaching the brain via the blood–brain barrier (BBB), further aggravated by the inflammatory response. Cumulative evidence documents that the non-coding single-stranded RNAs, micro RNAs (miRs) control the BBB functioning. However, miRs’ involvement in BBB breakdown in HE is still underexplored. Here, we hypothesized that in rats with acute liver failure (ALF) or rats subjected to hyperammonemia, altered circulating miRs affect BBB composing proteins.MethodsTransmission electron microscopy was employed to delineate structural alterations of the BBB in rats with ALF (thioacetamide (TAA) intraperitoneal (ip.) administration) or hyperammonemia (ammonium acetate (OA) ip. administration). The BBB permeability was determined with Evans blue dye and sodium fluorescein assay. Plasma MiRs were profiled by Next Generation Sequencing (NGS), followed by in silico analysis. Selected miRs, verified by qRT-PCR, were examined in cultured rat brain endothelial cells. Targeted protein alterations were elucidated with immunofluorescence, western blotting, and, after selected miR mimics transfection, through an in vitro resistance measurement.ResultsChanges in BBB structure and increased permeability were observed in the prefrontal cortex of TAA rats but not in the brains of OA rats. The NGS results revealed divergently changed miRNA-ome in the plasma of both rat models. The in silico analysis led to the selection of miR-122-5p and miR-183-5p with their target genes occludin and integrin β1, respectively, as potential contributors to BBB alterations. Both proteins were reduced in isolated brain vessels and cortical homogenates in TAA rats. We documented in cultured primary brain endothelial cells that ammonia alone and, in combination with TNFα increases the relative expression of NGS-selected miRs with a less pronounced effect of TNFα when added alone. The in vitro study also confirmed miR-122-5p-dependent decrease in occludin and miR-183-5p-related reduction in integrin β1 expression.ConclusionThis work identified, to our knowledge for the first time, potential functional links between alterations in miRs residing in brain endothelium and BBB dysfunction in ALF.

ACTN1 promotes HNSCC tumorigenesis and cisplatin resistance by enhancing MYH9-dependent degradation of GSK-3β and integrin β1-mediated phosphorylation of FAK

BackgroundHead and neck squamous cell carcinoma (HNSCC) is one of the most common malignant tumors globally. Understanding the molecular basis of tumor progression and drug resistance can offer innovative strategies to enhance clinical outcomes for HNSCC patients.MethodsThe cytoskeletal remodeling genes associated with cisplatin resistance were screened using a PCR array. The role of alpha-actinin 1 (ACTN1) in modulating cisplatin resistance and tumorigenesis in HNSCC was evaluated both in vitro and in vivo. Co-immunoprecipitation (Co-IP), IP-mass spectrometry (MS), western blotting, dual-luciferase assay, and bioinformatics analysis were performed to elucidate the underlying mechanisms involved.ResultsOur study identifies ACTN1 as a crucial contributor to cisplatin resistance and tumorigenesis in HNSCC, as evidenced across cellular, animal, and patient-derived xenograft models. From a clinical perspective, overexpression of ACTN1 significantly correlates with a suboptimal response to neoadjuvant chemotherapy and reduced overall survival in HNSCC patients. Mechanistically, ACTN1 predominantly activates β-catenin-mediated signaling by promoting the interaction between myosin heavy chain 9 (MYH9) and GSK-3β, leading to the ubiquitin-dependent degradation of GSK-3β. ACTN1 also interacts with integrin β1, subsequently activating the FAK/PI3K/AKT pathway, providing an additional avenue for the activation of β-catenin signaling. Our study also unveils that the β-catenin/c-Myc axis transcriptionally regulates ACTN1, thereby creating a positive feedback loop promoting HNSCC tumorigenesis and drug resistance.ConclusionsThese insights underscore the novel mechanisms that highlight ACTN1's pivotal role in driving HNSCC progression and resistance to chemotherapy, suggesting ACTN1 as a promising therapeutic target in HNSCC management.

Identification and characterization of β3 integrin in Crassostrea hongkongensis

Crassostrea hongkongensis is a main oyster species farmed in South China Sea offshore area, and commonly suffers from vibriosis. Vibrio parahaemolyticus is one of the principal pathogens that causes the oyster vibriosis. The integrin from C. hongkongensis was studied for its function against V. parahaemolyticus in the current work. The full-length cDNA sequence was 3274 bp with an open reading frame (ORF) of 2397 bp that encoded a polypeptide of 798 amino acids. The polypeptide included typical domains of β integrin (such as INB domain, three EGF-like domains, and highly conserved NPX[Y/F] motif), and phylogenetically clustered with β3 integrins of other invertebrates, thereby, designated as ChIntβ3. The mRNA transcripts of ChIntβ3 were significantly higher in the heart, hemocytes, adductor muscle and hepatopancreas than those in gills and mantle of C. hongkongensis. Following stimulation with V. parahaemolyticus, the mRNA expression level of ChIntβ3 was significantly up-regulated in both hemocytes and hepatopancreas. The recombinant ChIntβ3 had significantly agglutinating activity on Escherichia coli, V. parahaemolyticus, Bacillus amyloliquefaciens, Pichia pastoris and the red blood cells of rabbit. In the one hand, the survival rate (83.33 ± 5.77%) of the oyster after V. parahaemolyticus infection at 30 days was significantly higher in the ChIntβ3 overexpression group than those in the physiological saline control group (26.67 ± 20.82%) (p < 0.01) and the overexpressed GFP group (26.67 ± 5.77%) (p < 0.01); in the other hand, the cumulative survival duration of C. hongkongensis was significantly shorter in the RNAi treatment group (197.6 ± 2.77 h) than that in the control group (247.2 ± 36.24 h) (p < 0.01). The results indicated that ChIntβ3 was involved in the recognition and clearance of V. parahaemolyticus through cell adhesion, and contributed to the further understanding of the function of ChIntβ3 against infection of pathogens.

Integrin α6β4 Confers Doxorubicin Resistance in Cancer Cells by Suppressing Caspase-3-Mediated Apoptosis: Involvement of N-Glycans on β4 Integrin Subunit.

Drug resistance is a major obstacle to successful cancer treatment. Therefore, it is essential to understand the molecular mechanisms underlying drug resistance to develop successful therapeutic strategies. α6β4 integrin confers resistance to apoptosis and regulates the survival of cancer cells; however, it remains unclear whether α6β4 integrin is directly involved in chemoresistance. Here, we show that α6β4 integrin promotes doxorubicin resistance by decreasing caspase-3-mediated apoptosis. We found that the overexpression of α6β4 integrin by the β4 integrin gene rendered MDA-MB435S and Panc-1 cells more resistant to doxorubicin than control cells. The acquired resistance to doxorubicin by α6β4 integrin expression was abolished by the deletion of the cytoplasmic signal domain in β4 integrin. Similar results were found in MDA-MB435S and Panc-1 cells when N-glycan-defective β4 integrin mutants were overexpressed or bisecting GlcNAc residues were increased on β4 integrin by the co-expression of N-acetylglucosaminyltransferase III with β4 integrin. The abrogation of α6β4 integrin-mediated resistance to doxorubicin was accompanied by reduced cell viability and an increased caspase-3 activation. Taken together, our results clearly suggest that α6β4 integrin signaling plays a key role in the doxorubicin resistance of cancer cells, and N-glycans on β4 integrin are involved in the regulation of cancer cells.