Roles for epithelial integrin α3ß1 in regulation of the microenvironment during normal and pathological tissue remodeling.

Integrin receptors for the extracellular matrix activate intracellular signaling pathways that are critical for tissue development, homeostasis, and regeneration/repair, and their loss or dysregulation contributes to many developmental defects and tissue pathologies. This review will focus on tissue remodeling roles for integrin α3ß1, a receptor for laminins found in the basement membranes (BMs) that underlie epithelial cell layers. As a paradigm, we will discuss literature that supports a role for α3ß1 in promoting ability of epidermal keratinocytes to modify their tissue microenvironment during skin development, wound healing, or tumorigenesis. Preclinical and clinical studies have shown that this role depends largely on ability of α3ß1 to govern the keratinocyte's repertoire of secreted proteins, or the "secretome," including 1) matrix proteins and proteases involved in matrix remodeling and 2) paracrine-acting growth factors/cytokines that stimulate other cells with important tissue remodeling functions (e.g., endothelial cells, fibroblasts, inflammatory cells). Moreover, α3ß1 signaling controls gene expression that helps epithelial cells carry out these functions, including genes that encode secreted matrix proteins, proteases, growth factors, or cytokines. We will review what is known about α3ß1-dependent gene regulation through both transcription and posttranscriptional mRNA stability. Regarding the latter, we will discuss examples of α3ß1-dependent alternative splicing (AS) or alternative polyadenylation (APA) that prevents inclusion of cis-acting mRNA sequences that would otherwise target the transcript for degradation via nonsense-mediated decay or destabilizing AU-rich elements (AREs) in the 3'-untranslated region (3'-UTR). Finally, we will discuss prospects and anticipated challenges of exploiting α3ß1 as a clinical target for the treatment of cancer or wound healing.

Keratinocyte integrin α3ß1 induces expression of the macrophage stimulating factor, CSF-1, through a YAP/TEAD-dependent mechanism.

The development of wound therapy targeting integrins is hampered by inadequate understanding of integrin function in cutaneous wound healing and the wound microenvironment. Following cutaneous injury, keratinocytes migrate to restore the skin barrier, and macrophages aid in debris clearance. Thus, both keratinocytes and macrophages are critical to the coordination of tissue repair. Keratinocyte integrins have been shown to participate in this coordinated effort by regulating secreted factors, some of which crosstalk to distinct cells in the wound microenvironment. Epidermal integrin α3ß1 is a receptor for laminin-332 in the cutaneous basement membrane. Here we show that wounds deficient in epidermal α3ß1 express less epidermal-derived macrophage colony-stimulating factor 1 (CSF-1), the primary macrophage-stimulating growth factor. α3ß1-deficient wounds also have fewer wound-proximal macrophages, suggesting that keratinocyte α3ß1 may stimulate wound macrophages through the regulation of CSF-1. Indeed, using a set of immortalized keratinocytes, we demonstrate that keratinocyte-derived CSF-1 supports macrophage growth, and that α3ß1 regulates Csf1 expression through Src-dependent stimulation of Yes-associated protein (YAP)-Transcriptional enhanced associate domain (TEAD)-mediated transcription. Consistently, α3ß1-deficient wounds in vivo display a substantially reduced number of keratinocytes with YAP-positive nuclei. Overall, our current findings identify a novel role for epidermal integrin α3ß1 in regulating the cutaneous wound microenvironment by mediating paracrine crosstalk from keratinocytes to wound macrophages, implicating α3ß1 as a potential target of wound therapy.

INHIBITION OF INTEGRIN VLA-3 AND TETRASPANIN CD151 PROTECTS AGAINST NEUTROPHIL-MEDIATED ENDOTHELIAL DAMAGE.

ABSTRACT: Background: The recruitment of neutrophils to sites of localized injury or infection is initiated by changes on the surface of endothelial cells located in proximity to tissue damage. Inflammatory mediators, such as TNF-α, increase surface expression of adhesive ligands and receptors on the endothelial surface to which neutrophils tether and adhere. Neutrophils then transit through the activated endothelium to reach sites of tissue injury with little lasting vascular injury. However, in cases of sepsis, the interaction of endothelial cells with highly activated neutrophils can cause damage vascular damage. The identification of molecules that are essential for neutrophil diapedesis may reveal targets of therapeutic opportunity for preservation of endothelial function in the presence of critical illness. We tested the hypothesis that inhibition of neutrophil ß1 integrin very late antigen-3 (VLA-3; α3ß1) and/or inhibition of the tetraspanin (TM4) family member CD151 would protect against neutrophil-mediated loss of endothelial function. Methods: Blood was obtained from septic patients or healthy donors. Neutrophils were purified, and aliquots were treated with/without proinflammatory molecules. Confluent human umbilical vascular endothelial cells were activated with TNF-α. Electric cell impedance sensing was used to determine monolayer resistance over time after the addition of neutrophils that were treated with blocking antibodies against VLA-3 and/or CD151 or isotype controls. Groups (depending on relevancy) were analyzed by Mann-Whitney U test, Wilcoxon test, or repeated-measures one-way ANOVA. Results: Neutrophils from septic patients and neutrophils activated ex vivo reduced endothelial monolayer resistance to a greater extent than neutrophils from healthy donors. Antibody blockade of VLA-3 and/or CD151 significantly reduced activation-associated endothelial damage. Similar findings were demonstrated on fibronectin, collagen I, collagen IV, and laminin, suggesting that neutrophil surface VLA-3 and CD151 are responsible for endothelial damage regardless of substrata and are likely to be operative in all bodily tissues. Conclusion: This report identifies VLA-3 and CD151 on the activated human neutrophil, which are responsible for damage to endothelial function. Targeting these molecules in vivo may demonstrate preservation of organ function during critical illness.

Mechanism underlying the role of integrin α3ß1 in adhesive dysfunction between thyroid cells induced by diesel engine exhaust particles.

The role and mechanisms of DEP exposure on thyroid injury are not yet clear. This study explores thyroid damage induced by in vivo DEP exposure using a mouse model. This study has observed alterations in thyroid follicular architecture, including rupture, colloid overflow, and the formation of voids. Additionally, there was a significant decrease in the expression levels of proteins involved in thyroid hormone synthesis, such as thyroid peroxidase and thyroglobulin, their trend of change is consistent with the damage to the thyroid structure. Serum levels of triiodothyronine and tetraiodothyronine were raise. However, the decrease in TSH expression suggests that the function of the HPT axis is unaffected. To delve deeper into the intrinsic mechanisms of thyroid injury, we performed KEGG pathway enrichment analysis, which revealed notable alterations in the cell adhesion signaling pathway. Our immunofluorescence results show that DEP exposure impairs thyroid adhesion, and integrin α3ß1 plays an important role. CD151 binds to α3ß1, promoting multimolecular complex formation and activating adhesion-dependent small GTPases. Our in vitro model has confirmed the pivotal role of integrin α3ß1 in thyroid cell adhesion, which may be mediated by the CD151/α3ß1/Rac1 pathway. In summary, exposure to DEP disrupts the structure and function of the thyroid, a process that likely involves the regulation of cell adhesion through the CD151/α3ß1/Rac1 pathway, leading to glandular damage.

Nanodrug-bacteria conjugates-mediated oncogenic collagen depletion enhances immune checkpoint blockade therapy against pancreatic cancer.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) cancer cells specifically produce abnormal oncogenic collagen to bind with integrin α3ß1 receptor and activate the downstream focal adhesion kinase (FAK), protein kinase B (AKT), and mitogen-activated protein kinase (MAPK) signaling pathway. Collectively, this promotes immunosuppression and tumor proliferation and restricts the response rate of clinical cancer immunotherapies. METHODS: Here, by leveraging the hypoxia tropism and excellent motility of the probiotic Escherichia coli strain Nissle 1917 (ECN), we developed nanodrug-bacteria conjugates to penetrate the extracellular matrix (ECM) and shuttle the surface-conjugated protein cages composed of collagenases and anti-programmed death-ligand 1 (PD-L1) antibodies to PDAC tumor parenchyma. FINDINGS: We found the oncogenic collagen expression in human pancreatic cancer patients and demonstrated its interaction with integrin α3ß1. We proved that reactive oxygen species (ROS) in the microenvironment of PDAC triggered collagenase release to degrade oncogenic collagen and block integrin α3ß1-FAK signaling pathway, thus overcoming the immunosuppression and synergizing with anti-PD-L1 immunotherapy. CONCLUSIONS: Collectively, our study highlights the significance of oncogenic collagen in PDAC immunotherapy, and consequently, we developed a therapeutic strategy that can deplete oncogenic collagen to synergize with immune checkpoint blockade for enhanced PDAC treatment efficacy. FUNDING: This work was supported by the University of Wisconsin Carbone Cancer Center Research Collaborative and Pancreas Cancer Research Task Force, UWCCC Transdisciplinary Cancer Immunology-Immunotherapy Pilot Project, and the start-up package from the University of Wisconsin-Madison (to Q.H.).

The Role of α3ß1 Integrin Modulation on Fabry Disease Podocyte Injury and Kidney Impairment.

Podocyte dysfunction plays a crucial role in renal injury and is identified as a key contributor to proteinuria in Fabry disease (FD), primarily impacting glomerular filtration function (GFF). The α3ß1 integrins are important for podocyte adhesion to the glomerular basement membrane, and disturbances in these integrins can lead to podocyte injury. Therefore, this study aimed to assess the effects of chloroquine (CQ) on podocytes, as this drug can be used to obtain an in vitro condition analogous to the FD. Murine podocytes were employed in our experiments. The results revealed a dose-dependent reduction in cell viability. CQ at a sub-lethal concentration (1.0 µg/mL) induced lysosomal accumulation significantly (p < 0.0001). Morphological changes were evident through scanning electron microscopy and immunofluorescence, highlighting alterations in F-actin and nucleus morphology. No significant changes were observed in the gene expression of α3ß1 integrins via RT-qPCR. Protein expression of α3 integrin was evaluated with Western Blotting and immunofluorescence, demonstrating its lower detection in podocytes exposed to CQ. Our findings propose a novel in vitro model for exploring secondary Fabry nephropathy, indicating a modulation of α3ß1 integrin and morphological alterations in podocytes under the influence of CQ.