Mechanoimmune-Driven Backpack Sustains Dendritic Cell Maturation for Synergistic Tumor Radiotherapy.

Cell backpacks present significant potential in both therapeutic and diagnostic applications, making it essential to further explore their interactions with host cells. Current evidence indicates that backpacks can induce sustained immune responses. Our original objective was to incorporate a model antigen into the backpacks to promote dendritic cell maturation and facilitate antigen presentation, thereby inducing immune responses. However, we unexpectedly discovered that both antigen-loaded backpacks and empty backpacks demonstrated comparable abilities to induce dendritic cell maturation, resulting in nearly identical potency in T-cell proliferation. Our mechanistic studies suggest that the attachment of backpacks induces mechanical forces on dendritic cells via opening the PIEZO1 mechanical ion channel. This interaction leads to the remodeling of the intracellular cytoskeleton and facilitates the production of type I interferons by dendritic cells. Consequently, the mechano-immune-driven dendritic cell backpacks, when combined with radiotherapy, induce a robust antitumor effect. This research presents an avenue for leveraging mechanotransduction to enhance combination immunotherapeutic strategies, potentially leading to groundbreaking advancements in the field.

Research progress on the immunological functions of Piezo1 a receptor molecule that responds to mechanical force.

The human immune system is capable of defending against, monitoring, and self-stabilizing various immune cells. Differentiation, proliferation, and development of these cells are regulated by biochemical signals. Moreover, biophysical signals, such as mechanical forces, have been found to affect immune cell function, thus introducing a new area of immunological research. Piezo1, a mechanically sensitive ion channel, was awarded the Nobel Prize for Physiology and Medicine in 2021. This channel is present on the surface of many cells, and when stimulated by mechanical force, it controls calcium (Ca2+) inside the cells, leading to changes in downstream signals and thus regulating cell functions. Piezo1 is also expressed in various innate and adaptive immune cells and plays a major role in the immune function. In this review, we will explore the physiological functions and regulatory mechanisms of Piezo1 and its impact on innate and adaptive immunity. This may offer new insights into diagnostics and therapeutics for the prevention and treatment of diseases and surgical infections.

Tethering Piezo channels to the actin cytoskeleton for mechanogating via the cadherin-ß-catenin mechanotransduction complex.

The mechanically activated Piezo channel plays a versatile role in conferring mechanosensitivity to various cell types. However, how it incorporates its intrinsic mechanosensitivity and cellular components to effectively sense long-range mechanical perturbation across a cell remains elusive. Here we show that Piezo channels are biochemically and functionally tethered to the actin cytoskeleton via the cadherin-ß-catenin mechanotransduction complex, whose perturbation significantly impairs Piezo-mediated responses. Mechanistically, the adhesive extracellular domain of E-cadherin interacts with the cap domain of Piezo1, which controls the transmembrane gate, while its cytosolic tail might interact with the cytosolic domains of Piezo1, which are in close proximity to its intracellular gates, allowing a direct focus of adhesion-cytoskeleton-transmitted force for gating. Specific disruption of the intermolecular interactions prevents cytoskeleton-dependent gating of Piezo1. Thus, we propose a force-from-filament model to complement the previously suggested force-from-lipids model for mechanogating of Piezo channels, enabling them to serve as versatile and tunable mechanotransducers.

IQGAP1-mediated mechanical signaling promotes the foreign body response to biomedical implants.

The aim of this study was to further elucidate the molecular mechanisms that mediate pathologic foreign body response (FBR) to biomedical implants. The longevity of biomedical implants is limited by the FBR, which leads to implant failure and patient morbidity. Since the specific molecular mechanisms underlying fibrotic responses to biomedical implants have yet to be fully described, there are currently no targeted approaches to reduce pathologic FBR. We utilized proteomics analysis of human FBR samples to identify potential molecular targets for therapeutic inhibition of FBR. We then employed a murine model of FBR to further evaluate the role of this potential target. We performed histological and immunohistochemical analysis on the murine FBR capsule tissue, as well as single-cell RNA sequencing (scRNA-seq) on cells isolated from the capsules. We identified IQ motif containing GTPase activating protein 1 (IQGAP1) as the most promising of several targets, serving as a central molecular mediator in human and murine FBR compared to control subcutaneous tissue. IQGAP1-deficient mice displayed a significantly reduced FBR compared to wild-type mice as evidenced by lower levels of collagen deposition and maturity. Our scRNA-seq analysis revealed that decreasing IQGAP1 resulted in diminished transcription of mechanotransduction, inflammation, and fibrosis-related genes, which was confirmed on the protein level with immunofluorescent staining. The deficiency of IQGAP1 significantly attenuates FBR by deactivating downstream mechanotransduction signaling, inflammation, and fibrotic pathways. IQGAP1 may be a promising target for rational therapeutic design to mitigate pathologic FBR around biomedical implants.

Stretching the Function of Innate Immune Cells.

The importance of innate immune cells to sense and respond to their physical environment is becoming increasingly recognized. Innate immune cells (e.g. macrophages and neutrophils) are able to receive mechanical signals through several mechanisms. In this review, we discuss the role of mechanosensitive ion channels, such as Piezo1 and transient receptor potential vanilloid 4 (TRPV4), and cell adhesion molecules, such as integrins, selectins, and cadherins in biology and human disease. Furthermore, we explain that these mechanical stimuli activate intracellular signaling pathways, such as MAPK (p38, JNK), YAP/TAZ, EDN1, NF-kB, and HIF-1α, to induce protein conformation changes and modulate gene expression to drive cellular function. Understanding the mechanisms by which immune cells interpret mechanosensitive information presents potential targets to treat human disease. Important areas of future study in this area include autoimmune, allergic, infectious, and malignant conditions.

The Multiple Faces of Integrin-ECM Interactions in Inflammatory Bowel Disease.

Inflammatory Bowel Disease (IBD) comprises a series of chronic and relapsing intestinal diseases, with Crohn's disease and ulcerative colitis being the most common. The abundant and uncontrolled deposition of extracellular matrix, namely fibrosis, is one of the major hallmarks of IBD and is responsible for the progressive narrowing and closure of the intestine, defined as stenosis. Although fibrosis is usually considered the product of chronic inflammation, the substantial failure of anti-inflammatory therapies to target and reduce fibrosis in IBD suggests that fibrosis might be sustained in an inflammation-independent manner. Pharmacological therapies targeting integrins have recently shown great promise in the treatment of IBD. The efficacy of these therapies mainly relies on their capacity to target the integrin-mediated recruitment and functionality of the immune cells at the damage site. However, by nature, integrins also act as mechanosensitive molecules involved in the intracellular transduction of signals and modifications originating from the extracellular matrix. Therefore, understanding integrin signaling in the context of IBD may offer important insights into mechanisms of matrix remodeling, which are uncoupled from inflammation and could underlie the onset and persistency of intestinal fibrosis. In this review, we present the currently available knowledge on the role of integrins in the etiopathogenesis of IBD, highlighting their role in the context of immune-dependent and independent mechanisms.

Implant Fibrosis and the Underappreciated Role of Myofibroblasts in the Foreign Body Reaction.

Body implants and implantable medical devices have dramatically improved and prolonged the life of countless patients. However, our body repair mechanisms have evolved to isolate, reject, or destroy any object that is recognized as foreign to the organism and inevitably mounts a foreign body reaction (FBR). Depending on its severity and chronicity, the FBR can impair implant performance or create severe clinical complications that will require surgical removal and/or replacement of the faulty device. The number of review articles discussing the FBR seems to be proportional to the number of different implant materials and clinical applications and one wonders, what else is there to tell? We will here take the position of a fibrosis researcher (which, coincidentally, we are) to elaborate similarities and differences between the FBR, normal wound healing, and chronic healing conditions that result in the development of peri-implant fibrosis. After giving credit to macrophages in the inflammatory phase of the FBR, we will mainly focus on the activation of fibroblastic cells into matrix-producing and highly contractile myofibroblasts. While fibrosis has been discussed to be a consequence of the disturbed and chronic inflammatory milieu in the FBR, direct activation of myofibroblasts at the implant surface is less commonly considered. Thus, we will provide a perspective how physical properties of the implant surface control myofibroblast actions and accumulation of stiff scar tissue. Because formation of scar tissue at the surface and around implant materials is a major reason for device failure and extraction surgeries, providing implant surfaces with myofibroblast-suppressing features is a first step to enhance implant acceptance and functional lifetime. Alternative therapeutic targets are elements of the myofibroblast mechanotransduction and contractile machinery and we will end with a brief overview on such targets that are considered for the treatment of other organ fibroses.

Mechanically activated ion channel Piezo1 modulates macrophage polarization and stiffness sensing.

Macrophages perform diverse functions within tissues during immune responses to pathogens and injury, but molecular mechanisms by which physical properties of the tissue regulate macrophage behavior are less well understood. Here, we examine the role of the mechanically activated cation channel Piezo1 in macrophage polarization and sensing of microenvironmental stiffness. We show that macrophages lacking Piezo1 exhibit reduced inflammation and enhanced wound healing responses. Additionally, macrophages expressing the transgenic Ca2+ reporter, Salsa6f, reveal that Ca2+ influx is dependent on Piezo1, modulated by soluble signals, and enhanced on stiff substrates. Furthermore, stiffness-dependent changes in macrophage function, both in vitro and in response to subcutaneous implantation of biomaterials in vivo, require Piezo1. Finally, we show that positive feedback between Piezo1 and actin drives macrophage activation. Together, our studies reveal that Piezo1 is a mechanosensor of stiffness in macrophages, and that its activity modulates polarization responses.

Inflammatory signaling sensitizes Piezo1 mechanotransduction in articular chondrocytes as a pathogenic feed-forward mechanism in osteoarthritis.

Osteoarthritis (OA) is a painful and debilitating condition of synovial joints without any disease-modifying therapies [A. M. Valdes, T. D. Spector, Nat. Rev. Rheumatol. 7, 23-32 (2011)]. We previously identified mechanosensitive PIEZO channels, PIEZO1 and PIEZO2, both expressed in articular cartilage, to function in chondrocyte mechanotransduction in response to injury [W. Lee et al., Proc. Natl. Acad. Sci. U.S.A. 111, E5114-E5122 (2014); W. Lee, F. Guilak, W. Liedtke, Curr. Top. Membr. 79, 263-273 (2017)]. We therefore asked whether interleukin-1-mediated inflammatory signaling, as occurs in OA, influences Piezo gene expression and channel function, thus indicative of maladaptive reprogramming that can be rationally targeted. Primary porcine chondrocyte culture and human osteoarthritic cartilage tissue were studied. We found that interleukin-1α (IL-1α) up-regulated Piezo1 in porcine chondrocytes. Piezo1 expression was significantly increased in human osteoarthritic cartilage. Increased Piezo1 expression in chondrocytes resulted in a feed-forward pathomechanism whereby increased function of Piezo1 induced excess intracellular Ca2+ at baseline and in response to mechanical deformation. Elevated resting state Ca2+ in turn rarefied the F-actin cytoskeleton and amplified mechanically induced deformation microtrauma. As intracellular substrates of this OA-related inflammatory pathomechanism, in porcine articular chondrocytes exposed to IL-1α, we discovered that enhanced Piezo1 expression depended on p38 MAP-kinase and transcription factors HNF4 and ATF2/CREBP1. CREBP1 directly bound to the proximal PIEZO1 gene promoter. Taken together, these signaling and genetic reprogramming events represent a detrimental Ca2+-driven feed-forward mechanism that can be rationally targeted to stem the progression of OA.

Gateway Reflex and Mechanotransduction.

The gateway reflex explains how autoreactive CD4+ T cells cause inflammation in tissues that have blood-barriers, such as the central nervous system and retina. It depends on neural activations in response to specific external stimuli, such as gravity, pain, stress, and light, which lead to the secretion of noradrenaline at specific vessels in the tissues. Noradrenaline activates NFkB at these vessels, followed by an increase of chemokine expression as well as a reduction of tight junction molecules to accumulate autoreactive CD4+ T cells, which breach blood-barriers. Transient receptor potential vanilloid 1 (TRPV1) molecules on sensory neurons are critical for the gateway reflex, indicating the importance of mechano-sensing. In this review, we overview the gateway reflex with a special interest in mechanosensory transduction (mechanotransduction).

Alveolar cells under mechanical stressed niche: critical contributors to pulmonary fibrosis.

Pulmonary fibrosis arises from the repeated epithelial mild injuries and insufficient repair lead to over activation of fibroblasts and excessive deposition of extracellular matrix, which result in a mechanical stretched niche. However, increasing mechanical stress likely exists before the establishment of fibrosis since early micro injuries increase local vascular permeability and prompt cytoskeletal remodeling which alter cellular mechanical forces. It is noteworthy that COVID-19 patients with severe hypoxemia will receive mechanical ventilation as supportive treatment and subsequent pathology studies indicate lung fibrosis pattern. At advanced stages, mechanical stress originates mainly from the stiff matrix since boundaries between stiff and compliant parts of the tissue could generate mechanical stress. Therefore, mechanical stress has a significant role in the whole development process of pulmonary fibrosis. The alveoli are covered by abundant capillaries and function as the main gas exchange unit. Constantly subject to variety of damages, the alveolar epithelium injuries were recently recognized to play a vital role in the onset and development of idiopathic pulmonary fibrosis. In this review, we summarize the literature regarding the effects of mechanical stress on the fundamental cells constituting the alveoli in the process of pulmonary fibrosis, particularly on epithelial cells, capillary endothelial cells, fibroblasts, mast cells, macrophages and stem cells. Finally, we briefly review this issue from a more comprehensive perspective: the metabolic and epigenetic regulation.

Mechanosensing through YAP controls T cell activation and metabolism.

Upon immunogenic challenge, lymph nodes become mechanically stiff as immune cells activate and proliferate within their encapsulated environments, and with resolution, they reestablish a soft baseline state. Here we show that sensing these mechanical changes in the microenvironment requires the mechanosensor YAP. YAP is induced upon activation and suppresses metabolic reprogramming of effector T cells. Unlike in other cell types in which YAP promotes proliferation, YAP in T cells suppresses proliferation in a stiffness-dependent manner by directly restricting the translocation of NFAT1 into the nucleus. YAP slows T cell responses in systemic viral infections and retards effector T cells in autoimmune diabetes. Our work reveals a paradigm whereby tissue mechanics fine-tune adaptive immune responses in health and disease.

Tensile force transmitted through LFA-1 bonds mechanoregulate neutrophil inflammatory response.

Recruitment of leukocytes to sites of acute inflammation is guided by spatial and temporal cues that ensure appropriate cell numbers infiltrate the tissue at precise locations to protect it from infection and initiate repair. On inflamed endothelium, neutrophil rolling via selectins elicits cytosolic calcium release from endoplasmic reticulum (ER)-stores that are synergistic with chemokine signaling to activate formation of high affinity (HA) LFA-1 bonds to ICAM-1, which is necessary to anchor cells against the drag force of blood flow. Bond tension on LFA-1 within the area of adhesive contact with endothelium elicits calcium entry through calcium release-activated calcium channel protein 1 (Orai-1) membrane channels that in turn activate neutrophil shape change and migration. We hypothesized that mechanotransduction via LFA-1 is mediated by assembly of a cytosolic molecular complex consisting of Kindlin-3, receptor for activated C kinase 1 (RACK1), and Orai1. Initiation of Ca2+ flux at sites of adhesive contact required a threshold level of shear stress and increased with the magnitude of bond tension transduced across as few as 200 HA LFA-1. A sequential mechanism triggered by force acting on LFA-1/Kindlin-3 precipitated dissociation of RACK1, which formed a concentration gradient above LFA-1 bond clusters. This directed translocation of ER proximal to Orai1, where binding of inositol 1,4,5-triphosphate receptor type 1 and activation via stromal interaction molecule 1 elicited Ca flux and subsequent neutrophil shape change and motility. We conclude that neutrophils sense adhesive traction on LFA-1 bonds on a submicron scale to direct calcium influx, thereby ensuring sufficient shear stress of blood flow is present to trigger cell arrest and initiate transmigration at precise regions of vascular inflammation.

TRPV4-A Missing Link Between Mechanosensation and Immunity.

Transient receptor potential vanilloid-type 4 (TRPV4) cation channel is widely expressed in all tissues as well as in immune cells and its function as mechanosensitive Ca2+ channel seems to be conserved throughout all mammalian species. Of late, emerging evidence has implicated TRPV4 in the activation and differentiation of innate immune cells, especially in neutrophils, monocytes, and macrophages. As such, TRPV4 has been shown to mediate neutrophil adhesion and chemotaxis, as well as production of reactive oxygen species in response to pro-inflammatory stimuli. In macrophages, TRPV4 mediates formation of both reactive oxygen and nitrogen species, and regulates phagocytosis, thus facilitating bacterial clearance and resolution of infection. Importantly, TRPV4 may present a missing link between mechanical forces and immune responses. This connection has been exemplary highlighted by the demonstrated role of TRPV4 in macrophage activation and subsequent induction of lung injury following mechanical overventilation. Mechanosensation via TRPV4 is also expected to activate innate immune cells and establish a pro-inflammatory loop in fibrotic diseases with increased deposition of extracellular matrix (ECM) and substrate stiffness. Likewise, TRPV4 may be activated by cell migration through the endothelium or the extracellular matrix, or even by circulating immune cells squeezing through the narrow passages of the pulmonary or systemic capillary bed, a process that has recently been linked to neutrophil priming and depriming. Here, we provide an overview over the emerging role of TRPV4 in innate immune responses and highlight two distinct modes for the activation of TRPV4 by either mechanical forces ("mechanoTRPV4") or by pathogens ("immunoTRPV4").

B cell mechanosensing: A mechanistic overview.

B cells are essential to the adaptive immune system for providing the humoral immunity against cohorts of pathogens. The presentation of antigen to the B cell receptor (BCR) leads to the initiation of B cell activation, which is a process sensitive to the stiffness features of the substrates presenting the antigens. Mechanosensing of the B cells, potentiated through BCR signaling and the adhesion molecules, efficiently regulates B cell activation, proliferation and subsequent antibody responses. Defects in sensing of the antigen-presenting substrates can lead to the activation of autoreactive B cells in autoimmune diseases. The use of high-resolution, high-speed live-cell imaging along with the sophisticated biophysical materials, has uncovered the mechanisms underlying the initiation of B cell activation within seconds of its engagement with the antigen presenting substrates. In this chapter, we reviewed studies that have contributed to uncover the molecular mechanisms of B cell mechanosensing during the initiation of B cell activation.

Mechanosensing by Peyer's patch stroma regulates lymphocyte migration and mucosal antibody responses.

Fibroblastic reticular cells (FRCs) and their specialized collagen fibers termed 'conduits' form fundamental structural units supporting lymphoid tissues. In lymph nodes, conduits are known to transport interstitial fluid and small molecules from afferent lymphatics into the nodal parenchyma. However, the immunological contributions of conduit function have remained elusive. Here, we report that intestinal Peyer's patches (PPs) contain a specialized conduit system that directs the flow of water absorbed across the intestinal epithelium. Notably, PP FRCs responded to conduit fluid flow via the mechanosensitive ion channel Piezo1. Disruption of fluid flow or genetic deficiency of Piezo1 on CCL19-expressing stroma led to profound structural alterations in perivascular FRCs and associated high endothelial venules. This in turn impaired lymphocyte entry into PPs and initiation of mucosal antibody responses. These results identify a critical role for conduit-mediated fluid flow in the maintenance of PP homeostasis and mucosal immunity.

Ex vivo physiological compression of human osteoarthritis cartilage modulates cellular and matrix components.

Mechanical stimulation appears to play a key role in cartilage homeostasis maintenance, but it can also contribute to osteoarthritis (OA) pathogenesis. Accumulating evidence suggests that cartilage loading in the physiological range contributes to tissue integrity maintenance, whereas excessive or reduced loading have catabolic effects. However, how mechanical stimuli can regulate joint homeostasis is still not completely elucidated and few data are available on human cartilage. We aimed at investigating human OA cartilage response to ex vivo loading at physiological intensity. Cartilage explants from ten OA patients were subjected to ex vivo controlled compression, then recovered and used for gene and protein expression analysis of cartilage homeostasis markers. Compressed samples were compared to uncompressed ones in presence or without interleukin 1ß (IL-1ß) or interleukin 4 (IL-4). Cartilage explants compressed in combination with IL-4 treatment showed the best histological scores. Mechanical stimulation was able to significantly modify the expression of collagen type II (collagen 2), aggrecan, SOX9 transcription factor, cartilage oligomeric matrix protein (COMP), collagen degradation marker C2C and vascular endothelial growth factor (VEGF). Conversely, ADAMTS4 metallopeptidase, interleukin 4 receptor alpha (IL4Rα), chondroitin sulfate 846 epitope (CS846), procollagen type 2 C-propeptide (CPII) and glycosaminoglycans (GAG) appeared not modulated. Our data suggest that physiological compression of OA human cartilage modulates the inflammatory milieu by differently affecting the expression of components and homeostasis regulators of the cartilage extracellular matrix.

Mechanosensing through immunoreceptors.

The immune response is orchestrated by a variety of immune cells. The function of each cell is determined by the collective signals from various immunoreceptors, whose expression and activity depend on the developmental stages of the cell and its environmental context. Recent studies have highlighted the presence of mechanical force on several immunoreceptor-ligand pairs and the important role of force in regulating their interaction and function. In this Perspective, we use the T cell antigen receptor as an example with which to review the current understanding of the mechanosensing properties of immunoreceptors. We discuss the types of forces that immunoreceptors may encounter and the effects of force on ligand bonding, conformational change and the triggering of immunoreceptors, as well as the effects of force on the downstream signal transduction, cell-fate decisions and effector function of immune cells.