Local amplifiers of IL-4Rα-mediated macrophage activation promote repair in lung and liver

The type 2 immune response controls helminth infection and maintains tissue homeostasis but can lead to allergy and fibrosis if not adequately regulated. We have discovered local tissue-specific amplifiers of type 2-mediated macrophage activation. In the lung, surfactant protein A (SP-A) enhanced interleukin-4 (IL-4)-dependent macrophage proliferation and activation, accelerating parasite clearance and reducing pulmonary injury after infection with a lung-migrating helminth. In the peritoneal cavity and liver, C1q enhancement of type 2 macrophage activation was required for liver repair after bacterial infection, but resulted in fibrosis after peritoneal dialysis. IL-4 drives production of these structurally related defense collagens, SP-A and C1q, and the expression of their receptor, myosin 18A. These findings reveal the existence within different tissues of an amplification system needed for local type 2 responses

Long-term imaging of cellular forces with high precision by elastic resonator interference stress microscopy

This project has received funding from the Human Frontiers Science Program (RGY0074/2013), the Scottish Funding Council (via SUPA), the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (grant agreement no. 640012), the EPSRC DTP (EP/L505079/1), the RS MacDonald Charitable Trust and the MRC (G1100116 and G110312/1).Cellular forces are crucial for many biological processes but current methods to image them have limitations with respect to data analysis, resolution and throughput. Here, we present a robust approach to measure mechanical cell–substrate interactions in diverse biological systems by interferometrically detecting deformations of an elastic micro-cavity. Elastic resonator interference stress microscopy (ERISM) yields stress maps with exceptional precision and large dynamic range (2 nm displacement resolution over a >1 μm range, translating into 1 pN force sensitivity). This enables investigation of minute vertical stresses (<1 Pa) involved in podosome protrusion, protein-specific cell–substrate interaction and amoeboid migration through spatial confinement in real time. ERISM requires no zero-force reference and avoids phototoxic effects, which facilitates force monitoring over multiple days and at high frame rates and eliminates the need to detach cells after measurements. This allows observation of slow processes such as differentiation and further investigation of cells, for example, by immunostaining.PostprintPeer reviewe

Helminth-induced Th2 cell dysfunction is distinct from exhaustion and is maintained in the absence of antigen

T cell-intrinsic regulation, such as anergy, adaptive tolerance and exhaustion, is central to immune regulation. In contrast to Type 1 and Type 17 settings, knowledge of the intrinsic fate and function of Th2 cells in chronic Type 2 immune responses is lacking. We previously showed that Th2 cells develop a PD-1/PD-L2-dependent intrinsically hypo-responsive phenotype during infection with the filarial nematode Litomosoides sigmodontis, denoted by impaired functionality and parasite killing. This study aimed to elucidate the transcriptional changes underlying Th2 cell-intrinsic hypo-responsiveness, and whether it represents a unique and stable state of Th2 cell differentiation. We demonstrated that intrinsically hypo-responsive Th2 cells isolated from L. sigmodontis infected mice stably retained their dysfunctional Th2 phenotype upon transfer to naïve recipients, and had a divergent transcriptional profile to classical Th2 cells isolated prior to hypo-responsiveness and from mice exposed to acute Type 2 stimuli. Hypo-responsive Th2 cells displayed a distinct transcriptional profile to exhausted CD4+ T cells, but upregulated Blimp-1 and the anergy/regulatory-associated transcription factors Egr2 and c-Maf, and shared characteristics with tolerised T cells. Hypo-responsive Th2 cells increased mRNA expression of the soluble regulatory factors Fgl2, Cd38, Spp1, Areg, Metrnl, Lgals3, and Csf1, and a subset developed a T-bet+IFN-γ+ Th2/Th1 hybrid phenotype, indicating that they were not functionally inert. Contrasting with their lost ability to produce Th2 cytokines, hypo-responsive Th2 cells gained IL-21 production and IL-21R blockade enhanced resistance to L. sigmodontis. IL-21R blockade also increased the proportion of CD19+PNA+ germinal centre B cells and serum levels of parasite specific IgG1. This indicates a novel regulatory role for IL-21 during filarial infection, both in controlling protection and B cell responses. Thus, Th2 cell-intrinsic hypo-responsiveness is a distinct and stable state of Th2 cell differentiation associated with a switch from a classically active IL-4+IL-5+ Th2 phenotype, to a non-classical dysfunctional and potentially regulatory IL-21+Egr2+c-Maf+Blimp-1+IL-4loIL-5loT-bet+IFN-γ+ Th2 phenotype. This divergence towards alternate Th2 phenotypes during chronicity has broad implications for the outcomes and treatment of chronic Type 2-related infections and diseases

Tissue-specific contribution of macrophages to wound healing

AbstractMacrophages are present in all tissues, either as resident cells or monocyte-derived cells that infiltrate into tissues. The tissue site largely determines the phenotype of tissue-resident cells, which help to maintain tissue homeostasis and act as sentinels of injury. Both tissue resident and recruited macrophages make a substantial contribution to wound healing following injury. In this review, we evaluate how macrophages in two fundamentally distinct tissues, i.e. the lung and the skin, differentially contribute to the process of wound healing. We highlight the commonalities of macrophage functions during repair and contrast them with distinct, tissue-specific functions that macrophages fulfill during the different stages of wound healing

Abstracts from the 8th International Conference on cGMP Generators, Effectors and Therapeutic Implications

This work was supported by a restricted research grant of Bayer AG

CCR2 recruits an inflammatory macrophage subpopulation critical for angiogenesis in tissue repair

Monocytes/macrophages are critical in orchestrating the tissue-repair response. However, the mechanisms that govern macrophage regenerative activities during the sequential phases of repair are largely unknown. In the present study, we examined the dynamics and functions of diverse monocyte/macrophage phenotypes during the sequential stages of skin repair. By combining the analysis of a new CCR2-eGFP reporter mouse model with conditional mouse mutants defective in myeloid cell-restricted CCR2 signaling or VEGF-A synthesis, we show herein that among the large number of inflammatory CCR2(+)Ly6C(+) macrophages that dominate the early stage of repair, only a small fraction strongly expresses VEGF-A that has nonredundant functions for the induction of vascular sprouts. The switch of macrophage-derived VEGF-A during the early stage of tissue growth toward epidermal-derived VEGF-A during the late stage of tissue maturation was critical to achieving physiologic tissue vascularization and healing progression. The results of the present study provide new mechanistic insights into CCR2-mediated recruitment of blood monocyte subsets into damaged tissue, the dynamics and functional consequences of macrophage plasticity during the sequential repair phases, and the complementary role of macrophage-derived VEGF-A in coordinating effective tissue growth and vascularization in the context of tissue-resident wound cells. Our findings may be relevant for novel monocyte-based therapies to promote tissue vascularization

Interleukin-4 Receptor alpha Signaling in Myeloid Cells Controls Collagen Fibril Assembly in Skin Repair

Activation of the immune response during injury is a critical early event that determines whether the outcome of tissue restoration is regeneration or replacement of the damaged tissue with a scar. The mechanisms by which immune signals control these fundamentally different regenerative pathways are largely unknown. We have demonstrated that, during skin repair in mice, interleukin-4 receptor alpha (IL-4R alpha)-dependent macrophage activation controlled collagen fibril assembly and that this process was important for effective repair while having adverse pro-fibrotic effects. We identified Relm-alpha as one important player in the pathway from IL-4R alpha signaling in macrophages to the induction of lysyl hydroxylase 2 (LH2), an enzyme that directs persistent pro-fibrotic collagen cross-links, in fibroblasts. Notably, Relm-beta induced LH2 in human fibroblasts, and expression of both factors was increased in lipodermatosclerosis, a condition of excessive human skin fibrosis. Collectively, our findings provide mechanistic insights into the link between type 2 immunity and initiation of pro-fibrotic pathways

Data underpinning - “Long-Term Imaging of Cellular Forces with High Precision by Elastic Resonator Interference Stress Microscopy”

The attached data underpin the publication, “Long-Term Imaging of Cellular Forces with High Precision by Elastic Resonator Interference Stress Microscopy”. Data for each figure panel from the paper that displays numerical data are provided in the form of tagged image format file (.tif)