Yoshiya Ito

Background/Aim: Endometriosis is characterized by the accumulation of immune cells in endometrial lesions and the peritoneal cavity. Macrophages contribute to the growth and neovascularization of endometriotic lesions. Vascular endothelial growth factor receptor-1 (VEGFR1) is involved in neovascularization, while peritoneal macrophages (PMs) play a critical role in endometriosis development and establishment. We examined the role of VEGFR1 signaling in PMs during endometriosis development using a murine model of ectopic endometrial transplantation. Materials and Methods: Endometrial fragments from female wild-type (WT) or VEGFR1 tyrosine kinase-deficient (TK^−/−) donor mice were implanted into the peritoneal walls of recipient mice, either in a WT→WT or TK^−/−→TK^−/− combination. On day 14 after endometrial transplantation, the implant size, neovascular growth-promoting factors, macrophage accumulation in the implants and peritoneal cavity, and cytokine production were assessed. PMs from WT or TK^−/− mice were transferred into the peritoneal cavity of WT→WT mice and their effects were assessed. Results: Compared to WT→WT mice, TK^−/−→TK^−/− mice exhibited smaller implant sizes and reduced neovascularization, including angiogenesis and lymphangiogenesis. This was correlated with an increase in pro-inflammatory (M1) and a decrease in alternative (M2) large peritoneal macrophages (LPMs) within the peritoneal cavity. Transfer of TK^−/−-PMs into the peritoneal cavity of WT→WT mice reduced endometriosis development and macrophage accumulation. This led to increased expression of M1 macrophage genes and decreased expression of M2 phenotype genes, compared to WT-PMs transfer. PMs from TK^−/− mice exhibited increased M1-related and decreased M2-related gene expression. Conclusion: Deletion of VEGFR1 TK signaling in PMs suppressed endometriosis progression and neovascularization by increasing M1 LPMs. Specific inactivation of VEGFR1 TK signaling may represent a potential therapeutic target for the management of endometriosis.

Acute respiratory distress syndrome (ARDS) is a life-threatening lung injury that induces cytokine hypersecretion. Receptor activity-modifying protein (RAMP) 1, a subunit of the calcitonin gene-related peptide (CGRP) receptor, regulates the production of cytokines. This study examined the role of RAMP1 signaling during lipopolysaccharide (LPS)-induced acute lung injury (ALI). LPS administration to wild-type (WT) mice depleted alveolar macrophages (AMs) and recruited monocyte-derived macrophages (MDMs) and neutrophils. RAMP1-deficient (RAMP1^−/−) mice exhibited higher lung injury scores, cytokine levels, and cytokine-producing neutrophil infiltration. RAMP1-deficient AMs produced more cytokines in response to LPS than WT AMs. Adoptive transfer of RAMP1-deficient AMs to RAMP1^−/− mice increased cytokine levels and neutrophil accumulation compared to the transfer of WT AMs. RAMP1^−/− mice had reduced MDM recruitment and lower pro-inflammatory and reparative macrophage profiles. Cultured bone marrow (BM)-derived RAMP1-deficient macrophages stimulated with LPS showed decreased expression of pro-inflammatory and pro-repairing genes. CGRP administration to WT mice reduced cytokine production and neutrophil accumulation. These findings indicate that RAMP1 signaling mitigates LPS-induced ALI by inactivating AMs and promoting inflammatory and repair activities of MDMs. Targeting RAMP1 signaling presents a potential therapeutic approach for the treatment of ARDS.

Background/Aim: The liver effectively restores both size and function following partial hepatectomy (PHx). Angiogenesis is crucial for the repair and regeneration of liver tissue post-PHx. Calcitonin gene-related peptide (CGRP) released from sensory nerves and its receptor—receptor activity-modifying protein 1 (RAMP1) are involved in angiogenesis. This study aimed to assess the role of RAMP1 signaling in angiogenesis during liver regeneration following PHx. Materials and Methods: RAMP1 deficient (RAMP1^−/−) and wild-type (WT) mice were subjected to PHx. Results: RAMP1^−/− mice demonstrated delayed liver regeneration, indicated by lower liver-to-body weight ratios compared to WT mice. This was associated with lower levels of Ki67⁺ hepatocytes and hepatic trophic growth factors. Additionally, RAMP1^−/− mice exhibited lower levels of endothelial cell markers, including CD31, compared to WT mice. This reduction was associated with reduced levels of vascular endothelial growth factor (VEGF)-C, VEGF-D, and VEGF receptor 3 (VEGFR3). In WT mice with PHx, the administration of a VEGFR3 inhibitor reduced the liver-to-body weight ratio, Ki67⁺ hepatocytes, and VEGF-C/VEGFR3 expression levels in the liver compared to those in the vehicle-treated group. Conclusion: The deletion of RAMP1 signaling suppresses liver regeneration and angiogenesis through VEGFR3. Specific activation of RAMP1 signaling may represent a potential therapeutic strategy for liver regeneration following PHx.

Background/Aim: Intestinal lymphatic vessels (lacteals) play a critical role in the absorption and transport of dietary lipids into the circulation. Calcitonin gene-related peptide and receptor activity-modifying protein 1 (RAMP1) are involved in lymphatic vessel growth. This study aimed to examine the role of RAMP1 signaling in lacteal morphology and function in response to a high-fat diet (HFD). Materials and Methods: RAMP1 deficient (RAMP1^−/−) or wild-type (WT) mice were fed a normal diet (ND) or HFD for 8 weeks. Results: RAMP1^−/− mice fed a HFD had increased body weights compared to WT mice fed a HFD, which was associated with high levels of total cholesterol, triglycerides, and glucose. HFD-fed RAMP1^−/− mice had shorter and wider lacteals than HFD-fed WT mice. HFD-fed RAMP1^−/− mice had lower levels of lymphatic endothelial cell gene markers including vascular endothelial growth factor receptor 3 (VEGFR3) and lymphatic vascular growth factor VEGF-C than HFD-fed WT mice. The concentration of an absorbed lipid tracer in HFD-fed RAMP1^−/− mice was higher than that in HFD-fed WT mice. The zipper-like continuous junctions were predominant in HFD-fed WT mice, while the button-like discontinuous junctions were predominant in HFD-fed RAMP1^−/− mice. Conclusion: Deletion of RAMP1 signaling suppressed lacteal growth and VEGF-C/VEGFR3 expression but accelerated the uptake and transport of dietary fats through discontinuous junctions of lacteals, leading to excessive obesity. Specific activation of RAMP1 signaling may represent a target for the therapeutic management of diet-induced obesity.

Background: Accumulating evidence suggests that prostaglandin E₂, an arachidonic acid (AA) metabolite, enhances lymphangiogenesis in response to inflammation. However, thromboxane A₂ (TXA₂), another AA metabolite, is not well known. Thus, this study aimed to determine the role of thromboxane prostanoid (TP) signaling in lymphangiogenesis in secondary lymphedema. Methods and results: Lymphedema was induced by the ablation of lymphatic vessels in mouse tails. Compared with wild-type mice, tail lymphedema in Tp-deficient mice was enhanced, which was associated with suppressed lymphangiogenesis as indicated by decreased lymphatic vessel area and pro-lymphangiogenesis-stimulating factors. Numerous macrophages were found in the tail tissues of Tp-deficient mice. Furthermore, the deletion of TP in macrophages increased tail edema and decreased lymphangiogenesis and pro-lymphangiogenic cytokines, which was accompanied by increased numbers of macrophages and gene expression related to a pro-inflammatory macrophage phenotype in tail tissues. In vivo microscopic studies revealed fluorescent dye leakage in the lymphatic vessels in the wounded tissues. Conclusions: The results suggest that TP signaling in macrophages promotes lymphangiogenesis and prevents tail lymphedema. TP signaling may be a therapeutic target for improving lymphedema-related symptoms by enhancing lymphangiogenesis.

The liver displays a remarkable regenerative capacity in response to acute liver injury. In addition to the proliferation of hepatocytes during liver regeneration, non-parenchymal cells, including liver macrophages, liver sinusoidal endothelial cells (LSECs), and hepatic stellate cells (HSCs) play critical roles in liver repair and regeneration. Liver ischemia–reperfusion injury (IRI) is a major cause of increased liver damage during liver resection, transplantation, and trauma. Impaired liver repair increases postoperative morbidity and mortality of patients who underwent liver surgery. Successful liver repair and regeneration after liver IRI requires coordinated interplay and synergic actions between hepatic resident cells and recruited cell components. However, the underlying mechanisms of liver repair after liver IRI are not well understood. Recent technological advances have revealed the heterogeneity of each liver cell component in the steady state and diseased livers. In this review, we describe the progress in the biology of liver non-parenchymal cells obtained from novel technological advances. We address the functional role of each cell component in response to liver IRI and the interactions between diverse immune repertoires and non-hematopoietic cell populations during the course of liver repair after liver IRI. We also discuss how these findings can help in the design of novel therapeutic approaches. Growing insights into the cellular interactions during liver IRI would enhance the pathology of liver IRI understanding comprehensively and further develop the strategies for improvement of liver repair.

Macrophage polarization is critical for liver tissue repair following acute liver injury. However, the underlying mechanisms of macrophage phenotype switching are not well defined. Invariant natural killer T (iNKT) cells orchestrate tissue inflammation and tissue repair by regulating cytokine production. Herein, we examined whether iNKT cells played an important role in liver repair after hepatic ischemia-reperfusion (I/R) injury by affecting macrophage polarization. To this end, we subjected male C57BL/6 mice to hepatic I/R injury, and mice received an intraperitoneal (ip) injection of α-galactosylceramide (α-GalCer) or vehicle. Compared with that of the vehicle, α-GalCer administration resulted in the promotion of liver repair accompanied by acceleration of macrophage differentiation and by increases in the numbers of Ly6C^high pro-inflammatory macrophages and Ly6C^low reparative macrophages. iNKT cells activated with α-GalCer produced interleukin (IL)-4 and interferon (IFN)-γ. Treatment with anti-IL-4 antibodies delayed liver repair, which was associated with an increased number of Ly6C^high macrophages and a decreased number of Ly6C^low macrophages. Treatment with anti-IFN-γ antibodies promoted liver repair, associated with reduced the number of Ly6C^high macrophages, but did not change the number of Ly6C^low macrophages. Bone marrow-derived macrophages up-regulated the expression of genes related to both a pro-inflammatory and a reparative phenotype when co-cultured with activated iNKT cells. Anti-IL-4 antibodies increased the levels of pro-inflammatory macrophage-related genes and decreased those of reparative macrophage-related genes in cultured macrophages, while anti-IFN-γ antibodies reversed the polarization of macrophages. Cd1d-deficient mice showed delayed liver repair and suppressed macrophage switching, compared with that in wild-type mice. These results suggest that the activation of iNKT cells by α-GalCer facilitated liver repair after hepatic I/R injury by both IL-4-and IFN-γ-mediated acceleration of macrophage polarization. Therefore, the activation of iNKT cells may represent a therapeutic tool for liver repair after hepatic I/R injury.

Vascular endothelial growth factor (VEGF) is not only an important factor for angiogenesis but also lung development and homeostasis. VEGF-A binds three tyrosine kinase (TK) receptors VEGFR1–3. Idiopathic pulmonary fibrosis (IPF) is one of the poor prognoses of lung diseases. The relationship of VEGF and IPF remains to be clarified. Treatment with nintedanib used for the treatment of IPF reduced fibroblast proliferation, inhibited TK receptors, platelet-derived growth factor receptor (PDGFR), fibroblast growth factor receptor (FGFR), and VEGFR. Because the effect of that treatment is still not satisfactory, the emergence of new therapeutic agents is needed. This review describes the enhancement of pulmonary fibrosis by VEGFR1-TK signal and suggests that the blocking of the VEGFR1-TK signal may be useful for the treatment of pulmonary fibrosis.

Neuroimmune interactions are involved in the development of endometriosis. Here, we examined the role of a neuropeptide, calcitonin gene–related peptide (CGRP), and its receptor, receptor activity–modifying protein (RAMP) 1, in growth of endometrial tissues and the formation of blood and lymphatic vessels in a mouse ectopic endometrial transplantation model. Endometrial fragments from donor wild‐type (WT) mice transplanted into the peritoneal wall of recipient WT mice grew with increased density of blood and lymphatic vessels. When tissues from RAMP1‐deficient (RAMP1^−/−) mice were transplanted into RAMP1^−/− mice, implant growth and angiogenesis/lymphangiogenesis were decreased. CGRP was up‐regulated in dorsal root ganglia, and CGRP⁺ nerve fibres were distributed into the implants from the peritoneum. RAMP1 was co‐expressed with CD11b (macrophages) and S100A4 (fibroblasts), but did not co‐localize with blood vessel endothelial cell marker CD31 or lymphatic vessel endothelial hyaluronan receptor (LYVE)‐1. Cultured with CGRP, macrophages up‐regulated vascular endothelial growth factor (VEGF)‐A, VEGF‐C and VEGF‐D, whereas fibroblasts up‐regulated VEGF‐C, but not VEGF‐A or VEGF‐D, in a RAMP1‐dependent manner. CGRP receptor antagonist CGRP8‐37 inhibited growth of and angiogenesis/lymphangiogenesis within endometrial tissue implants. These results suggest that RAMP1 signalling is crucial for growth and angiogenesis/lymphangiogenesis in endometrial tissue. Blockade of RAMP1 is a potential tool for the treatment of endometriosis.