RESUMO
Background: The impediment to ß-amyloid (Aß) clearance caused by the invalid intracranial lymphatic drainage in Alzheimer's disease is pivotal to its pathogenesis, and finding reliable clinical available solutions to address this challenge remains elusive. Methods: The potential role and underlying mechanisms of intranasal oxytocin administration, an approved clinical intervention, in improving intracranial lymphatic drainage in middle-old-aged APP/PS1 mice were investigated by live mouse imaging, ASL/CEST-MRI scanning, in vivo two-photon imaging, immunofluorescence staining, ELISA, RT-qPCR, Western blotting, RNA-seq analysis, and cognitive behavioral tests. Results: Benefiting from multifaceted modulation of cerebral hemodynamics, aquaporin-4 polarization, meningeal lymphangiogenesis and transcriptional profiles, oxytocin administration normalized the structure and function of both the glymphatic and meningeal lymphatic systems severely impaired in middle-old-aged APP/PS1 mice. Consequently, this intervention facilitated the efficient drainage of Aß from the brain parenchyma to the cerebrospinal fluid and then to the deep cervical lymph nodes for efficient clearance, as well as improvements in cognitive deficits. Conclusion: This work broadens the underlying neuroprotective mechanisms and clinical applications of oxytocin medication, showcasing its promising therapeutic prospects in central nervous system diseases with intracranial lymphatic dysfunction.
Assuntos
Doença de Alzheimer , Peptídeos beta-Amiloides , Modelos Animais de Doenças , Sistema Glinfático , Camundongos Transgênicos , Ocitocina , Animais , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/metabolismo , Camundongos , Ocitocina/farmacologia , Ocitocina/administração & dosagem , Ocitocina/metabolismo , Sistema Glinfático/metabolismo , Sistema Glinfático/efeitos dos fármacos , Peptídeos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/diagnóstico por imagem , Administração Intranasal , Linfangiogênese/efeitos dos fármacos , Masculino , Aquaporina 4/metabolismo , Aquaporina 4/genética , Humanos , Imageamento por Ressonância Magnética , Meninges/metabolismo , Meninges/efeitos dos fármacos , Meninges/diagnóstico por imagemRESUMO
Both lymphatic vessels and macrophages are key factors influencing the inflammatory response. During the inflammatory response, lymphatic vessels undergo dilation and growth, playing a beneficial role in alleviating inflammation by facilitating the drainage of exudate, inflammatory mediators, and leukocytes. Consequently, the promotion of lymphangiogenesis has emerged as a novel therapeutic approach to treating inflammation. Macrophages play a crucial role in promoting lymphangiogenesis by secreting several pro-lymphatic growth factors, including vascular endothelial growth factor (VEGF)-C, and undergoing transdifferentiation into lymphatic endothelial cell progenitors (LECP), which integrate into newly formed lymphatic vessels. Macrophages exhibit heterogeneity and perform diverse functions based on their phenotypes. The regulation of macrophage polarization is crucial in inflammatory responses. Notably, macrophages promote lymphangiogenesis, while lymphatic vessels, in turn, serve as a conduit for macrophages to drain out inflamed tissue and also affect macrophage polarization. Thus, there is an interactive relationship between them. In this review, we discuss current work on the effects of macrophages on lymphangiogenesis as well as lymphatic vessel recruitment of macrophages and regulation of macrophage polarization. Furthermore, we explore the roles of lymphatic vessels and macrophages in various inflammation-related diseases, emphasizing potential therapeutic targets within the context of lymphatic-macrophage interactions.
Assuntos
Inflamação , Linfangiogênese , Vasos Linfáticos , Macrófagos , Macrófagos/imunologia , Macrófagos/metabolismo , Vasos Linfáticos/metabolismo , Vasos Linfáticos/patologia , Humanos , Inflamação/metabolismo , Inflamação/patologia , Animais , Linfangiogênese/fisiologia , Fator C de Crescimento do Endotélio Vascular/metabolismoRESUMO
BACKGROUND: Lymphatic vessels (LVs) play a crucial role in immune reactions by serving as the principal conduits for immune cells. However, to date, no study has analyzed the morphological changes in the LVs of patients with biliary atresia (BA). In this study, we aimed to determine the morphological changes in the LVs irrigating the liver in patients with BA, elucidate their correlations with the morphology of the portal vein (PV) branches, and discuss their etiopathogenetic significance. METHODS: Morphometric analyses of liver biopsy specimens from patients treated between 1986 and 2016 were performed. The parameters measured were as follows: the whole liver area of the specimen, fibrotic area, number of LVs, LVs without patent lumen (designated as Ly0) and PV branches, and diameters of the LVs with patent lumen and the PVs. RESULTS: The numbers of LVs, Ly0, and PV branches per unit area of the whole liver specimen were significantly higher in patients with BA than in control participants with liver disease and those with normal livers. However, no correlation was observed between the fibrotic area and the average diameter of LVs or PVs, and between the fibrotic area and the number of LVs or PV branches. Furthermore, no correlation was observed between the total number of LVs and the number of PV branches. CONCLUSIONS: The present study showed a significant increase in the number of total LVs and Ly0, characterized by a high Ly0 to total LVs ratio, suggesting that lymphangiogenesis occurs in the liver of patients with BA.
Assuntos
Atresia Biliar , Fígado , Linfangiogênese , Vasos Linfáticos , Veia Porta , Humanos , Atresia Biliar/patologia , Fígado/patologia , Fígado/irrigação sanguínea , Feminino , Masculino , Vasos Linfáticos/patologia , Veia Porta/patologia , Lactente , Pré-Escolar , Biópsia , CriançaRESUMO
BACKGROUND: Lymphatic valves are specialized structures in collecting lymphatic vessels and are crucial for preventing retrograde lymph flow. Mutations in valve-forming genes have been clinically implicated in the pathology of congenital lymphedema. Lymphatic valves form when oscillatory shear stress from lymph flow signals through the PI3K/AKT pathway to promote the transcription of valve-forming genes that trigger the growth and maintenance of lymphatic valves. Conventionally, in many cell types, AKT is phosphorylated at Ser473 by the mTORC2 (mammalian target of rapamycin complex 2). However, mTORC2 has not yet been implicated in lymphatic valve formation. METHODS: In vivo and in vitro techniques were used to investigate the role of Rictor, a critical component of mTORC2, in lymphatic endothelium. RESULTS: Here, we showed that embryonic and postnatal lymphatic deletion of Rictor, a critical component of mTORC2, led to a significant decrease in lymphatic valves and prevented the maturation of collecting lymphatic vessels. RICTOR knockdown in human dermal lymphatic endothelial cells not only reduced the level of activated AKT and the expression of valve-forming genes under no-flow conditions but also abolished the upregulation of AKT activity and valve-forming genes in response to oscillatory shear stress. We further showed that the AKT target, FOXO1 (forkhead box protein O1), a repressor of lymphatic valve formation, had increased nuclear activity in Rictor knockout mesenteric lymphatic endothelial cells in vivo. Deletion of Foxo1 in Rictor knockout mice restored the number of valves to control levels in lymphatic vessels of the ear and mesentery. CONCLUSIONS: Our work identifies a novel role for RICTOR in the mechanotransduction signaling pathway, wherein it activates AKT and prevents the nuclear accumulation of the valve repressor, FOXO1, which ultimately enables the formation and maintenance of lymphatic valves.
Assuntos
Proteínas de Transporte , Proteína Forkhead Box O1 , Linfangiogênese , Vasos Linfáticos , Alvo Mecanístico do Complexo 2 de Rapamicina , Mecanotransdução Celular , Camundongos Knockout , Proteínas Proto-Oncogênicas c-akt , Proteína Companheira de mTOR Insensível à Rapamicina , Transdução de Sinais , Animais , Proteína Companheira de mTOR Insensível à Rapamicina/metabolismo , Proteína Companheira de mTOR Insensível à Rapamicina/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteína Forkhead Box O1/metabolismo , Proteína Forkhead Box O1/genética , Vasos Linfáticos/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/genética , Humanos , Proteínas de Transporte/metabolismo , Proteínas de Transporte/genética , Células Endoteliais/metabolismo , Células Cultivadas , Serina-Treonina Quinases TOR/metabolismo , Fosforilação , Fatores de Transcrição Forkhead/metabolismo , Fatores de Transcrição Forkhead/genética , Camundongos , Complexos Multiproteicos/metabolismo , Complexos Multiproteicos/genética , Camundongos Endogâmicos C57BL , Interferência de RNA , TransfecçãoRESUMO
The meninges are critical for the brain functions, but the diversity of meningeal cell types and intercellular interactions have yet to be thoroughly examined. Here we identify a population of meningeal lymphatic supporting cells (mLSCs) in the zebrafish leptomeninges, which are specifically labeled by ependymin. Morphologically, mLSCs form membranous structures that enwrap the majority of leptomeningeal blood vessels and all the mural lymphatic endothelial cells (muLECs). Based on its unique cellular morphologies and transcriptional profile, mLSC is characterized as a unique cell type different from all the currently known meningeal cell types. Because of the formation of supportive structures and production of pro-lymphangiogenic factors, mLSCs not only promote muLEC development and maintain the dispersed distributions of muLECs in the leptomeninges, but also are required for muLEC regeneration after ablation. This study characterizes a newly identified cell type in leptomeninges, mLSC, which is required for muLEC development, maintenance, and regeneration.
Assuntos
Células Endoteliais , Meninges , Peixe-Zebra , Animais , Meninges/citologia , Meninges/metabolismo , Células Endoteliais/metabolismo , Células Endoteliais/citologia , Proteínas de Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Vasos Linfáticos/citologia , Vasos Linfáticos/metabolismo , Animais Geneticamente Modificados , Linfangiogênese/fisiologia , Regeneração/fisiologiaRESUMO
Thyroid carcinoma is a common endocrine malignancy,with most cases being indolent.Lymphatic metastasis as a representative metastasis type defines the clinical stage and prognosis of thyroid carcinoma.The mechanism of lymphatic metastasis in malignancies has been a research hotspot for years,and certain progress being achieved.This article reviews the molecular markers of lymphatic vessels and their application in diagnosis and treatment of neoplasms,the mechanism and role of lymphangiogenesis in lymphatic metastasis,the tracing methods for sentinel lymph nodes by lymphatic drainage,and the use of ultrasound in cervical lymph node metastasis of thyroid carcinoma.Especially,this paper details the application of conventional ultrasound,transvenous contrast-enhanced ultrasound,and trans-lymphatic contrast-enhanced ultrasound in cervical lymph node metastasis of thyroid carcinoma.
Assuntos
Linfangiogênese , Metástase Linfática , Neoplasias da Glândula Tireoide , Humanos , Neoplasias da Glândula Tireoide/patologiaRESUMO
Secondary lymphedema is caused by damage to the lymphatic system from surgery, cancer treatment, infection, trauma, or obesity. This damage induces stresses such as oxidative stress and hypoxia in lymphatic tissue, impairing the lymphatic system. In response to damage, vascular endothelial growth factor C (VEGF-C) levels increase to induce lymphangiogenesis. Unfortunately, VEGF-C often fails to repair the lymphatic damage in lymphedema. The underlying mechanism contributing to lymphedema is not well understood. In this study, we found that surgery-induced tail lymphedema in a mouse model increased oxidative damage and cell death over 16 days. This corresponded with increased VEGF-C levels in mouse tail lymphedema tissue associated with macrophage infiltration. Similarly, in the plasma of patients with secondary lymphedema, we found a positive correlation between VEGF-C levels and redox imbalance. To determine the effect of oxidative stress in the presence or absence of VEGF-C, we found that hydrogen peroxide (H2O2) induced cell death in human dermal lymphatic endothelial cells (HDLECs), which was potentiated by VEGF-C. The cell death induced by VEGF-C and H2O2 in HDLECs was accompanied by increased reactive oxygen species (ROS) levels and a loss of mitochondrial membrane potential. Antioxidant pre-treatment rescued HDLECs from VEGF-C-induced cell death and decreased ROS under oxidative stress. As expected, VEGF-C increased the number of viable and proliferating HDLECs. However, upon H2O2 treatment, VEGF-C failed to increase either viable or proliferating cells. Since oxidative stress leads to DNA damage, we also determined whether VEGF-C treatment induces DNA damage in HDLECs undergoing oxidative stress. Indeed, DNA damage, detected in the form of gamma H2AX (γH2AX), was increased by VEGF-C under oxidative stress. The potentiation of oxidative stress damage induced by VEFG-C in HDLECs was associated with p53 activation. Finally, the inhibition of vascular endothelial growth factor receptor-3 (VEGFR-3) activation blocked VEGF-C-induced cell death following H2O2 treatment. These results indicate that VEGF-C further sensitizes lymphatic endothelial cells to oxidative stress by increasing ROS and DNA damage, potentially compromising lymphangiogenesis.
Assuntos
Apoptose , Dano ao DNA , Células Endoteliais , Peróxido de Hidrogênio , Linfedema , Mitocôndrias , Estresse Oxidativo , Fator C de Crescimento do Endotélio Vascular , Fator C de Crescimento do Endotélio Vascular/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Animais , Humanos , Células Endoteliais/metabolismo , Células Endoteliais/efeitos dos fármacos , Linfedema/metabolismo , Linfedema/patologia , Linfedema/etiologia , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Camundongos , Apoptose/efeitos dos fármacos , Peróxido de Hidrogênio/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Linfangiogênese/efeitos dos fármacos , FemininoRESUMO
Tumours can obtain nutrients and oxygen required to progress and metastasize through the blood supply1. Inducing angiogenesis involves the sprouting of established vessel beds and their maturation into an organized network2,3. Here we generate a comprehensive atlas of tumour vasculature at single-cell resolution, encompassing approximately 200,000 cells from 372 donors representing 31 cancer types. Trajectory inference suggested that tumour angiogenesis was initiated from venous endothelial cells and extended towards arterial endothelial cells. As neovascularization elongates (through angiogenic stages SI, SII and SIII), APLN+ tip cells at the SI stage (APLN+ TipSI) advanced to TipSIII cells with increased Notch signalling. Meanwhile, stalk cells, following tip cells, transitioned from high chemokine expression to elevated TEK (also known as Tie2) expression. Moreover, APLN+ TipSI cells not only were associated with disease progression and poor prognosis but also hold promise for predicting response to anti-VEGF therapy. Lymphatic endothelial cells demonstrated two distinct differentiation lineages: one responsible for lymphangiogenesis and the other involved in antigen presentation. In pericytes, endoplasmic reticulum stress was associated with the proangiogenic BASP1+ matrix-producing pericytes. Furthermore, intercellular communication analysis showed that neovascular endothelial cells could shape an immunosuppressive microenvironment conducive to angiogenesis. This study depicts the complexity of tumour vasculature and has potential clinical significance for anti-angiogenic therapy.
Assuntos
Células Endoteliais , Neoplasias , Neovascularização Patológica , Análise de Célula Única , Humanos , Apresentação de Antígeno , Comunicação Celular , Diferenciação Celular , Linhagem da Célula , Progressão da Doença , Estresse do Retículo Endoplasmático , Células Endoteliais/citologia , Células Endoteliais/imunologia , Células Endoteliais/metabolismo , Linfangiogênese , Neoplasias/irrigação sanguínea , Neoplasias/classificação , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Neovascularização Patológica/patologia , Pericitos/patologia , Pericitos/citologia , Pericitos/metabolismo , Prognóstico , Receptores Notch/metabolismo , Transdução de Sinais , Microambiente Tumoral , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Animais , Peixe-ZebraRESUMO
Ubiquitination plays an essential role in protein stability, subcellular localization, and interactions. Crosstalk between different types of ubiquitination results in distinct biological outcomes for proteins. However, the role of ubiquitination-related crosstalk in lymph node (LN) metastasis and the key regulatory factors controlling this process have not been determined. Using high-throughput sequencing, we found that ubiquitin-conjugating enzyme E2 C (UBE2C) was overexpressed in bladder cancer (BCa) and was strongly associated with an unfavorable prognosis. Overexpression of UBE2C increased BCa lymphangiogenesis and promoted LN metastasis both in vitro and in vivo. Mechanistically, UBE2C mediated sodium-coupled neutral amino acid transporter 2 (SNAT2) monoubiquitination at lysine 59 to inhibit K63-linked polyubiquitination at lysine 33 of SNAT2. Crosstalk between monoubiquitination and K63-linked polyubiquitination increased SNAT2 membrane protein levels by suppressing epsin 1-mediated (EPN1-mediated) endocytosis. SNAT2 facilitated glutamine uptake and metabolism to promote VEGFC secretion, ultimately leading to lymphangiogenesis and LN metastasis in patients with BCa. Importantly, inhibition of UBE2C significantly attenuated BCa lymphangiogenesis in a patient-derived xenograft model. Our results reveal the mechanism by which UBE2C mediates crosstalk between the monoubiquitination and K63-linked polyubiquitination of SNAT2 to promote BCa metastasis and identify UBE2C as a promising target for treating LN-metastatic BCa.
Assuntos
Metástase Linfática , Enzimas de Conjugação de Ubiquitina , Ubiquitinação , Neoplasias da Bexiga Urinária , Animais , Feminino , Humanos , Masculino , Camundongos , Sistema ASC de Transporte de Aminoácidos , Linhagem Celular Tumoral , Linfangiogênese/genética , Antígenos de Histocompatibilidade Menor , Proteínas de Neoplasias/metabolismo , Proteínas de Neoplasias/genética , Enzimas de Conjugação de Ubiquitina/metabolismo , Enzimas de Conjugação de Ubiquitina/genética , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/genética , Fator C de Crescimento do Endotélio Vascular/metabolismo , Fator C de Crescimento do Endotélio Vascular/genéticaRESUMO
Salt-sensitive hypertension (SSHTN) is associated with M1 macrophage polarization and inflammatory responses, leading to inflammation-associated lymphangiogenesis and functional impairment across multiple organs, including kidneys and gonads. However, it remains unclear whether promoting M2 macrophage polarization can alleviate the hypertension, inflammation, and end organ damage in mice with salt sensitive hypertension (SSHTN). Male and female mice were made hypertensive by administering nitro-L-arginine methyl ester hydrochloride (L-NAME; 0.5 mg/ml) for 2 weeks in the drinking water, followed by a 2-week interval without any treatments, and a subsequent high salt diet for 3 weeks (SSHTN). AVE0991 (AVE) was intraperitoneally administered concurrently with the high salt diet. Control mice were provided standard diet and tap water. AVE treatment significantly attenuated BP and inflammation in mice with SSHTN. Notably, AVE promoted M2 macrophage polarization, decreased pro-inflammatory immune cell populations, and improved function in renal and gonadal tissues of mice with SSHTN. Additionally, AVE decreased lymphangiogenesis in the kidneys and testes of male SSHTN mice and the ovaries of female SSHTN mice. These findings highlight the effectiveness of AVE in mitigating SSHTN-induced elevated BP, inflammation, and end organ damage by promoting M2 macrophage polarization and suppressing pro-inflammatory immune responses. Targeting macrophage polarization emerges as a promising therapeutic approach for alleviating inflammation and organ damage in SSHTN. Further studies are warranted to elucidate the precise mechanisms underlying AVE-mediated effects and to assess its clinical potential in managing SSHTN.
Assuntos
Hipertensão , Inflamação , Rim , Macrófagos , Cloreto de Sódio na Dieta , Animais , Masculino , Macrófagos/imunologia , Macrófagos/efeitos dos fármacos , Feminino , Hipertensão/imunologia , Hipertensão/tratamento farmacológico , Hipertensão/fisiopatologia , Rim/efeitos dos fármacos , Rim/patologia , Rim/imunologia , Linfangiogênese/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Camundongos , Pressão Sanguínea/efeitos dos fármacos , Testículo/efeitos dos fármacos , Testículo/patologia , Modelos Animais de DoençasRESUMO
We reported that salt-sensitive hypertension (SSHTN) is associated with increased pro-inflammatory immune cells, inflammation, and inflammation-associated lymphangiogenesis in the kidneys and gonads of male and female mice. However, it is unknown whether these adverse end organ effects result from increased blood pressure (BP), elevated levels of salt, or both. We hypothesized that pharmaceutically lowering BP would not fully alleviate the renal and gonadal immune cell accumulation, inflammation, and lymphangiogenesis associated with SSHTN. SSHTN was induced in male and female C57BL6/J mice by administering nitro-L-arginine methyl ester hydrochloride (L-NAME; 0.5 mg/ml) in their drinking water for 2 weeks, followed by a 2-week washout period. Subsequently, the mice received a 3-week 4% high salt diet (SSHTN). The treatment group underwent the same SSHTN induction protocol but received hydralazine (HYD; 250 mg/L) in their drinking water during the diet phase (SSHTN+HYD). Control mice received tap water and a standard diet for 7 weeks. In addition to decreasing systolic BP, HYD treatment generally decreased pro-inflammatory immune cells and inflammation in the kidneys and gonads of SSHTN mice. Furthermore, the decrease in BP partially alleviated elevated renal and gonadal lymphatics and improved renal and gonadal function in mice with SSHTN. These data demonstrate that high systemic pressure and salt differentially act on end organ immune cells, contributing to the broader understanding of how BP and salt intake collectively shape immune responses and highlight implications for targeted therapeutic interventions.
Assuntos
Pressão Sanguínea , Hipertensão , Inflamação , Rim , Camundongos Endogâmicos C57BL , Cloreto de Sódio na Dieta , Animais , Hipertensão/imunologia , Hipertensão/fisiopatologia , Hipertensão/tratamento farmacológico , Hipertensão/induzido quimicamente , Masculino , Feminino , Pressão Sanguínea/efeitos dos fármacos , Cloreto de Sódio na Dieta/efeitos adversos , Rim/imunologia , Rim/efeitos dos fármacos , Inflamação/imunologia , Linfangiogênese/efeitos dos fármacos , Anti-Hipertensivos/farmacologia , Camundongos , Hidralazina/farmacologia , NG-Nitroarginina Metil Éster/farmacologia , Modelos Animais de Doenças , Gônadas/efeitos dos fármacosRESUMO
INTRODUCTION: Hypertension (HTN) is a major cardiovascular disease that can cause and be worsened by renal damage and inflammation. We previously reported that renal lymphatic endothelial cells (LECs) increase in response to HTN and that augmenting lymphangiogenesis in the kidneys reduces blood pressure and renal pro-inflammatory immune cells in mice with various forms of HTN. Our aim was to evaluate the specific changes that renal LECs undergo in HTN. METHODS: We performed single-cell RNA sequencing. Using the angiotensin II-induced and salt-sensitive mouse models of HTN, we isolated renal CD31+ and podoplanin+ cells. RESULTS: Sequencing of these cells revealed three distinct cell types with unique expression profiles, including LECs. The number and transcriptional diversity of LECs increased in samples from mice with HTN, as demonstrated by 597 differentially expressed genes (p < 0.01), 274 significantly enriched pathways (p < 0.01), and 331 regulons with specific enrichment in HTN LECs. These changes demonstrate a profound inflammatory response in renal LECs in HTN, leading to an increase in genes and pathways associated with inflammation-driven growth and immune checkpoint activity in LECs. CONCLUSION: These results reinforce and help to further explain the benefits of renal LECs and lymphangiogenesis in HTN.
Assuntos
Modelos Animais de Doenças , Células Endoteliais , Hipertensão , Inflamação , Rim , Animais , Camundongos , Hipertensão/genética , Hipertensão/patologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Rim/patologia , Rim/metabolismo , Expressão Gênica , Linfangiogênese/genéticaRESUMO
In zebrafish, brain lymphatic endothelial cells (BLECs) are essential for meningeal angiogenesis and cerebrovascular regeneration. Although epidermal growth factor-like domain 7 (Egfl7) has been reported to act as a pro-angiogenic factor, its roles in lymphangiogenesis remain unclear. Here, we show that Egfl7 is expressed in both blood and lymphatic endothelial cells. We generate an egfl7 cq180 mutant with a 13-bp-deletion in exon 3 leading to reduced expression of Egfl7. The egfl7 cq180 mutant zebrafish exhibit defective formation of BLEC bilateral loop-like structures, although trunk and facial lymphatic development remains unaffected. Moreover, while the egfl7 cq180 mutant displays normal BLEC lineage specification, the migration and proliferation of these cells are impaired. Additionally, we identify integrin αvß3 as the receptor for Egfl7. αvß3 is expressed in the CVP and sprouting BLECs, and blocking this integrin inhibits the formation of BLEC bilateral loop-like structures. Thus, this study identifies a role for Egfl7 in BLEC development that is mediated through the integrin αvß3.
Assuntos
Encéfalo , Células Endoteliais , Integrina alfaVbeta3 , Linfangiogênese , Proteínas de Peixe-Zebra , Peixe-Zebra , Animais , Humanos , Animais Geneticamente Modificados , Encéfalo/metabolismo , Movimento Celular/genética , Proliferação de Células , Família de Proteínas EGF/metabolismo , Família de Proteínas EGF/genética , Células Endoteliais/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Integrina alfaVbeta3/metabolismo , Integrina alfaVbeta3/genética , Linfangiogênese/genética , Mutação , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genéticaRESUMO
BACKGROUND: Mitral valve (MV) disease including myxomatous degeneration is the most common form of valvular heart disease with an age-dependent frequency. Genetic evidence indicates that mutations of the human transcription factor FOXC1 are associated with MV defects, including MV regurgitation. In this study, we sought to determine whether murine Foxc1 and its closely related factor, Foxc2, are required in valvular endothelial cells (VECs) for the maintenance of MV leaflets, including VEC junctions and the stratified trilaminar ECM (extracellular matrix). METHODS: Adult mice carrying tamoxifen-inducible, vascular endothelial cell (EC), and lymphatic EC-specific, compound Foxc1;Foxc2 mutations (ie, EC-Foxc-DKO and lymphatic EC-Foxc-DKO mice, respectively) were used to study the function of Foxc1 and Foxc2 in the maintenance of MVs. The EC and lymphatic EC mutations of Foxc1/c2 were induced at 7 to 8 weeks of age by tamoxifen treatment, and abnormalities in the MVs of these mutant mice were assessed via whole-mount immunostaining, immunohistochemistry/RNAscope, Movat pentachrome/Masson Trichrome staining, and Evans blue injection. RESULTS: EC deletions of Foxc1 and Foxc2 in mice resulted in abnormally extended and thicker MVs by causing defects in the regulation of ECM organization with increased proteoglycan and decreased collagen. Notably, reticular adherens junctions were found in VECs of control MV leaflets, and these reticular structures were severely disrupted in EC-Foxc-DKO mice. PROX1 (prospero homeobox protein 1), a key regulator in a subset of VECs on the fibrosa side of MVs, was downregulated in EC-Foxc1/c2 mutant VECs. Furthermore, we determined the precise location of lymphatic vessels in murine MVs, and these lymphatic vessels were aberrantly expanded and dysfunctional in EC-Foxc1/c2 mutant MVs. Lymphatic EC deletion of Foxc1/c2 also resulted in similar structural/ECM abnormalities as seen in EC-Foxc1/c2 mutant MVs. CONCLUSIONS: Our results indicate that Foxc1 and Foxc2 are required for maintaining the integrity of the MV, including VEC junctions, ECM organization, and lymphatic vessel formation/function to prevent myxomatous MV degeneration.
Assuntos
Modelos Animais de Doenças , Células Endoteliais , Fatores de Transcrição Forkhead , Linfangiogênese , Vasos Linfáticos , Camundongos Knockout , Animais , Fatores de Transcrição Forkhead/metabolismo , Fatores de Transcrição Forkhead/genética , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Vasos Linfáticos/metabolismo , Vasos Linfáticos/patologia , Valva Mitral/metabolismo , Valva Mitral/patologia , Mutação , Camundongos , Junções Intercelulares/metabolismo , Junções Intercelulares/patologia , Doenças das Valvas Cardíacas/metabolismo , Doenças das Valvas Cardíacas/patologia , Doenças das Valvas Cardíacas/genética , Fenótipo , Camundongos Endogâmicos C57BL , Prolapso da Valva Mitral/metabolismo , Prolapso da Valva Mitral/genética , Prolapso da Valva Mitral/patologia , Matriz Extracelular/metabolismo , Matriz Extracelular/patologiaRESUMO
Secondary lymphedema is a chronic and incurable disease lacking satisfactory therapeutic drugs. It primarily results from lymphatic vessel dysfunction resulting from factors such as tumor-related surgery, injury, or infection. Promoting lymphangiogenesis and lymphatic vessel remodeling is crucial for restoring tissue fluid drainage and treating secondary lymphedema. In this study, we discovered that the oral administration of a type-II arabinogalactan (CAPW-1, molecular weight: 64 kDa) significantly promoted lymphangiogenesis and alleviated edema in mice with secondary lymphedema. Notably, the tail diameter of the CAPW-1200 group considerably decreased in comparison to that of the lymphedema group, with an average diameter difference reaching 0.98 mm on day 14. CAPW-1 treatment also reduced the average thickness of the subcutaneous area in the CAPW-1200 group to 0.37 mm (compared with 0.73 mm in the lymphedema group). It also facilitated the return of injected indocyanine green (ICG) from the tail tip to the sciatic lymph nodes, indicating that CAPW-1 promoted lymphatic vessel remodeling at the injury site. In addition, CAPW-1 enhanced the proliferation and migration of lymphatic endothelial cells. This phenomenon was associated with the activation of the toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling pathway, thereby promoting the expression of vascular endothelial growth factor-C (VEGF-C), which can be abolished using a TLR4 antagonist. Despite these findings, CAPW-1 did not alleviate the symptoms of lymphedema or restore lymphatic drainage in VEGFR3flox/flox/Prox1-CreERT2 mice. In summary, CAPW-1 alleviates secondary lymphedema by promoting lymphangiogenesis and lymphatic vessel remodeling through the activation of the TLR4/NF-κB/VEGF-C signaling pathway, indicating its potential as a therapeutic lymphangiogenesis agent for patients with secondary lymphedema.
Assuntos
Galactanos , Linfangiogênese , Vasos Linfáticos , Linfedema , Receptor 4 Toll-Like , Animais , Linfangiogênese/efeitos dos fármacos , Camundongos , Linfedema/tratamento farmacológico , Linfedema/metabolismo , Linfedema/etiologia , Vasos Linfáticos/efeitos dos fármacos , Vasos Linfáticos/metabolismo , Galactanos/farmacologia , Galactanos/química , Receptor 4 Toll-Like/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , NF-kappa B/metabolismo , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Movimento Celular/efeitos dos fármacos , Modelos Animais de Doenças , MasculinoRESUMO
BACKGROUND: Currently, no perfect treatment for neovascularization and lymphangiogenesis exist, and each treatment method has its complications and side effects. This study aimed to investigate the anti-angiogenic and anti-inflammatory effects of cannabidiol and its mechanism of action. METHOD: An in vivo corneal neovascularization (CNV) model was established using the suture method to investigate the inhibitory effects of CBD on suture-induced corneal inflammation, pathological blood vessel formation, and lymphangiogenesis. Additionally, the impact of CBD on immune cells was studied. In vitro methodologies, including cell sorting and co-culture, were employed to elucidate its mechanism of action. RESULTS: Compared with the CNV group, CBD can inhibit CNV, lymphangiogenesis, and inflammation induced via the suture method. In addition, CBD specifically induced CD45+CD11b+Gr-1+ cell upregulation, which significantly inhibited the proliferation of CD4+ T lymphocytes in vitro and exhibited a CD31+ phenotype, proving that they were myeloid-derived suppressor cells (MDSCs). We administered anti-Gr-1 to mice to eliminate MDSCs in vivo and found that anti-Gr-1 partially reversed the anti-inflammatory and angiogenic effects of CBD. Furthermore, we found that compared with MDSCs in the normal group, CBD-induced MDSCs overexpress peroxisome proliferator-activated receptor-gamma (PPAR-γ). Administering PPAR-γ inhibitor in mice almost reversed the induction of MDSCs by CBD, demonstrating the role of PPAR-γ in the function of CBD. CONCLUSION: This study indicates that CBD may induce MDSCs upregulation by activating the nuclear receptor PPAR-γ, exerting anti-inflammatory, antiangiogenic, and lymphangiogenic effects, and revealing potential therapeutic targets for corneal neovascularization and lymphangiogenesis.
Assuntos
Anti-Inflamatórios , Canabidiol , Neovascularização da Córnea , Linfangiogênese , Camundongos Endogâmicos C57BL , Células Supressoras Mieloides , Animais , Canabidiol/farmacologia , Canabidiol/uso terapêutico , Neovascularização da Córnea/tratamento farmacológico , Neovascularização da Córnea/patologia , Células Supressoras Mieloides/efeitos dos fármacos , Células Supressoras Mieloides/imunologia , Camundongos , Linfangiogênese/efeitos dos fármacos , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Modelos Animais de Doenças , Suturas , PPAR gama/metabolismo , Humanos , Inflamação/tratamento farmacológico , Masculino , Córnea/patologia , Córnea/efeitos dos fármacos , Células CultivadasRESUMO
The lymphatic system is one of the most essential and complex systems in the human body. Disorders that affect the development or function of the lymphatic system can lead to multi-system complications and life-long morbidity. The past two decades have seen remarkable progress in our knowledge of the basic biology and function of the lymphatic system, the molecular regulators of lymphatic development, and description of disorders associated with disrupted lymphangiogensis. In this chapter we will touch on the clinical features of complex lymphatic anomalies, new molecular knowledge of the drivers of these disorders, and novel developmental therapeutics for lymphatic disease.
Assuntos
Anormalidades Linfáticas , Humanos , Anormalidades Linfáticas/terapia , Anormalidades Linfáticas/genética , Anormalidades Linfáticas/diagnóstico , Linfangiogênese/genéticaRESUMO
BACKGROUND: The contribution of the tumor microenvironment and extracellular matrix to the aggressive biology of Gastric Cancer (GC) has been recently characterized; however, the role of EMILIN-1 in this context is unknown. EMILIN-1 is an essential structural element for the maintenance of lymphatic vessel (LV) integrity and displays anti-proliferative properties as demonstrated in skin and colon cancer. Given the key role of LVs in GC progression, the aim of this study was to investigate the role of EMILIN-1 in GC mouse models. METHODS: We used the syngeneic YTN16 cells which were injected subcutaneously and intraperitoneally in genetically modified EMILIN-1 mice. In alternative, carcinogenesis was induced using N-Methyl-N-nitrosourea (MNU). Mouse-derived samples and human biopsies were analyzed by IHC and IF to the possible correlation between EMILIN-1 expression and LV pattern. RESULTS: Transgenic mice developed tumors earlier compared to WT animals. 20 days post-injection tumors developed in EMILIN-1 mutant mice were larger and displayed a significant increase of lymphangiogenesis. Treatment of transgenic mice with MNU associated with an increased number of tumors, exacerbated aggressive lesions and higher levels of LV abnormalities. A significant correlation between the levels of EMILIN-1 and podoplanin was detected also in human samples, confirming the results obtained with the pre-clinical models. CONCLUSIONS: This study demonstrates for the first time that loss of EMILIN-1 in GC leads to lymphatic dysfunction and proliferative advantages that sustain tumorigenesis, and assess the use of our animal model as a valuable tool to verify the fate of GC upon loss of EMILIN-1.
Assuntos
Progressão da Doença , Glicoproteínas de Membrana , Camundongos Transgênicos , Neoplasias Gástricas , Microambiente Tumoral , Animais , Neoplasias Gástricas/patologia , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/genética , Humanos , Camundongos , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/genética , Linfangiogênese , MetilnitrosoureiaRESUMO
The integrity of the lymphatic system is critical for preventing the dissemination of tumor cells, such as melanoma, to distant parts of the body. IFN-γ is well studied as a negative regulator for lymphangiogenesis, which is strongly associated with cancer metastasis. However, the exact mechanisms underlying this process remain unclear. In the present study, we investigated whether IFN-γ signaling in lymphatic endothelial cells (LECs) affects tumor cell dissemination by regulating the barrier function of tumor-associated lymphatic vessels. Using LEC-specific IFN-γ receptor (IFN-γR) knockout mice, we found that the loss of IFN-γR in LECs increased the dissemination of melanoma cells into the draining lymph nodes. Notably, IFN-γ signaling in LECs inhibited trans-lymphatic endothelial cell migration of melanoma cells, indicating its regulation of lymphatic barrier function. Further investigations revealed that IFN-γ upregulated the expression of the tight junction protein Claudin-3 in LECs, while knockdown of Claudin-3 in LECs abolished IFN-γ-induced inhibition of trans-lymphatic endothelial migration activity. Mechanistically, IFN-γ inhibits AMPK signaling activation, which is involved in the regulation of fatty acid metabolism. Modulating fatty acid metabolism and AMPK activation in LECs also affected the lymphatic dissemination of melanoma cells, further confirming that this process is involved in IFN-γ-induced regulation of lymphatic barrier function. These results provide novel insights into how IFN-γ modulates tight junctions in LECs, inhibiting the dissemination of melanoma cells via the lymphatic vessels.
Assuntos
Proteínas Quinases Ativadas por AMP , Células Endoteliais , Interferon gama , Melanoma , Camundongos Knockout , Animais , Interferon gama/metabolismo , Camundongos , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Proteínas Quinases Ativadas por AMP/metabolismo , Melanoma/patologia , Melanoma/metabolismo , Movimento Celular , Transdução de Sinais , Receptor de Interferon gama , Receptores de Interferon/metabolismo , Receptores de Interferon/genética , Metástase Linfática , Linhagem Celular Tumoral , Linfangiogênese , Humanos , Vasos Linfáticos/metabolismo , Vasos Linfáticos/patologia , Camundongos Endogâmicos C57BLRESUMO
During embryonic development, lymphatic endothelial cell (LEC) precursors are distinguished from blood endothelial cells by the expression of Prospero-related homeobox 1 (Prox1), which is essential for lymphatic vasculature formation in mouse and zebrafish. Prox1 expression initiation precedes LEC sprouting and migration, serving as the marker of specified LECs. Despite its crucial role in lymphatic development, Prox1 upstream regulation in LECs remains to be uncovered. SOX18 and COUP-TFII are thought to regulate Prox1 in mice by binding its promoter region. However, the specific regulation of Prox1 expression in LECs remains to be studied in detail. Here, we used evolutionary conservation and chromatin accessibility to identify enhancers located in the proximity of zebrafish prox1a active in developing LECs. We confirmed the functional role of the identified sequences through CRISPR/Cas9 mutagenesis of a lymphatic valve enhancer. The deletion of this region results in impaired valve morphology and function. Overall, our results reveal an intricate control of prox1a expression through a collection of enhancers. Ray-finned fish-specific distal enhancers drive pan-lymphatic expression, whereas vertebrate-conserved proximal enhancers refine expression in functionally distinct subsets of lymphatic endothelium.