Asunto(s)
Hemangioma , Neoplasias Cutáneas , Humanos , Hemangioma/patología , Hemangioma/metabolismo , Femenino , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/metabolismo , Receptores de Estrógenos/metabolismo , Receptores de Estrógenos/análisis , Receptores Androgénicos/metabolismo , Adulto , Progresión de la Enfermedad , Remisión Espontánea , Embarazo , Receptores de Progesterona/metabolismo , Receptores de Progesterona/análisis , Regresión Neoplásica EspontáneaRESUMEN
CONTEXT: Hemangioma (HA) is a benign vascular neoplasm that can lead to permanent scarring. C-C motif chemokine ligand 2 (CCL2) plays a crucial role in facilitating growth and angiogenesis during HA progression. However, the mechanism regulating CCL2 in HA remains poorly elucidated. OBJECTIVE: To elucidate the mechanism regulating CCL2 in HA. METHODS: Quantitative real-time polymerase chain reaction (RT-qPCR) was employed to determine the expression levels of CCL2, long noncoding RNA (lncRNA) CTBP1 divergent transcript (CTBP1-AS2), and microRNAs (miRNAs). Proliferation, migration, invasion, and angiogenic abilities of human HA endothelial cells (HemECs) were assessed using cell counting kit-8 (CCK-8), colony formation, flow cytometry, transwell, and tube formation assays. Bioinformatics analysis, RNA pull-down, and luciferase reporter assays were conducted to investigate whether CCL2 targets miR-335-5p. Additionally, rescue experiments were performed in this study. RESULTS: CCL2 expression was markedly upregulated in HemECs. CCL2 promoted HA cell proliferation, migration, invasion, and angiogenesis while inhibiting apoptosis. CCL2 was directly targeted by miR-335-5p. Additionally, we found that CTBP1-AS2 could function as a competing endogenous RNA (ceRNA) to sponge miR-335-5p, thereby upregulating CCL2. CONCLUSION: Our findings suggest that targeting the CTBP1-AS2/miR-335-5p/CCL2 axis may hold promise as a therapeutic strategy for HA.
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Quimiocina CCL2 , Hemangioma , MicroARNs , Neovascularización Patológica , Humanos , MicroARNs/genética , Neovascularización Patológica/genética , Neovascularización Patológica/patología , Neovascularización Patológica/metabolismo , Hemangioma/genética , Hemangioma/patología , Hemangioma/metabolismo , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Quimiocina CCL2/biosíntesis , Oxidorreductasas de Alcohol/genética , Proliferación Celular/fisiología , Movimiento Celular/genética , Progresión de la Enfermedad , ARN Largo no Codificante/genética , Proteínas de Unión al ADN/genética , AngiogénesisRESUMEN
We previously published that in patients with infantile hemangioma (IH) at the onset (T0) colony forming unit-fibroblasts (CFU-Fs) are present in in vitro cultures from PB. Herein, we characterize these CFU-Fs and investigate their potential role in IH pathogenesis, before and after propranolol therapy. The CFU-F phenotype (by flow cytometry), their differentiation capacity and ability to support angiogenesis (by in vitro cultures) and their gene expression (by RT-PCR) were evaluated. We found that CFU-Fs are actual circulating MSCs (cMSCs). In patients at T0, cMSCs had reduced adipogenic potential, supported the formation of tube-like structures in vitro and showed either inflammatory (IL1ß and ESM1) or angiogenic (F3) gene expression higher than that of cMSCs from CTRLs. In patients receiving one-year propranolol therapy, the cMSC differentiation in adipocytes improved, while their support in in vitro tube-like formation was lost; no difference was found between patient and CTRL cMSC gene expressions. In conclusion, in patients with IH at T0 the cMSC reduced adipogenic potential, their support in angiogenic activity and the inflammatory/angiogenic gene expression may fuel the tumor growth. One-year propranolol therapy modifies this picture, suggesting cMSCs as one of the drug targets.
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Hemangioma , Células Madre Mesenquimatosas , Humanos , Propranolol/farmacología , Propranolol/uso terapéutico , Propranolol/metabolismo , Transcriptoma , Células Madre Mesenquimatosas/metabolismo , Adipogénesis/genética , Hemangioma/genética , Hemangioma/tratamiento farmacológico , Hemangioma/metabolismoRESUMEN
Infantile hemangioma (IH) is the most prevalent vascular tumor during infancy, characterized by a rapid proliferation phase of disorganized blood vessels and spontaneous involution. IH possibly arises from a special type of multipotent stem cells called hemangioma stem cells (HemSCs), which could differentiate into endothelial cells, pericytes, and adipocytes. However, the underlying mechanisms that regulate the cell fate determination of HemSCs remain elusive. In this study, we unveil KLF2 as a candidate transcription factor involved in the control of HemSCs differentiation. KLF2 exhibits high expression in endothelial cells in proliferating IH but diminishes in adipocytes in involuting IH. Using a combination of in vitro culture of patient-derived HemSCs and HemSCs implantation mouse models, we show that KLF2 governs the proliferation, apoptosis, and cell cycle progression of HemSCs. Importantly, KLF2 acts as a crucial determinant of HemSC fate, directing their differentiation toward endothelial cells while inhibiting adipogenesis. Knockdown of KLF2 induces a proadipogenic transcriptome in HemSCs, leading to impaired blood vessel formation and accelerated adipocyte differentiation. Collectively, our findings highlight KLF2 as a critical regulator controlling the progression and involution of IH by modulating HemSC fate decisions.
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Diferenciación Celular , Progresión de la Enfermedad , Factores de Transcripción de Tipo Kruppel , Humanos , Factores de Transcripción de Tipo Kruppel/metabolismo , Factores de Transcripción de Tipo Kruppel/genética , Ratones , Animales , Lactante , Proliferación Celular/genética , Células Madre Neoplásicas/patología , Células Madre Neoplásicas/metabolismo , Hemangioma/patología , Hemangioma/metabolismo , Hemangioma/genética , Adipogénesis/genética , Adipocitos/metabolismo , Adipocitos/patología , Células Endoteliales/metabolismo , Células Endoteliales/patología , Apoptosis/genética , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismo , Masculino , Hemangioma Capilar/patología , Hemangioma Capilar/metabolismo , Hemangioma Capilar/genéticaRESUMEN
AIM: This study aims to explore the potential of Osmundacetone (OSC) as a new treatment for infantile hemangiomas (IH), the most common benign tumors in infancy. Currently, propranolol serves as the primary treatment for IH, but its effectiveness is limited, and it poses challenges of drug resistance and side effects. Therefore, there is a pressing need to identify alternative therapies for IH. METHODS: The effects of OSC on the proliferation and apoptosis of HemECs (endothelial cells from hemangiomas) were assessed using CCK-8 assay, colony formation assay, HOCHEST 33342 staining, and flow cytometry. Western blot analysis was performed to investigate OSC's influence on Caspases and angiogenesis-related proteins. Animal models were established using HemECs and BALB/c mice, and histological and immunohistochemical staining were conducted to evaluate the impact of OSC on mouse hemangiomas, VEGFR2, and MMP9 expression. RESULTS: OSC treatment significantly reduced HemECs' viability and colony-forming ability, while promoting apoptosis, as indicated by increased HOCHEST 33342 staining. OSC upregulated the protein expression of Bax, PARP, Caspase9, Caspase3, AIF, Cyto C, FADD, and Caspase8 in HemECs. In animal models, OSC treatment effectively reduced hemangioma size and improved histopathological changes. OSC also suppressed VEGFR2 and MMP9 expression while elevating Caspase3 levels in mouse hemangiomas. CONCLUSION: OSC demonstrated promising results in inhibiting HemECs' proliferation, inducing apoptosis, and ameliorating pathological changes in hemangiomas in mice. Moreover, it influenced the expression of crucial caspases and angiogenesis-related proteins. These findings suggest that OSC holds potential as a novel drug for clinical treatment of IH.
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Células Endoteliales , Hemangioma , Cetonas , Animales , Ratones , Caspasas/metabolismo , Transducción de Señal , Metaloproteinasa 9 de la Matriz/metabolismo , Angiogénesis , Proliferación Celular , Hemangioma/tratamiento farmacológico , Hemangioma/metabolismo , Hemangioma/patologíaRESUMEN
Infantile hemangioma (IH) is the most common benign tumor in children. Propranolol is the first-line treatment for IH, but the underlying mechanism of propranolol treatment in IH is not completely understood. Integrated transcriptional and metabolic analyses were performed to investigate the metabolic changes in hemangioma-derived endothelial cells (HemECs) after propranolol treatment. The findings were then further validated through independent cell experiments using a Seahorse XFp analyzer, Western blotting, immunohistochemistry and mitochondrial functional assays. Thirty-four differentially expressed metabolites, including the glycolysis metabolites glucose 6-phosphate, fructose 6-phosphate and fructose 1,6-bisphosphate, were identified by targeted metabolomics. A KEGG pathway enrichment analysis showed that the disturbances in these metabolites were highly related to glucose metabolism-related pathways, including the pentose phosphate pathway, the Warburg effect, glycolysis and the citric acid cycle. Transcriptional analysis revealed that metabolism-related pathways, including glycine, serine and threonine metabolism, tyrosine metabolism, and glutathione metabolism, were highly enriched. Moreover, integration of the metabolomic and transcriptomic data revealed that glucose metabolism-related pathways, particularly glycolysis, were altered after propranolol treatment. Cell experiments demonstrated that HemECs exhibited higher levels of glycolysis than human umbilical vein ECs (HUVECs) and that propranolol suppressed glycolysis in HemECs. In conclusion, propranolol inhibited glucose metabolism in HemECs by suppressing glucose metabolic pathways, particularly glycolysis.
Asunto(s)
Células Endoteliales , Hemangioma , Niño , Humanos , Células Endoteliales/metabolismo , Propranolol/farmacología , Propranolol/metabolismo , Transducción de Señal , Proliferación Celular , Hemangioma/tratamiento farmacológico , Hemangioma/metabolismo , Hemangioma/patología , Glucosa/metabolismo , Fosfatos/farmacologíaRESUMEN
BACKGROUND: Vascular anomalies and tumors are common in the head, neck, and craniofacial areas and are associated with abnormalities in the angiomatous architecture. However, the etiology and molecular basis for the pathogenesis of most vascular lesions are still unknown. Pericytes are mural cells that surround endothelial cells. Besides angiogenesis and other physiological functions, pericytes play an important role in vascularized tissue repair and as resident mesenchymal stem/progenitor cells. Perivascular cells demonstrate a distinct immunohistochemical profile, including expression of alpha-smooth muscle actin (α-SMA), CD146, CD105, and PDGFRß, without endothelial differentiation (absence of CD31 and CD34 immunoreactivity). These pericyte markers have been shown to be expressed in soft tissue hemangiomas. However, they have not been fully examined in intraosseous hemangiomas. METHODS: In this study, we compared mesenchymal stem cell (MSC) expression of CD146 and α-SMA markers in pericytes from hemangiomas from different tissues and malignant vascular tumors. RESULTS: The results demonstrated an increased expression of pericyte markers in perivascular cells of benign hemangiomas, especially intraosseous hemangiomas and a significantly reduced expression of pericyte markers in malignant angiosarcomas. CONCLUSION: The evidence provides insight into the function of pericytes in vascular tumors and suggests their role in vascular tumor disease types.
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Hemangioma , Neoplasias Vasculares , Humanos , Pericitos/metabolismo , Pericitos/patología , Neoplasias Vasculares/metabolismo , Neoplasias Vasculares/patología , Antígeno CD146/metabolismo , Células Endoteliales/metabolismo , Hemangioma/metabolismo , Hemangioma/patologíaRESUMEN
BACKGROUND: Infantile hemangioma (IH) is the most common tumor among infants, but the exact pathogenesis of IH is largely unknown. Our previous study revealed that glucose metabolism may play an important role in the pathogenesis of IH and that the inhibition of the glycolytic key enzyme phosphofructokinase-1 suppresses angiogenesis in IH. 6-Phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) is a metabolic enzyme that converts fructose-6-bisphosphate to fructose-2,6-bisphosphate (F-2,6-BP), which is the most potent allosteric activator of the rate-limiting enzyme phosphofructokinase-1. This study was performed to explore the role of PFKFB3 in IH. METHODS: Microarray analysis was performed to screen the differentially expressed genes (DEGs) between proliferating and involuting IH tissues. PFKFB3 expression was examined by western blot and immunohistochemistry analyses. Cell migration, apoptosis and tube formation were analyzed. Metabolic analyses were performed to investigate the effect of PFKFB3 inhibition by PFK15. Mouse models were established to examine the effect of PFKFB3 inhibition in vivo. RESULTS: PFKFB3 was identified as one of the most significant DEGs and was more highly expressed in proliferating IH tissues and hemangioma-derived endothelial cells (HemECs) than in involuting IH tissues and human umbilical vein endothelial cells, respectively. PFKFB3 inhibition by PFK15 suppressed HemEC glucose metabolism mainly by affecting glycolytic metabolite metabolism and decreasing the glycolytic flux. Moreover, PFK15 inhibited HemEC angiogenesis and migration and induced apoptosis via activation of the apoptosis pathway. Treatment with the combination of PFK15 with propranolol had a synergistic inhibitory effect on HemECs. Moreover, PFKFB3 knockdown markedly suppressed HemEC angiogenesis. Mechanistically, inhibition of PFKFB3 suppressed the PI3K-Akt signaling pathway and induced apoptotic cell death. More importantly, the suppression of PFKFB3 by PFK15 or shPFKFB3 led to markedly reduced tumor growth in vivo. CONCLUSIONS: Our findings suggest that PFKFB3 inhibition can suppress IH angiogenesis and induce apoptosis. Thus, targeting PFKFB3 may be a novel therapeutic strategy for IH.
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Hemangioma , Fosfatidilinositol 3-Quinasas , Lactante , Ratones , Animales , Humanos , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfofructoquinasa-2/metabolismo , Hemangioma/metabolismo , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Glucólisis , Glucosa/metabolismo , Proliferación CelularRESUMEN
OBJECTIVE: In this work, a propranolol hydrochloride (PRH) transfersomes loaded cutaneous hydrogel patch was developed for topical drug delivery in the affected area of infantile haemangioma. METHODS: Sodium cholate was used as the edge activator to prepare the transfersomes. Based on the central composite design, transfersomes hydrogel patch formulation was optimised with 48 h cumulative penetration and time lag as response values. Particle sizes and morphology of the prepared transfersomes were assessed. They were loaded in a cutaneous hydrogel patch, after which their skin permeation abilities were evaluated, and histopathological effects were investigated using guinea pigs. Moreover, in vivo pharmacokinetics studies were performed in rats. RESULTS: The transfersomes system had a encapsulation efficiency of 81.84 ± 0.53%, particle size of 186.8 ± 3.38 nm, polydispersity index of 0.186 ± 0.002, and a zeta potential of -28.6 ± 2.39 mV. Transmission electron microscopy images revealed sphericity of the particles. The ex vivo drug's penetration of the optimised transfersomes hydrogel patch was 111.05 ± 11.97 µg/cm2 through rat skin within 48 h. Assessment of skin tissue did not reveal any histopathological alterations in epidermal and dermal cells. Pharmacokinetic studies showed that skin Cmax (68.22 µg/cm2) and AUC0-24 (1007.33 µg/cm2 × h) for PRH transfersomes hydrogel patch were significantly higher than those of commercially available oral dosage form and hydrogel patch without transfersomes. These findings imply that the transfersomes hydrogel patch can prolong drug accumulation in the affected skin area, and reduce systemic drug distribution via the blood stream. CONCLUSIONS: The hydrogel patch-loaded PRH transfersomes is a potentially useful drug formulation for infantile haemangioma.
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Hemangioma , Propranolol , Ratas , Animales , Cobayas , Propranolol/metabolismo , Propranolol/farmacología , Absorción Cutánea , Hidrogeles/farmacología , Liposomas/metabolismo , Piel/metabolismo , Administración Cutánea , Sistemas de Liberación de Medicamentos , Hemangioma/metabolismo , Tamaño de la Partícula , Portadores de Fármacos/farmacologíaRESUMEN
Infantile hemangiomas are common vascular tumors with a specific natural history. The proliferation and regression mechanism of infantile hemangiomas may be related to the multilineage differentiation ability of hemangioma stem cells, but the specific mechanism is not well elucidated. KIAA1429 is an N6 -methyladenosine methylation-related protein that can also exert its role in a methylation-independent manner. This study aims to explore the function of KIAA1429 in infantile hemangiomas. qRT-PCR, western blotting, and immunostaining were performed to verify the expression of KIAA1429. The endothelial and fibroblast-like phenotypes of hemangioma endothelial cells were detected after KIAA1429 knockdown and overexpression. The stemness properties of hemangioma endothelial cells and the underlying mechanism of KIAA1429 in hemangiomas were also investigated. Nude mouse models of infantile hemangiomas were conducted to ascertain the effects of KIAA1429 in vivo. The results showed that KIAA1429 was highly expressed in infantile hemangiomas, particularly in involuting hemangiomas. In vitro experiments confirmed that KIAA1429 inhibited the endothelial phenotype, enhanced the differentiation ability, and promoted the fibroblast-like phenotype of hemangioma endothelial cells by inducing endothelial cell transition to facultative stem cells. However, the effect of KIAA1429 on the potential target was shown to be independent of N6 -methyladenosine methylation modification. Mouse models further revealed that KIAA1429 could inhibit the proliferation and promote the regression of hemangiomas. In conclusion, this study found that KIAA1429 played an important role in the regression of infantile hemangiomas by enhancing the stemness of hemangioma endothelial cells and could be a potential treatment target for infantile hemangiomas.
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Células Endoteliales , Hemangioma , Proteínas de Unión al ARN , Animales , Ratones , Western Blotting , Diferenciación Celular , Células Endoteliales/metabolismo , Hemangioma/genética , Hemangioma/metabolismo , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismoRESUMEN
OBJECTIVES: This study aimed to investigate the expression of TGFBI in infantile hemangioma (IH) of proliferative stage or involuting stage and detect the effects of TGFBI overexpression or knockdown on the biological beha-vior of hemangioma endothelial cells (HemECs) from proliferative IH by using plasmid and siRNA. METHODS: TGFBI expression levels in proliferative IH and involuting IH were detected by immunofluorescence. TGFBI overexpression plasmid and negative control plasmid were constructed and transfected into HemECs. siRNA for TGFBI and its negative control siRNA were constructed and transfected into HemECs. Western blot was used to detect the expression of TGFBI in the TGFI overexpression group (OE group) and its negative control (NC group), as well as TGFBI knockdown group (si-TGFBI group) and its negative control (si-NC group), to confirm the efficiency of transfection. CCK-8 assays were performed to assess the viability of HemECs. EdU assays were conducted to investigate the proliferation ability of HemECs. Transwell assays were used to detect the migration ability of HemECs. Tube formation assays were carried out to assess the angiogenic capacity of HemECs. Extracellular acidification rate (ECAR) assays were performed to investigate the glycolysis level of HemECs. RESULTS: The results of immunofluorescence showed that TGFBI expression was significantly elevated in proliferative IH compared with that in involuting IH. Western blot showed that TGFBI expression in the OE group was upregulated compared with that in the NC group, and TGFBI expression in si-TGFBI was downregulated compared with that in the si-NC group. The viability, cell proliferation, migration ability, and angiogenic capacity of HemECs were promoted in the OE group compared with those in the NC group, whereas these biological behaviors were inhibited in the si-TGFBI group compared with those in the si-NC group. In ECAR assays, the glycolysis level of HemECs in the OE group was enhanced compared with that in the NC group. CONCLUSIONS: TGFBI is upregulated in proliferative IH. TGFBI overexpression enhanced the viability, cell proliferation, migration ability, and angiogenic capacity of HemECs, which indicated that TGFBI might play a key role in IH progression by accelerating glycolysis. Thus, targeting TGFBI might be an effective therapeutic strategy for IH.
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Células Endoteliales , Hemangioma , Humanos , Proliferación Celular , Células Endoteliales/metabolismo , Hemangioma/genética , Hemangioma/metabolismo , ARN Interferente Pequeño/metabolismo , ARN Interferente Pequeño/farmacología , Transducción de SeñalRESUMEN
Purpose: Anti-vascular endothelial growth factor (anti-VEGF) treatment are now widely used in patients with circumscribed choroidal hemangioma (CCH), however the concentrations of VEGF and other cytokines in CCH patients have not been known before. The study was conducted to compare various cytokine concentrations in the aqueous humor of eyes with CCH and control. Methods: A total of 16 eyes of 16 patients with CCH, and 15 eyes of 15 patients with cataract as the control group were examined. Aqueous humor samples were assessed for 30 angiogenic and inflammatory cytokines by Luminex bead-based multiplex array. Results: Significantly, compared with control group, higher concentrations of VEGF-A and IP-10 were found in the CCH patients (P = 0.002 and P < 0.001). Conclusions: VEGF-A and IP-10 might be involved with the angiogenic and antiangiogenic process in CCH patients, which provides new insight into the pathophysiology of CCH and could be potential targets for treatment.
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Humor Acuoso , Hemangioma , Humor Acuoso/metabolismo , Quimiocina CXCL10/metabolismo , Citocinas/metabolismo , Hemangioma/metabolismo , Humanos , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Hepatic hemangioma (HH) is the most common benign primary liver tumor; however, despite its high prevalence, a stage-specific classification of this tumor is currently missing. For a spatial stage-specific classification, a tissue microarray (TMA) consisting of 98 HHs and 80 hemangioma margins and 78 distant liver tissues was digitally analyzed for the expression of 16 functional and vascular niche-specific markers. For cross-correlation of histopathology and functional characteristics, computed tomography/MRI imaging data of 28 patients were analyzed. Functional and morphological analyses revealed a high level of intra- and interpatient heterogeneity, and morphological heterogeneity was observed with regard to cellularity, vascular diameter, and endothelial cell subtype composition. While regressed hemangiomas were characterized by low blood vessel density, low beta-catenin levels, and a microvascular phenotype, non-regressed HHs showed a pronounced cellular and architectural heterogeneity. Functionally, cellular senescence-associated p16 expression identified an HH subgroup with high vascular density and increased lymphatic endothelial cell content. Histological HH regions may be grouped into spatially defined morphological compartments that may reflect the current region-specific disease stage. The stage-specific classification of HHs with signs of regression and vascular senescence may allow a better disease course-based and cell state-based subtyping of these benign vascular lesions.
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Hemangioma , Neoplasias Hepáticas , Biomarcadores/metabolismo , Células Endoteliales/metabolismo , Hemangioma/metabolismo , Humanos , Neoplasias Hepáticas/metabolismoRESUMEN
Peroxiredoxin (PRDX) is an antioxidant enzyme family with six isoforms (PRDX1-6). The main function of PRDXs is to decrease cellular oxidative stress by reducing reactive oxygen species, such as hydrogen peroxide, to H2O. Recently, it has been reported that PRDXs are overexpressed in various malignant tumors in humans, and are involved in the development, proliferation, and metastasis of tumors. However, studies on the expression of PRDXs in tumors of animals are limited. Therefore, in the present study, we immunohistochemically investigated the expression of PRDX1 and 2 in spontaneous canine hemangiosarcoma (HSA) and hemangioma (HA), as well as in selected normal tissue and granulation tissue, including newly formed blood vessels. Although there were some exceptions, immunolocalization of PRDX1 and 2 in normal canine tissues was similar to those in humans, rats, or mice. In granulation tissue, angiogenic endothelial cells were strongly positive for PRDX1 and 2, whereas quiescent endothelial cells in mature vessels were negative. Both PRDX1 and 2 were significantly highly expressed in HSA compared to HA. There were no significant differences in the expression of PRDX1 and 2 among the subtypes and primary sites of HSA. These results suggest that PRDX1 and 2 may be involved in the angiogenic phenotypes of endothelial cells in granulation tissue as well as in the behavior in the malignant endothelial tumors.
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Enfermedades de los Perros , Hemangioma , Enfermedades de los Roedores , Animales , Enfermedades de los Perros/patología , Perros , Células Endoteliales/metabolismo , Hemangioma/metabolismo , Hemangioma/patología , Hemangioma/veterinaria , Peróxido de Hidrógeno/metabolismo , Ratones , Oxidación-Reducción , Peroxirredoxinas , RatasRESUMEN
We have previously developed several kinds of rapamycin-encapsulated nanoparticles to achieve sustained release of rapamycin to treat hemangioma. However, lack of intrinsic targeting and easy clearance by the immune system are major hurdles that artificial fabricated nanoparticles must overcome. We constructed rapamycin-encapsulated macrophage-derived exosomes mimic nanoparticles-in-microspheres (RNM), to achieve the goal of continuous targeted therapy of hemangiomas. The rapamycin-encapsulated exosome mimic nanoparticles (RN) were firstly prepared by the extrusion-based method from the U937 cells (the human macrophage cell line). After then, RN was encapsulated with PLGA (poly(lactic-co-glycolic acid)) microspheres to obtain RNM. The release profile, targeting activity, and biological activity of RN and RNM were investigated on hemangioma stem cells (HemSCs). RN has a size of 100 nm in diameter, with a rapamycin encapsulation efficacy (EE) of 83%. The prepared microspheres RNM have a particle size of ~30 µm), and the drug EE of RNM is 34%. The sustained release of RNM can remarkably be achieved for 40 days. As expected, RN and RNM showed effective inhibition of cellular proliferation, significant cellular apoptosis, and remarkable repressed expression of angiogenesis factors in HemSCs. Our results showed that RNM is an effective approach for prolonged and effective delivery of rapamycin to hemangiomas.
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Exosomas/química , Hemangioma/tratamiento farmacológico , Nanopartículas/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico/química , Sirolimus/farmacología , Animales , Apoptosis/efectos de los fármacos , Biomimética/métodos , Proliferación Celular/efectos de los fármacos , Sistemas de Liberación de Medicamentos/métodos , Liberación de Fármacos , Femenino , Hemangioma/metabolismo , Humanos , Ratones , Microesferas , Tamaño de la Partícula , Sirolimus/química , Células U937RESUMEN
Our work aims to investigate long non-coding RNA (lncRNA) N6-methyladenosine (m6A) modification and its role in infantile hemangioma (IH). The mRNA and protein expression levels were assessed using quantitative real-time polymerase chain reaction, western blot and immunohistochemistry. Me-RIP assay was performed to evaluate lncRNA NEAT1 m6A levels. Cell proliferation, migration and invasion were evaluated using cell counting kit-8 assay, transwell migration and invasion assay, respectively. Photo-activatable ribonucleoside-enhanced crosslinking and immunoprecipitation assay was conducted to verify the binding relationship between lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) and ALKBH5 (an RNA demethylase). The binding relationship between lncRNA NEAT1, microRNA (miR)-378b and FOS-like antigen 1 (FOSL1) was verified using dual-luciferase reporter gene assay and/or RNA immunoprecipitation assay. ALKBH5, lncRNA NEAT1 and FOLS1 expression was elevated in IH tissues, while miR-378b was downregulated. ALKBH5 knockdown suppressed cell proliferation, migration and invasion of IH cells, while promoting cell apoptosis. ALKBH5 promoted lncRNA NEAT1 expression by reducing the m6A modification of lncRNA NEAT1. In addition, miR-378b was the target of lncRNA NEAT1, and its overexpression reversed the promotion effect of lncRNA NEAT1 overexpression on IH cell tumor-like behaviors. Moreover, FOLS1 was the target of miR-378b, and its overexpression reversed the inhibitory effect of miR-378b overexpression on IH cell tumor-like behaviors in vitro. ALKBH5 might have great potential as therapeutic target for IH, since ALKBH5 silencing suppressed IH progression by regulation of the NEAT1/miR-378b/FOSL1 axis.
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Hemangioma , MicroARNs , ARN Largo no Codificante , Desmetilasa de ARN, Homólogo 5 de AlkB/genética , Desmetilasa de ARN, Homólogo 5 de AlkB/metabolismo , Proliferación Celular/genética , Regulación Neoplásica de la Expresión Génica , Hemangioma/genética , Hemangioma/metabolismo , Humanos , MicroARNs/genética , MicroARNs/metabolismo , ARN Largo no Codificante/metabolismoRESUMEN
Infantile hemangioma (IH) is a common benign endothelial cell tumor in infants and young children, sometimes accompanied by potential complications, and may develop into malignant tumors. Hemangioma endothelial cells (HemECs) are one of the main components of IH. Podophyllotoxin (PPT) has been reported to have many pharmacological activities, especially anti-tumor, but its high toxicity, poor water solubility, and serious gastrointestinal side effects limit its clinical application. In this study, we have designed and synthesized 20 ester derivatives by introducing Boc-amino acids or organic acids at the C-4 position of podophyllotoxin through esterification reactions. The cytotoxicity of these compounds was evaluated on HemECs. Changes in cell proliferation and apoptotic signaling pathways were studied by DAPI staining, colony formation assay, migration assay, measurement of reactive oxygen species (ROS) levels flow cytometry, and Western blot analysis. We found that eight of the compounds were more potent than PPT. Of these, compound V-31 was the most active (IC50 = 0.079 ± 0.0049 µM). Further research indicated that compound V-31 inhibited its proliferation and migration, increased the level of ROS in cells, and induced apoptosis by downregulating p-PI3K, p-Akt, and Bcl-2, and upregulating cleaved caspase-3 and Bax. Our research provides the first insight into the application of PPT derivatives in HemECs, may provide an effective medicine for IH treatment.
Asunto(s)
Células Endoteliales , Hemangioma , Fosfatidilinositol 3-Quinasas , Podofilotoxina , Proteínas Proto-Oncogénicas c-akt , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Niño , Preescolar , Regulación hacia Abajo/efectos de los fármacos , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Células Endoteliales/patología , Ésteres/metabolismo , Hemangioma/tratamiento farmacológico , Hemangioma/metabolismo , Hemangioma/patología , Humanos , Fosfatidilinositol 3-Quinasas/metabolismo , Podofilotoxina/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Long non-coding RNA (lncRNA) breast cancer-associated transcript 54 (BRCAT54) and microRNA-1269b (miR-1269b) are two critical ncRNAs in cancer biology, while their roles in hemangioma are unknown. Our preliminary sequencing data revealed their altered expression in hemangioma and predicted they could interact with each other. This study was therefore carried out to investigate the roles of BRCAT54 and miR-1269b in hemangioma, with a focus on their interaction. In this study, hemangioma samples donated by 20 infantile hemangioma patients at proliferating-phase and 20 infantile hemangioma patients at involuting-phase were used. The expression of BRCAT54 and miR-1269b in hemangioma samples, as well as hemangioma-derived endothelial cells (HDECs) and human umbilical vein endothelial cells (HUVECs) were detected by RT-qPCR. IntaRNA 2.0 was applied to predict the interaction between BRCAT54 and miR-1269b, which was then confirmed by RNA-RNA pulldown assay. Accumulation of BRCAT54 in the subcellular location of HDECs was detected by subcellular fractionation assay. The role of BRCAT54 and miR-1269b in cell proliferation has been explored by the BrdU assay. Compared to proliferating-phase tissues, involuting-phase tissues exhibited decreased expression levels of BRCAT54 and increased expression levels of miR-1269b. HDECs had decreased expression levels of BRCAT54 and increased expression levels of miR-1269b compared to that of HUVECs. In HDECs, BRCAT54, which was detected in both nuclear and cytoplasm fractions, directly interacted with miR-1269b. BRCAT54 and miR-1269b did not affect the expression of each other, while BRCAT54 suppressed the role of miR-1269b in enhancing the proliferation of HDECs. BRCAT54 may sponge miR-1269b to suppress the proliferation of HDECs.
Asunto(s)
Neoplasias de la Mama , Hemangioma , MicroARNs , ARN Largo no Codificante , Neoplasias de la Mama/genética , Proliferación Celular/genética , Células Endoteliales/metabolismo , Femenino , Hemangioma/genética , Hemangioma/metabolismo , Humanos , MicroARNs/genética , MicroARNs/metabolismo , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismoRESUMEN
Propranolol and atenolol, current therapies for problematic infantile hemangioma (IH), are composed of R(+) and S(-) enantiomers: the R(+) enantiomer is largely devoid of beta blocker activity. We investigated the effect of R(+) enantiomers of propranolol and atenolol on the formation of IH-like blood vessels from hemangioma stem cells (HemSCs) in a murine xenograft model. Both R(+) enantiomers inhibited HemSC vessel formation in vivo. In vitro, similar to R(+) propranolol, both atenolol and its R(+) enantiomer inhibited HemSC to endothelial cell differentiation. As our previous work implicated the transcription factor sex-determining region Y (SRY) box transcription factor 18 (SOX18) in propranolol-mediated inhibition of HemSC to endothelial differentiation, we tested in parallel a known SOX18 small-molecule inhibitor (Sm4) and show that this compound inhibited HemSC vessel formation in vivo with efficacy similar to that seen with the R(+) enantiomers. We next examined how R(+) propranolol alters SOX18 transcriptional activity. Using a suite of biochemical, biophysical, and quantitative molecular imaging assays, we show that R(+) propranolol directly interfered with SOX18 target gene trans-activation, disrupted SOX18-chromatin binding dynamics, and reduced SOX18 dimer formation. We propose that the R(+) enantiomers of widely used beta blockers could be repurposed to increase the efficiency of current IH treatment and lower adverse associated side effects.
Asunto(s)
Atenolol/farmacología , Hemangioma , Células Madre Neoplásicas/metabolismo , Neovascularización Patológica , Propranolol/farmacología , Animales , Hemangioma/irrigación sanguínea , Hemangioma/tratamiento farmacológico , Hemangioma/metabolismo , Humanos , Ratones , Neovascularización Patológica/tratamiento farmacológico , Neovascularización Patológica/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
PURPOSE: To investigate the effect of propranolol on urine bFGF, MMP-2, MMP-9 expression of children with proliferative infantile hemangioma(IH), so as to clarify the mechanism of propranolol in treating IH. METHODS: From June 2018 to June 2019, thirty-four children with proliferative IH were treated with oral propranolol. In addition, twenty-one normal children (age <12 months) were chosen as the control group. 10 mL of sterile morning urine were collected before and 2 months after oral administration of propranolol in infants with IH. All blood samples were placed in ordinary disinfection test tubes, centrifuged at 1 000 r/min for 10 min, the supernatant of urine was collected and stored separately. The urine samples of normal control group were processed in the same way. The expression levels of bFGF in the urine of children with proliferative IH before and 2 months after oral administration of propranolol and in the normal control group were detected by enzyme-linked immunosorbent assay (ELISA). The expression levels of MMP-2 and MMP-9 in the urine of children with proliferative IH before and 2 months after treatment and in the control group were detected by gelatin zymography. SPSS 22.0 software package was used to analyze the data. RESULTS: Two months after oral propranolol treatment, the concentration of bFGF in urine was significantly lower than that before treatment (P<0.01), but still significantly higher than that in the control group (P<0.05). The expression levels of MMP-2 and MMP-9 were significantly lower than those before treatment(P<0.01), but still higher than those in the control group (P<0.05). CONCLUSIONS: One of the mechanisms of propranolol in the treatment of children with proliferative IH may be through inhibiting the expression levels of bFGF, MMP-2 and MMP-9, and then inhibiting the proliferation and angiogenesis of vascular endothelial cells in IH, so as to achieve the effect of treating hemangioma. The detection of the expression levels of bFGF, MMP-2 and MMP-9 in urine can be used as the index for oral propranolol treatment of children with proliferative IH.