EOMA Cells Research Articles

Oxidized cellulose microneedle patch combined with vascular embolization and local delivery of timolol maleate for hemangiomas

Hemangiomas are superficial tumors characterized by dense vascular structures that often affect the patient's aesthetic appearance due to the obvious red appearance on the skin. Current treatments, especially timolol maleate in the form of eye drops and hydrogels, suffer from low transdermal drug delivery rates, resulting in prolonged treatment time. To address this challenge, our study introduced a soluble microneedle patch with dextran as the main material to form microcatheters for sustained delivery of timolol maleate. In addition, we proposed a vascular embolization strategy to disrupt the blood supply in hemangiomas. Oxidized cellulose (C-cellulose) was selected for its excellent hemostatic properties. We incorporated C-cellulose into dextran microneedles to facilitate thrombosis in the vascular-rich areas of hemangiomas. The innovative microneedle patch we developed can penetrate the skin to a depth of 430 μm and dissolve rapidly within 3 minutes, ensuring direct drug delivery to the subcutaneous layer. Notably, the treated skin area regained its original appearance within two hours after treatment. In addition to excellent skin permeability and rapid dissolution, these patches significantly promoted apoptosis and inhibited cell migration in mouse hemangioendothelioma EOMA cells. Our results demonstrate that this approach not only achieves significant tumor inhibition in a mouse hemangioma model, but also represents a more effective, convenient, and non-invasive treatment option. Therefore, dextran/C-cellulose/timolol maleate microneedle patch (MNs/Timolol) has broad clinical application prospects in the treatment of hemangiomas, minimizing the risk of additional damage and improving treatment efficacy.

Interleukin-17F suppressed colon cancer by enhancing caspase 4 mediated pyroptosis of endothelial cells

The combination of anti-angiogenic treatment and immunotherapy presents a promising strategy against colon cancer. Interleukin-17F (IL-17F) emerges as a critical immune cell cytokine expressed in colonic epithelial cells, demonstrating potential in inhibiting angiogenesis. In order to clarify the roles of IL-17F in the colon cancer microenvironment and elucidate its mechanism, we established a mouse colon carcinoma cell line CT26 overexpressing IL-17F and transplanted it subcutaneously into syngeneic BALB/c mice. We also analyzed induced colon tumor in IL-17F knockout and wild type mice. Our results demonstrated that IL-17F could suppress colon tumor growth in vivo with inhibited angiogenesis and enhanced recruitment of cysteine-cysteine motif chemokine receptor 6 (CCR6) positive immune cells. Additionally, IL-17F suppressed the tube formation, cell growth and migration of endothelial cells EOMA in vitro. Comprehensive bioinformatics analysis of transcriptome profiles between EOMA cells and those treated with three different concentrations of IL-17F identified 109 differentially expressed genes. Notably, a potential new target, Caspase 4, showed increased expressions after IL-17F treatment in endothelial cells. Further molecular validation revealed a novel downstream signaling for IL-17F: IL-17F enhanced Caspase 4/GSDMD signaling of endothelial cells, CT26 cells and CT26 transplanted tumors, while IL-17F knockout colon tumors exhibited decreased Caspase 4/GSDMD signaling. The heightened expression of the GSDMD N-terminus, coupled with increased cellular propidium iodide (PI) uptake and lactate dehydrogenase (LDH) release, revealed that IL-17F promoted pyroptosis of endothelial cells. Altogether, IL-17F could modulate the colon tumor microenvironment with inhibited angiogenesis, underscoring its potential as a therapeutic target for colon cancer.

Abstract 112: Endothelial Mixed Lineage Leukemia 1 (kmt2a/MLL1) As A Central Mediator Of Immunothrombosis During SARS-CoV-2 Infection

Background: Endotheliitis in severe SARS-CoV-2 (CoV2) results in increased cellular adhesion molecules (CAMs) and procoagulant molecules (PCMs). Cell surface CAMs increase endothelial cell (EC) interactions with circulating immune cells (IC). Increased EC-IC interactions and a procoagulant EC phenotype augment immunothrombotic risk during CoV2 infection. Prophylactic anticoagulants during CoV2 infection are associated with unacceptably high bleeding risk. New targets to treat endotheliitis are required. Hypothesis: We hypothesize chromatin modifying enzymes (CME)s associated with inflammatory NF-kB signaling drive transcriptional changes in endothelial cells (ECs) leading to a prothrombotic and procoagulant phenotype. Aims: Identify potential CMEs that drive CoV-2 endotheliitis. Methods: Public human single cell RNA sequencing data evaluated the pulmonary endothelial epigenetic transcriptome (PMID34876692). Murine venous ECs (mVECs), EOMA cells, and human venous ECs (HUVECs) were treated with murine beta-coronavirus (MHV-A59) or CoV2 spike protein (S1), respectively. Pharmacologic and siRNA inhibition were used to accomplish MLL1 silencing. Results: Humans with fatal CoV2 infection demonstrated marked increase in the KMT2/MLL class of histone methyltransferases in pulmonary ECs. mVEC infection significantly increased transcription of CME kmt2a, CAM esel, and PCMs plau and f3. Significant enrichment of H3K4 methylation and MLL1 are seen on promotors of identified genes. Inhibition of MLL1 reduced CAM and PCM transcripts. Functional EC-IC adhesion testing revealed significant decreases in monocyte adhesion with MLL1 inhibition. Conclusion: Kmt2a/MLL1 positively regulates CoV-2 induction of CAMs and PCMs, endothelial inflammation via positive regulation of CAM gene transcription, and resultant EC-IC interactions. MLL1 may represent a novel therapeutic target to ameliorate immunothrombotic risk during CoV-2 infection.

Preliminary study on vascular-targeted polyphenol nanoparticles in photodynamic therapy of port-wine stains

SignificanceA combination of polyphenol with near infrared photodynamic therapy (PDT) and vascular-targeting nanoparticles holds significant promise as a potential treatment for port-wine stains (PWS). The anti-angiogenic properties of the polyphenol, along with the targeted approach of the nanoparticles and the ability of PDT to selectively damage tissue, make this treatment approach particularly compelling. ApproachUsing EOMA cells for invitro experiment to examine the anti-angiogenesis effects of PDT using ELC, which is self-assembly Epigallocatechin-3-gallate (EGCG) nanoparticles loading with Ce6 photosensitizer. ResultsIn comparison to free Ce6, ELC resulted in increased intracellular uptake of Ce6, leading to improved cytotoxicity when used with laser while maintaining good biocompatibility. Furthermore, ELC enhanced the generation of reactive oxygen species (ROS) within cells. However, ELC did not demonstrate any advantages in inducing apoptosis. ConclusionsPDT using ELC is a promising method for PWS.

Rapamycin and HPPH Co-Loaded Nanodrug Delivered via Dissolvable Microneedles to Treat Port-Wine Stains

Port-wine stains are a kind of dermatological disease of congenital capillary malformation. Based on the biological characteristics of port-wine stains and the advantages of microneedle transdermal administration, we intend to construct a nanodrug co-loaded with rapamycin (RPM), an anti-angiogenesis drug, and photochlor (HPPH), a photosensitizer, and integrate the nanodrug with dissolvable microneedles (MN) to achieve anti-angiogenesis and photodynamic combination therapy for port-wine stains. First, RPM and HPPH co-loaded nanoparticles (RPM-HPPH NP) were prepared by the emulsification solvent-volatilization method, and its ability to generate reactive oxygen species (ROS) was investigated under 660 nm laser irradiation. Mouse hemangioendothelioma endothelial cells (EOMA) were used as the subjects of the study. The cellular uptake behaviors were examined by fluorescence microscopy and flow cytometry. The cytotoxicity effects of RPM-HPPH NP with or without 660 nm laser irradiation on EOMA cells were examined by MTT assays (with free RPM serving as the control). Then, hyaluronic acid (HA) dissolvable microneedles loaded with RPM-HPPH NP (RPM-HPPH NP@HA MN) were obtained by compounding the nanodrug with HA dissolvable microneedle system through the molding method. The morphological characteristics and mechanical properties of RPM-HPPH NP@HA MN were investigated by scanning electron microscope and electronic universal testing machine. The penetration ability of RPM-HPPH NP@HA MN on the skin of nude mice was evaluated by trypan blue staining and H&E staining experiment. The RPM-HPPH NP prepared in the study had a particle size of 150 nm and generated large amounts of ROS under laser irradiation. At the cellular level, RPM-HPPH NP was taken up by EOMA cells in a time-dependent manner. The cytotoxicity of RPM-HPPH NP was higher than that of free RPM with or without laser irradiation. Under laser irradiation, RPM-HPPH NP exhibited stronger cytotoxic effects and the difference was statistically significant (P<0.05). The height of the needle tip of RPM-HPPH NP@HA MN was 600 µm and the mechanical property of a single needle was 0.75048 N. Trypan blue staining and HE staining showed that pressing on the microneedles could produce pores on the skin surface and penetration of the stratum corneum. RPM-HPPH NP@HA MN can deliver RPM-HPPH NP percutaneously to the lesion tissue and realize the synergistic treatment of port-wine stains with anti-angiogenic therapy and photodynamic therapy, providing a new strategy for the construction of nanodrug-loaded microneedle delivery system and the clinical treatment of port-wine stains.

Construction and applications of the EOMA spheroid model of Kaposiform hemangioendothelioma

BackgroundKaposiform hemangioendothelioma (KHE) is a rare intermediate vascular tumor with unclear pathogenesis. Recently, three dimensional (3D) cell spheroids and organoids have played an indispensable role in the study of many diseases, such as infantile hemangioma and non-involuting congenital hemangiomas. However, few research on KHE are based on the 3D model. This study aims to evaluate the 3D superiority, the similarity with KHE and the ability of drug evaluation of EOMA spheroids as an in vitro 3D KHE model.ResultsAfter two days, relatively uniform morphology and high viability of EOMA spheroids were generated by the rotating cell culture system (RCCS). Through transcriptome analysis, compared with 2D EOMA cells, focal adhesion-related genes such as Itgb4, Flt1, VEGFC, TNXB, LAMA3, VWF, and VEGFD were upregulated in EOMA spheroids. Meanwhile, the EOMA spheroids injected into the subcutaneous showed more obvious KMP than 2D EOMA cells. Furthermore, EOMA spheroids possessed the similar characteristics to the KHE tissues and subcutaneous tumors, such as diagnostic markers (CD31 and LYVE-1), cell proliferation (Ki67), hypoxia (HIF-1α) and cell adhesion (E-cadherin and N-cadherin). Based on the EOMA spheroid model, we discovered that sirolimus, the first-line drug for treating KHE, could inhibit EOMA cell proliferation and downregulate the VEGFC expression. Through the extra addition of VEGFC, the effect of sirolimus on EOMA spheroid could be weakened.ConclusionWith a high degree of similarity of the KHE, 3D EOMA spheroids generated by the RCCS can be used as a in vitro model for basic researches of KHE, generating subcutaneous tumors and drug screening.

Proangiogenic role of circRNA-007371 in liver fibrosis.

Circular RNAs (circRNAs) are crucially involved in cancers as competing endogenous RNA (ceRNA) or microRNA (miRNA) sponges. However, the function and mechanism of circRNAs in liver fibrosis remain unknown and are the focus of this study. Murine fibrotic models were induced by thioacetamide (TAA) or carbon tetrachloride (CCl4 ). Increased angiogenesis is accompanied by liver fibrosis in TAA- and CCl4 -induced murine fibrotic livers. circRNA microarray and argonaute 2 (AGO2)-RNA immunoprecipitation (RIP) sequencing (AGO2-RIP sequencing) were performed in murine livers to screen for functional circRNAs. Compared to control livers, 86 differentially expressed circRNAs were obtained in TAA-induced murine fibrotic livers using circRNA microarray. In addition, 551 circRNAs were explored by AGO2-RIP sequencing of murine fibrotic livers. The circRNA-007371 was then selected and verified for back-spliced junction, resistance to RNase R, and loop formation. In vitro, murine hemangioendothelioma endothelial (EOMA) cells were transfected with circRNA-007371 overexpressing plasmid or empty plasmid. circRNA-007371 overexpression promoted tube formation, migration, and cell proliferation of EOMA cells. RNA sequencing and miRNA sequencing were then performed to explore the mechanism of the proangiogenic effects of circRNA-007371. circRNA-007371 promotes liver fibrosis via miRNA sponges or ceRNA mechanisms. Stag1, the parent gene of circRNA-007371, may play a significant role in proangiogenic progression. In conclusion, circRNA-007371 enhances angiogenesis via a miRNA sponge mechanism in liver fibrosis. The antiangiogenic effect of circRNA-007371 inhibition may provide a new strategy for treating patients with liver cirrhosis.

P19. INHIBITION OF MICRORNA 126 LEADS TO COMPLETE REGRESSION OF HEMANGIOENDOTHELIOMA TUMORS IN MICE

PURPOSE: We have previously shown that urinary microRNA126 (miR126) levels are a biomarker for hemangioma growth based on a prospective longitudinal study of children with hemangiomas and healthy age matched controls. We sought to determine the significance of miR126 on the regulation of hemangioma growth using a validated mouse model. METHODS: qPCR measurement of miR126 levels was done in EOMA cells and non-tumor forming endothelial cells. 6 weeks old female 129P/3 mice received a subcutaneous injection of 5x106 EOMA cells. 4 days post injection tumors were visible and topical administration of control antagomiR (n=8) or miR126 antagomiR (n=15) delivered transcutaneously using tissue nanotransfection electroporation technique. Treatments were given twice weekly RESULTS: EOMA cells had 8,000-fold increase in miR126 levels compared to non-tumor forming endothelial cells. Control mice had significantly larger tumors with 100% mortality by 17 days post EOMA cell injection. miR126 antagomiR treated mice were significantly different from controls with decreased mortality (40%), smaller tumor size, prolonged survival with the last mouse death at 29 days post-injection, complete tumor regression by 6 weeks (n=9) and no observed recurrence at 90 days. CONCLUSION: These are the first results to demonstrate complete regression of hemangioendothelioma in response to miR26 inhibition. Elevation of miR126 levels in human hemangioma and murine hemangioendothelioma indicate shared mechanisms of growth. These findings demonstrate that miR126 is necessary for growth of both tumor types and represents a valid therapeutic target to treat hemangiomas.

Inhibitory effect of tetramethylpyrazine combined with propranolol on murine hemangioma endothelial cells

Purpose: To study the inhibitory effect of different doses of tetramethylpyrazine (TMP) combined with the beta-blocker, propranolol (Pro) on hemangioma endothelial (EOMA) cells.&#x0D; Methods: EOMA cells were cultured in vitro with varying doses of TMP and Pro (5, 10, 20 and 40 uM). The effect of treatments on cell proliferation was assessed by MTT assay, while cell apoptosis was assayed by flow cytometry. The expressions of Bcl-2, Bax, p-mTOR), total-mammalian target of rapamycin (t-mTOR, p-p70S6) and total-p70 ribosomal protein S6 (t-p70S6) proteins were determined using Western blot.&#x0D; Results: MTT data showed that when used alone, TMP had no significant inhibitory effect on EOMA cells (p &gt; 0.05). However, when TMP was combined with propranolol, there was significant inhibition of EOMA cells, and that the inhibition is dependent on TMP dose. Flow cytometry results showed that the combination of TMP and Pro induced EOMA cell apoptosis dose-dependently (p &lt; 0.05). Moreover, TMP dose-dependently inhibited the phosphorylation of mTOR and p70S6 in EOMA cells, and enhanced Bax expression, but downregulated Bcl-2 (p &lt; 0.05).&#x0D; Conclusion: These results suggest that TMP enhances the inhibitory influence of Pro p-mTOR and pp-70S6 in EOMA cells in a dose-dependent manner. Thus, TMP may enhance Pro-induced inhibition of the growth of endothelial cells, and promote apoptosis through suppression of activation of PI3K/AKT signal route. These findings provide a theoretical basis for the clinical application of TMP/Pro combination for the treatment of hemangioma.

Two new sesquiterpenes from Sonchus oleraceus and inhibitory mechanism on murine haemangioendothelioma (EOMA) cell lines

Sonchus oleraceus is a kind of medicinal and edible plant which is widely distributed. In this study, two new sesquiterpenes 1 and 2 along with three known compounds 3-5 were isolated from Sonchus oleraceus by the methods of column chromatography. The structures of the two novel compounds were constructed on the basis of HR-MS and NMR spectra. Cytotoxicities of 1 and 2 were assayed on EOMA cell lines and 1 exhibited no inhibitory effect while 2 elicited moderate inhibitory effect on EOMA cells with IC50 value of 26.5 μM. Western Blot assay indicated that 2 could suppress EOMA cell proliferation by inducing apoptosis through Bax/caspase-3 pathway.

15,16-dihydrotanshinone I inhibits EOMA cells proliferation by interfering in posttranscriptional processing of hypoxia-inducible factor 1.

Infantile hemangioma (IH), which threatens the physical and mental health of patients, is the most common benign tumor in infants. Previously, we found that 15,16-dihydrotanshinone I (DHTS) was significantly more effective at inhibiting hemangioma proliferation in vitro and in vivo than the first-line treatment propranolol. To investigate the underlying mechanism of DHTS, we used EOMA cells as a model to study the effect of DHTS. We compared the transcriptomes of control and DHTS-treated EOMA cells. In total, 2462 differentially expressed genes were detected between the groups. Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed downregulated activity of the hypoxia-inducible factor 1 alpha (HIF-1α) signaling pathway in EOMA cells following treatment with DHTS. Thus, we investigated HIF-1α expression at protein and mRNA levels. Our results revealed that DHTS downregulated HIF-1α expression by interfering in its posttranscriptional processing, and the RNA-binding protein HuR participated in this mechanism. Our findings provide a basis for clinical transformation of DHTS and insight into pathogenic mechanisms involved in IH.

Preparation and Evaluation of Cubic Nanoparticles for Improved Transdermal Delivery of Propranolol Hydrochloride

Transdermal drug delivery of propranolol hydrochloride (PRH) is promising for the treatment of infantile hemangioma (IH). Clinically used PRH hydrogel fails to reach the deep IH for complete recovery. In this study, the PRH-loaded cubic nanoparticles (CNPs) were prepared to promote the transdermal effect of PRH. A remote drug loading method was developed to prepare the PRH-CNPs. For the traditional passive drug loading method, the largest encapsulation efficiency (EE%) was around 50%. The remote drug loading was performed by increasing the pH of the mixture of blank CNPs and PRH solution. The optimal PRH-CNPs showed an EE% of 90.15 ± 2.44% at pH8.5. The permeation of the PRH solution was poor while the PRH-CNPs showed greatly enhanced skin permeation. It was found that smaller-sized PRH-CNPs contributed to increased skin permeation and retention. In addition, the PRH-CNPs had higher cytotoxicity towards the EOMA cells when compared with the PRH solution. During storage for 1month, the PRH-CNPs kept stable size distribution, pH, and EE%. In conclusion, results of this study suggested that the PRH-CNPs could be a potential candidate for the treatment of the IH by transdermal delivery.

Mitochondria as Target for Tumor Management of Hemangioendothelioma

Aims: Hemangioendothelioma (HE) may be benign or malignant. Mouse hemangioendothelioma endothelial (EOMA) cells are validated to study mechanisms in HE. This work demonstrates that EOMA cells heavily rely on mitochondria to thrive. Thus, a combination therapy, including weak X-ray therapy (XRT, 0.5 Gy) and a standardized natural berry extract (NBE) was tested. This NBE is known to be effective in managing experimental HE and has been awarded with the Food and Drug Administration Investigational New Drug (FDA-IND) number 140318 for clinical studies on infantile hemangioma. Results: NBE treatment alone selectively attenuated basal oxygen consumption rate of EOMA cells. NBE specifically sensitized EOMA, but not murine aortic endothelial cells to XRT-dependent attenuation of mitochondrial respiration and adenosine triphosphate (ATP) production. Combination treatment, selectively and potently, influenced mitochondrial dynamics in EOMA cells such that fission was augmented. This was achieved by lowering of mitochondrial sirtuin 3 (SIRT3) causing increased phosphorylation of AMP-activated protein kinase (AMPK). A key role of SIRT3 in loss of EOMA cell viability caused by the combination therapy was evident when pyrroloquinoline quinone, an inducer of SIRT3, pretreatment rescued these cells. Innovation and Conclusion: Mitochondria-targeting NBE significantly extended survival of HE-affected mice. The beneficial effect of NBE in combination with weak X-ray therapy was, however, far more potent with threefold increase in murine survival. The observation that safe natural products may target tumor cell mitochondria and sharply lower radiation dosage required for tumor management warrants clinical testing.

Advanced Glycation End Products Downregulate Placental Growth Factor Expression in Endothelial Cells Via a Macrophage‐Mediated Mechanism

In coronary artery disease, atherosclerosis chronically reduces blood flow. Placental growth factor (PLGF), a protein produced by endothelial cells (EC), can promote collateral artery remodeling to increase blood flow. However, collateral growth is impaired in people with type II diabetes. Our group has previously demonstrated that PLGF levels are reduced in a mouse model of diet‐induced type II diabetes. However, the mechanism for this reduction remains to be defined. Inflammatory cytokines and advanced glycation end products (AGE) are implicated in diabetic vascular dysfunction. We therefore hypothesized that AGE and TNFα signaling would inhibit PLGF expression in EC. Human coronary artery EC were treated with AGE, bovine serum albumin (BSA, as a control for AGE), or TNFα. AGE had no direct effect on PLGF in EC; however, TNFα reduced PLGF in EC (P&lt;0.001). Next, murine macrophages (RAW cell line) were treated with AGE or BSA. RAW media was then transferred to murine endothelial cells (EOMA cell line). Treatment with both RAW‐BSA and RAW‐AGE media significantly reduced PLGF levels in EOMA cells compared with control RAW media (BSA, 481.75 ±21.00 pg/mL; AGE, 631.29 ±41.88 pg/mL; control, 1022.48 ±115 pg/mL; P&lt;0.001, n=4). However, TNFα levels were only increased in RAW‐AGE media compared to RAW‐BSA media and RAW‐control media (AGE, 811.11 ±48.37 pg/mL; BSA, 305.68 ±13.95 pg/mL; control, 276.07 ± 10.40 pg/mL; P&lt; 0.001, n=3). This finding suggests that cytokines other than TNFα may also be inhibitory for PLGF. Analysis of RAW media using a Multiplex Pro mouse 6‐panel cytokine assay revealed that TNFα was increased in RAW‐AGE media compared to RAW‐BSA and RAW‐control media (P&lt;0.007). There was also a trend towards increased IL‐10 and IFN‐γ with AGE treatment, which approached significance (P&lt;0.06), suggesting that these cytokines may also be important targets for future investigation. Identifying signaling pathways by which AGE and inflammatory mediators affect PLGF will give new insights into the pathology of diabetic cardiovascular disease.Support or Funding InformationNIH R56 HL084494 (PL), OSU CVM

The regulatory effect of microRNA-21a-3p on the promotion of telocyte angiogenesis mediated by PI3K (p110\u03b1)/AKT/mTOR in LPS induced mice ARDS

BackgroundTelocytes (TCs) are newly identified interstitial cells that participate in tissue protection and repair. The present study investigated the mechanisms underlying the protective effect of TCs in a mouse model of respiratory distress.MethodsThe mouse model of acute respiratory distress syndrome (ARDS) was established by intratracheal instillation of lipopolysaccharide (LPS). After instillation of TCs culture medium, lung injury was assessed, and angiogenesis markers, including CD31 and endothelial nitric oxide synthase (eNOS), were detected by immunofluorescence. Bioinformatics analysis was used to screen significantly differentially expressed microRNAs (miRNAs) in cultured TCs stimulated with LPS, and the regulation of downstream angiogenesis genes by these miRNAs was analysed and verified. PI3K subunits and pathways were evaluated by using a PI3K p110α inhibitor to study the involved mechanisms.ResultsIn ARDS mice, instillation of TCs culture medium ameliorated LPS-induced inflammation and lung injury and increased the protein levels of CD31 and eNOS in the injured lungs. A total of 7 miRNAs and 1899 mRNAs were differentially regulated in TCs stimulated with LPS. Functional prediction analysis showed that the differentially expressed mRNAs were enriched in angiogenesis-related processes, which were highly correlated with miR-21a-3p. Culture medium from TCs with miR-21a-3p inhibition failed to promote angiogenesis in mouse models of LPS-induced ARDS. In cultured TCs, LPS stimulation upregulated the expression of miR-21a-3p, which further targeted the transcription factor E2F8 and decreased Notch2 protein expression. TCs culture medium enhanced hemangioendothelioma endothelial cells (EOMA cells) proliferation, which was blocked by the miR-21a-3p inhibitor. The PI3K p110α inhibitor decreased vascular endothelial growth factor levels in LPS-stimulated TCs and reversed the enhancing effect of TCs culture medium on EOMA cells proliferation.ConclusionsTCs exerted protective effects under inflammatory conditions by promoting angiogenesis via miR-21a-3p. The PI3K p110α subunit and transcriptional factor E2F8 could be involved in this process.

Blockade of angiogenin by thalidomide inhibits the tumorigenesis of murine hemangioendothelioma.

Thalidomide, a well-known immunomodulatory compound, has an anti-angiogenic activity, which may be utilized for the treatment of angiogenesis-related diseases such as hemangioendothelioma. The aim of the present study was to investigate both the antitumor role of thalidomide on hemangioendothelioma and the underlying mechanism. By using the xenograft mouse model, we found that thalidomide can inhibit the progression of hemangioendothelioma in vivo. Moreover, thalidomide shows no effect on the proliferation of hemangioendothelioma endothelial cell (EOMA), but significantly impairs the pro-angiogenic capacity of the EOMA cells in vitro. By qRT-PCR screening, we observed that the expression of angiogenin was downregulated by thalidomide treatment. We next performed tissue array analysis and found a positive correlation between angiogenin expression level and hemangioendothelioma occurrence in patients. Moreover, we confirmed that the antitumoral role of thalidomide is dependent on angiogenin expression both in vivo and in vitro. Taken together, we concluded that thalidomide can inhibit the progression of hemangioendothelioma by downregulating the expression of pro-angiogenic factor angiogenin and therefore can be used as a potent therapeutic to treat hemangioendothelioma.

MiR-291b-3p mediated ROS-induced endothelial cell dysfunction by targeting HUR

Endothelial dysfunction is an early marker of atherosclerosis. Previous studies have indicated that microRNA (miR)-291b-3p regulates the metabolism of lipids and glucose in the liver via targeting adenosine monophosphate-activated kinase α1 and transcription factor p65. The present study investigated whether miR-291b-3p mediated H2O2-mediated endothelial dysfunction. The level of apoptosis of EOMA mouse endothelial cells was analyzed by terminal deoxynucleotidyl-transferase-mediated dUTP nick end labelling staining. The mRNA levels of miR-291b-3p, intercellular adhesion molecule-1 (ICAM-1) and vascular adhesion molecule-1 (VCAM-1) were determined by quantitative polymerase chain reaction. The level of phosphorylated extracellular signal-regulated kinase, and levels of B-cell lymphoma 2 (Bcl-2)-associated X protein and Bcl-2 protein were detected by western blot analysis. The treatment of H2O2 induced the apoptosis and increased the mRNA levels of miR-291b-3p, ICAM-1 and VCAM-1 in EOMA cells. It was also demonstrated that the overexpression of miR-291b-3p promoted EOMA cell apoptosis and dysfunction. In contrast, the downregulation of miR-291b-3p rescued the effect of H2O2 on EOMA cell dysfunction. In addition, Hu antigen R (HuR) was identified as a target gene of miR-291b-3p in EOMA cells. The overexpression of HuR reversed the endothelial dysfunction induced by miR-291b-3p mimics. The present study provides novel insight into the critical role of miR-291b-3p on the endothelial dysfunction induced by H2O2. miR-291b-3p may mediate H2O2-induced endothelial dysfunction via targeting HuR.

DOWN REGULATION OF u-PA BY A NUTRIENT MIXTURE IN HEMANGIOMA (EOMA) CELLS BY INDUCING CASPASE-DEPENDENT APOPTOSIS

To examine, whether the antitumor effects of a specific nutrient mixture are due to induction of apoptosis by inhibition of u-PA. A nutrient mixture containing lysine, proline, ascorbic acid, and green tea extract which has showed anticancer activity against a number of cancer cell lines was used as an experimental composition. EOMA cells were grown in appropriate media with antibiotics in 24-well tissue culture plates. At near confluence, the cells were treated with nutrition mixture at 10, 100, 1000µg/ml in triplicate. Analysis of u-PA activity was carried out by fibrin zymography. Morphological changes and caspase activation associated with apoptosis induction was checked by H&amp;E staining and Live Green caspase assay, respectively. Apoptosis inducing anticancer drug camptothecin (10µM) was used as positive control. The nutrition mixture exhibited dose response toxicity with maximum toxicity 55% (p &lt; 0.001) at 1000µg/ml. EOMA cells expressed u-PA, which was inhibited by nutrition mixture in a dose-dependent manner. The caspase analysis revealed a dose dependent increase in apoptosis of EOMA hemangioma cells, with an increasing apoptosis observed at 100μg/ml, and maximum at 1000μg/ml. Cells treated with nutrition mixture showed significantly more apoptotic changes than the control or camptothecin-treated cells. These results suggest that NM may induce apoptosis of hemangioma cells in vitro thus warranting further investigation.

Differential effects of short chain fatty acids on endothelial Nlrp3 inflammasome activation and neointima formation: Antioxidant action of butyrate

Short chain fatty acids (SCFAs), a family of gut microbial metabolites, have been reported to promote preservation of endothelial function and thereby exert anti-atherosclerotic action. However, the precise mechanism mediating this protective action of SCFAs remains unknown. The present study investigated the effects of SCFAs (acetate, propionate and butyrate) on the activation of Nod-like receptor pyrin domain 3 (Nlrp3) inflammasome in endothelial cells (ECs) and associated carotid neointima formation. Using a partial ligated carotid artery (PLCA) mouse model fed with the Western diet (WD), we found that butyrate significantly decreased Nlrp3 inflammasome formation and activation in the carotid arterial wall of wild type mice (Asc+/+), which was comparable to the effect of gene deletion of the adaptor protein apoptosis-associated speck-like protein gene (Asc-/-). Nevertheless, both acetate and propionate markedly enhanced the formation and activation of the Nlrp3 inflammasome as well as carotid neointima formation in the carotid arteries with PLCA in Asc+/+, but not Asc-/- mice. In cultured ECs (EOMA cells), butyrate was found to significantly decrease the formation and activation of Nlrp3 inflammasomes induced by 7-ketocholesterol (7-Ket) or cholesterol crystals (CHC), while acetate did not inhibit Nlrp3 inflammasome activation induced by either 7-Ket or CHC, but itself even activated Nlrp3 inflammsomes. Mechanistically, the inhibitory action of butyrate on the Nlrp3 inflammasome was attributed to a blockade of lipid raft redox signaling platforms to produce O2•- upon 7-Ket or CHC stimulations. These results indicate that SCFAs have differential effects on endothelial Nlrp3 inflammasome activation and associated carotid neointima formation.

15, 16-Dihydrotanshinone I Inhibits Hemangiomas through Inducing Pro-apoptotic and Anti-angiogenic Mechanisms in Vitro and in Vivo.

Infantile hemangioma (IH) is a common and benign vascular neoplasms, which has a high incidence in children. Although IH is benign, some patients experience complications such as pain, functional impairment, and permanent disfigurement. Treatment options for IH include corticosteroids, surgery, vincristine, interferon or cyclophosphamide. However, none of these modalities are ideal due to restrictions or potential serious side effects. There is thus a great need to explore novel treatments for IH with less side effects. Angiogenesis, vasculogenesis and tumorigenesis are the main features of IH. Tanshen is mostly used in Chinese traditional medicine to treat hematological abnormalities. Therefore, the aim of our study was to evaluate anti-proliferation and anti-angiogenesis effects on hemangiomas cells by extracted Tanshen compounds compared with propranolol, the first-line treatment for IH currently, both in vitro and in vivo. Cell viability, apoptosis, protein expression and anti-angiogenesis were analyzed by CCK8, Annexin V staining, Western blot and tube formation, respectively. The anti-tumor activity in vivo was evaluated using a mouse xenograft model. Fourteen major compounds extracting from Tanshen were screened for their ability to inhibit hemangiomas cells. Of the 14 compounds investigated, 15,16-Dihydrotanshinone I (DHTS) was the most potent modulator of EOMA cell biology. DHTS could significantly decrease EOMA cells proliferation by inducing cell apoptosis, which is much more efficient than propranolol in vitro. DHTS increased the expression of several apoptosis-related proteins, including caspase9, caspase3, PARP, AIF, BAX, cytochrome c, caspase8 and FADD and significantly inhibited angiogenesis, as indicated by reduced tube formation and diminished expression of vascular endothelial cell growth factor receptor 2 and matrix metalloproteinase 9. In nude mice xenograft experiment, DHTS (10 mg/kg) could significantly inhibit the tumor growth of EOMA cells as well as propranolol (40 mg/kg). Our study showed that DHTS was much more effective than propranolol in inhibiting hemangiomas proliferation and angiogenesis in vitro and in vivo, which could have potential therapeutic applications for treatment of IH.