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Simple Summary: Angiogenesis is a synchronous mechanism of new blood vessel formation that is controlled by pro- and anti-angiogenic factors. Angiogenesis differs from tissue to tissue, especially in pathological conditions. The exact nature of the co-operation between angiogenic markers is still unknown. Herein, we describe the possible critical changes in VEGF-A, TGF-β1, ANG1, ANG2, and HIF-1α in ectopic endometriosis (with elevated pro-angiogenic factors in all studied fields), in ovarian endometrioid adenocarcinoma (showing typical upregulation on the TGF-β1 axis), and two different types of endometrial carcinoma (with an extreme pathological ANG2/ANG1 axis for mixed mesodermal tumors). Understanding the angiogenic mechanism within pathological tissues could help to improve angiogenic treatment strategies and lead to better prognoses. A characteristic feature of uterine pathologies is a specific change in cell metabolism, which predominantly manifests as a shift in the need for nutrients, thereby directing cells to engage in different angiogenic marker activities. Angiogenesis is one of the main signals supporting the survival and development of cells and tissues not only under physiological conditions. Therefore, it is necessary that we understand pathological hyperactivation in all uterine diseases, from endometriosis through ovarian endometrioid adenocarcinoma to malignant transformed cells of the uterine epithelium and body. This work presents the gene expression results of selected angiogenesis targets (VEGF-A, TGF-β1, ANG1/2, and HIF-1α), cell migration, and cell–cell interaction determined in vitro. Our results suggest that angiogenesis varies in the tested pathological conditions (ectopic endometriosis—12Z; ovarian endometrioid adenocarcinoma—A2780; tumors—SK-UT-1 and RL-95-2) compared to physiological angiogenesis (HME1). The differential expression of angiogenic factors may contribute (or is a contributing factor) to the observed differences to acknowledge an inherent variability in angiogenesis among cell lines. Determining the genomic phenomena responsible for processes associated with inadequate angiogenesis in the pelvic region could help us to develop individual treatment strategies and explain resistance to treatment. [ABSTRACT FROM AUTHOR]

Angiopoietins are crucial growth factors for maintaining a healthy, functional endothelium. Patients with type 2 diabetes (T2D) exhibit significant levels of angiogenic markers, particularly Angiopoietin-2, which compromises endothelial integrity and is connected to symptoms of endothelial injury and failure. This report examines the levels of circulating angiopoietins in people with T2D and diabetic nephropathy (DN) and explores its link with ANGPTL proteins. We quantified circulating ANGPTL3, ANGPTL4, ANGPTL8, Ang1, and Ang2 in the fasting plasma of 117 Kuwaiti participants, of which 50 had T2D and 67 participants had DN. The Ang2 levels increased with DN (4.34 ± 0.32 ng/mL) compared with T2D (3.42 ± 0.29 ng/mL). This increase correlated with clinical parameters including the albumin-to-creatinine ratio (ACR) (r = 0.244, p = 0.047), eGFR (r = −0.282, p = 0.021), and SBP (r = −0.28, p = 0.024). Furthermore, Ang2 correlated positively to both ANGPTL4 (r = 0.541, p < 0.001) and ANGPTL8 (r = 0.41, p = 0.001). Multiple regression analysis presented elevated ANGPTL8 and ACRs as predictors for Ang2's increase in people with DN. In people with T2D, ANGPTL4 positively predicted an Ang2 increase. The area under the curve (AUC) in receiver operating characteristic (ROC) analysis of the combination of Ang2 and ANGPTL8 was 0.77 with 80.7% specificity. In conclusion, significantly elevated Ang2 in people with DN correlated with clinical markers such as the ACR, eGFR, and SBP, ANGPTL4, and ANGPTL8 levels. Collectively, this study highlights a close association between Ang2 and ANGPTL8 in a population with DN, suggesting them as DN risk predictors. [ABSTRACT FROM AUTHOR]

Background: The angiopoietin/TIE system is one of the signalling pathways that regulate vascular development and remodelling during morphogenesis of the placental complex. Deregulation of the vascularization process is associated with primary placental insufficiency through destabilization of the maternal-fetal functional system. Disruption of expression of pro-angiogenic factors is associated with reduction of the microcirculatory bed and installation of circulatory dysfunction. The aim: Evaluation of Ang2 immunoexpression in early term compromised uterine pregnancies in the context of chorio-villous circulatory dysfunction in primary placental insufficiency. Material and methods: Abortion product from 67 patients with early term compromised pregnancies (stagnant pregnancies – 43 cases, early miscarriages – 11 cases, control batch that included 13 cases of pregnancies solved on social indications) were immunohistochemically evaluated with the marker for anti-Ang2 and anti-CD31. Results: The majority of sites analyzed were Ang2 negative, with the exception of syncytiotrophoblast, that reached the highest score (+3) in most cases. A maximum placental vascularisation index was achieved in the control group. The stagnant pregnacies batch has registered the lowest vascular density mean. Conclusions: The placentation period is characterized by a weak Ang2 cellular environment in the chorio-villous germinal site in the group of short-term compromised pregnancies, except a highly positive syncytiotrophoblast. The immunoexpression profile in vascular endothelium correlates statistically significantly with placental vascularization index and chorio-villous vascular density in stagnant pregnancies.

Objective: This study aims to investigate the role of BMAL1 in promoting angiogenesis in cerebral glioma among athletes by regulating vascular endothelial growth factor (VEGF) and angiopoietin-2 (ANG2). Methods: Human brain tumor cells, representative of gliomas often observed in athletes, were categorized into three groups: blank, control, and BMAL1-treated research group. Cell growth and apoptosis were analyzed using the MTT assay and flow cytometry, respectively. VEGF and ANG2 expressions, along with microvessel density (MVD), were assessed via immunohistochemistry. The Spearman rank test evaluated the correlation between VEGF and ANG2 levels. Further, groups A, B, and C (representing different pathological grades of human glioma tissues) and BMAL1-treated groups (D, E, and F) were compared to analyze BMAL1's role in modulating VEGF and ANG2. Results: The control group showed significantly lower cell survival and higher apoptosis rates compared to the blank group (P<0.05). Higher glioma grades correlated positively with increased apoptosis rates (P<0.05). VEGF, ANG2, and MVD expressions were significantly higher in the control group than in the blank group (P<0.05), with notable differences observed between the study and control groups (P<0.05). A positive correlation was found between glioma grade, MVD count, and the expressions of VEGF and ANG2 (P<0.05). Groups D, E, and F demonstrated significant differences in VEGF and ANG2 expressions, with group F exhibiting the highest levels (P<0.05), indicating BMAL1's regulatory effect on these angiogenic factors. Conclusion: The study confirms that BMAL1 regulates VEGF and ANG2 expression, promoting angiogenesis in cerebral glioma, a condition relevant to athletes due to their unique physiological stresses. This highlights the potential of targeting BMAL1 in developing therapeutic strategies for glioma angiogenesis in athletic populations, providing a novel approach to manage this challenging aspect of cerebral glioma treatment. [ABSTRACT FROM AUTHOR]

Defining reliable surrogate markers and overcoming drug resistance are the most challenging issues for improving therapeutic outcomes of antiangiogenic drugs (AADs) in cancer patients. At the time of this writing, no biomarkers are clinically available to predict AAD therapeutic benefits and drug resistance. Here, we uncovered a unique mechanism of AAD resistance in epithelial carcinomas with KRAS mutations that targeted angiopoietin 2 (ANG2) to circumvent antivascular endothelial growth factor (anti-VEGF) responses. Mechanistically, KRAS mutations up-regulated the FOXC2 transcription factor that directly elevated ANG2 expression at the transcriptional level. ANG2 bestowed anti-VEGF resistance as an alternative pathway to augment VEGF-independent tumor angiogenesis. Most colorectal and pancreatic cancers with KRAS mutations were intrinsically resistant to monotherapies of anti-VEGF or anti-ANG2 drugs. However, combination therapy with anti-VEGF and anti-ANG2 drugs produced synergistic and potent anticancer effects in KRAS-mutated cancers. Together, these data demonstrate that KRAS mutations in tumors serve as a predictive marker for anti-VEGF resistance and are susceptible to combination therapy with anti-VEGF and anti-ANG2 drugs. [ABSTRACT FROM AUTHOR]

Nowadays; intravitreal anti-vascular endothelial growth factor (VEGF) drugs are considered the first-line therapeutic strategy for treating macular exudative diseases; including wet age-related macular degeneration (w-AMD) and diabetic macular edema (DME). Despite the important clinical achievements obtained by anti-VEGF drugs in the management of w-AMD and DME; some limits still remain; including high treatment burden; the presence of unsatisfactory results in a certain percentage of patients and long-term visual acuity decline due to complications such as macular atrophy and fibrosis. Targeting the angiopoietin/Tie (Ang/Tie) pathway beyond the VEGF pathway may be a possible therapeutic strategy; which may has the potential to solve some of the previous mentioned challenges. Faricimab is a new; bispecific antibody targeting both VEGF-A and the Ang-Tie/pathway. It was approved by FDA and; more recently; by EMA for treating w-AMD and DME. Results from phase III trials TENAYA and LUCERNE (w-AMD) and RHINE and YOSEMITE (DME) have shown the potential of faricimab to maintain clinical efficacy with more prolonged treatment regimens compared to aflibercept (12 or 16 weeks) with a a good safety profile. [ABSTRACT FROM AUTHOR]

Angiopoietins are crucial growth factors for maintaining a healthy, functional endothelium. Patients with type 2 diabetes (T2D) exhibit significant levels of angiogenic markers, particularly Angiopoietin-2, which compromises endothelial integrity and is connected to symptoms of endothelial injury and failure. This report examines the levels of circulating angiopoietins in people with T2D and diabetic nephropathy (DN) and explores its link with ANGPTL proteins. We quantified circulating ANGPTL3, ANGPTL4, ANGPTL8, Ang1, and Ang2 in the fasting plasma of 117 Kuwaiti participants, of which 50 had T2D and 67 participants had DN. The Ang2 levels increased with DN (4.34 ± 0.32 ng/mL) compared with T2D (3.42 ± 0.29 ng/mL). This increase correlated with clinical parameters including the albumin-to-creatinine ratio (ACR) (r = 0.244, p = 0.047), eGFR (r = −0.282, p = 0.021), and SBP (r = −0.28, p = 0.024). Furthermore, Ang2 correlated positively to both ANGPTL4 (r = 0.541, p < 0.001) and ANGPTL8 (r = 0.41, p = 0.001). Multiple regression analysis presented elevated ANGPTL8 and ACRs as predictors for Ang2’s increase in people with DN. In people with T2D, ANGPTL4 positively predicted an Ang2 increase. The area under the curve (AUC) in receiver operating characteristic (ROC) analysis of the combination of Ang2 and ANGPTL8 was 0.77 with 80.7% specificity. In conclusion, significantly elevated Ang2 in people with DN correlated with clinical markers such as the ACR, eGFR, and SBP, ANGPTL4, and ANGPTL8 levels. Collectively, this study highlights a close association between Ang2 and ANGPTL8 in a population with DN, suggesting them as DN risk predictors.

The purpose of this study was to identify genes that mediate VEGF-induced permeability. We performed RNA-Seq analysis on primary human retinal endothelial cells (HRECs) cultured in normal (5 mM) and high glucose (30 mM) conditions that were treated with vehicle, VEGF, or VEGF then anti-VEGF. We filtered our RNA-Seq dataset to identify genes with the following four characteristics: (1) regulated by VEGF, (2) VEGF regulation reversed by anti-VEGF, (3) regulated by VEGF in both normal and high glucose conditions, and (4) known contribution to vascular homeostasis. Of the resultant 18 genes, members of the Notch signaling pathway and ANGPT2 (Ang2) were selected for further study. Permeability assays revealed that while the Notch pathway was dispensable for relaxing the barrier, it contributed to maintaining an open barrier. In contrast, Ang2 limited the extent of barrier relaxation in response to VEGF. These findings indicate that VEGF engages distinct sets of genes to induce and sustain barrier relaxation. Furthermore, VEGF induces expression of genes that limit the extent of barrier relaxation. Together, these observations begin to elucidate the elegance of VEGF-mediated transcriptional regulation of permeability. [ABSTRACT FROM AUTHOR]

Background: SARS-CoV-2-induced severe inflammatory response can be associated with severe medical consequences leading to multi-organ failure, including the liver. The main mechanism behind this assault is the aggressive cytokine storm that induces cytotoxicity in various organs. Of interest, hepatic stellate cells (HSC) respond acutely to liver injury through several molecular mechanisms, hence furthering the perpetuation of the cytokine storm and its resultant tissue damage. In addition, hepatocytes undergo apoptosis or necrosis resulting in the release of pro-inflammatory and pro-fibrogenic mediators that lead to chronic liver inflammation. Aims: The aim of this review is to summarize available data on SARS-CoV-2-induced liver inflammation in addition to evaluate the potential effect of anti-inflammatory drugs in attenuating SARS-CoV-2-induced liver inflammation. Methods: Thorough PubMed search was done to gather and summarize published data on SARS-CoV-2-induced liver inflammation. Additionally, various anti-inflammatory potential treatments were also documented. Results: Published data documented SARS-CoV-2 infection of liver tissues and is prominent in most liver cells. Also, histological analysis showed various features of tissues damage, e.g., hepatocellular necrosis, mitosis, cellular infiltration, and fatty degeneration in addition to microvesicular steatosis and inflammation. Finally, the efficacy of the different drugs used to treat SARS-CoV-2-induced liver injury, in particular the anti-inflammatory remedies, are likely to have some beneficial effect to treat liver injury in COVID-19. Conclusion: SARS-CoV-2-induced liver inflammation is a serious condition, and drugs with potent anti-inflammatory effect can play a major role in preventing irreversible liver damage in COVID-19. [ABSTRACT FROM AUTHOR]

Though survival rates for preterm infants are improving, the incidence of chronic lung disease of infancy, or bronchopulmonary dysplasia (BPD), remains high. Histologically, BPD is characterized by larger and fewer alveoli. Hypoxia-inducible factors (HIFs) may be protective in the context of hyperoxia-induced lung injury, but the cell-specific effects of HIF expression in neonatal lung injury remain unknown. Thus, we sought to determine whether HIF stabilization in SM22a-expressing cells can limit hyperoxiainduced neonatal lung injury. We generated SM22a-specific HIF-1a-stabilized mice (SM22a-PHD1/2/mice) by cross-breeding SM22a-promotor-driven Cre recombinase mice with prolyl hydroxylase PHD1flox/flox and PHD2flox/flox mice. Neonatal mice were randomized to 21% O2 (normoxia) or 80% O2 (hyperoxia) exposure for 14 days. For the hyperoxia recovery studies, neonatal mice were recovered from normoxia for an additional 10 wk. SM22a-specific HIF-1a stabilization mitigated hyperoxia-induced lung injury and preserved microvessel density compared with control mice for both neonates and adults. In SM22a-PHD1/2/mice, pulmonary artery endothelial cells (PAECs) were more proliferative and pulmonary arteries expressed more collagen IV compared with control mice, even under hyperoxic conditions. Angiopoietin-2 (Ang2) mRNA expression in pulmonary artery smooth muscle cells (PASMC) was greater in SM22a-PHD1/2/compared with control mice in both normoxia and hyperoxia. Pulmonary endothelial cells (PECs) cocultured with PASMC isolated from SM22a-PHD1/2/mice formed more tubes and branches with greater tube length compared with PEC cocultured with PASMC isolated from SM22a-PHD1/2þ/þ mice. Addition of Ang2 recombinant protein further augmented tube formation for both PHD1/2þ/þ and PHD1/2/PASMC. Cell-specific deletion of PHD1 and 2 selectively increases HIF-1a expression in SM22a-expressing cells and protects neonatal lung development despite prolonged hyperoxia exposure. HIF stabilization in SM22a-expressing cells preserved endothelial cell proliferation, microvascular density, increased angiopoietin-2 expression, and lung structure, suggesting a role for cell-specific HIF-1a stabilization to prevent neonatal lung injury. [ABSTRACT FROM AUTHOR]

Lymphangiogenesis is an essential physiological process but also a determining factor in vascular-related pathological conditions. Angiopoietin-2 (Ang2) plays an important role in lymphatic vascular development and function and its upregulation has been reported in several vascular-related diseases, including cancer. Given the established role of the small GTPase RhoA on cytoskeleton-dependent endothelial functions, we investigated the relationship between RhoA and Ang2-induced cellular activities. This study shows that Ang2-driven human dermal lymphatic endothelial cell migration depends on RhoA. We demonstrate that Ang2-induced migration is independent of the Tie receptors, but dependent on β1 integrin-mediated RhoA activation with knockdown, pharmacological approaches, and protein sequencing experiments. Although the key proteins downstream of RhoA, Rho kinase (ROCK) and myosin light chain, were activated, blockade of ROCK did not abrogate the Ang2-driven migratory effect. However, formins, an alternative target of RhoA, were identified as key players, and especially FHOD1. The Ang2-RhoA relationship was explored in vivo, where lymphatic endothelial RhoA deficiency blocked Ang2-induced lymphangiogenesis, highlighting RhoA as an important target for anti-lymphangiogenic treatments. [ABSTRACT FROM AUTHOR]

Angiopoietin-2 (Ang2), a member of the angiopoietin family, is widely involved in the process of vascular physiology, bone physiology, adipose tissue physiology and the occurrence and development of inflammation, cardiac hypertrophy, rheumatoid, tumor and other diseases under pathological conditions. Proliferation and metastasis of cancer largely depend on angiogenesis. Therefore, anti-angiogenesis has become the target of tumor therapy. Due to the Ang2 plays a key role in promoting angiogenesis and stability in vascular physiology, the imbalance of its expression is an important condition for the occurrence and development of cancer. It has been proved that blocking Ang2 can inhibit the growth, invasion and metastasis of cancer cells. In recent years, research has been constantly supplemented. We focus on the mechanisms that regulate the expression of Ang2 mRNA and protein levels in different cancers, contributing to a better understanding of how Ang2 exerts different effects in different cancers and stages, as well as facilitating more specific targeting of relevant molecules in cancer therapy. At the same time, the importance of Ang2 in cancer growth, metastasis, prognosis and combination therapy is pointed out. And finally, we will discuss the current investigations and future challenges of combining Ang2 inhibition with chemotherapy, immunotherapy, and radiotherapy to increase its efficacy in cancer patients. This review provides a theoretical reference for the development of new targets and effective combination therapy strategies for cancer treatment in the future. [ABSTRACT FROM AUTHOR]

Abstract Background Proper oxygen balance in the dental pulp is essential for cell metabolism. Angiogenesis in the pulp is a constant process during the life of the tooth. Hypoxia indicators in a tissue, such as HIF-1α, as well as vascular destabilization markers, such as ANG2 and its receptor TIE2, are necessary for angiogenesis. Therefore the purpose of this study is to evaluate the expression of HIF-1α, ANG1, ANG2 and TIE2 in dental pulp as early angiogenesis indicators in teeth with complete and incomplete root development. Methods Forty human dental pulps were obtained from freshly extracted third molars divided into two groups: incomplete (n = 20) and complete (n = 20) root development. Dental pulps were stored at − 80 °C, defrosted in an ice bath and re-frozen with their respective thaws to disintegrate the tissue. Three sonication cycles were performed until the tissues were homogenized, then thaw were centrifuged and the supernatant was collected for the detection of the markers to be studied. The samples were processed for the ELISA test using the ELISA-sandwich principle. Student t and Mann–Whitney U tests were performed to determine statistically significant differences between groups. Results In the complete root development, HIF-1α, ANG1, ANG2 and TIE2 expressions were significantly higher than their expression in the incomplete root development group. Conclusions The angiogenic process seems to be a physiological process in the dental pulp. Angiogenic activity is higher in teeth with mature than immature apex teeth.

Macrophages play important roles in angiogenesis; however, previous studies on macrophage angiogenesis have focused on traditional 2D cultures. In this study, we established a 3D culture system for macrophages using collagen microcarriers and assessed the effect of 3D culture on their angiogenic capabilities. Macrophages grown in 3D culture displayed a significantly different morphology and arrangement under electron microscopy compared to those grown in 2D culture. Tube formation assays and chick embryo chorioallantoic membrane assays further revealed that 3D-cultured macrophages were less angiogenic than those in 2D culture. Whole-transcriptome sequencing showed that nearly 40% of genes were significantly differently expressed, including nine important angiogenic factors of which seven had been downregulated. In addition, the expression of almost all genes related to two important angiogenic pathways was decreased in 3D-cultured macrophages, including the two key angiogenic factors, VEGFA and ANG2. Together, the findings of our study improve our understanding of angiogenesis and 3D macrophage culture in tissues, and provide new avenues and methods for future research on macrophages. [ABSTRACT FROM AUTHOR]

Macrophages play important roles in angiogenesis; however, previous studies on macrophage angiogenesis have focused on traditional 2D cultures. In this study, we established a 3D culture system for macrophages using collagen microcarriers and assessed the effect of 3D culture on their angiogenic capabilities. Macrophages grown in 3D culture displayed a significantly different morphology and arrangement under electron microscopy compared to those grown in 2D culture. Tube formation assays and chick embryo chorioallantoic membrane assays further revealed that 3D-cultured macrophages were less angiogenic than those in 2D culture. Whole-transcriptome sequencing showed that nearly 40% of genes were significantly differently expressed, including nine important angiogenic factors of which seven had been downregulated. In addition, the expression of almost all genes related to two important angiogenic pathways was decreased in 3D-cultured macrophages, including the two key angiogenic factors, VEGFA and ANG2. Together, the findings of our study improve our understanding of angiogenesis and 3D macrophage culture in tissues, and provide new avenues and methods for future research on macrophages.

The guarantee of cell survival under hypoxic conditions and rapid vascularization is a key in tissue engineering strategies for treating bone defects. Our study aimed to establish the protective role of bone marrow mesenchymal stem cells (BMSCs) and human umbilical vein endothelial cells (HUVECs) in hypoxic conditions and realize rapid vascularization in bone defects. Resveratrol (Res), a non-flavonoid polyphenolic compound, and angiopoietin-2 (ANG2), a vascular activating factor, were applied to enhance BMSC and HUVEC survival, osteogenesis, and angiogenesis. The morphology, autophagy, viability, apoptosis, cycle, and osteogenic differentiation of BMSCs treated with Res were analyzed. The results indicated that Res could improve BMSC survival and differentiation via the autophagy pathway under hypoxic conditions. In addition, Res maintained HUVEC growth and proliferation in a hypoxic and ANG2 double-adverse environment via the autophagy pathway. To simulate a relatively hypoxic environment, small-aperture PEGDA/TCS hydrogels containing Res and ANG2 were prepared. BMSCs were cultured in the PEGDA/TCS scaffold and transplanted into a large tibial defect. CD31 immunofluorescence showed that the density and size of new blood vessels in the bone defect were significantly enhanced by ANG2 and Res at 8 weeks after surgery. H&E, Masson, and immunohistochemical staining results indicated that ANG2 combined with Res could promote new bone formation in defects. All these results suggested that Res combined with ANG2 may be a novel strategy for the targeted therapy of hypoxic bone defects with tissue engineering scaffolds.

Abstract Background Cavity effusion is common in patients with infectious diseases. However, the incidence rate and characteristics of serous cavity effusions (SCE) in septic patients are not clear to date. The objective of this study was to investigate the incidence and characteristics of SCE in septic patients and to explore the correlations between the bloody effusions and the illness severity/prognosis in septic patients. Methods From January 2010 to January 2015, a total of 214 patients with severe sepsis and septic shock were enrolled in this retrospective observational study. Thoracentesis or abdominal paracentesis was performed in 45 septic patients because of massive pleural effusions or ascites. The serum concentrations of VEGF, VEGFR, Ang, sICAM-1, sVCAM-1, E-selectin, Serpine1 and VE-cadherin in 45 septic patients underwent paracentesis were measured by enzyme-linked immunosorbent assay (ELISA). Results Of the 214 septic patients, 155 (72.4%) had SCE according to imaging or ultrasound manifestations. 45 subjects with SCE underwent therapeutic thoracentesis or abdominal paracentesis. Effusion laboratory analysis showed that exudates were predominant when compared with transudates (95.6% vs. 4.4%), and 16 (35.6%) patients suffered bloody effusions. Compared with patients with non-bloody effusions, those with bloody effusions showed higher critical illness scores (13 vs. 17 for APACHE II; 7 vs. 9 for SOFA), and higher mortality (6.9% vs. 62.5%). Moreover, patients with bloody effusions had delayed TT and APTT, increased D-dimer concentration, and higher serum levels of CRP and PCT (P

Background: Choroidal neovascularization (CNV) is a major cause of blindness in patients with age-related macular degeneration. CNV is characterized by new blood vessel growth and subretinal fluid accumulation, which results in mechanical pressure on retinal pigment epithelial (RPE) cells. The overexpression of RPE-derived angiogenic factors plays an important role in inducing CNV. In this work, we investigated the effect of mechanical stress on the expression of angiogenic factors in porcine RPE cells and determined the impact of conditioned medium on in-vitro angiogenesis. Results: The goal of this study was to determine whether low levels of acute mechanical stress during early CNV can induce the expression of angiogenic factors in RPE cells and accelerate angiogenesis. Using a novel device, acute mechanical stress was applied to primary porcine RPE cells and the resulting changes in the expression of major angiogenic factors, VEGF, ANG2, HIF-1α, IL6, IL8 and TNF-α, were examined using immunocytochemistry, qRT-PCR, and ELISA. An in vitro tube formation assay was used to determine the effect of secreted angiogenic proteins due to mechanical stress on endothelial tube formation by human umbilical vein endothelial cells (HUVECs). Our results showed an increase in the expression of VEGF, ANG2, IL-6 and IL-8 in response to mechanical stress, resulting in increased in vitro angiogenesis. Abnormal epithelial-mesenchymal transition (EMT) in RPE cells is also associated with CNV and further retinal degeneration. Our qRT-PCR results verified an increase in the expression of EMT genes, CDH2, VIM and FN1, in RPE cells. Conclusions: In conclusion, we showed that acute mechanical stress induces the expression of major angiogenic and EMT factors and promotes in vitro angiogenesis, suggesting that mechanical stress plays a role in promoting aberrant angiogenesis in AMD. [ABSTRACT FROM AUTHOR]

Aim: To quantify the expression of angiogenic growth factors (ANG2, VEGFA, TGFß1) and their corresponding receptors (VEGFR1, VGFR2, NRP1 and TGFßR1) in human dental pulps from extracted third molars with complete and incomplete root development. Methodology: Fifty‐six dental pulp samples obtained from freshly extracted human third molars were divided equally into two groups according to their stage of root development; 28 third molars with complete root development and 28 third molars with incomplete root development. All samples were processed and total RNA was extracted, cDNA was then synthetized for each sample and the target genes expression profiles for ANG2, VEGFA, VEGFR1, VEGFR2, NRP1, TGFß1 and TGFßR1 were obtained by RT2‐PCR. The data was analysed with a Student's t‐test to compare the replicate ∆∆Ct values for each gene. Results: Teeth with incomplete root development were associated with a significantly greater gene expression of TGFβR1 (P = 0.03), whereas in teeth with complete root development the genes that had significantly greater expression were VEGFA (P = 0.04). Conclusion: The angiogenic growth factors (ANG2, VEGFA, TGFβ1) and their receptors (NRP1, VEGFR1, VEGFR2 and TGFßR1) were expressed in pulps of teeth with complete and incomplete root development measured by RT2‐PCR, with TGFBR1 genes being significantly different in teeth with incomplete root development and VEGFA genes in teeth with complete root development. [ABSTRACT FROM AUTHOR]

Ovarian cancer is the leading cause of death among all the gynecological cancers in the USA. Ovarian cancer employs a unique mode of metastasis, as exfoliated tumor cells disseminate within the peritoneal cavity, colonizing in several sites as well as accumulating ascites. Tumor recurrence and widespread metastasis are significant factors contributing to poor prognosis. PRSS21 is a metastasis-associated ovarian cancer gene that encodes the glycosyl-phosphatidylinositol-linked serine protease, testisin. Testisin expression is increased in multiple ovarian tumor types, with relatively little expression in normal tissues, but is differentially decreased in metastatic ovarian serous carcinomas compared to primary tumors. Here we explored the function of testisin in late-stage ovarian cancer progression using a murine xenograft model of ovarian intraperitoneal tumor metastasis. Increased tumor testisin expression inhibited intra-peritoneal tumor seeding and colonization, ascites accumulation, and metastatic tumor burden that was dependent on catalytically active testisin. The known testisin substrate, protease-activated receptor-2 (PAR-2), is a target of testisin activity. Gene profiling and mechanistic studies demonstrate that testisin activity suppresses the synthesis and secretion of pro-angiogenic angiopoietins, ANG2 and ANGPTL4, which normally promote vascular leak and edema. These observations support a model wherein testisin activates PAR-2 to antagonize proangiogenic angiopoietins that modulate vascular permeability and ascites accumulation associated with ovarian tumor metastasis. Key messages: Testisin inhibits metastatic ovarian tumor burden and ascites production. Testisin activity antagonizes ANG2 and ANGPTL4 synthesis and secretion. PAR-2 is a proteolytic target of testisin on the surface of ovarian cancer cells. [ABSTRACT FROM AUTHOR]

Cooption of the host vasculature is a strategy that some cancers use to sustain tumor progression without--or before--angiogenesis or in response to antiangiogenic therapy. Facilitated by certain growth factors, cooption can mediate tumor infiltration and confer resistance to antiangiogenic drugs. Unfortunately, this mode of tumor progression is difficult to target because the underlying mechanisms are not fully understood. Here, we analyzed the dynamics of vessel cooption during tumor progression and in response to antiangiogenic treatment in gliomas and brain metastases. We followed tumor evolution during escape from antiangiogenic treatment as cancer cells coopted, and apparently mechanically compressed, host vessels. To gain deeper understanding, we developed a mathematical model, which incorporated compression of coopted vessels, resulting in hypoxia and formation of new vessels by angiogenesis. Even if antiangiogenic therapy can block such secondary angiogenesis, the tumor can sustain itself by coopting existing vessels. Hence, tumor progression can only be stopped by combination therapies that judiciously block both angiogenesis and cooption. Furthermore, the model suggests that sequential blockade is likely to be more beneficial than simultaneous blockade. [ABSTRACT FROM AUTHOR]

Background/Aims: The pro-angiogenic factors vascular endothelial growth factor (VEGF) and angiopoietins (Angs) play a prominent role in synovial angiogenesis, an early and critical event in the pathogenesis of rheumatoid arthritis (RA). Interleukin (IL)-35 is an anti-inflammatory cytokine that attenuates collagen-induced arthritis, however, the mechanisms involved are not fully understood. Methods: The effects of IL-35 on endothelial cell migration, adhesion, and tube formation were examined using human umbilical vein endothelial cells (HUVEC) in vitro. The effects of IL-35 on vessel formation in vivo were examined using a murine Matrigel plugs model. MMP2/MMP9 and IL-6/IL-8 secretion were assessed by zymography and ELISA, respectively. The crosstalk between IL-35, VEGF, and Ang2 in HUVECs and RA synovial tissue explants was investigated. Results: IL-35 inhibited basal and VEGF-induced HUVEC migration and adhesion in vitro as well as tube formation in vitro and in vivo. VEGF increased Ang2 secretion by HUVECs and RA synovial tissue explants, and exogenous Ang2 promoted HUVEC migration, adhesion, and tube formation with similar potency to VEGF. Blocking the Ang/Tie2 pathway with a Tie2 kinase antibody inhibited the proangiogenic effects of exogenous Ang2 and VEGF in HUVECs. IL-35 inhibited basal and VEGF-induced Ang2 secretion by HUVECs and RA synovial tissue explants; it also antagonized the proangiogenic effects of exogenous Ang2 in HUVECs. Moreover, IL-35 reduced basal and VEGF/Ang2-induced MMP2/MMP9 and IL-6/IL-8 secretion. Conclusion: These results suggested that IL-35 restrains RA angiogenesis and inflammation by downregulating basal and VEGF-induced Ang2 secretion as well as disrupting Ang2/Tie2 signal transduction. Our findings extend current understanding of mechanisms regulating RA angiogenesis and may support development of novel angiogenesis-targeting therapeutics for RA treatment.

Endothelial cells that form the inner layer of blood and lymphatic vessels are important regulators of vascular functions and centrally involved in the pathogenesis of vascular diseases. In addition to the vascular endothelial growth factor (VEGF) receptor pathway, the angiopoietin (Ang)–Tie system is a second endothelial cell specific ligand–receptor signalling system necessary for embryonic cardiovascular and lymphatic development. The Ang–Tie system also regulates postnatal angiogenesis, vessel remodelling, vascular permeability and inflammation to maintain vascular homoeostasis in adult physiology. This system is implicated in numerous diseases where the vasculature has an important contribution, such as cancer, sepsis, diabetes, atherosclerosis and ocular diseases. Furthermore, mutations in the TIE2 signalling pathway cause defects in vascular morphogenesis, resulting in venous malformations and primary congenital glaucoma. Here, we review recent advances in the understanding of the Ang–Tie signalling system, including cross-talk with the vascular endothelial protein tyrosine phosphatase (VE-PTP) and the integrin cell adhesion receptors, focusing on the Ang–Tie system in vascular development and pathogenesis of vascular diseases. [ABSTRACT FROM AUTHOR]

Purpose: Angiopoietin 2 (Ang2) and soluble receptor for advanced glycation end products (sRAGE) are markers of endothelial and pulmonary epithelial damage with prognostic implications in adult acute respiratory distress syndrome (ARDS), but unclear significance in pediatric ARDS (PARDS).Methods: This was a prospective, observational study in children with PARDS (2012 Berlin and 2015 PALICC definitions) at the Children's Hospital of Philadelphia. Plasma was collected within 48 h of PARDS onset and biomarkers quantified by enzyme-linked immunosorbent assay.Results: In 82 children with PARDS (12 deaths, 15 %), Ang2 and sRAGE were higher in non-survivors than survivors (p < 0.01 for both). Mortality was highest in patients with Ang2 and sRAGE levels both above median values. Ang2 and sRAGE correlated with the number of non-pulmonary organ failures (both p < 0.001). Ang2 was higher in indirect lung injury and in immunocompromised children. In stratified analysis, both Ang2 and sRAGE were associated with mortality only in direct lung injury and in immunocompetent children, with no association evident in indirect lung injury or in immunocompromised children.Conclusions: Ang2 and sRAGE in early PARDS were higher in non-survivors than survivors and strongly correlated with number of non-pulmonary organ failures. When stratified by type of lung injury, Ang2 and sRAGE were associated with mortality only in direct lung injury. Similarly, when stratified by immunocompromised status, Ang2 and sRAGE were associated with mortality only in immunocompetent children. The utility of these biomarkers for prognostication and risk stratification requires investigation. [ABSTRACT FROM AUTHOR]

Despite encouraging response rate of bevacizumab (BVZ) in recurrent glioblastoma, many patients do not respond to this schedule and most of the responders develop an early relapse. Plasma concentrations of VEGF, PlGF, Ang2, and sTie2 were assessed by ELISA before and during BVZ treatment in seventy patients. Baseline levels of VEGF-A, and PlGF were higher in patients than in healthy volunteers, whereas no difference was found for Ang2, and sTie2. No biomarker at baseline was associated with response, PFS or OS. At recurrence, the authors observed an increase of Ang2 suggesting that Ang2/sTie2 could be involved in the resistance to BVZ. [ABSTRACT FROM AUTHOR]

Almost all cancers show intrinsic and/or evasive resistance to vascular endothelial growth factor ( VEGF) inhibitors by multiple mechanisms. Serum angiopoietin-2 (Ang2) level has been proposed as a potential biomarker of VEGF inhibitor response in several cancers. From these clinical observations, the Ang2 and Tie2 (its receptor) axis has been focused on as a promising target. Here, we show a novel strategy to circumvent the resistance by combining multi-tyrosine kinase inhibitors lenvatinib ( VEGF receptor, fibroblast growth factor receptor, and RET inhibitor) and golvatinib (E7050; c-Met, Tie2, and EphB4 inhibitor). Tie2 identifies a highly pro-angiogenic macrophage subset, Tie2-expressing macrophages ( TEM). Angi-Tie2 and EphB4-EphrinB2 signaling plays critical roles in pericyte-mediated vessel stabilization. In vitro analyses suggested that golvatinib combined with lenvatinib inhibited pericyte-mediated vessel stabilization and TEM differentiation. In thyroid and endometrial cancer models, golvatinib and lenvatinib inhibited pericyte network development and TEM infiltration, resulting in severe perfusion disorder and massive apoptosis. Body weight loss was tolerable, and no macroscopic change was observed. These preclinical studies suggest that modulation of the tumor microenvironment by a strategic and well-tolerated combination of multi-targeting tyrosine kinase inhibitors may sensitize cancer to VEGF inhibitors. [ABSTRACT FROM AUTHOR]

To investigate the intervention therapy effect on the growth of malignant melanoma, we have made an observation of expression levels of Ang2 in malignant melanoma cells, which was transduced by small interfering RNA (Ang2-siRNA) of Ang2 in vitro and in vivo. We successfully constructed Ang2-siRNA lent virus, and constructed nude mice model by transplanting malignant melanoma. Ang2-siRNA lent virus inhibited Ang2 mRNA of malignant melanoma in vitro and in vivo, and inhibited malignant melanoma tumor growth at the same time. Ang2-siRNA lent virus can interfere expression levels of Ang2 in malignant melanoma cells, inhibit tumor growth, and silent Ang2 gene expression, which may pave a new way for clinical gene therapy of malignant melanoma. [ABSTRACT FROM AUTHOR]

Abstract: Molecular mechanisms regulating the remodeling of the lymphatic vasculature from an immature plexus of vessels to a hierarchal network of initial and collecting lymphatics are not well understood. One gene thought to be important for this process is Angiopoietin-2 (Ang-2). Ang2−/− mice have previously been reported to exhibit an abnormal lymphatic phenotype but the precise nature of the lymphatic defects and the underlying mechanisms have yet to be defined. Here we demonstrate by whole-mount immunofluorescence staining of ear skin and mesentery that lymphatic vessels in Ang2−/− mice fail to mature and do not exhibit a collecting vessel phenotype. Furthermore, dermal lymphatic vessels in Ang2−/− pups prematurely recruit smooth muscle cells and do not undergo proper postnatal remodeling. In contrast, Ang2 knock-out Ang1 knock-in mice do develop a hierarchal lymphatic vasculature, suggesting that activation of Tie-2 is required for normal lymphatic development. Taken together, this work pinpoints a specific lymphatic defect of Ang2−/− mice and further defines the sequential steps in lymphatic vessel remodeling. [Copyright &y& Elsevier]

Decidualization is accompanied by extensive angiogenesis, which is an essential step in the maturation of new blood vessels in mammalian pregnancy. The purpose of this study was to determine a distribution of uNK cells (CD56+ uNK or CD56bright cells) in human decidua of the first-trimester pregnancy, and investigate whether uNK cells in human decidua could express vascular endothelial growth factor (VEGF-A) and/or angiopoietin2 (Ang2). Our immunohistochemical staining results demonstrated that a great amount of uNK (CD56+) cells scattered throughout the decidual stroma and near endometrial gland and spiral vessels in human decidua. The protein expression of VEGF-A and Ang2 was detected in decidual stroma cells, capillary endothelial cells and glandular cells in tissue specimens. There was a positive correlation between microvessel density (MVD) and the number of the CD56-positive uNK cells in decidual stroma, and also between the number of the CD56-positive uNK cells and VEGF-A protein expression in the tissue. In addition, we found that uNK cells in human decidua could express VEGF-A mRNA, but not Ang2 mRNA, in isolated uNK cells in human decidua of the first-trimester gestation by combination of LCM and Nested-PCR. Our study indicated that uNK cells, through expressing VEGF-A, may play an important role in the angiogenic response at the time of human decidualization and early placenta development. [ABSTRACT FROM AUTHOR]

The present study demonstrated that the vessel number in bone marrow biopsies from acute myeloid leukaemia (AML) patients (n = 23) was significantly increased at diagnosis compared with normal bone marrow (P = 0·019) and was restored to normal levels after achieving complete remission (P = 0·03). The in vitro angiogenic potential of culture supernatant of AML cells was assessed using endothelial cell (EC) migration and proliferation assays. Increased EC migration and EC proliferation was induced in 7/20 and 19/20 AML supernatents respectively. The degree of in vivo neovascularization did not correlate with the ability of AML cells to stimulate in vitro endothelial cell migration and/or proliferation. This might be in part a result of the heterogeneous pattern of angiogenic factors produced by AML cells. The expression of different angiogenic factors was studied using reverse transcription polymerase chain reaction. Cells from 17/20 AML patients showed wide variation in spontaneous vascular endothelial growth factor (VEGF) expression, 4/19 expressed varied spontaneous blastic fibroblast growth factor mRNA levels and all patient samples showed spontaneous interleukin 8 mRNA expression. All AML samples expressed matrix metalloproteinase (MMP)-2 and/or MMP-9. VEGF mRNA expression correlated well with protein level (P = 0·006). A correlation was found between the degree of VEGF expression and neoangiogenesis (correlation coefficient = 0·448, P = 0·05). These results suggest that malignant cell proliferation, angiogenesis and VEGF expression are linked in AML and might contribute to the growth advantage of the malignant counterpart as a result of the paracrine production of growth factors produced by the surrounding endothelial cells. [ABSTRACT FROM AUTHOR]

The guarantee of cell survival under hypoxic conditions and rapid vascularization is a key in tissue engineering strategies for treating bone defects. Our study aimed to establish the protective role of bone marrow mesenchymal stem cells (BMSCs) and human umbilical vein endothelial cells (HUVECs) in hypoxic conditions and realize rapid vascularization in bone defects. Resveratrol (Res), a non-flavonoid polyphenolic compound, and angiopoietin-2 (ANG2), a vascular activating factor, were applied to enhance BMSC and HUVEC survival, osteogenesis, and angiogenesis. The morphology, autophagy, viability, apoptosis, cycle, and osteogenic differentiation of BMSCs treated with Res were analyzed. The results indicated that Res could improve BMSC survival and differentiation via the autophagy pathway under hypoxic conditions. In addition, Res maintained HUVEC growth and proliferation in a hypoxic and ANG2 double-adverse environment via the autophagy pathway. To simulate a relatively hypoxic environment, small-aperture PEGDA/TCS hydrogels containing Res and ANG2 were prepared. BMSCs were cultured in the PEGDA/TCS scaffold and transplanted into a large tibial defect. CD31 immunofluorescence showed that the density and size of new blood vessels in the bone defect were significantly enhanced by ANG2 and Res at 8 weeks after surgery. H&E, Masson, and immunohistochemical staining results indicated that ANG2 combined with Res could promote new bone formation in defects. All these results suggested that Res combined with ANG2 may be a novel strategy for the targeted therapy of hypoxic bone defects with tissue engineering scaffolds. [ABSTRACT FROM AUTHOR]

Despite significant advances made in cancer treatment, the development of therapeutic resistance to anticancer drugs represents a major clinical problem that limits treatment efficacy for cancer patients. Herein, we focus on the response and resistance to current antiangiogenic drugs and immunotherapies and describe potential strategies for improved treatment outcomes. Antiangiogenic treatments that mainly target vascular endothelial growth factor (VEGF) signaling have shown efficacy in many types of cancer. However, drug resistance, characterized by disease recurrence, has limited therapeutic success and thus increased our urgency to better understand the mechanism of resistance to inhibitors of VEGF signaling. Moreover, cancer immunotherapies including immune checkpoint inhibitors (ICIs), which stimulate antitumor immunity, have also demonstrated a remarkable clinical benefit in the treatment of many aggressive malignancies. Nevertheless, the emergence of resistance to immunotherapies associated with an immunosuppressive tumor microenvironment has restricted therapeutic response, necessitating the development of better therapeutic strategies to increase treatment efficacy in patients. Angiopoietin-2 (ANG2), which binds to the receptor tyrosine kinase TIE2 in endothelial cells, is a cooperative driver of angiogenesis and vascular destabilization along with VEGF. It has been suggested in multiple preclinical studies that ANG2-mediated vascular changes contribute to the development and persistence of resistance to anti-VEGF therapy. Further, emerging evidence suggests a fundamental link between vascular abnormalities and tumor immune evasion, supporting the rationale for combination strategies of immunotherapy with antiangiogenic drugs. In this review, we discuss the recent mechanistic and clinical advances in targeting angiopoietin signaling, focusing on ANG2 inhibition, to enhance therapeutic efficacy of antiangiogenic and ICI therapies. In short, we propose that a better mechanistic understanding of ANG2-mediated vascular changes will provide insight into the significance of ANG2 in treatment response and resistance to current antiangiogenic and ICI therapies. These advances will ultimately improve therapeutic modalities for cancer treatment. [ABSTRACT FROM AUTHOR]

Background: Proper oxygen balance in the dental pulp is essential for cell metabolism. Angiogenesis in the pulp is a constant process during the life of the tooth. Hypoxia indicators in a tissue, such as HIF-1α, as well as vascular destabilization markers, such as ANG2 and its receptor TIE2, are necessary for angiogenesis. Therefore the purpose of this study is to evaluate the expression of HIF-1α, ANG1, ANG2 and TIE2 in dental pulp as early angiogenesis indicators in teeth with complete and incomplete root development. Methods: Forty human dental pulps were obtained from freshly extracted third molars divided into two groups: incomplete (n = 20) and complete (n = 20) root development. Dental pulps were stored at − 80 °C, defrosted in an ice bath and re-frozen with their respective thaws to disintegrate the tissue. Three sonication cycles were performed until the tissues were homogenized, then thaw were centrifuged and the supernatant was collected for the detection of the markers to be studied. The samples were processed for the ELISA test using the ELISA-sandwich principle. Student t and Mann–Whitney U tests were performed to determine statistically significant differences between groups. Results: In the complete root development, HIF-1α, ANG1, ANG2 and TIE2 expressions were significantly higher than their expression in the incomplete root development group. Conclusions: The angiogenic process seems to be a physiological process in the dental pulp. Angiogenic activity is higher in teeth with mature than immature apex teeth. [ABSTRACT FROM AUTHOR]

Sepsis is associated with significant mortality. Early diagnosis and prognosis of patients with sepsis is still a difficult clinical challenge. In this study, the ability of plasma PTX3 (pentraxin 3), MCP1 (monocyte chemoattractant protein 1) and Ang (angiopoietin)1/2 was investigated to evaluate the severity of sepsis. Blood samples were obtained from 43 patients with sepsis. A total of 33 post‐surgery patients with infections and 25 healthy individuals served as controls. The results showed that plasma PTX3, MCP1 and Ang2 significantly increased in patients on the first day of septic shock onset, while sepsis patients had significantly higher Ang2 level, compared with controls. Furthermore, PTX3, MCP1 and Ang2 had high AUROC values in patients with septic shock on the first day of sepsis onset. The findings suggest that PTX3, MCP1 and Ang2 maybe early predictors to evaluate the severity of sepsis and septic shock with the latest Sepsis 3.0 definitions. [ABSTRACT FROM AUTHOR]

Background: Cavity effusion is common in patients with infectious diseases. However, the incidence rate and characteristics of serous cavity effusions (SCE) in septic patients are not clear to date. The objective of this study was to investigate the incidence and characteristics of SCE in septic patients and to explore the correlations between the bloody effusions and the illness severity/prognosis in septic patients. Methods: From January 2010 to January 2015, a total of 214 patients with severe sepsis and septic shock were enrolled in this retrospective observational study. Thoracentesis or abdominal paracentesis was performed in 45 septic patients because of massive pleural effusions or ascites. The serum concentrations of VEGF, VEGFR, Ang, sICAM-1, sVCAM-1, E-selectin, Serpine1 and VE-cadherin in 45 septic patients underwent paracentesis were measured by enzyme-linked immunosorbent assay (ELISA). Results: Of the 214 septic patients, 155 (72.4%) had SCE according to imaging or ultrasound manifestations. 45 subjects with SCE underwent therapeutic thoracentesis or abdominal paracentesis. Effusion laboratory analysis showed that exudates were predominant when compared with transudates (95.6% vs. 4.4%), and 16 (35.6%) patients suffered bloody effusions. Compared with patients with non-bloody effusions, those with bloody effusions showed higher critical illness scores (13 vs. 17 for APACHE II; 7 vs. 9 for SOFA), and higher mortality (6.9% vs. 62.5%). Moreover, patients with bloody effusions had delayed TT and APTT, increased D-dimer concentration, and higher serum levels of CRP and PCT (P < 0.05). In addition, the serum levels of Ang2, sVCAM-1 and E-selectin were significantly higher in patients with bloody effusions than in those with non-bloody effusions (P < 0.05). However, the serum level of VEGFR2 was lower in patients with bloody fluids (P = 0.025). Conclusions: The incidence of serous cavity effusion is high in patients with sepsis. The septic patients with bloody effusions suffer a more inflammatory burden and a worse prognosis compared to septic patients with non-bloody effusions. [ABSTRACT FROM AUTHOR]