Research progress in the pathogenesis of hormone-induced femoral head necrosis based on microvessels: a systematic review.

Hormonal necrosis of the femoral head is caused by long-term use of glucocorticoids and other causes of abnormal bone metabolism, lipid metabolism imbalance and blood microcirculation disorders in the femoral head, resulting in bone trabecular fracture, bone tissue necrosis collapse, and hip dysfunction. It is the most common type of non-traumatic necrosis of the femoral head, and its pathogenesis is complex, while impaired blood circulation is considered to be the key to its occurrence. There are a large number of microvessels in the femoral head, among which H-type vessels play a decisive role in the "angiogenesis and osteogenesis coupling", and thus have an important impact on the occurrence and development of femoral head necrosis. Glucocorticoids can cause blood flow injury of the femoral head mainly through coagulation dysfunction, endothelial dysfunction and impaired angiogenesis. Glucocorticoids may inhibit the formation of H-type vessels by reducing the expression of HIF-1α, PDGF-BB, VGEF and other factors, thus causing damage to the "angiogenesis-osteogenesis coupling" and reducing the ability of necrosis reconstruction and repair of the femoral head. Leads to the occurrence of hormonal femoral head necrosis. Therefore, this paper reviewed the progress in the study of the mechanism of hormone-induced femoral head necrosis based on microvascular blood flow at home and abroad, hoping to provide new ideas for the study of the mechanism of femoral head necrosis and provide references for clinical treatment of femoral head necrosis.

CD44 promotes angiogenesis in myocardial infarction through regulating plasma exosome uptake and further enhancing FGFR2 signaling transduction.

BACKGROUND: Since angiogenesis occurs as the pathological process following myocardial infarction to alleviate ischemia, therapeutic angiogenesis has been proposed to be a cardioprotective strategy. CD44 has been implicated in endothelial cell functions and its role has been well established in angiogenesis for years. Although recent studies indicate the close correlation between CD44 and exosome, as well as the two being implicated in myocardial ischemia pathological processes, the effect and the underlying mechanism of CD44 and its regulated plasma exosome in pathological angiogenesis post-myocardial infarction have not been fully elucidated. METHODS: In this study, we used CD44 knockout mice to study the in vivo impacts of CD44 on ischemic angiogenesis in myocardial infarction. Mouse cardiac function was measured by echocardiography, histological changes were observed by Evans Blue and TTC-double staining and Masson's trichrome staining, and molecular changes were detected by immunofluorescence. In the in vitro study, CD44 knockout HUVECs were generated and CD44 inhibitor was used to study the mechanism of CD44 on angiogenesis. We performed the immunoprecipitation, proximity ligation assay, and super-resolution imaging to study the mechanistic regulation of FGFR2 signaling transduction by CD44. Importantly, we also isolated plasma exosomes from myocardial infarction model mice and studied the effect of plasma exosomes on the activation of the FGFR2 signaling pathway and the related phenotypic alterations, including exosomes uptake and angiogenic function in primary mouse microvascular endothelial cells, and further discovered the regulation mechanism of exosomal miRNAs. RESULTS: We observed that the expression of CD44 in the border zone of the infarcted heart was tightly related to pathological angiogenesis following myocardial ischemia. The depletion of CD44 impaired angiogenesis and impacts biogenesis and proangiogenic function of plasma exosomes. Subsequently, we found that CD44 mediated the activation of the FGFR2 signaling pathway as well as the caveolin 1-dependent uptake of exosomes in vascular endothelial cells. Most importantly, the proangiogenic therapeutic effect of plasma exosomal miRNAs depended upon the participation of CD44/FGFR2 signaling transduction in vascular endothelial cells. CONCLUSION: CD44 and its regulated plasma exosomes have crucial potent angiogenic activity. Our studies elucidate that CD44 plays a key role in plasma exosomal miRNA-enhanced angiogenic FGFR2 singling transduction and ischemic angiogenesis in the early stage of myocardial infarction.

Transarterial chemoembolization (TACE) plus apatinib vs. TACE alone for hepatocellular carcinoma.

OBJECTIVE: Transarterial chemoembolization (TACE) is a common therapy for hepatocellular carcinoma (HCC), while TACE-induced tumor angiogenesis would increase progression and metastasis risk. Besides, apatinib possesses the capability of inhibiting tumor angiogenesis. Thus, this study aimed to explore the efficacy and safety of TACE plus apatinib compared to TACE alone in HCC patients. METHODS: Ninety-six intermediate-advanced HCC patients were retrospectively enrolled and classified into TACE plus apatinib group (N = 45) and TACE group (N = 51) based on the treatment. RESULTS: Objective response rate (68.9% vs. 47.1%) was increased in TACE plus apatinib group than in TACE group (P = 0.031). However, no difference was found in disease-control rate between groups (95.6% vs. 86.3%) (P = 0.167). Progression-free survival (PFS) (median PFS (95% confidence interval (CI)): 20.0 (13.2-26.8) vs. 14.0 (8.3-19.7) months) was enhanced in TACE plus apatinib group compared with TACE group (P = 0.030), while no difference was found in overall survival between groups (P = 0.060). Additionally, multivariate Cox's analysis presented that TACE plus apatinib (vs. TACE alone) independently associated with prolonged PFS (P = 0.043, hazard ratio = 0.617). Regarding safety profile, no difference in liver function indexes (albumin, total bilirubin, alanine aminotransferase and aspartate aminotransferase) was found after treatment between two groups; meanwhile, only the incidence of hand-foot skin reaction (24.4% vs. 7.8%) was higher in TACE plus apatinib group compared to TACE group (P = 0.025), while no difference was found in other adverse events between two groups (all P > 0.05). CONCLUSION: TACE plus apatinib illustrates a superior efficacy with tolerable safety than TACE alone in intermediate-advanced HCC patients.

The Utilization of Bevacizumab in Patients with Advanced Ovarian Cancer: A Systematic Review of the Mechanisms and Effects.

Most ovarian cancer cases are diagnosed at an advanced stage (III or IV), in which a primary debulking surgery combined with adjuvant systemic chemotherapy is the standard management. Since targeted therapy is less toxic to human cells than systemic chemotherapy, it has drawn much attention and become more popular. Angiogenesis is a critical process during the proliferation of ovarian cancer cells. Currently, many studies have put emphases on anti-angiogenetic medication, such as bevacizumab, the first and most investigated angiogenesis inhibitor that can exert anti-neoplastic effects. Bevacizumab is a recombinant humanized monoclonal antibody that has been approved for first-line maintenance treatment of advanced ovarian cancer. This review is a summary of current literature about the molecular mechanisms of actions, safety, and effects of bevacizumab for use in advanced epithelial ovarian cancer. Some common side effects of bevacizumab will be also discussed. As an inhibitor of angiogenesis, bevacizumab binds to circulating vascular endothelial growth factor (VEGF) and thereby inhibits the binding of VEGF to its receptors on the surface of endothelial cells. Neutralization of VEGF prevents neovascularization and leads to apoptosis of tumor endothelial cells and a decrease in interstitial fluid pressure within the tumors, which allows greater capacity for chemotherapeutic drugs to reach specific targeted sites. Grossly, bevacizumab has demonstrated some significant therapeutic benefits in many randomized trials in combination with the standard chemotherapy for advanced epithelial ovarian cancer. Based on the available evidence, a higher dosage and a longer duration of bevacizumab appear to achieve better therapeutic effects and progression-free survival. On the other hand, patients with more severe diseases or at a higher risk of progression seem to benefit more from bevacizumab use. However, many unknown aspects of bevacizumab, including detailed mechanisms of actions, effectiveness, and safety for the treatment of ovarian cancer, warrant further investigation.

Risk factors for ocular neovascularization after central retinal artery occlusion.

BACKGROUND: To report the incidence and risk factors associated with ocular neovascularization (NV) in patients with central retinal artery occlusion (CRAO). METHODS: This retrospective study included patients diagnosed with acute CRAO in a single tertiary center. Medical charts were reviewed for ocular NV occurrences. We analyzed systemic and ocular conditions on first visit and demographic data. RESULTS: Eighty-seven eyes were eligible for this study. Among these, 13 eyes had ocular NV after CRAO, with an incidence of 15%. The prevalences of hypertension, diabetes mellitus, history of stroke, chronic kidney disease (CKD), and age at first visit were higher among patients with ocular NV than among patients without ocular NV after CRAO. Moreover, most patients with CKD in the ocular NV group had undergone dialysis. A multivariate regression analysis revealed that CKD (hazard ratio [HR]: 9.27, 95% CI, 1.87-46.05, p = 0.006) and glaucoma history (HR: 7.52, 95% CI, 1.14-49.46, p = 0.036) were significant risk factors for developing ocular NV among patients with CRAO. CONCLUSION: CKD and glaucoma history were significant risk factors for developing ocular NV after CRAO, particularly among patients that underwent dialysis.

Impact of Chemotherapy and Radiation Therapy on Inflammatory Response, Neovascularization, and Capsule Formation of Acellular Dermal Matrix in Breast Reconstruction: Analysis of the BREASTrial Biopsy Specimens.

BACKGROUND: The Breast Reconstruction Evaluation of Acellular Dermal Matrix as a Sling Trial is a single-center, blinded, prospective, randomized, controlled trial established to compare outcomes using two popular types of acellular dermal matrices, AlloDerm and DermaMatrix, in tissue expander breast reconstruction. This study used the acellular dermal matrix biopsy specimens from the trial to evaluate how adjuvant therapy influences inflammation, neovascularization, and capsule formation of the acellular dermal matrix. METHODS: Punch biopsy specimens were taken at the time of expander exchange and were analyzed by a blinded pathologist. The inflammatory response was quantified by the number of fibroblasts, giant cells, and lymphocytes. Neovascularization and capsule formation were similarly quantified by the number of new capillaries and capsule presence and thickness, respectively. RESULTS: Histology specimens were collected from 109 patients (170 breasts). In the absence of adjuvant therapy, there was no significant difference between AlloDerm and DermaMatrix in terms of inflammation, neovascularization, or capsule thickness. Both acellular dermal matrices showed a significant decrease in inflammation and neovascularization with adjuvant therapy. When chemotherapy and radiation therapy were used, the decrease in inflammation was greatest for the group reconstructed with DermaMatrix (p < 0.039). CONCLUSIONS: Adjuvant therapy influences the inflammatory response, neovascularization, and capsule formation in both acellular dermal matrices. Adjuvant therapy has a protective effect on the inflammatory response toward both acellular dermal matrices in breast reconstruction. In the setting of chemotherapy and radiation therapy, DermaMatrix produced the greatest reduction in inflammation. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, II.

Chemotherapy: a double-edged sword in cancer treatment.

Chemotherapy is a well-known and effective treatment for different cancers; unfortunately, it has not been as efficient in the eradication of all cancer cells as been expected. The mechanism of this failure was not fully clarified, yet. Meanwhile, alterations in the physiologic conditions of the tumor microenvironment (TME) were suggested as one of the underlying possibilities. Chemotherapy drugs can activate multiple signaling pathways and augment the secretion of inflammatory mediators. Inflammation may show two opposite roles in the TME. On the one hand, inflammation, as an innate immune response, tries to suppress tumor growth but on the other hand, it might be not powerful enough to eradicate the cancer cells and even it can provide appropriate conditions for cancer promotion and relapse as well. Therefore, the administration of mild anti-inflammatory drugs during chemotherapy might result in more successful clinical results. Here, we will review and discuss this hypothesis. Most chemotherapy agents are triggers of inflammation in the tumor microenvironment through inducing the production of senescence-associated secretory phenotype (SASP) molecules. Some chemotherapy agents can induce systematic inflammation by provoking TLR4 signaling or triggering IL-1B secretion through the inflammasome pathway. NF-kB and MAPK are key signaling pathways of inflammation and could be activated by several chemotherapy drugs. Furthermore, inflammation can play a key role in cancer development, metastasis and exacerbation.

Activation of the GPR35 pathway drives angiogenesis in the tumour microenvironment.

OBJECTIVE: Primary sclerosing cholangitis (PSC) is in 70% of cases associated with inflammatory bowel disease. The hypermorphic T108M variant of the orphan G protein-coupled receptor GPR35 increases risk for PSC and ulcerative colitis (UC), conditions strongly predisposing for inflammation-associated liver and colon cancer. Lack of GPR35 reduces tumour numbers in mouse models of spontaneous and colitis associated cancer. The tumour microenvironment substantially determines tumour growth, and tumour-associated macrophages are crucial for neovascularisation. We aim to understand the role of the GPR35 pathway in the tumour microenvironment of spontaneous and colitis-associated colon cancers. DESIGN: Mice lacking GPR35 on their macrophages underwent models of spontaneous colon cancer or colitis-associated cancer. The role of tumour-associated macrophages was then assessed in biochemical and functional assays. RESULTS: Here, we show that GPR35 on macrophages is a potent amplifier of tumour growth by stimulating neoangiogenesis and tumour tissue remodelling. Deletion of Gpr35 in macrophages profoundly reduces tumour growth in inflammation-associated and spontaneous tumour models caused by mutant tumour suppressor adenomatous polyposis coli. Neoangiogenesis and matrix metalloproteinase activity is promoted by GPR35 via Na/K-ATPase-dependent ion pumping and Src activation, and is selectively inhibited by a GPR35-specific pepducin. Supernatants from human inducible-pluripotent-stem-cell derived macrophages carrying the UC and PSC risk variant stimulate tube formation by enhancing the release of angiogenic factors. CONCLUSIONS: Activation of the GPR35 pathway promotes tumour growth via two separate routes, by directly augmenting proliferation in epithelial cells that express the receptor, and by coordinating macrophages' ability to create a tumour-permissive environment.

SIRT1-mediated deacetylation of FOXO3a transcription factor supports pro-angiogenic activity of interferon-deficient tumor-associated neutrophils.

Angiogenesis plays an important role during tumor growth and metastasis. We could previously show that Type I interferon (IFN)-deficient tumor-associated neutrophils (TANs) show strong pro-angiogenic activity, and stimulate tumor angiogenesis and growth. However, the exact mechanism responsible for their pro-angiogenic shift is not clear. Here, we set out to delineate the molecular mechanism and factors regulating pro-angiogenic properties of neutrophils in the context of Type I IFN availability. We demonstrate that neutrophils from IFN-deficient (Ifnar1-/- ) mice efficiently release pro-angiogenic factors, such as VEGF, MMP9 or BV8, and thus significantly support the vascular normalization of tumors by increasing the maturation of perivascular cells. Mechanistically, we could show here that the expression of pro-angiogenic factors in neutrophils is controlled by the transcription factor forkhead box protein O3a (FOXO3a), which activity depends on its post-translational modifications, such as deacetylation or phosphorylation. In TANs isolated from Ifnar1-/- mice, we observe significantly elevated SIRT1, resulting in SIRT1-mediated deacetylation of FOXO3a, its nuclear retention and activation. Activated FOXO3a supports in turn the transcription of pro-angiogenic genes in TANs. In the absence of SIRT1, or after its inhibition in neutrophils, elevated kinase MEK/ERK and PI3K/AKT activity is observed, leading to FOXO3a phosphorylation, cytoplasmic transfer and inactivation. In summary, we have found that FOXO3a is a key transcription factor controlling the angiogenic switch of neutrophils. Post-translational FOXO3a modifications regulate its transcriptional activity and, as a result, the expression of pro-angiogenic factors supporting development of vascular network in growing tumors. Therefore, targeting FOXO3a activity could provide a novel strategy of antiangiogenic targeted therapy for cancer.

Microbial metabolite deoxycholic acid promotes vasculogenic mimicry formation in intestinal carcinogenesis.

A high-fat diet (HFD) leads to long-term exposure to gut microbial metabolite secondary bile acids, such as deoxycholic acid (DCA), in the intestine, which is closely linked to colorectal cancer (CRC). Evidence reveals that vasculogenic mimicry (VM) is a critical event for the malignant transformation of cancer. Therefore, this study investigated the crucial roles of DCA in the regulation of VM and the progression of intestinal carcinogenesis. The effects of an HFD on VM formation and epithelial-mesenchymal transition (EMT) in human CRC tissues were investigated. The fecal DCA level was detected in HFD-treated Apcmin/+ mice. Then the effects of DCA on VM formation, EMT, and vascular endothelial growth factor receptor 2 (VEGFR2) signaling were evaluated in vitro and in vivo. Here we demonstrated that compared with a normal diet, an HFD exacerbated VM formation and EMT in CRC patients. An HFD could alter the composition of the gut microbiota and significantly increase the fecal DCA level in Apcmin/+ mice. More importantly, DCA promoted tumor cell proliferation, induced EMT, increased VM formation, and activated VEGFR2, which led to intestinal carcinogenesis. In addition, DCA enhanced the proliferation and migration of HCT-116 cells, and induced EMT process and vitro tube formation. Furthermore, the silence of VEGFR2 reduced DCA-induced EMT, VM formation, and migration. Collectively, our results indicated that microbial metabolite DCA promoted VM formation and EMT through VEGFR2 activation, which further exacerbated intestinal carcinogenesis.

Tumor-associated macrophages in osteosarcoma.

Osteosarcoma (OS) is the most common primary bone tumor in children and adolescents. It is an aggressive tumor with a tendency to spread to the lung, which is the most common site of metastasis. Patients with advanced OS with metastases have poor prognoses despite the application of chemotherapy, thus highlighting the need for novel therapeutic targets. The tumor microenvironment (TME) of OS is confirmed to be essential for and supportive of tumor growth and dissemination. The immune component of the OS microenvironment is mainly composed of tumor-associated macrophages (TAMs). In OS, TAMs promote tumor growth and angiogenesis and upregulate the cancer stem cell-like phenotype. However, TAMs inhibit the metastasis of OS. Therefore, much attention has been paid to investigating the mechanism of TAMs in OS development and the progression of immunotherapy for OS. In this article, we aim to summarize the roles of TAMs in OS and the major findings on the application of TAMs in OS treatment.

Secretory autophagy-induced bladder tumour-derived extracellular vesicle secretion promotes angiogenesis by activating the TPX2-mediated phosphorylation of the AURKA-PI3K-AKT axis.

Tumour angiogenesis is an independent risk factor for bladder cancer (BCa) progression, but viable and promising antiangiogenic targets are understudied. Secretory autophagy has received increasing interest recently, while the roles and executing mechanisms in the tumour microenvironment (TME) remain unclear. Herein, we found that active cathepsin B (CTSB) was upregulated in tumour tissues and serum EVs of 241 BCa patients from four cohorts and was significantly associated with poor prognosis. Starving TME (STME)-induced conventional autophagy in BCa cells elevated active CTSB levels by facilitating the expression and nuclear translocation of NFATC2. In addition, STME-induced secretory autophagy simultaneously led to markedly increased secretion of LC3-conjugated EVs loaded with active CTSB (EV-CTSB) into the TME. The increased exogenous active CTSB in endothelial cells by directly ingesting EV-CTSB prominently activated the TPX2-mediated phosphorylation of the AURKA-PI3K-AKT axis, increased VEGFA expression, and promoted angiogenesis. Our findings not only verify that EV-CTSB can be a promising target for antiangiogenic strategies in bladder cancer, but also reveal a novel action pattern based on secretory autophagy-induced EV secretion which is enlightening to explore crosstalk in the TME from various perspectives.

Vaccines targeting angiogenesis in melanoma.

Angiogenesis has a significant role in metastasis and progression of melanoma. Even small tumors may be susceptible to metastasis and hence lead to a worse outcome in patients with melanoma. One of the anti-angiogenic treatment approaches that is undergoing comprehensive study is specific immunotherapy. While tumor cells are challenging targets for immunotherapy due to their genetic instability and heterogeneity, endothelial cells (ECs) are genetically stable. Therefore, vaccines targeting angiogenesis in melanoma are appropriate choices that target both tumor cells and ECs while capable of inducing strong, anti-tumor immune responses with limited toxicity. The main targets of angiogenesis are VEGFs and their receptors but other potential targets have also been investigated, especially in preclinical studies. Various types of vaccines that target angiogenesis in melanoma have been studied including DNA, peptide, protein, dendritic cell-based, and endothelial cell vaccines. This review outlines a number of target antigens that are important for potential progress in developing vaccines for targeting angiogenesis in melanoma. We also discuss different types of vaccines that have been investigated, delivery mechanisms and popular adjuvants, and suggest ways to improve future clinical outcomes.

Role of Nectin­4 protein in cancer (Review).

The Nectin cell adhesion molecule (Nectin) family members are Ca2+­independent immunoglobulin­like cellular adhesion molecules (including Nectins 1­4), involved in cell adhesion via homophilic/heterophilic interplay. In addition, the Nectin family plays a significant role in enhancing cellular viability and movement ability. In contrast to enrichment of Nectins 1­3 in normal tissues, Nectin­4 is particularly overexpressed in a number of tumor types, including breast, lung, urothelial, colorectal, pancreatic and ovarian cancer. Moreover, the upregulation of Nectin­4 is an independent biomarker for overall survival in numerous cancer types. A large number of studies have revealed that high expression of Nectin­4 is closely related to tumor occurrence and development in various cancer types, but the manner in which Nectin­4 protein contributes to the onset and development of these malignancies is yet unknown. The present review summarizes the molecular mechanisms and functions of Nectin­4 protein in the biological processes and current advances with regard to its expression and regulation in various cancer types.

Hesperidin Inhibits UVB-Induced VEGF Production and Angiogenesis via the Inhibition of PI3K/Akt Pathway in HR-1 Hairless Mice.

Hesperidin is a citrus flavanone glycoside with potent anti-inflammatory effects that interferes with UVB-stimulated angiogenesis in skin, but its molecular mechanisms of action remain unclear. Here, we investigated the effects of hesperidin on UVB-induced angiogenesis in HR-1 hairless mice. We found hesperidin treatment inhibited skin neovascularization skin induced by repetitive UVB light exposure. Exposure to UVB radiation induces the expression of vascular endothelial growth factor (VEGF), matrix metalloproteinase-13 (MMP-13), and MMP-9, but we found all of these were inhibited by treatment with hesperidin. Using immunohistochemistry and Western blotting, we also found hesperidin inhibited the increase in hypoxia inducible factor-1 (HIF-1)α expression induced by UVB exposure. After discovering that UVB induces VEGF expression via the phosphoinositide 3-kinase (PI3K)/Akt signaling pathways, we found hesperidin reduces UVB-induced VEGF expression by inhibiting UVB-induced PI3K activity. This, in turn, reduces the UVB-induced Akt/p70S6K phosphorylation in human primary keratinocytes and fibroblast cells. Because it affects the mediators of angiogenesis, our data suggest hesperidin has an anti-angiogenic effect on the pathologic skin neovascularization induced by UVB light. Thus, hesperidin may prove useful in the treatment of skin injuries caused by UVB light exposure.

Resistance to anti-EGFR therapies in metastatic colorectal cancer: underlying mechanisms and reversal strategies.

Cetuximab and panitumumab are monoclonal antibodies (mAbs) against epidermal growth factor receptor (EGFR) that are effective agents for metastatic colorectal cancer (mCRC). Cetuximab can prolong survival by 8.2 months in RAS wild-type (WT) mCRC patients. Unfortunately, resistance to targeted therapy impairs clinical use and efficiency. The mechanisms of resistance refer to intrinsic and extrinsic alterations of tumours. Multiple therapeutic strategies have been investigated extensively to overcome resistance to anti-EGFR mAbs. The intrinsic mechanisms include EGFR ligand overexpression, EGFR alteration, RAS/RAF/PI3K gene mutations, ERBB2/MET/IGF-1R activation, metabolic remodelling, microsatellite instability and autophagy. For intrinsic mechanisms, therapies mainly cover the following: new EGFR-targeted inhibitors, a combination of multitargeted inhibitors, and metabolic regulators. In addition, new cytotoxic drugs and small molecule compounds increase the efficiency of cetuximab. Extrinsic alterations mainly disrupt the tumour microenvironment, specifically immune cells, cancer-associated fibroblasts (CAFs) and angiogenesis. The directions include the modification or activation of immune cells and suppression of CAFs and anti-VEGFR agents. In this review, we focus on the mechanisms of resistance to anti-EGFR monoclonal antibodies (anti-EGFR mAbs) and discuss diverse approaches to reverse resistance to this therapy in hopes of identifying more mCRC treatment possibilities.

Implication of N-Methyl-d-Aspartate Receptor in Homocysteine-Induced Age-Related Macular Degeneration.

Age-related macular degeneration (AMD) is a leading cause of vision loss. Elevated homocysteine (Hcy) (Hyperhomocysteinemia) (HHcy) has been reported in AMD. We previously reported that HHcy induces AMD-like features. This study suggests that N-Methyl-d-aspartate receptor (NMDAR) activation in the retinal pigment epithelium (RPE) is a mechanism for HHcy-induced AMD. Serum Hcy and cystathionine-ß-synthase (CBS) were assessed by ELISA. The involvement of NMDAR in Hcy-induced AMD features was evaluated (1) in vitro using ARPE-19 cells, primary RPE isolated from HHcy mice (CBS), and mouse choroidal endothelial cells (MCEC); (2) in vivo using wild-type mice and mice deficient in RPE NMDAR (NMDARR-/-) with/without Hcy injection. Isolectin-B4, Ki67, HIF-1α, VEGF, NMDAR1, and albumin were assessed by immunofluorescence (IF), Western blot (WB), Optical coherence tomography (OCT), and fluorescein angiography (FA) to evaluate retinal structure, fluorescein leakage, and choroidal neovascularization (CNV). A neovascular AMD patient's serum showed a significant increase in Hcy and a decrease in CBS. Hcy significantly increased HIF-1α, VEGF, and NMDAR in RPE cells, and Ki67 in MCEC. Hcy-injected WT mice showed disrupted retina and CNV. Knocking down RPE NMDAR improved retinal structure and CNV. Our findings underscore the role of RPE NMDAR in Hcy-induced AMD features; thus, NMDAR inhibition could serve as a promising therapeutic target for AMD.

A2E-induced inflammation and angiogenesis in RPE cells in vitro are modulated by PPAR-α, -ß/δ, -γ, and RXR antagonists and by norbixin.

N-retinylidene-N-retinylethanolamine (A2E) plays a central role in age-related macular degeneration (AMD) by inducing angiogenesis and inflammation. A2E effects are mediated at least partly via the retinoic acid receptor (RAR)-α. Here we show that A2E binds and transactivates also peroxisome proliferator-activated receptors (PPAR) and retinoid X receptors (RXR). 9'-cis-norbixin, a di-apocarotenoid is also a ligand of these nuclear receptors (NR). Norbixin inhibits PPAR and RXR transactivation induced by A2E. Moreover, norbixin reduces protein kinase B (AKT) phosphorylation, NF-κB and AP-1 transactivation and mRNA expression of the inflammatory interleukins (IL) -6 and -8 and of vascular endothelial growth factor (VEGF) enhanced by A2E. By contrast, norbixin increases matrix metalloproteinase 9 (MMP9) and C-C motif chemokine ligand 2 (CCL2) mRNA expression in response to A2E. Selective PPAR-α, -ß/δ and -γ antagonists inhibit the expression of IL-6 and IL-8 while only the antagonist of PPAR-γ inhibits the transactivation of NF-κB following A2E exposure. In addition, a cocktail of all three PPARs antagonists and also HX531, an antagonist of RXR reproduce norbixin effects on inflammation. Altogether, A2E's deleterious biological effects could be inhibited through PPAR and RXR regulation. Moreover, the modulation of these NR by norbixin may open new avenues for the treatment of AMD.

The Sodium-Glucose Cotransporter-2 Inhibitor Canagliflozin Alleviates Endothelial Dysfunction Following In Vitro Vascular Ischemia/Reperfusion Injury in Rats.

Vascular ischemia/reperfusion injury (IRI) contributes to graft failure and adverse clinical outcomes following coronary artery bypass grafting. Sodium-glucose-cotransporter (SGLT)-2-inhibitors have been shown to protect against myocardial IRI, irrespective of diabetes. We hypothesized that adding canagliflozin (CANA) (an SGLT-2-inhibitor) to saline protects vascular grafts from IRI. Aortic rings from non-diabetic rats were isolated and immediately mounted in organ bath chambers (control, n = 9-10 rats) or underwent cold ischemic preservation in saline, supplemented either with a DMSO vehicle (IR, n = 8-10 rats) or 50µM CANA (IR + CANA, n = 9-11 rats). Vascular function was measured, the expression of 88 genes using PCR-array was analyzed, and feature selection using machine learning was applied. Impaired maximal vasorelaxation to acetylcholine in the IR-group compared to controls was significantly ameliorated by CANA (IR 31.7 ± 3.2% vs. IR + CANA 51.9 ± 2.5%, p < 0.05). IR altered the expression of 17 genes. Ccl2, Ccl3, Ccl4, CxCr4, Fos, Icam1, Il10, Il1a and Il1b have been found to have the highest interaction. Compared to controls, IR significantly upregulated the mRNA expressions of Il1a and Il6, which were reduced by 1.5- and 1.75-fold with CANA, respectively. CANA significantly prevented the upregulation of Cd40, downregulated NoxO1 gene expression, decreased ICAM-1 and nitrotyrosine, and increased PECAM-1 immunoreactivity. CANA alleviates endothelial dysfunction following IRI.