Inhibition Of Vascular Endothelial Growth Factor Expression Research Articles

Regulation role of miR-204 on SIRT1/VEGF in metabolic memory induced by high glucose in human retinal pigment epithelial cells

AIM: To examine the regulatory role of microRNA-204 (miR-204) on silent information regulator 1 (SIRT1) and vascular endothelial growth factor (VEGF) under high-glucose-induced metabolic memory in human retinal pigment epithelial (hRPE) cells. METHODS: Cells were cultured with either normal (5 mmol/L) or high D-glucose (25 mmol/L) concentrations for 8d to establish control and high-glucose groups, respectively. To induce metabolic memory, cells were cultured with 25 mmol/L D-glucose for 4d followed by culture with 5 mmol/L D-glucose for 4d. In addition, exposed in 25 mmol/L D-glucose for 4d and then transfected with 100 nmol/L miR-204 control, miR-204 inhibitor or miR-204 mimic in 5 mmol/L D-glucose for 4d. Quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect miR-204 mRNA levels. SIRT1 and VEGF protein levels were assessed by immunohistochemical and Western blot. Flow cytometry was used to investigate apoptosis rate. RESULTS: It was found that high glucose promoted miR-204 and VEGF expression, and inhibited SIRT1 activity, even after the return to normal glucose culture conditions. Upregulation of miR-204 promoted apoptosis inhibiting SIRT1 and increasing VEGF expression. However, downregulation of miR-204 produced the opposite effects. CONCLUSION: The study identifies that miR-204 is the upstream target of SIRT1and VEGF, and that miR-204 can protect hRPE cells from the damage caused by metabolic memory through increasing SIRT1 and inhibiting VEGF expression.

Tumor-targeted and stimulus-responsive polymeric prodrug nanoparticles to enhance the anticancer therapeutic efficacy of doxorubicin

Polymeric prodrug nanoparticles have gained increasing attention in the field of anticancer drug delivery because of their dual functions as a drug carrier and a therapeutic agent. Doxorubicin (DOX) is a highly effective chemotherapeutic agent for various cancers but causes cardiotoxicity. In this work, we developed polymeric prodrug (pHU) nanoparticles that serve as both a drug carrier of DOX and a therapeutic agent. The composition of pHU includes antiangiogenic hydroxybenzyl alcohol (HBA) and ursodeoxycholic acid (UDCA), covalently incorporated through hydrogen peroxide (H2O2)-responsive peroxalate. To enhance cancer cell specificity, pHU nanoparticles were surface decorated with taurodeoxycholic acid (TUDCA) to facilitate p-selectin-mediated cancer targeting. TUDCA-coated and DOX-loaded pHU nanoparticles (t-pHUDs) exhibited controlled release of DOX triggered by H2O2, characteristic of the tumor microenvironment. t-pHUDs also effectively suppressed cancer cell migration and vascular endothelial growth factor (VEGF) expression in response to H2O2. In animal studies, t-pHUDs exhibited highly potent anticancer activity. Notably, t-pHUDs, with their ability to accumulate preferentially in tumors due to the p-selectin targeting, surpassed the therapeutic efficacy of equivalent DOX and pHU nanoparticles alone. What is more, t-pHUDs significantly suppressed VEGF expression in tumors and mitigated hepato- and cardiotoxicity of DOX. Given their cancer targeting ability, enhanced therapeutic efficacy and minimized off-target toxicity, t-pHUDs present an innovative and targeted approach with great translational potential as an anticancer therapeutic agent.

The Treatment of Diabetic Retinal Edema with Intravitreal Steroids: How and When.

Diabetic macular edema (DME) is a common complication of diabetes mellitus and a leading cause of visual impairment worldwide. It is defined as the diabetes-related accumulation of fluid, proteins, and lipids, with retinal thickening, within the macular area. DME affects a significant proportion of individuals with diabetes, with the prevalence increasing with disease duration and severity. It is estimated that approximately 25-30% of diabetic patients will develop DME during their lifetime. Poor glycemic control, hypertension, hyperlipidemia, diabetes duration, and genetic predisposition are recognized as risk factors for the development and progression of DME. Although the exact pathophysiology is still not completely understood, it has been demonstrated that chronic hyperglycemia triggers a cascade of biochemical processes, including increased oxidative stress, inflammation, activation of vascular endothelial growth factor (VEGF), cellular dysfunction, and apoptosis, with breakdown of the blood-retinal barriers and fluid accumulation within the macular area. Early diagnosis and appropriate management of DME are crucial for improving visual outcomes. Although the control of systemic risk factors still remains the most important strategy in DME treatment, intravitreal pharmacotherapy with anti-VEGF molecules or steroids is currently considered the first-line approach in DME patients, whereas macular laser photocoagulation and pars plana vitrectomy may be useful in selected cases. Available intravitreal steroids, including triamcinolone acetonide injections and dexamethasone and fluocinolone acetonide implants, exert their therapeutic effect by reducing inflammation, inhibiting VEGF expression, stabilizing the blood-retinal barrier and thus reducing vascular permeability. They have been demonstrated to be effective in reducing macular edema and improving visual outcomes in DME patients but are associated with a high risk of intraocular pressure elevation and cataract development, so their use requires an accurate patient selection. This manuscript aims to provide a comprehensive overview of the pathology, epidemiology, risk factors, physiopathology, clinical features, treatment mechanisms of actions, treatment options, prognosis, and ongoing clinical studies related to the treatment of DME, with particular consideration of intravitreal steroids therapy.

Platelet Rich Fibrin as a Potential Treatment for Corneal Neovascularization due to Infective Keratitis: An Experimental Study

Abstract
 Introduction & Objectives : Platelet-rich fibrin (PRF) as a source of growth factor has been widely used in various therapeutic fields so that it could accelerate the wound healing process. This study investigated the effect of platelet-rich fibrin PRF in inhibiting neovascularization, inhibiting the expression of vascular endothelial growth factor (VEGF) and stimulating pigment epithelial-derived factor (PEDF) in the infective keratitis animal model.
 Methods : A total of 12 New Zealand white rabbits (Oryctalogus cuniculus) were subjected to keratitis induction by inoculation of Pseudomonas aeruginosa. The rabbits were divided into two groups, the first group as control was treated antibiotic-only, and the second as treatment group was treated with antibiotic and PRF graft. The corneal neovascularization area was clinically measured before and after treatment. Expression of VEGF and PEDF in the corneal epithelium, stroma, and endothelium were evaluated by immunohistochemical examination on day 10 after keratitis induction.
 Results : There were significant differences in VEGF expression between groups in the epithelial layer (p=0.008), but no significant difference in the stromal (p=0.934) and endothelial (p=0.935) layers. PEDF expression showed a significant difference between groups in the epithelial (p = 0.004) and stroma layers (p = 0.029) but no significant difference in the endothelial layer (p = 0.157). However, there was no significant difference in clinical neovascularization between the two groups (p=0.526).
 Conclusion : PRF could inhibit VEGF expression and stimulate PEDF expression in the cornea of Pseudomonas aeruginosa keratitis animal model that potentially reduces the corneal neovascularization in the longer observation.

Osteopontin Promotes Angiogenesis in the Spinal Cord and Exerts a Protective Role Against Motor Function Impairment and Neuropathic Pain After Spinal Cord Injury.

Basic science study using a hemisection spinal cord injury (SCI) model. We sought to assess the effect of blocking osteopontin (OPN) upregulation on motor function recovery and pain behavior after SCI and to further investigate the possible downstream target of OPN in the injured spinal cord. OPN is a noncollagenous extracellular matrix protein widely expressed across different tissues. Its expression substantially increases following SCI. A previous study suggested that this protein might contribute to locomotor function recovery after SCI. However, its neuroprotective potential was not fully explored, nor were the underlying mechanisms. We constructed a SCI mouse model and analyzed the expression of OPN at different time points and the particular cell distribution in the injured spinal cord. Then, we blocked OPN upregulation with lentivirus-delivering siRNA targeting OPN specifically and examined its effect on motor function impairment and neuropathic pain after SCI. The underlying mechanisms were explored in the OPN-knockdown mice model and cultured vascular endothelial cells. The proteome study revealed that OPN was the most dramatically increased protein following SCI. OPN in the spinal cord was significantly increased three weeks after SCI. Suppressing OPN upregulation through siRNA exacerbated motor function impairment and neuropathic pain. In addition, SCI resulted in an increase in vascular endothelial growth factor (VEGF), AKT phosphorylation, and angiogenesis within the spinal cord, all of which were curbed by OPN reduction. Similarly, OPN knockdown suppressed VEGF expression, AKT phosphorylation, cell migration, invasion, and angiogenesis in cultured vascular endothelial cells. OPN demonstrates a protective influence against motor function impairment and neuropathic pain following SCI. This phenomenon may result from the proangiogenetic effect of OPN, possibly due to activation of the VEGF and/or AKT pathways.

Enhanced periodontal tissue healing via vascular endothelial growth factor expression following low-level erbium-doped: yttrium, aluminum, and garnet laser irradiation: In vitro and in vivo studies.

This study aimed to investigate the effects of low-level erbium-doped: yttrium, aluminum, and garnet (Er:YAG) laser irradiation on periodontal tissue healing and regeneration through angiogenesis in vivo and in vitro studies. Intrabony defects were surgically created in the bilateral maxilla molar of rats. The defects were treated by open flap debridement (OFD)with Er:YAG laser, including low-level laser irradiation (LLLI) to bone and blood clot surfaces, or conventional procedures. The mRNA expression of vascular endothelial growth factor (VEGF) in the surgical sites was quantified using real-time polymerase chain reaction. The decalcified specimens were prepared for histometric analysis. Also, LLLI was performed on human umbilical vein endothelial cells to evaluate the effects on angiogenesis. Cell proliferation, VEGF expression, and tube formation were assessed. In addition, capsazepine (CPZ), a selective inhibitor of transient receptor potential vanilloid 1 (TRPV1), treatment was performed before LLLI for the same assays. OFD using Er:YAG laser did not generate thermal damage on bone or root surfaces. LLLI accelerated hemostasis by coagulation of the superficial layers of blood clots in the laser-treated group. Postoperative healing was sound in all animals in both groups. VEGF expression and bone formation were significantly increased in the laser-treated group compared to those in the conventional treatment group. In vitro, cell proliferation and VEGF expression were significantly increased in the LLLI group compared to the control group. Tube-formation assays showed that LLLI significantly promoted angiogenesis. CPZ treatment significantly suppressed VEGF expression and tube formation following LLLI. This study suggests that Er:YAG laser irradiation may promote periodontal tissue healing by enhancing angiogenetic effect of endothelial cells via TRPV1.

Antiangiogenic Action of JZL184 on Endothelial Cells via Inhibition of VEGF Expression in Hypoxic Lung Cancer Cells.

JZL184, an inhibitor of monoacylglycerol lipase (MAGL) and thus of the degradation of the endocannabinoid 2-arachidonoylglycerol (2-AG), mediates various anticancer effects in preclinical studies. However, studies on the effect of this or other MAGL inhibitors under hypoxia, an important factor in tumor biology and response to cancer therapy, have not yet been performed in cancer cells. In the present study, the impact of the conditioned media (CM) of A549 and H358 lung cancer cells incubated with JZL184 under hypoxic conditions on the angiogenic properties of human umbilical vein endothelial cells (HUVECs) was investigated. Treatment of HUVECs with CM derived from cancer cells cultured for 48 h under hypoxic conditions was associated with a substantial increase in migration and tube formation compared with unconditioned medium, which was inhibited when cancer cells were incubated with JZL184. In this process, JZL184 led to a significant increase in 2-AG levels in both cell lines. Analysis of a panel of proangiogenic factors revealed inhibition of hypoxia-induced vascular endothelial growth factor (VEGF) expression by JZL184. Antiangiogenic and VEGF-lowering effects were also demonstrated for the MAGL inhibitor MJN110. Receptor antagonist experiments suggest partial involvement of the cannabinoid receptors CB1 and CB2 in the antiangiogenic and VEGF-lowering effects induced by JZL184. The functional importance of VEGF for angiogenesis in the selected system is supported by observations showing inhibition of VEGF receptor 2 (VEGFR2) phosphorylation in HUVECs by CM from hypoxic cancer cells treated with JZL184 or when hypoxic cancer cell-derived CM was spiked with a neutralizing VEGF antibody. On the other hand, JZL184 did not exert a direct effect on VEGFR2 activation induced by recombinant VEGF, so there seems to be no downstream effect on already released VEGF. In conclusion, these results reveal a novel mechanism of antiangiogenic action of JZL184 under conditions of hypoxic tumor-endothelial communication.

Effect of adalimumab on experimental corneal neovascularization model

To evaluate the efficacy of adalimumab (ADA) on inhibition of experimental corneal neovascularization (CNV) and compare the outcomes with bevacizumab (BEVA). Twenty-four female Winstar rats (48 eyes) were used. Silver/Potassium Nitrate sticks were used for creating CNV. Forty-eight eyes of the rats were separated into 6 groups. The eyes which only NaCl was injected subconjunctivally (SC) formed Group-1. The eyes which CNV was created and NaCl, BEVA (2.5mg/0.05mL), ADA (2.5mg/0.05mL), respectively, were injected SC formed group-2, 3 and 4. The eyes which only BEVA and ADA, respectively, were injected SC formed group-5 and 6. Five days later the animals were sacrificed. Hematoxylin and eosin staining, Masson trichrome staining, Vascular endothelial growth factor (VEGF), and Platelet-derived growth factor (PDGF) antibodies were performed. Histochemical results showed that there was no histopathological finding in group-1, 5, and 6. Collagen fiber irregularity was observed in group-2 and there was a significant improvement in collagen fiber irregularity in group-3 and 4. Collagen fiber proliferation was higher in group-2 than in group-3 and 4. VEGF and PDGF stainings were not observed in group-1, 5, and 6. VEGF and PDGF stainings were observed in group-2 and significantly decreased in group-3 and 4 compared to group-2. ADA was found to be superior to BEVA in terms of decreasing VEGF staining. Both BEVA and ADA were effective in inhibiting CNV. Subconjunctival ADA seems to be more effective than BEVA in terms of inhibiting VEGF expression. Further experimental studies about ADA and BEVA are needed.

Endostatin and Cancer Therapy: A Novel Potential Alternative to Anti-VEGF Monoclonal Antibodies

Angiogenesis is a physiological process that consists of the formation of new blood vessels from preexisting ones. Angiogenesis helps in growth, development, and wound healing through the formation of granulation tissue. However, this physiological process has also been linked to tumor growth and metastasis formation. Indeed, angiogenesis has to be considered as a fundamental step to the evolution of benign tumors into malignant neoplasms. The main mediator of angiogenesis is vascular endothelial growth factor (VEGF), which is overexpressed in certain cancers. Thus, there are anti-VEGF monoclonal antibodies, such as bevacizumab, used as anti-cancer therapies. However, bevacizumab has shown adverse events, such as hypertension and proteinuria, which in the most severe cases can lead to cessation of therapy, thus contributing to worsening patients' prognosis. On the other hand, endostatin is an endogenous protein that strongly inhibits VEGF expression and angiogenesis and shows a better safety profile. Moreover, endostatin has already given promising results on small scale clinical studies. Hence, in this review, we present data supporting the use of endostatin as a replacement for anti-VEGF monoclonal antibodies.

Combination of pimitespib (TAS-116) with sunitinib is an effective therapy for imatinib-resistant gastrointestinal stromal tumors.

Despite the effectiveness of imatinib, most gastrointestinal stromal tumors (GISTs) develop resistance to the treatment, mainly due to the reactivation of KIT tyrosine kinase activity. Sunitinib, which inhibits the phosphorylation of KIT and vascular endothelial growth factor (VEGF) receptor, has been established as second-line therapy for GISTs. The recently-developed heat shock protein 90 (HSP90) inhibitor pimitespib (PIM; TAS-116) demonstrated clinical benefits in some clinical trials; however, the effects were limited. The aim of our study was therefore to clarify the effectiveness and mechanism of the combination of PIM with sunitinib for imatinib-resistant GISTs. We evaluated the efficacy and mechanism of the combination of PIM with sunitinib against imatinib-resistant GIST using imatinib-resistant GIST cell lines and murine xenograft models. In vitro analysis demonstrated that PIM and sunitinib combination therapy strongly inhibited growth and induced apoptosis in imatinib-resistant GIST cell lines by inhibiting KIT signaling and decreasing auto-phosphorylated KIT in the Golgi apparatus. In addition, PIM and sunitinib combination therapy enhanced antitumor responses in the murine xenograft models compared to individual therapies. Further analysis of the xenograft models showed that the combination therapy not only downregulated the KIT signaling pathway but also decreased the tumor microvessel density. Furthermore, we found that PIM suppressed VEGF expression in GIST cells by suppressing protein kinase D2 and hypoxia-inducible factor-1 alpha, which are both HSP90 client proteins. In conclusion, the combination of PIM and sunitinib is effective against imatinib-resistant GIST via the downregulation of KIT signaling and angiogenic signaling pathways.

Treatment of Ovarian Hyperstimulation Syndrome in a Mouse Model by Cannabidiol, an Angiogenesis Pathway Inhibitor.

Studies suggest that ovarian hyperstimulation syndrome (OHSS) can be treated by reducing the level of vascular endothelial growth factor (VEGF). However, due to the side effects of commercially available VEGF-reducing drugs, they can be ruled out as a suitable treatment for OHSS; therefore, researchers are looking for new medications to treat OHSS. This study is aimed at investigating the effects of cannabidiol (CBD) in an OHSS model and to evaluate its efficacy in modulating the angiogenesis pathway and VEGF gene expression. For this purpose, 32 female mice were randomly divided into four groups (eight mice per group): control group, group 2 with OHSS induction, group 3 receiving 32 nmol of dimethyl sulfoxide after OHSS induction, and group 4 receiving 30 mg/kg of CBD after OHSS induction. The animals' body weight, ovarian weight, vascular permeability (VP), and ovarian follicle count were measured, and the levels of VEGF gene and protein expression in the peritoneal fluid were assessed. Based on the results, CBD decreased the body and ovarian weights, VP, and corpus luteum number compared to the OHSS group (p < 0.05). The peritoneal VEGF gene and protein expression levels reduced in the CBD group compared to the OHSS group (p < 0.05). Also, CBD caused OHSS alleviation by suppressing VEGF expression and VP. Overall, CBD downregulated VEGF gene expression and improved VP in OHSS.

Circular Sponge against miR-21 Enhances the Antitumor Activity of Doxorubicin against Breast Cancer Cells

Breast cancer is the most common type of cancer in women, with chemotherapy being the main strategy. However, its effectiveness is reduced by drug resistance mechanisms. miR-21 is upregulated in breast cancer that has been linked to drug resistance and carcinogenic processes. Our aim was to capture miR-21 with a circular sponge (Circ-21) and thus inhibit the carcinogenic processes and drug resistance mechanisms in which it participates. Proliferation, migration, colony formation, cell cycle, and poly [ADP-ribose] polymerase 1 (PARP-1) and vascular endothelial growth factor (VEGF) detection assays were performed with MCF7 breast cancer cells and MCF10A non-tumor cells. In addition, doxorubicin resistance tests and detection of drug resistance gene expression were performed in MCF7 cells. Reduction in proliferation, as well as migration and colony formation, increased PARP-1 expression, inhibition of VEGF expression and cell cycle arrest in G2/M phase were displayed in the Circ-21 MCF7, which were not observed in the MCF10A cells. Furthermore, in the MCF7 cells, the Circ-21 enhanced the antitumor activity of doxorubicin and decreased the expression of resistance genes: ABCA1, ABCC4, and ABCC5. Based on these results, the use of Circ-21 can be considered a first step for the establishment of an effective gene therapy in the treatment of breast cancer.

Treatment outcomes of injectable thermosensitive hydrogel containing bevacizumab in intervertebral disc degeneration.

Intervertebral disc (IVD) degeneration (IDD) is a common musculoskeletal disease and its treatment remains a clinical challenge. It is characterised by reduced cell numbers and degeneration of the extracellular matrix (ECM). Nucleus pulposus (NP) cells play a crucial role in this process. The purpose of this study is to explore the role of bevacizumab, a vascular endothelial growth factor (VEGF) inhibitor, in the treatment of IDD through local drug delivery. High expression of VEGF was observed in degenerating human and rat IVDs. We demonstrated that MMP3 expression was decreased and COL II synthesis was promoted, when VEGF expression was inhibited by bevacizumab, thereby improving the degree of disc degeneration. Thus, these findings provide strong evidence that inhibition of VEGF expression by local delivery of bevacizumab is safe and effective in ameliorating disc degeneration in rats. The injectable thermosensitive PLGA-PEG-PLGA hydrogels loaded with bevacizumab is a potential therapeutic option for disc degeneration.

Notch1 signaling in keratocytes maintains corneal transparency by suppressing VEGF expression.

SummaryThe cornea fends off chemicals, dirt, and infectious particles and provides most of the eye’s focusing power. Corneal transparency is of paramount importance to normal vision, yet how it is established and maintained remains unclear. Here, we ablated Notch1 in keratocytes using Twist2-Cre mice and found that Twist2-Cre; Notch1f/f mice developed stroma expansion and neovascularization, followed by hyperproliferation and metaplasia of corneal epithelial progenitor cells and plaque formation at central cornea, leading to loss of transparency. Development of these phenotypes does not involve bacteria-caused inflammation; instead, Notch1 deletion upregulates Vegfa and Vegfc via Hif1α in keratocytes. Vascular endothelial growth factor (VEGF) receptor inhibitor axitinib prevented development of these anomalies in Twist2-Cre; Notch1f/f mice, suggesting that VEGFs secreted by keratocytes promote not only neovascularization but also proliferation and metaplasia of epithelial progenitor cells at central cornea. This study uncovers a Notch1-Hif1α-VEGF pathway in keratocytes that maintains corneal transparency and represents a potential target for treatment of related corneal disorders.

MiR-146a suppresses the expression of vascular endothelial growth factor and inflammatory responses in diabetic retinopathy

This study was designed to explore the role of miR-146a in diabetic retinopathy (DR). 30 healthy control (HC), 50 patients with type 2 diabetes mellitus, and 48 DR patients were enrolled. Blood was collected and levels of miR-146a expression, vascular endothelial growth factor (VEGF), and three inflammatory cytokines (NF-κB, IL-1β, and TNF-α) were detected. Moreover, ARPE-19 cells were treated with miR-146a mimic or inhibitor in the presence of high glucose to evaluate its effect in vitro. DR patients had the lowest level of miR-146a and the highest level of VEGF as well as the most severe inflammation among the three groups. In addition, the miR-146a level was negatively correlated with the expression of VEGF and three inflammatory cytokines, respectively in DR patients. Moreover, VEGF expression was positively correlated with these three inflammatory cytokines in DR patients. In summary, miR-146a could inhibit VEGF expression and inflammation in DR.

Endostatin Inhibits Blood-Retinal Barrier Breakdown in Diabetic Rats by Increasing the Expression of ICAM-1 and VCAM-1 and Decreasing the Expression of VEGF.

Endostatin has become the strongest endogenous angiogenesis inhibitor due to suppressing VEGF expression. The purpose of this study was to assess the impact of endostatin on the blood-retinal barrier (BRB) in diabetic rats. SD rats were induced to develop diabetes by streptozotocin, and endostatin was administrated by intravitreal injection. The body weight, the level of blood glucose, the expressions of C-reactive protein (CRP), adhesion molecules intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), junction proteins (occludin, claudin-5, and zonula occluden-1), and VEGF were measured in rats' retinas of diabetes. The BRB breakdown was evaluated using Evans blue. The level of CRP and adhesion molecules (ICAM-1 and VCAM-1) was increased in retinas of diabetic rats, while endostatin significantly inhibited the upregulation of these. Diabetes increased the BRB permeability and retinal thickness. Diabetes also decreased the levels of occludin, claudin-5, and ZO-1 in retinals. These changes were inhibited by endostatin treatment. Upregulation of vascular endothelial growth factor (VEGF), transforming growth factor-β (TGF-β), and protein kinase C- (PKC-) β2 was also reversed by endostatin in retinas of diabetic rats. Endostatin provides protection against diabetic retinopathy, which may involve its barrier-enhancing effects.

Antcin K inhibits VEGF-dependent angiogenesis in human rheumatoid arthritis synovial fibroblasts.

Antrodia cinnamomea is a well-known medicinal mushroom in Taiwan that exhibits anti-inflammatory biological activities. In rheumatoid arthritis (RA), chronic inflammation and angiogenesis driven by proinflammatory cytokines reflect the severity of the disease. Although biological treatments have improved the outlook for RA, no healing exists. Moreover, the available pharmacotherapies do not work for all patients and drug safety is a major consideration. Investigations into plant-based medicines hope to reveal better, more tolerable agents. We examined whether Antcin K, a phytosterol isolated from A. cinnamomea, has anti-angiogenic activity in RA. The GSE12021 gene dataset from the Gene Expression Omnibus (GEO) database was examined for levels of vascular endothelial growth factor (VEGF) expression in 10 RA and 10 osteoarthritis (OA) synovial tissue samples. In clinical samples, VEGF expression was analyzed by immunohistochemical staining and ELISA in normal and RA synovial tissue, as well as OA and RA synovial fluid. Collagen-induced arthritis (CIA) and control tissue was stained with hematoxylin and eosin (H&E) for histological changes; Safranin O/Fast Green staining examined histopathological changes and evidence of bone erosion. Human RA synovial fibroblasts (RASFs) were incubated with Antcin K and cell viability was examined by the MTT assay. VEGF mRNA expression was detected in RASFs using qPCR. Antcin K significantly inhibited VEGF expression and ameliorates endothelial progenitor cell (EPC) migration and tube formation in RASFs by downregulating the phospholipase C-γ/protein kinase C-α pathway. Antcin K also induced anti-angiogenic effects in human RASFs without cytotoxicity. PRACTICAL APPLICATIONS: Analysis of GEO dataset samples and human synovial fluids or synovial tissues revealed higher VEGF levels in rheumatoid arthritis (RA) samples compared with osteoarthritis (OA) and healthy control samples. VEGF levels were also higher in mice with collagen-induced arthritis (CIA) than in healthy controls. Antcin K markedly suppressed VEGF expression in human RA synovial fibroblasts and inhibited the migration and tube formation of epithelial progenitor cells (EPCs) by downregulating the phospholipase C-γ/protein kinase C-α pathway. Further investigations are warranted to examine the effects of Antcin K in other angiogenesis-associated disorders.

Se-Methylselenocysteine Alleviates Liver Injury in Diethylnitrosamine (DEN)-Induced Hepatocellular Carcinoma Rat Model by Reducing Liver Enzymes, Inhibiting Angiogenesis, and Suppressing Nitric Oxide (NO)/Nitric Oxide Synthase (NOS) Signaling Pathway.

BackgroundHepatocellular carcinoma is the third leading cause of cancer-associated mortality. This study aimed to investigate the effects of se-methylselenocysteine (MSC) on oncogenesis of diethylnitrosamine (DEN)-induced hepatocellular carcinoma.Material/MethodsA hepatocellular carcinoma rat model was established by administering DEN. Rat models were divided into Model (0.1 mg/kg MSC), Model+0.3 mg/kg MSC, Model+1 mg/kg MSC, and Model+3 mg/kg MSC groups. A Normal control group consisted of mice not administered MSC. Hematoxylin and eosin staining was used to determine liver injury. Immunohistochemical analysis was conducted to identify CD34 and vascular endothelial growth factor (VEGF) expression. VEGF gene transcription was detected with RT-PCR. Biochemical analyses were performed to determine alanine aminotransferase, aspartate aminotransferase, total bilirubin, γ-glutamyl transpeptidase, alkaline phosphatase, and albumin levels in serum, and nitric oxide (NO)/nitric oxide synthase (NOS) levels in liver tissues. Transmission electron microscopy was used to observe the ultra-microstructures of hepatocytes.ResultsMSC treatment markedly alleviated liver injury and nuclear lesions in the treatment groups compared to the Model group. MSC treatment significantly improved liver functions in the treatment groups compared to the Model group (P<0.05). MSC treatment significantly decreased CD34 expression and NO and NOS levels in liver tissues and suppressed VEGF expression compared to the Model group (all P<0.05).ConclusionsMSC administration alleviated liver injury in a DEN-induced hepatocellular carcinoma rat model through reducing liver enzymes, inhibiting angiogenesis, and suppressing the NO/NOS signaling pathway.

Expression of hypoxia-inducible factor-1 in mice infected with Toxoplasma gondii during pregnancy

To investigate the expression and possible role of hypoxia-inducible factor-1 (HIF-1) at the maternal-fetal interface following Toxoplasma gondii infection during early pregnancy. Twenty pregnant C57BL/6 mice, each weighing 16 to 20 g, were randomly divided into 4 groups, including the 12-d control group, 12-d infection group, 18-d control group and 18-d infection group. Mice in the 12-d and 18-d infection groups were injected intraperitoneally with 150 tachyzoites of the T. gondii PRU strain on day 6 of pregnancy, while mice in the 12-d control and 18-d control groups were injected with the same volume of phosphate buffered saline (PBS). Mice in the control and infection groups were sacrificed on days 12 and 18 of pregnancy, and the placental and uterine specimens of the pregnant mice in each group were sampled for pathological examinations. The mRNA expression of HIF-1α, HIF-1β and vascular endothelial growth factor (VEGF) was quantified using quantitative fluorescent real-time PCR (qPCR) assay in the placental and uterine specimens, and the correlation between HIF-1α and VEGF mRNA expression was examined. In addition, and the HIF-1α expression was detected using immunohistochemical staining in the placental and uterine specimens of pregnant mice. Compared with the 12-d and 18-d control groups, adverse pregnant outcomes were observed in mice in 12-d and 18-d infection groups, such as teratism and placental dysplasia. HE staining showed swelling and blood stasis of cells, sinusoid reduction and inflammatory cell infiltration in the labyrinth area of the placenta specimens of mice in 12-d and 18-d infection groups relative to 12-d and 18-d control groups, and columnar epithelial cell injury and inflammatory cell infiltration were seen in the mouse uterine specimens in both infection groups. qPCR assay detected significantly higher HIF-1α (F = 132.6, P < 0.05) and HIF-1β mRNA expression (F = 286.9, P < 0.05) in the placental specimens and lower HIF-1α (F = 111.5, P < 0.05) and HIF-1β mRNA expression (F = 55.2, P < 0.05) in the uterine specimens in the 12-d infection group than in the 12-day control group, and significantly lower HIF-1α and HIF-1β mRNA expression was detected in the placental and uterine specimens in the 18-d infection group than in the 18-day control group (F = 215.8, 418.9, 156.8 and 200.1; all P values < 0.05). Significantly lower VEGF-A (F = 426.2, P < 0.05), VEGF-B (F = 104.6, P < 0.05) and VEGF-C mRNA expression (F = 566.9, P < 0.05) in the placental specimens and higher VEGF-A (F = 426.2, P < 0.05), VEGF-B (F = 104.6, P < 0.05) and VEGF-C mRNA expression (F = 566.9, P < 0.05) in the uterine specimens were detected in the 12-d infection group than in the 12-d control group, and higher VEGF-A, VEGF-B and VEGF-C mRNA expression was found in the placental and uterine specimens in the 18-d infection group than in the 18-d control group (F = 521.9, 100.6, 275.9, 224.6, 108.2 and 333.4; all P values < 0.05). Immunohistochemical staining showed strongly and mildly positive HIF-1α expression in the mouse placental labyrinth area in the 12-d and 18-d infection groups relative to 12-d and 18-d control groups, while no HIF-1α expression was detected in mouse uterine specimens. HIF-1α expression appears a tendency towards a rise in the second trimester and a reduction in the third trimester in mice following T. gondii infection during early pregnancy, which is contrary to the changing tendency of VEGF-A, VEGF-B, and VEGF-C expression. It is hypothesized that HIF-1α inhibits placental angiogenesis in mice during pregnancy through suppressing VEGF expression, resulting in adverse pregnant outcomes.

Serum exosomal miR-377-3p inhibits retinal pigment epithelium proliferation and offers a biomarker for diabetic macular edema.

ObjectivesDiabetic macular edema (DME) is a complication of diabetes mellitus that leads to diabetic retinopathy. Thus far, the role of serum exosomal microRNAs (miRNAs) in DME progression remains elusive. This study investigated serum exosomal miRNAs from patients with type 2 diabetes (T2D) and DME to identify miRNAs associated with expression of vascular endothelial growth factor (VEGF), a pivotal component in DME progression; it also evaluated the diagnostic values of these miRNAs for DME.MethodsSerum was collected from patients with T2D who did (n = 20) and did not have DME (n = 24). Exosomes were isolated from serum and subjected to real-time polymerase chain reaction, western blotting, luciferase reporter, and miRNA profiling analyses.ResultsVEGF was significantly upregulated in ARPE-19 cells treated with exosomes from patients with T2D and DME, compared with exosomes from patients with T2D alone. Among the top 10 downregulated miRNAs identified during exosomal miRNA profiling, miR-377-3p inhibited the expression of VEGF. Luciferase reporter assays confirmed that miR-377-3p could directly regulate VEGF expression. Receiver operating characteristic analysis identified serum exosomal miR-377-3p as a potential biomarker for DME.ConclusionSerum exosomal miR-377-3p inhibits VEGF expression to suppress retinal pigment epithelium proliferation and offers a diagnostic biomarker for DME.