Endothelial Growth Factor mRNA Expression Research Articles

Effect of EMB-FUBINACA on brain endothelial cell angiogenesis: Expression analysis of angiogenic markers.

Angiogenesis is the process by which blood vessels are generated from preexisting ones. Synthetic cannabinoids represent new psychoactive substances that bind to the cannabinoid receptor 1 (CB1R) and cannabinoid receptor 2 (CB2R) and simulate similar effects of tetrahydrocannabinol, the primary component found in cannabis. In the present study, we used the synthetic cannabinoid EMB-FUBINACA to study its impact on brain angiogenesis. Human brain microvascular endothelial cells (HBMECs) were cultivated in DMEM media before being subjected to different concentrations of EMB-FUBINACA and the control. Cell viability and the migration rates of HBMECs were evaluated using the viability and wound healing assays, respectively. An in vitro Matrigel Tube Formation Assay was carried out to measure the angiogenic capacity of endothelial cells. Angiopoietin-1 (ANG-1), Angiopoietin-2 (ANG-2), and vascular endothelial growth factor (VEGF) mRNA expression were detected using Real-Time PCR. The released VEGF, ANG-1, and ANG-2 concentrations were detected using ELISA. Western blotting was performed to measure the levels of phosphorylated GSK-3β, VEGF, ANG-1, and ANG-2. EMB-FUBINACA stimulated endothelial cell proliferation, migration, and capillary tube-like formation and promoted the expression of proangiogenic factors on RNA and protein levels. This study points out that the synthetic cannabinoid EMB-FUBINACA is a potential candidate for further investigations to confirm its potential as an inducer of brain angiogenesis. This could encourage researchers to create a new therapeutic approach for angiogenesis-related diseases.

Liquid plasma promotes angiogenesis through upregulation of endothelial nitric oxide synthase-induced extracellular matrix metabolism: potential applications of liquid plasma for vascular injuries

BackgroundApplications of nonthermal plasma have expanded beyond the biomedical field to include antibacterial, anti-inflammatory, wound healing, and tissue regeneration. Plasma enhances epithelial cell repair; however, the potential damage to deep tissues and vascular structures remains under investigation.ResultThis study assessed whether liquid plasma (LP) increased nitric oxide (NO) production in human umbilical vein endothelial cells by modulating endothelial NO synthase (eNOS) phosphorylation and potential signaling pathways. First, we developed a liquid plasma product and confirmed the angiogenic effect of LP using the Matrigel plug assay. We found that the NO content increased in plasma-treated water. NO in plasma-treated water promoted cell migration and angiogenesis in scratch and tube formation assays via vascular endothelial growth factor mRNA expression. In addition to endothelial cell proliferation and migration, LP influenced extracellular matrix metabolism and matrix metalloproteinase activity. These effects were abolished by treatment with NG-L-monomethyl arginine, a specific inhibitor of NO synthase. Furthermore, we investigated the signaling pathways mediating the phosphorylation and activation of eNOS in LP-treated cells and the role of LKB1-adenosine monophosphate-activated protein kinase in signaling. Downregulation of adenosine monophosphate-activated protein kinase by siRNA partially inhibited LP-induced eNOS phosphorylation, angiogenesis, and migration.ConclusionThe present study suggests that LP treatment may be a novel strategy for promoting angiogenesis in vascular damage.A1BBJsagnHJAo-Bf8DXnP6Video Graphical

The Effect of Riboflavin on Neurological Rehabilitation after Traumatic Brain Injury in Children.

Traumatic brain injury (TBI), which is the brain impairment and lesion caused by the external force injuring the head and the underlying brain, can cause pediatric death, disability, neurological disorders, and even lifelong disability. This study was to explore the effect of riboflavin (RF) on neurological rehabilitation and functional recovery after TBI. The rat models of TBI were constructed by treating rats with controlled cortical impact (CCI). By treating TBI rats with RF, we investigated whether the administration of RF would affect the sensorimotor function and cognitive ability recovery through adhesive removal test, modified neurological severity score (mNSS), corner test, wire-grip test and the Morris water maze. The effects of RF on lesion volume and water content were investigated using hematoxylin and eosin (H&E) staining and wet-dry method. The Nissl staining and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) staining were used to demonstrate the effect of RF on neural apoptosis. Inflammation-related cytokines of interleukin (IL)-6, IL-1β, tumor necrosis factor (TNF)-α, and transforming growth factor (TGF)-β1 were measured by enzyme-linked immunosorbent assay (ELISA) to evaluate the effect of RF on neuroinflammation. The impact of RF on oxidative stress was assessed by measuring malondialdehyde (MDA) content and superoxide dismutase (SOD) activity, and the platelet endothelial cell adhesion molecule-1 (CD31) staining for observing vessel density, the reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) for measuring vascular endothelial growth factor (VEGF) mRNA expression and western blot for VEGF protein expression were used for evaluated angiogenesis. The administration of RF could facilitate the recovery of neurological function by promoting the recovery of sensorimotor function and cognitive ability (p < 0.05). Furthermore, RF could reduce the lesion volume and water content after TBI and ameliorate neural apoptosis, neuroinflammation, and oxidative stress (p < 0.05). Finally, RF increased vessel density (p < 0.01) and VEGF levels (p < 0.01) in brain tissues after TBI, promoting angiogenesis. RF benefits neurological rehabilitation after TBI by promoting neurological function recovery, ameliorating the pathogenesis after TBI, and facilitating brain vascular remodeling. These findings provide a novel mechanism for RF treating pediatric TBI.

Suppressive Effects of Lactobacillus on Depression through Regulating the Gut Microbiota and Metabolites in C57BL/6J Mice Induced by Ampicillin

Depression is a medical and social problem. Multiple metabolites and neuroinflammation regulate it. Modifying the gut microbiota with probiotics to reduce depression through the gut-brain axis is a potential treatment strategy. In this study, three anti-depressive potentials of Lactobacillus spp. (LAB), including L. rhamnosus GMNL-74, L. acidophilus GMNL-185 and L. plantarum GMNL-141, which combined to produce low dosage LAB (1.6 × 108 CFU/mouse, LABL) and high dosage LAB (4.8 × 108 CFU/mouse, LABH), were administered to C57BL/6 mice induced depression by ampicillin (Amp). A behavioral test of depression, 16S ribosomal RNA gene amplicon sequencing, bioinformatic analysis, and short-chain fatty acid (SCFA) content measurement were executed to investigate the gut microbiota composition, activation of nutrient metabolism pathways, levels of inflammatory factors, gut-derived 5-HT biosynthesis genes, and SCFA levels in C57BL/6 mice. Results showed that after mice were induced by Amp, both LAB groups recovered from depressive behaviors, decreased the abundance of Firmicutes, and increased the abundance of Actinobacteria and Bacteroidetes in the mouse ileum. The prediction of metabolism pathways of microbes revealed the activation of arginine and proline metabolism, cyanoamino acid metabolism, and nicotinate and nicotinamide metabolism were increased, and fatty acid synthesis was decreased in both LAB groups. The LABH groups showed increased levels of acetic acid, propanoic acid, and iso-butyric acid and decreased butyric acid levels in the cecum. LABH treatment increased claudin-5 and reduced IL-6 mRNA expression. Both LAB groups also reduced monoamine oxidase, and the LABH group increased vascular endothelial growth factor mRNA expression. These results showed that the composite of three LAB exerts antidepressant effects by regulating the gut microbiota and modifying the levels of depression-related metabolites in C57BL/6J Amp-treated mice.

Safety and Potential Effects of Cannabidiol on Skin Cells

Objective: Cannabidiol (CBD), a phytochemical active compound from the Cannabis sativa L., has become a popular ingredient in many industries, especially skincare products. However, the scientific evidence supporting its potential skin benefits and safety concerns are still unclear. Therefore, the aim of this study was to investigate the short-term and long-term cytotoxic effects of CBD and its potential melanin-promoting effect on skin cells in order to deeply evaluate the safety of CBD for use in cosmetics. Material and Methods: HaCaT keratinocytes and B16F10 melanoma cell lines were cultured and investigated in regard to the cytotoxicity of cannabidiol in various concentrations (0-10 μg/ml) in the short term and long term by sulforhodamine B (SRB) assay and clonogenic assay, respectively. Next, the cellular melanin production was measured by melanin content assay. The expression of the related genes was accessed by qPCR. Results: The short-term and long-term cytotoxicity studies revealed that CBD at a low concentration was not toxic to skin cells. In addition, CBD could induce melanogenesis in melanocytes by increasing melanin content and upregulating tyrosinase expression. Also, CBD provoked cell proliferation and enhanced vascular endothelial growth factor (VEGF) mRNA expression in keratinocytes. Conclusion: Our study demonstrated that CBD at a low concentration (0.6 μg/ml) is safe for the skin cells in vitro and should thus also be safe if applied to skin. Additionally, CBD could significantly enhance melanogenesis and cell proliferation, which confirms its potential as a cosmeceutical product.

Activation of cAMP Signaling in Response to α-Phellandrene Promotes Vascular Endothelial Growth Factor Levels and Proliferation in Human Dermal Papilla Cells.

Dermal papilla cells (DPCs) are growth factor reservoirs that are specialized for hair morphogenesis and regeneration. Due to their essential role in hair growth, DPCs are commonly used as an in vitro model to investigate the effects of hair growth-regulating compounds and their molecular mechanisms of action. Cyclic adenosine monophosphate (cAMP), an intracellular second messenger, is currently employed as a growth-promoting target molecule. In a pilot test, we found that α-phellandrene, a naturally occurring phytochemical, increased cAMP levels in DPCs. Therefore, we sought to determine whether α-phellandrene increases growth factors and proliferation in human DPCs and to identify the underlying mechanisms. We demonstrated that α-phellandrene promotes cell proliferation concentration-dependently. In addition, it increases the cAMP downstream effectors, such as protein kinase A catalytic subunit (PKA Cα) and phosphorylated cAMP-responsive element-binding protein (CREB). Also, among the CREB-dependent growth factor candidates, we identified that α-phellandrene selectively upregulated vascular endothelial growth factor (VEGF) mRNA expression in DPCs. Notably, the beneficial effects of α-phellandrene were nullified by a cAMP inhibitor. This study demonstrated the cAMP-mediated growth effects in DPCs and the therapeutic potential of α-phellandrene for preventing hair loss.

Effects of curcumin nanoparticles on proliferation and VEGF expression of human retinal pigment epithelial cells.

To investigate the effects of curcumin (Cur) nanoparticles loaded with chitosan derivatives grafted by deoxycholic acid (Chit-DC) on human retinal pigment epithelial (hRPE) cell proliferation and vascular endothelial growth factor (VEGF) mRNA expression. Cur nanoparticles were synthesized with Chit-DC as the carrier and Cur as the supported drug. Cell counting kit-8 (CCK-8) method was used to detect the effects of different concentrations of Cur/Chit-DC, Chit-DC, and Cur on the proliferation of hRPE cells for different times. The changes of Cur/Chit-DC and Cur on hRPE cell cycle were determined by flow cytometry. Semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the mRNA expression levels of VEGF in hRPE cells treated with Cur, Chit-DC and Cur/Chit-DC at 10 µg/mL for 24h. Different concentrations of Chit-DC nanoparticle treated hRPE cells had no significant difference in terms of optical density (OD) values compared with the control group at 24h and 48h. Moreover, there was no change in the cell morphology under a light microscope. After 24h treatment with Cur/Chit-DC and Cur, the percentage of G0-G1 phase cells increased and the percentage of S phase cells decreased in all concentration groups. Cur/Chit-DC and Cur in all concentration groups inhibited the proliferation of hRPE cells in a time and dose dependent manner, and reduced the expression level of VEGF mRNA. The Cur/Chit-DC nanoparticles can release Cur continuously and have sustained release function. Both Cur/Chit-DC nanoparticles and Cur could inhibit hRPE cells cultured in vitro, and could reduce the expression level of VEGF mRNA in hRPE cells.

Enhancement of Farnesoid X Receptor Inhibits Migration, Adhesion and Angiogenesis through Proteasome Degradation and VEGF Reduction in Bladder Cancers.

(1) Background: Bladder cancer is a malignant tumor mainly caused by exposure to environmental chemicals, with a high recurrence rate. NR1H4, also known as Farnesoid X Receptor (FXR), acts as a nuclear receptor that can be activated by binding with bile acids, and FXR is highly correlated with the progression of cancers. The aim of this study was to verify the role of FXR in bladder cancer cells. (2) Methods: A FXR overexpressed system was established to investigate the effect of cell viability, migration, adhesion, and angiogenesis in low-grade TSGH8301 and high-grade T24 cells. (3) Results: After FXR overexpression, the ability of migration, adhesion, invasion and angiogenesis of bladder cancer cells declined significantly. Focal adhesive complex, MMP2, MMP9, and angiogenic-related proteins were decreased, while FXR was overexpressed in bladder cancer cells. Moreover, FXR overexpression reduced vascular endothelial growth factor mRNA and protein expression and secretion in bladder cancer cells. After treatment with the proteosome inhibitor MG132, the migration, adhesion and angiogenesis caused by FXR overexpression were all reversed in bladder cancer cells. (4) Conclusions: These results may provide evidence on the role of FXR in bladder cancer, and thus may improve the therapeutic efficacy of urothelial carcinoma in the future.

Regulatory role of miRNA-23a in diabetic retinopathy.

The present study aimed to investigate the expression of microRNA (miRNA)-23a in blood and tear samples from diabetic retinopathy (DR) patients. Blood and tear samples were obtained from 33 patients with proliferative DR. Additionally, a rat model of DR was established. Reverse transcription-quantitative PCR was used to determine vascular endothelial growth factor (VEGF) mRNA and miRNA-23a expression levels, while ELISA and western blot analysis were performed to determine protein expression levels. Bioinformatics analysis and dual luciferase reporter assay were used to predict and validate the interaction between miRNA-23a and VEGF and cell proliferative ability was assessed with the MTT assay. In comparison to control patients VEGF mRNA and protein expression levels were significantly elevated in the blood and tear samples from patients with DR, while the expression level of miRNA-23a was significantly reduced. In blood and retinal tissues from a rat model of DR, the mRNA and protein expression levels of VEGF were significantly increased, while the miRNA-23a expression level was significantly decreased relative to controls. Dual luciferase reporter assay showed that miRNA-23a bound to the 3'-untranslated region (UTR) of VEGF. Moreover, over-expression of miRNA-23a significantly reduced the expression level of VEGF and the proliferative activity of human retinal microvascular endothelial cells. The elevated VEGF expression in the blood and tears of patients with DR may be related to the reduced miRNA-23a expression. miRNA-23a may regulate microvascular growth at the retina via VEGF and contribute to DR progression.

Beta vulgaris rubra L. (Beetroot) Peel Methanol Extract Reduces Oxidative Stress and Stimulates Cell Proliferation via Increasing VEGF Expression in H2O2 Induced Oxidative Stressed Human Umbilical Vein Endothelial Cells.

The antioxidant capacity of polyphenols and flavonoids present in dietary agents aids in arresting the development of reactive oxygen species (ROS) and protecting endothelial smooth muscle cells from oxidative stress/induced necrosis. Beetroot (Beta vulgaris var. rubra L.; BVr) is a commonly consumed vegetable representing a rich source of antioxidants. Beetroot peel’s bioactive compounds and their role in human umbilical vein endothelial cells (HUVECs) are still under-researched. In the present study, beetroot peel methanol extract (BPME) was prepared, and its effect on the bio-efficacy, nuclear integrity, mitochondrial membrane potential and vascular cell growth, and immunoregulation-related gene expression levels in HUVECs with induced oxidative stress were analysed. Gas chromatography–mass spectroscopy (GC-MS) results confirmed that BPME contains 5-hydroxymethylfurfural (32.6%), methyl pyruvate (15.13%), furfural (9.98%), and 2,3-dihydro-3,5-dihydroxy-6-methyl-4H-Pyran-4-one (12.4%). BPME extract effectively enhanced cell proliferation and was confirmed by MTT assay; the nuclear integrity was confirmed by propidium iodide (PI) staining assay; the mitochondrial membrane potential (Δψm) was confirmed by JC-1 staining assay. Annexin V assay confirmed that BPME-treated HUVECs showed 99% viable cells, but only 39.8% viability was shown in HUVECs treated with H2O2 alone. In addition, BPME treatment of HUVECs for 48 h reduced mRNA expression of lipid peroxide (LPO) and increased NOS-3, Nrf-2, GSK-3β, GPX, endothelial nitric oxide synthase (eNOS) and vascular cell growth factor (VEGF) mRNA expression levels. We found that BPME treatment decreased proinflammatory (nuclear factor-κβ (F-κβ), tissue necrosis factor-α (TNF-α), toll-like receptor-4 (TLR-4), interleukin-1β (IL-1β)) and vascular inflammation (intracellular adhesion molecule (ICAM), vascular cell adhesion molecule (VCAM), EDN1, IL-1β)-related mRNA expressions. In conclusion, beetroot peel treatment effectively increased vascular smooth cell growth factors and microtubule development, whereas it decreased vascular inflammatory regulators. BPME may be beneficial for vascular smooth cell regeneration, tissue repair and anti-ageing potential.

CD146+Mesenchymal stem cells treatment improves vascularization, muscle contraction and VEGF expression, and reduces apoptosis in rat ischemic hind limb

Peripheral arterial disease (PAD) is an increasingly common narrowing of the peripheral arteries that can lead to lower limb ischemia, muscle weakness and gangrene. Surgical vein or arterial grafts could improve PAD, but may not be suitable in elderly patients, prompting research into less invasive approaches. Mesenchymal stem cells (MSCs) have been proposed as potential therapy, but their effectiveness and underlying mechanisms in limb ischemia are unclear. We tested the hypothesis that treatment with naive MSCs (nMSCs) or MSCs expressing CD146 (CD146+MSCs) could improve vascularity and muscle function in rat model of hind-limb ischemia. Sixteen month old Sprague-Dawley rats were randomly assigned to 4 groups: sham-operated control, ischemia, ischemia + nMSCs and ischemia+CD146+MSCs. After 4 weeks of respective treatment, rat groups were assessed for ischemic clinical score, Tarlov score, muscle capillary density, TUNEL apoptosis assay, contractile force, and vascular endothelial growth factor (VEGF) mRNA expression. CD146+MSCs showed greater CD146 mRNA expression than nMSCs. Treatment with nMSCs or CD146+MSCs improved clinical and Tarlov scores, muscle capillary density, contractile force and VEGF mRNA expression in ischemic limbs as compared to non-treated ischemia group. The improvements in muscle vascularity and function were particularly greater in ischemia+CD146+MSCs than ischemia + nMSCs group. TUNEL positive apoptotic cells were least abundant in ischemia+CD146+MSCs compared with ischemia + nMSCs and non-treated ischemia groups. Thus, MSCs particularly those expressing CD146 improve vascularity, muscle function and VEGF expression and reduce apoptosis in rat ischemic limb, and could represent a promising approach to improve angiogenesis and muscle function in PAD.

Changes in Vascular Endothelial Growth Factor (VEGF) mRNA Expression and Collagen Release in Cultured Human Dermal Fibroblasts by Electrical Stimulation

Changes in Vascular Endothelial Growth Factor (VEGF) mRNA Expression and Collagen Release in Cultured Human Dermal Fibroblasts by Electrical Stimulation

Effects of long non-coding RNA myocardial infarction-associated transcript on retinal neovascularization in a newborn mouse model of oxygen-induced retinopathy.

Whether long non-coding RNA myocardial infarction-associated transcript is involved in oxygen-induced retinopathy remains poorly understood. To validate this hypothesis, we established a newborn mouse model of oxygen-induced retinopathy by feeding in an oxygen concentration of 75 ± 2% from postnatal day 8 to postnatal day 12, followed by in normal air. On postnatal day 11, the mice were injected with the myocardial infarction-associated transcript siRNA plasmid via the vitreous cavity to knockdown long non-coding RNA myocardial infarction-associated transcript. Myocardial infarction-associated transcript siRNA transcription significantly inhibited myocardial infarction-associated transcript mRNA expression, reduced the phosphatidylinosital-3-kinase, phosphorylated Akt and vascular endothelial growth factor immunopositivities, protein and mRNA expression, and alleviated the pathological damage to the retina of oxygen-induced retinopathy mouse models. These findings suggest that myocardial infarction-associated transcript is likely involved in the retinal neovascularization in retinopathy of prematurity and that inhibition of myocardial infarction-associated transcript can downregulate phosphatidylinosital-3-kinase, phosphorylated Akt and vascular endothelial growth factor expression levels and inhibit neovascularization. This study was approved by the Animal Ethics Committee of Shengjing Hospital of China Medical University, China (approval No. 2016PS074K) on February 25, 2016.

Tanshinone IIA suppresses ovarian cancer growth through inhibiting malignant properties and angiogenesis.

BackgroundIn Chinese herbal medicine, Tanshinone IIA (Tan-IIA) is one of the main compounds extracted from Salvia miltiorrhiza Bunge. Tan-IIA has been demonstrated to inhibit the growth of various tumors. However, the detailed molecular and cellular mechanisms of the antitumor effect of Tan-IIA have yet to be fully illuminated.MethodsA2780 and ID-8 were treated with 0, 1.2, 2.4, 4.8, or 9.6 µg/mL Tan-IIA for 24 hours. Cell counting Kit-8 assay and EdU staining were used to evaluate cell proliferation. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and flow cytometry were performed to analyze apoptosis. Western blot was carried out to determine the protein levels. Flow cytometry was used for cell cycle analysis. The levels of mRNA expression were analyzed by real-time polymerase chain reaction. The anti-tumor effect of Tan-IIA was observed in a tumor-bearing mouse model.ResultsTan-IIA inhibited the proliferation of ovarian cancer cells in a dose-dependent manner by inducing G2/M phase arrest. It also down-regulated B-cell lymphoma 2 (Bcl-2) and up-regulated Bcl-2-associated X protein (Bax) in ovarian cancer cells to induce apoptosis, and suppressed cell migration by inhibiting focal adhesion kinase phosphorylation. Tan-IIA significantly reduced vascular endothelial growth factor (VEGF) and cyclooxygenase-2 (COX2) mRNA expression in ovarian cancer cells. In vivo, Tan-IIA significantly inhibited tumor growth by inducing apoptosis and promoting anti-angiogenesis.ConclusionsThe results of this study shed light on the molecular and cellular mechanisms for the antitumor effect of Tan-IIA.

A Type 2 Deiodinase-Dependent Increase in Vegfa Mediates Myoblast-Endothelial Cell Crosstalk During Skeletal Muscle Regeneration.

Background: The type 2 deiodinase (DIO2) converts thyroxine to 3,3',5-triiodothyronine (T3), modulating intracellular T3. An increase in DIO2 within muscle stem cells during skeletal muscle regeneration leads to T3-dependent potentiation of differentiation. The muscle stem cell niche comprises numerous cell types, which coordinate the regeneration process. For example, muscle stem cells provide secretory signals stimulating endothelial cell-mediated vascular repair, and, in turn, endothelial cells promote muscle stem differentiation. We hypothesized that Dio2 loss in muscle stem cells directly impairs muscle stem cell-endothelial cell communication, leading to downstream disruption of endothelial cell function. Methods: We assessed the production of proangiogenic factors in differentiated C2C12 cells and in a C2C12 cell line without Dio2 (D2KO C2C12) by real-time quantitative-polymerase chain reaction and enzyme-linked immunosorbent assay. Conditioned medium (CM) was collected daily in parallel to evaluate its effects on human umbilical vein endothelial cell (HUVEC) proliferation, migration and chemotaxis, and vascular network formation. The effects of T3-treatment on vascular endothelial growth factor (Vegfa) mRNA expression in C2C12 cells and mouse muscle were assessed. Chromatin immunoprecipitation (ChIP) identified thyroid hormone receptor (TR) binding to the Vegfa gene. Using mice with a targeted disruption of Dio2 (D2KO mice), we determined endothelial cell number by immunohistochemistry/flow cytometry and evaluated related gene expression in both uninjured and injured skeletal muscle. Results: In differentiated D2KO C2C12 cells, Vegfa expression was 46% of wildtype (WT) C2C12 cells, while secreted VEGF was 45%. D2KO C2C12 CM exhibited significantly less proangiogenic effects on HUVECs. In vitro and in vivo T3 treatment of C2C12 cells and WT mice, and ChIP using antibodies against TRα, indicated that Vegfa is a direct genomic T3 target. In uninjured D2KO soleus muscle, Vegfa expression was decreased by 28% compared with WT mice, while endothelial cell numbers were decreased by 48%. Seven days after skeletal muscle injury, D2KO mice had 36% fewer endothelial cells, coinciding with an 83% decrease in Vegfa expression in fluorescence-activated cell sorting purified muscle stem cells. Conclusion:Dio2 loss in the muscle stem cell impairs muscle stem cell-endothelial cell crosstalk via changes in the T3-responsive gene Vegfa, leading to downstream impairment of endothelial cell function both in vitro and in vivo.

Lipoxin A4 interferes with embryo implantation via suppression of epithelial-mesenchymal transition.

To test whether lipoxin A4 (LXA4) interferes with embryo implantation via suppression of epithelial-mesenchymal transition (EMT). We developed a mouse model of LXA4 blocking embryo implantation and detected the indicators of EMT to confirm that LXA4 inhibits EMT might be a mechanism of interfering with the embryo implantation. We detected integrin-linked kinase (ILK), N-formylpeptide receptor 2 (FPR2), vascular endothelial growth factor, matrix metalloproteinases (MMPs), Akt, GSK3β, NF-ĸB, twist, vimentin, fibronectin, and β-catenin mRNA expression using reverse transcriptase-polymerase chain reaction (RT-PCR) and real-time RT-PCR; localized protein expression using immunohistochemistry and Western blotting assay; MMPs activity assay by gelatin zymography; and the status of implantation in pregnant animals assessed by pontamine blue reaction test. Preimplantation administration of LXA4 resulted in implantation failure. LXA4 has a time- and dose-dependent effect on embryo implantation. Day 0.5 after fertilization is the most effective time to use LXA4 to block embryo implantation. (a) LXA4 reduced endometrial stroma edema; (b) LXA4 inhibited the activity of MMP9 and significantly upregulated the expression of β-catenin, and downregulated the expression of vimentin, fibronectin, twist, NF-κB, Akt, and Gsk-3β in the endometrium and TEV-1 cells; (c) LXA4 upregulated the expression of FPR2, and downregulated the expression of ILK; FPR2-overexpressing had an inhibitory effect on ILK in TEV-1 cells. LXA4 inhibits EMT which attenuates ILK action by enhancing FPR2; therefore, this might be a mechanism of interfering with embryo implantation.

Vascular Endothelial Growth Factor and Thrombospondin-1 mRNA Expression in Bladder Tumors: Correlation with Histopathology and Prognosis

Vascular Endothelial Growth Factor and Thrombospondin-1 mRNA Expression in Bladder Tumors: Correlation with Histopathology and Prognosis

Fructose 1,6-Bisphosphate as a Protective Agent for Experimental Fat Grafting.

Fat grafting procedures are considered to be a promising regenerative, cell‐directed therapy; however, their survival is mainly influenced by ischemia condition. Fructose 1,6‐bisphosphate (FBP), as an intermediate in energy metabolism, has the potential to rescue cells and tissues from hypoxic‐ischemic circumstances. In the present study, human lipoaspirates were grafted subcutaneously into nude mice followed by a daily intraperitoneal injection of FBP at different doses for 7 days. Next, the grafts were harvested at different time points till 12 weeks postimplantation and were evaluated for cell viability and function, tissue revascularization and inflammatory cell infiltration using histological analysis, whole‐mount living tissue imaging, glycerol 3‐phosphate dehydrogenase activity assays, and quantitative analysis of gene expression. The results demonstrated that exogenous FBP administration could attenuate the volume and weight reduction of fat graft; meanwhile, FBP enhanced adipocyte viability and function, increased blood vessel formation, and decreased inflammation. Moreover, in vitro cell experiments showed that FBP could promote adipose‐derived stem cell viability and vascular endothelial growth factor (VEGF) mRNA expression in ischemia conditions. Our study indicates that FBP can be used as a protective agent for fat grafting and may be applied in stem cell‐based regenerative medicine. stem cells translational medicine 2019;8:606–616

TNF-alpha increases angiogenic potential in a co-culture system of dental pulp cells and endothelial cells.

We recently demonstrated that a co-culture system of human umbilical vein endothelial cells (HUVECs) and human dental pulp stem cells (hDPSCs) could enhance angiogenesis ability in vitro. However, whether tumor necrosis factor α (TNF-α) could promote blood vessel formation during pulp regeneration remained unknown. The aim of this study was to investigate the effects of TNF-α on the formation of endothelial tubules and vascular networks in a co-culture system of hDPSCs and HUVECs. hDPSCs were co-cultured with HUVECs at a ratio of 1:5. The Matrigel assay was performed to detect the total tubule branching lengths and numbers of branches, and the Cell-Counting Kit 8 assay was performed to examine the effect of TNF-α on cell proliferation. Real-time polymerase chain reactions and western blot were used to detect vascular endothelial growth factor (VEGF) mRNA and protein expression. The Matrigel assay showed significantly greater total branching lengths and numbers of branches formed in the experimental groups treated with different concentrations of TNF-α compared with the control group. The decomposition times of the tubule structures were also significantly prolonged (P < 0.05). Treatment with 50 ng/ml TNF-α did not significantly change the proliferation of co-cultured cells, but it significantly increased the VEGF mRNA and protein expression levels (p < 0.05). In addition, the migration abilities of HUVECs and hDPSCs increased after co-culture with TNF-α (p < 0.05). TNF-α enhanced angiogenic ability in vitro in the co-culture system of hDPSCs and HUVECs.

PKM2 and HIF-1α regulation in prostate cancer cell lines.

Prostate cancer (PCA) is one of the most common cancer types in men, with cancer progression being linked to hypoxia and the induction of hypoxia-inducible factor (HIF).We investigated the expression of pyruvate kinase M2 (PKM2), its regulation by HIF isoforms 1α and 2α, and its role in HIF stabilization. We additionally examined cell survival in the prostate cancer cell lines PC3 and LNCaP under severe hypoxic (0.1% O2) and normoxic (20% O2) conditions. qRT-PCR showed higher up-regulation of PKM2 mRNA expression in LNCaP cells than in PC3 cells, while western blotting showed that PKM2 protein levels were up-regulated only in LNCaP cells. Inhibition of HIF-1α and HIF-2α by small interfering RNA (si-RNA) demonstrated HIF-1α dependent up-regulation of PKM2 at the mRNA and protein levels in LNCaP cells. PKM2 inhibition by si-RNA significantly decreased hypoxia-response element (HRE) activation in a gene reporter assay and down-regulated HIF-1α target vascular endothelial growth factor (VEGF) mRNA expression in PC3 cells, whereas HIF-1α protein levels were not significantly reduced. Additionally, PKM2 inhibition significantly reduced clonogenic survival in both cell lines in a colony formation assay. Prolyl hydroxylase 3 (PHD3) mRNA expression was up-regulated in both cell lines. It has been shown that PKM2 expression is regulated by HIF-1α and that PKM2 favors HIF-1α transactivation under mild (1% O2) but not severe (0.1% O2) hypoxic conditions, and some of our findings are consistent with these previous results. However, this mechanism was not fully observed in our studied cell lines, as PKM2 regulation and HIF-1α stabilization at the transactivation level occurred under severe hypoxic conditions. This discrepancy suggests that tumor tissue origin and cell type influence this model. Our findings expand the current knowledge of the mechanisms of PCA regulation, and would be important in developing novel therapeutic strategies.