Expression Of Angiogenic Growth Factors Research Articles

Effects of Ibuprofen and Diclofenac Pre-Treatment on Viability and Apoptosis Processes in Human Dental Pulp Stem Cells.

Background and Objectives: Stem cell-based regeneration strategies have shown therapeutic efficacy in various fields of regenerative medicine. These include bone healing after bone augmentation, often complicated by pain, which is managed by using nonsteroidal anti-inflammatory drugs (NSAIDs). However, information is limited about how NSAIDs affect the therapeutic potential of stem cells. Materials and Methods: We investigated the effects of ibuprofen and diclofenac on the characteristics, morphology, and immunophenotype of human mesenchymal stromal cells isolated from the dental pulp (DPSCs) and cultured in vitro, as well as their effects on the expression of angiogenic growth factors (VEGFA and HGF) and selected genes in apoptosis signalling pathways (BAX, BAK, CASP3, CASP9, and BCL2). Results: Ibuprofen and diclofenac significantly reduced the viability of DPSCs, while the expression of mesenchymal stem cell surface markers was unaffected. Both ibuprofen and diclofenac treatment significantly upregulated the expression of HGF, while the expression of VEGFA remained unchanged. Ibuprofen significantly altered the expression of several apoptosis-related genes, including the upregulation of CASP9 and BCL2, with decreased CASP3 expression. BAK, CASP3, CASP9, and BCL2 expressions were significantly increased in the diclofenac-treated DPSCs, while no difference was demonstrated in BAX expression. Conclusions: Our results suggest that concomitant use of the NSAIDs ibuprofen or diclofenac with stem cell therapy may negatively impact cell viability and alter the expression of apoptosis-related genes, affecting the efficacy of stem cell therapy.

Fe-Porphyrin Cross-Linked Hydrogel for Reactive Oxygen Species Scavenging and Oxygen Generation in Diabetic Wounds.

Healing chronic diabetic wounds is challenging because of excessive reactive oxygen species (ROS) and hypoxia in the wound microenvironment. To address this issue, we propose a hydrogel wound dressing composed of polyethylene glycol (PEG) cross-linked with a biomimetic catalase, Fe-containing porphyrin (FeP) (i.e., FeP hydrogel). The immobilized FeP can serve as a catalyst for both ROS scavenging and O2 generation. The properties of the hydrogels were optimized by varying the composition ratios of the two constituent materials based on their mechanical properties and catalytic activity. Our in vitro cell experiments revealed that the FeP-80 hydrogel enhanced the proliferation and migration of keratinocytes and dermal fibroblasts and promoted the expression of angiogenic growth factors in keratinocytes. When tested with an in vivo diabetic chronic wound model, the FeP-80 hydrogel promoted wound healing by facilitating re-epithelialization, promoting angiogenesis, and suppressing inflammation, compared with other control groups.

Evaluation of the genotoxicity, cytotoxicity, and bioactivity of calcium silicate-based cements

BackgroundAs calcium silicate-based cements (CSCs) have found success in various vital pulp therapy applications, several new CSC products have emerged. This study aimed to assess the genotoxicity, cytotoxicity, and bioactivity of four CSCs by comparing the newly introduced materials Bio MTA+ and MTA Cem with previously studied materials, Biodentine and NeoMTA.MethodsGenotoxicity was evaluated using the micronucleus (MN) assay in human peripheral blood lymphocyte cells, measuring MN frequency and nuclear division index (NDI). Cytotoxicity was assessed in human dental pulp stem cells through the Water-Soluble Tetrazolium Salt-1 (WST-1) colorimetric assay. Bioactivity was determined by ELISA, measuring the levels of angiogenic and odontogenic markers (BMP-2, FGF-2, VEGF, and ALP). Statistical analyses included ANOVA, Dunnet and Sidak tests, and Wald chi-square test. (p < .05).ResultsThe MN frequency in the groups was significantly lower than that in the positive control group (tetraconazole) (p < .05). NDI values decreased with increasing concentration (p < .05). Bio MTA+ and NeoMTA showed decreased cell viability at all concentrations in 7-day cultures (p < .01). All materials increased BMP-2, FGF-2, and VEGF levels, with Biodentine and NeoMTA showing the highest levels of BMP-2 and FGF-2 on day 7. Biodentine displayed the highest VEGF levels on day 7. Biodentine and NeoMTA groups exhibited significantly higher ALP activity than the Bio MTA+ and MTA Cem groups by day 7.ConclusionBio MTA+ and MTA Cem demonstrated no genotoxic or cytotoxic effects. Moreover, this study revealed bioactive potentials of Bio MTA+ and MTA Cem by enhancing the expression of angiogenic and osteogenic growth factors.

Modulation of fetoplacental growth, development and reproductive function by endocrine disrupters.

Maternal endocrine homeostasis is vital to a successful pregnancy, regulated by several hormones such as human chorionic gonadotropin, estrogen, leptin, glucocorticoid, insulin, prostaglandin, and others. Endocrine stress during pregnancy can modulate nutrient availability from mother to fetus, alter fetoplacental growth and reproductive functions. Endocrine disrupters such as bisphenols (BPs) and phthalates are exposed in our daily life's highest volume. Therefore, they are extensively scrutinized for their effects on metabolism, steroidogenesis, insulin signaling, and inflammation involving obesity, diabetes, and the reproductive system. BPs have their structural similarity to 17-β estradiol and their ability to bind as an agonist or antagonist to estrogen receptors to elicit an adverse response to the function of the endocrine and reproductive system. While adults can negate the adverse effects of these endocrine-disrupting chemicals (EDCs), fetuses do not equip themselves with enzymatic machinery to catabolize their conjugates. Therefore, EDC exposure makes the fetoplacental developmental window vulnerable to programming in utero. On the one hand prenatal BPs and phthalates exposure can impair the structure and function of the ovary and uterus, resulting in placental vascular defects, inappropriate placental expression of angiogenic growth factors due to altered hypothalamic response, expression of nutrient transporters, and epigenetic changes associated with maternal endocrine stress. On the other, their exposure during pregnancy can affect the offspring's metabolic, endocrine and reproductive functions by altering fetoplacental programming. This review highlights the latest development in maternal metabolic and endocrine modulations from exposure to estrogenic mimic chemicals on subcellular and transgenerational changes in placental development and its effects on fetal growth, size, and metabolic & reproductive functions.

Perfluorooctane sulfonate promotes the migration of colorectal cancer cells by inducing epithelial-mesenchymal transition

The potential association between colorectal cancer (CRC) and environmental pollutants is worrisome. Previous studies have found that some perfluoroalkyl acids, including perfluorooctane sulfonate (PFOS), induced colorectal tumors in experimental animals and promoted the migration of and invasion by CRC cells in vitro, but the underlying mechanism is unclear. Here, we investigated the effects of PFOS on the proliferation and migration of CRC cells and the potential mechanisms involving activating the PI3K/Akt-NF-κB signal pathway and epithelial-mesenchymal transition (EMT). It was found that PFOS promoted the growth and migration of HCT116 cells at non-cytotoxic concentrations and increased the mRNA expression of the migration-related angiogenic cytokines vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8). In a mechanistic investigation, the up-stream signal pathway PI3K/Akt-NF-κB was activated by PFOS, and the process was suppressed by LY294002 (PI3K/Akt inhibitor) and BAY11-7082 (NF-κB inhibitor) respectively, leading to less proliferation of HCT116 cells. Furthermore, matrix metalloproteinases (MMP) and EMT-related markers were up-regulated after PFOS exposure, and were also suppressed respectively by LY294002 and BAY11-7082. Moreover, the up-regulation of EMT markers was suppressed by a MMP inhibitor GM6001. Taken together, our results indicated that PFOS promotes colorectal cancer cell migration and proliferation by activating the PI3K/Akt-NF-κB signal pathway and epithelial-mesenchymal transition. This could be a potential toxicological mechanism of PFOS-induced malignant development of colorectal cancer.

Astragaloside IV promotes exosome secretion of endothelial progenitor cells to regulate PIK3R2/SPRED1 signaling and inhibit pyroptosis of diabetic endothelial cells

Background aimsTreating chronic non-healing diabetic wounds and achieving complete skin regeneration has always been a critical clinical challenge. MethodsIn order to address this issue, researchers conducted a study aiming to investigate the role of miR-126-3p in regulating the downstream gene PIK3R2 and promoting diabetic wound repair in endothelial progenitor cell (EPC)–derived extracellular vesicles. The study involved culturing EPCs with astragaloside IV, transfecting them with miR-126-3p inhibitor or mock plasmid, interfering with high glucose-induced damage in human umbilical vein endothelial cells (HUVECs) and treating diabetic skin wounds in rats. ResultsThe healing of rat skin wounds was observed through histological staining. The results revealed that treatment with miR-126-3p–overexpressing EPC-derived extracellular vesicles accelerated the healing of rat skin wounds and resulted in better tissue repair with slower scar formation. In addition, the transfer of EPC-derived extracellular vesicles with high expression of miR-126-3p to high glucose-damaged HUVECs increased their proliferation and invasion, reduced necrotic and apoptotic cell numbers and improved tube formation. In this process, the expression of angiogenic factors vascular endothelial growth factor (VEGF)A, VEGFB, VEGFC, basic fibroblast growth factor and Ang-1 significantly increased, whereas the expression of caspase-1, NRLP3, interleukin-1β, inteleukin-18, PIK3R2 and SPRED1 was suppressed. Furthermore, miR-126-3p was able to target and inhibit the expression of the PIK3R2 gene, thereby restoring the proliferation and migration ability of high glucose-damaged HUVEC. ConclusionsIn summary, these research findings demonstrate the important role of miR-126-3p in regulating downstream genes and promoting diabetic wound repair, providing a new approach for treating chronic non-healing diabetic wounds.

Engineered elastin-like polypeptide improves the efficiency of adipose-derived stem cell-mediated cutaneous wound healing in type II diabetes mellitus

Impaired cutaneous wound healing is a major complication in patients with diabetes mellitus (DM), leading to increased amputation and mortality rates in affected patients. Adipose-derived stem cells (ASCs) are widely used seed cells for promoted tissue regeneration to improve wound closure under diabetic conditions. However, ASCs-based therapies remain limited due to difficulties in maintaining cell quality during transplantation. To overcome this problem, extracellular matrix mimetic biomaterials have been developed for use in biomedical engineering field, including tissue engineering and regenerative medicine. Herein, a biosynthesized arginine–glycine–aspartate amino acid residues (RGD motif, known as a cell adhesion motif)-containing elastin-like polypeptides (REPs) improved the efficacy of ASCs in enhancing wound closure and skin elasticity in diabetic wounds by promoting the expression of angiogenic growth factors. Therefore, REPs can be used as potential supplements to stem cell-based therapeutic approach to accelerate diabetic wound repair.

O-266 Anti-angiogenic therapy as treatment for adenomyosis in mice

Abstract Study question What is the effect of anti-angiogenic therapy on adenomyosis in mice Summary answer Commencing treatment at six weeks with angiogenesis inhibitor axitinib successfully reduced the severity of adenomyosis by approximately 50% at 9 weeks. What is known already Adenomyosis is a gynecological disorder characterized by abnormal uterine bleeding, dysmenorrhea, pelvic pain and subfertility. Increased expression of angiogenic markers, as well as increased microvascular density (MVD) in adenomyosis suggest a pivotal role for angiogenesis in the pathophysiology of adenomyosis, which presents an opportunity for treatment. Study design, size, duration The effect of angiogenesis inhibition was studied in a tamoxifen-induced adenomyosis mouse model. 102 Mice received oral treatment with axitinib 3 mg/kg (‘dose I/AX3 treatment group’, n = 34), axitinib 25 mg/kg (‘dose II/AX25 treatment group’ n = 34), or with vehicle-only (‘placebo group’, n = 34) from week 6 until week 9. After termination of the mice at week 9, all uteri were analyzed for the presence of adenomyosis. Participants/materials, setting, methods The prevalence and severity of adenomyosis were assessed by; (i) grade of adenomyosis (0/1/2/3), based on the depth of endometrial gland infiltration into the myometrium in H&amp;E-stained sections, (ii) percentage of adenomyosis-affected tissue in vimentin-stained sections and (iii) degree of myometrium that is affected in α-SMA-stained sections. The adenomyosis severity index was calculated by multiplying mean grade/mouse with the percentage affected surface area. Changes in angiogenesis-related gene expression were evaluated using real-time quantitative PCR. Main results and the role of chance 101 mice completed adenomyosis induction and could be analyzed. The prevalence of adenomyosis was 30/33 (90.0%) in dose I (3mg/kg), 29/34 (85.3%) in dose II (25mg/kg) and 30/34 (88.2%) in placebo treated mice (p=0.78). The prevalence of high grade (2/3) adenomyosis was significantly lower in mice treated with axitinib dose II (N = 19, 55.9%) than in the placebo group (N = 27, 79.4%, p&amp;lt;0.05). The grade of adenomyosis per mouse after treatment was 0.7 point lower in the groups treated with axitinib dose I and dose II, than in the placebo treated group (dose I and II both median of 1.0 compared to 1.7, p &amp;lt; 0.05). The adenomyosis severity index was reduced by 48% in the axitinib-treated groups (dose I p&amp;lt;0.05). In the placebo-treated group, the myometrium adjacent to the affected site (as a marker for hypertrophy) was thicker in the placebo- than in the axitinib-treated groups. There were no signs of fibrosis. MVD was reduced in myometrium of dose I treated mice by 14% compared with Ax25-treated mice (p&amp;lt;0.05), as well as compared to 6-week-old tamoxifen-treated mice (p&amp;lt;0.05). Expression of angiogenic growth factors and their receptors was markedly reduced in the dose I and II axitinib-treated groups compared to the placebo-treated group. Limitations, reasons for caution Limitations of this study include the uncertainty on the optimal timing to start treatment, and the assessment of the presence and severity of adenomyosis. Further research should focus on commonality among different angiostatic drugs, as well as the method and timing of application. Wider implications of the findings Since there is currently no treatment option for premenopausal women who wish to retain their uterus and their fertility, treatment with an angiogenesis inhibitor might provide a solution. The major requirement in further development of anti-angiogenesis therapy is a safety and side-effect profile tolerable for this population. Trial registration number Not applicable

Vascular characterization and morphogenesis in porcine placenta at day 40 of gestation.

In porcine placenta, abnormal development of the placental vasculature leads to placental insufficiency. The aim of this study was to determine the mRNA expression of angiogenic growth factors and to determine the vascular characteristics in placenta at day 40 of pig gestation. Samples were collected from maternal-chorioallantoic interface (n = 21) for the measurement of mRNA expression of VEGFA, ANGPT1, ANGPT2, FGF2 and its receptors KDR, TEK, FGFR1IIIc, FGFR2IIIb respectively, and for immunohistochemistry analysis of CD31 and VEGFA. Immunohistochemical analysis of CD31 and VEGFA, morphometric measurement of blood vessels, high resolution light microscopy and transmission electron microscopy were performed. Capillary area density, number of blood vessels and capillary area were significantly higher on the maternal side than on the fetal side (P<0.05). The ultrastructural finding of blood vessels demonstrates close contact with the trophoblastic epithelium. The relative mRNA expression of VEGFA and its receptor KDR were higher compared with the other angiogenic genes. In conclusion, a high mRNA expression of VEGFA and its receptor KDR added to the immunohistochemical results suggest a potential role of these genes in this pathway associated with an increase in the density of the capillary area on the maternal side and a reduction in the hemotrophic diffusion distance at the interface for nutrient exchange.

Characterization of C-X-C chemokine receptor type 5 in the cornea and role in the inflammatory response after corneal injury

C-X-C chemokine receptor type 5 (CXCR5) regulates inflammatory responses in ocular and non-ocular tissues. However, its expression and role in the cornea are still unknown. Here, we report the expression of CXCR5 in human cornea in vitro and mouse corneas in vivo, and its functional role in corneal inflammation using C57BL/6J wild-type (CXCR5+/+) and CXCR5-deficient (CXCR5−/−) mice, topical alkali injury, clinical eye imaging, histology, immunofluorescence, PCR, qRT-PCR, and western blotting. Human corneal epithelial cells, stromal fibroblasts, and endothelial cells demonstrated CXCR5 mRNA and protein expression in PCR, and Western blot analyses, respectively. To study the functional role of CXCR5 in vivo, mice were divided into four groups: Group-1 (CXCR5+/+ alkali injured cornea; n = 30), Group-2 (CXCR5−/− alkali injured cornea; n = 30), Group-3 (CXCR5+/+ naïve cornea; n = 30), and Group-4 (CXCR5−/− naïve cornea; n = 30). Only one eye was wounded with alkali. Clinical corneal evaluation and imaging were performed before and after injury. Mice were euthanized 4 h, 3 days, or 7 days after injury, eyes were excised and used for histology, immunofluorescence, and qRT-PCR. In clinical eye examinations, CXCR5−/− mouse corneas showed ocular health akin to the naïve corneas. Alkali injured CXCR5+/+ mouse corneas showed significantly increased mRNA (p < 0.001) and protein (p < 0.01 or p < 0.0001) levels of the CXCR5 compared to the naïve corneas. Likewise, alkali injured CXCR5−/− mouse corneas showed remarkably amplified inflammation in clinical eye exams in live animals. The histological and molecular analyses of these corneas post euthanasia exhibited markedly augmented inflammatory cells in H&E staining and significant CD11b + cells in immunofluorescence (p < 0.01 or < 0.05); and tumor necrosis factor-alpha (TNFα; p < 0.05), cyclooxygenase 2 (COX-2; p < 0.0001), interleukin (IL)-1β (p < 0.0001), and IL-6 (p < 0.0001 or < 0.01) mRNA expression compared to the CXCR5+/+ mouse corneas. Interestingly, CXCR5−/− alkali injured corneas also showed altered mRNA expression of fibrotic alpha smooth muscle actin (α-SMA; p > 0.05) and angiogenic vascular endothelial growth factor (VEGF; p < 0.01) compared to the CXCR5+/+ alkali injured corneas. In summary, the CXCR5 gene is expressed in all three major layers of the cornea and appears to influence corneal inflammatory and repair events post-injury in vivo. More studies are warranted to tease the mechanistic role of CXCR5 in corneal inflammation and wound healing.

RNA Transfection with Hypoxia Inducible Factor-1alpha

Introduction: Ischemic necrosis of dermal flaps is a devastating complication of reconstructive surgery. The increasing prevalence of diabetes, obesity, and an aging population adds to this concern. Hypoxia-inducible factor-1α (HIF-1α), a master regulator of the adaptive response to hypoxia, controls the expression of angiogenic growth factors. The development of biologically active, gene-specific mRNAs, especially in COVID-19 vaccines, has shown the ability for intracellular protein expression. We sought to express HIF-1α through mRNA transfection and determined its biological activity by measuring the upregulation of selected downstream targets. Methods: 5’-methyl-capped poly-A tailed mRNA was generated using T7 RNA polymerase and verified by gel electrophoresis. Predominant and variant HIF-1α mRNA were transfected into primary human dermal fibroblasts via Lipofectamine in triplicate, and RNA levels were assessed using RT-qPCR. All gene expression levels were normalized to beta-actin expression levels Results: At one day after transfection, the levels of HIF-1α transcript were significantly higher in the cells transfected with predominant (p = 0.0104) and variant (p = 0.0007) HIF-1α transcripts relative to the control. Additionally, the expression of HIF-1α transcription product genes VEGF (p = 0.0274) and ANG-1 (p = 0.05) were significantly higher in the cells transfected with the HIF-1α transcripts than the control. Conclusion: Our approach led to the successful transfection of HIF-1α mRNA into human fibroblasts, resulting in upregulation of HIF-1α downstream angiogenic targets. Thus, the use of biologically active HIF-1α mRNA transfection offers a promising approach to inhibit ischemic necrosis.

PSI-9 Disrupting Cxcl12-Cxcr4 Chemokine Axis at the Fetal-Maternal Interface During Implantation Results in Altered Expression of Angiogenic and Insulin-Like Growth Factors in Ovine Placenta Midgestation

Abstract Placental dysfunction is a fundamental cause of intrauterine growth restriction, and fetal morbidity and mortality in livestock. Though causes of placental dysfunction are not well characterized, origin of disease transpires during placental development (placentation). The C-X-C motif chemokine ligand 12 (CXCL12) working through its receptor CXCR4 is implicated in placentation but precise roles of this axis are unclear. Using an in vivo sheep model, we demonstrated that suppressing CXCL12-CXCR4 signaling at the fetal-maternal interface reduces placental vascularization. We hypothesized these negative impacts early in gestation would result in compromised placental development and vascularization at midgestation. To test, onday 12 post-breeding, osmotic pumps were surgically installed in 22 ewes and delivered either CXCR4 inhibitor (AMD3100) at 1.5X dose, (n=8) or 3X dose (n=8) or saline (n=6) into the uterine lumen ipsilateral to the corpus luteum for 14 days. On day 90 placental tissues were collected and maternal and fetal placenta components separated and analyzed. The objectives were to determine if suppressing CXCL12-CXCR4 actions at the fetal-maternal interface during implantation results in reduced placental and vascular development at midgestation. Gene expression for angiogenic factors and members of the insulin-like growth factor family were analyzed using Real-time qPCR. Of the targets tested, mRNA expression was significantly altered by AMD3100 treatment for the vascular endothelial growth factor receptors (FLT-1 and KDR) primarily in maternal placenta. Expression of insulin-like growth factors 1 and 2, IGF-binding protein 3, and placental lactogen were significantly altered in fetal placenta with AMD3100 treatment compared to control. Our preliminary data underscore the importance of CXCL12-CXCR4 signaling during placentation and provide strong evidence that altering CXCL12-mediated signaling during early placentation induces enduring placental effects manifesting later in gestation. A greater understanding of CXCL12/CXCR4 functions may reveal methods to improve reproductive success and fetal health.

Wound microenvironment self-adaptive hydrogel with efficient angiogenesis for promoting diabetic wound healing.

Neovascularization is critical to improve the diabetic microenvironment, deliver abundant nutrients to the wound and promote wound closure. However, the excess of oxidative stress impedes the healing process. Herein, a self-adaptive multifunctional hydrogel with self-healing property and injectability is fabricated through a boronic ester-based reaction between the phenylboronic acid groups of the 3-carboxyl-4-fluorophenylboronic acid -grafted quaternized chitosan and the hydroxyl groups of the polyvinyl alcohol, in which pro-angiogenic drug of desferrioxamine (DFO) is loaded in the form of gelatin microspheres (DFO@G). The boronic ester bonds of the hydrogel can self-adaptively react with hyperglycemic and hydrogen peroxide to alleviate oxidative stress and release DFO@G in the early phase of wound healing. A sustained release of DFO is then realized by responding to overexpressed matrix metalloproteinases. In a full-thickness diabetic wound model, the DFO@G loaded hydrogel accelerates angiogenesis by upregulating expression of hypoxia-inducible factor-1 and angiogenic growth factors, resulting in collagen deposition and rapid wound closure. This multifunctional hydrogel can not only self-adaptively change the microenvironment to a pro-healing state by decreasing oxidative stress, but also respond to matrix metalloproteinases to release DFO. The self-adaptive multifunctional hydrogel has a potential for treating diabetic wounds.

Dl-3-N-Butylphthalide Promotes Angiogenesis in an Optimized Model of Transient Ischemic Attack in C57BL/6 Mice.

Transient ischemic attack (TIA) has been widely regarded as a clinical entity. Even though magnetic resonance imaging (MRI) results of TIA patients are negative, potential neurovascular damage might be present, and may account for long-term cognitive impairment. Animal models that simulate human diseases are essential tools for in-depth study of TIA. Previous studies have clarified that Dl-3-N-butylphthalide (NBP) promotes angiogenesis after stroke. However, the effects of NBP on TIA remain unknown. This study aims to develop an optimized TIA model in C57BL/6 mice to explore the microscopic evidence of ischemic injury after TIA, and investigate the therapeutic effects of NBP on TIA. C57BL/6 mice underwent varying durations (7, 8, 9 or 10 min) of middle cerebral artery occlusion (MCAO). Cerebral artery occlusion and reperfusion were assessed by laser speckle contrast imaging. TIA and ischemic stroke were distinguished by neurological testing and MRI examination at 24 h post-operation. Neuronal apoptosis was examined by TUNEL staining. Images of submicron cerebrovascular networks were obtained via micro-optical sectioning tomography. Subsequently, the mice were randomly assigned to a sham-operated group, a vehicle-treated TIA group or an NBP-treated TIA group. Vascular density was determined by immunofluorescent staining and fluorescein isothiocyanate method, and the expression of angiogenic growth factors were detected by western blot analysis. We found that an 8-min or shorter period of ischemia induced neither permanent neurological deficits nor MRI detectable brain lesions in C57BL/6 mice, but histologically caused neuronal apoptosis and cerebral vasculature abnormalities. NBP treatment increased the number of CD31+ microvessels and perfused microvessels after TIA. NBP also up-regulated the expression of VEGF, Ang-1 and Ang-2 and improved the cerebrovascular network. In conclusion, 8 min or shorter cerebral ischemia induced by the suture MCAO method is an appropriate TIA model in C57BL/6 mice, which conforms to the definition of human TIA, but causes microscopic neurovascular impairment. NBP treatment increased the expression of angiogenic growth factors, promoted angiogenesis and improved cerebral microvessels after TIA. Our study provides new insights on the pathogenesis and potential treatments of TIA.

The Antimicrobial Peptide Human β-Defensin-3 Accelerates Wound Healing by Promoting Angiogenesis, Cell Migration, and Proliferation Through the FGFR/JAK2/STAT3 Signaling Pathway.

In addition to its antimicrobial activity, the skin-derived antimicrobial peptide human β-defensin-3 (hBD-3) promotes keratinocyte proliferation and migration to initiate the wound healing process; however, its effects on fibroblasts, which are the major cell type responsible for wound healing, remain unclear. We investigated the role of hBD-3 in cell migration, proliferation and production of angiogenic growth factors in human fibroblasts and evaluated the in vivo effect of hBD-3 on promoting wound healing and angiogenesis. Following hBD-3 treatment, the mouse wounds healed faster and showed accumulation of neutrophils and macrophages in the early phase of wound healing and reduction of these phagocytes 4 days later. hBD-3-treated wounds also displayed an increased number of fibroblasts and newly formed vessels compared to those of the control mice. Furthermore, the expression of various angiogenic growth factors was increased in the hBD-3-treated wounds. Additionally, in vitro studies demonstrated that hBD-3 enhanced the secretion of angiogenic growth factors such as fibroblast growth factor, platelet-derived growth factor and vascular endothelial growth factor and induced the migration and proliferation of human fibroblasts. The hBD-3-mediated activation of fibroblasts involves the fibroblast growth factor receptor 1 (FGFR1)/Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathways, as evidenced by the inhibitory effects of pathway-specific inhibitors. We indeed confirmed that hBD-3 enhanced the phosphorylation of FGFR1, JAK2 and STAT3. Collectively, the current study provides novel evidence that hBD-3 might be a potential candidate for the treatment of wounds through its ability to promote wound healing, angiogenesis and fibroblast activation.

Hypoxic Pretreatment of Adipose-Derived Stem Cells Accelerates Diabetic Wound Healing via circ-Gcap14 and HIF-1α/VEGF Mediated Angiopoiesis.

Background and ObjectivesAdipose-derived stem cell (ADSC) transplantation improves stem cell paracrine function and can enhance wound healing. However, in diabetic patients, glucose-associated effects on this function and cell survival lead to impaired wound closure, thereby limiting ADSC transplantation efficiency. The hypoxia-inducible factor HIF-1α has an important protective function during wound healing. Here, we aim to clarify the regulatory mechanism of ADSCs.Methods and ResultsADSCs were isolated from BALB/C mice adipose samples. We then used high-throughput sequencing to assess abnormal expression of circular RNAs (circRNAs). We also used an in vivo full-thickness skin defect mouse model to assess the effects of transplanted ADSC on diabetic wound closure. Hypoxic pretreatment of ADSCs accelerated diabetic wound closure, which enhanced angiogenic growth factor expression in our mouse model. High-throughput sequencing and RT-qPCR indicated that circ-Gcap14 was upregulated in hypoxic pretreated ADSCs. Similarly, circ-Gcap14 downregulation also decreased the therapeutic effects of ADSCs; however, circ-Gcap14 overexpression increased the effects of ADSC by promoting angiopoiesis. We also used a luciferase reporter assay to confirm that miR-18a-5p and HIF-1α were downstream targets of circ-Gcap14. HIF-1α expression plays an important role in increased VEGF level.ConclusionsBased on our data, we suggest that circ-Gcap14 plays an important role in accelerating hypoxic ADSC-mediated diabetic wound closure, by enhancing mouse angiogenic growth factor expression and regulating downstream miR-18a-5p/HIF-1α expression.

Dental Pulp Mesenchymal Stem Cells Attenuate Limb Ischemia via Promoting Capillary Proliferation and Collateral Development in a Preclinical Model.

Critical limb ischemia (CLI), an end-stage manifestation of peripheral artery disease (PAD), still lacks effective therapeutic strategies. Recently, dental pulp-derived mesenchymal stem cells (DP-MSCs) have been attracting more and more attentions in therapeutic applications due to their high proliferation ability, powerful osteogenic differentiation potential, and effective anti-inflammatory effects. In this study, we compared the therapeutic effects of MSCs derived from different sources in a femoral artery-ligated preclinical ischemic model. We found that treatments with MSCs, including bone marrow- (BM-), adipose- (AD-), dental pulp- (DP-), and umbilical cord- (UC-) derived MSCs, improved limb functions, reduced inflammatory responses, increased angiogenesis, and promoted regeneration of muscle fiber. Among them, DP-MSCs and BM-MSCs produced much more impressive effects in restoring limb functions and promoting angiogenesis. The flow velocity restored to nearly 20% of the normal level at 3 weeks after treatments with DP-MSCs and BM-MSCs, and obvious capillary proliferation and collateral development could be observed. Although neovascularization was induced in the ischemic limb after ligation, MSCs, especially DP-MSCs, significantly enhanced the angiogenesis. In vitro experiments showed that serum deprivation improved the expression of angiogenic factors, growth factors, and chemokines in DP-MSCs and UC-MSCs, but not in BM-MSCs and AD-MSCs. However, DP-MSCs produced stronger therapeutic responses than UC-MSCs, which might be due to the higher expression of hepatocyte growth factor (HGF) and hypoxia-inducible factor-1 α (HIF-1α). We speculated that DP-MSCs might stimulate angiogenesis and promote tissue repair via expressing and secreting angiogenic factors, growth factors, and chemokines, especially HGF and HIF-1α. In conclusion, DP-MSCs might be a promising approach for treating CLI.

Probing the role of methyl methacrylate release from spacer materials in induced membrane bone healing.

In the induced membrane (IM) technique for bone reconstruction, a poly(methyl methacrylate) (PMMA) spacer is implanted to induce formation of a foreign body membrane around the defect site. Membrane development is essential for later bone grafting success, yet the mechanism by which the IM promotes bone regeneration remains unknown, as are the ways that spacer composition plays a role in the membrane's healing potential. This study investigated the impact of leached methyl methacrylate (MMA)-the major monomeric component of PMMA-on IM development. In vitro cell culture found that MMA elution did not impact endothelial cell or mesenchymal stem cell proliferation. For in vivo analysis, we advanced a streamlined rat femoral model to efficiently study the influence of spacer properties on IM characteristics. Comparison of membrane formation around polycaprolactone (PCL), MMA-eluting PCL (high-dose PCL-MMA and low-dose PCL-MMA), and surgical PMMA revealed robust membranes enveloped all groups after 4 weeks in vivo, with elevated expression of osteogenic bone morphogenetic protein-2 and angiogenic vascular endothelial growth factor compared with the surrounding muscle and bone tissues. Growth factor quantitation in IM tissue found no statistically significant difference between groups. New bone growth, vascularization, and CD163+ macrophage populations surrounding the polymer implants were also quantified; and blood vessel formation around high-dose PCL-MMA was found to be significantly decreased compared with PCL alone. To the best of our knowledge, these findings represent the first time that results have been obtained about the characteristics of membranes formed around PCL in the IM setting.

Effects of exercise training with weight loss on skeletal muscle expression of angiogenic factors in overweight and obese older men.

Low skeletal muscle capillarization is associated with impaired glucose tolerance (IGT); however, aerobic exercise training with weight loss (AEX + WL) increases skeletal muscle capillarization and improves glucose tolerance in adults with IGT. Given that the expression of angiogenic growth factors mediates skeletal muscle capillarization, we sought to determine whether angiogenic growth factor levels are associated with low capillarization in those with IGT versus normal glucose tolerance (NGT) or to the benefits of AEX + WL in both groups. Sixteen overweight or obese men 50-75 yr of age completed 6 mo of AEX + WL with oral glucose tolerance tests and vastus lateralis muscle biopsies for measurement of muscle vascular endothelial growth factor (VEGF), placental growth factor (PlGF), soluble fms-like tyrosine kinase receptor-1 (sFlt-1), and basic fibroblast growth factor (bFGF). At baseline, all growth factor levels were numerically lower in IGT than NGT, but these did not reach statistical significance (P = 0.06-0.33). Following AEX + WL, aerobic capacity [maximal oxygen consumption (V̇o2max)] increased by 16%, whereas body weight and 120-min postprandial glucose levels decreased by 10% and 15%, respectively (P ≤ 0.001 for all). There was a main effect of AEX + WL to increase VEGF (0.095 ± 0.016 vs. 0.114 ± 0.018 ng/µg, P < 0.05), PlGF (0.004 ± 0.001 vs. 0.005 ± 0.001 ng/µg, P < 0.05), and sFlt-1 (0.216 ± 0.029 vs. 0.264 ± 0.036 ng/µg, P < 0.01), with overall increases driven by the IGT group. These results suggest that 6 mo of AEX + WL increases skeletal muscle angiogenic growth factor levels in obese older adults with IGT and NGT, which may contribute to our previous findings that AEX + WL increases capillarization to improve glucose tolerance in those with IGT.NEW & NOTEWORTHY Skeletal muscle capillarization is lower in adults with impaired glucose tolerance than normal controls. This may, in part, be attributable to differential expression of angiogenic growth factors in skeletal muscle. Using a 6-mo aerobic exercise intervention with ∼10% body weight loss (AEX + WL), we show that the expression of angiogenic growth factors tends to be lower in adults with impaired glucose tolerance compared with normal controls and that AEX + WL increased expression of angiogenic growth factors in all participants.

Rosuvastatin inhibit the cell proliferation and the expression of angiogenic factors Vascular Endothelial Growth Factor and Matrix-Metalloproteinase-9 in DU-145 human prostate cancer cells

A few tumours are exceptionally stubborn to oral chemotherapy. The endurance of tumours in a few cases is helped by checkpoint immunomodulation to keep up the unevenness between resistant reconnaissance and disease cell division.