Release Of Angiogenic Factors Research Articles

Umbelliferone alleviates impaired wound healing and skin barrier dysfunction in high glucose-exposed dermal fibroblasts and diabetic skins.

Skin wound healing is a complex process involving various cellular and molecular events. However, chronic wounds, particularly in individuals with diabetes, often experience delayed wound healing, potentially leading to diabetic skin complications. In this study, we examined the effects of umbelliferone on skin wound healing using dermal fibroblasts and skin tissues from a type 2 diabetic mouse model. Our results demonstrate that umbelliferone enhances several crucial aspects of wound healing. It increases the synthesis of key extracellular matrix components such as collagen I and fibronectin, as well as proteins involved in cell migration like EVL and Fascin-1. Additionally, umbelliferone boosts the secretion of angiogenesis factors VEGF and HIF-1α, enhances the expression of cell adhesion proteins including E-cadherin, ZO-1, and Occludin, and elevates levels of skin hydration-related proteins like HAS2 and AQP3. Notably, umbelliferone reduces the expression of HYAL, thereby potentially decreasing tissue permeability. As a result, it promotes extracellular matrix deposition, activates cell migration and proliferation, and stimulates pro-angiogenic factors while maintaining skin barrier functions. In summary, these findings underscore the therapeutic potential of umbelliferone in diabetic wound care, suggesting its promise as a treatment for diabetic skin complications. KEY MESSAGES: Umbelliferone suppressed the breakdown of extracellular matrix components in the skin dermis while promoting their synthesis. Umbelliferone augmented the migratory and proliferative capacities of fibroblasts. Umbelliferone activated the release of angiogenic factors in diabetic wounds, leading to accelerated wound healing. Umbelliferone bolstered intercellular adhesion and reinforced the skin barrier by preventing moisture loss and preserving skin hydration.

ADAM8 deficiency in macrophages promotes cardiac repair after myocardial infarction via ANXA2-mTOR-autophagy pathway

IntroductionA disintegrin and metalloproteinase 8 (ADAM8), a crucial regulator in macrophages, is closely associated with cardiovascular disease progression. ObjectivesThis study aimed to explore how ADAM8 regulates macrophage function to inhibit cardiac repair after myocardial infarction (MI). MethodsMacrophage-specific ADAM8 knockout mice (ADAM8flox/flox, Lyz2-Cre, KO) and corresponding control mice (ADAM8flox/flox, Flox) were established using the CRISPR/Cas9 system. Bone marrow transplantation was performed, and macrophage-specific ADAM8-overexpressing adeno-associated virus (AAV6-CD68-Adam8) was produced. Finally, proteomics, RNA sequencing, and co-immunoprecipitation/mass spectrometry (COIP/MS) were used to explore the underlying mechanisms involved. ResultsADAM8 was highly expressed in the plasma of patients with acute myocardial infarction (AMI) and in cardiac macrophages derived from AMI mice. ADAM8 KO mice exhibited enhanced angiogenesis, suppressed inflammation, reduced cardiac fibrosis, and improved cardiac function during AMI, which were reversed by overexpressing macrophage-specific ADAM8 and intervention with the clinical anti-angiogenic biologic bevacizumab. Bone marrow transplantation experiments produced ADAM8 KO phenotypes. RNA sequencing showed that autophagy was activated in bone marrow-derived macrophages (BMDMs) with ADAM8 KO, which was confirmed via p-mTOR Ser2448/mTOR, p62, and LC3II/I detection. Autophagy inactivation suppressed angiogenic factor release and promoted inflammation in BMDMs with ADAM8 KO. Mechanistically, ADAM8 could bind to ANXA2 and promote phosphorylation of the ANXA2 Ser26 site. ADAM8 KO impeded ANXA2 phosphorylation, inhibited mTOR Ser2448 site phosphorylation, and activated autophagy, which were demonstrated using the activation or inactivation of ANXA2 phosphorylation. ConclusionsADAM8 was increased in cardiac macrophages after AMI. The ADAM8-ANXA2-mTOR-autophagy axis in macrophages is responsible for regulating angiogenesis and inflammation following MI. Thus, ADAM8 may be a new target in MI treatment.

Anti-tumor and anti-metastasis of water-soluble sulfated β-glucan derivatives from Saccharomyces cerevisiae

Traditional fungi β-glucan commonly possesses high molecular weight with poor water solubility, which remains significant challenge in the drug development and medical application. Water-soluble β-glucan with high molecular weight (dHSCG) of 560 kDa, low molecular weight (dLSCG) of 60 kDa, and sulfated derivative (SCGS) with a molecular weight of 146 kDa and sulfate degree at 2.04 were obtained through well-controlled degradation and sulfated modification from Saccharomyces cerevisiae in this study. The structural characteristics were confirmed as β-1,3/6-glucan by FT-IR and NMR spectroscopy. Carbohydrate microarrays and surface plasmon resonance revealed distinct and contrasting binding affinities between the natural β-glucans and sulfated derivatives. SCGS exhibited strong binding to FGF and VEGF, while natural β-glucan showed no response, suggesting its potential as a novel antitumor agent. Moreover, SCGS significantly inhibited the migration rate of the highly metastatic melanoma (B16F10) cells. The lung metastasis mouse model also demonstrated that SCGS significantly reduced and eliminated the nodules, achieving an inhibition rate of 86.7% in vivo, with a dramatic improvement in IFN-α, TNF-α, and IL-1β levels. Through analysis of protein content and distribution in lung tissues, the anti-tumor and anti-metastasis mechanism of SCGS involves the regulation of degrading enzymes to protect extracellular matrix (ECM), as well as the reduction of angiogenic factor release. These findings provide a foundation for exploring the potential of SCGS in the development of new anti-tumor and anti-metastasis drugs and open up a new field in cancer research.

Thymic Stromal Lymphopoietin (TSLP) Is Cleaved by Human Mast Cell Tryptase and Chymase.

Thymic stromal lymphopoietin (TSLP), mainly expressed by epithelial cells, plays a central role in asthma. In humans, TSLP exists in two variants: the long form TSLP (lfTSLP) and a shorter TSLP isoform (sfTSLP). Macrophages (HLMs) and mast cells (HLMCs) are in close proximity in the human lung and play key roles in asthma. We evaluated the early proteolytic effects of tryptase and chymase released by HLMCs on TSLP by mass spectrometry. We also investigated whether TSLP and its fragments generated by these enzymes induce angiogenic factor release from HLMs. Mass spectrometry (MS) allowed the identification of TSLP cleavage sites caused by tryptase and chymase. Recombinant human TSLP treated with recombinant tryptase showed the production of 1-97 and 98-132 fragments. Recombinant chymase treatment of TSLP generated two peptides, 1-36 and 37-132. lfTSLP induced the release of VEGF-A, the most potent angiogenic factor, from HLMs. By contrast, the four TSLP fragments generated by tryptase and chymase failed to activate HLMs. Long-term TSLP incubation with furin generated two peptides devoid of activating property on HLMs. These results unveil an intricate interplay between mast cell-derived proteases and TSLP. These findings have potential relevance in understanding novel aspects of asthma pathobiology.

Clinicopathological and immunohistochemical characteristics of bullous pilomatricoma: a retrospective, single-center study, and comparison with ordinary pilomatricoma

BackgroundBullous pilomatricoma is a rare variant of pilomatricoma. As it has been published in sporadic case reports, a limited understanding of its clinicopathological characteristics restricts its effective diagnosis and treatment. ObjectivesThis study aimed to analyze the clinicopathological and immunohistochemical characteristics of bullous pilomatricoma to better understand the bullous transformation of pilomatricoma. MethodsThe authors conducted a retrospective study of 12 patients with bullous pilomatricoma and compared their clinical, histopathological, and immunohistochemical data with those of patients with ordinary pilomatricoma. ResultsBullous pilomatricoma showed no sex preference, with a mean onset age of 31.2 years. The common sites were the upper extremities and trunk. Bullous pilomatricoma had a shorter disease duration, a larger diameter, and a greater tendency to increase in size than those of ordinary pilomatricoma. Histopathologically, bullous pilomatricoma had a shorter duration, lesser calcification, more mitotic figures, and distinct dermal features from those of ordinary pilomatricoma. Immunohistochemically, the expression of Matrix Metalloprotease (MMP)-2, MMP-9, vascular endothelial growth factor receptor-3 (VEGFR-3), and VEGF-C was elevated. Study limitationsThe study was retrospective, and the sample size was small. ConclusionThe distinctive features of bullous pilomatricoma potentially result from dermal changes associated with the release of angiogenic factors and proteolytic enzymes. This comprehensive analysis provides novel insights into the clinical features and pathogenesis of bullous pilomatricoma.

(358) Responder Analysis from a Multicentered Survey of Men’s Perception who underwent Low Intensity Extracorpeal Shock Wave Therapy (LIESWT), as a Novel and Experimental Intervention for the Treatment of Erectile Dysfunction (ED)

Abstract Introduction Low-intensity extracorporeal shock wave therapy (LI-ESWT) is an experimental, non-FDA approved yet novel treatment modality that has recently been developed for treating erectile dysfunction (ED). The basic science hypothesis concerning the mechanism of action is that LI-ESWT causes microtrauma at the cellular level which may stimulate the release of angiogenic factors, and cause neovascularization of the treated tissue. Completed research report improvement in erectile functioning, decreased reliance on phosphodiesterase inhibitor medication as well as the regained ability to attain and maintain natural or spontaneous erections. We report here a preliminary responder analysis of men who had LIESWT as part of their erectile dysfunction treatment paradigm. Objective We report here the preliminary responder analysis results of an internet based, random retrospective survey on past users of this novel technology for the treatment of ED. Methods A random survey was sent to a convenience sample from 10 national centers (Hawaii, Washington (2), Portland, Utah, Oklahoma (2), Arizona, California, Oregon) to men who had completed 6 sessions of LIESWT for the treatment of ED. The internet-based survey examined patient perception and experience of this intervention. We present the responders analysis ( n=47). All procedures were performed by sexual medicine health care professionals after a detailed and comprehensive history, physical examination, and laboratory/hormonal assessment. Results Surveys were completed anonymously over the course of a 2-week period by men who had undergone 6 sessions of LIESWT. 28% of participants were aged 50-59; whereas 63 percent were over the age of 60. Other adjunctive treatments included erectile dysfunction (ED) medication, testosterone replacement therapy (TRT), previous plasma rich protein injections and Intracavernosal injections. 83% of those who got the procedure noticed an improvement within the first 5 weeks. 89% of responders noticed an improvement in erection firmness whereas 81% noticed an improvement in quality of erections. 77% of responders noticed an improvement in their level of sexual satisfaction and 72% noticed an improvement in sexual pleasure. 60% noticed an improvement in the overall quality of the orgasm. Overall, the procedure was well tolerated with <0.5% (3/670 procedures) adverse events reported (1 pain in penis, 1 “knot in penis” and 1 pain in the head) reported. Overall, 65% of LIESWT users would recommend this procedure to a friend. Conclusions LIESWT is a novel and experimental intervention for the treatment of ED. While it shows excellent promise, most data contain some shortcomings, including: small numbers, limited follow-p, and various non-standardized protocols. Results from a double-blind, randomized, control trial confirm that LI-ESWT may result in significant clinical improvement of erectile function and a shift towards improvement in penile hemodynamics without any adverse effects. Our participant responder analysis demonstrates that LIESWT improves erectile function and other sexuality parameters. Further larger, randomized sham-controlled, and comparative clinical trials are warranted before LI-ESWT may be considered new standard of care for men with vasculogenic ED. Disclosure No

Mitochondrial Metabolism and EV Cargo of Endothelial Cells Is Affected in Presence of EVs Derived from MSCs on Which HIF Is Activated.

Small extracellular vesicles (sEVs) derived from mesenchymal stem cells (MSCs) have attracted growing interest as a possible novel therapeutic agent for the management of different cardiovascular diseases (CVDs). Hypoxia significantly enhances the secretion of angiogenic mediators from MSCs as well as sEVs. The iron-chelating deferoxamine mesylate (DFO) is a stabilizer of hypoxia-inducible factor 1 and consequently used as a substitute for environmental hypoxia. The improved regenerative potential of DFO-treated MSCs has been attributed to the increased release of angiogenic factors, but whether this effect is also mediated by the secreted sEVs has not yet been investigated. In this study, we treated adipose-derived stem cells (ASCs) with a nontoxic dose of DFO to harvest sEVs (DFO-sEVs). Human umbilical vein endothelial cells (HUVECs) treated with DFO-sEVs underwent mRNA sequencing and miRNA profiling of sEV cargo (HUVEC-sEVs). The transcriptomes revealed the upregulation of mitochondrial genes linked to oxidative phosphorylation. Functional enrichment analysis on miRNAs of HUVEC-sEVs showed a connection with the signaling pathways of cell proliferation and angiogenesis. In conclusion, mesenchymal cells treated with DFO release sEVs that induce in the recipient endothelial cells molecular pathways and biological processes strongly linked to proliferation and angiogenesis.

Utility of sFtl-1 and Placental Growth Factor Ratio for Adequate Preeclampsia Management.

Introduction: The pathophysiology of preeclampsia is represented by placental ischemia and the release of angiogenic factors. Recent research suggests that using the value of the sFtl-1/PIGF ratio is more accurate for monitoring angiogenic activity. The aim of this study consists in assessing the clinical utility of the sFtl-1/PIGF ratio in determining the diagnosis and severity of preeclampsia. Material and Methods: In our study a descriptive and prospective plan was used for analyzing the specific value of the sFtl-1/PIGF ratio in women with preeclampsia and in women with gestational hypertension, depending on the gestational age and severity. Results: The study included 59 women with preeclampsia and 25 women with gestational hypertension. The mean value of the sFtl-1/PIGF ratio of pregnant women with preeclampsia was 209.2 pg/mL, while in the gestational hypertension group, the mean value of the sFtl-1/PIGF ratio was 46.08 pg/mL. The difference between the value of the sFtl-1/PIGF ratio of the group with preeclampsia and the gestational hypertension group was > 67 (pg/mL), with a sensitivity of 86.44% and a specificity of 92.00%. Significant differences were found between the median values of the sFtl-1/PIGF ratio in pregnant women with severe preeclampsia in the early-onset subgroup compared to those in the late-onset subgroup (307 pg/mL, and 98 pg/mL, respectively, p = 0.009 < α = 0.05). Conclusions: The sFtl-1/PIGF ratio may be an alternative method for diagnosing preeclampsia and it can provide data about this condition's severity.

Gene Variations of Chemokine and Chemokine Receptor CXCL12/CXCR4 in Lung Cancer

Lung cancer is the most common type of cancer in the world and about 1 million people die from lung cancer every year in the world. Inflammation is an important factor in the onset, progression and metastasis of lung cancer. The most important regulators of inflammation are chemokines and chemokine receptors. Chemokines induce the proliferation of cancer cells and prevent their apoptosis. Chemokines may indirectly affect tumor growth by inducing growth and release of angiogenic factors from cells in the tumor microenvironment. CXCL12/CXCR4 are chemokine and chemokine receptors predicted to be involved in lung cancer pathogenesis. This study aimed to determine the relationship between CXCL12/CXCR4 gene variations and CXCL12 serum levels in disease pathogenesis in lung cancer. For this purpose, DNA samples isolated from 90 lung cancer patients (36 squamous cell carcinomas, 18 small cell carcinomas and 36 adenocarcinomas) and 90 control individuals were genotyped by PCR-RFLP method for CXCL12 (rs1801157) and CXCR4 (rs2228014). CXCL12 protein levels were determined from serum samples by the enzyme-linked immuno-sorbent assay (ELISA) method. Results were evaluated using IBM SPSS Statistics 21 software and FINNETI program. As a result, there was no significant difference between the genotype frequencies of the CXCL12 rs1801157 variant and the risk of lung cancer (P = 0.396). CXCR4 rs2228014 genotypes were significantly associated with lung cancer risk (P < 0.001). Lung cancer patients had significantly elevated serum CXCL12 levels than controls (P < 0.001). In conclusion, the rs2228014 variants localized on the chemokine receptors CXCR4 gene was found to be closely related to lung cancer risk.

CD70-Targeted Micelles Enhance HIF2α siRNA Delivery and Inhibit Oncogenic Functions in Patient-Derived Clear Cell Renal Carcinoma Cells.

The majority of clear cell renal cell carcinomas (ccRCCs) are characterized by mutations in the Von Hippel−Lindau (VHL) tumor suppressor gene, which leads to the stabilization and accumulation of the HIF2α transcription factor that upregulates key oncogenic pathways that promote glucose metabolism, cell cycle progression, angiogenesis, and cell migration. Although FDA-approved HIF2α inhibitors for treating VHL disease-related ccRCC are available, these therapies are associated with significant toxicities such as anemia and hypoxia. To improve ccRCC-specific drug delivery, peptide amphiphile micelles (PAMs) were synthesized incorporating peptides targeted to the CD70 marker expressed by ccRCs and anti-HIF2α siRNA, and the ability of HIF2α-CD27 PAMs to modulate HIF2α and its downstream targets was evaluated in human ccRCC patient-derived cells. Cell cultures were derived from eight human ccRCC tumors and the baseline mRNA expression of HIF2A and CD70, as well as the HIF2α target genes SLC2A1, CCND1, VEGFA, CXCR4, and CXCL12 were first determined. As expected, each gene was overexpressed by at least 63% of all samples compared to normal kidney proximal tubule cells. Upon incubation with HIF2α-CD27 PAMs, a 50% increase in ccRCC-binding was observed upon incorporation of a CD70-targeting peptide into the PAMs, and gel shift assays demonstrated the rapid release of siRNA (>80% in 1 h) under intracellular glutathione concentrations, which contributed to ~70% gene knockdown of HIF2α and its downstream genes. Further studies demonstrated that knockdown of the HIF2α target genes SLC2A1, CCND1, VEGFA, CXCR4, and CXCL12 led to inhibition of their oncogenic functions of glucose transport, cell proliferation, angiogenic factor release, and cell migration by 50−80%. Herein, the development of a nanotherapeutic strategy for ccRCC-specific siRNA delivery and its potential to interfere with key oncogenic pathways is presented.

ALDH1A1 overexpression in melanoma cells promotes tumor angiogenesis by activating the IL‑8/Notch signaling cascade.

ALDH1A1 is a cytosolic enzyme upregulated in tumor cells, involved in detoxifying cells from reactive aldehydes and in acquiring resistance to chemotherapeutic drugs. Its expression correlates with poor clinical outcomes in a number of cancers, including melanoma. The present study hypothesized that the increased ALDH1A1 expression and activity upregulated the release of proangiogenic factors from melanoma cells, which regulate angiogenic features in endothelial cells (ECs) through a rearrangement of the Notch pathway. In vivo, when subcutaneously implanted in immunodeficient mice, ALDH1A1 overexpressing melanoma cells displayed a higher microvessel density. In a 3D multicellular system, obtained co-culturing melanoma cancer cells with stromal cells, including ECs, melanoma ALDH1A1 overexpression induced the recruitment of ECs into the core of the tumorspheres. By using a genes array, overexpression of ALDH1A1 in tumor cells also promoted modulation of Notch cascade gene expression in ECs, suggesting an interaction between tumor cells and ECs mediated by enrichment of angiogenic factors in the tumor microenvironment. To confirm this hypothesis, inactivation of ALDH1A1 by the pharmacological inhibitor CM037 significantly affected the release of angiogenic factors, including IL-8, from melanoma cells. High levels of ALDH1A1, through the retinoic acid pathway, regulated the activation of NF-κB-p65 and IL-8. Further, in a 2D co-culture system, the addition of an IL-8 neutralizing antibody to ECs co-cultured with melanoma cells forced to express ALDH1A1 dampened endothelial angiogenic features, both at the molecular (in terms of gene and protein expression of mediators of the Notch pathway) and at the functional level (proliferation, scratch assay, tube formation and permeability). In conclusion, these findings demonstrated the existence of a link between melanoma ALDH1A1 expression and EC Notch signaling modification that results in a pro-angiogenic phenotype. Based on the crucial role of ALDH1A1 in melanoma control of the tumor microenvironment, the enzyme seems a promising target for the development of novel drugs able to interrupt the cross-talk between cancer (stem) cells and endothelial cells.

Preeclampsia associated changes in volume density of fetoplacental vessels in Chinese women and mouse model of preeclampsia

Preeclampsia (PE) is associated with abnormal placental vascular structure. However, the volume density of fetoplacental vessels in PE remains unclear. Additionally, manually annotated CT angiography, which is widely used to analyze placental vessels, has issues regarding inaccuracy. Thus, computer-aided CT angiography for analyzing the volume density of fetoplacental vessels would facilitate the understanding of PE pathogenesis. We performed computer-aided CT angiography to compare differences in placentas among 17 women with PE and 34 normotensive women. The contrast ratio in CT angiography was significantly enhanced using a three-dimensional (3-D) Hessian matrix algorithm. The PE-like mouse model was established by administration of 125mg/kg/day NG-nitro-l-arginine methyl ester (l-NAME) for 10 days. The presence of endothelial marker CD31 was confirmed by quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry (IHC). The expression of angiogenic factors (PlGF, VEGFA, and sFlt1) in placentas was detected using qRT-PCR and western blotting. The volume density in fetoplacental vessels and CD31 expression were significantly reduced in women with PE and l-NAME-induced mice. Additionally, the downregulation of angiogenic factors (PlGF/VEGFA) and upregulation of an anti-angiogenic factor (sFlt1) were determined in a mouse model. Contrast-enhanced CT angiography with the aid of a 3-D Hessian matrix algorithm was performed. PE significantly affects the formation of vascular vessels, resulting in a lower volume density of fetoplacental vessels in humans and mice. This may be explained by the abnormal release of angiogenic factors during PE.

Periosteum-derived podoplanin-expressing stromal cells regulate nascent vascularization during epiphyseal marrow development

Bone marrow development and endochondral bone formation occur simultaneously. During endochondral ossification, periosteal vasculatures and stromal progenitors invade the primary avascular cartilaginous anlage, which induces primitive marrow development. We previously determined that bone marrow podoplanin (PDPN)-expressing stromal cells exist in the perivascular microenvironment and promote megakaryopoiesis and erythropoiesis. In this study, we aimed to examine the involvement of PDPN-expressing stromal cells in postnatal bone marrow generation. Using histological analysis, we observed that periosteum-derived PDPN-expressing stromal cells infiltrated the cartilaginous anlage of the postnatal epiphysis and populated on the primitive vasculature of secondary ossification center. Furthermore, immunophenotyping and cellular characteristic analyses indicated that the PDPN-expressing stromal cells constituted a subpopulation of the skeletal stem cell lineage. In vitro xenovascular model cocultured with human umbilical vein endothelial cells and PDPN-expressing skeletal stem cell progenies showed that PDPN-expressing stromal cells maintained vascular integrity via the release of angiogenic factors and vascular basement membrane-related extracellular matrices. We show that in this process, Notch signal activation committed the PDPN-expressing stromal cells into a dominant state with basement membrane-related extracellular matrices, especially type IV collagens. Our findings suggest that the PDPN-expressing stromal cells regulate the integrity of the primitive vasculatures in the epiphyseal nascent marrow. To the best of our knowledge, this is the first study to comprehensively examine how PDPN-expressing stromal cells contribute to marrow development and homeostasis.

Targets Related to Vasculogenic Mimicry in Breast Cancer

Vasculogenic mimicry (VM) is the release of angiogenic factors from tumor cells, leading to morphological changes, migration and proliferation of vascular endothelial cells, ultimately leading to neovascularization. The presence of VM has been identified in breast cancer, the formation of VM also gives breast cancer a poor prognosis such as drug resistance and metastasis. The authors focus on the role of the PI3K/Akt signaling pathway, hypoxia-inducible factorα(HIFα), the P38/MAPK signaling pathway, cyclooxygenase (COX2), the epithelial-mesenchymal transition (EMT) pathway, non-coding RNAs, and the tumor phenotype in the development of VM in breast cancer, thus offering new ideas for the future treatment of breast cancer.

Adhesive protein-based angiogenesis-mimicking spatiotemporal sequential release of angiogenic factors for functional regenerative medicine

Damaged vascular structures after critical diseases are difficult to completely restore to their original conditions without specific treatments. Thus, therapeutic angiogenesis has been spotlighted as an attractive strategy. However, effective strategies for mimicking angiogenic processes in the body have not yet been developed. In the present work, we developed a bioengineered mussel adhesive protein (MAP)-based novel therapeutic angiogenesis platform capable of spatiotemporally releasing angiogenic growth factors to target disease sites with high viscosity and strong adhesiveness in a mucus-containing environment with curvature. Polycationic MAP formed complex coacervate liquid microdroplets with polyanionic hyaluronic acid and subsequently gelated into microparticles. Platelet-derived growth factor (PDGF), which is a late-phase angiogenic factor, was efficiently encapsulated during the process of coacervate microparticle formation. These PDGF-loaded microparticles were blended with vascular endothelial growth factor (VEGF), which is the initial-phase angiogenic factor, in MAP-based pregel solution and finally crosslinked in situ into a hydrogel at the desired site. The microparticle-based angiogenic-molecule spatiotemporal sequential (MASS) release platform showed good adhesion and underwater durability, and its elasticity was close to that of target tissue. Using two in vivo critical models, i.e., full-thickness excisional wound and myocardial infarction models, the MASS release platform was evaluated for its in vivo feasibility as an angiogenesis-inducing platform and demonstrated effective angiogenesis as well as functional regenerative efficacy. Based on these superior physicochemical characteristics, the developed MASS release platform could be successfully applied in many biomedical practices as a waterproof bioadhesive with the capability for the spatiotemporal delivery of angiogenic molecules in the treatment of ischemic diseases.

Size-controlled human adipose-derived stem cell spheroids hybridized with single-segmented nanofibers and their effect on viability and stem cell differentiation

BackgroundFabrication of three-dimensional stem cell spheroids have been studied to improve stem cell function, but the hypoxic core and limited penetration of nutrients and signaling cues to the interior of the spheroid were challenges. The incorporation of polymers such as silica and gelatin in spheroids resulted in relatively relaxed assembly of composite spheroids, and enhancing transport of nutrient and biological gas. However, because of the low surface area between cells and since the polymers were heterogeneously distributed throughout the spheroid, these polymers cannot increase the cell to extracellular matrix interactions needed to support differentiation.MethodsWe developed the stem cell spheroids that incorporate poly(ι-lactic acid) single-segmented fibers synthesized by electrospinning and physical and chemical fragmentation. The proper mixing ratio was 2000 cells/μg fibers (average length of the fibers was 50 μm - 100 μm). The SFs were coated with polydopamine to increase cell binding affinity and to synthesize various-sized spheroids. The function of spheroids was investigated by in vitro analysis depending on their sizes. For statistical analysis, Graphpad Prism 5 software (San Diego, CA, USA) was used to perform one-way analysis of variance ANOVA with Tukey’s honest significant difference test and a Student’s t-test (for two variables) (P < 0.05).ResultsSpheroids of different sizes were created by modulating the amount of cells and fibers (0.063 mm2–0.322 mm2). The fibers in the spheroid were homogenously distributed and increased cell viability, while cell-only spheroids showed a loss of DNA contents, internal degradation, and many apoptotic signals. Furthermore, we investigated stemness and various functions of various-sized fiber-incorporated spheroids. In conclusion, the spheroid with the largest size showed the greatest release of angiogenic factors (released VEGF: 0.111 ± 0.004 pg/ng DNA), while the smallest size showed greater effects of osteogenic differentiation (mineralized calcium: 18.099 ± 0.271 ng/ng DNA).ConclusionThe spheroids incorporating polydopamine coated single-segmented fibers showed enhanced viability regardless of sizes and increased their functionality by regulating the size of spheroids which may be used for various tissue reconstruction and therapeutic applications.

Therapeutic Potential of Tpl2 (Tumor Progression Locus 2) Inhibition on Diabetic Vasculopathy Through the Blockage of the Inflammasome Complex.

Diabetic retinopathy, one of retinal vasculopathy, is characterized by retinal inflammation, vascular leakage, blood-retinal barrier breakdown, and neovascularization. However, the molecular mechanisms that contribute to diabetic retinopathy progression remain unclear. Approach and Results: Tpl2 (tumor progression locus 2) is a protein kinase implicated in inflammation and pathological vascular angiogenesis. Nε-carboxymethyllysine (CML) and inflammatory cytokines levels in human sera and in several diabetic murine models were detected by ELISA, whereas liquid chromatography-tandem mass spectrometry analysis was used for whole eye tissues. The CML and p-Tpl2 expressions on the human retinal pigment epithelium (RPE) cells were determined by immunofluorescence. Intravitreal injection of pharmacological inhibitor or NA (neutralizing antibody) was used in a diabetic rat model. Retinal leukostasis, optical coherence tomography, and H&E staining were used to observe pathological features. Sera of diabetic retinopathy patients had significantly increased CML levels that positively correlated with diabetic retinopathy severity and foveal thickness. CML and p-Tpl2 expressions also significantly increased in the RPE of both T1DM and T2DM diabetes animal models. Mechanistic studies on RPE revealed that CML-induced Tpl2 activation and NADPH oxidase, and inflammasome complex activation were all effectively attenuated by Tpl2 inhibition. Tpl2 inhibition by NA also effectively reduced inflammatory/angiogenic factors, retinal leukostasis in streptozotocin-induced diabetic rats, and RPE secretion of inflammatory cytokines. The attenuated release of angiogenic factors led to inhibited vascular abnormalities in the diabetic animal model. The inhibition of Tpl2 can block the inflammasome signaling pathway in RPE and has potential clinical and therapeutic implications in diabetes-associated retinal microvascular dysfunction.

Jagged Ligands Enhance the Pro-Angiogenic Activity of Multiple Myeloma Cells.

Simple SummaryThe Jagged family of ligands are aberrantly expressed during multiple myeloma progression and contributes to activate Notch signaling both in myeloma cells and in the nearby bone marrow cell populations activating several pro-tumor effects. This work elucidates, in vitro, in vivo as well as in patients’ bone marrow biopsies, different mechanisms by which tumor cell-derived Jagged1 and 2 contribute to myeloma-associated angiogenesis. These include the ability to induce myeloma and bone marrow stromal cell secretion of VEGF along with a direct activation of the pro-angiogenic Notch signaling pathway in endothelial cells. This research provides a rational for a Jagged-directed therapy in multiple myeloma.Multiple myeloma (MM) is an incurable plasma cell malignancy arising primarily within the bone marrow (BM). During MM progression, different modifications occur in the tumor cells and BM microenvironment, including the angiogenic shift characterized by the increased capability of endothelial cells to organize a network, migrate and express angiogenic factors, including vascular endothelial growth factor (VEGF). Here, we studied the functional outcome of the dysregulation of Notch ligands, Jagged1 and Jagged2, occurring during disease progression, on the angiogenic potential of MM cells and BM stromal cells (BMSCs). Jagged1–2 expression was modulated by RNA interference or soluble peptide administration, and the effects on the MM cell lines’ ability to induce human pulmonary artery cells (HPAECs) angiogenesis or to indirectly increase the BMSC angiogenic potential was analyzed in vitro; in vivo validation was performed on a zebrafish model and MM patients’ BM biopsies. Overall, our results indicate that the MM-derived Jagged ligands (1) increase the tumor cell angiogenic potential by directly triggering Notch activation in the HPAECs or stimulating the release of angiogenic factors, i.e., VEGF; and (2) stimulate the BMSCs to promote angiogenesis through VEGF secretion. The observed pro-angiogenic effect of Notch activation in the BM during MM progression provides further evidence of the potential of a therapy targeting the Jagged ligands.

Co-Microencapsulation of Islets and MSC CellSaics, Mosaic-Like Aggregates of MSCs and Recombinant Peptide Pieces, and Therapeutic Effects of Their Subcutaneous Transplantation on Diabetes.

The subcutaneous transplantation of microencapsulated islets has been extensively studied as a therapeutic approach for type I diabetes. However, due to the lower vascular density and strong inflammatory response in the subcutaneous area, there have been few reports of successfully normalized blood glucose levels. To address this issue, we developed mosaic-like aggregates comprised of mesenchymal stem cells (MSCs) and recombinant peptide pieces called MSC CellSaics, which provide a continuous release of angiogenic factors and anti-inflammatory cytokines. Our previous report revealed that the diabetes of immunodeficient diabetic model mice was reversed by the subcutaneous co-transplantation of the MSC CellSaics and rat islets. In this study, we focused on the development of immune-isolating microcapsules to co-encapsulate the MSC CellSaics and rat islets, and their therapeutic efficiency via subcutaneous transplantation into immunocompetent diabetic model mice. As blood glucose level was monitored for 28 days following transplantation, the normalization rate of the new immuno-isolating microcapsules was confirmed to be significantly higher than those of the microcapsules without the MSC CellSaics, and the MSC CellSaics transplanted outside the microcapsules (p < 0.01). Furthermore, the number of islets required for the treatment was reduced. In the stained sections, a larger number/area of blood vessels was observed around the new immuno-isolating microcapsules, which suggests that angiogenic factors secreted by the MSC CellSaics through the microcapsules function locally for their enhanced efficacy.

The Application of Nanomaterials in Angiogenesis.

Induction of angiogenesis has enormous potential in the treatment of ischemic diseases and the promotion of bulk tissue regeneration. However, the poor activity of angiogenic cells and proangiogenic factors after transplantation is the main problem that imposes its wide applications. Recent studies have found that the development of nanomaterials has solved this problem to some extent. Nanomaterials can be mainly classified into inorganic nanomaterials represented by metals, metal oxides and metal hydroxides, and organic nanomaterials including DNA tetrahedrons, graphene, graphene oxide, and carbon nanotubes. These nanomaterials can induce the release of angiogenic factors either directly or indirectly, thereby initiating a series of signaling pathways to induce angiogenesis. Moreover, appropriate surface modifications of nanomaterial facilitate a variety of functions, such as enhancing its biocompatibility and biostability. In clinical applications, nanomaterials can promote the proliferation and differentiation of endothelial cells or mesenchymal stem cells, thereby promoting the migration of hemangioblast cells to form new blood vessels. This review outlines the role of nanomaterials in angiogenesis and is intended to provide new insights into the clinical treatment of systemic and ischemic diseases.