Migration Of Microvascular Endothelial Cells Research Articles

Bilayer Scaffolds Synergize Immunomodulation and Rejuvenation via Layer-Specific Release of CK2.1 and the "Exercise Hormone" Lac-Phe for Enhanced Osteochondral Regeneration.

Repairing osteochondral defects necessitates the intricate reestablishment of the microenvironment. The cartilage layer consists of a porous gelatin methacryloyl hydrogel (PGelMA) covalently crosslinked with the chondroinductive peptide CK2.1 via a "linker" acrylate-PEG-N-hydroxysuccinimide (AC-PEG-NHS). This layer is optimized for remodeling the senescent microenvironment in the cartilage region, thereby establishing a regenerative microenvironment that supports chondrogenesis. For the bone layer, silk fibroin methacryloyl (SilMA) is coated onto a three dimensional (3D)-printed 45S5 bioactive glass scaffold (BG scaffold). The "exercise hormone" N-lactoyl-phenylalanine (Lac-Phe) is loaded onto the SilMA, endowing it with diversified functions to regulate the osteogenic microenvironment. Systematic analysis in vitro reveals that PGelMA-CK2.1 shifts the microenvironment from a pro-inflammatory into an anti-inflammatory condition, and alleviates cellular senescence, thus modifying the cartilage microenvironment to improve the recruitment, proliferation and chondral differentiation of bone marrow mesenchymal stem cells (BMSCs). The scaffold bone layer enhances microvascular endothelial cell proliferation, migration, and angiogenic activities, which, couple with increased BMSC recruitment and regulatory mechanisms directing BMSC differentiation, favor a shift in the "osteogenesis-adipogenesis" balance toward enhanced osteogenesis. In vivo, it is found that this biphasic biomimetic scaffold favors simultaneous dual tissue regeneration. This approach facilitates the development of bioactive regenerative scaffolds and holds great potential for clinical application.

Study on the mechanism of OSM participating in myocardial fibrosis by inhibiting TGFβ-induced EndMT of cardiac microvascular endothelial cells through SPARC/SMAD signaling.

Cardiac fibrosis constitutes a crucial element in the progression of diverse chronic cardiac conditions. Notably, a significant correlation has been observed between the endothelial-to-mesenchymal transition (EndMT) and the emergence of cardiac fibrosis. To investigate mechanisms, we employed immunofluorescence for α-SMA and CD31 analysis, Western blotting for CD34, vimentin, and SPARC overexpression. CCK8, wound healing, and transwell assay-assessed cell viability, invasion, and migration. SPARC overexpression plasmid was constructed and validated by Western blotting. Fibrosis levels were quantified via Masson staining, and collagen 1 and 3 expressions were measured using ELISA assays. Notably, in TGF-β-induced H5V cells, the downregulation of CD31 and CD34 expression, along with the upregulation of α-SMA and vimentin, suggests the induction of EndMT in cardiac fibrosis. Interestingly, OSM treatment mitigated EndMT progression, cell invasion, migration, and the expression of p-SMAD2, p-SMAD3, and SPARC in TGF-β-treated H5V cells. Further analysis revealed that OSM alleviated TGFβ-induced EndMT, invasion, and migration of cardiac microvascular endothelial cells by suppressing SPARC/SMAD signaling. Moreover, OSM therapy notably mitigated myocardial tissue fibrosis, along with a reduction in the expression of collagen 1, collagen 3, α-SMA, and CD34, while augmenting CD31 and vimentin expression in ISO-induced myocardial tissue. Additionally, OSM exhibited the ability to suppress myocardial tissue fibrosis and the expression of EndMT markers as well as SPARC/SMAD signals in ISO-induced myocardial tissue. Our comprehensive analysis unveiled that OSM contributes significantly to myocardial fibrosis modulation by inhibiting TGFβ-mediated EndMT in myocardial microvascular endothelial cells via SPARC/SMAD signaling.

CircRNA_0003307 promoted brain microvascular endothelial cell angiogenesis, invasion, and migration in cerebral ischemia-reperfusion injury: Potential involvement of miRNA-191-5p/CDK6 pathway

BackgroundsThe role of miR-191-5p in cerebral ischemia–reperfusion (I/R) injury has been established, with its expression in endothelial cells demonstrating anti-angiogenic effects. A potential circular RNA, circRNA_0003307, has been identified through bioinformatics analysis as a candidate for interaction with miR-191-5p, yet its functional significance in brain I/R injury remains unexplored. We aimed to investigate whether circRNA_0003307 regulates brain microvascular endothelial cell (BMEC) vascular tube formation, invasion, and migration by regulating the miR-191-5p cascade. MethodsMouse BMECs (bEnd.3) were culturedand exposed to oxygen-glucose deprivation (OGD). The effects of circRNA_0003307 on vessel-like tube formation and cellular migration were examined. In addition, we investigated the protective effects of circRNA_0003307 on I/R injury in mice. ResultsThe results showed the level of circRNA_0003307 was concentration-dependently increased in OGD-induced bEnd.3 cells. ODG-induction enhanced angiogenesis, migration, and invasion of bEnd.3 cells, which were further promoted by the transfection of pcDNA-0003307. Silencing circRNA_0003307 expression showed the opposite results. The dual luciferase assay demonstrated miRNA-191-5p interacted with circRNA_00033073′ UTR, and miRNA-191-5p could bind with CDK6. Meanwhile, circRNA_0003307 promoted the expression of CDK6 by sponging miRNA-191-5p. The overexpression of circRNA_0003307 activated the angiogenesis, migration, and invasion of OGD-induced bEnd.3 cells, which were hindered by miRNA-191-5p mimic or siRNA-CDK6. Thus, circRNA_0003307 promoted ODG-induced angiogenesis, migration, and invasion of bEnd.3 cells by targeting miR-191-5p/CDK6 axis. In vivo, circRNA_0003307 had protective effects on brain I/R injury, including neuroprotection, anti-apoptosis and angiogenesis. ConclusionCircRNA_0003307 may be a promisingtherapeutictarget forthe treatment of cerebral I/R injury.

Role of Gpr124 in the Migration and Proliferation of Retinal Microvascular Endothelial Cells and Microangiopathies in Diabetic Retinopathy.

Retinal microangiopathies, such as neovascularization and preretinal and vitreous hemorrhages, are the primary pathological features of diabetic retinopathy (DR). These conditions can worsen visual impairment and may result in blindness. Furthermore, multiple metabolic pathways are associated with microangiopathy in DR. However, the specific underlying pathological mechanisms remain unclear. Several studies have demonstrated the important role of G protein-coupled receptor 124 (Gpr124) in cerebral vascular endothelial cells, but its effect on the retinal endothelium has not been elucidated. In this study, we found that Gpr124 is expressed in both pathological retinal fibrous vascular membranes of DR patients and retinal blood vessels of mice, with elevated protein expression specifically observed in the retinas of DR model mice. Furthermore, Gpr124 expression was elevated after high-glucose treatment of human retinal microvascular endothelial cells (HRMECs). Inhibition of Gpr124 expression affected the high glucose-induced proliferation, migration, and tube-forming ability of HRMECs. We concluded that Gpr124 expression was upregulated in DR and promoted HRMECs angiogenesis in a high-glucose environment. This finding may help to elucidate the pathogenesis of DR and provide a critical research basis for identifying effective treatments.

Quemeiteng granule relieves goiter by suppressing thyroid microvascular endothelial cell proliferation and angiogenesis via miR-217-5p-mediated targeting of FGF2-induced regulation of the ERK pathway

Ethnopharmacological relevanceGoiters are enlargements of the thyroid gland and are a global public issue. Quemeiteng granule (QMTG) is a traditional Chinese medicine (TCM) formula used to treat goiter in Yunnan Province. However, the effectiveness and underlying mechanism of these treatments have not been fully elucidated. Aim of the studyThis study aimed to investigate the therapeutic effects of QMTG on goiter and the downstream regulatory mechanisms. Materials and methodsIn this study, we first evaluated the antigoiter efficacy of QMTG through biochemical indices [body weight, thyroid coefficient, triiodothyronine (T3), thyroxine (T4), free triiodothyronine (FT3), free thyroxine (FT4), and thyroid stimulating hormone (TSH)] and hematoxylin-eosin (HE) staining in a Propylthiouracil (PTU)-induced model. Based on microRNA sequencing (miRNA-seq) and bioinformatics analysis, key miRNA was screened out. A dual-luciferase reporter assay was performed to confirm the transcriptional regulation of the target gene by the miRNA. The viability of rat thyroid microvascular endothelial cells (RTMECs) and human thyroid microvascular endothelial cells (HTMECs) was assessed using the CCK-8 assays. The migration and angiogenesis of RTMECs and HTMECs were visualized through tube formation and wound scratch assays. Proteins involved in angiogenesis and the ERK pathway were assessed via Western blotting. ResultsQMTG significantly increased body weight, decreased the thyroid coefficient, increased the levels of T3, T4, FT3 and FT4 and reduced TSH levels in rats with goiter. QMTG also promoted the morphological recovery of thyroid follicles. MiR-217-5p was identified as a key miRNA. Our studies revealed that miR-217-5p directly targets FGF2 and that QMTG promotes the recovery of thyroid hormone (TH) levels and morphological changes in the thyroid, suppresses thyroid microvascular endothelial cell vitality, tube formation and migration, and reduces the expression of VEGF, Ang-1 and VCAM-1 triggered by miR-217-5p, thereby inhibiting the Ras/MEK/ERK cascade through FGF2. ConclusionsOur experiments demonstrated that the QMTG had therapeutic effects on goiter. These effects were attributed to the inhibition of ERK pathway-induced proliferation and angiogenesis through the targeting of FGF2 by miR-217-5p.

Neural stem cell-derived exosomes regulate cell proliferation, migration, and cell death of brain microvascular endothelial cells via the miR-9/Hes1 axis under hypoxia.

Our previous study found that mouse embryonic neural stem cell (NSC)-derived exosomes (EXOs) regulated NSC differentiation via the miR-9/Hes1 axis. However, the effects of EXOs on brain microvascular endothelial cell (BMEC) dysfunction via the miR-9/Hes1 axis remain unknown. Therefore, the current study aimed to determine the effects of EXOs on BMEC proliferation, migration, and death via the miR-9/Hes1 axis. Immunofluorescence, quantitative real-time polymerase chain reaction, cell counting kit-8 assay, wound healing assay, calcein-acetoxymethyl/propidium iodide staining, and hematoxylin and eosin staining were used to determine the role and mechanism of EXOs on BMECs. EXOs promoted BMEC proliferation and migration and reduced cell death under hypoxic conditions. The overexpression of miR-9 promoted BMEC proliferation and migration and reduced cell death under hypoxic conditions. Moreover, miR-9 downregulation inhibited BMEC proliferation and migration and also promoted cell death. Hes1 silencing ameliorated the effect of amtagomiR-9 on BMEC proliferation and migration and cell death. Hyperemic structures were observed in the regions of the hippocampus and cortex in hypoxia-induced mice. Meanwhile, EXO treatment improved cerebrovascular alterations. NSC-derived EXOs can promote BMEC proliferation and migration and reduce cell death via the miR-9/Hes1 axis under hypoxic conditions. Therefore, EXO therapeutic strategies could be considered for hypoxia-induced vascular injury.

Extended siponimod release via low-porosity PLGA fibres: a comprehensive three-month in vitro evaluation for neovascular ocular diseases.

Neovascular ocular diseases are among the most common causes of preventable or treatable vision loss. Their management involves lifelong, intravitreal injections of anti-vascular endothelial growth factor (VEGF) therapeutics to inhibit neovascularization, the key pathological step in these diseases. Anti-VEGF products approved for ocular administration are expensive biological agents with limited stability and short half-life. Additionally, their therapeutic advantages are hindered by high treatment resistance, poor patient compliance and the need for frequent, invasive administration. Herein, we used electrospinning to develop a unique, non-porous, PLGA implant for the ocular delivery of siponimod to improve ocular neovascular disease management. Siponimod is an FDA-approved drug for multiple sclerosis with a novel indication as a potential ocular angiogenesis inhibitor. The electrospinning conditions were optimised to produce a microfibrous, PLGA matte that was cut and rolled into the desired implant size. Physical characterisation techniques (Raman, PXRD, DSC and FTIR) indicated siponimod was distributed uniformly within the electrospun fibres as a stabilised, amorphous, solid dispersion with a character modifying drug-polymer interaction. Siponimod dispersion and drug-polymer interactions contributed to the formation of smooth fibres, with reduced porous structures. The apparent reduced porosity, coupled with the drug's hydrophobic dispersion, afforded resistance to water penetration. This led to a slow, controlled, Higuchi-type drug diffusion, with ∼30% of the siponimod load released over 90 days. The released drug inhibited human retinal microvascular endothelial cell migration and did not affect the cells' metabolic activity at different time points. The electrospun implant was physically stable after incubation under stress conditions for three months. This novel siponimod intravitreal implant broadens the therapeutic possibilities for neovascular ocular diseases, representing a potential alternative to biological, anti-VEGF treatments due to lower financial and stability burdens. Additionally, siponimod interaction with PLGA provides a unique opportunity to sustain the drug release from the electrospun fibres, thereby reducing the frequency of intravitreal injection and improving patient adherence.

ANNEXIN A2 FACILITATES NEOVASCULARIZATION TO PROTECT AGAINST MYOCARDIAL INFARCTION INJURY VIA INTERACTING WITH MACROPHAGE YAP AND ENDOTHELIAL INTEGRIN Β3.

Cardiac macrophages with different polarization phenotypes regulate ventricular remodeling and neovascularization after myocardial infarction (MI). Annexin A2 (ANXA2) promotes macrophage polarization to the repair phenotype and regulates neovascularization. However, whether ANXA2 plays any role in post-MI remodeling and its underlying mechanism remains obscure. In this study, we observed that expression levels of ANXA2 were dynamically altered in mouse hearts upon MI and peaked on the second day post-MI. Using adeno-associated virus vector-mediated overexpression or silencing of ANXA2 in the heart, we also found that elevation of ANXA2 in the infarcted myocardium significantly improved cardiac function, reduced cardiac fibrosis, and promoted peri-infarct angiogenesis, compared with controls. By contrast, reduction of cardiac ANXA2 exhibited opposite effects. Furthermore, using in vitro coculture system, we found that ANXA2-engineered macrophages promoted cardiac microvascular endothelial cell (CMEC) proliferation, migration, and neovascularization. Mechanistically, we identified that ANXA2 interacted with yes-associated protein (YAP) in macrophages and skewed them toward pro-angiogenic phenotype by inhibiting YAP activity. In addition, ANXA2 directly interacted with integrin β3 in CMECs and enhanced their growth, migration, and tubule formation. Our results indicate that increased expression of ANXA2 could confer protection against MI-induced injury by promoting neovascularization in the infarcted area, partly through the inhibition of YAP in macrophages and activation of integrin β3 in endothelial cells. Our study provides new therapeutic strategies for the treatment of MI injury.

Light-induced circadian rhythm disorder leads to microvascular dysfunction via up-regulating NETs

Circadian rhythm is a physical, mental, and behavioral pattern over the course of 24-hour cycle, and its disturbance is associated with increased risk of cardiovascular diseases. Microvascular dysfunction serves as an important cause of cardiovascular disease, but the relationship between rhythm disturbances and microcirculation remains elusive. Herein, we constructed the mice model of circadian rhythm disturbance and investigated the alterations of microvascular conditions. It was revealed that coronary microcirculatory function and cardiac diastolic function were significantly reduced, along with endothelium-dependent diastolic function of microvessels remarkably impaired in the rhythm-disordered group of mice compared to the control group. Notably, rhythm disturbance led to a significant upregulation of neutrophil extracellular traps (NETs) levels in mice, which cause endothelial dysfunction by inhibiting microvascular endothelial cell activity and migration capacity as well as inducing apoptosis. Additionally, intraperitoneal injection of Cl-amidine suppressed the production of NETs, which further improved coronary microcirculatory function and endothelium-dependent diastolic function. In conclusion, this study demonstrated that circadian rhythm disorders could induce the development of coronary microvascular dysfunction (CMD) through the up-regulation of NETs, providing a potential therapeutic direction for the treatment of CMD.

Metabolomic Profiling of Long-Chain Polyunsaturated Fatty Acid Oxidation in Adults with Retinal Vein Occlusion: A Case-Control Study

BackgroundLong-chain polyunsaturated fatty acids (LCPUFAs) and their metabolites are closely related to neovascular eye diseases. However, the clinical significance of their oxylipins in retinal vein occlusion (RVO) remains inconclusive. ObjectivesThis case-control study aimed to explore metabolomic profiles of LCPUFA oxidation in RVO and to identify potential indicators for diagnosis and pathologic progression. MethodsThe plasma concentrations of ω-3 (n–3) and ω-6 (n–6) LCPUFA and their oxylipins in 44 adults with RVO and 36 normal controls were analyzed using ultraperformance liquid chromatography tandem mass spectrometry. Univariate analysis combined with principal component and orthogonal projections to latent structure discriminant analysis was used to screen differential metabolites. Aortic ring and choroidal explant sprouting assays were used to investigate the effects of 5-oxo-eicosatetraenoic acids (ETE) on angiogenesis ex vivo. Tubule formation and wound healing assays were performed to verify its effects on human retinal microvascular endothelial cell functions. ResultsHigher ω-6 and lower ω-3 LCPUFA plasma concentrations were measured in the adults with RVO compared with control (odds ratio [OR]: 2.34; 95% confidence interval [CI]: 1.42, 3.86; P < 0.001; OR: 0.28; 95% CI: 0.15, 0.51; P < 0.001). Metabolomic analysis revealed 20 LCPUFA and their oxylipins dysregulated in RVO, including increased arachidonic acid (ω-6, OR: 1.85; 95% CI: 1.18, 2.90; P < 0.001) and its lipoxygenase product 5-oxo-ETE (OR: 11.76; 95% CI: 3.73, 37.11; P < 0.001), as well as decreased docosahexaenoic acid (ω-3, OR: 0.13; 95% CI: 0.05, 0.33; P < 0.001). Interestingly, 5-oxo-ETE was downregulated in ischemic compared with nonischemic central RVO. Exogenous 5-oxo-ETE attenuated aortic ring and choroidal explant sprouting and inhibited tubule formation and migration of human retinal microvascular endothelial cells in a dose-dependent manner, possibly via suppressing the vascular endothelial growth factor signaling pathway. ConclusionsThe plasma concentrations of ω-6 and ω-3 LCPUFA and their oxylipins were associated with RVO. The ω-6 LCPUFA-derived metabolite 5-oxo-ETE was a potential marker of RVO development and progression.

The endothelial-to-mesenchymal transition changes the focal adhesion site proteins levels and the SLRP-lumican level in HMEC-1 cell line

Scleroderma, the chronic autoimmune disease is a consequence of inflammation in the connective tissue. Prolonged duration affects formation of compact connective tissue strands (scarring) within the target organ. Endothelial cells undergoing endothelial-to-mesenchymal transition (EndMT) are the source of fibroblast phenotype-resembling cells. EndMT contributes to reorganization of the focal adhesion proteins (FA), including integrins, and intensive extracellular matrix (ECM) remodelling. However, in endothelial cells, the relationship between EndMT and the interaction of integrin receptors with lumican - a component of ECM, is still unclear. Our findings indicate that at the early stages of EndMT caused by Snail-1 transcription factor overexpression, the level of the β1 integrin subunit and its phosphorylation are elevated. Simultaneously, the changes in the level of proteins that build FAs and promote activation of integrin receptors as well as a decrease in lumican quantity were observed. These modulations contributed to increased migration of human microvascular endothelial cells, HMEC-1. Our findings were achieved by WB, ELISA and wound healing assay. Taken altogether, transfection of HMEC-1 cells with Snail-1 plasmids inducing the early stages of EndMT results in the increase of total FAK and integrin β1 phosphorylation as well as cell migration: phenomena which are modulated by interaction with lumican.

Overexpression and potential roles of midkine via regulation of vascular endothelial growth factor A in psoriasis.

Midkine plays a critical role in angiogenesis by regulating the vascular endothelial growth factor (VEGF) signalling pathway, which is known to be associated with psoriasis pathogenesis. However, research on midkine-psoriasis relationship remains limited. The objective of this study was to detect midkine expression in psoriasis and investigate its potential role in the disease. Midkine expression was measured using immunohistochemistry and ELISA. Effects of midkine on HaCaT cell proliferation, VEGF-A production and signalling pathways were assessed using CCK8, RT-PCR and WB. Scratch and in vitro tube formation tests were used to evaluate the effects of HaCaT-cell-activated midkine on the migration and tube formation of human dermal microvascular endothelial cells. Murine psoriasiform models were injected with midkine recombinant protein and midkine monoclonal antibody to investigate skin lesions, tissue sections and dermal microvessel density. Levels of midkine significantly increased in both lesions and serum of patients with psoriasis. Serum expression of midkine decreased after treatment and a positive correlation was found between midkine and disease severity. Midkine promoted HaCaT cell proliferation and VEGF-A production. The Notch2/HES1/JAK2-STAT5A pathway expression increased after midkine treatment of HaCaT cells. The supernatant of HaCaT cells treated with midkine promoted HMEC-1 migration and angiogenesis in vitro. Recombinant midkine protein exacerbated psoriasiform lesions with increased expressions of VEGF-A and microvessel density, while midkine monoclonal antibody alleviated psoriasis lesions. Midkine may have a significant impact on psoriasis angiogenesis by regulating VEGF-A expression through the Notch2/HES1/JAK2-STAT5A pathway, highlighting a potential therapeutic target for psoriasis treatment.

The 14-Kilodalton Human Growth Hormone Fragment a Potent Inhibitor of Angiogenesis and Tumor Metastasis.

The 14-kilodalton human growth hormone (14 kDa hGH) N-terminal fragment derived from the proteolytic cleavage of its full-length counterpart has been shown to sustain antiangiogenic potentials. This study investigated the antitumoral and antimetastatic effects of 14 kDa hGH on B16-F10 murine melanoma cells. B16-F10 murine melanoma cells transfected with 14 kDa hGH expression vectors showed a significant reduction in cellular proliferation and migration associated with an increase in cell apoptosis in vitro. In vivo, 14 kDa hGH mitigated tumor growth and metastasis of B16-F10 cells and was associated with a significant reduction in tumor angiogenesis. Similarly, 14 kDa hGH expression reduced human brain microvascular endothelial (HBME) cell proliferation, migration, and tube formation abilities and triggered apoptosis in vitro. The antiangiogenic effects of 14 kDa hGH on HBME cells were abolished when we stably downregulated plasminogen activator inhibitor-1 (PAI-1) expression in vitro. In this study, we showed the potential anticancer role of 14 kDa hGH, its ability to inhibit primary tumor growth and metastasis establishment, and the possible involvement of PAI-1 in promoting its antiangiogenic effects. Therefore, these results suggest that the 14 kDa hGH fragment can be used as a therapeutic molecule to inhibit angiogenesis and cancer progression.

L-ascorbic acid could ameliorate the damage of myocardial microvascular endothelial cell caused by hypoxia-reoxygenation via targeting HMGB1

In this study, we intend to explore the potential function of l-ascorbic acid in hypoxia-reoxygenation (H/R)-induced damage of CMECs and its related molecular mechanism. With different concentrations of l-ascorbic acid treatment, the proliferation, migration, inflammation and autophagy of cardiac microvascular endothelial cells (CMECs) were determined by several biological experiments. Si-HMGB1 transfection was used to reduce HMGB1 expression and to detect the function of HMGB1 in H/R-induced damage of CMECs. Under H/R condition, the proliferation and migration abilities of CMECs were reduced, and the inflammation and autophagy of CMECs were increased. Whereas, after l-ascorbic acid treatment, the reduction in the proliferation and migration of CMECs, as well as the increase in the inflammation and autophagy of CMECs induced by H/R were reversely altered. HMGB1 was confirmed as a specific target of l-ascorbic acid, and si-HMGB1 treatment strengthened the beneficial effect of l-ascorbic acid on H/R-induced damage of CMECs, followed by further reduction in the proliferation and migration abilities of CMECs, as well as the increase in the inflammation and autophagy of CMECs. Few studies have reported the function of l-ascorbic acid in myocardial ischemia on CMECs, but our experimental data showed that l-ascorbic acid treatment could ameliorate the H/R-induced damage of CMECs by regulating HMGB1 expression.

Up-regulation of miR-192-5p inhibits the ELAVL1/PI3Kδ axis and attenuates microvascular endothelial cell proliferation, migration and angiogenesis in diabetic retinopathy.

Diabetic retinopathy (DR) is a common complication of diabetes mellitus that poses a threat to adults. MicroRNAs (miRNAs) play a key role in DR progression. However, the role and mechanism of miR-192-5p in DR remain unclear. We aimed to investigate the effect of miR-192-5p on cell proliferation, migration and angiogenesis in DR. Expression of miR-192-5p, ELAV-like RNA binding protein 1 (ELAVL1) and phosphoinositide 3-kinase delta (PI3Kδ) in human retinal fibrovascular membrane (FVM) samples and human retinal microvascular endothelial cells (HRMECs) was assessed using RT-qPCR. ELAVL1 and PI3Kδ protein levels were evaluated by Western blot. RIP and dual luciferase reporter assays were performed to confirm the miR-192-5p/ELAVL1/PI3Kδ regulatory networks. Cell proliferation, migration and angiogenesis were assessed by CCK8, transwell and tube formation assays. MiR-192-5p was decreased in FVM samples from DR patients and high glucose (HG)-treated HRMECs. Functionally, overexpressed miR-192-5p inhibited cell proliferation, migration and angiogenesis in HG-treated HRMECs. Mechanically, miR-192-5p directly targeted ELAVL1 and decreased its expression. We further verified that ELAVL1 bound to PI3Kδ and maintained PI3Kδ mRNA stability. Rescue analysis demonstrated that the suppressive effects of HG-treated HRMECs caused by miR-192-5p up-regulation were overturned by overexpressed ELAVL1 or PI3Kδ. MiR-192-5p attenuates DR progression by targeting ELAVL1 and reducing PI3Kδ expression, suggesting a biomarker for the treatment of DR.

USP14 Regulates ATF2/PIK3CD Axis to Promote Microvascular Endothelial Cell Proliferation, Migration, and Angiogenesis in Diabetic Retinopathy.

Diabetic retinopathy (DR) is one of the leading causes of blindness in diabetic patients. However, the pathogenesis of DR is complex, and no firm conclusions have been drawn so far. It has become a hot spot in ophthalmology research to deeply study the mechanism of DR pathological changes and find effective treatment options. Human retinal microvascular endothelial cells (HRMECs) were induced by high glucose (HG) to construct DR cell model. CCK-8 assay was used to detect the viability of HRMECs. Transwell assay was used to detect the migration ability of HRMECs. Tube formation assay was used to identify the tube formation ability of HRMECs. The expressions of USP14, ATF2 and PIK3CD were detected by Western blot analysis and qRT-PCR assay. Immunoprecipitation (IP) was used to ascertain the relationship of USP14 and ATF2. To explore the regulatory relationship between ATF2 and PIK3CD by dual-luciferase reporter gene assay and Chromatin immunoprecipitation (ChIP) assay. High glucose treatment promoted the proliferation, migration, and tube formation of HRMEC, and the expressions of USP14, ATF2 and PIK3CD were significantly up-regulated. USP14 or ATF2 knockdown inhibited HG-induced HRMECs proliferation, migration, and tube formation. USP14 regulated the expression of ATF2, and ATF2 promoted PIK3CD expression. PIK3CD overexpression attenuated the inhibitory effectiveness of USP14 knockdown on proliferation, migration and tube formation of DR cell model. Here, we revealed that USP14 regulated the ATF2/PIK3CD axis to promote proliferation, migration, and tube formation in HG-induced HRMECs.

Microfibril-Associated Glycoprotein-2 Promoted Fracture Healing via Integrin αvβ3/PTK2/AKT Signaling

Fracture healing is a complex physiological process in which angiogenesis plays an essential role. Microfibril-associated glycoprotein-2 (MAGP2) has been reported to possess a proangiogenic activity via integrin αvβ3, yet its role in bone repair is unexplored. In this study, a critical-sized femoral defect (2 mm) was created in mice, followed by the delivery of an adenovirus-based MAGP2 overexpression vector or its negative control at the fracture site. At days 7, 14, 21, and 28 postfracture, bone fracture healing was evaluated by radiography, micro-computed tomography, and histopathologic analysis. Adenovirus-based MAGP2 overexpression vector–treated mice exhibited increased bone mineral density and bone volume fraction. MAGP2 overexpression contributed to an advanced stage of endochondral ossification and induced cartilage callus into the bony callus. Further analysis indicated that MAGP2 was associated with enhanced angiogenesis, as evidenced by marked MAGP2 and integrin αvβ3 costaining and increased endothelial cell markers such as endomucin and CD31 levls, as well as elevated phosphorylation of protein tyrosine kinase 2 (PTK2) and AKT serine/threonine kinase 1 (AKT) in the callus. In vitro, recombinant human MAGP2 treatment enhanced the viability, migration, and tube formation ability of human microvascular endothelial cells, which was partially reversed by integrin αvβ3 inhibition or MK-2206, a specific AKT inhibitor. Inhibition of integrin αvβ3 abolished MAGP2-induced PTK2 and AKT activation. Taken together, our data provide the first evidence that MAGP2 promotes angiogenesis and bone formation by activating the integrin αvβ3/PTK2/AKT signaling pathway.

ADSC-derived exosomes attenuate myocardial infarction injury by promoting miR-205-mediated cardiac angiogenesis

BackgroundAcute myocardial infarction is a major health problem and is the leading cause of death worldwide. Myocardial apoptosis induced by myocardial infarction injury is involved in the pathophysiology of heart failure. Therapeutic stem cell therapy has the potential to be an effective and favorable treatment for ischemic heart disease. Exosomes derived from stem cells have been shown to effectively repair MI injury-induced cardiomyocyte damage. However, the cardioprotective benefits of adipose tissue-derived mesenchymal stem cell (ADSC)-Exos remain unknown. This study aimed to investigate the protective effects of exosomes from ADSC on the hearts of MI-treated mice and to explore the underlying mechanisms.MethodsCellular and molecular mechanisms were investigated using cultured ADSCs. On C57BL/6J mice, we performed myocardial MI or sham operations and assessed cardiac function, fibrosis, and angiogenesis 4 weeks later. Mice were intramyocardially injected with ADSC-Exos or vehicle-treated ADSCs after 25 min following the MI operation.ResultsEchocardiographic experiments showed that ADSC-Exos could significantly improve left ventricular ejection fraction, whereas ADSC-Exos administration could significantly alleviate MI-induced cardiac fibrosis. Additionally, ADSC-Exos treatment has been shown to reduce cardiomyocyte apoptosis while increasing angiogenesis. Molecular experiments found that exosomes extracted from ADSCs can promote the proliferation and migration of microvascular endothelial cells, facilitate angiogenesis, and inhibit cardiomyocytes apoptosis through miRNA-205. We then transferred isolated exosomes from ADSCs into MI-induced mice and observed decreased cardiac fibrosis, increased angiogenesis, and improved cardiac function. We also observed increased apoptosis and decreased expression of hypoxia-inducible factor-1α and vascular endothelial growth factor in HMEC-1 transfected with a miRNA-205 inhibitor.ConclusionIn summary, these findings show that ADSC-Exos can alleviate cardiac injury and promote cardiac function recovery in MI-treated mice via the miRNA-205 signaling pathway. ADSC-Exos containing miRNA205 have a promising therapeutic potential in MI-induced cardiac injury.

Boosting UPR transcriptional activator XBP1 accelerates acute wound healing.

Patients' suffering from large or deep wounds caused by traumatic and/or thermal injuries have significantly lower chances of recapitulating lost skin function through natural healing. We tested whether enhanced unfolded protein response (UPR) by expression of a UPR transcriptional activator, X-box-binding protein 1 (XBP1) can significantly promote wound repair through stimulating growth factor production and promoting angiogenesis. In mouse models of a second-degree thermal wound, a full-thickness traumatic wound, and a full-thickness diabetic wound, the topical gene transfer of the activated form of XBP1 (spliced XBP1, XBP1s) can significantly enhance re-epithelialization and increase angiogenesis, leading to rapid, nearly complete wound closure with intact regenerated epidermis and dermis. Overexpression of XBP1s stimulated the transcription of growth factors in fibroblasts critical to proliferation and remodeling during wound repair, including platelet-derived growth factor BB, basic fibroblast growth factor, and transforming growth factor beta 3. Meanwhile, the overexpression of XBP1s boosted the migration and tube formation of dermal microvascular endothelial cells in vitro. Our functional and mechanistic investigations of XBP1-mediated regulation of wound healing processes provide novel insights into the previously undermined physiological role of the UPR in skin injuries. The finding opens an avenue to developing potential XBP1-based therapeutic strategies in clinical wound care protocols.

BIBF1120 Protects against Diabetic Retinopathy through Neovascularization-Related Molecules and the MAPK Signaling Pathway.

Diabetic retinopathy (DR) is one of the microvascular complications of diabetes mellitus and a major pathological feature of neovascular DR. These patients potentially experience vision impairment and blindness. Platelet-derived growth factor receptor β (PDGFRβ), fibroblast growth factor receptor 1 (FGFR1), and vascular endothelial growth factor receptor 2 (VEGFR2) are implicated in the DR pathogenesis. Nintedanib (BIBF1120) is an oral selective dual receptor tyrosine kinase (RTK) inhibitor of VEGFR2, FGFR1, and PDGFRβ. In this study, intravitreal injection of BIBF1120 blocked the phosphorylation of VEGFR2, FGFR1, PDGFRβ, and MAPK signaling pathway proteins in a streptozotocin (STZ)-induced diabetic retinopathy mouse model. In in vitro cell experiments, BIBF1120 did not change cellular activity under normal conditions, while it further suppressed the tube formation, migration, and proliferation of high glucose-induced human retinal microvascular endothelial cells (HRMECs). Additionally, BIBF1120 blocked the phosphorylation of p38, JNK, and ERK1/2 in high glucose-treating HRMECs. Our results indicate that the BIBF1120 treatment can be a novel potential drug to protect against DR.