Vascular Endothelial Growth Factor Release Research Articles

Fibrin clot and Leukocyte-rich platelet-rich fibrin show similar release kinetics and amount of growth factors: a pilot study

BackgroundIn knee arthroscopic surgery, fibrin clot (FC) and leukocyte-rich platelet-rich fibrin (L-PRF) may be used in augmentation for meniscal repair. Studies have investigated growth factors released from FC and L-PRF; however, it is difficult to compare FC and L-PRF between different studies. Direct comparison of growth factors that may support meniscal healing released from FC and L-PRF may be beneficial in deciding whether to use FC or L-PRF. If no significant difference is seen, the surgeon may decide to use FC which is easier to prepare compared to L-PRF. The purpose of this pilot study is to investigate the release amount and pattern of basic fibroblast growth factor (bFGF), platelet-derived growth factor AB (PDGF-AB), transforming growth factor β1 (TGF-β1), vascular endothelial growth factor (VEGF), and stromal cell-derived factor 1 (SDF-1) from FC and L-PRF.MethodTwenty milliliters (ml) of whole blood was collected from each of the four volunteers. Ten milliliters of whole blood was allocated for preparation of FC and 10 ml for L-PRF. FC and L-PRF were separately placed in 5 ml of culture media. Five milliliters of the culture media was sampled and refilled at 15 min, 1 day, 3 days, 1 week and 2 weeks. The collected culture was used to quantify bFGF, PDGF-AB, TGF-β1, VEGF, and SDF-1 release by Enzyme-linked immune-sorbent assay (ELISA). Mann–Whitney U test was performed to assess significance of differences in amount of each growth factor released between FC and L-PRF. Significance was accepted at P value less than 0.05.ResultsAt two weeks, the cumulative release of TGF-β1 was the highest among all the growth factors in both FC and L-PRF (FC:19,738.21 pg/ml, L-PRF: 16,229.79 pg/ml). PDGF-AB (FC: 2328 pg/ml, L-PRF 1513.57 pg/ml) had the second largest amount, followed by VEGF (FC: 702.06 pg/ml, L-PRF 595.99 pg/ml) and bFGF (FC: 23.48 pg/ml, L-PRF 18.2 pg/ml), which order was also common in both FC and L-PRF. No significant difference in final release amount and pattern was seen between FC and L-PRF.ConclusionThe current pilot study showed that cumulative release amount and release pattern of PDGF-AB, VEGF, TGF-β1, and bFGF did not significantly differ between FC and L-PRF during the two weeks of observation.

Effects of hypoxia on bronchial and alveolar epithelial cells linked to pathogenesis in chronic lung disorders.

Introduction: Chronic lung disorders involve pathological alterations in the lung tissue with hypoxia as a consequence. Hypoxia may influence the release of inflammatory mediators and growth factors including vascular endothelial growth factor (VEGF) and prostaglandin (PG)E2. The aim of this work was to investigate how hypoxia affects human lung epithelial cells in combination with profibrotic stimuli and its correlation to pathogenesis. Methods: Human bronchial (BEAS-2B) and alveolar (hAELVi) epithelial cells were exposed to either hypoxia (1% O2) or normoxia (21% O2) during 24h, with or without transforming growth factor (TGF)-β1. mRNA expression of genes and proteins related to disease pathology were analysed with qPCR, ELISA or immunocytochemistry. Alterations in cell viability and metabolic activity were determined. Results: In BEAS-2B and hAELVi, hypoxia significantly dowregulated genes related to fibrosis, mitochondrial stress, oxidative stress, apoptosis and inflammation whereas VEGF receptor 2 increased. Hypoxia increased the expression of Tenascin-C, whereas both hypoxia and TGF-β1 stimuli increased the release of VEGF, IL-6, IL-8 and MCP-1 in BEAS-2B. In hAELVi, hypoxia reduced the release of fibroblast growth factor, epidermal growth factor, PGE2, IL-6 and IL-8, whereas TGF-β1 stimulus significantly increased the release of PGE2 and IL-6. TGF-β1 stimulated BEAS-2B cells showed a decreased release of VEGF-A and IL-8, while TGF-β1 stimulated hAELVi cells showed a decreased release of PGE2 and IL-8 during hypoxia compared to normoxia. Metabolic activity was significantly increased by hypoxia in both epithelial cell types. Discussion: In conclusion, our data indicate that bronchial and alveolar epithelial cells respond differently to hypoxia and profibrotic stimuli. The bronchial epithelium appears more responsive to changes in oxygen levels and remodelling processes compared to the alveoli, suggesting that hypoxia may be a driver of pathogenesis in chronic lung disorders.

A 3D printed polylactic acid-Baghdadite nanocomposite scaffold coated with microporous chitosan-VEGF for bone regeneration applications

Three-dimensional (3D) printing technology has become an advanced approach for fabricating patient-specific scaffolds with complex geometric shapes to replace damaged or diseased tissue. Herein, polylactic acid (PLA)-Baghdadite (Bgh) scaffold were made through the fused deposition modeling (FDM) 3D printing method and subjected to alkaline treatment. Following fabrication, the scaffolds were coated with either chitosan (Cs)-vascular endothelial growth factor (VEGF) or lyophilized Cs-VEGF known as PLA-Bgh/Cs-VEGF and PLA-Bgh/L.(Cs-VEGF), respectively. Based on the results, it was found that the coated scaffolds had higher porosity, compressive strength and elastic modulus than PLA and PLA-Bgh samples. Also, the osteogenic differentiation potential of scaffolds following culture with rat bone marrow-derived mesenchymal stem cells (rMSCs) was evaluated through crystal violet and Alizarin-red staining, alkaline phosphatase (ALP) activity and calcium content assays, osteocalcin measurements, and gene expression analysis. The release of VEGF from the coated scaffolds was assessed and also the angiogenic potential of scaffolds was evaluated. The sum of results presented in the current study strongly suggests that the PLA-Bgh/L.(Cs-VEGF) scaffold can be a proper candidate for bone healing applications.

3D-bioprinted double-crosslinked angiogenic alginate/chondroitin sulfate patch for diabetic wound healing

Improving chronic wound healing remains a challenge in the clinical practice. In this study, we developed double-crosslinked angiogenic 3D-bioprinted patches for diabetic wound healing by the photocovalent crosslinking of vascular endothelial growth factor (VEGF) using ultraviolet (UV) irradiation. 3D printing technology can precisely customize the structure and composition of patches to meet different clinical requirements. The biological polysaccharide alginate and chondroitin sulfate methacryloyl were used as biomaterials to construct the biological patch, which could be crosslinked using calcium ion crosslinking and photocrosslinking, thereby improving its mechanical properties. More importantly, acrylylated VEGF could be easily and rapidly photocrosslinked under UV irradiation, which simplified the step of chemically coupling growth factors and prolonged VEGF release time. These characteristics suggest that 3D-bioprinted double-crosslinked angiogenic patches are ideal candidates for diabetic wound healing and other tissue engineering applications.

Cellular infiltration in an injectable sulfated cellulose nanocrystal hydrogel and efficient angiogenesis by VEGF loading

BackgroundCellular infiltration and angiogenesis into implanted biomaterial scaffolds are crucial for successful host tissue integration and tissue regeneration. Cellulose nanocrystal (CNC) is a nano-sized cellulose derivative, which can form an injectable physical gel with salts. Sulfate groups of sulfated CNC (CNC-S) can act as a binding domain to various growth factors and cytokines with a heparin-binding domain for sustained release of them. Vascular endothelial growth factor (VEGF) can promote the proliferation of endothelial cells and angiogenesis. In this study, VEGF-loaded CNC-S hydrogel was evaluated as an injectable scaffold that can induce cellular infiltration and angiogenesis.MethodsCNC-S was hydrolyzed to get desulfated CNC (CNC-DS), which was used as a negative control group against CNC-S. Both CNC-S and CNC-DS hydrogels were prepared and compared in terms of biocompatibility and VEGF release. The hydrogels with or without VEGF loading were subcutaneously injected into mice to evaluate the biocompatibility, cellular infiltration, and angiogenesis induction of the hydrogels.ResultsBoth hydrogels possessed similar stability and shear-thinning behavior to be applicable as injectable hydrogels. However, CNC-S hydrogel showed sustained release (until 8 weeks) of VEGF whereas CNC-DS showed a very fast release of VEGF with a large burst. Subcutaneously injected CNC-S hydrogel showed much enhanced cellular infiltration as well as better biocompatibility with milder foreign body response than CNC-DS hydrogel. Furthermore, VEGF-loaded CNC-S hydrogel induced significant angiogenesis inside the hydrogel whereas VEGF-loaded CNC-DS did not.ConclusionCNC-S possesses good properties as a biomaterial including injectability, biocompatibility, and allowing cellular infiltration and sustained release of growth factors. VEGF-loaded CNC-S hydrogel exhibited efficient angiogenesis inside the hydrogel. The sulfate group of CNC-S was a key for good biocompatibility and the biological activities of VEGF-loaded CNC hydrogel.

Replicating endothelial shear stress in organ-on-a-chip for predictive hypericin photodynamic efficiency

Photodynamic therapy using Hypericin (Hy-PDT) is an alternative non-invasive treatment that enables selective tumor inhibition and angiogenesis derived from the differential recruitment of endothelial cells in the tumor microenvironment. Most PDT studies were performed on in vitro models without vascular biomechanical simulation. Our work strives to develop a microchip that generates a constant shear stress force to investigate the Hy-PDT efficiency on human umbilical vein endothelial cells (HUVECs). The microchip with a single straight microchannel was composed of the bottom layer (polystyrene), the middle layer (double-sided biocompatible adhesive tape), and the top layer (polyester film) and could produce shear stress in the range of 1.4 – 7.0 dyn cm−2. The quantification of vascular endothelial growth factor (VEGF), cell viability, and activities of caspases 3 and 7 were assayed to validate the microchip and Hy-PDT efficacy. After the endothelization, static and dynamic cell incubations with Hy were conducted in microchips. Compared to static systems, the shear stress displayed its effect on the increasing release of VEGF and promoted more cell damage and cell death via necrosis during Hy-PDT. In conclusion, the expressive shear stress-dependent manner during PDT treatments suggests that the microchip could be an essential approach in preclinical tests to evaluate the therapeutic outcome considering the endothelial shear stress microenvironment.

Multifunctional nanoparticle-VEGF modification for tissue-engineered vascular graft to promote sustained anti-thrombosis and rapid endothelialization.

Purpose: The absence of a complete endothelial cell layer is a well-recognized reason leading to small-diameter tissue-engineered vascular graft failure. Here we reported a multifunctional system consisting of chitosan (CS), Arg-Glu-Asp-Val (REDV) peptide, heparin, and vascular endothelial growth factor (VEGF) to achieve sustained anti-thrombosis and rapid endothelialization for decellularized and photo-oxidized bovine internal mammary arteries (DP-BIMA). Methods: CS-REDV copolymers were synthesized via a transglutaminase (TGase) catalyzed reaction. CS-REDV-Hep nanoparticles were formed by electrostatic self-assembly and loaded on the DP-BIMA. The quantification of released heparin and vascular endothelial growth factor was detected. Hemolysis rate, platelets adhesion, endothelial cell (EC) adhesion and proliferation, and MTT assay were performed in vitro. The grafts were then tested in a rabbit abdominal aorta interposition model for 3months. The patency rates were calculated and the ECs regeneration was investigated by immunofluorescence staining of CD31, CD144, and eNOS antibodies. Results: The nanoparticle-VEGF system (particle size: 61.8 ± 18.3nm, zeta-potential: +13.2mV, PDI: .108) showed a sustained and controlled release of heparin and VEGF for as long as 1month and exhibited good biocompatibility, a lower affinity for platelets, and a higher affinity for ECs in vitro. The nanoparticle-VEGF immobilized BIMA achieved 100% and 83.3% patency in a rabbit abdominal interposition model during 1 and 3months, respectively, without any thrombogenicity and showed CD31, CD144, eNOS positive cell adhesion as early as 1day. After 3months, CD31, CD144, and eNOS positive cells covered almost the whole luminal surface of the grafts. Conclusion: The results demonstrated that the multifunctional nanoparticle-VEGF system can enhance the anti-thrombosis property and promote rapid endothelialization of small-diameter tissue-engineered vascular grafts. Utilizing nanoparticles to combine different kinds of biomolecules is an appropriate technology to improve the long-term patency of small-diameter tissue-engineered vascular grafts.

VEGF combined with DAPT promotes tissue regeneration and remodeling in vascular grafts.

Previous research on tissue-engineered blood vessels (TEBVs) has mainly focused on the intima or adventitia unilaterally, neglecting the equal importance of both layers. Meanwhile, the efficacy of grafts modified with vascular endothelial growth factor (VEGF) merely has been limited. Here, we developed a small-diameter graft that can gradually release VEGF and γ secretase inhibitor IX (DAPT) to enhance tissue regeneration and remodeling in both the intima and adventitia. In vitro, experiments revealed that the combination of VEGF and DAPT had superior pro-proliferation and pro-migration effects on endothelial cells. In vivo, the sustained release of VEGF and DAPT from the grafts resulted in improved regeneration and remodeling. Specifically, in the intima, faster endothelialization and regeneration of smooth muscle cells led to higher patency rates and better remodeling. In the adventitia, a higher density of neovascularization, M2 macrophages and fibroblasts promoted cellular ingrowth and replacement of the implant with autologous neo-tissue. Furthermore, western blot analysis confirmed that the regenerated ECs were functional and the effect of DAPT was associated with increased expression of vascular endothelial growth factor receptor 2. Our study demonstrated that the sustained release of VEGF and DAPT from the graft can effectively promote tissue regeneration and remodeling in both the intima and adventitia. This development has the potential to significantly accelerate the clinical application of small-diameter TEBVs.

COX-2/PGE2/VEGF signaling promotes ERK-mediated BMSCs osteogenic differentiation under hypoxia by the paracrine action of ECs.

Understanding the crosstalk between endothelial cells (ECs) and bone-marrow mesenchymal stem cells (BMSCs) in response to hypoxic environments and deciphering of the underlying mechanisms are of great relevance for better application of BMSCs in tissue engineering. Here, we demonstrated that hypoxia promoted BMSCs proliferation, colony formation, osteogenic markers expression, mineralization, and extracellular signal-regulated protein kinase (ERK) phosphorylation, and that PD98059 (ERK inhibitor) blocked hypoxia-induced osteogenic differentiation. Hypoxia enhanced ECs migration, cyclooxygenase 2 (COX-2) and integrin αvβ3 expression, and prostaglandin E2 (PGE2), vascular endothelial growth factor (VEGF) secretion. NS398 (selective COX-2 inhibitor) and LM609 (integrin αvβ3 specific inhibitor) impaired the ECs response to hypoxia, and exogenous PGE2 partially reversed the effects of NS398. BMSCs: ECs co-culture under hypoxia upregulated BMSCs osteogenesis and ERK phosphorylation, as well as ECs migration, integrin αvβ3 expression, and PGE2 and VEGF secretion. NS398 (pretreated ECs) lessened PGE2, VEGF concentrations of the co-culture system. NS398-treated ECs and AH6809 (combined EP1/2 antagonist)/L-798106 (selective EP3 antagonist)/L-161982 (selective EP4 antagonist)/SU5416 [VEGF receptor (VEGFR) inhibitor]-treated BMSCs impaired the co-cultured ECs-induced enhancement of BMSCs osteogenic differentiation. In conclusion, hypoxia enhances BMSCs proliferation and ERK-mediated osteogenic differentiation, and augments the COX-2-dependent PGE2 and VEGF release, integrin αvβ3 expression, and migration of ECs. COX-2/PGE2/VEGF signaling is involved in intercellular BMSCs: ECs communication under hypoxia.

Multifunctional microneedle patches with aligned carbon nanotube sheet basement for promoting wound healing

Wound healing has become a health concern and economic burden worldwide for its high incidence and complexity. Attempts to improve wound treatment tend to develop biomedical patches with delicate structure design and functionality. In this paper, we present novel multifunctional microneedle patches with aligned carbon nanotube (CNT) sheet basement for promoting wound healing. Because of the outstanding biocompatibility, biodegradability and non-immunogenicity, hyaluronic acid (HA) is utilized to construct the CNT-integrated microneedle patch with vascular endothelial growth factor (VEGF) encapsulation. It was found that the aligned CNT layer basement could not only impart the composite microneedle patch with orientation morphology, but also endow the patch with controllable release performance, due to its photo-thermal or electro-thermal conversion capacity. Based on these features, we have demonstrated that the highly ordered microstructure of CNT could effectively induce the orientation of fibroblasts; while VEGF release could facilitate tubular formation of endothelial cells. Thus, the aligned CNT and VEGF-loaded multifunctional patch is beneficial to the wound repair process in a typical animal experiment. These results indicate that the proposed microneedle patch is potentially promising for regeneration medicine.

Functional recovery of myocardial infarction by specific EBP-PR1P peptides bridging injectable cardiac extracellular matrix and vascular endothelial growth factor.

Vascular endothelial growth factor (VEGF) is the most potent angiogenic factor and plays an important role in therapy of myocardial infarction (MI). Currently, how to retain regional concentration and decrease rapid diffusion is critical for its clinical application of VEGF. In recent years, the application of targeting peptides has been developed rapidly and provides new strategies for the sustained release of VEGF. In present study, a bi-functional EBP-PR1P peptide was designed and bridged VEGF to injectable cardiac extracellular matrix (c-ECM). Through EBP-PR1P peptides, VEGF could specifically bind with c-ECM to realize the sustained release, without impacting the bioactivity of VEGF. Then VEGF/EBP-PR1P/c-ECM scaffolds were constructed and administrated into rats with MI. The results showed VEGF/EBP-PR1P/c-ECM could promote angiogenesis, protect cardiomyocytes survival against apoptosis, and improve the recovery of cardiac function. In addition, the mechanism of EBP-PR1P/VEGF was also investigated which canonical downstream of VEGF-Akt signaling pathway was activated. These results showed specific VEGF/EBP-PR1P/c-ECM scaffolds served as promising delivery system for VEGF that facilitated the functional recovery of MI.

Nanoparticles based composite coatings with tunable vascular endothelial growth factor and bone morphogenetic protein-2 release for bone regeneration.

Bone healing is a complex cascade involving precisely coordinated spatiotemporal presentation of multiple growth factors (GFs), including osteogenic and angiogenic GFs, and each stage of bone healing requires varying types and content of GFs. In this study, we fabricated a composite nanocoating with tunable vascular endothelial growth factor (VEGF) and bone morphogenetic protein-2 (BMP-2) that was coated on the surface of a polydopamine (PDA)-decorated tertiary calcium phosphate (TCP) scaffold using VEGF-loaded chitosan/bovine serum albumin nanoparticles (CS/BSA-NPs) and BMP-2-loaded poly-L-lysine/oxidized alginate nanoparticles (PLL/OALG-NPs). It was found that VEGF could be efficiently released to promote vascularization in early bone repair stages due to the rapid biodegradation of CS/BSA-NPs, while bone formation can be promoted by a sustained release of BMP-2 from the slowly degrading PLL/OALG-NPs. The composite coating and TCP scaffold can be conjugated due to the excellent adhesive property of PDA. The composite coating can achieve the rapid release of VEGF and sustained release of BMP-2, which can activate GFs for accelerating bone healing.

Pericytes of Stria Vascularis Are Targets of Cisplatin-Induced Ototoxicity: New Insights into the Molecular Mechanisms Involved in Blood-Labyrinth Barrier Breakdown.

The stria vascularis (SV) contributes to cochlear homeostasis and consists of three layers, one of which contains the blood-labyrinthic barrier (BLB), with a large number of bovine cochlear pericytes (BCPs). Cisplatin is a chemotherapeutic drug that can damage the SV and cause hearing loss. In this study, cell viability, proliferation rate, cytotoxicity and reactive oxygen species production were evaluated. The protein content of phospho-extracellular signal-regulated kinases (ERK) 1/2, total ERK 1/2, phospho-cytosolic phospholipase A2 (cPLA2), total cPLA2 and cyclooxygenase 2 (COX-2) and the release of prostaglandin E2 (PGE2) and vascular endothelial growth factor (VEGF) from BCPs were analyzed. Finally, the protective effect of platelet-derived growth factor (PDGF-BB) on BCPs treated with cisplatin was investigated. Cisplatin reduced viability and proliferation, activated ERK 1/2, cPLA2 and COX-2 expression and increased PGE2 and VEGF release; these effects were reversed by Dexamethasone. The presence of PDGF-BB during the treatment with cisplatin significantly increased the proliferation rate. No studies on cell regeneration in ear tissue evaluated the effect of the PDGF/Dex combination. The aim of this study was to investigate the effects of cisplatin on cochlear pericytes and propose new otoprotective agents aimed at preventing the reduction of their vitality and thus maintaining the BLB structure.

Novel ROS-scavenging hydrogel with enhanced anti-inflammation and angiogenic properties for promoting diabetic wound healing

Accelerating angiogenesis of diabetic wounds is crucial to promoting wound healing. Currently, vascular endothelial growth factor (VEGF), an angiogenesis-related bioactive molecule, is widely used in clinic to enhance wound angiogenesis, but it faces problems of inactivation and low utilization due to harsh microenvironment. Here, we developed a novel reactive oxygen species (ROS)-scavenging hydrogel aimed to polarize macrophages toward an anti-inflammatory phenotype, inducing efficient angiogenesis in diabetic wounds. This composite hydrogel with good biosafety and mechanical properties showed sustainable release of bioactive VEGF. Importantly, it could significantly reduce ROS level and rapidly improve wound microenvironment, which ensured the activity of VEGF in vitro and in vivo and successful healing eventually. At the same time, the composite hydrogel exhibited excellent antibacterial properties. In vivo results confirmed good anti-inflammatory, stimulated vascularization and accelerated wound healing attributed to the novel ROS-scavenging hydrogel, which might serve as a promising wound dressing in diabetic wound healing.

Amfenac together with simvastatin reduces IL‐8 and MCP‐1 and increases VEGF release from ARPE‐19 cells upon inflammatory conditions

AbstractPurpose: Rhegmatogenous retinal detachment (RRD) is a sight‐threatening condition with the separation of the neurosensory retina from the retinal pigment epithelium (RPE). Clinical data have associated statin medication with lower risk of re‐vitrectomy in patients operated for RRD. We have previously shown statins to have anti‐inflammatory properties in lipopolysaccharide (LPS)‐treated RPE cells. The purpose of the present study was to examine whether the nonsteroidal anti‐inflammatory drug (NSAID) amfenac could complement the anti‐inflammatory effect of simvastatin in human RPE cells under pathophysiological conditions induced by IL‐1a. In addition to inflammatory markers, we studied the effects of amfenac and simvastatin on growth factors regulating blood vessel formation.Methods: ARPE‐19 cells were pre‐treated with 0.1 μM amfenac and 5 μM simvastatin for 24 h after which 10 pg/ml of IL‐1α was added for 24 h. Enzyme‐linked immunosorbent assay (ELISA) was used to measure secretion of cytokines (IL‐8, MCP‐1), vascular endothelial growth factor (VEGF) and pigment epithelium derived factor (PEDF). Cytotoxicity was measured using the lactate dehydrogenase (LDH) assay.Results: Amfenac and simvastatin were well tolerated upon inflammatory conditions. Both medicines alone and together reduced IL‐1a‐activated IL‐8 release. Simvastatin significantly reduced also the release of MCP‐1. The effect of amfenac alone was not significant but together with simvastatin, significant reduction in MCP‐1 levels was still reached. IL‐1a with the DMSO solvent increased release of PEDF and VEGF compared to DMSO group. Exposure of IL‐1α‐treated cells to amfenac and simvastatin together increased VEGF levels with no changes in the PEDF.Conclusions: Amfenac and simvastatin showed potential to reduce inflammation in the retina upon retinal detachment. Increased VEGF levels may have neuroprotective effects by improving connections between RPE cells and photoreceptors.

Preparation and Biocompatibility of Core-Shell Microspheres for Sequential, Sustained Release of BMP-2 and VEGF

Bone defect repair remains a challenge in orthopedics. This study describes the development and potential effectiveness of vascular endothelial growth factor (VEGF)/bone morphogenetic protein-2 (BMP-2) shell-core microspheres for promoting bone regeneration. Poly(L-lactic acid)/polylactic-co-glycolic acid (PLLA/PLGA) core-shell microspheres loaded with VEGF and BMP-2 were prepared by a coaxial electrospray technique, and their surface morphology, core-shell distribution, and particle size were examined. Different groups of microspheres were prepared with different placement of the growth factors, and the encapsulation efficiency and in vitro release curves were measured. Additionally, the effects of the different groups of microspheres on the proliferation and differentiation of osteoblasts and vascular endothelial cells were investigated. The prepared microspheres had a core-shell structure with good homogeneity and dispersion, a clear boundary, and a smooth surface. On scanning electron microscopy, the mean diameter of the microspheres was similar for all six preparations (P > 0.05). During in vitro release, growth factor was initially released via a brief burst release from the outer shell of the microsphere followed by a slower sustained release. The release of growth factors from the inner core remained relatively slow and sustained. Sequential release of different growth factors was achieved through the inconsistent release rates from the microsphere shell and inner core. All groups of microspheres showed no cytotoxicity, good biocompatibility, and the ability to promote osteoblast proliferation. The microspheres loaded with BMP-2 also promoted osteoblast differentiation, and VEGF-loaded microspheres promoted the proliferation and differentiation of vascular endothelial cells. The BMP-2 (core)/VEGF (shell) microsphere group best promoted osteoblast differentiation. The microspheres prepared in this study exhibited slow sequential release of BMP-2 and VEGF and showed good biocompatibility along with the ability to promote osteoblast differentiation and vascular endothelial cell proliferation.

Construction and osteogenic effects of 3D-printed porous titanium alloy loaded with VEGF/BMP-2 shell-core microspheres in a sustained-release system.

The repair and reconstruction of bone defects remain a challenge in orthopedics. The present study offers a solution to this problem by developing a vascular endothelial growth factor (VEGF)/bone morphogenetic protein 2 (BMP-2) shell-core microspheres loaded on 3D-printed porous titanium alloy via gelatin coating to prepare a titanium-alloy microsphere scaffold release system. The composite scaffold was characterized via scanning electron microscope (SEM) and energy disperse spectroscopy (EDS), and the effect of the composite scaffold on the adhesion, proliferation, and differentiation of osteoblasts were determined in vitro. Furthermore, a rabbit femoral defect model was established to verify the effect of the composite scaffold on osteogenesis and bone formation in vivo. The results demonstrated that the composite scaffold could release VEGF and BMP-2 sequentially. Meanwhile, the composite scaffold significantly promoted osteoblast adhesion, proliferation, and differentiation (p < 0.05) compared to pure titanium alloy scaffolds in vitro. Furthermore, the composite scaffold can exhibit significant osteogenic differentiation (p < 0.05) than gelatin-coated titanium alloy scaffolds. The in vivo X-rays demonstrated that the implanted scaffolds were in a good position, without inflammation and infection. Micro-CT and quantitative results of new bone growth illustrated that the amount of new bone in the composite scaffold is significantly higher than that of the gelatin-coated and pure titanium alloy scaffolds (p < 0.05). Similarly, the fluorescence labeling and V-G staining of hard tissue sections indicated that the bone integration capacity of the composite scaffold was significantly higher than the other two groups (p < 0.05). This research suggests that VEGF/BMP-2 shell-core microspheres loaded on 3D-printed titanium alloy porous scaffold through gelatin hydrogel coating achieved the sequential release of VEGF and BMP-2. Most importantly, the in vitro and in vivo study findings have proven that the system could effectively promote osteogenic differentiation and osseointegration.

Plasma rich in growth factors (PRGF) and leukocyte-platelet rich fibrin (L-PRF): comparative release of growth factors and biological effect on osteoblasts

PurposeTo compare the release of platelet-derived growth factor (PDGF), vascular endothelial growth factor (VEGF), insulin-like growth factor (IGF-I) and interleukin 1β (IL-1β) of plasma rich in growth factors (PRGF) and leucocyte platelet-rich fibrin (L-PRF) and to evaluate their biological implication in osteoblasts.MethodsBlood from 3 healthy volunteers was processed into PRGF, immediate L-PRF (L-PRF 0ʹ) and L-PRF 30 min after collection (L-PRF-30ʹ) and a control group. Growth factors release were analyzed at 7 times by ELISA. Cell proliferation, collagen-I synthesis and alkaline phosphatase activity were assessed in primary cultures of human osteoblasts.ResultsA slower controlled release of IGF-I, VEGF and PDGF was observed in the PRGF group at day 14. A higher synthesis of type I collagen was also quantified in PRGF. L-PRF released significantly higher amounts of IL-1β, that was almost absent in the PRGF.ConclusionsThe addition of leukocytes dramatically increases the secretion of proinflammatory cytokines, which are likely to negatively influence the synthesis of type I collagen and alkaline phosphatase (ALP) by osteoblasts.

Heat Shock Protein 27 Affects Myeloid Cell Activation and Interaction with Prostate Cancer Cells

Heat shock proteins are cytoprotective molecules induced by environmental stresses. The small heat shock protein 27 (Hsp27) is highly expressed under oxidative stress conditions, mediating anti-oxidative effects and blocking apoptosis. Since medical gas plasma treatment subjects cancer cells to a multitude of reactive oxygen species (ROS), inducing apoptosis and immunomodulation, probable effects of Hsp27 should be investigated. To this end, we quantified the extracellular Hsp27 in two prostate cancer cell lines (LNCaP, PC-3) after gas plasma-induced oxidative stress, showing a significantly enhanced release. To investigate immunomodulatory effects, two myeloid cell lines (THP-1 and HL-60) were also exposed to Hsp27. Only negligible effects on viability, intracellular oxidative milieu, and secretion profiles of the myeloid cells were found when cultured alone. Interestingly, prostate cancer-myeloid cell co-cultures showed altered secretion profiles with a significant decrease in vascular endothelial growth factor (VEGF) release. Furthermore, the myeloid surface marker profiles were changed, indicating an enhanced differentiation in co-culture upon Hsp27 treatment. Finally, we investigated morphological changes, proliferation, and interaction with prostate cancer cells, and found significant alterations in the myeloid cells, supporting the tendency to differentiate. Collectively, our results suggest an ambiguous effect of Hsp27 on myeloid cells in the presence of prostate cancer cells which needs to be further investigated.

Effect of Ultrasound Therapy on Expression of Vascular Endothelial Growth Factor, Vascular Endothelial Growth Factor Receptors, CD31 and Functional Recovery After Facial Nerve Injury.

Functional recovery is provided by some neurotrophic factors released from the near vicinity of the injury site. Ultrasound treatment is known to increase neurotrophic factor expression. This study was aimed at determining the effect of ultrasound treatment on the expression of vascular endothelial growth factor (VEGF), its receptors and new vessel formation after facial nerve injury. Sixty-four Wistar rats were divided into four groups: control (group 1), sham (group 2), facial-facial coaptation (group 3), and facial-facial coaptation and ultrasound treatment (group 4). Animals in each group were evaluated on the 14th and 28th days. Immunohistochemical staining and electrophysiological and gene-level evaluations were performed for the expression of VEGF and its receptors. When the results were evaluated, it was determined that VEGF, VEGFR1 (VEGF receptor 1), VEGFR2 (VEGF receptor 2) and CD31 levels were significantly higher in groups 3 and 4 compared with the control and sham groups. The increase in these values was more prominent after 28 d of ultrasound treatment than all groups. Electrophysiological results revealed similar evident functional improvement in group 4 with decreased latency and increased amplitudes compared with group 3. Our findings suggest that ultrasound treatment might promote injured facial nerve regeneration by stimulating release of VEGF and its receptors and may result in functional improvement.