scholarly journals Healing and Angiogenic Properties of Collagen/Chitosan Scaffolds Enriched with Hyperstable FGF2-STAB® Protein: In Vitro, Ex Ovo and In Vivo Comprehensive Evaluation

Biomedicines ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 590
Author(s):  
Lucy Vojtová ◽  
Veronika Pavliňáková ◽  
Johana Muchová ◽  
Katarína Kacvinská ◽  
Jana Brtníková ◽  
...  
Keyword(s):  
Wound Healing ◽  
Comprehensive Evaluation ◽  
Chorioallantoic Membrane ◽  
Histological Evaluation ◽  
Skin Defect ◽  
Angiogenic Potential ◽  
Freeze Dried ◽  
Chitosan Scaffolds

Wound healing is a process regulated by a complex interaction of multiple growth factors including fibroblast growth factor 2 (FGF2). Although FGF2 appears in several tissue engineered studies, its applications are limited due to its low stability both in vitro and in vivo. Here, this shortcoming is overcome by a unique nine-point mutant of the low molecular weight isoform FGF2 retaining full biological activity even after twenty days at 37 °C. Crosslinked freeze-dried 3D porous collagen/chitosan scaffolds enriched with this hyper stable recombinant human protein named FGF2-STAB® were tested for in vitro biocompatibility and cytotoxicity using murine 3T3-A31 fibroblasts, for angiogenic potential using an ex ovo chick chorioallantoic membrane assay and for wound healing in vivo with 3-month old white New Zealand rabbits. Metabolic activity assays indicated the positive effect of FGF2-STAB® already at very low concentrations (0.01 µg/mL). The angiogenic properties examined ex ovo showed enhanced vascularization of the tested scaffolds. Histological evaluation and gene expression analysis by RT-qPCR proved newly formed granulation tissue at the place of a previous skin defect without significant inflammation infiltration in vivo. This work highlights the safety and biocompatibility of newly developed crosslinked collagen/chitosan scaffolds involving FGF2-STAB® protein. Moreover, these sponges could be used as scaffolds for growing cells for dermis replacement, where neovascularization is a crucial parameter for successful skin regeneration.

scholarly journals In VitroEvaluation of Scaffolds for the Delivery of Mesenchymal Stem Cells to Wounds

2015 ◽  
Vol 2015 ◽  
pp. 1-14 ◽  
Author(s):  
Elizabeth A. Wahl ◽  
Fernando A. Fierro ◽  
Thomas R. Peavy ◽  
Ursula Hopfner ◽  
Julian F. Dye ◽  
...  
Keyword(s):  
Stem Cells ◽  
Wound Healing ◽  
Mesenchymal Stem Cells ◽  
Three Dimensional ◽  
Chorioallantoic Membrane ◽  
Angiogenic Potential ◽  
Chick Chorioallantoic Membrane ◽  
Bovine Collagen

Mesenchymal stem cells (MSCs) have been shown to improve tissue regeneration in several preclinical and clinical trials. These cells have been used in combination with three-dimensional scaffolds as a promising approach in the field of regenerative medicine. We compare the behavior of human adipose-derived MSCs (AdMSCs) on four different biomaterials that are awaiting or have already received FDA approval to determine a suitable regenerative scaffold for delivering these cells to dermal wounds and increasing healing potential. AdMSCs were isolated, characterized, and seeded onto scaffolds based on chitosan, fibrin, bovine collagen, and decellularized porcine dermis.In vitroresults demonstrated that the scaffolds strongly influence key parameters, such as seeding efficiency, cellular distribution, attachment, survival, metabolic activity, and paracrine release. Chick chorioallantoic membrane assays revealed that the scaffold composition similarly influences the angiogenic potential of AdMSCsin vivo. The wound healing potential of scaffolds increases by means of a synergistic relationship between AdMSCs and biomaterial resulting in the release of proangiogenic and cytokine factors, which is currently lacking when a scaffold alone is utilized. Furthermore, the methods used herein can be utilized to test other scaffold materials to increase their wound healing potential with AdMSCs.

scholarly journals Mutual influence of selenium nanoparticles and FGF2-STAB® on biocompatible properties of collagen/chitosan 3D scaffolds: in vitro and ex ovo evaluation

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Johana Muchová ◽  
Vanessa Hearnden ◽  
Lenka Michlovská ◽  
Lucie Vištejnová ◽  
Anna Zavaďáková ◽  
...  
Keyword(s):  
Antibacterial Activity ◽  
Metabolic Activity ◽  
Mutual Influence ◽  
Chorioallantoic Membrane ◽  
Antagonistic Effect ◽  
Dermal Fibroblasts ◽  
Selenium Nanoparticles ◽  
Freeze Dried ◽  
Chitosan Scaffolds

AbstractIn a biological system, nanoparticles (NPs) may interact with biomolecules. Specifically, the adsorption of proteins on the nanoparticle surface may influence both the nanoparticles' and proteins' overall bio-reactivity. Nevertheless, our knowledge of the biocompatibility and risk of exposure to nanomaterials is limited. Here, in vitro and ex ovo biocompatibility of naturally based crosslinked freeze-dried 3D porous collagen/chitosan scaffolds, modified with thermostable fibroblast growth factor 2 (FGF2-STAB®), to enhance healing and selenium nanoparticles (SeNPs) to provide antibacterial activity, were evaluated. Biocompatibility and cytotoxicity were tested in vitro using normal human dermal fibroblasts (NHDF) with scaffolds and SeNPs and FGF2-STAB® solutions. Metabolic activity assays indicated an antagonistic effect of SeNPs and FGF2-STAB® at high concentrations of SeNPs. The half-maximal inhibitory concentration (IC50) of SeNPs for NHDF was 18.9 µg/ml and IC80 was 5.6 µg/ml. The angiogenic properties of the scaffolds were monitored ex ovo using a chick chorioallantoic membrane (CAM) assay and the cytotoxicity of SeNPs over IC80 value was confirmed. Furthermore, the positive effect of FGF2-STAB® at very low concentrations (0.01 µg/ml) on NHDF metabolic activity was observed. Based on detailed in vitro testing, the optimal concentrations of additives in the scaffolds were determined, specifically 1 µg/ml of FGF2-STAB® and 1 µg/ml of SeNPs. The scaffolds were further subjected to antimicrobial tests, where an increase in selenium concentration in the collagen/chitosan scaffolds increased the antibacterial activity. This work highlights the antimicrobial ability and biocompatibility of newly developed crosslinked collagen/chitosan scaffolds involving FGF2-STAB® and SeNPs. Moreover, we suggest that these sponges could be used as scaffolds for growing cells in systems with low mechanical loading in tissue engineering, especially in dermis replacement, where neovascularization is a crucial parameter for successful skin regeneration. Due to their antimicrobial properties, these scaffolds are also highly promising for tissue replacement requiring the prevention of infection.

scholarly journals A Novel Gold Calreticulin Nanocomposite Based on Chitosan for Wound Healing in a Diabetic Mice Model

Nanomaterials ◽  
2019 ◽  
Vol 9 (1) ◽  
pp. 75 ◽  
Author(s):  
Sara Paola Hernández Martínez ◽  
Teodoro Iván Rivera González ◽  
Moisés Armides Franco Molina ◽  
Juan José Bollain y Goytia ◽  
Juan José Martínez Sanmiguel ◽  
...  
Keyword(s):  
Wound Healing ◽  
Topical Administration ◽  
Nuclear Antigen ◽  
Histological Evaluation ◽  
Mice Model ◽  
Fibroblast Migration ◽  
And Migration ◽  
Proliferating Cell

The development of new nanomaterials to promote wound healing is rising, because of their topical administration and easy functionalization with molecules that can improve and accelerate the process of healing. A nanocomposite of gold nanoparticles (AuNPs) functionalized with calreticulin was synthetized and evaluated. The ability of the nanocomposite to promote proliferation and migration was determined in vitro, and in vivo wound healing was evaluated using a mice model of diabetes established with streptozotocin (STZ). In vitro, the nanocomposite not affect the cell viability and the expression of proliferating cell nuclear antigen (PCNA). Moreover, the nanocomposite promotes the clonogenicity of keratinocytes, endothelial cells, and fibroblasts, and accelerates fibroblast migration. In vivo, mice treated with the nanocomposite presented significantly faster wound healing. The histological evaluation showed re-epithelization and the formation of granular tissue, as well as an increase of collagen deposition. Therefore, these results confirm the utility of AuNPs–calreticulin nanocomposites as potential treatment for wound healing of diabetic ulcers.

Human Erythropoietin Induces a Pro-Angiogenic Phenotype in Cultured Endothelial Cells and Stimulates Neovascularization In Vivo

Blood ◽  
1999 ◽  
Vol 93 (8) ◽  
pp. 2627-2636 ◽  
Author(s):  
Domenico Ribatti ◽  
Marco Presta ◽  
Angelo Vacca ◽  
Roberto Ria ◽  
Roberta Giuliani ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Chorioallantoic Membrane ◽  
Janus Kinase ◽  
Angiogenic Factor ◽  
Matrix Metalloproteinase 2 ◽  
Angiogenic Potential ◽  
Angiogenic Response

Abstract Hematopoietic and endothelial cell lineages share common progenitors. Accordingly, cytokines formerly thought to be specific for the hematopoietic system have been shown to affect several functions in endothelial cells, including angiogenesis. In this study, we investigated the angiogenic potential of erythropoietin (Epo), the main hormone regulating proliferation, differentiation, and survival of erythroid cells. Epo receptors (EpoRs) have been identified in the human EA.hy926 endothelial cell line by Western blot analysis. Also, recombinant human Epo (rHuEpo) stimulates Janus Kinase-2 (JAK-2) phosphorylation, cell proliferation, and matrix metalloproteinase-2 (MMP-2) production in EA.hy926 cells and significantly enhances their differentiation into vascular structures when seeded on Matrigel. In vivo, rHuEpo induces a potent angiogenic response in the chick embryo chorioallantoic membrane (CAM). Accordingly, endothelial cells of the CAM vasculature express EpoRs, as shown by immunostaining with an anti-EpoR antibody. The angiogenic response of CAM blood vessels to rHuEpo was comparable to that elicited by the prototypic angiogenic cytokine basic fibroblast growth factor (FGF2), it occurred in the absence of a significant mononuclear cell infiltrate, and it was not mimicked by endothelin-1 (ET-1) treatment. Taken together, these data demonstrate the ability of Epo to interact directly with endothelial cells and to elicit an angiogenic response in vitro and in vivo and thus act as a bona fide direct angiogenic factor.

scholarly journals Spheroid Coculture of Human Gingiva-Derived Progenitor Cells With Endothelial Cells in Modified Platelet Lysate Hydrogels

2021 ◽  
Vol 9 ◽  
Author(s):  
Siddharth Shanbhag ◽  
Ahmad Rashad ◽  
Ellen Helgeland Nymark ◽  
Salwa Suliman ◽  
Catharina de Lange Davies ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Progenitor Cells ◽  
Umbilical Vein ◽  
Chorioallantoic Membrane ◽  
Platelet Lysate ◽  
Angiogenic Potential ◽  
Hydrogel Matrix ◽  
Free Environment

Cell coculture strategies can promote angiogenesis within tissue engineering constructs. This study aimed to test the angiogenic potential of human umbilical vein endothelial cells (HUVEC) cocultured with gingiva-derived progenitor cells (GPC) as spheroids in a xeno-free environment. Human platelet lysate (HPL) was used as a cell culture supplement and as a hydrogel matrix (HPLG) for spheroid encapsulation. HUVEC and HUVEC + GPC (1:1 or 5:1) spheroids were encapsulated in various HPLG formulations. Angiogenesis was assessed via in vitro sprouting and in vivo chick chorioallantoic membrane (CAM) assays. HUVEC revealed characteristic in vitro sprouting in HPL/HPLG and this was significantly enhanced in cocultures with GPC (p < 0.05). A trend for greater sprouting was observed in 5:1 vs 1:1 HUVEC + GPC spheroids and in certain HPLG formulations (p > 0.05). Both HUVEC and HUVEC + GPC spheroids in HPLG revealed abundant and comparable neoangiogenesis in the CAM assay (p > 0.05). Spheroid coculture of HUVEC + GPC in HPLG represents a promising strategy to promote angiogenesis.

scholarly journals Atropa Belladonna L. Water Extract: Modulator of Extracellular Matrix Formation in Vitro and in Vivo

2012 ◽  
pp. 241-250 ◽  
Author(s):  
P. GÁL ◽  
T. VASILENKO ◽  
I. KOVÁČ ◽  
M. KOSTELNÍKOVÁ ◽  
J. JAKUBČO ◽  
...  
Keyword(s):  
Wound Healing ◽  
Skin Wound ◽  
Dermal Fibroblasts ◽  
Histological Evaluation ◽  
Control Group ◽  
Atropa Belladonna ◽  
Skin Wound Healing ◽  
Collagen Iii

Previously, we found that treatment of cutaneous wounds with Atropa belladonna L. (AB) revealed shortened process of acute inflammation as well as increased tensile strength and collagen deposition in healing skin wounds (Gál et al. 2009). To better understand AB effect on skin wound healing male Sprague-Dawley rats were submitted to one round full thickness skin wound on the back. In two experimental groups two different concentrations of AB extract were daily applied whereas the control group remained untreated. For histological evaluation samples were removed on day 21 after surgery and stained for wide spectrum cytokeratin, collagen III, fibronectin, galectin-1, and vimentin. In addition, in the in vitro study different concentration of AB extract were used to evaluate differences in HaCaT keratinocytes proliferation and differentiation by detection of Ki67 and keratin-19 expressions. Furthermore, to assess ECM formation of human dermal fibroblasts on the in vitro level fibronectin and galectin-1 were visualized. Our study showed that AB induces fibronectin and galectin-1 rich ECM formation in vitro and in vivo. In addition, the proliferation of keratinocytes was also increased. In conclusion, AB is an effective modulator of skin wound healing. Nevertheless, further research is needed to find optimal therapeutic concentration and exact underlying mechanism of action.

scholarly journals RELEVANCE AND PERSPECTIVES OF EXPERIMENTAL WOUND MODELS IN WOUND HEALING RESEARCH

2017 ◽  
Vol 10 (7) ◽  
pp. 57 ◽  
Author(s):  
Santram Lodhi ◽  
Gautam P Vadnere
Keyword(s):  
Wound Healing ◽  
Search Engines ◽  
Wound Repair ◽  
Healing Process ◽  
Chorioallantoic Membrane ◽  
Tube Formation ◽  
Wound Healing Process ◽  
Burn Wound

The wound healing process consists of four highly integrated and overlapping phases: Hemostasis, inflammation, proliferation, and tissue remodeling. These phases and their biophysiological functions must occur in the proper sequence, at a specific time and continue for a specific duration at an optimal intensity. There are many factors that can affect wound healing which interferes with one or more phases in this process, thus causing improper or impaired tissue repair. This review was aimed to collect data and made a critical analysis. This will provide concise information regarding different models and parameters used for wound healing study. The data related to different wound models are collected using popular search engines as well as relevant science search engines and database including Google Scholar, Science Direct, and PubMed. A new drug substance can be evaluated for wound healing activity using different in vitro models such as cell culture, chick chorioallantoic membrane model, tube formation on metrigel and capillary growth model. The in vivo wound models such as incision, excision, dead space, burn wound, ischemic wound, and diabetic wound models are frequently used. Each model has specific importance. The limitations and advantages of each are described in this review. Although animal wound repair is an imperfect reflection of human wound healing and its clinical challenges, these models can be fundamental tools for the development of new approaches to rational wound therapy. 

Human Erythropoietin Induces a Pro-Angiogenic Phenotype in Cultured Endothelial Cells and Stimulates Neovascularization In Vivo

Blood ◽  
1999 ◽  
Vol 93 (8) ◽  
pp. 2627-2636 ◽  
Author(s):  
Domenico Ribatti ◽  
Marco Presta ◽  
Angelo Vacca ◽  
Roberto Ria ◽  
Roberta Giuliani ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Chorioallantoic Membrane ◽  
Janus Kinase ◽  
Angiogenic Factor ◽  
Matrix Metalloproteinase 2 ◽  
Angiogenic Potential ◽  
Angiogenic Response

Hematopoietic and endothelial cell lineages share common progenitors. Accordingly, cytokines formerly thought to be specific for the hematopoietic system have been shown to affect several functions in endothelial cells, including angiogenesis. In this study, we investigated the angiogenic potential of erythropoietin (Epo), the main hormone regulating proliferation, differentiation, and survival of erythroid cells. Epo receptors (EpoRs) have been identified in the human EA.hy926 endothelial cell line by Western blot analysis. Also, recombinant human Epo (rHuEpo) stimulates Janus Kinase-2 (JAK-2) phosphorylation, cell proliferation, and matrix metalloproteinase-2 (MMP-2) production in EA.hy926 cells and significantly enhances their differentiation into vascular structures when seeded on Matrigel. In vivo, rHuEpo induces a potent angiogenic response in the chick embryo chorioallantoic membrane (CAM). Accordingly, endothelial cells of the CAM vasculature express EpoRs, as shown by immunostaining with an anti-EpoR antibody. The angiogenic response of CAM blood vessels to rHuEpo was comparable to that elicited by the prototypic angiogenic cytokine basic fibroblast growth factor (FGF2), it occurred in the absence of a significant mononuclear cell infiltrate, and it was not mimicked by endothelin-1 (ET-1) treatment. Taken together, these data demonstrate the ability of Epo to interact directly with endothelial cells and to elicit an angiogenic response in vitro and in vivo and thus act as a bona fide direct angiogenic factor.

scholarly journals Discovery and Characterization of a High-Affinity Small Peptide Ligand, H1, Targeting FGFR2IIIc for Skin Wound Healing

2018 ◽  
Vol 49 (3) ◽  
pp. 1074-1089 ◽  
Author(s):  
Ying Zhao ◽  
Qiang Wang ◽  
Yuan Jin ◽  
Yadan Li ◽  
Changjun Nie ◽  
...  
Keyword(s):  
Wound Healing ◽  
Growth Factor ◽  
Wound Repair ◽  
Chorioallantoic Membrane ◽  
Skin Wound ◽  
Small Peptide ◽  
Skin Wound Healing ◽  
Wound Model

Background/Aims: How to aid recovery from severe skin injuries, such as burns, chronic or radiation ulcers, and trauma, is a critical clinical problem. Current treatment methods remain limited, and the discovery of ideal wound-healing therapeutics has been a focus of research. Functional recombinant proteins such as basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) have been developed for skin repair, however, some disadvantages in their use remain. This study reports the discovery of a novel small peptide targeting fibroblast growth factor receptor 2 IIIc (FGFR2IIIc) as a potential candidate for skin wound healing. Methods: A phage-displayed peptide library was used for biopanning FGFR2IIIc-targeting small peptides. The selected small peptides binding to FGFR2IIIc were qualitatively evaluated by an enzyme-linked immunosorbent assay. Their biological function was detected by a cell proliferation assay. Among them, an optimized small peptide named H1 was selected for further study. The affinity of the H1 peptide and FGFR2IIIc was determined by an isothermal titration calorimetry device. The ability of theH1 peptide to promote skin wound repair was investigated using an endothelial cell tube formation assay and wound healing scratch assay in vitro. Subsequently, the H1 peptide was assessed using a rat skin full-thickness wound model and chorioallantoic membrane (CAM) assays in vivo. To explore its molecular mechanisms, RNA-Seq, quantitative real-time PCR, and western blot assays were performed. Computer molecular simulations were also conducted to analyze the binding model. Results: We identified a novel FGFR2IIIc-targeting small peptide, called H1, with 7 amino acid residues using phage display. H1 had high binding affinity with FGFR2IIIc. The H1 peptide promoted the proliferation and motility of fibroblasts and vascular endothelial cells in vitro. In addition, the H1 peptide enhanced angiogenesis in the chick chorioallantoic membrane and accelerated wound healing in a rat full-thickness wound model in vivo. The H1 peptide activated both the PI3K-AKT and MAPK-ERK1/2 pathways and simultaneously increased the secretion of vascular endothelial growth factor. Computer analysis demonstrated that the model of H1 peptide binding to FGFR2IIIc was similar to that of FGF2 and FGFR2IIIc. Conclusion: The H1 peptide has a high affinity for FGFR2IIIc and shows potential as a wound healing agent. As a substitute for bFGF, it could be developed into a novel therapeutic candidate for skin wound repair in the future.

Topical application of the phospholipid growth factor lysophosphatidic acid promotes wound healing in vivo

2001 ◽  
Vol 280 (2) ◽  
pp. R466-R472 ◽  
Author(s):  
Louisa Balazs ◽  
Juraj Okolicany ◽  
Mike Ferrebee ◽  
Betsy Tolley ◽  
Gabor Tigyi
Keyword(s):  
Wound Healing ◽  
Lysophosphatidic Acid ◽  
Histological Evaluation ◽  
Lipid Mediator ◽  
Proliferating Cells ◽  
Capillary Endothelial Cells ◽  
Keloid Formation ◽  
Cytological Features

The lipid mediator lysophosphatidic acid (LPA) regulates cell proliferation and enhances cell motility in vitro, both of which are important events in wound healing. To evaluate the effects of LPA in vivo, it was applied to a full-thickness wound of rat skin. LPA in micromolar concentrations, or solvent, was applied daily. Animals were killed at 1, 3, 6, and 9 days after wounding and processed for histological evaluation, including hematoxylin-eosin staining and histochemical markers for macrophage-histiocytes, proliferating cells, and capillary endothelial cells. LPA treatment accelerated wound closing and increased neoepithelial thickness. Cytological evaluation showed no evidence for a secondary inflammation-mediated injury, infection, or increased keloid formation. Whereas LPA caused only a modest dose-dependent increase in proliferating cells, a marked increase in the immigration of histiocyte-macrophage cells was observed as early as day 1. The peaks of several cytological features and immunohistological markers preceded those of the untreated side. Our data suggest that exogenously applied LPA in this model promotes healing and that macrophage-histiocytes are the primary LPA-responsive cells in vivo.

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