Growth Factors VEGF-A165 and FGF-2 as Multifunctional Biomolecules Governing Cell Adhesion and Proliferation

Vascular endothelial growth factor-A165 (VEGF-A165) and fibroblast growth factor-2 (FGF-2) are currently used for the functionalization of biomaterials designed for tissue engineering. We have developed a new simple method for heterologous expression and purification of VEGF-A165 and FGF-2 in the yeast expression system of Pichia pastoris. The biological activity of the growth factors was assessed in cultures of human and porcine adipose tissue-derived stem cells (ADSCs) and human umbilical vein endothelial cells (HUVECs). When added into the culture medium, VEGF-A165 stimulated proliferation only in HUVECs, while FGF-2 stimulated the proliferation of both cell types. A similar effect was achieved when the growth factors were pre-adsorbed to polystyrene wells. The effect of our recombinant growth factors was slightly lower than that of commercially available factors, which was attributed to the presence of some impurities. The stimulatory effect of the VEGF-A165 on cell adhesion was rather weak, especially in ADSCs. FGF-2 was a potent stimulator of the adhesion of ADSCs but had no to negative effect on the adhesion of HUVECs. In sum, FGF-2 and VEGF-A165 have diverse effects on the behavior of different cell types, which maybe utilized in tissue engineering.

Download Full-text

Control of proliferation of human vascular endothelial cells. Characterization of the response of human umbilical vein endothelial cells to fibroblast growth factor, epidermal growth factor, and thrombin

The Journal of Cell Biology ◽

10.1083/jcb.77.3.774 ◽

1978 ◽

Vol 77 (3) ◽

pp. 774-788 ◽

Cited By ~ 220

Author(s):

D Gospodarowicz ◽

KD Brown ◽

CR Birdwell ◽

BR Zetter

Keyword(s):

Endothelial Cells ◽

Growth Factors ◽

Growth Factor ◽

Vascular Endothelial Cells ◽

Umbilical Vein ◽

Physiological Role ◽

Cell Types ◽

Vascular Endothelial ◽

Human Endothelial Cells ◽

Epidermal Growth

Because the response of human endothelial cells to growth factors and conditioning agents has broad implications for our understanding of wound healing angiogenesis, and human atherogenesis, we have investigated the responses of these cells to the fibroblast (FGF) and epidermal growth factors (EGF), as well as to the protease thrombin, which has been previously shown to potentiate the growth response of other cell types of FGF and EGF. Because the vascular endothelial cells that form the inner lining of blood vessels may be expected to be exposed to high thrombin concentrations after trauma or in pathological states associated with thrombosis, they are of particular interest with respect to the physiological role of this protease in potentiating cell proliferation. Our results indicate that human vascular endothelial cells respond poorly to either FGF or thrombin alone. In contrast, when cells are maintained in the presence of thrombin, their proliferative response to FGF is greatly increased even in cultures seeded at a density as low as 3 cells/mm2. Human vascular endothelial cells also respond to EGF and thrombin, although their rate of proliferation is much slower than when maintained with FGF and thrombin. In contrast, bovine vascular endothelial cells derived from vascular territories as diverse as the bovine heart, aortic arch, and umbilical vein respond maximally to FGF alone and neither respond to nor bind EGF. Furthermore, the response of bovine vascular endothelial cells to FGF was not potentiated by thrombin, indicating that the set of factors controlling the proliferation of vascular endothelial cells could be species-dependent. The requirement of cultured human vascular endothelial cells for thrombin could explain why the human cells, in contrast to bovine endothelial cells, are so difficult to maintain in tissue culture. Our results demonstrate that by using FGF and thrombin one can develop cultures of human vascular endothelial cells capable of being passage repeatedly while maintaining a high mitotic index. The stock cultures used for these studies have been passed weekly with a split ratio of 1 to 10 and are currently in their 30th passage. These cultures are indistinguishable from earlier passages when examined for the presence of Weibel-Palade bodies or Factor VIII antigen. We conclude that the use of FGF and thrombin can prevent the precocious senescence observed in most human endothelial cells cultures previously described.

Download Full-text

Hormonal control of programmed cell death/apoptosis

Acta Endocrinologica ◽

10.1530/eje.0.1380482 ◽

1998 ◽

pp. 482-491 ◽

Cited By ~ 75

Author(s):

W Kiess ◽

B Gallaher

Keyword(s):

Cell Death ◽

Mammary Gland ◽

Growth Factors ◽

Growth Factor ◽

Programmed Cell Death ◽

Cell Types ◽

Hormonal Control ◽

Endocrine Glands ◽

Survival Factors ◽

Endocrine Tissues

Apoptosis or programmed cell death is a physiological form of cell death that occurs in embryonic development and during involution of organs. It is characterized by distinct biochemical and morphological changes such as DNA fragmentation, plasma membrane blebbing and cell volume shrinkage. Many hormones, cytokines and growth factors are known to act as general and/or tissue-specific survival factors preventing the onset of apoptosis. In addition, many hormones and growth factors are also capable of inducing or facilitating programmed cell death under physiological or pathological conditions, or both. Steroid hormones are potent regulators of apoptosis in steroid-dependent cell types and tissues such as the mammary gland, the prostate, the ovary and the testis. Growth factors such as epidermal growth factor, nerve growth factor, platelet-derived growth factor (PDGF) and insulin-like growth factor-I act as survival factors and inhibit apoptosis in a number of cell types such as haematopoietic cells, preovulatory follicles, the mammary gland, phaeochromocytoma cells and neurones. Conversely, apoptosis modulates the functioning and the functional integrity of many endocrine glands and of many cells that are capable of synthesizing and secreting hormones. In addition, exaggeration of the primarily natural process of apoptosis has a key role in the pathogenesis of diseases involving endocrine tissues. Most importantly, in autoimmune diseases such as autoimmune thyroid disease and type 1 diabetes mellitus, new data suggest that the immune system itself may not carry the final act of organ injury: rather, the target cells (i.e. thyrocytes and beta cells of the islets) commit suicide through apoptosis. The understanding of how hormones influence programmed cell death and, conversely, of how apoptosis affects endocrine glands, is central to further design strategies to prevent and treat diseases that affect endocrine tissues. This short review summarizes the available evidence showing where and how hormones control apoptosis and where and how programmed cell death exerts modulating effects upon hormonally active tissues.

Download Full-text

Hydrogel-based microfluidics for vascular tissue engineering

BioNanoMaterials ◽

10.1515/bnm-2015-0026 ◽

2016 ◽

Vol 17 (1-2) ◽

Cited By ~ 6

Author(s):

Anastasia Koroleva ◽

Andrea Deiwick ◽

Alexander Nguyen ◽

Roger Narayan ◽

Anastasia Shpichka ◽

...

Keyword(s):

Tissue Engineering ◽

Vascular Tissue ◽

Umbilical Vein ◽

Gelatin Zymography ◽

Cell Types ◽

Microfluidic System ◽

Vascular Tissue Engineering ◽

Immunofluorescent Staining ◽

Pcr Analysis ◽

Morphological Development

AbstractIn this work, we have explored 3-D co-culture of vasculogenic cells within a synthetically modified fibrin hydrogel. Fibrinogen was covalently linked with PEG-NHS in order to improve its degradability resistance and physico-optical properties. We have studied influences of the degree of protein PEGylation and the concentration of enzyme thrombin used for the gel preparation on cellular responses. Scanning electron microscopy analysis of prepared gels revealed that the degree of PEGylation and the concentration of thrombin strongly influenced microstructural characteristics of the protein hydrogel. Human umbilical vein endothelial cells (HUVECs) and human adipose-derived stem cells (hASCs), used as vasculogenic co-culture, could grow in 5:1 PEGylated fibrin gels prepared using 1:0.2 protein to thrombin ratio. This gel formulation supported hASCs and HUVECs spreading and the formation of cell extensions and cell-to-cell contacts. Expression of specific ECM proteins and vasculogenic process inherent cellular enzymatic activity were investigated by immunofluorescent staining, gelatin zymography, western blot and RT-PCR analysis. After evaluation of the optimal gel composition and PEGylation ratio, the hydrogel was utilized for investigation of vascular tube formation within a perfusable microfluidic system. The morphological development of this co-culture within a perfused hydrogel over 12 days led to the formation of interconnected HUVEC-hASC network. The demonstrated PEGylated fibrin microfluidic approach can be used for incorporating other cell types, thus representing a unique experimental platform for basic vascular tissue engineering and drug screening applications.

Download Full-text

The Response of Tissue Engineered Cartilage to the Temporal Application of Transforming and Insulin-Like Growth Factors

ASME 2007 Summer Bioengineering Conference ◽

10.1115/sbc2007-176523 ◽

2007 ◽

Author(s):

Christopher J. O’Conor ◽

Kenneth W. Ng ◽

Lindsay E. Kugler ◽

Gerard A. Ateshian ◽

Clark T. Hung

Keyword(s):

Tissue Engineering ◽

Growth Factors ◽

Growth Factor ◽

Transforming Growth Factor ◽

Cartilage Tissue Engineering ◽

Cartilage Tissue ◽

Mechanical Stimuli ◽

Tissue Development ◽

Engineering Research ◽

Engineered Cartilage

Agarose has been used as an experimental scaffold for cartilage tissue engineering research due to its biocompatibility with chondrocytes, support of cartilage tissue development, and ability to transmit mechanical stimuli [1–3]. Tissue engineering studies have demonstrated that the temporal application of transforming growth factor (TGF) β3 for only 2 weeks elicits rapid tissue development that results in mechanical properties approaching native values [4]. However, it is not known whether this response to a 2-week exposure to growth factors is unique to TGF-β3. Therefore, the present study characterizes the response of tissue engineered cartilage to the temporal application of the anabolic growth factors TGF-β1, TGF-β3, and insulin-like growth factor I (IGF-I).

Download Full-text

Growth factor loading on aliphatic polyester scaffolds

RSC Advances ◽

10.1039/d0ra10232f ◽

2021 ◽

Vol 11 (12) ◽

pp. 6735-6747

Author(s):

Hong Shen ◽

Xixue Hu

Keyword(s):

Tissue Engineering ◽

Growth Factors ◽

Growth Factor ◽

Factor Loading ◽

Aliphatic Polyester ◽

P Cells

Cells, scaffolds and growth factors are three elements of tissue engineering. Growth factors precisely controlled by a scaffold will be used in conjunction with the scaffolds and cells to repair and regenerate defect tissue.

Download Full-text

Multiplexed digital quantification of binge-like alcohol-mediated alterations in maternal uterine angiogenic mRNA transcriptome

Physiological Genomics ◽

10.1152/physiolgenomics.00009.2012 ◽

2012 ◽

Vol 44 (11) ◽

pp. 622-628 ◽

Cited By ~ 8

Author(s):

Jayanth Ramadoss ◽

Ronald R. Magness

Keyword(s):

Growth Factors ◽

Growth Factor ◽

Ribosomal Protein ◽

Transforming Growth Factor ◽

Fetal Alcohol Spectrum Disorders ◽

Expression System ◽

Digital Gene Expression ◽

Angiogenic Growth Factors ◽

Platelet Factor ◽

Factor Α

Genomic studies on fetal alcohol spectrum disorders (FASD) have utilized either genome-wide microarrays/bioinformatics or targeted real-time PCR (RT-PCR). We utilized herein for the first time a novel digital approach with high throughput as well as the capability to focus on one physiological system. The aim of the present study was to investigate alcohol-induced alterations in uterine angiogenesis-related mRNA abundance using digital mRNA technology. Four biological and three technical replicates of uterine arterial endothelial cells from third-trimester ewes were fluorescence-activated cell sorted, validated, and treated without or with binge-like alcohol. A capture probe covalently bound to an oligonucleotide containing biotin and a color-coded reporter probe were designed for 85 angiogenesis-related genes and analyzed with the Nanostring nCounter system. Twenty genes were downregulated (↓) and two upregulated (↑), including angiogenic growth factors/receptors (↓placental growth factor), adhesion molecules (↓angiopoietin-like-3; ↓collagen-18A1; ↓endoglin), proteases/matrix proteins/inhibitors (↓alanyl aminopeptidase; ↓collagen-4A3; ↓heparanase; ↓plasminogen, ↑plasminogen activator urokinase; ↓platelet factor-4; ↓plexin domain containing-1; ↓tissue inhibitor of metalloproteinases-3), transcription/signaling molecules (↓heart and neural crest derivatives-2; ↓DNA-binding protein inhibitor; ↓NOTCH-4; ↓ribosomal protein-L13a1; ↓ribosomal protein large-P1), cytokines/chemokines (↓interleukin-1B), and miscellaneous growth factors (↓leptin; ↓platelet-derived growth factor-α); ↓transforming growth factor (TGF-α; ↑TGF-β receptor-1). These novel data show significant detrimental alcohol effects on genes controlling angiogenesis supporting a mechanistic role for abnormal uteroplacental vascular development in FASD. The tripartite digital gene expression system is therefore a valuable tool to answer many additional questions about FASD from both mechanistic as well as ameliorative perspectives.

Download Full-text

Engineering Craniofacial Structures: Facing the Challenge

Journal of Dental Research ◽

10.1177/0022034509349926 ◽

2009 ◽

Vol 88 (12) ◽

pp. 1077-1091 ◽

Cited By ~ 60

Author(s):

S.H. Zaky ◽

R. Cancedda

Keyword(s):

Tissue Engineering ◽

Growth Factors ◽

Progenitor Cells ◽

Cell Types ◽

Future Treatment ◽

Current Review ◽

Regenerative Ability ◽

Selection Of ◽

Irreparable Damage

The human innate regenerative ability is known to be limited by the intensity of the insult together with the availability of progenitor cells, which may cause certain irreparable damage. It is only recently that the paradigm of tissue engineering found its way to the treatment of irreversibly affected body structures with the challenge of reconstructing the lost part. In the current review, we underline recent trials that target engineering of human craniofacial structures, mainly bone, cartilage, and teeth. We analyze the applied engineering strategies relative to the selection of cell types to lay down a specific targeted tissue, together with their association with an escorting scaffold for a particular engineered site, and discuss their necessity to be sustained by growth factors. Challenges and expectations for facial skeletal engineering are discussed in the context of future treatment.

Download Full-text

Current insights on use of growth factors as therapy for Intervertebral Disc Degeneration

BioMolecular Concepts ◽

10.1515/bmc-2018-0003 ◽

2018 ◽

Vol 9 (1) ◽

pp. 43-52 ◽

Cited By ~ 16

Author(s):

Justin C. Kennon ◽

Mohamed E. Awad ◽

Norman Chutkan ◽

John DeVine ◽

Sadanand Fulzele

Keyword(s):

Tissue Engineering ◽

Low Back Pain ◽

Back Pain ◽

Growth Factors ◽

Growth Factor ◽

Intervertebral Disc ◽

Transforming Growth Factor ◽

Low Back

AbstractChronic low back pain is a critical health problem and a leading cause of disability in aging populations. A major cause of low back pain is considered to be the degeneration of the intervertebral disc (IVD). Recent advances in therapeutics, particularly cell and tissue engineering, offer potential methods for inhibiting or reversing IVD degeneration, which have previously been impossible. The use of growth factors is under serious consideration as a potential therapy to enhance IVD tissue regeneration. We reviewed the role of chosen prototypical growth factors and growth factor combinations that have the capacity to improve IVD restoration. A number of growth factors have demonstrated potential to modulate the anabolic and anticatabolic effects in both in vitro and animal studies of IVD tissue engineering. Members of the transforming growth factor-β superfamily, IGF-1, GDF-5, BMP-2, BMP-7, and platelet-derived growth factor have all been investigated as possible therapeutic options for IVD regeneration. The role of growth factors in IVD tissue engineering appears promising; however, further extensive research is needed at both basic science and clinical levels before its application is appropriate for clinical use.

Download Full-text

Choline Phospholipid Metabolites of Human Vascular Endothelial Cells Altered by Cyclooxygenase Inhibition, Growth Factor Depletion, and Paracrine Factors Secreted by Cancer Cells

Molecular Imaging ◽

10.1162/15353500200303127 ◽

2003 ◽

Vol 2 (2) ◽

pp. 153535002003031

Author(s):

Noriko Mori ◽

Kshama Natarajan ◽

V. P. Chacko ◽

Dmitri Artemov ◽

Zaver M. Bhujwalla

Keyword(s):

Endothelial Cells ◽

Growth Factors ◽

Growth Factor ◽

Cancer Cells ◽

Umbilical Vein ◽

Phospholipid Metabolism ◽

Paracrine Factors ◽

Inflammatory Agent ◽

Choline Phospholipid ◽

Anti Inflammatory

Magnetic resonance studies have previously shown that solid tumors and cancer cells in culture typically exhibit high phosphocholine and total choline. Treatment of cancer cells with the anti-inflammatory agent, indomethacin (INDO), reverted the phenotype of choline phospholipid metabolites in cancer cells towards a less malignant phenotype. Since endothelial cells form a key component of tumor vasculature, in this study, we used MR spectroscopy to characterize the phenotype of choline phospholipid metabolites in human umbilical vein endothelial cells (HUVECs). We determined the effect of growth factors, the anti-inflammatory agent INDO, and conditioned media obtained from a malignant cell line, on choline phospholipid metabolites. Growth factor depletion or treatment with INDO induced similar changes in the choline phospholipid metabolites of HUVECs. Treatment with conditioned medium obtained from MDA-MB-231 cancer cells induced changes similar to the presence of growth factor supplements. These results suggest that cancer cells secrete growth factors and/or other molecules that influence the choline phospholipid metabolism of HUVECs. The ability of INDO to alter choline phospholipid metabolism in the presence of growth factor supplements suggests that the inflammatory response pathways of HUVECs may play a role in cancer cell-HUVEC interaction and in the response of HUVECs to growth factors.

Download Full-text

Growth factor- and heparin-dependent regulation of constitutive and agonist-mediated human endothelial barrier function

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00185.2006 ◽

2006 ◽

Vol 291 (5) ◽

pp. H2126-H2135 ◽

Cited By ~ 3

Author(s):

Alan B. Moy ◽

Ken Blackwell ◽

Mack H. Wu ◽

Harris J. Granger

Keyword(s):

Growth Factors ◽

Growth Factor ◽

Numerical Model ◽

Barrier Function ◽

Umbilical Vein ◽

Membrane Capacitance ◽

Endothelial Barrier ◽

Endothelial Barrier Function ◽

Barrier Dysfunction ◽

Umbilical Vein Endothelial Cells

We report functional differences in constitutive and agonist-mediated endothelial barrier function between cultured primary and Clonetics human umbilical vein endothelial cells (pHUVEC and cHUVEC) grown in soluble growth factors and heparin. Basal transendothelial resistance (TER) was much lower in pHUVEC than in cHUVEC grown in medium supplemented with growth factors, such as basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), and human epithelial growth factor (EGF), and heparin. On the basis of a numerical model of TER, the increased basal TER in cHUVEC was due to effects on cell-matrix adhesion and membrane capacitance. Heparin and bFGF increased constitutive TER in cultured pHUVEC, and heparin mediated additional increases in constitutive TER in pHUVEC supplemented with bFGF. EGF attenuated bFGF-mediated increases in TER. On the basis of the numerical model, in contrast to cHUVEC, heparin and bFGF augmented TER through effects on cell-cell adhesion and membrane capacitance in pHUVEC. Thrombin mediated quantitatively greater amplitude and a more sustained decline in TER in cultured cHUVEC than pHUVEC. Thrombin-mediated barrier dysfunction was attenuated in pHUVEC conditioned in EGF in the presence or absence of heparin. Thrombin-mediated barrier dysfunction was also attenuated when monolayers were exposed to low concentrations of heparin and further attenuated in the presence of bFGF. cAMP stimulation mediated differential attenuation of thrombin-mediated barrier dysfunction between pHUVEC and cHUVEC. VEGF displayed differential effects in TER in serum-free medium. Taken together, these data demonstrate marked differential regulation of constitutive and agonist-mediated endothelial barrier function in response to mitogens and heparin stimulation.

Download Full-text