Fibrinogen Concentration Influence on VEGF-Induced Differentiation of Adipose Tissue-Derived Stem Cells into Endothelial Cells

2007 ◽  
Vol 342-343 ◽  
pp. 17-20
Author(s):  
Hyeong In Kim ◽  
Ji Yeon Seo ◽  
Seung Jo Jeung ◽  
Sae Gwang Park ◽  
Young Il Yang
Keyword(s):  
Stem Cells ◽  
Endothelial Cells ◽  
Adipose Tissue ◽  
Network Formation ◽  
Capillary Tube ◽  
Specific Gene ◽  
Fibrinogen Concentration ◽  
And Migration ◽  
Von Willebrand

Fibrin is a natural substrate for growth, adhesion, and migration of mature endothelial cells (ECs) and a candidate coating material in approaches to graft endothelialization. Adipose tissue represents an abundant, practical source of donor tissue for stem cells which may be a useful source for engineering of vascular grafts. However, the optimal substrates that promote differentiation of adipose tissue-derived stem cells (ASCs) into ECs remain to be elucidated. In the present study, we investigated whether fibrin can be used as a substratum to support in vitro ECs differentiation of ASCs and whether fibrinogen concentration can be affect on ECs differentiation of ASCs. For determination of phenotypic characteristics of ASCs used in this experiment, we performed flow cytometry analysis. ASCs were plated on fibrin composed of varying concentrations of fibrinogen and induced into ECs differentiation in presence of VEGF. Before inducing into ECs, ASCs did not express any markers of hematopoietic cells (CD34, CD45), ECs (CD31, CD34), and endothelial progenitor cells (CD34, CD133, Flk-1). The degree of ECs differentiation was determined by capillary network formation, ECs-specific gene expression, and F-actin assembly. During the first 12 h after seeding, cells spread randomly, moved and formed small interconnected clusters. These clusters decreased in size and formed a capillary tube at 48 h. During the further incubation in presence of VEGF for 7 days, ASCs expressed mRNA and protein of von Willebrand factor (vWF). The degree of ECs differentiation of ASCs was consistently decreased as fibrinogen concentration increase. Fibrin may be used as biomatrix to promote differentiation of ASCs into ECs for tissue engineering.

scholarly journals Topography-Mediated Myotube and Endothelial Alignment, Differentiation, and Extracellular Matrix Organization for Skeletal Muscle Engineering

Polymers ◽  
2020 ◽  
Vol 12 (9) ◽  
pp. 1948
Author(s):  
Ana Maria Almonacid Suarez ◽  
Marja G. L. Brinker ◽  
Linda A. Brouwer ◽  
Iris van der Ham ◽  
Martin C. Harmsen ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Network Formation ◽  
Basal Membrane ◽  
Physical Factors ◽  
Cell Alignment ◽  
Human Myotubes ◽  
And Migration

Understanding the response of endothelial cells to aligned myotubes is important to create an appropriate environment for tissue-engineered vascularized skeletal muscle. Part of the native tissue environment is the extracellular matrix (ECM). The ECM is a supportive scaffold for cells and allows cellular processes such as proliferation, differentiation, and migration. Interstitial matrix and basal membrane both comprise proteinaceous and polysaccharide components for strength, architecture, and volume retention. Virtually all cells are anchored to their basal lamina. One of the physical factors that affects cell behavior is topography, which plays an important role on cell alignment. We tested the hypothesis that topography-driven aligned human myotubes promote and support vascular network formation as a prelude to in vitro engineered vascularized skeletal muscle. Therefore, we used a PDMS-based topography substrate to investigate the influence of pre-aligned myotubes on the network formation of microvascular endothelial cells. The aligned myotubes produced a network of collagen fibers and laminin. This network supported early stages of endothelial network formation.

scholarly journals Identification of Angiogenic Cargo in Extracellular Vesicles Secreted from Human Adipose Tissue-Derived Stem Cells and Induction of Angiogenesis In Vitro and In Vivo

Pharmaceutics ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 495
Author(s):  
Prakash Gangadaran ◽  
Ramya Lakshmi Rajendran ◽  
Ji Min Oh ◽  
Eun Jung Oh ◽  
Chae Moon Hong ◽  
...  
Keyword(s):  
Stem Cells ◽  
Endothelial Cells ◽  
Adipose Tissue ◽  
Blood Vessels ◽  
Extracellular Vesicles ◽  
Human Adipose Tissue ◽  
Antibody Array ◽  
Angiogenic Proteins

Angiogenesis is defined as the generation of new blood vessels or the sprouting of endothelial cells from a pre-existing vascular network. Angiogenesis occurs during the growth and development of an organism, the response of organs or tissues to injury, and during cancer development and progression. The majority of studies on stem-cell-derived extracellular vesicles (EVs) have used cell lines, and have primarily focused on well-known solitary proteins. Here, we isolated stem cells from human adipose tissue (ADSCs), and we isolated EVs from them (ADSC-EVs). The ADSC-EVs were characterised and 20 angiogenic proteins were analysed using an angiogenic antibody array. Furthermore, we analysed the ability of ADSC-EVs to induce angiogenesis in vitro and in vivo. ADSC-EVs were positive for CD81 and negative for GM130, calnexin, and cytochrome-C. ADSC-EVs showed typical EV spherical morphology and were ~200 nm in size. ADSC-EVs were found to contain angiogenic proteins as cargo, among which interleukin 8 (IL-8) was the most abundant, followed by chemokine (C-C motif) ligand 2 (CCL2), a tissue inhibitor of metalloproteinases 1 (TIMP-1), TIMP-2, and vascular endothelial growth factor-D (VEGF-D). ADSC-EVs treatment increased the proliferation, migration, total vessel length, total number of junctions, and junction density of endothelial cells in vitro. The results of an in vivo Matrigel plug assay revealed that ADSC-EVs induced more blood vessels in the Matrigel compared with the control. These results demonstrate that ADSC-EVs contain angiogenic proteins as cargo and promote angiogenesis in vitro and in vivo. Therefore, ADSC-EVs have potential for therapeutic use in ischaemia.

scholarly journals Deducing insulin producing cells in goat adipose tissue derived mesenchymal stem cells

2021 ◽  
Author(s):  
Amit Dubey ◽  
Hruda Nanda Malik ◽  
Dinesh Kumar ◽  
Sikander Saini ◽  
Satish Kumar ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Regenerative Medicine ◽  
Basic Mechanism ◽  
Specific Protein ◽  
Specific Gene ◽  
Pancreatic Cells ◽  
Insulin Producing Cells

Abstract Background: Mesenchymal stem cells (MSCs) is a new herald for regenerative medicine for control of incurable diseases in human and animals. Diabetes occurs when the blood glucose is high due to lack of insulin hormone secreted by the pancreatic cells. The global diabetes in 2019 is estimated 463 million people and rising to 578 million by 2030. Methods: Here we differentiated goat adipose tissue derived MSCs into insulin producing cells. To achieve this, the goat MSCs were cultured in serum-free DMEM/F12 medium containing glucose, nicotinamide, activin-A, exendin-4, pentagastrin, retinoic acid and mercaptoethanol for three weeks. The in vitro differentiation ADSCs into insulin-producing cells was confirmed by detecting the pancreatic endoderm specific markers i.e. Igf-1, Sst, Ngn3, Pdx-1, Isl-1, c-Kit, Thy-1, and Glut-2 in differentiating cells. Results: There was a significant increase in insulin specific gene expression with respect to duration of differentiation. Pancreatic insulin-producing cells were further characterized by immunolocalization of Pdx-1, insulin, and Islets-1 specific protein. The release of insulin in response to a glucose challenge was also evaluated. Conclusions: The study provides new opportunities for deciphering the basic mechanism of in vitro genesis of pancreatic cells and basic properties, availability, and abundance of ADSCs render them well-suited for applications in regenerative medicine.

Adverse Influence of Cigarette Smoking on the Endothelium

1993 ◽  
Vol 70 (04) ◽  
pp. 707-711 ◽  
Author(s):  
Andrew D Blann ◽  
Charles N McCollum
Keyword(s):  
Endothelial Cells ◽  
Von Willebrand Factor ◽  
Tobacco Smoke ◽  
Lipid Peroxides ◽  
Short Exposure ◽  
Acute Effects ◽  
Adverse Influence ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryThe effect of smoking on the blood vessel intima was examined by comparing indices of endothelial activity in serum from smokers with that from non-smokers. Serum from smokers contained higher levels of von Willebrand factor (p <0.01), the smoking markers cotinine (p <0.02) and thiocyanate (p <0.01), and was more cytotoxic to endothelial cells in vitro (p <0.02) than serum from non-smokers. The acute effects of smoking two unfiltered medium tar cigarettes was to briefly increase von Willebrand factor (p <0.001) and cytotoxicity of serum to endothelial cells in vitro (p <0.005), but lipid peroxides or thiocyanate were not increased by this short exposure to tobacco smoke. Although there were correlations between von Willebrand factor and smokers consumption of cigarettes (r = 0.28, p <0.02), number of years smoking (r = 0.41, p <0.001) and cotinine (r = 0.45, p <0.01), the tissue culture of endothelial cells with physiological levels of thiocyanate or nicotine suggested that these two smoking markers were not cytotoxic. They are therefore unlikely to be directly responsible for increased von Willebrand factor in the serum of smokers. We suggest that smoking exerts a deleterious influence on the endothelium and that the mechanism is complex.

Epinephrine Induces von Willebrand Factor Release from Cultured Endothelial Cells: Involvement of Cyclic AMP-dependent Signalling in Exocytosis

1997 ◽  
Vol 77 (06) ◽  
pp. 1182-1188 ◽  
Author(s):  
Ulrich M Vischer ◽  
Claes B Wollheinn
Keyword(s):  
Endothelial Cells ◽  
Adenylate Cyclase ◽  
Von Willebrand Factor ◽  
Umbilical Vein ◽  
Free Calcium ◽  
Camp Content ◽  
Von Willebrand ◽  
Willebrand Factor

Summaryvon Willebrand factor (vWf) is released from endothelial cell storage granules after stimulation with thrombin, histamine and several other agents that induce an increase in cytosolic free calcium ([Ca2+]i). In vivo, epinephrine and the vasopressin analog DDAVP increase vWf plasma levels, although they are thought not to induce vWf release from endothelial cells in vitro. Since these agents act via a cAMP-dependent pathway in responsive cells, we examined the role of cAMP in vWf secretion from cultured human umbilical vein endothelial cells. vWf release increased by 50% in response to forskolin, which activates adenylate cyclase. The response to forskolin was much stronger when cAMP degradation was blocked with IBMX, an inhibitor of phosphodiesterases (+200%), whereas IBMX alone had no effect. vWf release could also be induced by the cAMP analogs dibutyryl-cAMP (+40%) and 8-bromo-cAMP (+25%); although their effect was weak, they clearly potentiated the response to thrombin. Epinephrine (together with IBMX) caused a small, dose-dependent increase in vWf release, maximal at 10-6 M (+50%), and also potentiated the response to thrombin. This effect is mediated by adenylate cyclase-coupled β-adrenergic receptors, since it is inhibited by propranolol and mimicked by isoproterenol. In contrast to thrombin, neither forskolin nor epinephrine caused an increase in [Ca2+]j as measured by fura-2 fluorescence. In addition, the effects of forskolin and thrombin were additive, suggesting that they act through distinct signaling pathways. We found a close correlation between cellular cAMP content and vWf release after stimulation with epinephrine and forskolin. These results demonstrate that cAMP-dependent signaling events are involved in the control of exocytosis from endothelial cells (an effect not mediated by an increase in [Ca2+]i) and provide an explanation for epinephrine-induced vWf release.

scholarly journals Comparative analysis of mouse bone marrow and adipose tissue mesenchymal stem cells for critical limb ischemia cell therapy

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Pegah Nammian ◽  
Seyedeh-Leili Asadi-Yousefabad ◽  
Sajad Daneshi ◽  
Mohammad Hasan Sheikhha ◽  
Seyed Mohammad Bagher Tabei ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Cell Therapy ◽  
Femoral Artery ◽  
Critical Limb Ischemia ◽  
Limb Ischemia

Abstract Introduction Critical limb ischemia (CLI) is the most advanced form of peripheral arterial disease (PAD) characterized by ischemic rest pain and non-healing ulcers. Currently, the standard therapy for CLI is the surgical reconstruction and endovascular therapy or limb amputation for patients with no treatment options. Neovasculogenesis induced by mesenchymal stem cells (MSCs) therapy is a promising approach to improve CLI. Owing to their angiogenic and immunomodulatory potential, MSCs are perfect candidates for the treatment of CLI. The purpose of this study was to determine and compare the in vitro and in vivo effects of allogeneic bone marrow mesenchymal stem cells (BM-MSCs) and adipose tissue mesenchymal stem cells (AT-MSCs) on CLI treatment. Methods For the first step, BM-MSCs and AT-MSCs were isolated and characterized for the characteristic MSC phenotypes. Then, femoral artery ligation and total excision of the femoral artery were performed on C57BL/6 mice to create a CLI model. The cells were evaluated for their in vitro and in vivo biological characteristics for CLI cell therapy. In order to determine these characteristics, the following tests were performed: morphology, flow cytometry, differentiation to osteocyte and adipocyte, wound healing assay, and behavioral tests including Tarlov, Ischemia, Modified ischemia, Function and the grade of limb necrosis scores, donor cell survival assay, and histological analysis. Results Our cellular and functional tests indicated that during 28 days after cell transplantation, BM-MSCs had a great effect on endothelial cell migration, muscle restructure, functional improvements, and neovascularization in ischemic tissues compared with AT-MSCs and control groups. Conclusions Allogeneic BM-MSC transplantation resulted in a more effective recovery from critical limb ischemia compared to AT-MSCs transplantation. In fact, BM-MSC transplantation could be considered as a promising therapy for diseases with insufficient angiogenesis including hindlimb ischemia.

scholarly journals Derivation, characterization, and in vitro cell regeneration of canine white adipose tissue-derived mesenchymal stem cells obtained from a mesenteric region

2021 ◽  
Vol 82 (1) ◽  
Author(s):  
Anirban Mandal ◽  
Ajeet Kumar Jha ◽  
Dew Biswas ◽  
Shyamal Kanti Guha
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
White Adipose Tissue ◽  
Stromal Vascular Fraction ◽  
Cell Regeneration ◽  
Wound Healing Assay ◽  
Chain Reaction ◽  
Polymerase Chain

Abstract Background The study was conducted to assess the characterization, differentiation, and in vitro cell regeneration potential of canine mesenteric white adipose tissue-derived mesenchymal stem cells (AD-MSCs). The tissue was harvested through surgical incision and digested with collagenase to obtain a stromal vascular fraction. Mesenchymal stem cells isolated from the stromal vascular fraction were characterized through flow cytometry and reverse transcription-polymerase chain reaction. Assessment of cell viability, in vitro cell regeneration, and cell senescence were carried out through MTT assay, wound healing assay, and β-galactosidase assay, respectively. To ascertain the trilineage differentiation potential, MSCs were stained with alizarin red for osteocytes, alcian blue for chondrocytes, and oil o red for adipocytes. In addition, differentiated cells were characterized through a reverse transcription-polymerase chain reaction. Results We observed the elongated, spindle-shaped, and fibroblast-like appearance of cells after 72 h of initial culture. Flow cytometry results showed positive expression for CD44, CD90, and negative expression for CD45 surface markers. Population doubling time was found 18–24 h for up to the fourth passage and 30±0.5 h for the fifth passage. A wound-healing assay was used to determine cell migration rate which was found 136.9 ± 4.7 μm/h. We observed long-term in vitro cell proliferation resulted in MSC senescence. Furthermore, we also found that the isolated cells were capable of differentiating into osteogenic, chondrogenic, and adipogenic lineages. Conclusions Mesenteric white adipose tissue was found to be a potential source for isolation, characterization, and differentiation of MSCs. This study might be helpful for resolving the problems regarding the paucity of information concerning the basic biology of stem cells. The large-scale use of AD-MSCs might be a remedial measure in regenerative medicine.

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