Angiogenic properties of aged adipose derived mesenchymal stem cells after hypoxic conditioning

Abstract

BackgroundMesenchymal stem cells derived from adipose tissue (ADSC) are multipotent stem cells, originated from the vascular-stromal compartment of fat tissue. ADSC are used as an alternative cell source for many different cell therapies, however in ischemic cardiovascular diseases the therapeutic benefit was modest. One of the reasons could be the use of autologous aged ADSC, which recently were found to have impaired functions. We therefore analysed the effects of age on age markers and angiogenic properties of ADSC. Hypoxic conditioning was investigated as a form of angiogenic stimulation.MethodsADSC were harvested from young (1-3 month), adult (12 month) and aged (18-24 month) mice and cultured under normoxic (20%) and hypoxic (1%) conditions for 48 h. Differences in proliferation, apoptosis and telomere length were assessed in addition to angiogenic properties of ADSC.ResultsProliferation potential and telomere length were decreased in aged ADSC compared to young ADSC. Frequency of apoptotic cells was higher in aged ADSC. Gene expression of pro-angiogenic factors including vascular endothelial growth factor (VEGF), placental growth factor (PlGF) and hepatic growth factor (HGF) were down-regulated with age, which could be restored by hypoxia. Transforming growth factor (TGF-β) increased in the old ADSC but was reduced by hypoxia.Expression of anti-angiogenic factors including thrombospondin-1 (TBS1) and plasminogen activator inhibitor-1 (PAI-1) did increase in old ADSC, but could be reduced by hypoxic stimulation. Endostatin (ENDS) was the highest in aged ADSC and was also down-regulated by hypoxia. We noted higher gene expression of proteases system factors like urokinase-type plasminogen activator receptor (uPAR), matrix metalloproteinases (MMP2 and MMP9) and PAI-1 in aged ADSC compared to young ADSC, but they decreased in old ADSC. Tube formation on matrigel was higher in the presence of conditioned medium from young ADSC in comparison to aged ADSC.ConclusionsADSC isolated from older animals show changes, including impaired proliferation and angiogenic stimulation. Angiogenic gene expression can be partially be improved by hypoxic preconditioning, however the effect is age-dependent. This supports the hypothesis that autologous ADSC from aged subjects might have an impaired therapeutic potential.

Adipose-Derived Mesenchymal Stromal Cells From Aged Patients With Coronary Artery Disease Keep Mesenchymal Stromal Cell Properties but Exhibit Characteristics of Aging and Have Impaired Angiogenic Potential

“…In analyzing the expression of factors involved in ECM remodeling and vascular cell migration and invasion, we revealed that mRNA levels of uPA, uPAR, and PAI-1 as well as uPAR surface expression were higher in ADSCs from older patients. These results are consistent with those that we previously obtained with ADSCs from young and old mice [42]. Urokinase, a multidomain protein, not only specifically cleaves plasminogen and converts it into plasmin, particularly causing activation of different MMPs, but also initiates intracellular signaling on binding to its receptor on the cell surface and therefore plays multiple roles in vascular remodeling and angiogenesis [52].…”

“…We have previously shown that ADSCs isolated from old mice (18 months), along with acquiring the properties of aged cells (shorter telomeres, higher rate of apoptotic cells, less proliferative capacity, enhanced oxidative damage), have an impaired ability to stimulate blood vessel growth in in vitro and in vivo models of angiogenesis compared with ADSCs from young animals (1-2 months) [42]. In the current study, we analyzed age-associated changes in human ADSCs obtained from both patients without cardiovascular pathology and patients with CAD as a cohort of the most obvious candidates for cell therapy.…”

PlGF: A Multitasking Cytokine with Disease-Restricted Activity

“…Conversely, PlGF gene or protein transfer in infarcted mice stimulates angiogenesis in the infarct border and vessel enlargement in the remote myocardium, improving vascular perfusion, cardiomyocyte hypertrophy, and cardiac recovery (Luttun et al 2002;Roncal et al 2008). Unlike systemic treatment with Adipose tissue-derived MSC Viglietto et al 1997;Efimenko et al 2011 Cell types are listed with reported information on inducers of PlGF expression where available. AngII, Angiopoietin II; BMP, bone morphogenic protein; HSC, hepatic stellate cell; IL, interleukin; RPE, retinal pigment epithelial; MSC, mesenchymal stem cell; SMC, smooth muscle cell; TAM, tumor-associated macrophage; TGF, transforming growth factor; TNF, tumor necrosis factor.…”

Influence of Donor Age and Passage Number on Angiogenic Activity in Human Adipose-Derived Stem Cell-Conditioned Media

“…Bone marrow MSCs from patients aged over 40 years show increased levels of reactive oxygen species (ROS) and nitric oxide (NO) and impaired stress responses [37]. In mice, aged ASCs also exhibit increases in ROS and NO production and a decrease in the antioxidative defense enzyme glutathione peroxide with significant telomere shortening [26]. Our study results using 10% FBS in DMEM as a growth medium imply that ASCs can be cultured until passage 6 for clinical applications.…”

“…On the other hand, limited information is currently available regarding the angiogenic potential of aged ASCs [26][27][28]. In the setting of ASC-based therapeutic angiogenesis, clarification of how in vivo and in vitro aging of culturedculture flasks.…”