Porcine ovarian cortex-derived putative stem cells can differentiate into endothelial cells in vitro

Abstract

Endothelial cells (ECs), the primary component of the vasculature, play a crucial role in neovascularization. However, the number of endogenous ECs is inadequate for both experimental purposes and clinical applications. Porcine ovarian putative stem cells (poPSCs), although not pluripotent, are characterized by great plasticity. Therefore, this study aimed to investigate whether poPSCs have the potential to differentiate into cells of endothelial lineage. poPSCs were immunomagnetically isolated from postnatal pig ovaries based on the presence of SSEA-4 protein. Expression of mesenchymal stem cells (MSCs) markers after pre-culture, both at the level of mRNA: ITGB1, THY, and ENG and corresponding protein: CD29, CD90, and CD105 were significantly higher compared to the control ovarian cortex cells. To differentiate poPSCs into ECs, inducing medium containing vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), insulin-like growth factor (IGF), epidermal growth factor (EGF), ascorbic acid, and heparin was applied. After 14 days, poPSC differentiation into ECs was confirmed by immunofluorescence staining for vascular endothelial cadherin (VECad) and vascular endothelial growth factor receptor-2 (VEGFR-2). Semi-quantitative WB analysis of these proteins confirmed their high abundance. Additionally, qRT-PCR showed that mRNA expression of corresponding marker genes: CDH5, KDR was significantly higher compared with undifferentiated poPSCs. Finally, EC functional status was confirmed by the migration test that revealed that they were capable of positive chemotaxis, while tube formation assay demonstrated their ability to develop capillary networks. In conclusion, our results provided evidence that poPSCs may constitute the MSC population in the ovary and confirmed that they might be a potential source of ECs for tissue engineering.

Anabolic Steroids-Driven Regulation of Porcine Ovarian Putative Stem Cells Favors the Onset of Their Neoplastic Transformation

“…Among various animal experimental models, pigs share many similarities with humans in the form of organ size, physiology, and functioning [ 31 ]. The limited ethical dilemmas and importantly, successful isolation of putative stem cells from the ovarian cortex (poPSCs) [ 32 , 33 , 34 ], make pigs the valuable experimental model for not only preclinical assessment for stem cell therapy but also for the purposes of somatic cell nuclear transfer (SCNT)-based cloning in mammals. The efforts undertaken to use poPSCs, which have been subjected to extrinsic anabolic steroid-dependent oncogenic transformation (carcinogenesis), as a completely new source of nuclear donor cells (NDCs) for future research focused on SCNT, have not yet been accomplished.…”

Anabolic Steroids Activate the NF-κB Pathway in Porcine Ovarian Putative Stem Cells Independently of the ZIP-9 Receptor

“…Western blot analysis was performed according to a technique developed and modified in our laboratory [ 41 ]. Briefly, after the termination of both poPSC cultures treated and not treated with AASs for 7 or 14 days, they were washed twice with cold PBS.…”

“…Due to the large diversity of ovarian ASCs, they are also called putative stem cells (PSCs) [ 37 , 38 , 39 ]. The results of our previous studies [ 37 , 40 , 41 ] have demonstrated that porcine ovarian putative stem cells (poPSCs) isolated from the ovarian cortex (against the presence of SSEA-4 antigen), though the non-specific expression of selected pluripotency markers are not pluripotent but rather multipotent. We have shown that poPSCs can differentiate into functional neural-like and endothelial cells, poPSCs likely represent just multipotent mesenchymal stem cell (MSCs) populations in the ovary.…”

“…We have shown that poPSCs can differentiate into functional neural-like and endothelial cells, poPSCs likely represent just multipotent mesenchymal stem cell (MSCs) populations in the ovary. This fact is evidenced by the expression of MSC-related markers such as ITGB1 (CD29), THY1 (CD90) and ENG (CD105), which is exhibited in poPSCs [ 41 ]. Additionally, in our latest report, we have proved that poPSCs display the expression of ARs by which AASs, including Ndn and Bdn, can influence them [ 29 ].…”

In focus in HCB

“…Given the need for a source of endothelial cells for tissue engineering and human regenerative medicine, the porcine model, which as mentioned above has been used previously for many xenogeneic procedures (see also for instance Gasek et al 2021), appeared to represent a potential source for these cells. This led Wartalski et al (2021) to extend their recent studies to investigate whether the multipotent high-plasticity poPSCs could also be directed to differentiate into endothelial-lineage cells. They isolated poPSCs from porcine ovarian cortex using an immunomagnetic approach with anti-SSEA-4 conjugated to the magnetic beads.…”