Isolation of fresh endothelial cells from porcine heart for cardiovascular studies: a new fast protocol suitable for genomic, transcriptomic and cell biology studies.
BACKGROUND
Endothelial cells (ECs) play a key role in tissue homeostasis, in several pathological conditions, and specifically in the control of vascular functions. ECs are frequently used as in vitro model systems for cardiovascular studies and vascular biology. The porcine model is commonly used in human clinical cardiovascular studies. Currently, however, there is no robust protocol for the isolation of porcine heart ECs. We have developed a fast isolation protocol, which is cost effective, takes only 1-2 h, and produces EC purity of over 97%. This protocol is optimized for porcine hearts but can be adapted for use with other large animals.


METHODS
Heart is washed by flushing with PBS, whereafter endothelial cells are detached by collagenase incubation and the cells can then be collected immediately after the incubation and plated within an hour after the heart is isolated from a pig.


RESULTS
The swiftness of the protocol limits changes in the phenotype and RNA expression profile of the cells. Cells were identified as ECs with CD31 (PECAM-1) antibody immunostaining. Functionality of ECs were ensured with in vitro angiogenesis assay. The purity of the ECs was verified by using fluorescence assisted cell sorting (FACS) with the CD31 antibody.


CONCLUSION
We developed a new, fast, and cost-effective isolation method for pig heart ECs. Successful isolation of pure ECs is a prerequisite for several cardiovascular and vascular biology studies.
['Hätinen|Olli-Pekka A|OA|', 'Lähteenvuo|Johanna E|JE|', 'Korpela|Henna J|HJ|', 'Pajula|Juho J|JJ|', 'Ylä-Herttuala|Seppo|S|0000-0001-7593-2708']
D000818:Animals D002318:Cardiovascular Diseases D003585:Cell Biology D002469:Cell Separation D002478:Cells, Cultured D042783:Endothelial Cells D005260:Female D023281:Genomics D009206:Myocardium D013552:Swine D059467:Transcriptome
Isolation Endothelial cell Porcine heart Pig Fresh tissue Myocardial