Isolation and cultivation of aortic endothelial cells from spontaneously hypertensive rat with a modified tissue explant technique.
In the present study, we successfully established a "tissue explant technique" to obtain high yield and purity of endothelial cells from the aorta of hypertensive and normotensive rats (SHR and WKY). Small pieces of aorta were placed on fibronectin precoated petri dishes. The effects of oxygenation in the tissue preparation stage, tilting of the petri dish during the explanting period and timing of the removal of tissue blocks from petri dishes were evaluated. These procedures appeared to be critical for cell survival, tissue adhesion and minimizing of non-endothelial cell contamination. The cultured endothelial cells were characterized by morphological, immunohistochemical and biochemical examination. The cultured cells from both SHR and WKY rats showed similar endothelial cell character, positive immunofluorescence staining for the von Willebrand factor, and uptake of acetylated low-density lipoprotein (DiI-ac-LDL). The secretory function of prostacylcin I2 (PGI2), thromboxane A2 and endothelin of cultured endothelial cells was measured. The results showed that the secretion of both PGI2 and endothelin was greater in SHR than in WKY rats, but that there was no difference in thromboxane A2 secretion. Therefore, our "tissue explant technique" can provide high yield and purity of endothelial cells with their specific biological function in vitro. It will permit us to further study the role of endothelial cells in the development of hypertension.
Chao
C F
CF
Department of Biology and Anatomy, National Defense Medical Center, Taipei, Taiwan, Republic of China.
Chao
Z C
ZC
Lu
M H
MH
Lu
J Y
JY
Chen
H I
HI
D000818:Animals; D001013:Aorta, Thoracic; D002455:Cell Division; D002478:Cells, Cultured; D046508:Culture Techniques / Q000379:methods; D016232:Endothelins / Q000032:analysis; D004730:Endothelium, Vascular / Q000166:cytology* / Q000378:metabolism / Q000473:pathology; D011464:Epoprostenol / Q000032:analysis; D005169:Factor VIII / Q000032:analysis; D005455:Fluorescent Antibody Technique; D006973:Hypertension / Q000473:pathology; D007202:Indicators and Reagents; D008077:Lipoproteins, LDL / Q000378:metabolism; D008297:Male; D008855:Microscopy, Electron, Scanning; D009924:Organ Culture Techniques / Q000379:methods; D051381:Rats; D011918:Rats, Inbred SHR; D011921:Rats, Inbred WKY; D013928:Thromboxane A2 / Q000032:analysis; D017539:Tunica Intima / Q000166:cytology* / Q000473:pathology / Q000648:ultrastructure; D014841:von Willebrand Factor / Q000032:analysis
D016232:Endothelins; D007202:Indicators and Reagents; D008077:Lipoproteins, LDL; D014841:von Willebrand Factor; D013928:Thromboxane A2; D005169:Factor VIII; D011464:Epoprostenol