Jg. Derhaag et al., PRODUCTION AND CHARACTERIZATION OF SPONTANEOUS RAT-HEART ENDOTHELIAL-CELL LINES, Laboratory investigation, 74(2), 1996, pp. 437-451
Endothelial cells (EC) are important regulatory cells in physiology an
d pathology, in vitro studies with rat EC from heart tissue are hamper
ed by laborious isolation and purification procedures, low yield, and
limited lifespan of the cells. Therefore, it is essential to obtain lo
ng-term heart EC lines that offer a more adequate in vitro system for
studying rat heart EC. An ex vivo perfusion model was used to isolate
EC from rat heart. Isolation and culture conditions were modified to a
llow spontaneous development of immortalized rat heart EC (RHEC) lines
. Produced cell lines were tested for endothelial nature using a panel
of markers. The selected RHEC lines were subsequently tested for a se
ries of phenotypic and functional properties representative of EC in t
he context of physiologic and inflammatory functions in vivo. A series
of three spontaneous RHEC lines was produced from 13 isolations from
Lewis rat hearts: RHEC-3, RHEC-10, and RHEC-11. These lines were stabl
e for more than 30 passages (RHEC-3 for more than 100). The cell lines
were tumorigenic and developed hemangiomas on in vivo injection. All
three lines expressed major histocompatibility complex (MHC) class I b
ut no MHC class II. Intercellular adhesion molecule 1 was only express
ed by RHEC-3. Cytokine stimulation induced vascular cell adhesion mole
cule 1 in RHEC-3 and RHEC-11 as well as MHC class II in all three line
s in different quantities. The phenotypic characteristics of the diffe
rent RHEC lines resembled the myocardial microvascular endothelium in
situ. The three lines expressed angiotensin-converting enzyme, and the
y responded to histamine and ATP but not to thrombin and bradykinin. T
hey constitutively produced small amounts of endothelin and high level
s of tissue plasminogen activator; they produced little (after stimula
tion with phorbol-ester PMA) or no von Willebrand factor. The RHEC lin
es produced thromboxane A2 but no prostacyclin; on stimulation with ar
achidonic acid and A23187, they produced prostaglandin E2. Therefore,
we conclude the following. 1) The described isolation and culture tech
nique is successful for production of spontaneous stable EC lines from
rat heart. 2) RHEC-3, -10, and -11 can be considered a series of diff
erent lines representative of the heterogeneity of heart microvascular
endothelium in vivo. 3) The RHEC lines offer a series of valuable too
ls to study various heart EC functions and mechanisms in physiology an
d pathology.