Isolation and functional characterization of pericytes derived from hamster skeletal muscle.
OBJECTIVE
At the interface of tissue and capillaries, pericytes (PC) may generate electrical signals to be conducted along the skeletal muscle vascular network, but they are functionally not well characterized. We aimed to isolate and cultivate muscle PC allowing to analyse functional properties considered important for signal generation and conduction.


METHODS
Pericytes were enzymatically isolated from hamster thigh muscles and further selected during a 16-30 days' cultivation period. PC markers were studied by fluorescence activated cell scanning (FACS) and immunocytochemistry. Electrical properties of the cultured PC were investigated by patch clamp technique as well as the membrane potential sensitive dye DiBAC(4) (3).


RESULTS
The cultured cells showed typical PC morphology and were positive for NG2, alpha smooth muscle actin, PDGFR-β and the gap junction protein Cx43. Expressions of at least one single or combinations of several markers were found in 80-90% of subpopulations. A subset of the patched cells expressed channel activities consistent with a Kv1.5 channel. In vivo presence of the channels was confirmed in sections of hamster thigh muscles. Interleukin-8, a myokine known to be released from exercising muscle, increased the expression but not the activity of this channel. Pharmacologic stimulation of the channel activity by flufenamic acid induced hyperpolarization of PC alone but not of endothelial cells [human umbilical vein endothelial cells (HUVEC)] alone. However, hyperpolarization was observed in HUVEC adjacent to PC when kept in co-culture.


CONCLUSIONS
We established a culture method for PC from skeletal muscle. A first functional characterization revealed properties which potentially enable these cells to generate hyperpolarizing signals and to communicate them to endothelial cells.
Walter-Brendel-Centre of Experimental Medicine, Ludwig-Maximilians-Universität München, Munich, Germany.
C
C
Mogensen
B
B
Bergner
S
S
Wallner
A
A
Ritter
S
S
d'Avis
V
V
Ninichuk
P
P
Kameritsch
T
T
Gloe
W
W
Nagel
U
U
Pohl
D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D018031:Connexin 43 / Q000378:metabolism; D006224:Cricetinae; D017629:Gap Junctions / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods*; D016209:Interleukin-8 / Q000494:pharmacology; D051666:Kv1.5 Potassium Channel / Q000378:metabolism; D008564:Membrane Potentials / Q000502:physiology; D008647:Mesocricetus; D018482:Muscle, Skeletal / Q000098:blood supply / Q000166:cytology*; D018408:Patch-Clamp Techniques; D020286:Pericytes / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*
D015415:Biomarkers; D018031:Connexin 43; D016209:Interleukin-8; D051666:Kv1.5 Potassium Channel