Myoblasts survive intracardiac transfer and divide further after transplantation.
OBJECTIVE
Skeletal myoblasts have been proven to survive transplantation into myocardial scar tissue. The objective of this study was to evaluate whether these cells can also be transferred into vital myocardium and maintain their ability for cell division after transplantation. In addition, an intravital fluorescence dye for marking these cells was evaluated.


METHODS
Skeletal myoblasts were harvested from male Lewis rats (n = 6) and then expanded in culture. Before implantation, these cells were trypsinized and labeled using an intravital fluorescence dye (PKH-26). Syngenic myoblast transfer to recipient female Lewis rats (n = 36) was used to simulate autologous transplantation. Under general anesthesia, the rats received injections of 106 myoblasts via a subxyphoidal approach into the apex of the heart. The animals were then divided into 3 groups (n = 10 each). The animals were sacrificed at several time points, and the hearts were harvested for histologic examination: group A, 7 days postoperatively; group B, 14 days postoperatively; and group C, 28 days postoperatively. An additional group, group D (n = 6), served as a control group; these animals were injected with only cell medium. Corresponding to the study groups, 2 animals of this control group were sacrificed at each time point, and the hearts were explanted. At histological examination, 8- m sections were investigated to identify surviving stained cells. For further evaluation, the sections were stained using monoclonal antibodies against n-cam, desmin, and a- actin.


RESULTS
No fluorescing cells were found in any hearts of rats in the control group. Surviving fluorescing myoblasts were found in 9 of 10 hearts of groups A and C and 8 of 10 hearts of group B. Labeled myoblasts were located in the intercellular spaces between the myocardial fibers. Fibrotic or inflammatory reactions could not be identified around the injection site in any hearts of the study groups. Immunohistochemical staining results showed that the labeled cells expressed n-cam, desmin, and a-actin. The myoblasts had regained their physiologic structures and had started to form myofibers. In groups B and C, more n-cam-positive cells than labeled cells were found, indicating further cell division.


CONCLUSIONS
Intravital fluorescence staining with PKH- 26 dye proved to be an easy and reliable method for identifying cells after cellular transplantation. Myoblasts survived an intracardiac transfer, regaining their physiologic structures and maintaining their ability for further cell division.
Gulbins
Helmut
H
Department of Cardiac Surgery, University Hospital Grosshadern, LMU Munich, D-81366 Munich, Germany. H.Gulbins@hch.med.uni-muenchen.de
Schrepfer
Sonja
S
Uhlig
Antje
A
Goldemund
Angelika
A
Oberhoffer
Martin
M
Reichenspurner
Hermann
H
Meiser
Bruno
B
Reichart
Bruno
B
D000818:Animals; D002455:Cell Division / Q000502:physiology*; D002470:Cell Survival / Q000502:physiology*; D004195:Disease Models, Animal; D005260:Female; D006085:Graft Survival / Q000502:physiology*; D006331:Heart Diseases / Q000503:physiopathology* / Q000601:surgery*; D016027:Heart Transplantation; D008297:Male; D032448:Myoblasts, Skeletal / Q000502:physiology* / Q000637:transplantation*; D051381:Rats; D011917:Rats, Inbred Lew