Apoptosis in fresh and cryopreserved cardiac valves of pig samples.
To analyse the influence of cold ischemic time (CIT) (2-24 h) and of cryopreservation (liquid phase) on the viability of the valvular fibroblasts and in the presence of apoptosis. Cardiac valves from 10 pigs were evaluated by anatomo-pathological study of the wall, muscle and leaflet. At the same time, the presence of cellular death due to apoptosis was investigated in two ways; directly on tissue by Apodetec system and by two-colour flow cytometry assay analyzing a suspension of fibroblast from valve leaflets using Anexina V and propidium iodure (PI). We established three groups of samples to compare different experimental conditions: 2 h of ischemia (group 1), 24 h of ischemia (group 2), and a programme of cryopreservation (-1 degrees C/min) after 2 h of ischemia, followed by storage in liquid nitrogen during a week and thawing was performed (group 3). The analysis of viabilities showed slight differences between all three groups. The results indicated CIT of 24 h undergoing more structural affectation than CIT of 2 h. Flow cytometry analysis did not show important differences between groups; however cryopreserved samples (group 3) slightly less viability and a higher percentage of death by apoptosis than group 1 and 2 using flow cytometry. Apoptosis was confirmed on tissue from all valves but mainly in samples of group 2 and group 3. In summary, the viability of the valves in the case of ischemic times of 2 h, 24 h or after cryopreservation/thawing differs slightly. The death of the cells is mainly mediated by necrosis and not by apoptosis.
Rendal Vázquez
M Esther
ME
Unit of Cryobiology, Complejo Hospitalario Juan Canalejo, Carretera del Pasaje s/n, Hospital Teresa Herrera, 15006 La Coruna, Spain. Esther_Rendal@canalejo.org
Díaz Román
T M
TM
Rodríguez Cabarcos
M
M
Zavanella Botta
C
C
Domenech García
N
N
González Cuesta
M
M
Sánchez Dopico
M J
MJ
Pértega Díaz
S
S
Andión Núñez
C
C
D000818:Animals; D017209:Apoptosis; D002470:Cell Survival; D050377:Cold Ischemia; D015925:Cryopreservation / Q000379:methods*; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology*; D006351:Heart Valves / Q000166:cytology* / Q000502:physiology; D009336:Necrosis; D009926:Organ Preservation / Q000379:methods*; D013552:Swine; D014015:Tissue Banks; D014022:Tissue Survival