A study of the effects of warm ischaemic times on harvested homografts

Thesis (M. Tech.) -- Central University of Technology, Free State, 200

Does prolonged post-mortem cold ischemic harvesting time influence cryopreserved pulmonary homograft tissue integrity?

Published ArticleThis study investigated cryopreserved pulmonary homograft (CPA) structural integrity after prolonged cold ischemic harvesting times in a juvenile sheep model. Three groups with different post-mortem cold ischemic harvesting times were studied, i.e. Group 1 (24 h, n = 10); group 2 (48 h, n = 10); group 3 (72 h, n = 10). In each group, 5 CPAs were studied in vitro after cryopreservation and thawing. The other 5 CPAs were implanted in juvenile sheep for a minimum of 180 days. Serology samples were obtained and echocardiography was performed before euthanasia. Hematoxylin and eosin (H&E), scanning electron microscopy (SEM), von Kossa, Picrosirius red, α-actin, immunohistochemistry [von Willebrand factor (vWF), CD4, CD31 and CD34] and calcium content analyses were performed on explanted CPAs. The in vitro and in vivo studies failed to demonstrate any change in tensile strength, Young's Modulus and thermal denaturation (Td) results between the groups. SEM demonstrated a reduction in endothelial cells (50 % at 24 h, 60.9 % at 48 h and 40.9 % at 72 h), but H&E could not demonstrate autolysis in any CPA in vitro. All cultures were negative. In the explanted groups, IgE, IgM and IgG results were inconclusive. Echocardiography demonstrated normal valve function in all groups. H&E and Picrosirius red staining confirmed tissue integrity. vWF, CD31 and CD34 staining confirmed a monolayer of endothelial cells in all explanted valves. Calcium content of explanted CPA leaflets was similar. This experimental study supports the concept of prolonging the cold ischemic harvesting time of cryopreserved homografts to reduce homograft shortage

Cadaver donation: structural integrity of pulmonary homografts harvested 48 h post mortem in the juvenile ovine model

Published ArticleAbstract Cryopreserved pulmonary homograft (CPH) implantation remains the gold standard for reconstruction of the right ventricular outflow tract (RVOT). Harvesting homografts\24-h post mortem is the international norm, thereby largely excluding cadaveric donors. This study examines the structural integrity and stability of ovine pulmonary homografts harvested after a 48-h post mortem period, cryopreserved and then implanted for up to 180 days. Fifteen ovine pulmonary homografts were harvested 48-h post mortem and cryopreserved. Five CPH served as a control group (group 1; n = 5). CPH were implanted in the RVOT of juvenile sheep and explanted after 14 days (group 2; n = 5) and 180 days (group 3; n = 5). Leaflet integrity was evaluated by strength analysis, using tensile strength (TS), Young’s modulus (YM) and thermal denaturation temperature (Td), and morphology, including haematoxylin and eosin (H&E), Picrosirius red staining, scanning electron microscopy (SEM), transmission electron microscopy (TEM) and von Kossa stains. Echocardiography confirmed normal function in all implants. In explants, no reduction in TS, YM or Td could be demonstrated and H&E showed mostly acellular leaflet tissue with no difference on Picrosirius red. TEM demonstrated consistent collagen disruption after cryopreservation in all three groups, with no morphological deterioration during the study period. von Kossa stains showed mild calcification in group 3. No deterioration of structural integrity could be demonstrated using strength or morphological evaluations between the controls and implant groups over the study period. Extending the post mortem harvesting time of homografts beyond 24 h did not appear to negatively affect the long-term performance of such transplanted valves in this study