Ricardo Vilani

Results of a decellularized porcine heart valve implanted into the juvenile sheep model.

OBJECTIVE This study was performed to evaluate the possibility of creating a glutaraldehyde-free porcine xenograft to improve long-term durability. METHODS A decellularized porcine pulmonary valve was implanted into the right ventricular outflow tract of 7 juvenile sheep. Valves were explanted after 3 months (n = 4) and 6 months (n = 3). Evaluation was performed by gross examination, radiography, histology (hematoxylin-eosin and Sirius red staining), and immunohistochemistry. Quantitative determination of calcium content was investigated by atomic absorption spectrometry. RESULTS All animals showed fast recovery without complications. At explantation, all decellularized valves showed smooth and pliable leaflets without evidence of thrombosis. The valve wall was also smooth and pliable without hardness. Light microscopy showed a monolayer of host endothelial cells covering the inner surface of the heart valves and repopulation of host fibroblasts into the deeper layers. Sirius red staining enabled visualization of the production of new collagen. Radiographic results showed an absence of calcification, confirmed by the low calcium levels (1.08 +/- 0.28 microg/g and 0.73 +/- 0.31 microg/g at 3 and 6 months, respectively) revealed by atomic absorption spectrometry. CONCLUSIONS The results with the juvenile sheep model showed that decellularized heart valves are recellularized in vivo. Host endothelial cells form a monolayer on the inner surface of the valve matrix. Furthermore, host fibroblasts repopulate the valve matrix and produce collagen; thus, a remodeling potential can be expected.

Decellularized heterografts versus cryopreserved homografts: experimental study in sheep model

OBJECTIVES The aim of this study is to assess the biological behaviour of porcine decellularized heterografts (Desc group) compared with cryopreserved homografts (Crio group) implanted in juvenile sheep. METHODS Decellularized porcine pulmonary heterografts were implanted in five animals and cryopreserved pulmonary homografts in another five. The animals were followed-up for a mean of 280 +/- 14 days. The valve diameter was measured by echocardiography, which was performed at the 30th postoperative day, and before the explantation. The valves were also assessed macroscopically. Histological evaluation was performed using H.E., Gomori and Weigert staining. Immunohistochemistry specified different cell types (Factor VIII, CD3, Vimentin and CD68). Calcium quantity was analyzed using atomic absortion spectometry. RESULTS There was one death in the Desc group due to endocarditis. The valves of Crio group showed decrease in the cellularity whereas the valves of Desc group showed matrix repopulation with endothelial and interstitial cells. Loss of collagen density and disarrangement of the normal fiber architecture was observed in Crio group. Calcium content demonstrated higher levels on the cusps and conduits in Crio group comparatively with Desc group. (P=0.016). The mean valvular diameter at the explantation was significantly increased (P=0.025) in the Desc group. CONCLUSIONS Decellularized heterografts had a different biological behaviour when compared to cryopreserved homografts and become repopulated by cells with fibroblasts and endothelial cells characteristics. The matrix was preserved and some regenerative potential was present.

Successful implantation of a decellularized equine pericardial patch into the systemic circulation.

Background In the past, successful use of decellularized xenogenic tissue was shown in the pulmonary circulation. This study, however, evaluates a newly developed decellularized equine pericardial patch under high pressure circumstances. Material/Methods Seven decellularized equine pericardial scaffolds were implanted into the descending aorta of the juvenile sheep. The implanted patches were oversized to evaluate the durability of the decellularized tissue under high surface tension (Law of Laplace). After 4 months of implantation, all decellularized patches were inspected by gross examination, light microscopy (H&E, Serius red, Gomori, Weigert, and von Kossa straining), and immunohistochemical staining. Results The juvenile sheep showed fast recovery after surgery. There was no mortality during follow-up. At explantation, only limited adhesion was seen at the surgical site. Gross examination showed a smooth and pliable surface without degeneration, as well as absence of aneurysmatic dilatation. Light microscopy showed a well preserved extracellular scaffold with a monolayer of endothelial cells covering the luminal side of the patch. On the outside part of the patch, a well developed neo-vascularization was seen. Host fibroblasts were seen in all layers of the scaffolds. There was no evidence for structural deterioration or calcification of the decellularized equine pericardial scaffolds. Conclusions In the juvenile sheep, decellularized equine tissue showed no structural deterioration, but regeneration and remodeling processes at systemic circulation.

Experimental study with deccelularized porcine heterografts: the prosthesis of the future

OBJETIVO: Avaliar, comparativamente, o comportamento de homoenxertos valvares criopreservados e de heteroenxertos descelularizados implantados na via de saida do ventriculo direito de carneiros jovens, assim como relatar a experiencia clinica inicial com homoenxertos descelularizados em pacientes submetidos a operacao de Ross. METODO: No grupo A, foram implantados quatro homoenxertos pulmonares criopreservados e no Grupo B, quatro heteroenxertos porcinos valvares descelularizados com acido deoxicolico. Em cada grupo, dois animais foram sacrificados no 3o mes e dois no 5o mes de pos-operatorio. As pecas foram avaliadas macroscopicamente e por radiografias, alem de exame microscopico com coloracoes HE, Tricromico de Mallory e Sirius Red. A experiencia clinica com quatro pacientes submetidos a operacao de Ross com emprego de homoenxertos valvares descelularizados e relatada. RESULTADOS: Todos os animais sobreviveram. Os ecocardiogramas demonstraram boa funcao valvar nos dois grupos. Embora as cuspides valvares em ambos os grupos tivessem aspecto macroscopico normal, a mensuracao de calcio e o estudo radiologico demonstraram inicio de calcificacao focal nos homoenxertos criopreservados, mas nao nos heteroenxertos descelularizados. Esses achados foram confirmados por exames microscopicos. Os homoenxertos criopreservados demonstraram perda de sua celularidade, enquanto que os heteroenxertos descelularizados demonstraram repopulacao progressiva da matriz colagena com fibroblastos, assim como reendotelizacao. Os quatro pacientes operados tiveram evolucao pos-operatoria imediata satisfatoria, com funcao normal dos homoenxertos. CONCLUSOES: Heteroenxertos valvares descelularizados foram progressivamente repopulados por celulas autogenas e exibiram minima tendencia a calcificacao no modelo estudado. Esses resultados sugerem que homoenxertos descelularizados possam ter alguma capacidade regenerativa e com isso ter durabilidade superior aos homoenxertos criopreservados convencionais.

An Experimental Study of Decellularized Xenografts Implanted Into the Aortic Position with 4 Months of Follow Up

This study evaluated decellularized stentless porcine xenografts into the aortic position of seven juvenile sheep. The hemodynamic performance were analyzed by means of echocardiographic examination. Post mortum, specimens were evaluated by gross examination, light microscopy, and immunohistochemical staining. Explantations were performed at up to four months. Echocardiographic evaluation examination showed a maximum flow velocity of 0.94m/s with absence of regurgitation. Gross examination showed smooth and pliable leaflets endothelial cells covered the valve wall and leaflets, deeper layers presented ingrowth of host interstitial cells. There was no evidence for calcification. Our preliminary results showed excellent hemodynamics. Regeneration by host cells and absence of calcifications was observed at 4 months follow-up within the systemic circulation.

Utilização do etanol na prevenção da calcificação em heteroenxerto valvar pulmonar porcino: estudo experimental em ovinos

Objective: The objective of this work was to comparatively evaluate of the use of ethanol (E) with glutaraldehyde (GDA) to inhibit the calcification of porcine pulmonary valvar heterografts (PPVH) implanted in the right ventricle outflow tract of young sheep. Method: Ten young sheep were used, divided into two similar groups: the GDA (Control) Group and the Ethanol Group (in which the PPVH were pre-treated with ethanol80). The surgery consisted of implanting the PPVH in the right ventricle outflow tract using a cardiopulmonary bypass. Echocardiography was performed to measure the transvalvar gradient and macroscopic analyses of the PPVH wall and microscopic analysis for calcification were performed. The amount of calcium was measured in the valve cusp at 210 days. Results: The quantity of calcium in the PPVH of the GDA Group was 7.98 ± 6.82µg calcium/mg tissue and of the E Group it was 0.31 ± 0.33µg calcium/ mg tissue. In respect to the maximum transvalvar gradient at 30 days, there was no significant difference between the groups. At 90, 180 and 210 days however, the GDA Group showed a greater transvalvar gradient than the E Group. The macroscopic analyses of the heterograft wall evidenced calcification in both groups but it was more pronounced in the GDA Group. The microscopic analysis of the cusp valve revealed the presence of calcification in all animals of the GDA Group, while in the E Group no calcification was seen. Conclusion: In conclusion ethanol-80 inhibits calcification on the cusps during the period of the analysis but does not inhibit the calcification of the PPVH.