A new organ preservation solution for static cold storage of the liver. Amniotic fluid

Abstract Purpose: To evaluate the effect of amniotic fluid in liver preservation in organ transplantation, and compare it with standard preservation solutions. Methods: The groups consisted of Group 1: Ringer Lactate (RL) group, Group 2: HTK group, Group 3: UW group, Group 4: AF group. The livers of rats from Group 1, 2, 3, and 4 were perfused and placed into falcon tubes containing RL, HTK, UW, and AF solutions at +4‎°C, respectively. The tubes were stored for 12 hours in the refrigerator at +4°C. Tissue samples were taken at the 6th and 12th hours for histopathological examinations of the perfused livers, and storage solutions for biochemical analyzes at 6th and 12th hours. Results: AF was shown to maintain organ viability by reducing the number of cells undergoing apoptosis. Histopathological changes such as sinusoidal dilatation, hydropic degeneration, and focal necrosis were found to be similar to the groups in which the standard organ preservation solutions were used. Additionally, the results of INOS, IL-10, and TNF-α,which were evaluated immunohistochemically, have been shown to be similar to the UW and HTK groups. Conclusions: AF provided conservation similar to UW and HTK in the 12-hour liver SCS process. The fact that apoptosis values are comparable to standard preservation solutions supports the success of AF in the cold storage of the liver.

The impact of topically applied preservation solutions on the respiratory epithelium of tracheal grafts submitted to cold ischemia: functional and morphological analysis

OBJECTIVE: Advances in graft reepithelialization and revascularization have renewed interest in airway transplantation. This study aims to determine whether topically applied preservation solutions can ameliorate ischemic injury to tracheal grafts. We analyzed 1) the effects of cold ischemia on the mucociliary clearance of tracheal grafts and 2) the impact of topically applied preservation solutions on the effects of cold ischemia on mucociliary clearance. METHOD: Tracheal segments (n=217) from 109 male Wistar rats were harvested, submerged in low-potassium-dextran-glucose, histidine-tryptophan-ketoglutarate, or saline solution (saline group), and stored at 4°C for 6, 10, 16, or 24 hours. A control group (not submerged) was analyzed immediately after harvesting. In situ mucociliary transport and ciliary beating frequency were measured using a stroboscope. Epithelial integrity, cellular infiltration, and mucus storage were quantified by light microscopy and image analysis software, along with transmission electron microscopy. RESULTS: 1) The effects of cold ischemia: in situ mucociliary transport and ciliary beating frequency were greater in the control group than after cold ischemia. Microscopic analysis results were similar between groups. 2) The effects of preservation solutions: there was no difference between the low-potassium-dextran-glucose, histidine-tryptophan-ketoglutarate, and saline groups in functional or light microscopy analysis. The saline group presented stronger signs of ischemic injury with transmission electron microscopy. CONCLUSIONS: Cold ischemia diminished the mucociliary clearance of the tracheal respiratory epithelium. Topically applied preservation solutions did not ameliorate the injury caused by cold ischemia to the tracheal respiratory epithelium. .

Modelo experimental de perfusão pulmonar ex vivo em ratos: avaliação histopatológica e de apoptose celular em pulmões preservados com solução de baixo potássio dextrana vs. solução histidina-triptofano-cetoglutarato / An experimental rat model of ex vivo lung perfusion for the assessment of lungs regarding histopathological findings and apoptosis: low-potassium dextran vs. histidine-tryptophan-ketoglutarate

OBJETIVO: Comparar os achados histopatológicos e de apoptose em pulmões de ratos preservados em soluções low-potassium dextran (LPD, baixo potássio dextrana), histidine-tryptophan-ketoglutarate (HTK, histidina-triptofano-cetoglutarato) ou salina normal (SN) em 6 h e 12 h de isquemia pela utilização de um modelo experimental de perfusão pulmonar ex vivo. MÉTODOS: Sessenta ratos Wistar foram anestesiados, randomizados e submetidos à perfusão anterógrada pela artéria pulmonar com uma das soluções preservadoras. Após a extração, os blocos cardiopulmonares foram preservados por 6 ou 12 h a 4ºC, sendo então reperfundidos com sangue homólogo em um sistema de perfusão ex vivo durante 60 min. Ao final da reperfusão, fragmentos do lobo médio foram extraídos e processados para histopatologia, sendo avaliados os seguintes parâmetros: congestão, edema alveolar, hemorragia alveolar, hemorragia, infiltrado inflamatório e infiltrado intersticial. O grau de apoptose foi avaliado pelo método TdT-mediated dUTP nick end labeling. RESULTADOS: A histopatologia demonstrou que todos os pulmões preservados com SN apresentaram edema alveolar após 12 h de isquemia. Não houve diferenças em relação ao grau de apoptose nos grupos estudados. CONCLUSÕES: No presente estudo, os achados histopatológicos e de apoptose foram semelhantes com o uso das soluções LPD e HTK, enquanto a presença de edema foi significativamente maior com o uso de SN.

OBJECTIVE: To compare histopathological findings and the degree of apoptosis among rat lungs preserved with low-potassium dextran (LPD) solution, histidine-tryptophan-ketoglutarate (HTK) solution, or normal saline (NS) at two ischemia periods (6 h and 12 h) using an experimental rat model of ex vivo lung perfusion. METHODS: Sixty Wistar rats were anesthetized, randomized, and submitted to antegrade perfusion via pulmonary artery with one of the preservation solutions. Following en bloc extraction, the heart-lung blocks were preserved for 6 h or 12 h at 4ºC and then reperfused with homologous blood for 60 min in an ex vivo lung perfusion system. At the end of the reperfusion, fragments of the middle lobe were extracted and processed for histopathological examination. The parameters evaluated were congestion, alveolar edema, alveolar hemorrhage, inflammatory infiltrate, and interstitial infiltrate. The degree of apoptosis was assessed using the TdT-mediated dUTP nick end labeling method. RESULTS: The histopathological examination showed that all of the lungs preserved with NS presented alveolar edema after 12 h of ischemia. There were no statistically significant differences among the groups in terms of the degree of apoptosis. CONCLUSIONS: In this study, the histopathological and apoptosis findings were similar with the use of either LPD or HTK solutions, whereas the occurrence of edema was significantly more common with the use of NS.

Effects of ischemic preconditioning associated to different preservation solutions in protecting the intestinal graft / Efeitos do precondicionamento isquêmico associado a diferentes soluções de preservação na proteção do enxerto intestinal

PURPOSE: To evaluate the effects of ischemic preconditioning (IPC) associate with different preservation solutions, in the protecting of gut. METHODS: Four groups of 14 rats underwent laparotomy and collecting 20 cm of ileum, for preservation, at 4ºC, in Belzer (Belz), Ringer (RL), Celsior (Cs) and Custodiol (Cust) solutions, for 24 hours. Prior to collection, half of the animals in each group were subjected to IPC. During preservation, in the periods of zero, 12, 18 and 24 hours, were conducted evaluating the degree of mucosal injury and dosage of malondialdehyde acid (MDA). RESULTS: In all periods the RL group, with and without IPC, presented MDA values higher than the Belz and Cs. The degree of mucosal injury in the non-ipc RLgroup with 12h preservation was higher than the others; with 18 and 24h, the RL and Cust had higher degrees of damage than Cs and Belz. With IPC, in all periods, the group Cs and Belz had lower degrees of injury. CONCLUSION: The Celsior and Belzer solutions had better protective effects on the gut and these effects were enhanced by IPC.

OBJETIVO: Avaliar os efeitos do precondicionamento isquêmico (PCI) associado a diferentes soluções de preservação, na proteção do intestino delgado. MÉTODOS: Quatro grupos de 14 ratos Wistar, foram submetidos à laparotomia e coleta de 20 cm de íleo, para preservação, a 4ºC, nas soluções de Belzer (Belz), Ringer (RL), Celsior (Cs) e Custodiol (Cust) por 24 horas. Previamente à coleta, em metade dos animais de cada grupo, o intestino foi submetido ao PCI. Durante a preservação, nos períodos de Zero, 12, 18 e 24 horas, foram realizados avaliação do grau de lesão da mucosa e dosagem do ácido malondialdeído (MDA). RESULTADOS: Em todos os períodos o grupo RL, sem e com pci, apresentou valores maiores de MDA do que o Belz e Cs. O grau de lesão da mucosa nos grupos sem pci com preservação de 12h, no grupo RL, foi maior que nos demais; com 18h e 24h o grupo RL e Cust apresentaram maiores graus de lesão do que Cs e Belz. Com o pci, em todos os períodos, os grupos Belz e Cs apresentaram menores graus de lesão CONCLUSÃO: As Soluções Celsior e Belzer tiveram melhores efeitos na proteção do intestino e estes efeitos foram incrementados pelo precondicionamento isquêmico.

Histologic alterations of rat kidneys perfused with a Euro-Collins diltiazem solution / Alterações histológicas ocorridas em rins de ratos perfundidos com solução de Euro-Collins associada à diltiazem

PURPOSE: Analyse the histologic changes of rat kidneys perfused with isotonic saline solution (ISS), Euro-Collins solution (ECS) and Euro-Collins solution with diltiazem (ECSD). METHODS: Thirty-six Wistar rats were used divided equally, as follow: group A (ISS), group B (ECS) and group C (ECSD). Through a catheter placed into the abdominal aorta, a renal perfusion was performed using a solution according to the group to which the animal belonged. After the complete perfusion, bilateral nephrectomy was performed and the organs were preserved under hypothermia for five distinct periods of time. Glomerulus and tubule were evaluated through optical microscopy. RESULTS: Renal perfusion with ECS and ECSD proved effectiveness in the preservation of the organs up to 36 hours and an increase in the percentage of injured glomeruli was noticed only in the period of 48 hours. CONCLUSIONS: The results showed that exists an association between the tubular injury and the glomeruli lesion degree; kidneys with a higher degree of tubular damage were related to severe glomerular lesion. Also, the addition of a calcium channel blocker, diltiazem, to the ECS for the renal perfusion does not decrease the percentage of glomerular lesion.

OBJETIVO: Analisar as alterações histológicas nos rins de ratos perfundidos com solução salina isotônica (ISS), solução Euro-Collins (ECS) e solução Euro-Collins com diltiazem (ECSD). MÉTODOS: Foram divididos, de forma igual, 36 ratos Wistar, como se segue: grupo A (ISS), grupo B (ECS), grupo C (ECSD). Através de um cateter localizado na aorta abdominal, foi realizada a perfusão renal com a solução de acordo com o grupo ao qual o animal pertencia. Após a perfusão total, realizou-se nefrectomia bilateral com a preservação dos órgãos sob hipotermia por cinco períodos distintos de tempo. Glomérulos e túbulos foram avaliados por microscopia óptica. RESULTADOS: Tanto a perfusão renal com ECS quanto a com ECSD provaram sua efetividade na preservação dos órgãos em até 36 horas e aumento da porcentagem de glomérulos injuriados foi notada apenas no período de 48 horas. CONCLUSÕES: Os resultados mostraram haver uma correlação entre a injúria tubular e o grau de lesão glomerular; rins com um maior grau de dano tubular foram relacionados com lesão glomerular severa. Além disso, a adição de um bloqueador de canal de cálcio, diltiazem, à ECS para a perfusão renal não diminui a porcentagem de lesão glomerular.

Biomechanical and histological evaluation of glycerol-preserved human sclerae

PURPOSE: To determine the histological and biomechanical characteristics of glycerol-preserved human sclera. METHODS: A total of 114 paired human sclerae were cleaned and preserved with 98 percent glycerol under refrigeration at 4 to 8ºC. The samples were divided into a control group with no preservation and 5 groups of 19 sclerae in 7, 15, 30, 90 and 180 days of preservation. Each specimen was submitted to histological examination and tested for traction distensibility functions. RESULTS: Preservation in glycerol did not cause alterations in the histological architecture of the scleral tissue. The mean load required to break the scleral tissue increased according to preservation time as a sigmoid function. A significant increase in mechanical resistance and decrease in distension of scleral tissue occurred after 90 days of preservation. CONCLUSIONS: Scleral preservation in glycerol keeps tissue integrity. The preserved material is less distensible after 90 days. Surgeons who use sclera in ophthalmic procedures should be aware of the mechanical characteristics of glycerol-preserved sclera and take into account tissue preservation time.

OBJETIVO: Determinar as características histológicas e biomecânicas de escleras humanas preservadas em glicerol. MÉTODOS: Escleras de ambos os olhos de 55 doadores foram limpas e preservadas com glicerol a 98 por cento sob refrigeração (4 a 8ºC). A amostra foi dividida em grupo controle sem preservação e 5 grupos de 19 escleras com 7, 15, 30, 90 e 180 dias de preservação. Todas as amostras foram submetidas à avaliação histológica e aos testes de tração e distensão. RESULTADOS: A preservação em glicerol não provocou alterações na arquitetura histológica do tecido escleral. A carga média necessária para romper o tecido escleral aumentou com o tempo de preservação segundo uma função sigmóide. Um incremento significativo na resistência mecânica e diminuição da elasticidade do tecido ocorreram após 90 dias de preservação. CONCLUSÕES: A preservação escleral com glicerol mantém a integridade tecidual. O material preservado torna-se menos distensível após 90 dias de preservação. Cirurgiões que usam esclera preservada em procedimentos oftalmológicos devem estar conscientes das propriedades mecânicas do material e levar em conta o tempo de preservação do material.

A Non-Frozen Living Tissue Bank for Allotransplantation Using Green Tea Polyphenols

Generally, mammalian cells and living tissues can be cryopreserved in a frozen state at very low temperatures over a long storage term. The survival rate of cell suspensions is often acceptable however, living tissues suffer a variety of injuries. In this paper, it was demonstrated that the addition of polyphenols extracted from green tea to conventional cell culture medium and tissue compatible liquid, can control cell proliferation and also preserve tissues for several months at ordinary room temperature, including such tissues as blood vessels, cartilage, islet cells and corneas. This protocol allows a non-frozen living tissue bank to be established using the preservation fluid described.

Preservación de córneas para trasplante en medio de cultivo MCB / Corneal preservation for transplant in MCB culture

Se compara la viabilidad endotelial de córneas de conejo, conservadas en tres medios diferentes durante 48, 72 y 96 horas valoradas mediante la tinción con azul de tripano, microscopia de luz y microscopia electrónica. Material y métodos: Fueron analizadas 20 córneas divididas de la siguiente manera: ocho se conservaron en Optisol, 10 en MCB y dos en solución salina balanceada, mantenidas a 4o C. Resultados: En las córneas mantenidas en solución salina balanceada se observó destrucción de las células endoteliales a las 48 horas. En córneas preservadas en MCB y Optisol se apreció destrucción de un 5-10 por ciento de las células a las 72 horas. La destrucción fue del 20 por ciento después de 96 horas y de 30 por ciento luego de 105 horas en las células preservadas en el medio MCB. El estudio de microscopia electrónica de transmisión también confirmó que las células estructuralmente se encuentran preservadas hasta las 90 horas tanto en Optisol como en MCB. Conclusión: El medio de preservación para córneas producido en México (MCB) mantiene el tejido corneal hasta por 90 horas, de manera similar al Optisol, pero a un costo considerablemente menor.

Influência do pH na preservaçäo de córneas em Optisol / pH variation in Optical and its effectt in corneal storage

Os autores analizaram a influência de pequenas variaçöes do pH na preservaçäo de córneas em Optisol (Chiron Ophthalmics, Irvine, California). Dez pares de córneas humanas foram armazenadas por cinco dias, com Optisol, em pH 7,0 7,2 e 7,5 e depois analisados com lâmpada de fenda, microscópio especular e microscopia eletrônica. Näo foram encontradas diferenças significativas nem na contagem endotelial,nem nas caacterísticasmorfológicas do endotélio corneano nas córneas preservadas nesses pHs