No evidence of porcine endogenous retrovirus in patients with type 1 diabetes after long-term porcine islet xenotransplantation.

Xenotransplantation is a promising alternative for donor shortage to ameliorate physiologic and metabolic disorders. The major concern for xenotransplant is the risk of zoonosis mainly by the porcine endogenous retrovirus (PERV), presentation in the piglet genome. Twenty-three patients with type 1 diabetes were transplanted with porcine islets using collagen-generating devices which were implanted subcutaneously in the anterior wall of the abdomen. Clinical characteristics and metabolic tests were recorded in each visit. They were tested for PERV using PCR and RT-PCR from blood pretransplantation and every 3 months during a 4.6- to 8-year follow-up after their first xenotransplant. Tests by PCR of every DNA sample (780 samples) revealed that there was no PERV infection in the DNA of white cells. No evidence of PERV activation was found in this group of patients with type 1 diabetes during clinical long-term follow-up.

Unexpected immunoresponse to Gal and APA antigens in diabetic type 1 patients receiving neonatal pig islets after 6 years.

Cotransplantation of porcine islets and Sertoli cells into preimplanted subcutaneous devices improve metabolic control in type 1 diabetic patients, and survive grafted for more than 4 years. We report here, further assessment of the endocrine and porcine nature of the surviving cells and the immune responses elicited toward Gal alpha(1,3)-Gal beta(1,4)-GlcNAc (Gal) antigen in patients who received a second and third transplants. No immunosuppressive drugs were administered. We were able to immunostain insulin- and glucagon-positive cells in all biopsies of patients and Sertoli cell markers in 60.9% of biopsies. Additionally, all biopsies tested, amplified the porcine COII gene. Patients demonstrated an increase in antipig antibodies in response to the first transplant with a decreasing response toward the second and third transplants. In all transplants, the IgG levels promptly returned to basal values after 3-4 months. The long-term survival of porcine cells and the reduced humoral immune response to multiple transplants indicate a form of tolerance. We have not been able to find CD25-positive cells, indicating that it is probably an immune accommodation of the graft.

Xenotransplantation of porcine neonatal islets of Langerhans and Sertoli cells: a 4-year study.

OBJECTIVE: Porcine islets of Langerhans for xenotransplantation into humans have been proposed as a solution to the shortage of human donors. Rejection is one of the main constraints. This study presents the results of a clinical trial using a novel method for transplanting and immunoprotecting porcine islets in type 1 diabetic patients. DESIGN: A 4-year follow up of a clinical trial involving 12 patients, with no immunosuppressive drugs at any point. Eleven age matched untransplanted diabetics served as controls. METHODS: We have developed a procedure for protecting neonatal porcine islets by combining them with Sertoli cells and placing them in a novel subcutaneous autologous collagen-covered device. RESULTS: In the patients in the treatment group, no complications arose and no porcine endogenous retrovirus infection was detected. Half of the patients showed a significant reduction in insulin requirements compared with both their pre transplant levels and controls, and this reduction was maintained for up to 4 years. Two patients became insulin-independent for several months. Porcine insulin was detected in three patients' sera following glucose stimulation up to 4 years post transplant. Three years post transplant, one of four devices was removed from four patients, and the presence of insulin-positive cells in the transplant was demonstrated by immunohistology in all 4 patients. CONCLUSIONS: Long-term cell survival with concurrent positive effects on metabolic control are possible by this technique.

Method for evaluating quality of cultured neonatal pig Sertoli cells.

BACKGROUND: Sertoli cells (SC) in the testis secrete factors that nourish and immunoprotect developing spermatozoa, which have made them the focus of studies that aim to generate localized tolerance, particularly for transplantation and perhaps autoimmunity. Several methods have been described to isolate these cells, which include a two-step enzymatic digestion with limited assessment of the culture. Here we describe a one-step method, and a series of tests for determining purity, viability, and function of the cultured cells. METHODS: We isolated SC from neonatal pigs using Liberase HI digestion. Viability and apoptosis of cultured cells were measured by flow cytometry with propidium iodide and annexin, respectively. Specific identification of the Sertoli type was made by immunodetection of Sox9, vimentin, and Mullerian inhibiting substance. Moreover, for functionality we were able to detect clusterin in the cultured cells by Western blot. RESULTS: Our isolation method had a yield and purity similar to previous reports measured with two-step methods. Viability was 95.22 +/- 0.57% and apoptotic cells were 10.5 +/- 0.32% after 48 h in culture. At 7 days, practically all cells expressed Sox9, Mullerian inhibiting substance, clusterin, and vimentin. CONCLUSIONS: We describe an alternative strategy for preparing and identifying cultured SC for further assays of metabolic activity or in transplantation models. Establishing a one-step Liberase-digestion method for isolation, evaluating viability and apoptosis by more sensitive methods, and detecting specific markers in culture can help to evaluate the quality of cultured cells. Specific cell markers for identifying SC may be critical when identifying SC outside the testis, in contrast with vimentin which is useful only for in situ cells.

Improved method for isolation of porcine neonatal pancreatic cell clusters.

UNLABELLED: Based on the described methods for the isolation of neonatal pancreatic cell clusters (NPCCs), we have developed modifications in order to improve their quality, functionality, and process reproducibility in the isolation technique, for potential use in research. In addition, we indicate techniques for describing yield, functionality, viability and purity of the NPCCs. METHODS: Purity of the NPCCs was determined through dithizone staining and subjected to image analysis. Viability and apoptosis was measured through flow cytometry with propidium iodide and annexin, respectively. NPCC functionality was measured through a static glucose stimulation test. RESULTS: We developed a high-yield reproducible technique that had 81 279.55 +/- 18 257.05 IEQ/g of pancreas at 4 days of culture, with a 94% viability and an 88 +/- 2.73% purity. Stimulation index from the glucose stimulation test was >10. CONCLUSION: The technique allowed us to obtain NPCC with optimal viability, functionality, purity, and endurance for use in research.

Neoplasia epitelial papilar sólida y quística de páncreas: imágenes del tumor y seguimiento / Solid Cystic papillary epithelial neoplasia of the pancreas: tumor images and follow up

Se presenta el caso clínico de una niña de 8 años de edad, que presenta dolor abdominal de 2 meses de evolución. Se estudió como dolor abdominal crónico encontrándose en el ultrasonido abdominal un tumor dependiente de páncreas: La tomografía axial computada (TAC) y la resonancia magnética (RM) permitieron una adecuada localización de la tumoración (cuerpo y cola del páncreas). Se sometió a resección total de la tumoración por laparotomía. El reporte histopatológico fue el de neoplasia epitelial papilar sólida y quística de páncreas. El seguimiento postoperatorio ha sido de 6 meses; la paciente está asintomática y el control tomográfico postoperatorio no muestra actividad tumoral

Symposium Trasplante de órganos en México: Trasplante de páncreas / Transplantation of pancreas

En el presente artículo se hace una revisión de la literatura en relación con los trasplantes de páncreas. Se discute acerca de los dos tipos que en la actualidad se emplean para resolver la patología pancreática como son el trasplante total y el de células. Se comenta sobre los resultados obtenidos en los principales centros internacionales, donde se realiza este tipo de cirugía

Micropinzas vasculares: desarrollo tecnológico universitario / Microsurgicals clamps. a university technolocigal development

Un problema importante en el entrenamiento y la enseñanza de la microcirugía es la disponibilidad de micropinzas vasculares debido a su alto costo y la dificultad para su importación. Por estas razones, en nuestro laboratorio, desarrollamos unas micropinzas vasculares, equiparables a las que se producen en otros países. Las pinzas fueron sometidas a pruebas de control. Su empleo resultó satisfactorio en el laboratorio de entrenamiento y enseñanza

Trasplante de páncreas y riñón: informe de un caso / Pancreas and kidney transplant: report of one case

El presente informe es de una paciente con diabetes tipo I que fue sometida a doble trasplante de riñón y pancreático en un solo tiempo en el cual se constata por un lado la efectividad del látex como bloqueador de la secreción exocrina y, por el otro, la gran afección de arterias por su enfermedad primaria.

Ciclosporina-A en el transplante renal en niños y sus efectos sobre el crecimiento / Ciclosporin-A in renal transplantation in children and its effects on growth

Los corticosteroides del esquema inmunosupresor que reciben convencionalmente los niños con trasplante renal constituyen un factor importante en la detención de su crecimiento. El empleo de otros esquemas que eviten o disminuyan la dosis de corticosteroides ofrecería mejores perspectivas pero sería conveniente balancear este benefício contra otros efectos secundarios y el costo de los fármacos. Este estudio consistió en emplear como esquema postrasplante: (I) ciclosporina-A más prednisona (a muy bajas dosis) y comparar sus resultados a un año, con los obtenidos con azathioprina más prednisona (II) (a dosis triple de la anterior). Se estudiaron cuatro parejas de niños urémicos con características similares en cuanto a edad, retardo en el crecimiento y tiempo de evolución de la nefropatía. A cuatro se les administró el esquema I y a los otros cuatro el esquema II. Al año se encontró que los niños del grupo I habían recibido 160 mg/kg/año dosis promedio de prednisona, su velocidad de crecimiento fue de 5.4 ñ 3.2 cm/año y la diferencia de la desviación estándar de la talla para la edad cronológica fue de -0.12 ñ 0.09. Los niños del grupo II habían recibido 428 mg/kg/año, su velocidad de crecimiento fue 1.7 ñ 1.4 cm y la desviación estándar de la talla para la edad cronológica fue + 0.34 ñ 0.25. Los niveles séricos de creatinina fueron 121 ñ 6.4 micron-mol/L en los niños del grupo I contra 62 ñ 12.7 en los niños del grupo II. Solo uno de los casos del grupo I logró revertir su disfunción renal a lo normal al cambiar su tratamiento al esquema convencional. Se sugiere que por su costo y nefrotoxicidad, la ciclosporina-A debiera estar indicada para casos donde la detención del crecimiento fuera un problema psicosocial importante para el receptor, cuando la histocompatibilidad con un donante vivo no sea óptimo o cuando el receptor haya tenido uno o más trasplantes previos