Molecular Pathways Leading to Induction of Cell Death and Anti-Proliferative Properties by Tacrolimus and mTOR Inhibitors in Liver Cancer Cells.

BACKGROUND/AIMS: Orthotopic liver transplantation (OLT) is the recommended treatment for patients at early stages of hepatocarcinoma (HCC) with portal hypertension and/or increased bilirubinemia, but without vascular-associated diseases. Tumor recurrence, which is the main drawback for the survival of patients submitted to OLT for HCC, has been related to tumor-related variables and the immunosuppressive therapies. We have previously shown that Tacrolimus (FK506) exerts a more potent pro-apoptotic and anti-proliferative effects than the mammalian target of rapamycin (mTOR) inhibitors (Sirolimus and Everolimus) in liver cancer cells. This study identified the role of the immunosuppressant partners such as FK506-binding proteins (FKBPs) in the induction of cell death and arrest of cell proliferation by immunosuppressants in two representative liver cancer cells. METHODS: The regulation of endoplasmic reticulum (ER) stress, apoptosis/autophagy, cell proliferation, and FKBPs expression was determined in Tacrolimus-, Sirolimus- and Everolimus-treated primary human hepatocytes, and hepatoma HepG2 and Huh7 cell lines. The functional repercussion of FKBPs on cell death and proliferation was also addressed using the siRNA technology. The assessed antitumoral properties of the immunosuppressants were associated to microRNAs (miRNAs) pattern. RESULTS: The enhanced pro-apoptotic and anti-proliferative properties of Tacrolimus versus mTOR inhibitors were associated with increased protein kinase RNA-like endoplasmic reticulum kinase (PERK)-related ER stress, Ser15P-p53/p53 ratio and p21 protein expression that may counterbalance the risk of proliferative upregulation caused by enhanced Thr172P-Cdk4/Cdk4 activation in liver cancer cells. The inhibition of the mTOR pathway by Sirolimus and Everolimus was related to an induction of autophagy; and at a high dose, these drugs impaired translation likely at a very early step of the elongation phase. Tacrolimus and mTOR inhibitors increased the protein expression of FKBP12 and FKBP51 that appeared to play pro-survival role. Interestingly, the administration of immunosuppressants yields a specific pattern of miRNAs. Tacrolimus and mTOR inhibitors decreased miR-92a-1-5p, miR-197-3p, miR-483-3p and miR-720, and increased miR-22-3p, miR-376a-3p, miR-663b, miR-886-5p, miR-1300 and miR-1303 expressions in HepG2 cells. CONCLUSION: The more potent pro-apoptotic and anti-proliferative properties of Tacrolimus versus mTOR inhibitors were associated with an increased activation of PERK and p53 signaling, and p21 protein expression. FKBP12 and FKBP51 appeared to be the most relevant partners of Tacrolimus and mTOR inhibitors exerting a pro-survival effect in HepG2 cells. The observed effects of immunosuppressants were related to a specific miRNA signature in liver cancer cells.

Molecular characterization of autophagic and apoptotic signaling induced by sorafenib in liver cancer cells.

Sorafenib is the unique accepted molecular targeted drug for the treatment of patients in advanced stage of hepatocellular carcinoma. The current study evaluated cell signaling regulation of endoplasmic reticulum (ER) stress, c-Jun-N-terminal kinase (JNK), Akt, and 5'AMP-activated protein kinase (AMPK) leading to autophagy and apoptosis induced by sorafenib. Sorafenib induced early (3-12 hr) ER stress characterized by an increase of Ser51 P-eIF2α/eIF2α, C/EBP homologous protein (CHOP), IRE1α, and sXBP1, but a decrease of activating transcription factor 6 expression, overall temporally associated with the increase of Thr183,Tyr185 P-JNK1/2/JNK1/2, Thr172 P-AMPKα, Ser413 P-Foxo3a, Thr308 P-AKt/AKt and Thr32 P-Foxo3a/Foxo3a ratios, and reduction of Ser2481 P-mammalian target of rapamycin (mTOR)/mTOR and protein translation. This pattern was related to a transient increase of tBid, Bim EL , Beclin-1, Bcl-xL, Bcl-2, autophagy markers, and reduction of myeloid cell leukemia-1 (Mcl-1) expression. The progressive increase of CHOP expression, and reduction of Thr308 P-AKt/AKt and Ser473 P-AKt/AKt ratios were associated with the reduction of autophagic flux and an additional upregulation of Bim EL expression and caspase-3 activity (24 hr). Small interfering-RNA (si-RNA) assays showed that Bim, but not Bak and Bax, was involved in the induction of caspase-3 in sorafenib-treated HepG2 cells. Sorafenib increased autophagic and apoptotic markers in tumor-derived xenograft model. In conclusion, the early sorafenib-induced ER stress and regulation of JNK and AMPK-dependent signaling were related to the induction of survival autophagic process. The sustained drug treatment induced a progressive increase of ER stress and PERK-CHOP-dependent rise of Bim EL , which was associated with the shift from autophagy to apoptosis. The kinetic of Bim EL expression profile might also be related to the tight balance between AKt- and AMPK-related signaling leading to Foxo3a-dependent BIM EL upregulation.

Development of a liver graft assessment expert machine-learning system: when the artificial intelligence helps liver transplant surgeons.

Background: The complex process of liver graft assessment is one point for improvement in liver transplantation. The main objective of this study is to develop a tool that supports the surgeon who is responsible for liver donation in the decision-making process whether to accept a graft or not using the initial variables available to it. Material and method: Liver graft samples candidate for liver transplantation after donor brain death were studied. All of them were evaluated "in situ" for transplantation, and those discarded after the "in situ" evaluation were considered as no transplantable liver grafts, while those grafts transplanted after "in situ" evaluation were considered as transplantable liver grafts. First, a single-center, retrospective and cohort study identifying the risk factors associated with the no transplantable group was performed. Then, a prediction model decision support system based on machine learning, and using a tree ensemble boosting classifier that is capable of helping to decide whether to accept or decline a donor liver graft, was developed. Results: A total of 350 liver grafts that were evaluated for liver transplantation were studied. Steatosis was the most frequent reason for classifying grafts as no transplantable, and the main risk factors identified in the univariant study were age, dyslipidemia, personal medical history, personal surgical history, bilirubinemia, and the result of previous liver ultrasound (p < 0.05). When studying the developed model, we observe that the best performance reordering in terms of accuracy corresponds to 76.29% with an area under the curve of 0.79. Furthermore, the model provides a classification together with a confidence index of reliability, for most cases in our data, with the probability of success in the prediction being above 0.85. Conclusion: The tool presented in this study obtains a high accuracy in predicting whether a liver graft will be transplanted or deemed non-transplantable based on the initial variables assigned to it. The inherent capacity for improvement in the system causes the rate of correct predictions to increase as new data are entered. Therefore, we believe it is a tool that can help optimize the graft pool for liver transplantation.

Differential Antitumoral Properties and Renal-Associated Tissue Damage Induced by Tacrolimus and Mammalian Target of Rapamycin Inhibitors in Hepatocarcinoma: In Vitro and In Vivo Studies.

Orthotopic liver transplantation (OLT) is the recommended treatment for patients at early stages of hepatocarcinoma (HCC) with potential portal hypertension and/or bilirubinemia, but without vascular-associated diseases. The patients are receiving immunosuppressive therapy to reduce graft rejection, but differential side effects have been related to calcineurin and mTOR inhibitor administration regarding tumor recurrence and nephrotoxicity. The in vitro studies showed that Tacrolimus exerted a more potent pro-apoptotic effect than Everolimus (Huh 7>Hep 3B>HepG2), being sirolimus only active in Hep3B cell line. Tacrolimus and Everolimus exerted potent antiproliferative properties in Huh 7 and Hep3B in which cells Sirolimus was inactive. Interestingly, Tacrolimus- and Everolimus-dependent G0/G1 cell accumulation occurred as a consequence of drastic reduction in S, as well as in S and G2+M phases, respectively. The in vivo studies support data on the more effective antitumoral properties of Everolimus, eventual risk of pro-angiogenic tumoral properties and nephrotoxicity of Tacrolimus, and pro-proliferative properties of Sirolimus in tumors developed in nude mice.

[Prevention of peritoneal adhesions to intraperitoneal prostheses. An experimental study in pigs]. / Prevención de adherencias peritoneales a las prótesis intraperitoneales. Estudio experimental en cerdos.

INTRODUCTION: The aim of this study was to investigate the effect of fibrin glue and hyaluronidase gel on the prevention of postoperative peritoneal adhesions to intraperitoneal prostheses. MATERIAL AND METHOD: Twenty pigs, divided in two groups, were included. In all animals, four implants (4 x 4 cm) were placed: two polypropylene mesh implants were placed in an upper location and two polytetrafluoroethylene (PTFE) implants (Dualmesh Plus Corduroy) were placed in a lower position. Implants located in the right side of the animals were painted with fibrin glue (group A, n = 10) or with hyaluronidase gel (group B, n = 10). After 5 weeks, the animals were sacrificed and the results (number and grade of intraperitoneal adhesions, histological data on prosthesis integration, such as mesothelialization, fibroblast infiltration, vessel neoformation, etc.) were evaluated. RESULTS: Intraperitoneal adhesions decreased in implants painted with fibrin glue and hyaluronidase gel compared with untreated implants. When right-sided adhesions formed, they were looser and in many animals, the implants were completely peritonized. Integration of the prostheses was not affected by either fibrin glue or hyaluronidase gel. CONCLUSIONS: Adhesion formation can be reduced after abdominal surgery. The reduction achieved in this study was greater in the quantity than in the consistency of the adhesions. The results with hyaluronidase gel were moderately superior to those obtained with fibrin glue. Hyaluronidase gel has the advantage of being inexpensive.

Trasplante hepático Split para 2 adultos / No disponible

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Fístula arteriovenosa postraumática / Posttraumatic arteriovenous fistula

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Prevención de adherencias peritoneales a las prótesis intraperitoneales. Estudio experimental en cerdos / Prevention of peritoneal adhesions to intraperitoneal prostheses. An experimental study in pigs

Introducción. El objetivo de este estudio es investigar el efecto de la cola de fibrina y del gel de hialuronidasa en la prevención de adherencias peritoneales a las prótesis intraperitoneales. Material y método. En este estudio hemos utilizado 20 cerdos, divididos en 2 grupos: en todos los animales se procedió a colocar implantes de 4 x 4 cm: 2 de malla de polipropileno en una posición más cefálica, y otros dos de politetrafluoroetileno (Dualmesh® Plus Corduroy) en una posición más caudal. Los implantes situados en el lado derecho del animal se impregnaron de inhibidores de la producción de adherencias (en 10 animales se utilizó cola de fibrina, serie A, y en otros 10 se utilizó gel de hialuronidasa, serie B). Después de 5 semanas, se procedió al sacrificio de los animales y se evaluaron los resultados (cantidad y calidad de las adherencias formadas, así como datos histológicos de integración de las prótesis, como mesotelización infiltración por fibroblastos, vasos neoformados, etc.). Resultados. Al cabo de 5 semanas se apreciaba que los implantes impregnados de sustancias inhibidoras de la producción de adherencias presentaban menos adherencias, éstas (cuando existían) eran más laxas, e incluso en muchos casos los implantes estaban perfectamente peritonizados. La integración de las prótesis no estaba afectada por la presencia de los inhibidores. Conclusiones. La formación de adherencias puede disminuirse tras la cirugía abdominal. La disminución conseguida es mayor en la cantidad que en la consistencia de adherencias. Los resultados son algo mejores en la serie en la que se utilizó hialuronidasa que en la que se utilizó cola de fibrina. La hialuronidasa tiene la ventaja de tener un menor coste (AU)

Introduction. The aim of this study was to investigate the effect of fibrin glue and hyaluronidase gel on the prevention of postoperative peritoneal adhesions to intraperitoneal prostheses. Material and method. Twenty pigs, divided in two groups, were included. In all animals, four implants (4 x 4 cm) were placed: two polypropylene mesh implants were placed in an upper location and two polytetrafluoroethylene (PTFE) implants (Dualmesh Plus Corduroy) were placed in a lower position. Implants located in the right side of the animals were painted with fibrin glue (group A, n = 10) or with hyaluronidase gel (group B, n = 10). After 5 weeks, the animals were sacrificed and the results (number and grade of intraperitoneal adhesions, histological data on prosthesis integration, such as mesothelialization, fibroblast infiltration, vessel neoformation, etc.) were evaluated. Results. Intraperitoneal adhesions decreased in implants painted with fibrin glue and hyaluronidase gel compared with untreated implants. When right-sided adhesions formed, they were looser and in many animals, the implants were completely peritonized. Integration of the prostheses was not affected by either fibrin glue or hyaluronidase gel. Conclusions. Adhesion formation can be reduced after abdominal surgery. The reduction achieved in this study was greater in the quantity than in the consistency of the adhesions. The results with hyaluronidase gel were moderately superior to those obtained with fibrin glue. Hyaluronidase gel has the advantage of being inexpensive (AU)

Hemorragia digestiva baja por angiomatosis mesentérica masiva como inicio del síndrome de Klippel-Trénaunay-Weber / Lower digestive hemorrhage by masive mesenterial angiomatosis as an onset of Klippel-Trénaunay-Weber syndrome

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