Antitumoral gene-based strategy involving nitric oxide synthase type III overexpression in hepatocellular carcinoma.

Hepatocellular carcinoma develops in cirrhotic liver. The nitric oxide (NO) synthase type III (NOS-3) overexpression induces cell death in hepatoblastoma cells. The study developed gene therapy designed to specifically overexpress NOS-3 in cultured hepatoma cells, and in tumors derived from orthotopically implanted tumor cells in fibrotic livers. Liver fibrosis was induced by CCl4 administration in mice. The first-generation adenoviruses were designed to overexpress NOS-3 or green fluorescent protein, and luciferase complementary DNA under the regulation of murine alpha-fetoprotein (AFP) and Rous Sarcoma Virus (RSV) promoters, respectively. Both adenovirus and Hepa 1-6 cells were used for in vitro and in vivo experiments. Adenoviruses were administered through the tail vein 2 weeks after orthotopic tumor cell implantation. AFP-NOS-3/RSV-luciferase increased oxidative-related DNA damage, p53, CD95/CD95L expression and caspase-8, -9 and -3 activities in cultured Hepa 1-6 cells. The increased expression of CD95/CD95L and caspase-8 activity was abolished by Nω-nitro-l-arginine methyl ester hydrochloride, p53 and CD95 small interfering RNA. AFP-NOS-3/RSV-luciferase adenovirus increased cell death markers, and reduced cell proliferation of established tumors in fibrotic livers. The increase of oxidative/nitrosative stress induced by NOS-3 overexpression induced DNA damage, p53, CD95/CD95L expression and cell death in hepatocellular carcinoma cells. The effectiveness of the gene therapy has been demonstrated in vitro and in vivo.

CD34+ circulating progenitor cells after different training programs.

Circulating progenitor cells (CPC) are bone marrow-derived cells that are mobilized into the circulation. While exercise is a powerful mediator of hematopoiesis, CPC levels increase, and reports of their activation after different types of exercise are contradictory. Moreover, few studies have compared the possible effects of different training programs on CPC concentrations. 43 physically active healthy male subjects (age 22±2.4 years) were assigned to 4 different training groups: aerobic, resistance, mixed and control. Except for the control group, all participants trained for 6 weeks. Peripheral blood samples were collected through an antecubital vein, and CPC CD34(+) was analyzed on different days: pre-training, post-training, and 3 weeks after finishing the training period. While no significant differences in CPC were observed either within or between the different training groups, there was a tendency towards higher values post-training and large intra- and intergroup dispersion. We detected an inverse linear relationship between pre-training values and % of CPC changes post-training (p<0.001). In the CPC values 3 weeks after training this inverse relationship was maintained, though to a lower extent (p<0.001). No changes in CPC CD34(+) were detected after 6 weeks of different training groups, or after 3 weeks of follow-up.

Radiofrequency-assisted transection of the pancreas versus stapler in distal pancreatectomy: study protocol for a multicentric randomised clinical trial (TRANSPAIRE).

INTRODUCTION: To date, no pancreatic stump closure technique has been shown to be superior to any other in distal pancreatectomy. Although several studies have shown a trend towards better results in transection using a radiofrequency device (radiofrequency-assisted transection (RFT)), no randomised trial for this purpose has been performed to date. Therefore, we designed a randomised clinical trial, with the hypothesis that this technique used in distal pancreatectomies is superior in reducing clinically relevant postoperative pancreatic fistula (CR-POPF) than mechanical closures. METHODS AND ANALYSIS: TRANSPAIRE is a multicentre randomised controlled trial conducted in seven Spanish pancreatic centres that includes 112 patients undergoing elective distal pancreatectomy for any indication who will be randomly assigned to RFT or classic stapler transections (control group) in a ratio of 1:1. The primary outcome is the CR-POPF percentage. Sample size is calculated with the following assumptions: 5% one-sided significance level (α), 80% power (1-ß), expected POPF in control group of 32%, expected POPF in RFT group of 10% and a clinically relevant difference of 22%. Secondary outcomes include postoperative results, complications, radiological evaluation of the pancreatic stump, metabolomic profile of postoperative peritoneal fluid, survival and quality of life. Follow-ups will be carried out in the external consultation at 1, 6 and 12 months postoperatively. ETHICS AND DISSEMINATION: TRANSPAIRE has been approved by the CEIM-PSMAR Ethics Committee. This project is being carried out in accordance with national and international guidelines, the basic principles of protection of human rights and dignity established in the Declaration of Helsinki (64th General Assembly, Fortaleza, Brazil, October 2013), and in accordance with regulations in studies with biological samples, Law 14/2007 on Biomedical Research will be followed. We have defined a dissemination strategy, whose main objective is the participation of stakeholders and the transfer of knowledge to support the exploitation of activities. REGISTRATION DETAILS: ClinicalTrials.gov Registry (NCT04402346).

Prevalence of hepatitis A and B markers and vaccine indication in cirrhotic patients evaluated for liver transplantation in Spain.

Vaccination against hepatitis A virus (HAV) and hepatitis B virus (HBV) is generally recommended for patients with chronic liver disease and those evaluated for liver transplantation in the absence of immunity. HAV and HBV infections after liver transplantation are frequent and associated with a worse prognosis. The data suggest that the number of patients with chronic liver disease without naturally acquired immunity against HAV and HBV is substantial, and that new vaccination strategies are needed. The aim of this study was to determine the level of immunity from hepatitis A and B infections and the need for HBV and HAV vaccination among cirrhotic patients evaluated for liver transplantation. We studied HBV and HAV serological markers (HbsAg, anti-HBc, anti-HBs, IgG anti-HAV) in 451 cirrhotic patients evaluated for liver transplantation to investigate the association with gender, age, and etiology of cirrhosis. Negative HBV markers were observed in 57% of patients with 43% displaying one positive HBV marker: HBsAg (+), 9.5%; anti-HBc (+)/anti-HBs (-), 11.5%; anti-HBc (-)/anti-HBs(+), 4.2%; anti-HBc(+)/anti-HBs(+), 17.7%. HBV vaccine indication established in 68.5% of patients was greater among women and hepatitis C virus-negative patients. No differences were observed in age or cause of cirrhosis. HAV vaccination indicated in 6.7% of patients (IgG anti-HVA-negative) was greater among patients with negative HBV markers (9.3% vs 3.3%, P = .018) and younger patients (25.3% of patients

Evaluation of two commercial enzyme immunoassays for the detection of norovirus in faecal samples from hospitalised children with sporadic acute gastroenteritis.

Two commercially available enzyme immunoassays (EIAs), IDEIA and Ridascreen, for norovirus antigen detection were evaluated with 117 faecal samples from hospitalised children with acute gastroenteritis. Eighteen of 39 samples positive by RT-PCR were characterised by sequence analysis, and 17 of these were related to norovirus genogroup II. When compared with RT-PCR, the sensitivity and specificity values were 76.9% and 85.9%, respectively, for the IDEIA assay, and 59.0% and 73.1%, respectively, for the Ridascreen assay. The sensitivity and specificity of both EIA tests require improvement, but they could both eventually be of use in the diagnosis of norovirus diarrhoea in clinical laboratories.

[Role of malnutrition in intestinal anastomosis collagenization: an analysis of procollagen (PINP) and carboxyterminal telopeptide (ICTP) by radioimmunoassay]. / Efectos de la desnutrición sobre la colagenización de anastomosisintestinales: análisis de procolágeno y telopéptido carboxiterminalmediante radioinmunoanálisis.

INTRODUCTION: some clinical, anatomo-pathological, and technical factors influence the correct healing of intestinal suture following an intestinal resection. One of the most influential factors is patient nutritional status. OBJECTIVES: to evaluate the influence of malnutrition on the viability of primary intestinal anastomosis by the analysis of collagen I deposition. METHODS: 40 Wistar rats, radioimmunoassay material. We used 2 groups of rats, 20 animals in each group: a control group (A) and a "malnutrition" group (B). RESULTS: there was a decrease in PINP (procollagen) deposition in the colon of group B rats as compared to the colon of group A (0.3620 and 0.4340 mg/g respectively) (p = 0.032). There is an increase in ICTP (carboxyterminal telopeptide) in the colon of group B (0.9545 as against 0.8460 mg/g in group A) (p = 0.875). In anastomoses of group B there was a decrease in PINP synthesis as compared to group A (0.376 and 0.468 mg/g respectively, p = 0.002). As regards ICTP, there was an increase in group B (p = 0.330). In relation to the control group no differences were observed in ICTP increases in group B (p = 1). CONCLUSIONS: colonic anastomosis increases the levels of PINP and ICTP in healed tissue (p = 0.000); malnutrition reduces collagenization in anastomoses (p = 0.000).

Morbidity and mortality in liver retransplantation.

INTRODUCTION: The incidence of orthotopic liver retransplantation (re-OLT) ranges from 6% to 11%. The most frequent causes of early re-OLT are allograft failure, uncontrolled acute rejection, and vascular complications. MATERIALS AND METHODS: A retrospective study of 512 orthotopic liver transplants (OLTs) in 482 patients over 15 years. RESULTS: The incidence of re-OLT was 6.6%, with a higher percentage of men requiring re-OLT than first-time OLT (75.0% vs 63.0%, P < .05). The reasons for re-OLT were thrombosis 21.7%, aneurysm 6.5%, stenosis 3.2%, primary nonfunction (PNF) 21.7%, and chronic rejection or recurrence of the initial disease 40.4%. Complications included PNF (22.0%), acute renal failure (65.6%), postoperative infection (87.5%), and adult respiratory distress syndrome (9.4%; P < .05). No differences were seen in the incidence of septicemia or postoperative hemorrhage. The average survival was much lower in re-OLT (21.8 days) compared with OLT (194.5 days; P < .05). The mortality rates in re-OLT were 100% for primary biliary cirrhosis, 85.7% for HCV, 50% for alcoholic cirrhosis, and 20% for HBV. A direct association between the Model for End-stage Liver Disease (MELD) score and the number of complications was present. DISCUSSION: There was a greater requirement for re-OLT in men and those patients transplanted due to hepatitis B virus cirrhosis and fulminant hepatitis (P < .05). The re-OLT patients had no greater incidence of sepsis compared with the OLT patients, although they did have a greater incidence of primary graft dysfunction, acute renal failure, adult respiratory distress syndrome, and postoperative infection (P < .05). The MELD was a good parameter for predicting graft evolution. Re-OLT in patients with primary biliary cirrhosis and hepatitis C virus was associated with a high degree of mortality.

Survey of hemostasis management and transfusion in liver transplantation.

OBJECTIVE: To determine the management of haemostasis and transfusion practice in the field of liver transplantation in Spain. METHODS: A questionnaire was developed for physicians in anaesthesiology of all centres performing liver transplantation in Spain. The information required made reference to the 12 months prior to its distribution, from January 1 to December 31, 2011. RESULTS: Data were collected from 24 centres in which liver transplantation is performed in Spain. Only 46% reported that they had protocols or practice guidelines for the management of haemostasis, and 83% of hospitals responded that they knew the percentage of transfused patients, but only 57% knew the mean transfusion. Regarding the degree of satisfaction with the management of haemostasis/coagulation, 50% said they were not satisfied. Thromboelastometry was used as an additional method of preoperative monitoring in only 8% of the centres and intra-operatively in one-third. Less than half (46%) of the centres performed preoperative correction of coagulation deficits based on conventional tests. The mean number of packed red cells used was ≤4 in 57% of centres. Consumption of fresh frozen plasma was highly variable, while 100% of centres consumed less than 4 pools of platelets per patient. CONCLUSIONS: There is a wide variability in the management of haemostasis and transfusion practice among Spanish centres. There are no guidelines or they are not widely used. The mean use of transfused blood products remain high. There was a decrease in centres using new methods of monitoring.

Mycotic pseudoaneurysms after liver transplantation.

The most frequent etiology of visceral artery aneurysms is arteriosclerosis, but vascular manipulation during hepatic transplantation may also cause a mycotic pseudoaneurysm. Treatment with embolization, stents or percutaneous thrombin injection have been recommended but surgical revascularization is indicated when interventional techniques fail. A 43-year-old man with hepatitis C virus cirrhosis who underwent orthotopic liver transplantation from a cadaveric donor was treated with cyclosporine, mycophenolate, and steroids and was discharged from hospital at 35 days. Two months later he was readmitted with a febrile syndrome. Abdominal computed tomography showed necrosis of hepatic segments IV, V, and VI. Magnetic resonance imaging and angiography revealed partial thrombosis of the hepatic artery and stenosis of the portal anastomosis secondary to an aneurysm of the hepatic artery. A few hours after the radiological diagnosis, the patient suffered a bout of upper gastrointestinal bleeding and shock. Emergency surgery revealed a mycotic pseudoaneurysm of the common hepatic artery, which had ruptured into the bile tract with hemobilia. The liver graft was removed because of severe necrosis of the right liver. The patient died awaiting a new liver transplantation.

Is "intra-operating room" thromboelastometry useful in liver transplantation? A case-control study in 303 patients.

Coagulation monitoring during liver transplantation (LT) is, even today, fundamental to reduce blood loss during surgery. Thromboelastometry (TEM) is a proven technique for controlling the various parameters that influence coagulation. However, there are no studies linking "intra-operating room" TEM (orTEM) with LT outcomes. We describe a case-control study in 303 liver graft recipients analyzing variables associated with operative complications and long-term LT outcomes. The results showed that orTEM reduced the use of blood products in patients with Model for End-Stage Liver Disease scores of ≥ 21, retransplantation, and high surgical difficulty and important intraoperative bleeding. In addition, results in survival and postoperative complications were better when orTEM was used. In conclusion, we confirm that use of orTEM is associated with less use of blood products and a lower rate of complications after LT.

Donor characteristics that are associated with survival in liver transplant recipients older than 70 years with grafts.

INTRODUCTION: The use of grafts from donors older than 70 years of age is increasing due to the decrease in the number of donors and the increase in waiting list patients. MATERIAL AND METHODS: We undertook a univariate and multivariate analysis of 980 adult recipients of whole liver grafts, 129 of them from donors aged 70 years or older. RESULTS: No differences were found in patient survival compared with recipients of younger grafts. There were no higher rates of rejection, vascular or biliary complications, postoperative bleeding, or infections, but older grafts were associated with graft dysfunction (P = .01) and a higher frequency of postoperative refractory ascites (P = .007), but without a greater need for retransplantation. As graft-associated factors, the joint presence in the donor of diabetes (P = .00; confidence interval [CI] = 0.04-0.117), hypertension (P = .00; CI = 0.22-0.39), and weight of more than 90 kg (P = .031; CI = 0.05-0.104) were suggestive of poor prognostic factors in recipient survival. Survival in hepatitis C virus (HCV) recipients or recipients aged older than 60 years was worse with donors aged older than 70 years, although not significantly so. With grafts from donors aged older than 80 years (n = 15), although patient survival rate was good (70% at 10 years), there was a higher rate of retransplantation (20%) and the early mortality rate was 13.3%. CONCLUSIONS: Use of grafts from donors aged older than 70 years is safe, with similar survival to patients with younger grafts. The appearance of initial dysfunction with prolonged ascites may be due to a delay in reaching a correct functionality, but was not associated with increased mortality, complications, or need for retransplantation. It should also be avoided in recipients older than 60 years or with HCV. Grafts older than 80 years were associated with a good long-term patient survival but at the expense of a higher rate of retransplantation. However, it helps to reduce the time on the waiting list and, thus, mortality. We noted decreased survival associated with donor hypertension, diabetes, and obesity, so these donors should be selected more rigorously.

Antitumor efficacy of mammalian target of rapamycin inhibitor therapy in liver transplant recipients with oncological disease: a case-control study.

INTRODUCTION: The reported incidences of de novo malignancy following orthotopic liver transplantation (OLT) are significantly greater than those in the general population. We have analyzed the efficacy of mammalian target of rapamycin inhibitor (mTORi) as immunosuppressant therapy in patients with de novo malignancies or those engrafted because of a primary liver cancer. METHODS: We performed a case-control study of patients with hepatocellular carcinoma (HCC; n = 119), cholangiocarcinoma (n = 1) or de novo malignancies (n = 73). Thirty-seven patients with these tumors were treated with mTORi, and 167, with calcineurin inhibitors (CNI). Switching to mTORi was performed progressively, withdrawing the CNI over 15 days, until obtaining levels of 5-10 ng/dL. RESULTS: No incidence of rejection, serious adverse events, or death was observed with an overall actuarial survival of 68.5% in the mTORi group versus 45.7% among the CNI group. Overall rates of tumor recurrence were 15.2% and 36.8%, respectively (P < .05). Among patients with HCC, survival was 100% of mTORi with and 61.5% among CNI patients, with tumor recurrence rates of 6.2% and 19.1%, respectively (P < .05). DISCUSSION: Surprising differences in survival and tumor recurrence rates were observed among the mTORi-treated group compared with controls. Switching from CNI to mTORi immunosuppressant therapy appeared to be safe. It seems to be reasonable to employ this strategy in liver transplant patients with primary hepatic or "de novo" neoplasms.

Results of liver transplantation with donors older than 70 years: a case-control study.

Older donors are a growing part of the total pool but no definite consensus exists on the age limit for their acceptance. This retrospective case-control unicenter study compared the outcomes of 72 orthotopic liver transplantations (OLTs) from April 1990 to April 2010 using donors older than 70 years versus 738 chronologically correlated OLTs performed with donors younger than 60 years. The percentage of refusal was greater among older than younger donors (48.2 vs 14.3%; P < .001). No difference was observed in mean cold ischemia times between older (370.5 minutes) versus younger groups (389.2 minutes). or in postoperative complications of rejection or renal insufficiency except for sepsis and mortality. Long-term survival was lower among transplant recipients from donors older than 70 years (P = .001) and these cases showed more blood requirements associated with prolonged cold ischemia (P = .02). Multivariate analysis revealed graft dysfunction, mortality, and reduced survival to be associated with donor weight and recipient MELD (Model for End-stage Liver Disease) (P < .05). Interestingly, the mortality related to hepatitis C virus recurrence was not greater among patients whose donors were older than 70. Septuagenarians' livers can be used safely, but careful donor and recipient evaluation are required to avoid additional risk factors.

Liver transplantation in patients with cryptogenic cirrhosis: long-term follow-up.

OBJECTIVE: The objective of this study was to evaluate long-term survival, histological diagnoses, and mobility of patients with cryptogenic cirrhosis (CC) treated with orthotopic liver transplantation (OLT). PATIENTS AND METHODS: We performed a retrospective analysis of 35 patients who underwent transplantation with CC among 800 OLT patients. There were no differences in gender, mean age of 47 years, average MELD (Model for End-stage Liver Disease) of 16, and hepatocellular carcinoma incidence (8%). RESULTS: In 28.6% of patients, the diagnosis of CC was wrong. There was no incidence of an acute rejection episode and a low incidence of complications, although the postoperative mortality rate was 20%, of chronic rejection was 25%, and recurrence of disease was 4%. Cumulative at 3-, 5-, and 10-year survivals were lower than the other OLT. Survival was lower in patients receiving suboptimal grafts. CONCLUSIONS: One of 3 patients who underwent transplantation for CC had a specific etiologic diagnosis. The chronic rejection rate and postoperative mortality rate were higher than other etiologies, and survivals at 5, 10, and 15 years were lower than other OLT.

Efficacy, tolerance, and safety of mammalian target of rapamycin inhibitors as rescue immunosuppressants in liver transplantation.

BACKGROUND: Mammalian target of rapamycin (mTOR) inhibitors behave as potent immunosuppressants which have the advantages, with respect to calcineurin inhibitors (CNI: cyclosporine or tacrolimus), of no nephrotoxicity and inhibition of cell proliferation. They are particularly suitable for patients with renal insufficiency or neoplasias. MATERIALS AND METHODS: Twenty-two liver transplant patients were immunosuppressed with everolimus or sirolimus as rescue therapy after CNI treatment: 7 hepatocellular carcinomas; 5 de novo malignancies; 4 renal insufficiencies; 4 chronic rejections; and 2 acute rejection episodes. RESULTS: There were 16.7% tumor recurrences, and 25% improvements in renal function, 75% in chronic rejection, and 50% in acute rejection. There was no incidence of rejection, kidney failure, gastrointestinal intolerance, hydrocarbon intolerance, hypertension, or arterial or venous thrombosis. We observed incidences of 50% for hypercholesterolemia, 31.8% for hypertriglyceridemia, 22.7% for thrombocytopenia, 18.2% for leukopenia, and 9.1% for anemia. The intercurrent infection rate was 13.6%, including oral thrush in 13.6%. Lower limb edema occurred in 13.6%, with 1 case of facial edema and 1 of alopecia. CONCLUSIONS: mTOR inhibitors were safe immunosuppressive drugs whose side effects were controlled and easily managed. They have advantages with respect to CNI due to their slight effects on kidney function and lack of promotion of diabetes mellitus. Although their long-term effectiveness for control of neoplastic diseases is yet to be seen, they can be used safely in these patients with no incidence of rejection. Their effectiveness to control chronic rejection seems significant, but it is doubtful for steroid-resistant acute rejection episodes.

Ictericia secundaria a metástasis pancreática de sarcoma de partes blandas / Obstructive jaundice secondary to pancreatic metastases of a soft-tissue sarcoma

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Efectos de la desnutrición sobre la colagenización de anastomosis intestinales: análisis de procolágeno y telopéptido carboxiterminal mediante radioinmunoanálisis / Role of malnutrition in intestinal anastomosis collagenization: an analysis of procollagen (PINP) andcarboxyterminal telopeptide (ICTP) by radioimmunoassay

Introducción: diversos factores influyen en la cicatrización correcta de las suturas intestinales tras la práctica de una resección intestinal. Uno de los factores más implicados es el estado nutricional del paciente. Objetivos: evaluar la influencia de la desnutrición inducida sobre la viabilidad de una anastomosis intestinal primaria mediante el análisis del procolágeno (PINP) como marcador de la síntesis de colágeno I, y del telopéptido carboxiterminal del colágeno I (ICTP) como marcador de la destrucción del mismo. Métodos: 40 ratas Wistar y material de radioinmunoensayo. Métodos: diseñamos 2 grupos de ratas, 20 animales por cada grupo: grupo control (A) y grupo “desnutrición” (B). Se analiza PINP e ICTP mediante RIA sobre tejido colónico homogeneizado, preanastomótico y anastomótico. Resultados: existen unos niveles menores de PINP en el colon de las ratas del grupo B comparado con el colon del grupo A (0,3620 y 0,4340 μg/g respectivamente) (p = 0,032). Hay un mayor nivel de ICTP analizado en el colon del grupo B (0,9545 en contraposición a 0,8460 μg/g en el grupo A) (p = 0,875). En las anastomosis del grupo B existe una menor síntesis de PINP en comparación con el grupo A (0,376 y 0,468 μg/g respectivamente, p = 0,002). Conclusiones: la anastomosis colónica incrementa los niveles de PINP e ICTP en el tejido cicatricial (p = 0,000); la malnutrición reduce la colagenización de las anastomosis (p = 0,000)

Introduction: some clinical, anatomo-pathological, and technical factors influence the correct healing of intestinal suture following an intestinal resection. One of the most influential factors is patient nutritional status. Objectives: to evaluate the influence of malnutrition on the viability of primary intestinal anastomosis by the analysis of collagen I deposition. Methods: 40 Wistar rats, radioimmunoassay material. We used 2 groups of rats, 20 animals in each group: a control group (A) and a “malnutrition” group (B). Results: there was a decrease in PINP (procollagen) deposition in the colon of group B rats as compared to the colon of group A (0.3620 and 0.4340 μg/g respectively) (p = 0.032). There is an increase in ICTP (carboxyterminal telopeptide) in the colon of group B (0.9545 as against 0.8460 μg/g in group A) (p = 0.875). In anastomoses of group B there was a decrease in PINP synthesis as compared to group A (0.376 and 0.468 mg/g respectively, p = 0.002). As regards ICTP, there was an increase in group B (p = 0.330). In relation to the control group no differences were observed in ICTP increases in group B (p = 1). Conclusions: colonic anastomosis increases the levels of PINP and ICTP in healed tissue (p = 0.000); malnutrition reduces collagenization in anastomoses (p = 0.000)