Leticia Fernández

Is Ischemic Preconditioning a Useful Strategy in Steatotic Liver Transplantation

This study examined the effect of preconditioning on steatotic livers for transplantation and attempted to identify the underlying protective mechanisms. Blood flow alterations, neutrophil accumulation, tumor necrosis factor α release and lipid peroxidation were observed in nonsteatotic livers after transplantation. Steatotic and nonsteatotic liver grafts were similar in their blood flow, neutrophil accumulation, and TNF release after transplantation. However, in the presence of steatosis, lipid peroxidation and hepatic injury increased. In addition, recipients of steatotic liver grafts were more vulnerable to lung damage associated with transplantation. The conversion of xanthine dehydrogenase to xanthine oxidase and the accumulation of xanthine during cold ischemia was greater in steatotic than in nonsteatotic liver grafts. The results obtained with xanthine oxidase inhibitors indicated that xanthine/xanthine oxidase could be responsible for the increased lipid peroxidation as well as the exacerbated liver and lung damage associated with transplantation of steatotic livers. Preconditioning reduced the xanthine accumulation and percentage of xanthine oxidase seen in steatotic liver grafts during cold ischemia, and conferred protection against liver and lung damage following transplantation. The benefits of preconditioning could be mediated by nitric oxide. These findings suggest that preconditioning could be a relevant new strategy to protect against the inherent risk of steatotic liver failure following transplantation.

The Combination of Ischemic Preconditioning and Liver Bcl-2 Overexpression Is a Suitable Strategy to Prevent Liver and Lung Damage after Hepatic Ischemia-Reperfusion

The present study evaluates the effectiveness of ischemic preconditioning and Bcl-2 overexpression against the liver and lung damage that follow hepatic ischemia-reperfusion and investigates the underlying protective mechanisms. Preconditioning and Bcl-2, respectively, reduced the increased tumor necrosis factor (TNF) and macrophage inflammatory protein-2 (MIP)-2 levels observed after hepatic reperfusion. Bcl-2 overexpression or anti-MIP-2 pretreatment seems to be more effective than preconditioning or anti-TNF pretreatment against inflammatory response, microcirculatory disorders, and subsequent hepatic ischemia-reperfusion injury. Furthermore, each one of these strategies individually was unable to completely inhibit hepatic injury. The combination of preconditioning and Bcl-2 overexpression as well as the combined anti-TNF and anti-MIP-2 pretreatment totally prevented hepatic injury, whereas the benefits of preconditioning and Bcl-2 were abolished by TNF and MIP-2. In contrast to preconditioning, Bcl-2 did not modify lung damage induced by hepatic reperfusion. This could be explained by the differential effect of both treatments on TNF release. Anti-TNF therapy or preconditioning, by reducing TNF release, reduced pulmonary inflammatory response, whereas the benefits of preconditioning on lung damage were abolished by TNF. Thus, the induction of both Bcl-2 overexpression in liver and preconditioning, as well as pharmacological strategies that simulated their benefits, such as anti-TNF and anti-MIP-2 therapies, could be new strategies aimed to reduce lung damage and inhibit the hepatic injury associated with hepatic ischemia-reperfusion.

Role of ischemic preconditioning and the portosystemic shunt in the prevention of liver and lung damage after rat liver transplantation.

Background. This study evaluates whether surgical strategies such as the portosystemic shunt and ischemic preconditioning can protect against hepatic and pulmonary injury associated with liver transplantation. Methods. The effect of the portosystemic shunt, ischemic preconditioning, and both surgical procedures together were evaluated in rat liver transplantation. Alanine aminotransferase, hyaluronic acid levels in plasma, adenosine triphosphate and nucleotide levels in liver and edema, malondialdehyde levels, and myeloperoxidase activity were measured 24 hr posttransplantation. Plasmatic tumor necrosis factor (TNF) levels were measured as a possible proinflammatory factor responsible for hepatic and pulmonary damage associated with liver transplantation Results. Hepatocyte and cell endothelial damage were observed in liver grafts subjected to 8 hr of cold ischemia. This was associated with increased plasma TNF levels and lung inflammatory response. Portosystemic shunt application in the recipient protected endothelial cells but did not confer an effective protection from hepatocyte damage or reduce the increased plasma TNF levels and lung damage after liver transplantation. However, preconditioning of the donor liver conferred protection against both the endothelial cell and hepatocyte damage observed after liver transplantation. Preconditioning also attenuated the increased plasma TNF release and pulmonary damage. The combination of both surgical strategies resulted in levels of liver injury, TNF, and lung damage similar to those seen after liver transplantation. Conclusions. These findings indicate that ischemic preconditioning could be a preferred treatment to reduce hepatic and pulmonary damage associated with liver transplantation. However, this strategy may not be effective in several clinical situations requiring a portosystemic shunt.

Assessment of synthetic chimeric multiple antigenic peptides for diagnosis of GB virus C infection

The use of synthetic peptides of both structural and nonstructural proteins of GB virus C (GBV-C) has been studied for the development of new systems to diagnose infection caused by this virus. In an attempt to increase the antigenicity of linear peptide sequences, chimeric multiple antigenic peptides (MAPs) containing epitopes from E2, NS4, and NS5 GBV-C proteins have been synthesized. The synthetic constructs were evaluated by ELISA to establish whether the epitopes in chimeric branched peptides are more efficiently recognized by the specific antibodies compared to the monomeric linear sequences. Moreover, we have investigated the application of a commercial biosensor instrument for the detection of antibodies against the GBV-C in human serum samples. The results of the immunoassays reported in this work highlight the usefulness of synthetic tetrameric branched peptides containing sequences from envelope and nonstructural GBV-C proteins for the diagnosis of GBV-C infection. The potential clinical value of the MAP(4)(E2-NS5a) for the serodiagnosis of GBV-C infection was demonstrated, thus providing the basis for performing prevalence studies of the infection among the hemodialyzed and hepatitis C virus (HCV)-infected population.

The TNF family member APRIL dampens collagen-induced arthritis

Background The tumour necrosis factor (TNF)-family members B cell activating factor (BAFF) and A PRoliferation-Inducing Ligand (APRIL) play important roles in B cell biology, and share binding to B cell maturation antigen and transmembrane activator and cyclophilin ligand interactor, both receptors of the TNF-family. However, while it is reported that BAFF can break B cell tolerance, the role of APRIL in autoimmunity remains elusive. Objective To evaluate the role of APRIL on collagen-induced arthritis (CIA). Methods CIA was induced in APRIL-transgenic (Tg) DBA/1 mice and littermates. Disease progression was evaluated by clinical and histological signs of arthritis. In another experimental setting mice were exposed to the collagen antibody-induced arthritis. In addition, we tested T cell dependent humoral responses in APRIL-Tg mice. Results We found that APRIL-Tg displayed a strongly reduced incidence and severity of CIA compared with littermates, with decreases in collagen-specific autoantibody titres, immune complex deposition and downstream mast cell activation in joints. Notably, ectopic APRIL-expression was also found to negatively regulate T cell dependent humoral responses. The lower autoantibody production in APRIL-Tg mice during CIA appears to be crucial, as arthritis induced by administration of anti-collagen antibodies developed similar in APRIL-Tg and control mice, thus demonstrating that the downstream effector pathways induced by anti-collagen antibodies remain intact in APRIL-Tg mice. This protective effect was specifically mediated by APRIL, as adenoviral delivery of APRIL decreased CIA in a therapeutic setting. Conclusions Collectively, our data identify APRIL as a negative regulator of CIA by regulating autoantibody production. These findings are of important clinical relevance, as the therapeutic potential of transmembrane activator and cyclophilin ligand interactor-Fc (atacicept) is presently evaluated in clinical trials.

Diagnostic Value of Anti-GBV-C Antibodies in HIV-Infected Patients

The beneficial effect of co‐infection by GB virus C (GBV‐C) in the course of the disease in human immunodeficiency virus (HIV)‐infected patients has been described, although its mechanism of action is yet to be determined. The role of anti‐GBV‐C antibodies in HIV‐infected patients also remains unknown. At present, there are no commercial systems to detect specific markers of GBV‐C infection. The research presented follows our previous work from which we obtained chimeric molecules formed by two domains of different GBV‐C proteins with good sensitivity/specificity balances in the detection of anti‐GBV‐C antibodies in hemodialyzed and chronic hepatitis patient samples. It has been investigated the ability of the synthetic peptides to recognize specific anti‐GBV‐C antibodies in HIV and HCV/HIV co‐infected patients by a peptide‐based ELISA immunoassay. The results showed that human immunodeficiency virus‐infected patients have a significantly higher frequency of anti‐GBV‐C antibodies than healthy controls. A comparison between HCV+/HIV+ and HCV−/HIV+ was analyzed. Although a higher percentage of HCV/HIV‐positive sera were positive for antibodies against GBV‐C peptides, the difference was not significant. The presence of anti‐GBV‐C antibodies could represent a good marker of exposure to GBV‐C in HIV‐infected patients to facilitate a further analysis of the effect of this exposure in the progression of illness caused by HIV infection.

APRIL facilitates viral-induced erythroleukemia but is dispensable for T cell immunity and lymphomagenesis

The TNF family member, a proliferation‐inducing ligand (APRIL), has been suggested to act as a costimulatory molecule in T cell responses. However, studies addressing this role in vivo are largely lacking. Here, we evaluated the effects of APRIL on physiological T cell responses in vivo. Although receptors for APRIL are expressed on a subset of T cells, neither TCR transgenic (Tg) T cell responses nor endogenous TCR responses were affected by Tg APRIL expression in vivo. Moreover, APRIL did not significantly enhance the induction of T cell lymphomas upon Moloney murine leukemia virus (MLV) infection. This clearly contrasts current belief and indicates that APRIL does not serve a major role in T cell immunity or lymphomagenesis. However, we did observe a strong increase in erythroleukemia formation after MLV inoculation of APRIL Tg mice. Strikingly, this erythroleukemia‐facilitating property of APRIL was confirmed using the erythroleukemogenic Friend‐MLV. Erythroleukemia in APRIL Tg mice was characterized by low hematocrits and grossly enlarged spleens with an increased percentage of erythroid precursors. Altogether, these results unveil new proerythroleukemogenic properties of APRIL.

Synthetic peptides derived from an N-terminal domain of the E2 protein of GB virus C in the study of GBV-C/HIV-1 co-infection.

Synthetic peptides derived from GB virus C (GBV‐C) have previously been studied in our group for the development of new systems capable of diagnosing diseases caused by this humanotropic virus. We also recently described specific peptide domains of the E2 envelop protein of GBV‐C that have the capacity to interfere with the HIV‐1 fusion peptide, produce a notable decrease in cellular membrane fusion, and perturb HIV‐1 infectivity in a dose‐dependent manner.

The TNF-ligand APRIL can control CIA by regulating antibody-production and stimulating anti-inflammatory IL-10 producing B cells

Background Increased levels of the tumour necrosis factor (TNF) -ligand APRIL (A Proliferation Inducing Ligand) were found in synovial fluid and serum of patients with inflammatory arthritis pointing to a pro-inflammatory role of APRIL. APRIL can bind to BCMA and TACI, two receptors of the TNF family, which can also bind the B cell activating factor (BAFF). In the collagen-induced arthritis (CIA), administration of TACI-Ig was found to prevent disease progression and to lower disease scores, compared with controls. As TACI binds both APRIL and BAFF, it remained to be determined, whether this effect was due to the capacity of TACI to block just one or both ligands. Methods CIA was induced in APRIL-transgenic (Tg) DBA/1 mice and littermates. Severity of disease was scored for each paw using a scale 0–4. In addition, mice were analysed for histological signs of arthritis. Anticollagen antibody titers were determined by ELISA. Lymphocyte populations in draining lymph nodes, spleen and peritoneum were analysed by FACS. In another experimental setting mice were exposed to the collagen antibodies induced arthritis (CAIA). In addition, we employed models fort contact hypersensitivity (CHS) and experimental autoimmune encephalomyelitis (EAE). For the former, APRIL Tg and control mice were sensitised and challenged at the ear with oxazolone. EAE was induced by immunising mice with MOG, a myelin peptide, in adjuvant. Results APRIL Tg mice displayed in contrast to littermates a lower disease score and incidence of arthritis, and also produced less collagen specific antibodies. Joints of littermates had higher IgG levels and increased number of degranulating, thus activated, mast cells. To confirm that the decreased IgG levels developed in the CIA model in APRIL Tg mice are directly linked to the less severe disease development, we employed the model of CAIA. In fact, disease development in CAIA was similar in APRIL Tg and control mice. In addition, we detected a significantly increased IL-10 production of peritoneal B cells in APRIL Tg mice in the CIA model. Evidence is accumulating that IL-10 producing B cells can regulate autoimmune diseases including arthritis. The regulatory role of APRIL by modulating activity IL-10-producing B was confirmed in the CHS and EAE model. Conclusion Our results show that APRIL can control inflammation in the three tested disease models CIA, CHS and EAE. This suggests a therapeutic potential of APRIL agonists to downregulate inflammatory diseases such as rheumatoid arthritis.