Serono Research Grants, 2002
Field of Endocrinology and Metabolism
Research Project:
Ex vivo expansion of pancreatic autoantigen-specific CD4+ CD25+ immunoregulatory t lymphocytes. Study of their potential as natural immunosuppressants in the treatment of autoimmune diabetes after allogeneic islet transplant.
Main researcher:
Dra. Concepción Mora Giral
Project Summary:
Type 1 diabetes mellitus is an autoimmune disease caused by selective destruction of insulin pancreatic beta cells by the immune system. The animal model for Type 1 diabetes mellitus that is closest to the same type of diabetes in humans is the NOD (Non Obese Diabetic) murine model. In any case, the initial factor or factors that trigger this immunological attack of beta cells is unknown. Recovering glucidic homeostasis in these patients requires exogenous supply of insulin. Nevertheless, the desired normoglycemia is not achieved at all times of day and this treatment is insufficient in order to avoid the occurrence of chronic degenerative complications. Replacement of the beta cells destroyed is necessary, and the most viable method in order to achieve beta cell mass restoration today is by means of a pancreas transplant. Transplanting isolated beta cells or islets offers theoretical advantages with respect to whole organ transplant, such as reduced morbidity and mortality, since no major surgery is required, and above all it allows designing strategies in order to avoid graft rejection or autoimmune destruction (with prior islet microencapsulation, antigen-specific immunosuppression with autologous regulatory T-cells, as proposed in the present application).
The average beta cell mass in order for a type 1 diabetic patient to become normoglycemic after transplant is around 1 million islets. This amount of islets is too high with respect to a normal pancreas (600000 islets), very probably due to factors related to replication, apoptosis, revascularization and post-transplant innervation.
After the transplant, in the absence of immunosuppressants, there is a high probability of a fresh outbreak of diabetes due to graft rejection and/or to a recurrence of the autoimmune process. In order to avoid this, the most common treatment consists in administering non-organ-specific immunosuppressant drugs that act by preventing the different polyclonal responses to autoantigens and alloantigens (including allografts and pathogenic antigens, etc). However, certain immunosuppressant drugs (corticoids, cyclosporine and tacrolimus) have a clearly defined diabetogenic effect. Selective elimination of certain T cell subpopulations in vivo may break the tolerance towards autoantigens (autotolerance) and consequently trigger autoimmune diseases. Tolerance towards autoantigens in healthy individuals can be understood as the result of 4 mechanisms acting simultaneously in order to counteract the existence of autoaggressive T cells: clonal deletion, anergy, immunological ignorance and immunosuppression or immunoregulation of autoaggressive T cells. Autoimmunity appears when the balance is broken between these four mechanisms.
The regulatory (or immunosuppressant) T cell subpopulation best characterized in mice and humans has been the CD4+CD25+ T cell subpopulation. These surface markers define a population of T cells that constitutively expresses the IL2 receptor α chain (CD25) on its surface. This CD4+CD25+ T lymphocyte subpopulation seems to appear as a different lineage. It does not proliferate in response to TCR (anergy) activation and it inhibits the proliferation of CD4+CD8- or CD4-CD6+ cells. The immunosuppressant behavior of CD4+CD25+ lymphocytes requires the presence of the costimulating molecule CTLA-4 on the cell surface and can be reversed in vitro using high doses if IL-2 or using anti-CD28 agonist antibodies. It is interesting to note that in between 5% and 10% of TCD4+ peripheral cells in non-immunized healthy mice correspond to this regulatory phenotype. When this subpopulation of CD4+CD25+ T lymphocytes is eliminated in certain mouse strains predisposed to autoimmune processes, these mice develop several types of autoimmune diseases.
Interestingly, the existence of CD4+CD25+ regulatory T cells has also been demonstrated in humans.
The aim of the present project is to eliminate the need for non-specific immunosuppressant treatment after transplant by the infusion of autologous (one’s own) pancreatic antigen-specific regulatory T cells.
Given the important harmful effects associated with the long term non-specific immunosuppressant treatment that currently takes place after a transplant, we propose a strategy for developing selective immunosuppression for islet antigen-specific T lymphocytes which prevents recurrence of the autoimmune process without forcing a generalized immunosuppression. One of the best alternatives to immunosuppressive drugs is the use of autologous (one’s own) regulatory T cells that are naturally present in humans and mice and therefore also in candidates to islet transplants. The best characterized regulator cells are CD4+ T lymphocytes, which constitutively express marker CD25 on the surface. We propose isolating this lymphocyte subpopulation from NOD mice thymus, spleen and pancreatic lymph nodes, expanding in vitro the CD4+CD25+ T cells specific for insulin (beta cell specific autoantigen) and verifying that they keep their immunosuppressive properties after in vitro expansion. Insulin-specific CD4+CD25+ cells thus expanded will be reinfused in pre-diabetic NOD mice or mice that have undergone an islet transplant under the renal capsule. These insulin specific CD4+CD25+ lymphocytes should be housed only in the pancreatic islets and should cause immunosuppression in the insulin environment, thus avoiding the autoimmune process.
Working hypothesis: We propose that the CD4+CD25+ regulatory T cells isolated from thymus, spleen and nodes of NOD mice can be cultivated in vitro in the presence of IL-2 and isogenic antigen-presenting cells (APCs) loaded with pancreatic autoantigens (insulin). Once expanded in vitro and having verified their immunomodulatory properties, the enrichment with these regulatory T cells that recognize the antigens in beta cells will favor their organ-specific action in the pancreatic organ or under the kidney capsule in which the islet graft is located after the transplant. This organ (islet) and antigen-specific location of the immunoregulatory cells will allow specific immunosuppression of autoaggressive lymphocytes strictly limited to the pancreatic medium-graft and would avoid all the problems derived from the generalized immunosuppression that is currently used.
Jury:
Field of Clinical Research in Endocrinology and Metabolism
Research Project:
Research Project:
Study on the relation between the deficit in insulin-like growth factor (IGF-1) and alterations in the hypothalamo-hypophysogonadal axis. Insulin-sensibility and bone metabolism parameters in males with cirrhosis before and after the hepatic transplant.
Main researcher:
Dr. Francisco Javier Salvador Rodríguez
Project Summary:
Project summary:
Severe hepatic failure occurring with clinical and analytical stigmas of hypogonadism associated with osteoporosis and disorders in hydrocarbonated metabolism. The elements implied in the physiopathology of these disorders are not well known. Both alterations in sex hormone metabolism and in the factors regulating phosphocalcium metabolism are potential candidates. There is experimental evidence according to which reduction in the IGF-1 concentration that accompanies hepatic cirrhosis is related with these alterations that can be corrected by means of exogenous administration of IGF-1. The influence of IGF-1 on steroidogenesis, spermatogenesis and bone mineralization is well known. Furthermore, this hormone whose circulatory concentration comes mainly from its hepatic synthesis has an anabolizing effect with respect to the hydrocarbonated metabolism. Even when a partial improvement of hypogonadism has been demonstrated after performing a hepatic transplant, there are no studies in humans that have demonstrated what element is implicated in this variation.
The comparison between the pre- and post transplant situations represents a superb study model in order to know what mechanisms are involved in the alterations derived from severe hepatic failure. The present study is designed for evaluating the role of IGF-1 and its transporting proteins as a mediator in the recovery of post-hepatic transplant biological functions. The results may constitute the base for contemplating the administration of IGF-1 as a replacement treatment of hepatic failure.
Jury:
Field of Reproduction
Research Project:
Study of the expression of sex hormone transport protein (SHBG) and estrogen receptor beta (ER-B) during spermatogenesis and its possible implication in masculine fertility disorders.
Main researcher:
Dra. Francina Munell Casadesús
Project Summary:
The regulation of spermatogenesis progression by androgens has been widely demonstrated although the mechanism by which they exert their action is not yet known. The absence of androgen receptors in germ cells along with the demonstration that these cells have P450 aromatase activity and beta estrogen receptors (ERβ) suggests that part of the actions of androgens in spermatogenesis are mediated by estrogens. On the other hand, the study of transgenic mice showing overexpression in human sex hormone transport protein (SHBG) has shown the existence of an alternative SVG protein (Alt-SHGB) in the human testicle produced from an intracellularly located alternative promoter. The alternative SVG protein has been identified in the acrosome of the mature sperm both in the transgenic mouse for human SHBG as in human semen. The fact that this alternative protein keeps the hormone binding region suggests that it may also be involved in the action of estrogens in germ cells.
The aims of the present project are, on the one hand, the study of ERβ and alternative SHBG isoform expression in the human testicle and, on the other hand, the analysis of their expression in males with fertility problems. Specifically, we propose identifying and localizing the expression of messenger RNH and Alt SHBG, ERβ and ERβ co proteins(ERβ variant produced by alternative splicing) in normal adult testicles and during their development and in human semen, as well as the cell compartment in which they perform their action, verifying the binding of Alt SHBG and ERβ proteins to estradiol in the spermatozoid and studying the variations of the expression of Alt SHBG, ERβ and ERβ co in males with fertility problems.
Jury:
Field of Multiple Sclerosis Clinical Research
Research Project:
Determination of the expression of type I interferon receptor (IFNAR) in patients with multiple sclerosis treated with interferon beta and its relationship with the reduction of interferon therapeutic efficacy.
Main researcher:
Dr. Oscar Fernández Fernández
Project Summary:
Multiple sclerosis (MS) is a neurodegenerative disease for which different treatments are being tested, amongst which IFN-beta has shown to be effective in a high percentage of patients. However, there are patients in which this beneficial effect is not produced and all the efforts of our group in recent years are directed towards discovering the possible causes of this reduction of therapeutic efficacy of IFN-beta. One of the reasons could be the presence of neutralizing antibodies against IFN-beta, but these studies performed by us and other groups have not been able to clearly demonstrate their participation in a loss of effectiveness.
Aim: To determine in a cohort of patients with multiple sclerosis treated with different types of IFN-beta whether the reduction in therapeutic effectiveness of IFN-beta is due to differences in the expression of the cell receptor for the FINAR interferon on the lymphocyte surface. Furthermore, it will be determined whether there are differences in the expression of each one of the subunits (IFNAR1 and IFNAR2) conforming the receptor. All this correlates with the clinical state of the patient and the presence of neutralizing antibodies against IFN-beta in samples from said patients.
Scope of the study: Neurology Department of the Hospital Complex Carlos Haya in Malaga. Laboratory attached to the Investigation Unit of H.C. Carlos Haya.
Methodology: The clinical activity of the disease will be registered in a protocol along the follow-up period. Determination of the IFN type 1 receptor will be performed by quantification of the expression of their mRNA in mononuclear cells by real time RT-PCR.
Jury:
