Role of Fas/FasL signaling in regulation of anti-viral response during HSV-2 vaginal infection in mice

Fas receptor–Fas ligand (FasL) signaling is involved in apoptosis of virus-infected cells but increasing evidence accumulates on Fas receptor as a mediator of apoptosis-independent processes such as induction of activating and pro-inflammatory signals. In this study, we examined the role of Fas/FasL pathway in regulation of anti-viral response to genital HSV-2 infection using a murine model of HSV-2 infection applied to C57BL6/J, B6. MRL-Faslpr/J and B6Smn.C3-Faslgld/J mice. HSV-2 infection of Fas- and FasL-deficient mice led to decreased migration of IFN-γ expressing NK cells and CD4+ T cells, but not of γδ T cells, into the vaginal tissue. The vaginal tissues of HSV-2 infected Fas- and FasL-deficient mice showed increased production of IL-10, followed by low expression of the early CD69 activation marker on CD4+ and CD8+ T cells and increased numbers of regulatory T cells (Tregs). Experiments in co-cultures of CD4+ T cells and bone marrow derived dendritic cells showed that lack of bilateral Fas–FasL signaling led to expansion of Tregs and increased production of IL-10 and TGF-β1. Our results demonstrate that Fas/FasL can regulate development of tolerogenic dendritic cells and expansion of Tregs early during HSV-2 infection, which further influences effective anti-viral response.     

Interplay between immune responses to HLA and non-HLA self-antigens in allograft rejection

Recent studies strongly suggest an increasing role for immune responses against self-antigens (Ags) which are not encoded by the major histocompatibility complex in the immunopathogenesis of allograft rejection. Although, improved surgical techniques coupled with improved methods to detect and avoid sensitization against donor human leukocyte antigen (HLA) have improved the immediate and short term function of transplanted organs. However, acute and chronic rejection still remains a vexing problem for the long term function of the transplanted organ. Immediately following organ transplantation, several factors both immune and non immune mechanisms lead to the development of local inflammatory milieu which sets the stage for allograft rejection. Traditionally, development of antibodies (Abs) against mismatched donor HLA have been implicated in the development of Ab mediated rejection. However, recent studies from our laboratory and others have demonstrated that development of humoral and cellular immune responses against non-HLA self-Ags may contribute in the pathogenesis of allograft rejection. There are reports demonstrating that immune responses to self-Ags especially Abs to the self-Ags as well as cellular immune responses especially through IL17 has significant pro-fibrotic properties leading to chronic allograft failure. This review summarizes recent studies demonstrating the role for immune responses to self-Ags in allograft immunity leading to rejection as well as present recent evidence suggesting there is interplay between allo- and autoimmunity leading to allograft dysfunction.     

Leukoreduction system chambers are an efficient,valid, and economic source of functional monocyte-derived dendritic cells and lymphocytes

The demand for human monocyte-derived dendritic cells (moDCs), as well as for primary human B and T lymphocytes for immunological research purposes has been increased in recent years. Classically, these monocytes are isolated from blood, leukapheresis products or buffy coats of healthy donors by plastic adherence of peripheral blood mononuclear cells (PBMCs), followed by stimulation with granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-4, while lymphocytes are usually isolated from the non-adherent fraction (NAF) by magnetic cell sorting. However, donor-blood is a limited resource and not every blood bank offers leukapheresis products or buffy coats for laboratory use. Additionally, a leukapheresis is very expensive and also the generation/isolation of cells is time- and cost-intensive. To overcome some of these obstacles, we evaluated if low-cost leukoreduction system chambers (LRSCs), which arise after routine donor plateletpheresis procedures, and are usually discarded, would be an alternative and appropriate source of PBMCs to generate moDCs and to isolate lymphocytes. By analyzing the number and phenotype of immature and mature dendritic cells (DCs), as well as of B and T lymphocytes derived from LRSCs, we found all cells to be of high quantity and quality. Further investigations on DCs comprising transwell migration assays, allogeneic mixed lymphocyte reactions (MLR), cytokine secretion assays, and cytotoxic T cell induction assays revealed high migratory, as well as stimulatory capacity of these cells. In addition, DCs and T cells were efficiently electroporated with mRNA and showed characteristic cytokine production after co-culture, demonstrating LRSCs as an efficient, valid, and economic source for generation of moDCs and lymphocytes for research purposes.     

Antigen presentation by proliferating thymic macrophages to A (T,G)-A-L specific T cell line in an H-2 restricted manner

Long-term cultures of murine homogeneous populations of Ia-bearing thymus-derived murine macrophages were tested for their ability to present antigen to a (T,G)-A-L specific IL-2-dependent continuous T cell line. Thymus-derived macrophages, with and without pretreatment for Ia induction, triggered efficiently antigen-specific T cell proliferation in an MHC restricted way. This experimental system, consisting of two normal proliferating homogeneous populations of macrophages and antigen specific T cells, provides an ideal tool for studying the mechanism of antigen presentation to T cells and for elucidation of the role of macrophages in T-B cell collaboration for antibody production.     

Physiological organization of immune response based on the homeostatic mechanism of matrix reprogramming: Implication in tumor and biotechnology

It is accepted that the immune system responds to pathogens with activation of antigen-independent innate and antigen-dependent adaptive immunity. However many immune events do not fit or are even inconsistent with this notion. We developed a new homeostatic model of the immune response. This model consists of four units: a sensor, a regulator, an effector and a rehabilitator. The sensor, macrophages or lymphocytes, recognize pathogenic cells and generate alarm signals. The regulator, antigen-presenting cells, Тregs and myeloid-derived suppressor cells, evaluate the signals and together with sensor cells program the effector. The effector, programmed macrophages and lymphocytes, eliminate the pathogenic cells. The rehabilitator, M2 macrophages, restrict inflammation, provide angiogenesis and reparation of tissue damage, and restore the homeostasis. We suggest the terms “immune matrix” for a biological template of immune responses to pathogens and “matrix reprogramming” for the interdependent reprogramming of different cells in the matrix. In an adequate immune response, the matrix forms a negative feedback mechanism to support the homeostasis. We defined the cellular and phenotypic composition of a tumor immune matrix. A tumor reprograms the homeostatic negative feedback mechanism of matrix into a pathogenic positive feedback mechanism. M2 macrophages play a key role in this transformation. Therefore, macrophages are an attractive target for biotechnology. Based on our hypotheses, we are developing a cell biotechnology method for creation of macrophages with a stable antitumor phenotype. We have shown that such macrophages almost doubled the survival time of mice with tumor.