Browsing by Subject "tetramer"

Now showing 1 - 3 of 3
  • Thumbnail Image
    Item
    Examining Helper T-cell Recovery After Sepsis
    (2015-08) Cabrera-Perez, Javier
    Sepsis strikes 750,000 Americans every year with ~ 210,000 of these patients dying – far more than the number of deaths from prostate cancer, breast cancer, and AIDS combined. Some of these deaths occur during the acute, inflammatory stages of sepsis, but ~70% of these patients survive the initial infection, only to perish due to hospital-acquired infections. Most sepsis research has focused on understanding the acute, inflammatory stage of sepsis, but the increased susceptibility to secondary infections has led clinicians and researchers to believe that the chronic stage of sepsis is important and is characterized by immunosuppression. CD4 T-cells, essential for coordinating immune responses to opportunistic pathogens, are severely depleted during the acute stage of sepsis, but gradually recover throughout the immunosuppressive phase of sepsis. Despite the well-characterized immune cell apoptosis during sepsis, the impact of sepsis on protective T-cell responses (especially CD4 T-cells) against secondary pathogen challenge remains poorly understood. This dissertation presents a previously unappreciated mechanism of CD4 T-cell impairment during the immunosuppressive stage of sepsis. In the present study, we have studied sepsis immunosuppression by using Class II major histocompatibility complex tetramers to track endogenous, antigen specific CD4 T-cells, in order to examine a hypothesis: that the uneven recovery of the Ag-specific CD4 T-cell repertoire contributes to the alarming rate of infections in sepsis survivors. In addition, we have examined the impact of enteric microbial populations in the recovery of CD4 T-cells after sepsis. The results described present a previously unappreciated mechanism of CD4 T-cell impairment during the immunosuppressive stage of sepsis.
  • Thumbnail Image
    Item
    The porcine memory B cell in conferring long term adaptive immunity to viral pathogens
    (2017-06) Rahe, Michael
    Porcine reproductive and respiratory syndrome virus (PRRSV) is the most important pathogen of swine health and well-being worldwide. Discovered nearly thirty years ago, there is still no vaccine capable of producing a broadly protective immune response against the virus. This deficiency is due in large part to a failure to understand how the adaptive immune system responds to vaccination or infection. Specifically, there is little to no knowledge regarding the all-important memory immune response to PRRSV. The objective of this dissertation was to fill in this significant gap in knowledge by identifying and characterizing the memory B cell response to PRRSV vaccination. First, we identified the presence of memory B cells against PRRSV non-structural protein 7 (nsp7) through the use of a novel in vitro B cell culture system. Next, we created and validated a novel reagent, a B cell tetramer, against nsp7 to enhance the speed and sensitivity of memory cell identification. Finally, through the utilization of the nsp7 tetramer, we evaluated the regional specificity and dynamic nsp7 specific B cell response to PRRSV MLV vaccination within select secondary lymphoid organs. These results constitute the first evidence of regional specialization of the B cell response to vaccination in an outbred animal species. Furthermore, the presented methods are a blueprint for the study of antigen specific cellular responses to any significant pathogen of animals important for food or fiber.
  • Thumbnail Image
    Item
    Programmed death-1 regulates islet-specific lymphocytes in type 1 diabetes
    (2018-12) Martinov, Tijana
    Programmed death-1 (PD-1) is a T cell inhibitory receptor important for tolerance maintenance. PD-1 is highly expressed on chronically stimulated T cells, such as those specific for persistent viral or tumor antigens. PD-1 pathway blockade revolutionized cancer therapy in recent years. While response rates are higher than with chemotherapy, not all patients respond, and some develop autoimmune-like symptoms, or even overt autoimmunity. Herein, I sought to understand how the PD-1 pathway regulated islet-specific CD4+ T cells during type 1 diabetes (T1D) progression in non-obese diabetic (NOD) mice. Since insulin itself is one of the main antigens driving T1D, we developed insulin peptide:MHCII tetramer reagents to track insulin-reactive CD4+ T cells. Insulin-specific CD4+ T cells that expressed the most PD-1 also had the highest affinity for self, suggesting that PD-1 preferentially regulated those cells with the highest autoimmune potential. In NOD mice, the majority of insulin-specific CD4+ T cells had an anergic (tolerant) phenotype, but surprisingly, PD-1 blockade did not override the anergy program. These findings suggested that the differentiation state of the CD4+ T cell pre-determine its susceptibility to PD-1 blockade. Autoantibody production is a hallmark of autoimmunity, and has also been reported in patients treated with PD-1 blockade, suggesting that PD-1 might regulate this process. Autoantibody production results from B cell:CD4+ T cell interactions in the germinal center of the lymph node. The dynamics and regulation of the germinal center in spontaneous autoimmunity and after PD-1 blockade are not well understood, primarily due to an inability to track self-specific lymphocytes. To bridge this knowledge gap, we used tetramers to phenotype islet-specific CD4+ T cells and B cells in mice. PD-1- or PD-L1-deficient mice, as well as NOD mice treated with anti-PD-1, had increased insulin autoantibodies, as well as increased insulin-specific T follicular helper CD4+ T cells and germinal center B cells compared to controls. This increase was dependent on CD4+ T cell-intrinsic PD-1 signaling and relied on peptide:MHCII recognition. Taken together, my thesis work provides a mechanistic explanation for autoantibody onset following PD-1 blockade in the clinic, and has important implications for cancer immunotherapy and autoimmunity.