title: Characterisation of T Follicular Helper Cell (TFH) in Nasopharynx-Associated Lymphoid tissue and Its effect on regulation of immune response to influenza virus
Creators: name: Aljurayyan, Abdullah Nasser
Publisher: name: University of Liverpool Repository
Year: 2015

description: TFH cells have been identified as a new T helper subset specialized to regulate the development of effector and memory B cells and long-lived plasma cells. The interaction between TFH and B cells leads to the activation of B cells and germinal centers (GC) formation. Considering the importance of TFH for B cell antibody response, novel vaccine adjuvants and intranasal vaccines to boost TFH number or function may be an attractive vaccination strategy to enhance vaccine efficacy in humans. Adenotonsillar tissues are major parts of nasopharyngeal associated lymphoid tissues (NALT) and they are important in response to upper respiratory tract pathogens and intranasal vaccination. This PhD project investigated the frequencies of TFH in human NALT and PBMC in children and adults. The effects of CpG-DNA and live attenuated influenza vaccine (LAIV) on TFH in human NALT and the TFH-mediated B cell immunity to influenza virus were studied. The importance of the cytokine IL-21 and plasmacytoid dendritic cells (pDC) in TFH cell-mediated B cell antibody production was also investigated. Adenotonsillar MNC and PBMC were isolated from adenotonsillar tissues and peripheral blood respectively. TFH (CD4+ CXCR5high ICOShigh) numbers and function were analysed by flowcytometry and intracellular cytokine staining. Purified TFH (CD4+ CXCR5hi) and non-TFH cells (CD4+ CXCR5-) were co-cultured with B cells in the presence of influenza virus antigen and CpG-DNA or of LAIV. Purified pDC were added to the TFH-B cell co-culture to study their importance in TFH -mediated B cell antibody production. Haemagglutinin (HA)-specific antibody production was analysed by ELISA and ELISpot assay. IL-21 receptor blocking by neutralization was used to study the importance of IL-21 in TFH-mediated B cell antibody production. A prominent number of TFH were found in human NALT which were considerably higher than in PBMC. There was an age-associated difference in TFH numbers in NALT and BPMC, i.e. the mean TFH number was higher in children than in adults. TFH in NALT were shown to express high levels of IL-4, IL-10 and IL-21 and that were important for B cell antibody production. A good correlation between the numbers of GC B cell and TFH in NALT was seen. Co-culture of purified TFH but not non-TFH with B cells promoted antibody production. Stimulation of adenotonsillar MNC by CpG-DNA significantly increased TFH number and that was correlated with HA-specific antibody production following influenza antigen stimulation. Co-incubation of TFH-B cell with pDC enhanced the CpG-DNA-mediated antibody production. We also found that stimulation with LAIV significantly increased TFH number and that was correlated with HA-specific antibody production. Blocking the IL-21R significantly reduced the number of TFH that was correlated with a significant reduction of HA-specific antibody production. Enhancing vaccine immunogenicity through modulation of TFH numbers or function in human NALT using immunological adjuvants such as CpG-DNA and through intranasal vaccination may be an effective vaccination strategy against respiratory pathogens.