Memory B cells are found in lymphoid and non–lymphoid tissues, suggesting that some may be tissue–resident cells. Here we show that pulmonary influenza infection elicited lung–resident memory B cells (BRM cells) that were phenotypically and functionally distinct from their systemic counterparts. BRM cells were established in the lung early after infection, in part because their placement required local antigen encounter. Lung BRM cells, but not systemic memory B cells, contributed to early plasmablast responses following challenge infection. Following secondary infection, antigen–specific BRM cells differentiated in situ , whereas antigen–non–specific BRM cells were maintained as memory cells. These data demonstrate that BRM cells are an important component of immunity to respiratory viruses like influenza and suggest that vaccines designed to elicit BRM cells must deliver antigen to the lung.

MicroRNAs are key regulators of the immune response, but their role in CD8 T cell differentiation in vivo is not known. We show that miR-155 is important in both effector and memory antiviral CD8 T cell responses. Without miR-155, there was a weaker effector response and a skewing toward memory precursor cells. At the memory stage, miR-155-deficient CD8 T cells preferentially differentiated into central memory cells and were capable of mounting a potent secondary response. For intracellular pathogens, a crucial part of long-term immunity is the generation of memory CD8 T cells. Naïve T cells can be directed toward at least two different fate-determining pathways after antigen (Ag) exposure. One pathway, driven by IL-12 and IL-2 signaling, upregulates T box expressed in T cell (T-bet) expression, resulting in terminal differentiation of short-lived effector cells (SLECs). Alternatively, a pathway driven by lower levels of IL-12 but stimulated by IL-21 or IL-10 leads to production of memory precursor effector cells (MPECs), which display less effector activity than SLECs (1). The ultimate repository for longterm memory is the central memory T cell (T cm ), and understanding how T cells can be directed toward this fate has profound implications for vaccine design. Given the distinct transcriptional profiles of effector and memory cells (2), it is an open question if there are master regulators of gene expression in these cells. MicroRNAs (miRNAs) are important posttranscriptional regulators in many developmental processes. Several miRNAs have been shown to be critical in many stages of hematopoietic development (3). While miRNAs have been shown to influence CD4 T cell and regulatory T cell differentiation, it is currently unclear which miRNAs influence CD8 T cell differentiation.Studies using CD8 T cells lacking dicer, an enzyme essential for the processing of all miRNAs, showed deficits in CD8 T cell accumulation and survival (4). Other studies profiled miRNA expression following CD8 T cell activation in vitro (5, 6). Upregulation of miR-21 and miR-155 was observed, which was interesting due to previous studies implicating roles for them in immune cell function (7,8). In another study, CD8 T cells cultured in vitro to resemble central memory cells downregulated miR-155 relative to effector CD8 T cells (9). While these studies implied miR-155 involvement in CD8 T cell differentiation and memory development, it is still unclear which processes are affected by miR-155 during T cell differentiation in vivo. Here we report that miR-155 plays a critical role in the generation of memory CD8 T cells during virus infection in vivo.Discussion. To determine the kinetics of miR-155 expression after CD8 T cell activation, we simulated OT-I cells with antigen in vitro and measured miR-155 expression by PCR. miR-155 expression peaked at 3 days poststimulation and then declined at all subsequent time points (Fig. 1). The decline in miR-155 was much more marked in cells cultured with IL-15 than in cultures supplemented with IL-2, ...

CD4 help is crucial for memory CD8+ T cell development, yet the mechanisms of CD4 help and why (CD4) helpless memory CD8+ T cells elicit poor recall responses are currently not well understood. In this study we investigated these questions using an in vivo acute virus infection model. We show herein that CD4 help during priming is required for memory CD8+ T cell differentiation, and that stimulation of CD40 during priming rescues the helpless defects in the absence of CD4+ T cells. The defective recall response by helpless memory cells did not correlate with the amount of cell death and was independent of TRAIL. However, helpless memory cells excessively up-regulated the inhibitory receptor PD-1 (programmed cell death-1), and PD-1 blockade enhanced the recall response of helpless memory cells. Furthermore, providing IL-2 signaling in vivo during the recall response reduced PD-1 expression and rescued the recall response of helpless memory cells. Our study identifies molecular pathways involved in CD4 help for memory CD8+ T cell generation that are independent of TRAIL, and it provides therapeutic implications that helpless memory cell function can be restored at multiple stages through various immunological interventions.

The coronavirus disease 2019 (COVID-19) pandemic has highlighted the urgent need for effective prophylactic vaccination to prevent the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Intranasal vaccination is an attractive strategy to prevent COVID-19 as the nasal mucosa represents the first-line barrier to SARS-CoV-2 entry. The current intramuscular vaccines elicit systemic immunity but not necessarily high-level mucosal immunity. Here, we tested a single intranasal dose of our candidate adenovirus type 5-vectored vaccine encoding the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein (AdCOVID) in inbred, outbred, and transgenic mice. A single intranasal vaccination with AdCOVID elicited a strong and focused immune response against RBD through the induction of mucosal IgA in the respiratory tract, serum neutralizing antibodies, and CD4+ and CD8+ T cells with a Th1-like cytokine expression profile. A single AdCOVID dose resulted in immunity that was sustained for over six months. Moreover, a single intranasal dose completely protected K18-hACE2 mice from lethal SARS-CoV-2 challenge, preventing weight loss and mortality. These data show that AdCOVID promotes concomitant systemic and mucosal immunity and represents a promising vaccine candidate.

The T cell response possesses a number of inhibitory receptors to regulate the extent of the antiviral response and prevent immune pathology. These receptors are generally transiently upregulated during an effector response and then downregulated during memory. Some inhibitory receptors, such as programmed death 1 (PD-1) and LAG-3, were shown to be aberrantly upregulated during memory to chronic lymphocytic choriomeningitis virus infection, limiting functional capabilities. However, little is known about the impact of inhibitory receptors on memory development during a normal CD8 T cell response to acute virus infection. Our previous data showed that PD-1 is aberrantly upregulated during a secondary response by memory CD8 T cells that were generated without CD4 T cell help. Therefore, we examined the role of PD-1 in memory differentiation during acute vaccinia virus infection in intact mice. In the absence of PD-1, the primary and memory CD8 T cell responses were enhanced. Moreover, there were distinct phenotypic and functional changes in the memory PD-1−/− CD8 T cells. Higher levels of CD62L, CD27, and CCR7 were detected; cells produced more IL-2 and made an enhanced secondary response. These changes indicate a skewing of the memory population toward the central memory phenotype in the absence of PD-1 signaling.