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Blocking pyrimidine de novo synthesis by inhibiting dihydroorotate dehydrogenase is used to treat autoimmunity and prevent expansion of rapidly dividing cell populations including activated T cells. Here we show memory T cell precursors are resistant to pyrimidine starvation. Although the treatment effectively blocked effector T cells, the number, function and transcriptional profile of memory T cells and their precursors were unaffected. This effect occurred in a narrow time window in the early T cell expansion phase when developing effector, but not memory precursor, T cells are vulnerable to pyrimidine starvation. This vulnerability stems from a higher proliferative rate of early effector T cells as well as lower pyrimidine synthesis capacity when compared with memory precursors. This differential sensitivity is a drug-targetable checkpoint that efficiently diminishes effector T cells without affecting the memory compartment. This cell fate checkpoint might therefore lead to new methods to safely manipulate effector T cell responses.

Resident T lymphocytes (T

Prolonged antigen exposure in chronic viral infections reduces the effector capacity of cytotoxic T cells - a phenomenon known as T cell exhaustion. High viral titer, strong TCR stimulation, and high antigen concentrations associated with strong inflammatory signals are known factors that promote the development of T cell exhaustion. A largely unexplored factor has been the influence of the microbiome on the development of T cell exhaustion. We report that T cell exhaustion progresses independently of the presence or absence of a bacterial microbiome in chronic lymphocytic choriomeningitis virus (LCMV) infections. Virus-specific CD8 T cells in germ-free mice showed high expression of the inhibitory receptor PD-1 and decreased cytokine production. Moreover, their global gene expression patterns, as determined by single-cell sequencing, were similar to those of cells in specific pathogen-free mice. In stark contrast to the phenotypic similarities, the absence of the microbiome delayed the partial viral control typically seen in chronic LCMV infections. Thus, our study demonstrates that the microbiome is dispensable for the induction of T cell exhaustion but critical for effector T cell function in chronic LCMV infections.

T cell receptor (TCR) stimulation leads to expression of the transcription factor TOX. Prolonged TCR signaling, such as encountered during chronic infections or in tumors, leads to sustained TOX expression, which induces a state of exhaustion or dysfunction. While CD8 memory T cells (Tmem) in specific pathogen-free laboratory mice typically do not express TOX, functional human Tmem show heterogeneous TOX expression levels. Whether TCR-independent mechanisms can alter TOX expression in human and murine Tmem has not been defined. We report that human and mouse Tmem increase TOX expression following stimulation with inflammatory cytokines IL-12, IL-15, and IL-18. TOX and PD-1 expression patterns often appear to be directly correlated, however, we found that TOX is not necessary for cytokine-driven expression of PD-1. Together, these observations highlight that inflammation is sufficient to alter TOX and PD-1 expression and that the signals regulating TOX expression appear well conserved in human and murine Tmem.

T cell receptor (TCR) stimulation leads to the expression of the transcription factor thymocyte selection-associated high-mobility group box (TOX). Prolonged TCR signaling, such as encountered during chronic infections or in tumors, leads to sustained TOX expression, which is required for the induction of a state of exhaustion or dysfunction. Although CD8+ memory T (Tmem) cells in mice typically do not express TOX at steady state, some human Tmem cells express TOX but appear fully functional. This seeming discrepancy between mouse and human T cells has led to the speculation that TOX is differentially regulated between these species, which could complicate the interpretation of preclinical mouse model studies. We report here that, similar to TCR-mediated signals, inflammatory cytokines are also sufficient to increase TOX expression in human and mouse Tmem cells. Thus, TOX expression is controlled by the environment, which provides an explanation for the different TOX expression patterns encountered in T cells isolated from specific pathogen-free laboratory mice versus humans. Finally, we report that TOX is not necessary for cytokine-driven expression of programmed cell death 1. Overall, our data highlight that the mechanisms regulating TOX expression are conserved across species and indicate that TOX expression reflects a T cell's activation state and does not necessarily correlate with T cell dysfunction.

T cell memory relies on the generation of antigen-specific progenitors with stem-like properties. However, the identity of these progenitors has remained unclear, precluding a full understanding of the differentiation trajectories that underpin the heterogeneity of antigen-experienced T cells. We used a systematic approach guided by single-cell RNA sequencing data to map the organizational structure of the human CD8+ memory T cell pool under physiological conditions. We identified two previously unrecognized subsets of clonally, epigenetically, functionally, phenotypically, and transcriptionally distinct stem-like CD8+ memory T cells. Progenitors lacking the inhibitory receptors programmed death-1 (PD-1) and T cell immunoreceptor with Ig and ITIM domains (TIGIT) were committed to a functional lineage, whereas progenitors expressing PD-1 and TIGIT were committed to a dysfunctional, exhausted-like lineage. Collectively, these data revealed the existence of parallel differentiation programs in the human CD8+ memory T cell pool, with potentially broad implications for the development of immunotherapies and vaccines.

miR-23~27~24 regulates TFH cells through targeting multiple genes including TOX, a key transcription factor for TFH cell biology., Follicular helper T (TFH) cells are essential for generating protective humoral immunity. To date, microRNAs (miRNAs) have emerged as important players in regulating TFH cell biology. Here, we show that loss of miR-23~27~24 clusters in T cells resulted in elevated TFH cell frequencies upon different immune challenges, whereas overexpression of this miRNA family led to reduced TFH cell responses. Mechanistically, miR-23~27~24 clusters coordinately control TFH cells through targeting a network of genes that are crucial for TFH cell biology. Among them, thymocyte selection–associated HMG-box protein (TOX) was identified as a central transcription regulator in TFH cell development. TOX is highly up-regulated in both mouse and human TFH cells in a BCL6-dependent manner. In turn, TOX promotes the expression of multiple molecules that play critical roles in TFH cell differentiation and function. Collectively, our results establish a key miRNA regulon that maintains optimal TFH cell responses for resultant humoral immunity.

This is a high-fidelity simulation reference guide for clinicians teaching inpatient pediatrics to third-year medical students. This guide involves four high-fidelity simulation cases where the students obtain a brief history and physical exam. Through history and physical exam, including recognizing abnormal pediatric vital signs, the students determine a diagnosis and provide initial stabilization. The scenarios include asthma, bronchiolitis, anaphylaxiss, and hypovolemic shock. There is a knowledge test and confidence survey included to evaluate for improvement after the simulation exercises. This module began as a research project involving 80 medical students. Students participated throughout one afternoon of the pediatric clerkship. Students significantly improved their confidence with pediatric patients after the simulation experience. Of a six-question survey, all six questions showed significant improvement in confidence. Many students commented on the quality of education provided by this simulation exercise and requested more simulation in their curriculum. This module is now implemented into the curriculum of the pediatric clerkship and the medical school is improving simulation in other aspects of medical education.