Activation of a naive T cell is a highly energetic event, which requires a substantial increase in nutrient metabolism. Upon stimulation, T cells increase in size, rapidly proliferate, and differentiate, all of which lead to a high demand for energetic and biosynthetic precursors. Although amino acids are the basic building blocks of protein biosynthesis and contribute to many other metabolic processes, the role of amino acid metabolism in T cell activation has not been well characterized. We have found that glutamine in particular is required for T cell function. Depletion of glutamine blocks proliferation and cytokine production, and this cannot be rescued by supplying biosynthetic precursors of glutamine. Correlating with the absolute requirement for glutamine, T cell activation induces a large increase in glutamine import, but not glutamate import, and this increase is CD28-dependent. Activation coordinately enhances expression of glutamine transporters and activities of enzymes required to allow the use of glutamine as a Krebs cycle substrate in T cells. The induction of glutamine uptake and metabolism requires ERK function, providing a link to TCR signaling. Together, these data indicate that regulation of glutamine use is an important component of T cell activation. Thus, a better understanding of glutamine sensing and use in T cells may reveal novel targets for immunomodulation.

Key Points Question Are cerebrospinal fluid (CSF) SARS-CoV-2 antigens associated with central nervous system inflammation in patients with COVID-19? Findings Of 44 patients with COVID-19 (23 neurosymptomatic) included in this hospital-based cross-sectional study, CSF nucleocapsid antigen was detectable in 89% of patients with available data and was significantly correlated with immune activation markers (neopterin and interferon γ). Moreover, neurosymptomatic patients had a more pronounced inflammatory CSF profile compared with neuroasymptomatic patients that could not be attributed to differences in COVID-19 severity. Meaning These results suggest that viral components may contribute to central nervous system immune responses without direct viral invasion and highlight the clinical importance of neurologic symptoms.

BackgroundThe last few years have seen major advances in blood biomarkers for Alzheimer's Disease (AD) with the development of ultrasensitive immunoassays, promising to transform how we diagnose, prognose, and track progression of neurodegenerative dementias.MethodsWe evaluated a panel of four novel ultrasensitive electrochemiluminescence (ECL) immunoassays against presumed CNS derived proteins of interest in AD in plasma [phosphorylated-Tau181 (pTau181), total Tau (tTau), neurofilament light (NfL), and glial fibrillary acidic protein (GFAP)]. Two sets of banked plasma samples from the Massachusetts Alzheimer's Disease Research Center's longitudinal cohort study were examined: A longitudinal prognostic sample (n = 85) consisting of individuals with mild cognitive impairment (MCI) and 4 years of follow-up and a cross-sectional sample (n = 238) consisting of individuals with AD, other neurodegenerative diseases (OND), and normal cognition (CN).ResultsParticipants with MCI who progressed to dementia due to probable AD during follow-up had higher baseline plasma concentrations of pTau181, NfL, and GFAP compared to non-progressors. The best prognostic discrimination was observed with pTau181 (AUC = 0.83, 1.7-fold increase) and GFAP (AUC = 0.83, 1.6-fold increase). Participants with autopsy- and/or biomarker verified AD had higher plasma levels of pTau181, tTau and GFAP compared to CN and OND, while NfL was elevated in AD and further increased in OND. The best diagnostic discrimination was observed with pTau181 (AD vs CN: AUC = 0.90, 2-fold increase; AD vs. OND: AUC = 0.84, 1.5-fold increase) but tTau, NfL, and GFAP also showed good discrimination between AD and CN (AUC = 0.81–0.85; 1.5–2.2 fold increase).ConclusionsThese new ultrasensitive ECL plasma assays for pTau181, tTau, NfL, and GFAP demonstrated diagnostic utility for detection of AD. Moreover, the absolute baseline plasma levels of pTau181 and GFAP reflect cognitive decline over the next 4 years, providing prognostic information that may have utility in both clinical practice and clinical trial populations.

Background: The last few years have seen major advances in blood biomarkers for Alzheimer's Disease (AD) with the development of ultrasensitive immunoassays, promising to transform how we diagnose, prognose, and track progression of neurodegenerative dementias. Methods: We evaluated a panel of four novel ultrasensitive electrochemiluminescence (ECL) immunoassays against presumed CNS derived proteins of interest in AD in plasma [phosphorylated-Tau181 (pTau181), total Tau (tTau), neurofilament light (NfL), and glial fibrillary acidic protein (GFAP)]. 366 plasma samples from the Massachusetts Alzheimer's Disease Research Center's longitudinal cohort study were examined to differentiate definite AD, other neurodegenerative diseases (OND), and cognitively normal (CN) individuals. A subset of samples were selected to have longitudinal follow up to also determine the utility of this plasma biomarker panel in predicting 4-year risk for cognitive decline in individuals with different levels of cognitive impairment. Results: pTau181, tTau and GFAP were higher in AD compared to CN and OND, while NfL was elevated in AD and further increased in OND. pTau181 performed the best (AD vs CN: AUC=0.88, 2-fold increase; AD vs OND: AUC=0.78, 1.5-fold increase) but tTau also showed excellent discrimination (AD vs CN: AUC=0.79, 1.5-fold increase; AD vs OND: AUC=0.72, 1.3-fold increase). Participants with MCI who progressed to AD dementia had higher baseline plasma concentrations of pTau181, NfL, and GFAP compared to non-progressors with the best discrimination for pTau181 (AUC=0.82, 1.7-fold increase) and GFAP (AUC=0.81, 1.6-fold increase). Conclusions: These new ultrasensitive ECL plasma assays for pTau181, tTau, NfL, and GFAP detect CNS disease with high specificity and accuracy. Moreover, the absolute baseline plasma levels of pTau and GFAP reflect clinical disease aggressiveness over the next 4 years, providing diagnostic and prognostic information that may have utility in both clinical and clinical trial populations. Classification of Evidence: This study provides Class II evidence that plasma levels of pTau181, tTau, NfL, and GFAP are associated with AD and that pTau181 and GFAP are associated with progression from MCI to AD dementia.

Most immunoassays quantify cytokines with pg/mL sensitivity, which is insufficient to detect those present at low levels. MSD’s next-generation S-PLEXSM assay format was developed to quantify cytokines with fg/mL sensitivity. Here we describe measuring IL-2, IL-4, IL-6, IL-10, and IL-17A levels in normal sera and the supernatants of model cell lines using S-PLEX assays. The assays demonstrated a dynamic range of approximately four orders of magnitude. Standard intra-plate coefficients of variation (CVs) ranged from 3%-15% and inter-plate CVs ranged from 8%–18%. The lower limit of detection (LLOD) was <1 fg/mL for all assays except IL-17A (11 fg/mL). IL-2, IL-6, IL-10, and IL-17A were detectable in 100% of normal sera samples (n=36–75) and IL-4 was detectable in 40% of normal sera samples (n=75). The average concentrations were <1 pg/mL for normal samples. To confirm the sensitivity of these assays and their ability to detect native analytes, we characterized a panel of 23 cell lines that are models for cytokine secretion. As an example, the MOLT-4 cell line derived from acute lymphoblastic leukemia natively expressed detectable levels of IL-2 (1,031 fg/mL), IL-4 (221 fg/mL), IL-10 (60 fg/mL), and IL-17A (30 fg/mL). The expression profiles of these cell lines confirmed the sensitivity of the S-PLEX assays. S-PLEX cytokine assays provide a 100 to 1,000-fold lower LOD than standard ELISAs, allowing the determination of baseline cytokine levels in many sample types.