The administration route is decisive for the ability of the vaccine adjuvant CAF09 to induce antigen-specific CD8+ T-cell responses:the immunological consequences of the biodistribution profile

A prerequisite for vaccine-mediated induction of CD8+ T-cell responses is the targeting of dendritic cell (DC) subsets specifically capable of cross-presenting antigen epitopes to CD8+ T cells. Administration of a number of cationic adjuvants via the intraperitoneal (i.p.) route has been shown to result in strong CD8+ T-cell responses, whereas immunization via e.g. the intramuscular (i.m.) or subcutaneous (s.c.) routes often stimulate weak CD8+ T-cell responses. The hypothesis for this is that self-drainage of the adjuvant/antigen to the lymphoid organs, which takes place upon i.p. immunization, is required for the subsequent activation of cross-presenting lymphoid organ-resident CD8α+ DCs. In contrast, s.c. or i.m. immunization usually results in the formation of a depot at the site of injection (SOI), which hinders the self-drainage and targeting of the vaccine to cross-presenting CD8α+ DCs. We investigated this hypothesis by correlating the biodistribution pattern and the adjuvanticity of the strong CD8+ T-cell inducing liposomal cationic adjuvant formulation 09 (CAF09), which is composed of dimethyldioctadecylammonium bromide/monomycoloyl glycerol liposomes with polyinosinic:polycytidylic acid electrostatically adsorbed to the surface. Biodistribution studies with radiolabeled CAF09 and a surface-adsorbed model antigen [ovalbumin (OVA)] showed that a significantly larger fraction of the vaccine dose localized in the draining lymph nodes (dLNs) and the spleen 6 h after i.p. immunization, as compared to after i.m. immunization. Studies with fluorescently labelled OVA + CAF09 demonstrated a preferential association of OVA + CAF09 to DCs/monocytes, as compared to macrophages and B cells, following i.p. immunization. Administration of OVA + CAF09 via the i.p. route did also result in DC activation, whereas no DC activation could be measured within the same period with unadjuvanted OVA and OVA + CAF09 administered via the s.c. or i.m. routes. In the dLNs, the highest level of activated, cross-presenting CD8α+ DCs was detected at 24 h post immunization, whereas an influx of activated, migrating and cross-presenting CD103+ DCs to the dLNs could be measured after 48 h. This suggests that the CD8α+ DCs are activated by self-draining OVA + CAF09 in the lymphoid organs, whereas the CD103+ DCs are stimulated by the OVA + CAF09 at the SOI. These results support the hypothesis that the self-drainage of OVA + CAF09 to the draining LNs is required for the activation of CD8α+ DCs, while the migratory CD103+ DCs may play a role in sustaining the subsequent induction of strong CD8+ T-cell responses

RNAModMapper: RNA Modification Mapping Software for Analysis of Liquid Chromatography Tandem Mass Spectrometry Data

Liquid chromatography tandem mass spectrometry (LC-MS/MS) has proven to be a powerful analytical tool for the characterization of modified ribonucleic acids (RNAs). The typical approach for analyzing modified nucleosides within RNA sequences by mass spectrometry involves ribonuclease digestion followed by LC-MS/MS analysis and data interpretation. Here we describe a new software tool, RNAModMapper (RAMM), to assist in the interpretation of LC-MS/MS data. RAMM is a stand-alone package that requires user-submitted DNA or RNA sequences to create a local database against which collision-induced dissociation (CID) data of modified oligonucleotides can be compared. RAMM can interpret MS/MS data containing modified nucleosides in two modes: fixed and variable. In addition, RAMM can also utilize interpreted MS/MS data for RNA modification mapping back against the input sequence(s). The applicability of RAMM was first tested using total tRNA isolated from Escherichia coli. It was then applied to map modifications found in 16S and 23S rRNA from Streptomyces griseus

Differentiating Positional Isomers of Nucleoside Modifications by Higher-Energy Collisional Dissociation Mass Spectrometry (HCD MS)

The analytical identification of positional isomers (e.g., 3-, N4-, 5-methylcytidine) within the > 160 different post-transcriptional modifications found in RNA can be challenging. Conventional liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) approaches rely on chromatographic separation for accurate identification because the collision-induced dissociation (CID) mass spectra of these isomers nearly exclusively yield identical nucleobase ions (BH2+) from the same molecular ion (MH+). Here, we have explored higher-energy collisional dissociation (HCD) as an alternative fragmentation technique to generate more informative product ions that can be used to differentiate positional isomers. LC-MS/MS of modified nucleosides characterized using HCD led to the creation of structure- and HCD energy-specific fragmentation patterns that generated unique fingerprints, which can be used to identify individual positional isomers even when they cannot be separated chromatographically. While particularly useful for identifying positional isomers, the fingerprinting capabilities enabled by HCD also offer the potential to generate HPLC-independent spectral libraries for the rapid analysis of modified ribonucleosides. Graphical Abstract ᅟ

Mild Traumatic Brain Injury Is Related to Elevated Cerebrospinal Fluid Tau in Alzheimer\u27s Disease Dementia.

Few studies have examined an association between mild traumatic brain injury (mTBI) and Alzheimer\u27s disease (AD). For this reason, we compared an AD dementia group with an mTBI history (n = 10) to a matched AD control group (n = 20) on measures of cognitive function, cerebral glucose metabolism, and markers of amyloid and tau deposition. Only a trend and medium-to-large effect size for higher phosphorylated and total tau was identified for the mTBI group. A history of mTBI may be associated with greater tau in AD, indicating a potential pathway for increasing risk for AD, though further evaluation with larger samples is needed