Dectin-1 ligands produce distinct training phenotypes in human monocytes through differential activation of signaling networks.

Cells of the innate immune system retain memory of prior exposures through a process known as innate immune training. ß-glucan, a Dectin-1 ligand purified from the Candida albicans cell wall, has been one of the most widely utilized ligands for inducing innate immune training. However, many Dectin-1 ligands exist, and it is not known whether these all produce the same phenotype. Using a well-established in vitro model of innate immune training, we compared two commercially available Dectin-1 agonists, zymosan and depleted zymosan, with the gold standard ß-glucan in the literature. We found that depleted zymosan, a ß-glucan purified from Saccharomyces cerevisiae cell wall through alkali treatment, produced near identical effects as C. albicans ß-glucan. However, untreated zymosan produced a distinct training effect from ß-glucans at both the transcript and cytokine level. Training with zymosan diminished, rather than potentiated, induction of cytokines such as TNF and IL-6. Zymosan activated NFκB and AP-1 transcription factors more strongly than ß-glucans. The addition of the toll-like receptor (TLR) ligand Pam3CSK4 was sufficient to convert the training effect of ß-glucans to a phenotype resembling zymosan. We conclude that differential activation of TLR signaling pathways determines the phenotype of innate immune training induced by Dectin-1 ligands.

Latency reversal plus natural killer cells diminish HIV reservoir in vivo.

HIV is difficult to eradicate due to the persistence of a long-lived reservoir of latently infected cells. Previous studies have shown that natural killer cells are important to inhibiting HIV infection, but it is unclear whether the administration of natural killer cells can reduce rebound viremia when anti-retroviral therapy is discontinued. Here we show the administration of allogeneic human peripheral blood natural killer cells delays viral rebound following interruption of anti-retroviral therapy in humanized mice infected with HIV-1. Utilizing genetically barcoded virus technology, we show these natural killer cells efficiently reduced viral clones rebounding from latency. Moreover, a kick and kill strategy comprised of the protein kinase C modulator and latency reversing agent SUW133 and allogeneic human peripheral blood natural killer cells during anti-retroviral therapy eliminated the viral reservoir in a subset of mice. Therefore, combinations utilizing latency reversal agents with targeted cellular killing agents may be an effective approach to eradicating the viral reservoir.

MiniBioReactor Arrays (MBRAs) as a Tool for Studying C. difficile Physiology in the Presence of a Complex Community.

The commensal microbiome plays an important role in the dynamics of Clostridium difficile infection. In this chapter, we describe minibioreactor arrays (MBRAs), an in vitro cultivation system that we developed that allows for C. difficile physiology to be assayed in the presence of complex fecal microbial communities. The small size of the bioreactors within the MBRAs allows for dozens of reactors to be run simultaneously and therefore several different variables can be tested with limited time and cost. When coupled with experiments in animal models of C. difficile infection, MBRAs can provide important insights into C. difficile physiology and pathogenesis.