Scaffolding and peer review methods were combined on an online platform to develop scientific writing skills in an undergraduate organic chemistry lab.Collaboration was encouraged throughout the semester.First students anonymously critiqued lab report sections from their peers through the online platform. Later, small groups produced complete reports built up from the sections covered by peer review. Each student served a different role in the group, moving from a collaborative approach towards independent writing. The culmination of the semester was a complete report, produced independent of peer or instructor guidance. Implementation was accomplished through a series of rubrics, worksheets, and instructor feedback. Preliminary assessment of these curriculum changes indicates that student writing skills improved and that student feedback is mixed, but generally positive. The ultimate goal is the development of writing skills in lower-level chemistry labs in preparation for writing-intensive upper-level labs.

The ontogeny of exhausted CD8 T cells as well as the underlying mechanisms that account for the functional inactivation of these cells remains ill-defined. We have utilized cytokine reporter mice, which mark the synthesis of interferon-γ by the expression of Thy1.1, to decipher how activation events during the early stages of a chronic infection dictate the development of exhaustion. We show that, during chronic lymphocytic choriomeningitis virus infection, the precursors of exhausted T cells all upregulate interferon-γ mRNA and become susceptible to depletion with anti-Thy1.1 antibodies. This potent phase of hyperactivation which precedes exhaustion is antigen-dependent and is dictated by viral levels and T cell precursor frequencies. Unlike acute infections, which result in massive expansion of the responding T cells, during chronic infections further expansion of the initial response becomes rapidly attenuated. The exhausted T cells which subsequently emerge in chronically infected hosts attain a distinct CD127lo, KLRG-1lo phenotype but do not silence interferon-γ transcription even though protein expression is abolished. Thus ablation of interferon-γ production by exhausted cells is not due to transcriptional silencing, implicating post-transcription regulatory mechanisms in disabling this effector module.

B cells with potent IL-10-dependent regulatory functions (B10 cells) expand in mice and humans during inflammation and autoimmunity. In this study, the fate of B10 cells following in vivo expansion was examined using two distinct il-10 reporter strains: Tiger mice that express cytoplasmic GFP and 10BiT mice that express cell surface Thy1.1. B10 cells from Tiger mice express cytoplasmic IL-10 and GFP with similar kinetics following stimulation in vitro and in vivo. By contrast, B10 cell Thy1.1 expression in 10BiT mice was delayed relative to IL-10 production and persisted even after B10 cells had lost the capacity to produce IL-10. Thy1.1 also served as a marker for B10 cells that expressed plasma cell-associated transcription factors and had the capacity to differentiate into CD138+B220LO antibody-secreting plasma cells in vivo and in vitro. Antibodies secreted by B10 cells included broadly reactive “natural antibodies” and autoantibodies, and were antigen-specific in immunized mice. At the molecular level, B10 cells predominantly expressed a germline-encoded IgM antibody repertoire with diverse VH and VL gene utilization. These results indicate that B10 cells can differentiate following IL-10 secretion to produce broadly-reactive germline-encoded antibodies that may function to rapidly clear antigens and further reduce inflammation and immunopathology.

Exposure to the opportunistic mold Aspergillus fumigatus may result in a wide range of infectious diseases. Among these infections, invasive aspergillosis is the most common and the most lethal. Our lab has previously shown a protective role for Dectin-dependent IL-22 production. In our current studies we sought to identify potential cell source(s) of IL-22 as well as factors critical for its production during invasive aspergillosis. Employing human CD4 IL-22 reporter mice, we show that iNKT cells and γδ T cells are sources of IL-22 48 h post-A. fumigatus exposure. In subsequent studies, we observed an early impairment in IL-22 production by lung cells from CD1d deficient (iNKT deficient) mice that returned to wild type levels by 48 h. In contrast, mice deficient in γδ T cells had relatively intact IL-22 production early after A. fumigatus exposure that became significantly attenuated by 48 h post-challenge. In order to identify factors critical for IL-22 production we interrogated the role of several pertinent transcription factors including: related orphan receptor-γ (RORγ), the aryl hydrocarbon receptor (Ahr), MyD88, and TRIF. Collectively, our data shows that iNKT cells and γδ T cells support early vs. late IL-22 production, which is fully dependent on RORγ and MyD88, partially dependent on Ahr, and independent of TRIF.

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