This study examined the response of various forms and sources of glucans toward two different Limulus amebocyte lysate (LAL) methods, the modified LAL, and Glucatell. The glucans studied were curdlan, laminarin, yeast glucan, barley glucan, paramylon, pullulan, pustulan, mannan, and pachyman (as part of the Glucatell kit). Both methods provided largely similar results for each of the glucans; however, the Glucatell method yielded slightly higher responses to certain structures that may not necessarily be of fungal origin, leading to falsely greater positive results. The performance of each method to measure fungal glucan concentration specifically was then assessed.
In view of the role interleukin (IL)-7 plays in T cell survival, homeostasis and function it is no surprise expression of the IL-7 receptor-α (CD127) is tightly regulated. Dysregulation of IL-7 signaling and CD127 expression has been implicated in a number of diseases such as multiple sclerosis, breast cancer, arthritis as well as in HIV infection. Thus it is important to understand how IL-7 regulates its own receptor expression. Here we elucidate the mechanism by which IL-7 suppresses CD127 transcripts in primary human CD8 T cells. We show by qPCR and nuclear run-on assays that IL-7 suppresses CD127 gene transcription in a time- and dose-dependent manner. The IL-7 mediated suppression of CD127 transcripts is dependent on JAK/STAT5 signaling. Notably, cycloheximide blocked IL-7’s ability to down-regulate CD127 transcripts suggesting IL-7 stimulates the de novo synthesis of a transcriptional repressor which in turn suppresses CD127 gene transcription. Through DNA microarrays and PCR arrays, the IL-7 inducible transcription factor c-Myb was identified. The region within the CD127 gene promoter required for IL-7 mediated transcriptional suppression was identified through progressive truncations using firefly luciferase as a reporter gene and is located from -1760 to -2406 bp upstream of the TATA box and contains c-Myb binding sites. Using siRNA-mediated knockdown and transient over-expression, we illustrate c-Myb suppresses CD127 gene transcription in primary human CD8 T cells.
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