IGEM:Stanford/2009/Project Homeostasis/Inflammation Device

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Summary of Device


The anti-inflammatory device consists of two components: a sensor and a responder. The sensor portion detects an input molecule whose concentrations reflect the degree of local inflammation. When the concentration of the input rises above a certain baseline threshold, the sensor will activate the responder portion and induce the production of an output molecule. The output will attenuate local inflammation by enhancing the expansion of the immunosuppressive Treg lineage while curbing the proliferation of the inflammatory Th17 lineage.

We have chosen the soxRS regulon, a genetic defense mechanism native to E. coli, to serve as our sensor. The soxRS regulon is responsive to nitric oxide (NO), a free radical that exists at high concentrations in the inflamed lumen of patients with active inflammatory bowel diseases. In our machine, the presence of NO activates the constitutively expressed soxR protein, forming a complex that binds to the soxS promoter and induces production of the output molecule, all-trans retinoic acid (RA). Synthesis of RA requires the Crt cluster and the blh gene, which together constitute a carotenoid biosynthetic pathway that metabolizes isoprenoid precursors and yields RA. RA diffuses across the E. coli membranes and negatively regulates the Th17 lineage while enhancing the Treg response. Thus, in response to high concentrations of the input signal NO, the soxRS sensor will activate the anti-inflammatory responder and induce production of RA.

Parts Designs
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Other Considerations
NO toxicity to E. coli: As a free radical, NO is toxic to cells and has been shown to disrupt cellular respiration of E. coli at high concentrations. However, we do not anticipate this to be a problem given the sensitivity of the protective mechanisms that have evolved in E. coli to protect against growth inhibition by NO. In particular, the enzymes cytochrome bd and flavohemoglobin have been implicated in this protection and are sensitive to NO at micromolar concentrations.

RA toxicity to host: High concentrations of vitamin A are toxic and chronic overdose (hypervitaminosis A) is associated with pathologic liver conditions and osteoporosis. However, the recommended daily allowance for vitamin A is 900 micrograms, an amount well above the threshold needed to polarize T cell lineages. Acute toxicity is associated with an intake of > 100x the RDA in the course of a few days for adults and chronic toxicity correlates with a daily intake of preformed vitamin A 7500 micrograms for six years.

Other potential "input" molecules: PGE2

Other possible “output” molecules: vitamin D, Y-320 pyrazoleanilide derivative, 17beta-oestradiol

Challenges

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Results

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