Introduction
Although the physical Build and Test steps are ongoing, our system’s design and expected functionality are supported by both computational modeling and literature evidence.
Promoter Activation Modeling
Hill's Equation-based simulations predict that the PnorV and PoxyS promoters will respond appropriately to nitric oxide (through the norR transcriptional factor) and hydrogen peroxide (through the oxyR transcriptional factor), respectively, which supports the validity of the AND-gate logic in our system.
PnorV Promoter
The PnorV promoter senses nitric oxide and produces a steep response curve, which means it has a sharp transition from inactive to fully active once NO reaches its threshold concentration. Its binary-like behavior minimizes any unwanted background expression while ensuring the activation is strong in NO’s presence (see Figure 1).
PoxyS Promoter
The PoxyS promoter senses hydrogen peroxide and produces a much more broad response curve, indicating a much more gradual activation. Since its detection window is wider, it is able to respond proportionally across the range of H₂O₂ levels in inflamed skin and proves the evidence of oxidative stress (see Figure 2).
Plux Promoter (AND Gate)
The Plux promoter functions as an AND gate dependent on both LuxR and 30C6HSL production. The standard Hill’s Equation was modified to account for multiple inputs instead of just one. The output of Plux is proportional to the product of both signals, which creates strict activation requirements for the promoter and ensures that the biosensor only responds when both NO and H₂O₂ are present together (see Figure 3).
This modeling demonstrates that our system can successfully distinguish true inflammatory conditions (when both NO and H₂O₂ are elevated) to normal, healthy skin, which may include lower levels of elevation.
Figure 1: PnorV promoter activation curve showing response to varying NO concentrations
Figure 2: PoxyS promoter activation curve showing response to varying H₂O₂ concentrations
Figure 3: Plux promoter AND gate response showing activation patterns dependent on NO and H₂O₂ concentrations
Literature Support
Indole-3-acetic acid (IAA), the intended anti-inflammatory output, has been shown in published studies to inhibit p65 nuclear translocation, suppress NF-κB signaling, reduce HO-1 and TNF-α expression, and neutralize reactive oxygen species.
p65 Subunit
The p65 subunit (encoded for by RelA gene) is part of the NF-κB transcription factor, a master regulator of inflammation and immune responses. NF-κB is usually a p50/p65 heterodimer that is held inactive in the cytoplasm by IκB. When a cell receives a stress or inflammatory signal, IκB is degraded, freeing p65 to enter the nucleus.
Once in the nucleus, p65 binds DNA and activates transcription of target genes. Its transactivation domain drives expression of pro-inflammatory cytokines, chemokines, and immune regulators.
In a recent study, IAA has been proven to reduce nuclear translocation of p65 and directly reduce NF-κB levels with this mechanism, aiding in reducing inflammation.
Indole-3-acetic acid Photodynamic Therapy
Indole-3-acetic acid has been used on acne therapies before, the most recent application being Indole-3-Acetic Acid Photodynamic Therapy (IAA-PDT). In IAA PDT, IAA acts as a photosensitizer that produces free radicals when exposed to green light irradiation. These free radicals, called reactive oxygen species (ROS), damage and kill acne-causing bacteria, without harming healthy skin tissue. A major advantage of IAA PDT is that once IAA has been exposed to light, it loses its photosensitizing ability, meaning that the patients do not require post-procedure photo protection. Previous clinical studies done by the National Library of Medicine have shown that IAA-PDT effectively suppresses the growth of two bacteria that are commonly associated with acne (P. acnes and S. aureus). Additionally, the study has shown that IAA PDT reduces both inflammatory lesions and sebum secretion, while also causing the destruction of the follicular ostia epithelium, which prevents clogged pores. However, this approach may be expensive and inaccessible for many people due to the need for visiting a healthcare provider to get this treatment done, which is why we looked into a more widely available delivery option.
Logical Circuit Design
Our dual-plasmid system directly links detection (NO/H₂O₂ sensing) to response (IAA production), demonstrating a rationally designed pathway from inflammatory signal to anti-inflammatory output.
Conclusion
Together, these modeling and literature-based results provide conceptual proof of concept, confirming that, theoretically, our engineered system is capable of detecting inflammation and producing a targeted anti-inflammatory response, even though experimental confirmation is still in progress.
References
Jung‐Im Na, et al. “Indole‐3‐Acetic Acid: A Potential New Photosensitizer for Photodynamic Therapy of Acne Vulgaris.” Lasers in Surgery and Medicine, vol. 43, no. 3, 1 Mar. 2011, pp. 200–205, https://doi.org/10.1002/lsm.21029.
Lin, Jennifer, and Marilyn T. Wan. “Current Evidence and Applications of Photodynamic Therapy in Dermatology.” Clinical, Cosmetic and Investigational Dermatology, May 2014, p. 145, www.ncbi.nlm.nih.gov/pmc/articles/PMC4038525/, https://doi.org/10.2147/ccid.s35334.