Adaptive Wound-Responsive Gene Circuit for Fibrosis Prevention Abstract Chronic wounds and fibrotic scarring represent a significant unmet clinical need, affecting millions of patients annually and resulting in impaired tissue function, pain, and reduced quality of life. Current therapeutic approaches lack the spatiotemporal precision needed to modulate the wound microenvironment dynamically — delivering anti-inflammatory signals early and anti-fibrotic signals later, in response to the wound’s own molecular cues. This project proposes the design and experimental validation of a two-stage, NF-κB/STAT3-responsive synthetic gene circuit encoded in a piggyBac transposon vector, engineered for stable integration into dermal fibroblasts. The circuit is designed to first sense early inflammatory NF-κB signaling and secrete IL-10 (an anti-inflammatory cytokine), then switch to a STAT3/NF-κB dual-input logic gate that drives decorin secretion (an anti-fibrotic proteoglycan) as the wound transitions to the proliferative phase. A Bxb1 serine integrase-based irreversible switching mechanism ensures the circuit commits to Stage 2 and does not revert. Dual fluorescent reporters — mCherry (Stage 1) and EGFP (Stage 2) — enable real-time monitoring of circuit state. The construct (~8–9 kb) will be synthesized as a whole plasmid by Twist Bioscience and validated in an automated 384-well fibroblast stimulation assay using the Echo 525, Tempest, and Spark Plate Reader. This work establishes a programmable, cell-autonomous therapeutic platform with broad implications for wound care, fibrosis, and synthetic immunology.