Decellularized Matrix Sheets versus Micro-Patterned Synthetic Films: Divergent Re-Engagement of Positional Gene Networks During Limb Regeneration
June 27
It remains unclear how the physical medium used to encode positional information—ranging from desiccated extracellular-matrix (ECM) sheets to digitally micro-patterned polymers—shapes the subsequent recall of that information during regeneration. We compared three cohorts of axolotl limb stumps that received positional cues by overlaying the wound surface with (i) a pliable, decellularized matrix sheet (Sheet), (ii) a photo-lithographically patterned hydrogel (Pattern), or (iii) a flexible, conductive polymer film (Film). After a one-hour consolidation interval that included an unrelated mechanical stimulation task, we interrogated the blastema for recall of positional identity using light-sheet imaging combined with spatial transcriptomics.
We obtained three principal findings. First, the latency to full epithelial closure was significantly shorter in the Sheet cohort than in the Pattern and Film cohorts, and the fidelity of proximodistal marker expression was markedly higher in the Sheet cohort for morphologically straightforward segments. Because cell density, cytokine composition, and exposure time were matched between the Sheet and Pattern cohorts, these results indicate that the Sheet condition induced deeper, more stable encoding of positional cues.
Second, across all cohorts, regeneration-stage gene activation localized to the bilateral blastema cores, the epidermal cap, the presumptive apical growth zone, and flanking mesenchyme, confirming the engagement of canonical positional-memory circuits (e.g., Hoxa9, Meis1, Fgf8, and Wnt5a).
Third, signal intensities within these territories were significantly higher for the Sheet cohort than for the Pattern and Film cohorts. These augmented activations could not be attributed to generalized wound stress or procedural complexity, as overall regeneration rates were comparable across groups.
The simultaneous superiority in both pattern fidelity and gene-network activation for the Sheet cohort suggests that biologic ECM sheets provide richer molecular and spatial encoding—possibly via native glycosaminoglycan topographies and integrin-mediated mechanotransduction—which can later be leveraged as effective recall cues, culminating in heightened activation of positional gene networks during regeneration.