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The Next Big Fat Thing: A Cleaner, More Potent Injectable Fat Allograft Processed Using Supercritical CO₂
Sophia Salingaros^*1, Abby Chopoorian Fuchsman¹, Kate Manley¹, Jini Jeon¹, Samuel Medina¹, Matthew W. Liao¹, Ankita Sarkar², Eric Eisenhut², Tony Eisenhut², David M. Bednarski², Xue Dong¹, Jason A. Spector^1
1Weill Cornell Medicine, New York, NY; ²NovaSterilis, Inc, Lansing, NY

BACKGROUND: Autologous fat grafting is widely used for soft tissue reconstruction and augmentation, but unpredictable volume retention and donor site morbidity remain a concern. Decellularized adipose extracellular matrix (adECM) is a promising alternative. While conventional decellularization employs harsh chemicals that can denature proteins and leave toxic residues, we have engineered a human-derived adECM using supercritical carbon dioxide (scCO₂), promising a gentler, eco-friendly decellularization and enhanced tissue regeneration. METHODS: Abdominoplasty adipose tissue was mechanically homogenized, delipidated, and underwent a non-wetting scCO₂ cleansing step, two cycles of scCO₂ decellularization, cryomilling, and scCO₂ terminal sterilization. Resultant adECM (0.4mL of 0.5g sample/mL PBS) was injected into dorsal and cranial subcutaneous pockets of wild type C57BL/6 mice through a 20-gauge catheter. Commercial cross-linked hyaluronic acid (HA) was injected (0.4mL) as a control. Samples were explanted after 1-, 3- and 6-months. RESULTS: Residual DNA in processed adECM was 44ng/mg, below the accepted decellularization threshold of 50ng/mg. After 6-months, adipose flank volume was 307mm³ (77% of initial volume), adipose cranial 273mm³ (68%), and HA 1393mm³ (348%) due to its hygroscopic nature. Cell density was significantly higher in adECM versus HA grafts (p<0.0001) and declined over time as inflammation subsided. In adECM grafts, 6-month M1/M2 macrophage ratio was 0.30 (predominately pro-regenerative M2 phenotype over pro-inflammatory M1), perilipin expression revealed adipocytes increasing in number and size over time (p<0.0001), CD31 expression demonstrated neo-vascularization, and Picrosirius red staining demonstrated new collagen deposition which matured over time. HA explants had sparse cells and no adipocytes or new collagen. CONCLUSION: We present an innovative scCO₂-decellularized "off-the-shelf" adECM with 73% volume retention after 6-months in vivo, with robust architectural remodeling including adipogenesis, angiogenesis, and collagen deposition.