Development of a large‐volume human‐derived adipose acellular allogenic flap by perfusion decellularization

Abstract

In reconstructive surgery, transfer of patients' tissue (autologous flaps) is routinely used to repair large soft tissue defects caused by surgery, trauma, chronic diseases, or malformations; unfortunately, this strategy is not always possible and often creates a secondary defect in the donor site of the tissue. Tissue-engineered synthetic flaps are currently unable to repair clinically-relevant, large-volume defects; allogenic flaps from cadaveric donors could provide a ready-to-use biological alternative if treated with methods to avoid the immune-rejection of the donor's cells. Here, we describe the successful decellularization of a large (> 800 cc) human-derived adipose flap through a perfusion apparatus; we demonstrate the complete removal of the immunogenic cellular components of the flap with the retention of its structural components and vascular network. Our aim is to obtain a universally compatible, off-the-shelf acellular allogenic flap that could be recellularized with cells from recipient patients to provide a tissue-engineered allogenic/autologous alternative for reconstruction of large-volume soft-tissue defects.

Characterization and Proteomic Analysis of Decellularized Adipose Tissue Hydrogels Derived from Lean and Overweight/Obese Human Donors

“…The amount of fibrotic collagen bundles was calculated as a percentage of total collagen content per each image. Images were loaded into ImageJ software [ 37 ] and total collagen content was isolated by color using the “Color Deconvolution” plug‐in and fibrotic collagen bundles were further isolated using a band‐pass filter in the “Threshold Color” plug‐in. [ 38 ] Images were then subjected to a threshold such that colored pixels were converted to white and noncolored pixels were converted to black.…”

“…Hydrogels were then dried using DCP1 critical point drying apparatus (Denton vacuum Inc; Moorestown, NJ) and sputter coated (Electron Microscopy Sciences; Hatfield, PA) with platinum for 4 min. Hydrogels were imaged with FEI quanta 3D FEG FIB/SEM (5.0 KV) and analyzed using ImageJ software (DiameterJ plugin) [ 37 ] as described previously. [ 39 ] Fiber diameter, pore area, and fiber intersection density were quantified from six independent fields of view (FOV) from three hydrogels ( n = 18 per group) at 50 000× magnification for the lDAT and oDAT groups (data presented as mean ± S.D.…”

Automated Decellularization of the Rodent Epigastric Free Flap: A Comparison of Sodium Dodecyl Sulfate–Based Protocols

“…For similar perfusion decellularization protocols for vascularized composite tissues, Giatsidis et al successfully utilized a 100-hour (∼4 days) protocol for human abdominal panniculectomy tissue. 23 Jank et al applied 10 days of SDS perfusion for fasciocutaneous groin flap in miniature swine with success. 20 Qu et al utilized varying durations as 12, 24, and 72 hours of SDS perfusion combined with agitation for rat fasciocutaneous epigastric free flap and suggested the use of a combined protocol with SDS exposure for 24 hours or longer.…”

“…13 14 15 16 17 18 However, these approaches have seen minimal crossover into the microsurgical field with composite tissues. 19 20 21 22 23…”

“…[13][14][15][16][17][18] However, these approaches have seen minimal crossover into the microsurgical field with composite tissues. [19][20][21][22][23] Sodium dodecyl sulfate (SDS) is the predominant choice of detergent for decellularization protocols. 18,24 However, SDS also has deleterious effects on the ultrastructure and capillary network of the scaffold.…”