[Construction of tissue-engineered skin flap in vitro]

Tissue engineering aims to replace or repair damaged tissues using biological substitutes. Prior research has shown that adipose-derived stem cells (ASCs) can differentiate into multiple lineages, including adipogenic, osteogenic, and chondrogenic. However, the integration of ASCs with skin components to form a functional skin flap remains an open question. Existing methods focus on separate tissue constructs but lack a combined approach mimicking skin structure. This gap motivated the development of a composite skin flap integrating both skin and adipose tissues. No prior work had resolved how to assemble these tissues into a layered structure resembling native skin. The challenge lies in maintaining cell viability and differentiation during in vitro culture. This study addresses the need for a reproducible in vitro model of skin flap construction. The novelty lies in using ASCs as a source for adipose tissue and combining it with cultured keratinocytes and fibroblasts.

The aim of this research was to develop a tissue-engineered skin flap by combining adipose tissue and composite skin. The specific problem addressed is the lack of a reliable in vitro model that mimics the layered structure of native skin with an underlying adipose layer. The motivation stems from the need for better skin substitutes in reconstructive surgery and wound healing. The study sought to determine if ASCs could be induced to form adipose tissue and integrated with cultured skin components. The researchers aimed to test the feasibility of assembling these tissues into a three-layered structure. They also wanted to assess whether the cultured tissues retained their differentiated characteristics. The study focused on methods to culture and assemble ASCs, keratinocytes, and fibroblasts into a functional skin flap. The goal was to provide a reproducible protocol for tissue-engineered skin flap construction.

The study used human adipose-derived stem cells (ASCs) isolated from adipose tissue. These cells were cultured and tested for their ability to differentiate into adipogenic, osteogenic, and chondrogenic lineages. ASCs were mixed with collagen gel and induced for adipogenic differentiation over 15 days. The differentiation was confirmed using inverted microscopy, oil-red O staining, and HE staining. For the composite skin, keratinocytes and fibroblasts were isolated from human foreskin. Fibroblasts were mixed with collagen gel and cultured for 5 days. Keratinocytes were seeded on the gel and cultured for 4 days before being transferred to an air-liquid interface for 10 more days. The adipose tissue and composite skin were then assembled into a three-layered structure and cultured for 3 days. The assembled flap was analyzed using HE staining to confirm its layered structure and cell distribution.

The cultured ASCs demonstrated adipogenic, osteogenic, and chondrogenic differentiation potential. Adipogenic induction of ASCs in collagen gel for 15 days resulted in visible lipid accumulation confirmed by oil-red O staining. The composite skin consisted of a suprabasal layer of stratified keratinocytes, a middle layer with numerous cells, and a sublayer of adipogenic ASCs. HE staining showed distinct layers in the assembled skin flap, resembling normal skin structure. The adipose-derived ASCs formed a sublayer beneath the skin components. The three-layered structure was maintained after 3 days of co-culture. The results suggest that ASCs can be effectively used to generate adipose tissue for skin flap construction. The study provides evidence that in vitro skin flaps can be constructed by combining ASCs, keratinocytes, and fibroblasts.

The authors propose that tissue-engineered skin flaps can be successfully constructed by assembling adipose tissue and composite skin. Their findings suggest that ASCs can be induced to form adipose tissue and integrated with cultured skin components. The assembled flap exhibited a three-layered structure similar to native skin. The results indicate that ASCs are a viable source for adipose tissue in skin flap construction. The study provides a reproducible method for in vitro skin flap engineering. The authors suggest that this approach could be useful for reconstructive surgery and wound healing applications. The findings support the potential of ASCs in tissue engineering. The study concludes that combining ASCs with skin components is a feasible strategy for skin flap construction.