Human-Derived Scaffold Components and Stem Cells Creating Immunocompatible Dermal Tissue Ensuing Regulated Nonfibrotic Cellular Phenotypes

Abstract

Regeneration of large-sized acute and chronic wounds provoked by severe burns and diabetes is a major concern worldwide. The availability of immunocompatible matrix with a wide range of regenerative medical applications, more specifically, for nonhealing chronic wounds is an unmet clinical need. Extrapolating the in vitro tissue engineering knowledge for in vivo guided wound regeneration could be a meaningful approach. This study aimed to develop a completely human-derived and minimally immune-responsive scaffold comprising of acellular amniotic membrane (AM), fibrin (FIB) and hyaluronic acid (HA), termed AMFIBHA. The potential for in vivo guidance of skin regeneration was validated through in vitro dermal tissue assembly on the combination scaffold by growing human fibroblasts, differentiated from human adipose tissue-derived mesenchymal stem cells (hADMSCs). An effective method was standardized for obtaining decellularized amnion (dAM) for assuring better immuno-compatibility. The biochemical stability of dAM upon plasma sterilization (pdAM) confirms its suitability for both in vitro and in vivo tissue engineering. The problem of poor handling characteristics was solved by combining the dried dAM with fibrin derived from a clinically used fibrin sealant kit. An additional constituent HA, derived from human umbilical cord tissue, imparts the required water absorption and retention property for better cell migration and growth. Post sterilization, the combination scaffold AMFIBHA demonstrated hemo-/cytocompatibility, confirming the absence of detergent residuals. Upon long-term (20 days/40 days) culture of hADMSC-derived fibroblasts, the suppleness of generated tissue was established by demonstrating regulated deposition of collagen, elastin, and glycosaminoglycans using both qualitative and quantitative measurements. Regulated expressions of transforming growth factors-beta 1 (TGF-β1) & TGF-β3, alpha smooth muscle actin (α-SMA), fibrillin-1, collagen subtypes, and elastin suggest non-fibrotic fibroblast phenotype, which could be an effect of microenvironment endowed by the AM, FIB, and HA. In burn wound model experiments, immune response to cellular AM was prominent as compared to untreated/sham control wounds and decellularized AM-treated and AMFIBHA-treated wounds, ensuring biocompatibility. Wound regeneration with complete epithelialization, angiogenesis, development of rete pegs, and other skin appendages were clearly visualized in 28 days after treating large-sized (4 × 4 cm2), debrided, full-thickness third-degree burn wounds, indicating guided wound regeneration potential of AMFIBHA dermal substitute.

Optimization of decellularized human placental macroporous scaffolds for spermatogonial stem cells homing

“…As mentioned earlier in introduction, decellularization and preservation methods can remarkably affect the biomechanical and biological properties of tissue [ 17 , 18 ]. Therefore, it is very important to use an optimal concentration and incubation time point of detergents with successful removal of cells and minimal negative effects on ECM proteins and growth factors [ 17 , 29 ]. In our previous study, we optimized a decellularization procedure for decellularization of amniotic membrane using chemical agents such as EDTA and NaOH [ 28 ].…”

Extracellular matrix‐based combination scaffold for guided regeneration of large‐area full‐thickness rabbit burn wounds upon a single application

“…Exogenous HA can provide a moist wound healing environment allowing autolytic debridement 23‐25 . We have reported in vitro tissue engineering of a combination wound care product comprising cell‐less AM, human FIB, and HA, termed as AMFIBHA 26 . The previous study confirmed its hemo‐/cyto‐compatible nature with optimum physicochemical properties and supple dermal tissue regeneration.…”

“…The study design ensured the implantation of four samples of each specific type on four different animals. Full‐thickness burn associated damage of skin was confirmed using histological analysis of burnt tissue, as published earlier 26 . For the experiment, prepared the wounds and exposed fresh tissue by surgical debridement of the burnt full‐thickness skin along the wound edges, extending to the panniculus carnosus layer using a sterile scalpel blade before application of the scaffolds.…”

“…For this study, the AM preparation followed a published protocol 26 . The final processed AM was in hydrogen peroxide (H 2 O 2 )‐plasma sterilized (Sterrad Plasma Sterilizer, USA) lyophilized form, stored at 4°C until use, and was trimmed into 16 cm 2 (4 cm x 4cm) size.…”

Tissue Engineering for Mimics and Modulations of Immune Functions

“…developed a stable dermal tissue graft with minimal immune responses by combining decellularized AM together with fibrin glue and HA. [ 99 ] For bone tissue engineering, Wu et al. aimed to transform the commonly used bone ECM filler particles derived from porcine thighbones to a hydrogel form that was more likely to reduce host immune responses.…”