Development of acellular dermis from porcine skin using periodic pressurized technique

Abstract

In this work, a new method for producing acellular dermis (ADM), a natural scaffold used for dermal replacement, from porcine skin was developed. Fresh porcine skin from local slaughterhouse was dehaired by sodium sulphide following by epidermis removal using glycerol. After fat removal by chloroform/methanol (2/1 v/v) solvent, cellular components were removed using enzymatic treatment incorporated with a periodic pressurized technique. The effects of enzyme type (trypsin and dispase II) and periodic pressurized conditions on the efficiency of cell removal were investigated. When periodic pressure was applied, enzymatic treatment time could be shorten since the enzyme solution was able to penetrate into tight dermis. As a result, cells could be easily removed from porcine skin as noticed quantitatively by DNA assay and qualitatively by H&E staining. When enzyme refreshment was introduced into the decellularized process, the percentage of cell removal was further enhanced. This ensured that no inhibitions effect from the removed cells on enzyme-substrate interaction. Moreover, short-time enzymatic treatment with periodic pressurized technique could prevent the disruption of dermal structure, as observed by SEM. Dispase II can be used to remove cell better than trypsin in the periodic pressurized technique. However, in vivo study indicated that numerous fibroblast from the host tissue infiltrated into ADM prepared using both enzymes. Neo-collagen and neo-capillaries were produced in both implanted ADMs. The result elucidated that the use of periodic pressurized technique with enzymatic treatment has a high potential to be a new method to produce ADM for skin tissue engineering.

Porcine skin as a source of biodegradable matrices: alkaline treatment and glutaraldehyde crosslinking

“…Their unlimited availability is one of the advantages in using heterologous matrices. Porcine skin is one of the several heterologous matrices under study, mainly on account of its high similarity to human skin and its biocompatibility, mechanical resistance and low antigenicity [6] . Porcine skin is a well-suited temporary wound dressing for seconddegree burns and usually promotes scar-free healing with an average healing time of about 10 days [5] .…”

Biomaterials from Decellularized Tissues

“…Like the ECM from most tissues, SIS-ECM and UBM-ECM are composed of more than 90% collagen, the majority of which is type I [46,75] with varying amounts of collagen type III, IV, V, VI, and VII also present. The ECM derived from the esophageal mucosa (EM-ECM) [76], skeletal muscle (SM-ECM), and dermis (D-ECM) show similar composition [76][77][78][79]. The collagen composition of ECMs originating from different tissues will vary depending on whether or not a basement membrane is present.…”

Heart Valve Matrices: State of the Art and Perspectives

“…Спирты, например глицерол, вызывают дегидратацию и лизис клеток [40]. Он способен удалять фосфолипиды из клапанных кондуитов, усиливающие кальци-фикацию протеза [41,42].…”

“…Липазы, как было показано, само-стоятельно не способны удалить все жиры, поэтому используются только как одна их составляющих сложных протоколов [43,44]. Диспаза -нейтральная протеа-за -позволяет более эффективно, чем трипсин, удалять клеточные компоненты из ткани, одновременно сильнее повреждая матрикс, но при этом способствуя ускоренной репопуляции [40]. Также трипсин и диспаза используются в качестве дополнения к детергентам при децеллюляризации сложных тканей [25].…”

“…Многократное же повторение цикла не приводит к зна-чимой потере белков, но отрицательно сказывается на механических свойствах матрикса [59,40,43,51,60].…”