Decellularization of caprine esophagus using fruit pericarp extract of Sapindus mukorossi.

Biological detergents like sodium deoxycholate, sodium dodecyl sulphate and Triton X-100 impairs the collagenous and non-collagenous proteins, glycosaminoglycans and growth factors. Further, certain chemical and enzymes are responsible for residual cytotoxicity in the decellularized extracellular matrix. The main focus of this study was to explore the decellularization property of soap nut pericarp extract (SPE) for development of decellularized tubular esophageal scaffold. For this 2.5, 5.0 and 10% concentrations of SPE were used for decellularization of caprine esophageal tissues. Histological analysis of hematoxylin and eosin and Masson's trichrome stained tissue samples confirmed decellularization with preservation of extracellular matrix microarchitecture. Scanning electron microscopic images of luminal surface of decellularized esophageal matrix showed randomly oriented collagen fibres with large interconnected pores and cells were absent. However, the external surface was more textured with fibrous structures and collagen fibres were well preserved. DAPI stained decellularized tissues revealed complete removal of nuclear components, verified by DNA content measurement and SDS-PAGE. The FTIR spectra of decellularized esophagus shows absorption peaks of amide A, B, I, II and III. Elastic modulus of the decellularized esophagus scaffolds increased (P > 0.05) as compared to native tissues. Histological and scanning electron microscopic evaluation of in vitro seeded scaffolds showed attachment and growth of primary chicken embryo fibroblasts over and within the decellularized scaffolds. It was concluded that 5% SPE is ideal for preparation of cytocompatible decellularized caprine esophageal scaffold with well-preserved extracellular matrix architecture and, may be used as an alternative to biological detergents and other chemicals.

Development of decellularized aortic scaffold for regenerative medicine using Sapindus mukorossi fruit pericarp extract.

The aim of this study is to develop a novel decellularization method using aqueous extract of soap nut pericarp (SPE) and its evaluation using hematoxylin-eosin staining, scanning electron microscopy, diamidino-2-phenylindol (DAPI) staining, mechanical testing, sodium dodecyl sulfate polyacrylamide gel electrophoresis and DNA quantification. The presently available decellularization agent raises some concerns due to the potential for presence of residual cytotoxic agents in the extracellular matrix. Histological analysis of hematoxylin and eosin and masson's trichrome stained processed aortic samples shows complete decellularization with preservation of extracellular matrix microarchitecture at 120 h. Further, staining of tissue samples with DAPI demonstrates complete removal of DNA fragments. Quantitative evaluation of DNA in the decellularized aorta tissues demonstrated a significant (P < 0.01) decrease in DNA content as compared to native tissues. Collagen quantification assay indicate no significant (P> 0.05) difference in its content between native and decellularized caprine aorta. Tensile strength of the decellularized scaffolds decreased non-significantly (P > 0.05) when compared to native tissues. There was no significant (P > 0.05) difference in young's modulus of elasticity, stiffness and stretch ratio between native aortic tissues and decellularized aortic scaffolds. Histological and scanning electron microscopic examination of in vitro cultured scaffold demonstrated the cell viability and proliferation of primary chicken embryo fibroblasts. SPE treatment is thus capable of producing cytocompatible decellularized caprine aorta scaffold with preservation of extracellular matrix architecture for vascular tissue engineering and could be applied widely as one of the decellularization agent.

Primary chicken embryo fibroblasts seeded acellular dermal matrix (3-D ADM) improve regeneration of full thickness skin wounds in rats.

Rat skins were deepithelialized and decellularized by hypertonic saline and sodium deoxycholate (SDC), respectively. Primary chicken embryo fibroblasts (P-CEF) were cultured and seeded on prepared acellular dermal matrix (ADM). A full thickness skin defect (20×20 mm(2)) was created in thirty-six rats and randomly divided into three equal groups. Defect was left open, repaired with ADM and ADM seeded with P-CEF (3-D ADM) in groups 1, 2 and 3, respectively. By day 28, the treated wounds healed completely without scar. By day 7 hydroxyproline contents was higher in group 3 as compared to groups 1 and 2. There was slightly more B cell response in animals implanted with ADM and 3-D ADM. At day 21, stimulation index was lower with acellular dermis antigen as compared to 3-D ADM antigen. In group 1 on day 3, the granulation tissue showed more inflammatory reaction, fibroplasia and neovascularization as compared to group 2 and 3. By day 28, there was complete epithelization was observed in all groups over. However, a large scar was observed in group 1. The graft was completely absorbed and replaced with densely thick and best arranged collagen fibers. On day 7, malonyldialdehyde and superoxide dismutase levels were significantly (P<0.05) increased in group 1. Reduced glutathione values increased and reached to near normal in groups 2 and 3. Catalase values were significantly (P<0.05) higher in group 1 at different time intervals. SEM samples of group 2 showed ingrowth of fibroblasts into acellular matrix at host graft junction. However, in group 3 fibroblasts were infiltrated within the pores of graft. It was concluded that P-CEF cells seeded ADM facilitated early and better healing.