a-decellularization-methodology-for-production-natural-acellular

A Decellularization Methodology for the Production of a Natural Acellular Intestinal Matrix

Successful tissue engineering involves the  combination of scaffolds with appropriate cells in vitro or in vivo.  Scaffolds may be synthetic, ...

University College London

In recent years, various types of stem cells have been characterized and their potential for cardiac regeneration has been investigated. We have previously described the isolation of broadly multipotent cells from amniotic fluid, defined as amniotic fluid stem (AFS) cells. The aim of this study was to investigate the therapeutic potential of human AFS cells (hAFS) in a model of acute myocardial infarction. Wistar rats underwent 30â€‰min of ischemia by ligation of the left anterior descending coronary artery, followed by administration of hAFS cells and 2â€‰h of reperfusion. Infarct size was assessed by 2,3,5-triphenyltetrazolium chloride staining and planimetry. hAFS cells were also analyzed by enzyme-linked immunosorbent assay to detect secretion of putative paracrine factors, such as the actin monomer-binding protein thymosin Î²4 (TÎ²4). The systemic injection of hAFS cells and their conditioned medium (hAFS-CM) was cardioprotective, improving myocardial cell survival and decreasing the infarct size from 53.9%Â±2.3% (control animals receiving phosphate-buffered saline injection) to 40.0%Â±3.0% (hAFS cells) and 39.7%Â±2.5% (hAFS-CM, P<0.01). In addition, hAFS cells were demonstrated to secrete TÎ²4, previously shown to be both cardioprotective and proangiogenic. Our results suggest that AFS cells have therapeutic potential in the setting of acute myocardial infarction, which may be mediated through paracrine effectors such as TÎ²4. Therefore, AFS cells might represent a novel source for cell therapy and cell transplantation strategies in repair following ischemic heart disease, with a possible paracrine mechanism of action and a potential molecular candidate for acute cardioprotection.

Amniotic fluid stem cells are cardioprotective following acute myocardial infarction.

Management of intestinal failure remains a clinical challenge and total parenteral nutrition, intestinal elongation and/or transplantation are partial solutions. In this study, using a detergent-enzymatic treatment (DET), we optimize in rats a new protocol that creates a natural intestinal scaffold, as a base for developing functional intestinal tissue. After 1 cycle of DET, histological examination and SEM and TEM analyses showed removal of cellular elements with preservation of the native architecture and connective tissue components. Maintenance of biomechanical, adhesion and angiogenic properties were also demonstrated strengthen the idea that matrices obtained using DET may represent a valid support for intestinal regeneration.

A rat decellularized small bowel scaffold that preserves villus-crypt architecture for intestinal regeneration.

Tissue engineering of the oesophagus has been proposed as a therapeutic alternative to organ transplantation. We previously demonstrated that a detergent enzymatic treatment (DET) is a valid method to obtain an acellular matrix with preservation of the native architecture. In this study, we aimed to develop a natural acellular matrix from pig oesophagus, as a valid framework for oesophageal replacement.

Detergent enzymatic treatment for the development of a natural acellular matrix for oesophageal regeneration.

A number of congenital and acquired disorders require esophageal tissue replacement. Various surgical techniques, such as gastric and colonic interposition, are standards of treatment, but frequently complicated by stenosis and other problems. Regenerative medicine approaches facilitate the use of biological constructs to replace or regenerate normal tissue function. We review the literature of esophageal tissue engineering, discuss its implications, compare the methodologies that have been employed and suggest possible directions for the future. Medline, Embase, the Cochrane Library, National Research Register and ClinicalTrials.gov databases were searched with the following search terms: stem cell and esophagus, esophageal replacement, esophageal tissue engineering, esophageal substitution. Reference lists of papers identified were also examined and experts in this field contacted for further information. All full-text articles in English of all potentially relevant abstracts were reviewed. Tissue engineering has involved acellular scaffolds that were either transplanted with the aim of being repopulated by host cells or seeded prior to transplantation. When acellular scaffolds were used to replace patch and short tubular defects they allowed epithelial and partial muscular migration whereas when employed for long tubular defects the results were poor leading to an increased rate of stenosis and mortality. Stenting has been shown as an effective means to reduce stenotic changes and promote cell migration, whilst omental wrapping to induce vascularization of the construct has an uncertain benefit. Decellularized matrices have been recently suggested as the optimal choice for scaffolds, but smart polymers that will incorporate signalling to promote cell-scaffold interaction may provide a more reproducible and available solution. Results in animal models that have used seeded scaffolds strongly suggest that seeding of both muscle and epithelial cells on scaffolds prior to implantation is a prerequisite for complete esophageal replacement. Novel approaches need to be designed to allow for peristalsis and vascularization in the engineered esophagus. Although esophageal tissue engineering potentially offers a real alternative to conventional treatments for severe esophageal disease, important barriers remain that need to be addressed.

Esophageal tissue engineering: a new approach for esophageal replacement.

Tissue-engineered skeletal muscle is urgently required to treat a wide array of devastating congenital and acquired conditions. Selection of the appropriate cell type requires consideration of several factors which amongst others include, accessibility of the cell source, in vitro myogenicity at high efficiency with the ability to maintain differentiation over extended periods of time, susceptibility to genetic manipulation, a suitable mode of delivery and finally in vivo differentiation giving rise to restoration of structural morphology and function. Potential stem-progenitor cell sources include and are not limited to satellite cells, myoblasts, mesoangioblasts, pericytes, muscle side-population cells, CD133(+) cells, in addition to embryonic stem cells, mesenchymal stem cells, amniotic fluid stem cells and induced pluripotent stem (iPS) cells. The relative merits and inherent limitations of these cell types within the field of tissue-engineering are discussed in the light of current research. Recent advances in the field of iPS cells should bear the fruits for some exciting developments within the field in the forthcoming years.

Skeletal Muscle Tissue Engineering: Which Cell to Use?

In the United States, more than 2600 kidneys are discarded annually, from the total number of kidneys procured for transplant. We hypothesized that this organ pool may be used as a platform for renal bioengineering and regeneration research. We previously showed that decellularization of porcine kidneys yields renal extracellular matrix (ECM) scaffolds that maintain their basic components, support cell growth and welfare in vitro and in vivo, and show an intact vasculature that, when such scaffolds are implanted in vivo, is able to sustain physiological blood pressure. The purpose of the current study was to test if the same strategy can be applied to discarded human kidneys in order to obtain human renal ECM scaffolds. The results show that the sodium dodecylsulfate-based decellularization protocol completely cleared the cellular compartment in these kidneys, while the innate ECM framework retained its architecture and biochemical properties. Samples of human renal ECM scaffolds stimulated angiogenesis in a chick chorioallantoic membrane assay. Importantly, the innate vascular network in the human renal ECM scaffolds retained its compliance. Collectively, these results indicate that discarded human kidneys are a suitable source of renal scaffolds and their use for tissue engineering applications may be more clinically applicable than kidneys derived from animals.

Discarded human kidneys as a source of ECM scaffold for kidney regeneration technologies.

Tissue engineering of autologous lung tissue aims to become a therapeutic alternative to transplantation. Efforts published so far in creating scaffolds have used harsh decellularization techniques that damage the extracellular matrix (ECM), deplete its components and take up to 5 weeks to perform. The aim of this study was to create a lung natural acellular scaffold using a method that will reduce the time of production and better preserve scaffold architecture and ECM components. Decellularization of rat lungs via the intratracheal route removed most of the nuclear material when compared to the other entry points. An intermittent inflation approach that mimics lung respiration yielded an acellular scaffold in a shorter time with an improved preservation of pulmonary micro-architecture. Electron microscopy demonstrated the maintenance of an intact alveolar network, with no evidence of collapse or tearing. Pulsatile dye injection via the vasculature indicated an intact capillary network in the scaffold. Morphometry analysis demonstrated a significant increase in alveolar fractional volume, with alveolar size analysis confirming that alveolar dimensions were maintained. Biomechanical testing of the scaffolds indicated an increase in resistance and elastance when compared to fresh lungs. Staining and quantification for ECM components showed a presence of collagen, elastin, GAG and laminin. The intratracheal intermittent decellularization methodology could be translated to sheep lungs, demonstrating a preservation of ECM components, alveolar and vascular architecture. Decellularization treatment and methodology preserves lung architecture and ECM whilst reducing the production time to 3 h. Cell seeding and in vivo experiments are necessary to proceed towards clinical translation.

Preservation of micro-architecture and angiogenic potential in a pulmonary acellular matrix obtained using intermittent intra-tracheal flow of detergent enzymatic treatment.

Article	Year
Amniotic fluid stem cells are cardioprotective following acute myocardial infarction. Stem cells and development\| PubMed ID: 21534857	2011
A rat decellularized small bowel scaffold that preserves villus-crypt architecture for intestinal regeneration. Biomaterials\| PubMed ID: 22305104	2012
Detergent enzymatic treatment for the development of a natural acellular matrix for oesophageal regeneration. Pediatric surgery international\| PubMed ID: 23124129	2013
Esophageal tissue engineering: a new approach for esophageal replacement. World journal of gastroenterology : WJG\| PubMed ID: 23322987	2012
Skeletal Muscle Tissue Engineering: Which Cell to Use? Tissue engineering. Part B, Reviews\| PubMed ID: 23679017	2013
Discarded human kidneys as a source of ECM scaffold for kidney regeneration technologies. Biomaterials\| PubMed ID: 23680364	2013
Preservation of micro-architecture and angiogenic potential in a pulmonary acellular matrix obtained using intermittent intra-tracheal flow of detergent enzymatic treatment. Biomaterials\| PubMed ID: 23727263	2013

Panagiotis Maghsoudlou

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