Preparation and characterization of 3D plant-based scaffold based on decellularization method in liver tissue engineering / 中国组织工程研究

BACKGROUND:Tissue engineering has brought new hope to the clinical challenge of liver failure,and the preparation of plant-derived decellularized fiber scaffolds holds significant importance in liver tissue engineering. OBJECTIVE:To prepare apple tissue decellularized scaffold material by using fresh apple slices and a solution of sodium dodecyl sulfate,and assess its biocompatibility. METHODS:Fresh apples were subjected to decellularization using phosphate buffer saline and sodium dodecyl sulfate solution,separately.Afterwards,the decellularized apple tissues and apple decellularized scaffold materials were decontaminated with phosphate buffer saline.Subsequently,scanning electron microscopy was used to assess the effectiveness of decellularization of the apple materials.Adipose-derived mesenchymal stem cells were extracted from the inguinal fat BALB/C of mice,and their expression of stem cell-related markers(CD45,CD34,CD73,CD90,and CD105)was identified through flow cytometry.The cells were then divided into a scaffold-free control group and a scaffold group.Equal amounts of adipose-derived mesenchymal stem cells were seeded onto both groups.The biocompatibility of the decellularized scaffold with adipose-derived mesenchymal stem cells was evaluated using CCK-8 assay,hematoxylin-eosin staining,and phalloidine staining.Cell adhesion and growth on the scaffold were observed under light microscopy and scanning electron microscopy.Furthermore,the scaffold was subdivided into the non-induced group and the hepatogenic-induced group.Adipose-derived mesenchymal stem cells were cultured on the decellularized apple scaffold,and they were cultured for 14 days in regular culture medium or hepatogenic induction medium for comparison.Immunofluorescent staining using liver cell markers,including albumin,cytokeratin 18,and CYP1A1,was performed.Enzyme-linked immunosorbent assay was used to detect the secretion of alpha fetoprotein and albumin.Additionally,scanning electron microscopy was employed to observe the morphology of the induced cells on the scaffold,verifying the expression of liver cell-related genes on the decellularized scaffold material.Finally,the cobalt-60 irradiated and sterilized decellularized apple scaffolds were transplanted onto the surface of mouse liver and the degradation of the scaffold was observed by gross observation and hematoxylin-eosin staining after 28 days. RESULTS AND CONCLUSION:(1)The scanning electron microscopy results revealed that the decellularized apple scaffold material retained a porous structure of approximately 100 μm in size,with no residual cells observed.(2)Through flow cytometry analysis,the cultured cells were identified as adipose-derived mesenchymal stem cells.(3)CCK-8 assay results demonstrated that the prepared decellularized apple tissue scaffold material exhibited no cytotoxicity.Hematoxylin-eosin staining and phalloidine staining showed that adipose-derived mesenchymal stem cells were capable of adhering and proliferating on the decellularized apple tissue scaffold.(4)The results obtained from immunofluorescence staining and enzyme-linked immunosorbent assay revealed that adipose-derived mesenchymal stem cells cultured on the decellularized apple scaffolds exhibited elevated expression of liver-specific proteins,including albumin,alpha-fetoprotein,cytokeratin 18,and CYP1A1.These results suggested that they were induced differentiation into hepatocyte-like cells possessing functional characteristics of liver cells.(5)The decellularized apple scaffold implanted at 7 days has integrated with the liver,with partial degradation of the scaffold observed.By 28 days,the decellularized apple scaffold has completely degraded and has been replaced by newly-formed tissue.(6)The results indicate that the decellularized scaffold material derived from apple tissue demonstrates favorable biocompatibility,promoting the proliferation,adhesion,and hepatic differentiation of adipose-derived mesenchymal stem cells.

Current status of applying matrigel to in vitro models for cancer reseach / 肿瘤

The components of the tumor microenvironment have a complex composition and play an important regulatory role in tumor evolution.The extracellular matrix(ECM)serves as a vital mediator in regulating cancer advancement within the TME.Incorporating ECM into tumor modeling allows for a more comprehensive simulation of the TME in vitro.As a medium for organoid growth,the matrix gel fulfills the functions of ECM.ECM is normally derived from the recombinant basement membrane of mouse sarcoma(Matrigel);however,batch-to-batch variations in Matrigel affect experiment reproducibility.To overcome such issues,researchers have designed synthetic matrix gels.This paper provides an overview of the current research in the application of matrix gels for tumor organoid modeling,offering insights for continuous optimization of organoid culture,thereby achieving more efficient and cost-effective organoid construction and advancing the translation of basic research to clinical applications.

The study of the whole bladder acellular matrix prepared by the perfusion decellularization system combined with the adipose-derived stem cells to construct the tissue engineering bladder / 中华泌尿外科杂志

Objective:To prepare the whole bladder acellular matrix (BAM) using the self-designed perfusion decellularization system, and evaluate the feasibility of constructing the tissue engineering bladder with the adipose-derived stem cells (ADSCs).Methods:This study was conducted from October 2020 to April 2021. The self-designed perfusion decellularization system was used, and four different decellularization protocols (group A, group B, group C and group D) were formulated, according to the flow direction of the perfusate and the action time of different decellularization solutions. Among them, the urethral orifice of the bladder tissue was used as the outflow tract of the perfusion fluid in groups A and B. The top of the bladder was cut off and used as the outflow tract of the perfusion fluid in groups C and D. In groups A and C, 1% Triton X-100 was treated for 6 h, and 1% sodium dodecyl sulfate (SDS) was treated for 2 h. In groups B and D, 1% Triton X-100 was treated for 7 h, and 1% sodium dodecyl sulfate (SDS) was treated for 1 h. In addition, the tissue in the normal bladder group was directly obtained from the natural bladder tissue, which did not require perfusion, cryopreservation and thawing. The fast and efficient decellularization protocol was screened out through HE, DAPI, Masson trichrome and Alcian Blue staining and quantitative analyses to prepare the whole bladder scaffold. The prepared BAM was used as the scaffold material, and the ADSCs were used as the seeding cells to construct the tissue engineering bladder. HE and DAPI staining were used to observe the distribution of ADSCs on the BAM.Results:HE and DAPI staining showed that there was no obvious nuclear residue in the group C. Masson trichrome and Alcian Blue staining showed that the collagen structure and glycosaminoglycan were well preserved in the group C. There was no significant difference in bladder wall thickness between the group C and the normal bladder group [(975.44±158.62)μm vs.(1 064.49±168.52)μm, P > 0.05]. The DNA content in the group C [(43.59 ±4.59) ng/mg] was lower than that in the normal bladder group, group A, group B and group D [(532.50±26.69), (135.17±6.99), (182.49±13.69) and(84.00±4.38)ng/mg], and the difference was statistically significant ( P<0.05). The collagen content [(10.98 ± 0.29)μg/mg] and glycosaminoglycan content [(2.30±0.18)μg/mg] in group C were not significantly different with those in the normal bladder group [(11.69±0.49) and (2.36±0.09)μg/mg, P>0.05]. Scanning electron microscopy showed that a large number of pore structures could be observed on the surface of the prepared BAM in groups A-D and were facilitated to cell adhesion. The isolated and cultured ADSCs were identified by flow cytometry to confirm the positive expression of CD90 and CD29, and the negative expression of CD45 and CD106. Live/dead staining and CCK-8 detection confirmed that the prepared BAM in the group C had no cytotoxicity. HE and DAPI staining showed that a large number of ADSCs were distributed on the surface and inside of the tissue engineering bladder. Conclusions:The whole bladder shape BAM prepared by the self-designed perfusion decellularization system could be used as the scaffold material for bladder tissue engineering, and the constructed tissue engineering bladder could be used for bladder repair and reconstruction.

Preparation and histological evaluation of human decellularized adipose tissue / 解剖学报

Objective To decellularize adipose tissue derived from human liposuction, evaluate the histological composition of human decellularized adipose tissue, and explore the possibility for it as tissue engineering scaffold materials. Methods The adipose tissue derived from five human liposuction was decellularized by a series of physical chemistry and enzymology method . The general morphology of the decellularization process was observed. HE staining, Masson staining, Gomori staining, oil red 0 staining and DAPI staining were performed and the content of DNA, collagens and glyeosaminoglycans remaining in the decellularized adipose tissue were quantitatively detected. Results The decellularized adipose tissue was some kind of milky, soft, variable insoluble material. HE staining and DAPI staining showed no residual cell components. Masson staining showed the presence of collagen fibers. Gomori staining showed the presence of reticular fibers, and oil red 0 staining showed no visible lipid residues. The residual DNA content in the tissue is very tiny and the collagen and glycosaminoglycan contents were retained. Conclusion The adipose tissue derived from the body' s liposuction can be decellularized by physical, chemical, and enzymatic method . The cells and lipid in the decellularized adipose tissue can be effectively removed, and the components of the extracellular matrix of the adipose tissues are mostly retained.

Research progress of intervertebral disc repair with decellularized matrix materials / 中国组织工程研究

BACKGROUND: With the development of tissue engineering, the repair and regeneration of disc becomes possible. Decellularized extracellular matrix is an important means for disc regeneration. OBJECTIVE: To review and summarize the processing, quality control and application of decellularized matrix materials applied in intervertebral disc regeneration in recent years and put forward the prospect. METHODS: PubMed, Web of Science and CNKI databases were searched for the articles concerning decellularized methods and decellularized matrix repairing intervertebral disc with the search terms of “intervertebral disc, decellularization, extracellular matrix, scaffold material, tissue engineering” in English and Chinese, respectively. After screening based on the inclusion and exclusion criteria, the articles with high relevance were included for review. RESULTS AND CONCLUSION: The decellularized tissue-engineered intervertebral disc aims to maintain the physiologically relevant bioactivators to a great extent, improve mechanical properties and biocompatibility, and reduce immunogenicity. The decellularized matrix material can simulate the microenvironment of the extracellular matrix in the intervertebral disc. As a cell carrier, it can well induce the differentiation of seed cells, which has achieved certain progress in the repair of intervertebral discs. However, further studies need to address the following issues: proper porosity of decellularized matrix materials, immunological rejection, implant ways in vivo and repair effect.

Extraction of porcine cancellous bone by supercritical carbon dioxide / 中国组织工程研究

BACKGROUND: In recent years, there have been many studies on the use of supercritical fluids for biological material treatment in countries outside China. However, little is reported on application of supercritical fluids to bone tissue extraction, in particular in China. OBJECTIVE: To evaluate the effectiveness of supercritical carbon dioxide extraction technology in the treatment of porcine femur cancellous bone and its effect on bone biological properties. METHODS: Porcine femoral bone blocks that were subjected to supercritical carbon dioxide extraction (study group) or not (control group) were prepared to determine bone mineral density, microstructure, maximum compressive strength, elastic modulus, bone tissue composition, collagen content and perform histological analysis. Bone marrow mesenchymal stem cells (BMSCs) were inoculated into two groups of bone blocks, and cultured for 1 day. The microporous structure of trabecular bone and cell adhesion and growth in bone material-cell composite were observed by scanning electron microscopy. The two groups of bone blocks were implanted subcutaneously in SD rats. The inflammatory reaction of subcutaneous tissue was observed histologically at 1, 2 and 4 weeks after surgery. The experimental protocol had been approved by the Animal Ethics Committee of Chinese PLA General Hospital, China. RESULTS AND CONCLUSION: There were no significant differences in pore size, bone mineral density, maximum compressive strength, elastic modulus and collagen content between the study and control groups (P>0.05). Scanning electron microscopy showed that in the control group, the material pores had poor connectivity and there was soft tissue residue; in the study group, material pores were connected to each other and the structure was intact. Fourier transform infrared spectroscopy and X-ray diffraction analysis showed that the two groups of bone tissue materials had similar absorption and diffraction peaks. Thermogravimetric analysis showed that supercritical carbon dioxide extraction could reduce water content in bone tissue. Hematoxylin-eosin staining showed that there were no soft tissue residues in the bone, and the cell residues in the bone pit were significantly reduced in the study group, while soft tissue and cell residues were observed in the control group. Sirius red staining and modified Masson staining showed that the structure of bone collagen in the study group was intact, the cytoplasmic components reduced, and the cytoplasmic components in the control group remained significantly. Scanning electron microscopy showed that there was no obvious cell adhesion in the control group, but cell adhesion growth was obvious in the study group. Perivascular inflammatory response in the bone tissue implantation region was obviously weaker in the study group than in the study group. These results suggest that supercritical carbon dioxide extraction technology is an effective and environment-friendly bone tissue processing technology. It can effectively remove porcine cancellous bone cells and soft tissue without affecting its collagen structure and content and mechanical properties, retaining intact bone pore structure, increasing cell adhesion and growth, and effectively reducing inflammatory rejection.

Preparation method of acellular matrix and its application in tissue engineering of parotid gland / 中国组织工程研究

BACKGROUND: Bioscaffolds composed of acellular matrix are widely used In animal and clinical research to repair and reconstruct tissues and organs, but all decellularlzatlon methods will destroy matrix structure and function to some extent. OBJECTIVE: To review the preparation methods, advantages and disadvantages of acellular matrix and its application in the study of parotid gland and other tissue engineering. METHODS: A computer-based search of CNKI, CBM, PubMed and Elsevier databases was performed to retrieve related articles published from 2008 to 2019. The search term were “decellular matrix, preparation method, parotid gland, tissue engineering, regeneration, decellularizatlon, preparation method, parotid gland tissue engineering” In Chinese and English, respectively. Seventy-four eligible articles were Included. RESULTS AND CONCLUSION: Acellular matrix preparation methods of most tissues and organs require chemical, biological (enzyme), physical methods and the their combination, which depend on the thickness, compositions and nature. Although not all decellularization methods can remove cellular components from tissues and organs, tissues and organs that completely remove cells have the advantage of remodeling tissue specificity, providing a favorable microenvironment for the proliferation and differentiation of inoculated cells. Due to the complex structure of the parotid gland and the challenges of tissue engineering In clinical applications, clinical transplantation In patients Is limited. In vivo studies In this field are limited to animals, and applications based on submandibular gland acellular matrix bioscaffold materials are expected to be a favorable source of organizational parotid glands.

Biophysical methods for quality evaluation of decellularized and recellularized tissue-engineered constructs of organs and tissues

Tissue engineering is rapidly growing now and can become a promising alternative to transplantation of organs and tissues,as it is devoid of major shortcomings of transplantology, such as acute shortage, complexity of selection, delivery andstorage of donor material, lifelong immunosuppressive therapy. One of the most widely known methods of obtainingbiological scaffolds for the subsequent creation of tissue-engineered constructs of organs and tissues is decellularization.The evaluation of the quality of the obtained scaffolds, based on the study of the viability of cell structures in decellularizedand recellularized matrices, is one of the priorities of modern regenerative medicine worldwide. In this investigation, thebiophysical criteria of decellularization and recellularization of tissue-engineered constructs based on the evaluation of thegeneration of free radicals in native, decellularized and recellularized tissues by EPR spectroscopy and chemoluminescencein a complex assessment of the quality of biological matrixes obtained are considered using intrathoracic organs and tissuesof rats. It has been established that the intensity indices of free radical generation in native and recellularized tissues ofanimal organs, as well as in decellularized matrices, can serve as one of the express criteria for quantitative assessment ofcell structures viability

A Comparative Study of the Effects of Different Decellularization Methods and Genipin-Cross-Linking on the Properties of Tracheal Matrices

BACKGROUND: Different decellularization methods can affect the integrity and the biomechanical and biocompatible properties of the tracheal matrix. Natural cross-linking with genipin can be applied to improve those properties. The goals of this study were to evaluate the effects of different decellularization methods on the properties of genipin-cross-linked decellularized tracheal matrices in rabbits. METHODS: The tracheas of New Zealand rabbits were decellularized by the Triton-X 100-processed method (TPM) and the detergent-enzymatic method (DEM) and were then cross-linked with genipin. Mechanical tests, haematoxylin-eosin staining, Masson trichrome staining, Safranin O staining, DAPI staining, scanning electronic microscopy (SEM), and biocompatibility tests were used to evaluate the treatment. The bioengineered trachea and control trachea were then implanted into allogeneic rabbits for 30 days. The structural and functional analyses were performed after transplantation. RESULTS: The biomechanical tests demonstrated that the biomechanical properties of the decellularized tracheas decreased and that genipin improved them (p < 0.05). The histological staining results revealed that most of the mucosal epithelial cells were removed and that the decellularized trachea had lower immunogenicity than the control group. The analysis of SEM revealed that the decellularized trachea retained the micro- and ultra-structural architectures of the trachea and that the matrices cross-linked with genipin were denser. The biocompatibility evaluation and in vivo implantation experiments showed that the decellularized trachea treated with the DEM had better biocompatibility than that treated with the TPM and that immunogenicity in the cross-linked tissues was lower than that in the uncross-linked tissues (p < 0.05). CONCLUSION: Compared with the trachea treated with the TPM, the rabbit trachea processed by the DEM had better biocompatibility and lower immunogenicity, and its structural and mechanical characteristics were effectively improved after the genipin treatment, which is suitable for engineering replacement tracheal tissue.

Decellularization of human umbilical arteries / Descelularización de las arterias umbilicales humanas

SUMMARY: Arterial obstruction in small diameter (<6 mm) vessels are many times treated with grafts, however autologous aren't always available and synthetic have a high rate of complications. Decellularization of umbilical arteries may provide a solution, but the ideal method is debatable. We compare effectiveness between SDS and Triton X-100. Umbilical cords obtained from full term pregnancies with normal development and no evident complications in the newborn, were micro-dissected within 12 h and stored in phosphate buffered saline without freezing. Arteries were then processed for decellularization using 0.1 % and 1 % SDS, and 1 % Triton X100 protocols. Evaluation of cellular and nuclear material, collagen fibers, elastic fibers, and glycosoaminoglycans of the extracellular matrix (ECM) were evaluated as well as morphometric analysis under histological and immunohistochemical techniques. Triton X-100 was ineffective, preserving nuclear remains identified by immunofluorescence, had the most notable damage to elastic fibers, and decrease in collagen. SDS effectively eliminated the nuclei and had a less decrease in elastic fibers and collagen. Laminin was preserved in all groups. No significant differences were identified in luminal diameters; however the middle layer decreased due to decellularization of muscle cells. In conclusion, 0.1 % SDS decellularization was the most effective in eliminating cells and preserving the main components of the ECM.

RESUMEN: La obstrucción arterial en vasos de pequeño diámetro (<6 mm) se trata muchas veces con injertos, sin embargo, los autólogos no siempre están disponibles y los sintéticos tienen una alta tasa de complicaciones. La descelularización de las arterias umbilicales puede proporcionar una solución, pero el método ideal es discutible. Comparamos la efectividad entre los métodos SDS y Triton X-100. Cordones umbilicales obtenidos a partir de embarazos a término con evolución normal y sin complicaciones evidentes del recién nacido, se microdiseccionaron en 12 horas y se almacenaron en solución salina con fosfato sin congelación. Las arterias se procesaron luego para la descelularización usando los protocolos de SDS al 0,1 % y 1 %, y Triton X-100 al 1 %. Se realizó la evaluación de material celular y nuclear, fibras de colágeno, fibras elásticas y glucosoaminoglicanos de la matriz extracelular (MEC), así como el análisis morfométrico bajo técnicas histológicas e inmunohistoquímicas. Triton X-100 fue ineficaz, conservando los restos nucleares identificados por inmunofluorescencia, tuvo el daño más notable a las fibras elásticas y la disminución del colágeno. SDS efectivamente eliminó los núcleos y tuvo una disminución menor en las fibras elásticas y el colágeno. Laminina fue preservado en todos los grupos. No se identificaron diferencias significativas en los diámetros luminales; sin embargo, la capa media disminuyó debido a la descelularización de las células musculares. la descelularización con SDS al 0,1 % fue la más efectiva para eliminar células y preservar los principales componentes de la MEC.

Effect of cells in the epimysium conduit on the regeneration of peripheral nerve / 中国修复重建外科杂志

Objective: To investigate the effect of cells in the epimysium conduit (EMC) on the regeneration of sciatic nerve of mice. Methods: The epimysium of the 8-week-old male C57BL/6J enhanced green fluorescent protein (EGFP) mouse was trimmed to a size of 5 mm×3 mm, and prepared in a tubular shape (ie, EMC). Some epimysia were treated with different irradiation doses (0, 15, 20, 25, 30, 35 Gy) to inhibit cells migration. Then the number of migrating cells were counted, and the epimysia with the least migrating cells were selected to prepare EMC. Some epimysia were subjected to decellularization treatment and prepared EMC. HE and Masson staining were used to identify the decellularization effect. Twenty-four C57BL/6J wild-type mice were used to prepare a 3-mm-long sciatic nerve defect of right hind limb model and randomly divided into 3 groups ( n=8). EMC (group A), EMC after cell migration inhibition treatment (group B), and decellularized EMC (group C) were used to repair defects. At 16 weeks after operation, the midline of the regenerating nerve was taken for gross, toluidine blue staining, immunofluorescence staining, and transmission electron microscopy. Results: At 15 days, the number of migrating cells gradually decreased with the increase of irradiation dose. There was no significant difference between 30 Gy group and 35 Gy group ( P>0.05); there were significant differences between the other groups ( P0.05). Conclusion: The cellular components of the epimysium participate in and promote the regeneration of the sciatic nerve in mice.

3D tissue engineering, an emerging technique for pharmaceutical research

Tissue engineering and the tissue engineering model have shown promise in improving methods of drug delivery, drug action, and drug discovery in pharmaceutical research for the attenuation of the central nervous system inflammatory response. Such inflammation contributes to the lack of regenerative ability of neural cells, as well as the temporary and permanent loss of function associated with neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and traumatic brain injury. This review is focused specifically on the recent advances in the tissue engineering model made by altering scaffold biophysical and biochemical properties for use in the treatment of neurodegenerative diseases. A portion of this article will also be spent on the review of recent progress made in extracellular matrix decellularization as a new and innovative scaffold for disease treatment.

Properties of decellularized rabbit carotid artery with two different cross-linked technologies: A comparative study / 中南大学学报(医学版)

Objective:To compare the properties between decellularized rabbit carotid artery with different cross-linked technologies.Methods:The decellularized rabbit carotid arteries were randomly divided into a photo-oxidation group and a procyanidins group.One group was cross-linked with photo-oxidation and the other group was cross-linked with procyanidins.The in vitro or in vivo properties of the two groups were evaluated by testing heat-shrinking temperature,max tensile strength and the max elongation or by testing tissue structure,inflammatory reaction and calcification degree.Results:The heat-shrinking temperature,max tensile strength and the max elongation were similar in the two groups (P＞0.05).The tissue structure and inflammatory reaction were also similar in the two groups.Mthough the result of Von-Kossa calcium salt stain was slightly different,the calcium content was lower in the procyanidins group than that in the photo-oxidation group (P＜0.05).Conclusion:The grafts by two cross-linked technologies show excellent mechanical capability,lower immunogenicity,good biological stability and anti-calcification ability.The procyanidins group shows a better anti-calcification property than the photo-oxidation group.

Development of new technology in periodontal tissue engineering / 中华口腔医学杂志

The loss of periodontal support tissues might cause movement or finally loss of the teeth affected, impair furthermore the pronunciation and mastication functions, and even result in a series of physiological and psychological problems. Tissue engineering, as a technology to remodel missing tissues or organs and functional reconstruction, has achieved gratifying progress in regeneration of periodontal tissues. However, conventional construction methods have some deficiencies for functional periodontal reconstruction. In recent years, with the progress of tissue engineering technology, a series of new techniques and methods, such as cell sheet technology, decellular technology, electrospinning technology and three-dimensional printing, has been applied in tissue engineering bringing new hope for the regeneration of periodontal tissues. In this review article, the recent progress achieved in the field of periodontal tissue engineering and application of modern technology are summarized to make a brief exposition and to explore the future development of periodontal regeneration.

Decellularization of placentas: establishing a protocol

Biological biomaterials for tissue engineering purposes can be produced through tissue and/or organ decellularization. The remaining extracellular matrix (ECM) must be acellular and preserve its proteins and physical features. Placentas are organs of great interest because they are discarded after birth and present large amounts of ECM. Protocols for decellularization are tissue-specific and have not been established for canine placentas yet. This study aimed at analyzing a favorable method for decellularization of maternal and fetal portions of canine placentas. Canine placentas were subjected to ten preliminary tests to analyze the efficacy of parameters such as the type of detergents, freezing temperatures and perfusion. Two protocols were chosen for further analyses using histology, scanning electron microscopy, immunofluorescence and DNA quantification. Sodium dodecyl sulfate (SDS) was the most effective detergent for cell removal. Freezing placentas before decellularization required longer periods of incubation in different detergents. Both perfusion and immersion methods were capable of removing cells. Placentas decellularized using Protocol I (1% SDS, 5 mM EDTA, 50 mM TRIS, and 0.5% antibiotic) preserved the ECM structure better, but Protocol I was less efficient to remove cells and DNA content from the ECM than Protocol II (1% SDS, 5 mM EDTA, 0.05% trypsin, and 0.5% antibiotic).

Novel Approach Toward the Generation of Tissue Engineered Heart Valve by Using Combination of Antioxidant and Detergent: A Potential Therapy in Cardiovascular Tissue Engineering

To develop decellularized heart valve scaffold from porcine for heart valve regeneration. Porcine heart valves were decellularized with unique optimized approach by using 1% sodium dodecyl sulfate solution and 5% dimethyl sulfoxide for the first time. Effect of decellularization process on scaffold were characterized by hematoxylin-eosin, 4′,6-diamidino-2-phenylindole, Masson's trichrome, alcian blue staining and scanning electron microscopy for extracellular matrix (ECM) analysis in scaffold. The results showed that developed protocol for decellularization of heart valve scaffold shown complete removal of all cellular components, without changing the properties of ECM. The developed protocol was successfully used for heart valve ECM scaffolds development from porcine. The developed protocol seems to be promising solution for the heart valve tissue engineering application.

Study of tissue-engineered small vascular graft sheathed with nano poly (1,3-diamino-2-hydroxypropaneco-polyolsebacate) on adventitia and coated with heparin on intima / 上海交通大学学报（医学版）

Objective · To develop a new hybrid tissue-engineered vascular graft (HTEV) with excellent mechanical properties and biological functions. Methods · Decellularized rat aortas (DRAs) were prepared. Then, electrospinning nano poly (1,3-diamino-2-hydroxypropane-co-polyolsebacate) (ES-APS) was used to sheathe DRAs in order to improve the mechanical properties. After that, the intima of HTEV scaffold was modified with heparin coating. HTEVs were implanted in rat models in vivo to evaluate their biological functions. Six weeks later, vascular ultrasound and micro-CT angiography were carried out. Results · The donor aortic vessels were successfully decellularized. The total DNA content of DRA group [(115.4±10.9) ng/mg] significantly decreased compared with natural aorta group [(398.6±14.6) ng/mg] (P=0.000). But collagenous fibers and elastic fibers of decellularized vessels were severely injured. Mechanical tests of scaffolds showed that ES-APS significantly enhanced the mechanical properties. The wall thickness [(187±11) μm], suture retention strength [(0.51±0.06) N] and burst pressure [(2103±232) mmHg] of HTEV group all significantly increased compared with DRA group (P<0.01). Heparin coating modification of HTEV significantly reduced the number of adhesive platelets. Vascular ultrasound and micro-CT angiography showed all grafts were totally patent 6 weeks after implantation in rat models. ES-APS sheath successfully prevented the occurrence of vasodilation and aneurysm formation. Conclusion · DRA sheathed with ES-APS on adventitia and coated with heparin on intima is a new kind of HTEV, which possesses increased tensile strength and improved biocompatibility.

Updates of application of decellularized liver bioscaffold / 中华消化外科杂志

Replacement therapy is the most effective method for the treatment of end-stage liver disease,and decellularized liver bioscaffold broadens the research field of the replacement therapy.The present liver bioscaffold preparation is to perfuse chemical reagents (detergents,enzymes,et al) into the vascular structure of the liver under certain physical conditions,so as to remove cellular components and retain extracellular matrix and microvascular structure.Cells were reseeded into the decellularized liver scaffold to obtain the recellularized liver,which can be cultured and evaluated in vitro or in vivo by observing the adhesion of seeded cells,detecting the synthesis and secretion of the recellularized liver.Currently,the selection of seed cells,recellularization protocol and recellularized liver transplantation are still under exploration.In this review,the preparation,evaluation,detection and application of the decellularized liver bioscaffold are introduced for the further experimental study and clinical research.

Research progress on different methods of decellularization in tissues and organs / 国际生物医学工程杂志

Biologic scaffold materials derived from decellularized tissues and organs have been successfully used in tissue engineering and regeneration medicine.Decellularization methodology varies according to the differences of the origin,tissue density,composition,geometric and biologic properties of the tissue.Optimal decellularization method should be developed on the basis of complete cell content removal with the preservation of extracellular matrix (ECM) integrity and bioactivity.In this article,an overview of decellularization methods and their effect upon acellular bracket structure and composition are presented.

Determining the ideal decellularization protocol applied for preparing for liver biological scaffold / 重庆医学

Objective To develop an appropriate and simple decellularization protocol for preparing decellularized liver bio-logical scaffold(DLBS) .Methods Through immune-ofluorescence analysis ,scanning electron microscope ,quantification of Glyco-samin-oglycans(GAGs) and DNA ,we evaluated the effectiveness of low concentrations of SDS(0 .25% ,0 .50% ) and 1% Triton X-100 in the preservation of extracellular matrix of rat livers .Then ,we assessed Cytotoxicity of DLBS treated with three methods by MTT assay .Results Residual DNA after SDS(0 .25% ,0 .50% ) treatments were below 50 ng/mg dry weight ,which were less than Triton X-100 treatments .The content of GAGs in the 0 .25% SDS and 1% Triton X-100-treated scaffolds were higher than other detergent-treated scaffolds .The result of MTT assay showed that the liver scaffolds treated with three methods were not cytotoxic to proliferation of C3A cells .Conclusion The protocol containing 0 .25% SDS and a perfusion rate of 5 mL/min could be a rapid and effective decellularization protocol .DLBS constructed using this protocol could be an ideal material for preparing a transplant-able organ .