Potential of human amniotic membrane application for articular cartilage regeneration: a review.

Objectives: Repairing of articular cartilage damage is challenging. Clinically, tissue engineering technology is used to induce stem cell differentiation and proliferation on biological scaffolds to repair defective joints. However, no ideal biological scaffold has been identified. This study investigated the effects of amniotic membrane/collagen scaffold on the differentiation of adipose-derived stem cells (ADSCs) and articular cartilage repair. Methods: Adipose tissue of New Zealand rabbits was excised, and ADSCs were isolated and induced for differentiation . An articular cartilage defect model was constructed to identify the effect of amniotic membrane/collagen scaffolding on cartilage repair. Cartilage formation was analyzed by imaging and toluene blue staining. Knee joint recovery in rabbits was examined using hematoxylin and eosin, toluidine, safranine, and immunohistochemistry at 12 weeks post-operation. Gene expression was examined using Elisa, RT-PCR, western blotting, and immunofluorescence. Results: The adipose tissue was effectively differentiated into ADSCs, which further differentiated into chondrogenic, osteogenic, and lipogenic lineages after three weeks' culture in vitro. Compared with platelet-rich plasmon (PRP) scaffold, the amniotic membrane scaffold better promoted the growth and differentiation of ADSCs. Additionally, scaffolds containing the PRP and amniotic membrane efficiently enhanced the osteogenic differentiation of ADSCs. The levels of COL1A1, COL2A1, COL10A1, SOX9 and ACAN in ADSCs + amniotic membrane + PRP group were significantly higher than the other groups both in vitro and in vivo. The Wakitani scores of the ADSC + amniotic membrane + PRP group were lower than that in ADSC + PRP (4.4 ± 0.44**), ADSC + amniotic membrane (2.63±0.38**), and control groups (6.733±0.21) at week 12 post-operation. Osteogenesis in rabbits of the ADSC + amniotic membrane + PRP group was significantly upregulated when compared with other groups. Amniotic membranes significantly promoted the expression of cartilage regeneration related factors (SOX6, SOX9, RUNX2, NKX3-2, MEF2C, and GATA4). The ADSC + PRP + amniotic membrane group exhibited the highest levels of TGF-β, PDGF and FGF while exhibited the loweset level of IL-1β, IL6, and TNF-αin articular cavity. Conclusion: Amniotic membrane/collagen combination-based scaffolds promoted the proliferation and cartilage differentiation of ADSCs, and may provide a new treatment paradigm for patients with cartilage injury.

The application of human amniotic membrane in the surgical management of limbal stem cell deficiency

Objectives: Repairing articular cartilage damage is challenging. Clinically, tissue engineering technology is used to induce stem cell differentiation and proliferation on biological scaffolds to repair defective joints. However, no ideal biological scaffolds have been identified. This study investigated the effects of amniotic membrane/collagen scaffolds on the differentiation of adipose-derived stem cells (ADSCs) and articular cartilage repair. Methods: Adipose tissue of New Zealand rabbits was excised, and ADSCs were isolated and induced for differentiation. An articular cartilage defect model was constructed to identify the effect of amniotic membrane/collagen scaffolds on cartilage repair. Cartilage formation was analyzed by imaging and toluene blue staining. Knee joint recovery in rabbits was examined using hematoxylin and eosin, toluidine, safranine, and immunohistochemistry at 12 weeks post-operation. Gene expression was examined using ELISA, RT-PCR, Western blotting, and immunofluorescence. Results: The adipose tissue was effectively differentiated into ADSCs, which further differentiated into chondrogenic, osteogenic, and lipogenic lineages after 3 weeks’ culture in vitro. Compared with platelet-rich plasmon (PRP) scaffolds, the amniotic membrane scaffolds better promoted the growth and differentiation of ADSCs. Additionally, scaffolds containing the PRP and amniotic membrane efficiently enhanced the osteogenic differentiation of ADSCs. The levels of COL1A1, COL2A1, COL10A1, SOX9, and ACAN in ADSCs + amniotic membrane + PRP group were significantly higher than the other groups both in vitro and in vivo. The Wakitani scores of the ADSC + amniotic membrane + PRP group were lower than that in ADSC + PRP (4.4 ± 0.44**), ADSC + amniotic membrane (2.63 ± 0.38**), and control groups (6.733 ± 0.21) at week 12 post-operation. Osteogenesis in rabbits of the ADSC + amniotic membrane + PRP group was significantly upregulated when compared with other groups. Amniotic membranes significantly promoted the expression of cartilage regeneration-related factors (SOX6, SOX9, RUNX2, NKX3-2, MEF2C, and GATA4). The ADSC + PRP + amniotic membrane group exhibited the highest levels of TGF-β, PDGF, and FGF while exhibiting the lowest level of IL-1β, IL6, and TNF-α in articular cavity. Conclusion: Amniotic membrane/collagen combination-based scaffolds promoted the proliferation and cartilage differentiation of ADSCs, and may provide a new treatment paradigm for patients with cartilage injury.

Amniotic membrane (AM) has great potential as a scaffold for tissue regeneration in reconstructive surgery. To date, no systematic review of the literature has been performed for the applications of AM in wound closure of internal organs. Therefore, in this systematic review and meta‐analysis, we summarize the literature on the safety and efficacy of AM for the closure of internal organs. A systematic search was performed in MEDLINE‐PubMed database and OVID Embase to retrieve human and controlled animal studies on wound closure of internal organs. The Cochrane Risk of Bias tool for randomized clinical trials and the SYRCLE risk of bias tool for animal studies were used. Meta‐analyses (MAs) were conducted for controlled animal studies to assess efficacy of closure, mortality and complications in subjects who underwent surgical wound closure in internal organs with the application of AM. Sixty references containing 26 human experiments and 36 animal experiments were included. The MAs of the controlled animal studies showed comparable results with regard to closure, mortality and complications, and suggested improved mechanical strength and lower inflammation scores after AM application when compared to standard surgical closure techniques. This systematic review and MAs demonstrate that the application of AM to promote wound healing of internal organs appears to be safe, efficacious, and feasible.

Human amniotic membrane has been widely used as substrate for ex vivo expansion and transplantation of limbal epithelial cells. To further clarify its suitability as a surrogate niche for limbal stem cells and progenitor cells, we analyzed the composition of the amniotic epithelial basement membrane, with special focus on the expression of limbus-specific matrix components. Cryosections of corneoscleral specimens obtained from 10 human donor eyes and of 6 amniotic membrane specimens obtained at cesarean section were stained by indirect immunofluorescence using a broad panel of antibodies against basement membrane components. Both amniotic and limbal epithelial basement membranes showed positive immunoreactivity for collagen type IV α1, α2, α5, and α6 chains; collagens type VII, XV, XVI, XVII, and XVIII; laminin α3, β1, β2, β3, γ1, and γ2 chains; laminin-111 and laminin-332; nidogen-1 and nidogen-2; fibronectin; fibulin-2; fibrillin-2; perlecan; and agrin. Both types of basement membrane were negative for collagen type IV α3 and α4 chains, collagen type V, and laminin α4 chain. Limbal basement membrane components, which were not detected in amniotic membrane, included laminin α1, α2, α5, and γ3 chains; BM40/SPARC; tenascin-C; matrilin-2; endostatin; and collagen type XVIII. Despite extensive similarities in basement membrane composition between amniotic and corneolimbal epithelia, the lack of limbus-specific environmental factors argues against the potential of denuded amniotic membrane as a surrogate niche for limbal stem cells but supports its suitability as a substrate to promote the formation of a well-differentiated stratified corneal epithelial equivalent for tissue engineering strategies.

The special structure and biological viability of amniotic membrane (AM) allow it to be an ideal biological material. Biological improvement of the AM opens a new research field and allows it to be used broadly in clinical practice. The histological structure, physiological property, immunological behavior, the history and current status of the application of AM, the mechanism and the classification of various ophthalmic applications of AM and the mechanism of AM transplantation, as well as the biological improvement of AM were summarized and discussed.

Ex vivo culturing of limbal stem cells on human amniotic membranes can be accelerated if all amniotic epithelial cells have been removed beforehand. A common application of acellular amniotic membranes is their use in cultivating autologous oral mucosal epithelial cells for transplantation in cases of bilateral stem cell insufficiency. Amniotic epithelial cells can be eliminated with enzymatic-chemical or mechanical methods or with a combination of both. The efficacy of a waterjet cutter to eliminate amniotic epithelial cells from the amniotic membrane was investigated. Deep frozen placentas from healthy mothers were defrosted and a well-defined surface of the amniotic membrane (d = 15mm) was treated with the waterjet in a standardized way. The waterjet used two different nozzles (pin-point and narrow stream nozzles). The applied system pressures with the pin-point stream nozzle (aperture 120μm) were 30, 40 and 50bar and the narrow stream nozzle was operated with pressures of 70, 80 and 90bar on the amniotic membrane. A total number of 42 tissue samples were examined with an optical microscope using native trypan blue staining. For each type of nozzle and each application pressure two amnion samples were examined with a scanning electron microscope to analyze the efficacy of the mechanical epithelial cell elimination from the amniotic membrane. After medical imaging and histopathological examination the efficacy was graded using the following scale: 0 = no amniotic epithelial cells, 1 = no cells, low amounts of cell debris, 2 = single amniotic epithelial cells, large amounts of cell debris, 3 = loose cell layer, 4 = continuous sheet of epithelial cells. To eliminate epithelial cells from the surface of the amniotic membrane with the waterjet pinpoint stream nozzle (aperture: 120μm) an application pressure of 30-50bar was needed. The use of the narrow stream nozzle required a pressure of 70-90bar. The preparation of amniotic membranes with the waterjet represents a precise option to mechanically eliminate amniotic epithelial cells from the amniotic membrane. The use of a waterjet cutter as an exclusively mechanical method without enzymatic-chemical substances may be a benefit, as cytotoxic effects on culturing limbal stem cells caused by chemical substances are not present.

<p>Introduction. The amniotic membrane is used in transplant surgery, ophthalmology and dermatology. Various methods have been developed to preserve amniotic membrane: hypothermic storage, cryopreservation, lyophilization. Transplantation of fresh amniotic membrane showed low inflammatory response. The efficient antibiotic solutions are carefully chosen for the hypothermic storage of amniotic membranes. The aim of this study was to compare the efficacy of two antibiotic solutions for the hypothermic amniotic membrane preservation and the structure of the amniotic membrane after the preservation process. Methods. Fifteen amniotic membranes were prepared and hypothermically stored in penicillin solution in BSS (2000 IU/ml) while the remaining fifteen in an antibiotic solution containing: benzylpenicillin (50 µg/ml), gentamicin (100 µg/ml), ciprofloxacin (200 µg/ml) and fluconazole (100 µg/ml). All amniotic membranes were microbiologically tested after preparation and after hypothermic storage for two weeks. Histological analysis of thirty amniotic membranes was performed after the process of preservation. Results. Fifteen amniotic membranes were sterile after hypothermic preservation in the penicillin solution. Also, fifteen amniotic membranes were sterile after hypothermic preservation in the solution of antibacterial agents (penicillin, gentamicin, ciprofloxacin) and antifungal agent fluconazole. The amniotic membrane had a normal structure and thickness of 35.33±11.03 mm. Conclusion. Both antibiotic solutions, one that contains only penicillin and one that contains multiple antibacterial agents and fluconazole, provide sterility of fresh hypothermically stored amniotic membranes for two weeks. In the preparation of fresh hypothermically stored amniotic membrane, a solution with multiple antibiotics is preferred. The normal tissue structure of the amniotic membrane was histologically confirmed after the process of preservation.</p>

To report the current clinical applications and trends of scleral and amniotic membrane use in ophthalmology. Review of annual reports from the Catalan Transplant Organization (OCATT), on scleral patch and amniotic membrane eye indications in Catalonia region (Spain) over a 6-year period from 2013 to 2018. A total of 874 scleral and 1665 amniotic membranes patches were implanted, from January 2013 to December 2018. The most frequent indication over the 6-year period for scleral patch was glaucoma surgery (77.5%), eyelid reconstruction (5.2%) and corneal or scleral ulcer (5%). Regarding amniotic membrane, corneal ulcer (26.9%), conjunctival reconstruction (23.8%) and corneal epithelial defect (22.7%) were the most common indications. During the study period, an increasing trend was found on sclera patches for eyelid reconstruction (p = 0.0032) and amniotic membrane for inflammation management (p = 0.0198). Glaucoma surgery and corneal ulcers have represented the top indications for scleral patch and amniotic membrane use, over the period, respectively. A significant trend has also been found towards eyelid reconstruction using scleral patches and amniotic membrane for anterior segment inflammation management. This evolving scenario in tissue use for ocular surgery has to be taken into consideration, especially regarding eye banks facing current and futures changes in tissue preservation, storage and indications.

Carbodiimide cross-linked amniotic membranes for cultivation of limbal epithelial cells

Periodontal disease is characterized by the destruction of tooth supporting structures, and continuous destruction of these structures may lead to alveolar bone defects and tooth loss. Periodontal therapy aims to arrest the disease progression as well as to regenerate the loss of structures. Since, the regeneration of these structures is a complex process, cell-based tissue engineering has become one of the methods for periodontal tissue regeneration. In order to give mechanical support to the cells, an amniotic membrane has been proposed as one type of periodontal scaffold, due to its predictable properties. In this review, the integral structure, properties and the recent research in application of amniotic membranes, basically in medical and dental surgeries, along with its potential as a scaffold in periodontal regeneration are highlighted. Amniotic membranes have shown great potential as a suitable substrate/scaffold in in vitro and animal studies; thus provide an alternative for scaffolds materials nowadays. Nevertheless, further studies are required to establish its role and efficacy in periodontal tissue engineering.

The amniotic membranes were collected from the placentae of selected and screened donors. Processing was done by washing the fresh amniotic membrane successively in sterile saline, 0.05% sodium hypochlorite solution and sterile distilled water until it was completely cleared of blood particles. The membranes were sterilized by gamma irradiation at 25 kGy. The processed amniotic membranes were applied to 50 open wounds comprising of 42 full thickness defects and eight partial thickness defects. These included leprotic, diabetic, traumatic, gravitational ulcers and superficial burn in the form of scald and corrosive burn. The radiation processed amniotic membranes favoured healing of unresponsive and non-healing ulcers of different etiologies. Ulcers with duration of minimum 3 weeks to maximum 12 months were found to heal in 2-6 weeks by the application of amniotic membranes.

Menstrual blood is a unique body fluid that contains mesenchymal stem cells (MSCs). These cells have attracted a great deal of attention due to their exceptional advantages including easy access and frequently accessible sample source and no need for complex ethical and surgical interventions, as compared to other tissues. Menstrual blood-derived MSCs possess all the major stem cell properties and even have a greater proliferation and differentiation potential as compared to bone marrow-derived MSCs, making them a perspective tool in a further clinical practice. Although the potential of menstrual blood stem cells to differentiate into a large variety of tissue cells has been studied in many studies, their chondrogenic properties have not been extensively explored and investigated. Articular cartilage is susceptible to traumas and degenerative diseases, such as osteoarthritis, and has poor self-regeneration capacity and therefore requires more effective therapeutic technique. MSCs seem promising candidates for cartilage regeneration; however, no clinically effective stem cell-based repair method has yet emerged. This chapter focuses on studies in the field of menstrual blood-derived MSCs and their chondrogenic differentiation potential and suitability for application in cartilage regeneration. Although a very limited number of studies have been made in this field thus far, these cells might emerge as an efficient and easily accessible source of multipotent cells for cartilage engineering and cell-based chondroprotective therapy.

ABSTRACTIntroduction: In reconstructive urology, autologous tissues such as intestinal segments, skin, and oral mucosa are used. Due to their limitations, reconstructive urologists are waiting for a novel material, which would be suitable for urinary tract wall replacement. Human amniotic membrane (AM) is a naturally derived biomaterial with a capacity to support reepithelization and inhibit scar formation. AM has a potential to become a considerable asset for reconstructive urology, i.e., reconstruction of ureters, urinary bladder, and urethrae.Areas covered: This review aims to discuss the potential application of human AM in reconstructive urology. The environment for urinary tract healing is particularly unfavorable due to the presence of urine. Due to its fetal origin, the bioactivity of AM is orientated to induce intrinsic regeneration mechanisms and inhibit scarring. This review introduces the concept of applying human AM in reconstructive urology procedures to improve their outcomes and future tissue engineering based strategies.Expert opinion: Many fields of medicine that have accomplished translational research have proven the usefulness of AM in clinical practice. There is an urgent need for studies to be conducted on large animal models that might convincingly demonstrate the underestimated potential of AM to urologists around the world.

Large articular cartilage defects remain an immense challenge in the field of regenerative medicine because of their poor intrinsic repair capacity. Currently, the available medical interventions can relieve clinical symptoms to some extent, but fail to repair the cartilaginous injuries with authentic hyaline cartilage. There has been a surge of interest in developing cell-based therapies, focused particularly on the use of mesenchymal stem/progenitor cells with or without scaffolds. Mesenchymal stem/progenitor cells are promising graft cells for tissue regeneration, but the most suitable source of cells for cartilage repair remains controversial. The tissue origin of mesenchymal stem/progenitor cells notably influences the biological properties and therapeutic potential. It is well known that mesenchymal stem/progenitor cells derived from synovial joint tissues exhibit superior chondrogenic ability compared with those derived from non-joint tissues; thus, these cell populations are considered ideal sources for cartilage regeneration. In addition to the progress in research and promising preclinical results, many important research questions must be answered before widespread success in cartilage regeneration is achieved. This review outlines the biology of stem/progenitor cells derived from the articular cartilage, the synovial membrane, and the synovial fluid, including their tissue distribution, function and biological characteristics. Furthermore, preclinical and clinical trials focusing on their applications for cartilage regeneration are summarized, and future research perspectives are discussed.