Comment on "Advancing Myocardial Infarction Treatment: Harnessing Multi-Layered Recellularized Cardiac Patches with Fetal Myocardial Scaffolds and Acellular Amniotic Membrane".

The effects of acellular amniotic membrane matrix on osteogenic differentiation and ERK1/2 signaling in human dental apical papilla cells

Decellularized Amniotic Membrane Scaffold as a Pericardial Substitute: An InVivo Study.

Wound healing is a complex process of repair that involves the interaction between different cell types and involves coordinated interactions between intracellular and extracellular signaling. Bone Marrow Mesenchymal Stem Cells (BMSCs) based and acellular amniotic membrane (AM) therapeutic strategies with the potential for treatment and regeneration of tissue. We aimed to evaluate the involvement of paracrine effects in tissue repair after the flap skin lesion rat model. In the full-thickness flap skin experiment of forty Wistar rats: A total of 40 male Wistar rats were randomized into four groups: group I: control (C; n = 10), with full-thickness lesions on the back, without (BMSCs) or AM (n = 10); group II: injected (BMSCs; n = 10); group III: covered by AM; group IV–injected (AM + BMSCs; n = 10). Cytokine levels, IL-1, and IL-10 assay kits, superoxide dismutase (SOD), glutathione reductase (GRs) and carbonyl activity levels were measured by ELISA 28th day, and TGF-β was evaluated by immunohistochemical, the expression collagen expression was evaluated by Picrosirius staining. Our results showed that the IL-1 interleukin was higher in the control group, and the IL-10 presented a higher mean when compared to the control group. The groups with BMSCs and AM showed the lowest expression levels of TGF-β. SOD, GRs, and carbonyl activity analysis showed a predominance in groups that received treatment from 80%. The collagen fiber type I was predominant in all groups; however, the AM + BMSCs group obtained a higher average when compared to the control group. Our findings suggest that the AM+ BMSCs promote skin wound healing, probably owing to their paracrine effect attributed to the promotion of new collagen for tissue repair.

The aim of this study was to determine the biocompatibility of an acellular human amniotic membrane biomaterial, which may have clinical utility for cell delivery. Human amniotic membrane was decellularized using 0.03% (w/v) sodium dodecyl sulfate (SDS), with hypotonic tris buffer and protease inhibitors and nuclease treatment. The membrane was terminally sterilized using an optimal concentration of peracetic acid. Residual SDS present within the acellular membrane was quantified using radio-labeled C14 SDS. In vivo biocompatibility was assessed by implantation of acellular human amniotic membrane subcutaneously into mice for 3 months and comparison with fresh and glutaraldehyde-fixed tissue. Cellular infiltrate into the explanted tissues was characterized using monoclonal antibodies against the following cell surface markers: CD3, CD4, CD34, and F4/80. Calcification was determined using the Von Kossa's stain. The potential of acellular human amniotic membrane to support the attachment and proliferation, and maintain viability of primary human dermal fibroblasts and primary human dermal keratinocytes was assessed in vitro, using a static culture system. Peracetic acid at a concentration of 0.1% (v/v) was sufficient for the sterilization of acellular amniotic membrane. Levels of SDS present within the acellular tissue were 0.62 +/- 0.13 microg/mg. Analysis of explanted samples from the mice indicated that acellular amniotic membrane contained low numbers of T-cells and high numbers of fibroblastic cells, macrophages, and endothelial cells, indicative of a wound-healing response. There was no evidence of calcification present within explanted acellular amniotic membrane compared to explanted glutaraldehyde-fixed amniotic membrane. Acellular amniotic membrane was shown to be capable of supporting the attachment and proliferation of primary human fibroblasts and keratinocytes. The viability of the cells was maintained for up to 4 weeks. Cell-seeded acellular amniotic membrane has the potential for delivering autologous or allogeneic cells to treat a variety of conditions, including diabetic foot ulcers, corneal defects, and severe skin burns.

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BackgroundThe necrotic skin flap represents a great challenge in plastic and reconstructive surgery. In this study, we evaluated the effect of bioscaffolds, acellular amniotic membranes (AAMs), and bone marrow-derived mesenchymal stem cells (BM-MSCs) on random skin flap (RSF) survival in rats by applying a cell-free extracellular matrix scaffold as a supportive component for the growth and proliferation of BM-MSCs on RSFs. AAM matrix scaffolds were created by incubating AMs in ethylenediaminetetraacetic acid 0.05% at 37°C, and cell scrapers were used.ObjectivesThe aim of the present study was to assess the effect of AAM as a scaffold in TE, and combined with transplanted BM-MSCs, on the survival of RSFs and on the biomechanical parameters of the incision-wound flap margins 7 days after flap elevation.Materials and MethodsBM-MSCs and AAMs were transplanted into subcutaneous tissue in the flap area. On the 7th postoperative day, the surviving flap areas were measured using digital imaging software, and the flap tissue was collected for evaluation. Forty rats were randomly divided into four groups of 10 each: group 1 received an AAM injection; group 2 underwent BM-MSC transplantation; group 3 received both AAM injection + BM-MSC transplantation; and group 4 was the control group, receiving only saline.ResultsThe survival area in the AAM/BM-MSC group was significantly higher than in the control group (18.49 ± 1.58 versus 7.51 ± 2.42, P < 0.05). The biomechanical assessment showed no significant differences between the experimental groups and the control group (P > 0.05), and there was no correlation with flap survival.ConclusionsOur findings showed that the treatment of flaps with BM-MSC and AAM transplantations significantly promoted flap survival compared to a control group. The viability of the flap was improved by combining BM-MSCs with AAM matrix scaffolds.

Interposition of acellular amniotic membrane at the tendon to bone interface would be better for healing than overlaying above the tendon to bone junction in the repair of rotator cuff injury

A decellularized amniotic membrane matrix is an ideal scaffolding system. This study compared the effectiveness of amniotic membrane and amniotic membrane pretreated with monophosphoryl lipid A (MPLA) for repairing the fascia of the abdomen with tension sutures. A total of 24 rats were randomly divided into three groups of eight each. In the control group, the fascia was repaired only with non-absorbable monofilament yarn under tension. In the second and third groups, in addition to the control treatment, acellular amniotic membrane was used and MPLA pretreated amniotic membrane was used, respectively. At 3, 7 and 14 days after treatment, the fascia repair was examined macroscopically and microscopically in all groups. Macroscopic examination showed that the use of MPLA pretreated significantly different from the other groups only in the fibrin exudate. Changes in the fascia rupture pressure showed significant differences between groups. Group three, which was sutured with MPLA pretreated acellular membrane, showed greater amounts of collagen, monocytes and neovascularization, especially at days 7 and 14. The results show that MPLA pretreated acellular amniotic membrane helped to repair abdominal fascia to some extent.

Tissue engineered skin constructed by cultured seed cells provides a new way to accelerate skin wound healing and improve the quality of tissue repair. Conventional models with dermal scaffold material based on collagen hydrogels, however, have poor stability and unsatisfactory longevity. Here, we describe an improved acellular amniotic membrane as a scafford with epidermal stem cells for tissue engineering skins and observe their clinical effect on deep burn wounds. The human epidermal stem cells was isolated from the skin samples by trypsin digesting method and purified by collagen adhering method, and then seeded on the acellular amniotic membrane to form the engineered skin. The biological attachment and growth of cultured epidermal stem cells were observed. Twelve cases of deep burn wounds were applied with the engineered skin. The results showed that the epidermal stem cells adhered to the surface of acellular amniotic membrane quickly after being seeded and exhibited a high colony formation efficiency. The wounds treated with engineered skin healed rapidly with good clinical take. The average time of wound closure in engineered skin group was shortened than that of control group. There were no obvious evidence of immunological rejection and inflammatory reaction during the observation. This indicated that the acellular amniotic membrane with good histocompatibility may be a suitable scaffold with epidermal stem cells in the construction of tissue engineering skin to repair deep burn wounds.

Tissue engineered skins have the potential to overcome the limitations of present large area of skin replacements. To achieve this, we investigated the effect and fate of tissue engineered skins comprising human epidermal stem cells and acellular amniotic membrane after their transplantation to full-thickness skin defect wound, so as to lay a foundation for clinic application. The human epidermal stem cells was isolated from the skin samples by trypsin digesting method and purified by collagen adhering method, and then seeded on the acellular amniotic membrane and cultured in the keratinocyte serum free medium in present of the epidermal growth factor to form the tissue engineered skin. Eighteen New Zealand white rabbit with full thickness skin defect on the back were grafted with the tissue engineered skin (experimental grafted group), single acellular amniotic membrane(control group 1) and no treated (control group 2),respectively. The wound healing conditions were observed and wound contraction rate were calculated. Histological changes were undertaken by tissue sampling from the grafted wound. The expression of CK19 and HLA-DR were detected by the immunohistochemistry methods. The results showed that the grafted wounds in experimental group healed very well with good external appearance and elasticity. The wound contraction rate in experimental grafted group, control group 1 and control group 2 was 22.6 plusmn 1.54%, 29.3 plusmn 1.41 % and 30.4 plusmn 1.58 %, respectively. There were significantly different between experimental grafted groups and control group(P<0.05). No obvious immune rejection and any bleeding, suppuration or infection under all the grafted skins. There were full differentiation of epithelium and orderly collagen proliferation in experimental grafted wounds by histological examination. The expression of CK19 and HLA-DR were positive in the experimental grafted wounds, which were negative in the control group. This study suggest that the skin grafting with tissue engineered skins comprising human epidermal stem cells and acellular amniotic membrane could improve wound healing quality prominently, which maybe an ideal skin substitute with good histocompatibility.

Acellular amniotic membrane (AM) has been studied, with promising results on the reconstruction of lesioned tissues, and has become an attractive approach for tracheal repair. This study aimed to evaluate the repair of the trachea with human umbilical cord mesenchymal stem cells (hucMSCs) differentiated in chondrocytes, grown on an experimental model. Tracheal defects were induced by surgical tracheostomy in 30 New Zealand rabbits, and the acellular amniotic membrane, with or without cells, was covering the defect. The hucMSCs were isolated and cultivated with chondrogenic differentiation over the culture of 14 days, and then grown on the AM. In this study, the AM was biocompatible and hucMSCs differentiated into chondrocytes. Our results demonstrated an important role for AM with cultured cells in the promotion of immature collagen, known to produce tissue regeneration. In addition, cartilaginous tissue was found at the tracheal defects, demonstrated by immunohistology results. This study suggests that this biomaterial implantation can be an effective future therapeutic alternative for patients with tracheal injury.

Early repair of skin injury and maximal restoration of the function and appearance have become important targets of clinical treatment. In the present study, we observed the healing process of skin defects in nude mice and structural characteristics of the new skin after transplantation of isolated and cultured adipose derived mesenchymal stem cells (ADMSCs) onto the human acellular amniotic membrane (AAM). The result showed that ADMSCs were closely attached to the surface of AAM and grew well 24 h after seeding. Comparison of the wound healing rate at days 7, 14, and 28 after transplantation showed that ADMSCs seeded on AAM facilitated the healing of full-thickness skin wounds more effectively as compared with either hAM or AAM alone, indicating that ADMSCs participated in skin regeneration. More importantly, we noticed a phenomenon of hair follicle development during the process of skin repair. Composite ADMSCs and AAM not only promoted the healing of the mouse full-thickness defects but also facilitated generation of the appendages of the affected skin, thus promoting restoration of the skin function. Our results provide a new possible therapy idea for the treatment of skin wounds with respect to both anatomical regeneration and functional restoration.

In this study, we used human amniotic membrane (AM) to prepare a dermal scaffold with intact basement membrane (BM) and good biostability for quick expansion and transplantation of epidermal keratinocytes (EKs). Fresh AM was treated by repeated freeze-thaw cycles and DNase digestion. This new method was able to cleanse the cell components effectively and retain the BM structure with continuous distributions of laminin, collagen IV, VI, and VII. Subsequently, the acellular amniotic membrane (AAM) was cross-linked with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) for 5 min, 30 min, and 6 h. With the time of cross-linking prolonging, the mechanical strength and biostability of AAM increased gradually, while its cytotoxicity to EKs also increased. The 5-min cross-linked AAM (5min-AAM) had no significant cytotoxicity with good histocompatibility. The relative cell viability of EKs seeded on the 5min-AAM surface was 367 ± 33% and 631 ± 43% at 7 and 14 days of culture, respectively, both higher than 294 ± 30% and 503 ± 41% of the conventional cell culture dish (CCD) group, and the proportion of P63-positive cells was significantly higher than that of the CCD group on day 7 (54.32 ± 4.27% vs. 33.32 ± 3.18%, p < 0.05). When the 5min-AAM loaded with EKs (EK-AAM) was grafted onto full-thickness skin defects in nude mice, the cells survived well and formed an epidermis similar to normal skin. The new epidermis was thicker, and reconstruction of the dermal structure was good with an intact BM. Four weeks after transplantation, the wound contraction rate in the EK-AAM group was 43.09 ± 7.05%, significantly lower than that in the EK sheet group (57.49 ± 5.93%) and control group (69.94 ± 9.47%) (p < 0.05). In conclusion, repeated freeze-thaw treatment with appropriate EDC cross-linking offers AAM an intact BM structure with good operability and biostability. It may prove to be an ideal dermal scaffold to promote expansion of EKs in vitro and be transplanted for reconstruction of the dermal structure.

Background:Skin flap procedures are employed in plastic surgery, but failure can lead to necrosis of the flap. Studies have used bone marrow mesenchymal stem cells (BM-MSCs) to improve flap viability. BM-MSCs and acellular amniotic membrane (AAM) have been introduced as alternatives. The objective of this study was to evaluate the effect of BM-MSCs and AAM on mast cells of random skin flaps (RSF) in rats.Methods:RSFs (80 × 30 mm) were created on 40 rats that were randomly assigned to one of four groups, including (I) AAM, (II) BM-MSCs, (III) BM-MSCs/AAM, and (IV) saline (control). Transplantation was carried out during the procedure (zero day). Flap necrosis was observed on day 7, and skin samples were collected from the transition line of the flap to evaluate the total number and types of mast cells. The development and the total number of mast cells were related to the development of capillaries.Results:The results of one-way ANOVA indicated that there was no statistically significant difference between the mean numbers of mast cell types for different study groups. However, the difference between the total number of mast cells in the study groups was statistically significant (p = 0.001).Conclusion:The present study suggests that the use of AAM/BM-MSCs can improve the total number of mast cells and accelerate the growth of capillaries at the transient site in RSFs in rats.

Objective:To compare the healing effects of dried and acellular human amniotic membrane and Mepitel for coverage of split-thickness graft donor site (STGDS). Methods:Twenty patients who underwent STGDS regeneration surgery in identical anatomic regions were enrolled in this randomized controlled clinical trial conducted in Hazrate Fatemeh hospital (Iran). Patients were randomly assigned in 3 groups of wound dressing; group A by Mepitel, group B AmiCare (Dried amniotic membrane) and group C OcuReg-A (Acellular amniotic membrane). Re-epithelization rate (healing time), pain sensation, scar formation and infection rate were assessed till complete healing was achieved. Results: Our results showed no significant difference between Amicare, OcuReg-A and Mepitel in the features analyzed by us including: Re-epithelization rate (healing time) P value; 0.573, Pain sensation P value: day 4 th: 0.131, day8 th: 0.93 and day 12 th: 0.365, Scar formation P value>0.05and Infection rate. Conclusion: Our findings confirmed the safety and efficacy of AmiCare (dried amniotic membrane) and OcuReg-A (Acellular amniotic membrane) in treatment of split-thickness donor site in comparison with Mepitel as a standard wound dressing. Trial registration number: IRCT201511118177N12