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Denudation of cryopreserved amniotic membrane. AM with intact epithelium was illustrated by antipancytokeratin staining (green) (A) before thermolysin treatment and (B) after treatment. Cell nuclei were counterstained with DAPI (blue). SEM images of denuded AM showing the (C) denuded basement membrane and (D) a high-magnification image of the individual fibers on the stromal layer.
Source publication
To evaluate a silk fibroin (SF) biomaterial as a substrate for corneal epithelial cell proliferation, differentiation, and stratification in vitro compared with denuded human amniotic membrane (AM).
Primary human and rabbit corneal epithelial cells and immortalized human corneal limbal epithelial cells were cultured on the SF and denuded AM, respec...
Contexts in source publication
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... thermolysin treatment, uniform staining of pankeratin recognizing the cytoskeleton of cells was observed within the entire amniotic epithelial layer (Fig. 1A). In contrast, after treatment, only residual cellular debris on the basement membrane stained by pankeratin can be observed (Fig. 1B). SEM results confirmed the total removal of the epithelial cells with intact basement membranes (Figs. 1C, 1D), which is consistent with previous findings. 22 Denuded AM showed a semitransparent ...
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... thermolysin treatment, uniform staining of pankeratin recognizing the cytoskeleton of cells was observed within the entire amniotic epithelial layer (Fig. 1A). In contrast, after treatment, only residual cellular debris on the basement membrane stained by pankeratin can be observed (Fig. 1B). SEM results confirmed the total removal of the epithelial cells with intact basement membranes (Figs. 1C, 1D), which is consistent with previous findings. 22 Denuded AM showed a semitransparent appearance and the tendency of sticking together when lifted by two fine forceps (Figs. 2A, 2B). An Ultracell surgical sponge (Aspen Surgical, ...
Context 3
... of pankeratin recognizing the cytoskeleton of cells was observed within the entire amniotic epithelial layer (Fig. 1A). In contrast, after treatment, only residual cellular debris on the basement membrane stained by pankeratin can be observed (Fig. 1B). SEM results confirmed the total removal of the epithelial cells with intact basement membranes (Figs. 1C, 1D), which is consistent with previous findings. 22 Denuded AM showed a semitransparent appearance and the tendency of sticking together when lifted by two fine forceps (Figs. 2A, 2B). An Ultracell surgical sponge (Aspen Surgical, Caledonia, MI) was used to distinguish the orientation of the denuded AM, as the adherent side indicated the ...
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... thermolysin treatment, uniform staining of pankeratin recognizing the cytoskeleton of cells was observed within the entire amniotic epithelial layer (Fig. 1A). In contrast, after treatment, only residual cellular debris on the basement membrane stained by pankeratin can be observed (Fig. 1B). SEM results confirmed the total removal of the epithelial cells with intact basement membranes (Figs. 1C, 1D), which is consistent with previous findings. 22 Denuded AM showed a semitransparent ...
Context 5
... thermolysin treatment, uniform staining of pankeratin recognizing the cytoskeleton of cells was observed within the entire amniotic epithelial layer (Fig. 1A). In contrast, after treatment, only residual cellular debris on the basement membrane stained by pankeratin can be observed (Fig. 1B). SEM results confirmed the total removal of the epithelial cells with intact basement membranes (Figs. 1C, 1D), which is consistent with previous findings. 22 Denuded AM showed a semitransparent appearance and the tendency of sticking together when lifted by two fine forceps (Figs. 2A, 2B). An Ultracell surgical sponge (Aspen Surgical, ...
Context 6
... of pankeratin recognizing the cytoskeleton of cells was observed within the entire amniotic epithelial layer (Fig. 1A). In contrast, after treatment, only residual cellular debris on the basement membrane stained by pankeratin can be observed (Fig. 1B). SEM results confirmed the total removal of the epithelial cells with intact basement membranes (Figs. 1C, 1D), which is consistent with previous findings. 22 Denuded AM showed a semitransparent appearance and the tendency of sticking together when lifted by two fine forceps (Figs. 2A, 2B). An Ultracell surgical sponge (Aspen Surgical, Caledonia, MI) was used to distinguish the orientation of the denuded AM, as the adherent side indicated the ...
Citations
... Silk fibroin is a insoluble protein obtained from the mulberry silkworm Bombyx mori (Bm) [51][52][53][54][55]. Transparent thin silk films supports the generation of a corneal epithelial cell sheet in vitro [56]. For damaged corneal surface the silk fibroin film obtained from nonmulberry Antheraea mylitta (Am) offers to be a promising which supports cell growth and differentiation [57][58][59]. Rigidity associated with other cellular events has been offered by the scaffold. Ocular abnormalities does not shown any markable changes. ...
... Before CALEC, techniques for culturing autologous and allogeneic limbal epithelial cells for treatment of LSCD used a variety of manufacturing procedures and substrates on which the cells are grown, ranging from petrolatum gauze (12), contact lenses (12,(21)(22)(23), fibrin (13,15,(24)(25)(26), keratin (27), silk fibroin (28), and collagen (29). However, despite landmark publications (13,30,31) describing this approach decades ago, no previous clinical studies have evaluated cultivated LSC grafting in the United States. ...
To treat unilateral limbal stem cell (LSC) deficiency, we developed cultivated autologous limbal epithelial cells (CALEC) using an innovative xenobiotic-free, serum-free, antibiotic-free, two-step manufacturing process for LSC isolation and expansion onto human amniotic membrane with rigorous quality control in a good manufacturing practices facility. Limbal biopsies were used to generate CALEC constructs, and final grafts were evaluated by noninvasive scanning microscopy and tested for viability and sterility. Cultivated cells maintained epithelial cell phenotype with colony-forming and proliferative capacities. Analysis of LSC biomarkers showed preservation of "stemness." After preclinical development, a phase 1 clinical trial enrolled five patients with unilateral LSC deficiency. Four of these patients received CALEC transplants, establishing preliminary feasibility. Clinical case histories are reported, with no primary safety events. On the basis of these results, a second recruitment phase of the trial was opened to provide longer term safety and efficacy data on more patients.
... As a natural polymer with good biocompatibility, SF has been widely used in biomedical applications, including ophthalmic surgical sutures, artificial corneas, artificial tendons, orthopedic ligaments, cartilage engineering, artificial skin for wound surfaces in traumatology, and anticoagulation stents in cardiology [174][175][176]. Although there is extensive research into the application of SFs in different base materials, most of it is still at the laboratory research stage, and few products have been successfully commercialized and are actually used in clinical treatment (Table 3). ...
Osteoarthritis (OA) is a common joint disease characterized by cartilage damage and degeneration. Traditional treatments such as NSAIDs and joint replacement surgery only relieve pain and do not achieve complete cartilage regeneration. Silk fibroin (SF) biomaterials are novel materials that have been widely studied and applied to cartilage regeneration. By mimicking the fibrous structure and biological activity of collagen, SF biomaterials can promote the proliferation and differentiation of chondrocytes and contribute to the formation of new cartilage tissue. In addition, SF biomaterials have good biocompatibility and biodegradability and can be gradually absorbed and metabolized by the human body. Studies in recent years have shown that SF biomaterials have great potential in treating OA and show good clinical efficacy. Therefore, SF biomaterials are expected to be an effective treatment option for promoting cartilage regeneration and repair in patients with OA. This article provides an overview of the biological characteristics of SF, its role in bone and cartilage injuries, and its prospects in clinical applications to provide new perspectives and references for the field of bone and cartilage repair.
... 20 In a similar report by Liu et al., the authors reported 90% transmission in SF films which were explored for corneal epithelial tissue regeneration. 21 As for the mechanical properties, native corneal tissue is a mechanically tough biological material with Young's modulus ranging between 0.1 and 57 MPa. In this report, the results displayed comparable tensile properties among all three sources, with a tensile strength ranging between 25 and 32 MPa. ...
The demand‐to‐supply gap, rejection rates, and the chances of infection associated with organ/tissue transplantation prompted researchers to find alternative solutions such as tissue engineering. Here, healthy cells are cultured over a biomaterial framework supplemented with growth factors to create bioartificial tissues. As a scaffolding biomaterial, silk fibroin (SF), a biopolymer obtained from Bombyx mori silk cocoons, offers unique properties. However, natural polymers, including SF, were criticized for preconceived source‐dependent batch‐to‐batch variations. Therefore, this study aims to prepare B. mori SF‐based films and investigate source‐dependent variations, if any. For this purpose, we have sourced silk cocoons from three geographical locations in India and processed them into films with a solvent‐casting approach. As a whole, our results indicate that there were slight variations in the morphological features in the raw cocoon stage; however, once processed, there were no significant differences in their topological, physical, chemical, optical, mechanical, or degradable properties with respect to the source. Further, all the films were found to be noncytotoxic and cytocompatible with corneal cells in vitro. Therefore, the study indicates no source‐dependent variations in biopolymers and suggested that SF from any source can be processed into biomaterials for potential biomedical applications.
... Silk has lately gained attention as a bioinspired material of interest for tissue engineering applications requiring regeneration, repair, or replacement of different ocular components [139]. The benefit of silk's transparency prompted Liu et al. to use silk fibroin in place of the human amniotic membrane, which may avoid these potential challenges of transparency, mechanical stability, and disease transmission [140]. Transparency depends on a number of factors, one of which is the orientation and arrangement of individual cells [141,142]. ...
Biomaterial research has led to revolutionary healthcare advances. Natural biological macromolecules can impact high-performance, multipurpose materials. This has prompted the quest for affordable healthcare solutions, with a focus on renewable biomaterials with a wide variety of applications and ecologically friendly techniques. Imitating their chemical compositions and hierarchical structures, bioinspired based materials have elevated rapidly over the past few decades. Bio-inspired strategies entail extracting fundamental components and reassembling them into programmable biomaterials. This method may improve its processability and modifiability, allowing it to meet the biological application criteria. Silk is a desirable biosourced raw material due to its high mechanical properties, flexibility, bioactive component sequestration, controlled biodegradability, remarkable biocompatibility, and inexpensiveness. Silk regulates temporo-spatial, biochemical and biophysical reactions. Extracellular biophysical factors regulate cellular destiny dynamically. This review examines the bioinspired structural and functional properties of silk material based scaffolds. We explored silk types, chemical composition, architecture, mechanical properties, topography, and 3D geometry to unlock the body’s innate regenerative potential, keeping in mind the novel biophysical properties of silk in film, fiber, and other potential forms, coupled with facile chemical changes, and its ability to match functional requirements for specific tissues.
... A small portion of limbal stem cells is collected from the healthy fellow eye (auto-CLET) or, in case of bilateral LSCD, from a living related donor or cadaveric unrelated donor (allo-CLET) [48,49]. Limbal cells are cultured and expanded in vitro using an amniotic membrane [50] or a tissue-engineered scaffold [51,52]. The anterior lens capsule (ALC) can be easily obtained during cataract surgery and, due to the optical transparency and widespread availability, proved to be an excellent scaffold for limbal stem cell proliferation and ocular surface restoration [53,54]. ...
Corneal blindness is the fifth leading cause of blindness worldwide, and therapeutic options are still often limited to corneal transplantation. The corneal epithelium has a strong barrier function, and regeneration is highly dependent on limbal stem cell proliferation and basement membrane remodeling. As a result of the lack of corneal donor tissues, regenerative medicine for corneal diseases affecting the epithelium is an area with quite advanced basic and clinical research. Surgery still plays a prominent role in the treatment of epithelial diseases; indeed, innovative surgical techniques have been developed to transplant corneal and non-corneal stem cells onto diseased corneas for epithelial regeneration applications. The main goal of applying regenerative medicine to clinical practice is to restore function by providing viable cells based on the use of a novel therapeutic approach to generate biological substitutes and improve tissue functions. Interest in corneal epithelium rehabilitation medicine is rapidly growing, given the exposure of the corneal outer layers to external insults. Here, we performed a review of basic, clinical and surgical research reports on regenerative medicine for corneal epithelial disorders, classifying therapeutic approaches according to their macro- or microscopic target, i.e., into cellular or subcellular therapies, respectively.
... As a natural polymer material with good biocompatibility, silk fibroin has been widely used in biomedical fields, including ophthalmic surgical sutures, artificial corneas, artificial tendons, and ligaments in orthopedics, cartilage engineering, artificial skin on the wound surface in the trauma department, and anticoagulant scaffold in the cardiology department (Liu et al., 2012;Jeong et al., 2014;Nakayama et al., 2020). China is a big silk producer with abundant silk fibroin and good basic scientific research on silk fibroin. ...
Background: In this study, a new composite biological mesh named SFP was prepared by combining silk fibroin with polypropylene mesh. The mechanism and clinical application value of the SFP composite mesh were explored.
Methods: The fibrous membrane was prepared by electrospinning of silk fibroin. The silk fibrous membrane was adhered to the polypropylene mesh by fibrin hydrogel to make a new composite mesh. The characterizations were verified by structural analysis and in vitro cell experiments. A total of 40 Sprague–Dawley rats were randomly divided into two groups, and 20 rats in each group were implanted with the SFP mesh and pure polypropylene mesh, respectively. The rats were sacrificed in batches on the 3rd, 7th, 14th, and 90th days after surgery. The adhesion degree and adhesion area on the mesh surface were compared, and a histopathological examination was carried out.
Results: In vitro cell function experiments confirmed that the SFP mesh had good cell viability. The control group had different degrees of adhesion on the 3rd, 7th, 14th, and 90th days after surgery. However, there was almost no intraperitoneal adhesions on the 3rd and 7th days after surgery, and some rats only had mild adhesions on the 14th and 90th days after surgery in the SFP group. There were statistically significant differences in the postoperative intraperitoneal adhesion area and adhesion degree between the two groups (p < 0.05). Histopathological examination confirmed that the mesenchymal cells were well arranged and continuous, and there were more new capillaries and adipocyte proliferation under the mesenchymal cells in the SFP group.
Conclusion: The SFP mesh shows good biocompatibility and biofunction in vitro and in vivo. It can promote the growth of peritoneal mesenchymal cells. The formation of a new mesenchymal cell layer can effectively reduce the extent and scope of adhesion between the mesh and abdominal organs. The SFP mesh will have a good application prospect in the field of abdominal wall hernia repair.
... Additionally, fibrous silk fibroin has also been used as a substrate to induce corneal epithelial differentiation from iPSCs. A cell sheet with tight cell-cell interactions was further formed from differentiated epithelial cells on the same substrate [126]. Epithelial cells differentiated from iPSCs, ESCs, and intestinal stem cells have been used for the formation of tight junctions in studies on gut epithelium transport [127]. ...
Tremendous progress has been made in the past decade regarding our understanding of the gut microbiome’s role in human health. Currently, however, a comprehensive and focused review marrying the two distinct fields of gut microbiome and material research is lacking. To bridge the gap, the current paper discusses critical aspects of the rapidly emerging research topic of “material engineering in the gut microbiome and human health.” By engaging scientists with diverse backgrounds in biomaterials, gut-microbiome axis, neuroscience, synthetic biology, tissue engineering, and biosensing in a dialogue, our goal is to accelerate the development of research tools for gut microbiome research and the development of therapeutics that target the gut microbiome. For this purpose, state-of-the-art knowledge is presented here on biomaterial technologies that facilitate the study, analysis, and manipulation of the gut microbiome, including intestinal organoids, gut-on-chip models, hydrogels for spatial mapping of gut microbiome compositions, microbiome biosensors, and oral bacteria delivery systems. In addition, a discussion is provided regarding the microbiome-gut-brain axis and the critical roles that biomaterials can play to investigate and regulate the axis. Lastly, perspectives are provided regarding future directions on how to develop and use novel biomaterials in gut microbiome research, as well as essential regulatory rules in clinical translation. In this way, we hope to inspire research into future biomaterial technologies to advance gut microbiome research and gut microbiome-based theragnostics.
... In TE silk fibroin films were adopted in several applications. In the corneal epithelium regeneration, culturing in-vitro on rabbits and human Corneal epithelial cells, fibroin was reported to both stimulate the epithelial proliferation and to positively impact the cell migration rate [166,167]. Fibroin films has been also used as a skin patch for wound healing [168], this application was tested in 2017 in a randomized single-blind parallel controlled clinical trial with 71 patients [169] compared to a commercial dressing silk fibroin films were proved to reduce both the healing time and the rate of occurrence of adverse effects. As cardiac patch fibroin was used in combination with other materials [170][171][172]. ...
Silk fibroin has become a prominent material in tissue engineering (TE) over the last 20 years with almost 10,000 published works spanning in all the TE applications, from skeleton to neuronal regeneration. Fibroin is an extremely versatile biopolymer that, due to its ease of processing, has enabled the development of an entire plethora of materials whose properties and architectures can be tailored to suit target applications. Although the research and development of fibroin TE materials and devices is mature, apart from sutures, only a few medical products made of fibroin are used in the clinical routines. <40 clinical trials of Bombyx mori silk-related products have been reported by the FDA and few of them resulted in a commercialized device. In this review, after explaining the structure and properties of silk fibroin, we provide an overview of both fibroin constructs existing in the literature and fibroin devices used in clinic. Through the comparison of these two categories, we identified the burning issues faced by fibroin products during their translation to the market. Two main aspects will be considered. The first is the standardization of production processes, which leads both to the standardization of the characteristics of the issued device and the correct assessment of its failure. The second is the FDA regulations, which allow new devices to be marketed through the 510(k) clearance by demonstrating their equivalence to a commercialized medical product. The history of some fibroin medical devices will be taken as a case study. Finally, we will outline a roadmap outlining what actions we believe are needed to bring fibroin products to the market.
... The potential of SF as a biomaterial for corneal epithelial cell sheet generation has further been evaluated by Liu et al. (2012) [159], who compared the biological cell behavior of human and rabbit primary and immortalized corneal epithelial cells on SF and denuded human amniotic membrane (AM). The cells adhered to and proliferated on both SF and AM, and expression of DNp63a, a progenitor cell marker [160], and keratin 3/12, a differentiation marker [161], existed on both cell culture types. ...
... The potential of SF as a biomaterial for corneal epithelial cell sheet generation has further been evaluated by Liu et al. (2012) [159], who compared the biological cell behavior of human and rabbit primary and immortalized corneal epithelial cells on SF and denuded human amniotic membrane (AM). The cells adhered to and proliferated on both SF and AM, and expression of DNp63a, a progenitor cell marker [160], and keratin 3/12, a differentiation marker [161], existed on both cell culture types. ...
... The cells adhered to and proliferated on both SF and AM, and expression of DNp63a, a progenitor cell marker [160], and keratin 3/12, a differentiation marker [161], existed on both cell culture types. Moreover, after being cultured at the airliquid interface for 7 days, cells on the SF formed a stratified graft with a 2-to 3-cell layering with compact columnar cells on the basal layer and squamous cells present on the apical layers [159]. Since no cytotoxic response or inhibition of cell growth was observed in the SF cultures, their study strongly supports the high biocompatibility of SF biomaterial to corneal epithelial cells and their potential for corneal regeneration. ...
Silk fibroin (SF) is a natural protein (biopolymer) extracted from the cocoons of Bombyx mori L. (silkworm). It has many properties of interest in the field of biotechnology, the most important being biodegradability, biocompatibility and robust mechanical strength with high tensile strength. SF is usually dissolved in water-based solvents and can be easily reconstructed into a variety of material formats, including films, mats, hydrogels, and sponges, by various fabrication techniques (spin coating, electrospinning, freeze-drying, and physical or chemical crosslinking). Furthermore, SF is a feasible material used in many biomedical applications, including tissue engineering (3D scaffolds, wounds dressing), cancer therapy (mimicking the tumor microenvironment), controlled drug delivery (SF-based complexes), and bone, eye and skin regeneration. In this review, we describe the structure, composition, general properties, and structure–properties relationship of SF. In addition, the main methods used for ecological extraction and processing of SF that make it a green material are discussed. Lastly, technological advances in the use of SF-based materials are addressed, especially in healthcare applications such as tissue engineering and cancer therapeutics.
