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Several previous studies in the field of assisted reproduction have focused on the use of blood gel derivatives, such as platelet-rich fibrin (PRF), as a treatment for endometrial rehabilitation. However, the ability to release growth factors and the gel form of this product led to the evolution of platelet lysates. In this study, blood gel derivatives, including PRF lysate, which was in liquid form, and PRF gel, were collected and evaluated for growth factors. It was shown to be effective in endometrial wound healing and regeneration in mouse injured uterine tissue models through structure and function (pinopode expression, embryo implantation) evaluation. The results demonstrated that the concentrations of growth factors, including PDGF-AB and VEGF-A, were higher in the PRF lysate compared to the PRF gel (p < 0.05). PRF lysate could release these growth factors for 8 days. Furthermore, both PRF gel and PRF lysate restored the morphology of injured endometrial tissues in terms of luminal and glandular epithelia, as well as uterine gland secretory activity. However, the presence of pinopodes and embryonic implantation were only observed in the PRF lysate group. It can be concluded that PRF lysate promotes wound healing in mouse injured tissue models in vitro, which can act as healing products in tissue repair.
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Background: Owing to the growing global demand for organ replacement and tissue regeneration, three-dimensional (3D) printing is widely recognized as an essential technology in tissue engineering. Biomaterials become a potential source of raw materials for printing ink by containing factors that promote tissue regeneration. Platelet concentrates are autologous biological products that are capable of doing that. Objectives: This study was carried out to create bioinks capable of providing biological signals by combining gelatin-alginate with platelet concentrates. Methods: This study combined platelet concentrates, including platelet-rich plasma (PRP) and platelet-rich fibrin (PRF), with gelatin and alginate to create bioinks. Bioink properties, including gelatinization and pH, were assessed before printing. After that, the scaffolds were done, and the growth factor (GF) release and cytotoxicity from these scaffolds were performed. Results: Results showed that all the three bioinks, including alginate-gelatin (AG), alginate-gelatin-PRP (AGP), and alginate-gelatin-PRF (AGF) were gelatinized right at the end of bioink fabrication and had a pH around 7. The scaffolds from bioinks supplemented with platelet concentrates secreted GFs that remained for 12 d, and the extracts from them were not cytotoxic for the L929 cell line. Conclusion: In summary, bioinks were made by combining AG with platelet concentrates and had properties suitable for creating scaffolds with cell-oriented grafts in the development of artificial tissues and organs.
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(1) Background: Hemocompatibility is a critical challenge for tissue-derived biomaterial when directly contacting the bloodstream. In addition to surface modification with heparin, endothelialization of the grafted material is suggested to improve long-term clinical efficacy. This study aimed to evaluate the ability to endothelialize in vitro of heparinized bovine pericardial scaffolds. (2) Methods: bovine pericardial scaffolds were fabricated and heparinized using a layer-by-layer assembly technique. The heparinized scaffolds were characterized for heparin content, surface morphology, and blood compatibility. Liquid extraction of the samples was prepared for cytotoxicity testing on human endothelial cells. The in-vitro endothelialization was determined via human endothelial cell attachment and proliferation on the scaffold. (3) Results: The heparinized bovine pericardial scaffold exhibited a heparin coating within its microfiber network. The scaffold surface immobilized with heparin performed good anti-thrombosis and prevented platelet adherence. The proper cytotoxicity impact was observed for a freshly used heparinized sample. After 24 h washing in PBS 1X, the cell compatibility of the heparinized scaffolds was improved. In-vitro examination results exhibited human endothelial cell attachment and proliferation for 7 days of culture. (4) Conclusions: Our in-vitro analysis provided evidence for the scaffold's ability to support endothelialization, which benefits long-term thromboresistance.
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OBJECTIVES: In this study, we aimed to investigate the effects of a mixture of advanced platelet-rich fibrin (A-PRF) and xenogenic bone substitute material (XBSM) on the proliferation and migration of human periodontal ligament stem cells (hPDLSCs) based on the in vitro release of growth factors. MATERIAL AND METHODS: The concentrations of platelet-derived growth factor-AB (PDGF-AB) and vascular endothelial growth factor (VEGF) released by the A-PRF-XBSM mixture were estimated using enzyme-linked immunoassay for up to 7 d. The A-PRF-XBSM mixture exudate was incubated with hPDLSCs. At Days 1, 3, 5, and 7, cell proliferation and migration were investigated by cell counting and wound-healing assays. RESULTS: PDGD-AB and VEGF were released from the A-PRF-XBSM mixture exudate for up to 7 days. hPDLSCs were cultured in media with various concentrations of the A-PRF-XBSM mixture exudate and exhibited their proliferation and migration ability. Furthermore, the factors released from the 100% A-PRF-XBSM mixture exudate had a substantial effect on cell migration, whereas those released from 4% and 20% A-PRF-XBSM mixture exudates stimulated hPDLSC proliferation. CONCLUSIONS: A-PRF-XBSM mixture continuously released growth factors over 7 days and enhanced hPDLSC proliferation and migration. Therefore, A-PRF in combination with XBSM might provide potential advantages for periodontal tissue regeneration.
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Substitutos Ósseos , Fibrina Rica em Plaquetas , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Ligamento Periodontal , Fibrina Rica em Plaquetas/metabolismo , Células-Tronco , Fator A de Crescimento do Endotélio Vascular/metabolismo , Fator A de Crescimento do Endotélio Vascular/farmacologiaRESUMO
Nanosized multi-drug delivery systems provide synergistic effects between drugs and bioactive compounds, resulting in increased overall efficiency and restricted side effects compared to conventional single-drug chemotherapy. In this study, we develop an amphiphilic heparin-poloxamer P403 (HP403) nanogel that could effectively co-load curcuminoid (Cur) and cisplatin hydrate (CisOH) (HP403@CisOH@Cur) via two loading mechanisms. The HP403 nanogels and HP403@CisOH@Cur nanogels were closely analyzed with 1H-NMR spectroscopy, FT-IR spectroscopy, TEM, and DLS, exhibiting high stability in spherical forms. In drug release profiles, accelerated behavior of Cur and CisOH at pH 5.5 compared with neutral pH was observed, suggesting effective delivery of the compounds in tumor sites. In vitro studies showed high antitumor activity of HP403@CisOH@Cur nanogels, while in vivo assays showed that the dual-drug platform prolonged the survival time of mice and prevented tail necrosis. In summary, HP403@CisOH@Cur offers an intriguing strategy to achieve the cisplatin and curcumin synergistic effect in a well-designed delivery platform that increases antitumor effectiveness and overcomes undesired consequences caused by cisplatin in breast cancer treatment.
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Decellularization technology is a process that uses different methods such as physical, chemical or enzymatic methods in order to eliminate cellular remnants from original tissues or organs while minimizing any adverse effect on the structural properties, biological activity, and mechanical integrity of the remaining ECM. Regenerative medicine uses the most promising therapies to replace or regenerate tissues and organs in human, restore or establish normal functions lost due to disease or injury. By the combination between new biomaterials and cells, one of the goals of regenerative medicine is to create autologous grafts for transplantation therapies in the future.Various decellularization methods have been developed include chemical treatment, biological treatment and physical treatment. The aim of this chapter is to evaluate the decellularization method and all available materials that preserves the matrix without structural disruption.
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Engenharia Tecidual , Alicerces Teciduais , Osso e Ossos , Matriz Extracelular , Humanos , Medicina RegenerativaRESUMO
Prosthetic vascular graft infection is one of the most severe vascular surgery complications. Fibrin gel (FG) has many useful characteristics as biocompatibility, biodegradation, adhesion, and haemostasis to develop the local antibiotic delivery system. In this study, human plasma was collected from peripheral blood that was used to create fibrin gel by supplement ion Ca2+. Antibiotic-containing fibrin gel was then evaluated in some characteristics such as surface structure, biodegradation, antibiotic delivery, cytotoxicity, and bacterial biofilm prevention in vitro and in vivo. The results showed that fibrin gel was excellent material for the extended delivery of antibiotics. Most importantly, antibiotic-containing fibrin gel was not toxic for human fibroblast cells in vitro and inhibited bacterial biofilm growth in vitro and in vivo. This research is the first step in developing an antibiotic delivery system for effective graft infection treatments.
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Biofilmes/efeitos dos fármacos , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus/efeitos dos fármacos , Vancomicina/farmacologia , Antibacterianos/química , Antibacterianos/farmacologia , Biofilmes/crescimento & desenvolvimento , Sistemas de Liberação de Medicamentos/tendências , Fibrina/química , Fibrina/farmacologia , Géis/química , Géis/farmacologia , Humanos , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/patogenicidade , Vancomicina/químicaRESUMO
OBJECTIVE: To access the effects of platelet-rich plasma (PRP) on the behaviour of human bone marrow-derived mesenchymal stem cells (hBMSCs), including proliferation and migration. METHODS: PRP was diluted with DMEM/F12, resulting in concentrations of 1%, 2%, and 5%. The proliferation of hBMSCs was examined by 2 methods: cell-number counting with the haemocytometer method and the colony-forming unit-fibroblast (CFU-F) assay. Cell migration was evaluated using the scratch wound healing (SWH) assay; after that, the recorded digital images were analysed by the Image-Analysis J 1.51j8 software to compare the cell-free areas between groups after 0, 24, and 48 hours. RESULTS: hBMSCs cultured in DMEM/F12 at PRP concentrations of 1%, 2%, and 5% were all able to proliferate and migrate. In the 5% PRP group, hBMSCs proliferated greatly with a significantly higher cell number than reported for all other groups on days 5, 7, and 9. CFU-Fs were observed in all groups, except for the negative control group. The SWH assay demonstrated that hBMSCs cultured in 2% and 5% PRP almost filled the artificial wound scratch and significantly migrated more than those of all other groups at both 24 h and 48 h. CONCLUSION: This study indicated that, due to the significant enhancement of cell proliferation and migration, 5% PRP might be the optimal concentration that should be used to promote the potential of hBMSCs in wound healing.
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Bovine pericardium has been proposed as an available material for tissue engineering and bioprosthetic reconstruction. In this study, bovine pericardium was fabricated into a scaffold for culturing and chondrogenic differentiation of human adipose-derived stem cells (hADSCs). Bovine pericardium was treated in 10 mM Tris-HCl and 0.15% SDS, followed by crosslinking in 0.1% glutaraldehyde. Treated bovine pericardium (tBP) was characterized as a slight yellowish thin membrane with enhanced tensile strength and strain property. The membrane maintained stability under enzymatic conditions for up to 16 days of incubation. The results confirmed tBP as a cell-friendly scaffold for hADSCs due to low cytotoxicity and its ability to support an appropriate attachment and proliferation of hADSCs. Moreover, there was an accumulation of the extracellular matrix proteoglycan in tBP seeded with hADSCs after 7 and 14 days of chondrogenic induction. COMP as a specific marker of chondrogenesis was detected after 7 days, whereas type X-a1 collagen (Col10a1) expression was stable up to day 14. However, minor expression of aggrecan was found. Taken together, these results indicate that tBP is a potential scaffold for hADSCs for cartilage tissue engineering.Key words: Bovine pericardium, scaffold, adipose-derived stem cells, chondrogenic differentiation, cartilage regeneration, augmentation rhinoplasty.
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Nanogel-based systems loaded with single anticancer drugs display miscellaneous effectiveness in tumor remission, gradually circumventing mutation and resistance in chemotherapy. Hence, the existence of dual-drug delivered nano-sized systems has been contemporaneous with drug development and preceded the conventional-dose chemotherapy. Among outstanding synergistic drug nanoplatforms, thermosensitive copolymer heparin-Pluronic F127 (Hep-F127) co-delivering cisplatin (CDDP) and curcumins (Cur) (Hep-F127/CDDP/Cur) has emerged as a notable candidate for temperature-responsive drug delivery. The procedure was based on the entrapment of curcumin into the hydrophobic core of bio-degradable co-polymer Hep-F127 while the hydrophilic drug CDDP subsequently conjugated to the backbone heparin to form the core-shell structure. The copolymer was characterized by Fourier transform infrared (FT-IR) spectrophotometry, Transmission Electron Microscopy (TEM), and Dynamic Light Scattering (DLS), to corroborate the successful synthesis and via HPLC along with AES-ICP to evaluate the high drug loading along with a controllable release from the nano-gels. A well-defined nano-shell with size in the 129.3 ± 3.8 nm size range could enhance higher the efficacy of the conjugated-CDDP to Hep-F127 than that of single doses. Moreover, the considerable amount of dual-drug released from thermosensitive nanogels between different conditions (pH = 7.4 and pH = 5.5) in comparison to CDDP from Hep-F127 partially indicated the significantly anti-proliferative ability of Hep-F127/CDDP/Cur to the MCF-7 cell line. Remarkably, drug testing in a xenograft model elucidates the intricate synergism of co-delivery in suppressing tumor growth, which remedies some of the problems affecting in cancer chemotherapy.
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Cisplatino/química , Curcumina/química , Sistemas de Liberação de Medicamentos/métodos , Nanopartículas/química , Difusão Dinâmica da Luz , Humanos , Concentração de Íons de Hidrogênio , Células MCF-7 , Microscopia Eletrônica de Transmissão , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
Burn wound healing is a complex multifactorial process that relies on coordinated signaling molecules to succeed. Curcumin is believed to be a potent antioxidant and anti-inflammatory agent; therefore, it can prevent the prolonged presence of oxygen free radicals which is a significant factor causing inhabitation of optimum healing process. This study describes an extension of study about the biofunctional nanocomposite hydrogel platform that was prepared by using curcumin and an amphiphilic chitosan-g-pluronic copolymer specialized in burn wound healing application. This formular (nCur-CP, nanocomposite hydrogel) was a free-flowing sol at ambient temperature and instantly converted into a nonflowing gel at body temperature. In addition, the storage study determined the great stability level of nCur-CP in long time using UV-Vis and DLS. Morphology and distribution of nCur in its nanocomposite hydrogels were observed by SEM and TEM, respectively. In vitro studies suggested that nCur-CP exhibited well fibroblast proliferation and ability in antimicrobacteria. Furthermore, second- and third-degree burn wound models were employed to evaluate the in vivo wound healing activity of the nCur-CP. In the second-degree wound model, the nanocomposite hydrogel group showed a higher regenerated collagen density and thicker epidermis layer formation. In third degree, the nCur-CP group also exhibited enhancement of wound closure. Besides, in both models, the nanocomposite material-treated groups showed higher collagen content, better granulation, and higher wound maturity. Histopathologic examination also implied that the nanocomposite hydrogel based on nanocurcumin and chitosan could enhance burn wound repair. In conclusion, the biocompatible and injectable nanocomposite scaffold might have great potential to apply for wound healing.
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Queimaduras/terapia , Quitosana/análogos & derivados , Curcumina/administração & dosagem , Hidrogéis/administração & dosagem , Nanocompostos/administração & dosagem , Cicatrização , Alginatos/administração & dosagem , Animais , Antibacterianos/administração & dosagem , Anti-Inflamatórios/administração & dosagem , Antioxidantes/administração & dosagem , Temperatura Corporal , Proliferação de Células , Quitosana/administração & dosagem , Liofilização , Luz , Masculino , Camundongos , Testes de Sensibilidade Microbiana , Poloxâmero/química , Polímeros , Espalhamento de Radiação , Espectrofotometria UltravioletaRESUMO
Supportive membranes have recently been applied to treat periodontal disease in order to achieve periodontal tissue regeneration. The crucial role of these membranes is to facilitate the restoration of the structural and functional periodontium. Bovine pericardium (BP) is mainly composed of collagen type I, which was demonstrated to have good mechanical properties and biological regenerative potential. Our research aimed to extend the application of membrane derived from BP to periodontal disease treatment. However, the fabrication method to achieve a xenogenic-free membrane with the mechanical properties required for periodontal treatment is rarely mentioned. Therefore, a procedure for the extraction and modification of BP using sodium dodecyl sulfate (SDS) and glutaraldehyde (GA) was developed. BP was harvested and decellularized using different SDS concentrations (0.05-0.3%). GA was used to further modify the membranes to achieve suitable thickness, mechanical strength, and pore size. A combination protocol of 0.15% SDS treatment for 12 h with continuous agitation combined with 0.1% GA for 6 h for membrane fabricating was applied. The modified BP (mBP) had the targeted characteristics, such as 0.2-0.5 mm thickness, approximately 10 MPa in tensile strength, 30% in strain force, and a pore size <5 µm, which is comparable to commercially available collagen membranes. Findings from this study demonstrated that the established method was effective in preparing BP membrane for periodontal treatment while decreasing the concentration of reagents and processing time. Moreover, our modified membrane was found to have no cytotoxicity but supports the migration, attachment, and proliferation of human gingival fibroblasts in vitro. Taken together, these results confirmed that mBP is suitable for application in periodontal disease treatment and regeneration.
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OBJECTIVES: This study aimed to simplify the collection, isolation and cryopreservation procedure of human dental pulp stem cells (DPSCs) to ease the establishment of dental stem cell banking. DESIGN: Extracted third molars were collected and stored either in growth medium or in gentamicin-saline (480µg/ml) for 6, 9 or 12h. DPSCs were isolated and subjected to cryopreservation by a controlled-rate or rapid freezing method in 5 or 10% DMSO. Flow cytometry and growth pattern of DPSCs before and after cryopreservation were conducted. RESULTS: Rate of contamination by which the extracted teeth were stored in control and gentamicin-saline were 9.1% (N=33) and 2.3% (N=43), respectively. Successful cell isolation rate of teeth preserved in gentamicin-saline at 6h (92.9%) was comparable to those of growth media group (90.3%). At 9 and 12h, the rates dropped significantly to 75% and 54%, respectively. Cryopreservation by controlled-rate freezing either in 5 or 10% DMSO resulted in a significantly higher percentage of viable cells than by rapid freezing. Cells conserved by controlled-rate freezing in 5% DMSO showed a pattern of growth similar to control unfrozen cells; 10% DMSO significantly deteriorated the growth pattern of the cells. After thawing, DPSCs conserved by controlled-rate freezing still expressed stemness characteristics, although hematopoietic stem cell markers were slightly increased. CONCLUSION: Gentamicin-saline was effective in preserving human teeth for DPSC isolation. Controlled-rate freezing in 5% DMSO gave the highest rate of cell viability. This study simplifies the storage conditions and proposes a simple method for cryopreservation of DPSCs.
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Criopreservação/métodos , Polpa Dentária/citologia , Células-Tronco Mesenquimais/fisiologia , Adolescente , Adulto , Sobrevivência Celular , Citometria de Fluxo , Gentamicinas/farmacologia , Humanos , Dente Serotino , Cloreto de Sódio/farmacologia , Extração DentáriaRESUMO
Regeneration of dentin tissues in the pulp space of teeth serves the ultimate goal of preserving teeth via endodontic approaches. In recent times, many studies suggested that human dentin scaffolds combined with dental stem cells was a potential strategy for the complete dentin tissue regeneration. In this study, human dental pulp stem cells (DPSCs) were isolated and cultured. Dentin specimens were prepared from human third molars and treated with ethylene diamine tetra-acetic acid and citric acid to remove the smear layer. Then, DPSCs were cultured onto human treated dentin (hTD) and implanted in mouse model for 4, 6 and 8 weeks. The resulting grafts were assessed by hematoxylin and eosin stain and immunohistochemical stains. As a result, DPSCs were supported and induced to regenerate of dentin-like tissues which expressed specific dentin markers such as dentin sialophosphoprotein and dentin matrix protein 1 by combination with hTD in vivo. Furthermore, cells existed in the newly-formed dentin-like tissues also expressed typical human mitochondria antibodies, demonstrated that new tissues originated from human. In conclusion, the obtain results extend hopefully newly-established therapy to apply in endodontics and traumatic dental hard tissues.
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Polpa Dentária/citologia , Dentina/crescimento & desenvolvimento , Regeneração Tecidual Guiada Periodontal/instrumentação , Transplante de Células-Tronco/métodos , Células-Tronco/citologia , Alicerces Teciduais , Dentina/química , Dentina/citologia , Desenho de Equipamento , Análise de Falha de Equipamento , Humanos , Regeneração/fisiologiaRESUMO
Periodontal ligament (PDL) is a specialized connective tissue that connects cementum and alveolar bone to maintain and support the teeth in situ and preserve tissue homeostasis. Recent studies have revealed the existence of stem cells in human dental tissues including periodontal ligament that play an important role, not only in the maintenance of the periodontium but also in promoting periodontal regeneration. In this study, human periodontal ligament cells (hPDLCs) were isolated by outgrowth and enzymatic dissociation methods. Expression of surface markers on PDLCs as human mesenchymal stem cells (MSCs) was identified by flow cytometry. In addition, proliferation and differentiation capacity of cultured cells to osteoblasts, adipocytes were evaluated. As a result, we successfully cultured cells from the human periodontal ligament tissues. PDLCs express mesenchymal stem cell (MSC) markers such as CD44, CD73, and CD90 and do not express CD34, CD45, and HLA-DR. PDLCs also possess the multipotential to differentiate into various types of cells, such as osteoblast and adipocytes, in vitro. Therefore, these cells have high potential to serve as materials for tissue engineering, especially dental tissue engineering.