Injectable nanohydroxyapatite-chitosan-gelatin micro-scaffolds induce regeneration of knee subchondral bone lesions.
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ABSTRACT:
SUBMITTER: Wang B
PROVIDER: S-EPMC5711958 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:UnlabelledHuman embryonic stem cells and adult stem cells have always been the cell source for bone tissue engineering. However, their limitations are obvious, including ethical concerns and/or a short lifespan. The use of human induced pluripotent stem cells (hiPSCs) could avoid these problems. Nanohydroxyapatite (nHA) is an important component of natural bone and bone tissue engineering scaffolds. However, its regulation on osteogenic differentiation with hiPSCs from human gingival fibroblasts (hGFs) is unknown. The purpose of the present study was to investigate the osteogenic differentiation of hiPSCs from patient-derived hGFs regulated by nHA/chitosan/gelatin (HCG) scaffolds with different nHA ratios, such as HCG-111 (1 wt/vol% nHA) and HCG-311 (3 wt/vol% nHA). First, hGFs were reprogrammed into hiPSCs, which have enhanced osteogenic differentiation capability. Second, HCG-111 and HCG-311 scaffolds were successfully synthesized. Finally, hiPSC/HCG complexes were cultured in vitro or subcutaneously transplanted into immunocompromised mice in vivo. The osteogenic differentiation effects of two types of HCG scaffolds on hiPSCs were assessed for up to 12 weeks. The results showed that HCG-311 increased osteogenic-related gene expression of hiPSCs in vitro proved by quantitative real-time polymerase chain reaction, and hiPSC/HCG-311 complexes formed much bone-like tissue in vivo, indicated by cone-beam computed tomography imaging, H&E staining, Masson staining, and RUNX-2, OCN immunohistochemistry staining. In conclusion, our study has shown that osteogenic differentiation of hiPSCs from hGFs was improved by HCG-311. The mechanism might be that the nHA addition stimulates osteogenic marker expression of hiPSCs from hGFs. Our work has provided an innovative autologous cell-based bone tissue engineering approach with soft tissues such as clinically abundant gingiva.SignificanceThe present study focused on patient-personalized bone tissue engineering. Human induced pluripotent stem cells (hiPSCs) were established from clinically easily derived human gingival fibroblasts (hGFs) and defined nanohydroxyapatite/chitosan/gelatin (HCG) scaffolds. hiPSCs derived from hGFs had better osteogenesis capability than that of hGFs. More interestingly, osteogenic differentiation of hiPSCs from hGFs was elevated significantly when composited with HCG-311 scaffolds in vitro and in vivo. The present study has uncovered the important role of different nHA ratios in HCG scaffolds in osteogenesis induction of hiPSCs derived from hGFs. This technique could serve as a potential innovative approach for bone tissue engineering, especially large bone regeneration clinically.
| S-EPMC4704877 | biostudies-literature
Project description:Incorporation of nanohydroxyapatite (nHAP) within a chitosan (CS)/silk fibroin (SF) nanofibrous membrane scaffold (NMS) may provide a favorable microenvironment that more closely mimics the natural bone tissue physiology and facilitates enhanced osteogensis of the implanted cell population. In this study, we prepared pristine CS/SF NMS, composite CS/SF/nHAP NMS containing intrafibrillar nHAP by in situ blending of 10% or 30% nHAP before the electrospinning step, and composite CS/SF/nHAP NMS containing extrafibrillar nHAP by depositing 30% nHAP through alternative soaking surface mineralization. We investigated the effect of the incorporation of HAP nanoparticles on the physicochemical properties of pristine and composite NMS. We confirmed the presence of ~30 nm nHAP in the composite nanofibrous membranes by thermogravimetry analysis (TGA), X-ray diffraction (XRD), and scanning electron microscopy (SEM), either embedded in or exposed on the nanofiber. Nonetheless, the alternative soaking surface mineralization method drastically influenced the mechanical properties of the NMS with 88% and 94% drop in Young's modulus and ultimate maximum stress. Using in vitro cell culture experiments, we investigated the effects of nHAP content and location on proliferation and osteogenic differentiation of human bone marrow mesenchymal stem cells (hMSCs). The proliferation of hMSCs showed no significant difference among pristine and composite NMS. However, the extent of osteogenic differentiation of hMSCs was found to be positively correlated with the content of nHAP in the NMS, while its location within the nanofiber played a less significant role. In vivo experiments were carried out with hMSCs seeded in CS/SF/30%nHAP NMS prepared by in situ blending and subcutaneous implantation in nude mice. Micro-computed tomography images as well as histological and immunohistochemical analysis of the retrieved hMSCs/NMS construct 1 and 2 months postimplantation indicated that NMS had the potential for bone regeneration and can be suggested as a promising scaffold for bone tissue engineering.
| S-EPMC4298333 | biostudies-literature
Project description:In this work, physical cross-linking was used to create nanocomposite hydrogels composed of unmodified gum arabic (GA), chitosan (Ch), and natural nanohydroxyapatite (nHA), using an acrylic acid (AA) solvent. Different GA/chitosan contents (15%, 25%, and 35% of the used AA) as well as different nHA contents (2, 5, and 10 wt.%), were used and studied. The natural nHA and the fabricated GA/Ch/nHA nanocomposite hydrogels were characterized using different analysis techniques. Using acrylic acid solvent produced novel hydrogels with compressive strength of 15.43-22.20 MPa which is similar to that of natural cortical bone. The addition of natural nHA to the hydrogels resulted in a significant improvement in the compressive strength of the fabricated hydrogels. In vitro studies of water absorption and degradation-and in vivo studies-confirmed that the nanocomposite hydrogels described here are biodegradable, biocompatible, and facilitate apatite formation while immersed in the simulated body fluid (SBF). In light of these findings, the GA/Ch/nHA nanocomposite hydrogels are recommended for preparing bioactive nanoscaffolds for testing in bone regeneration applications.
| S-EPMC9370697 | biostudies-literature
Project description:Chitosan is a natural polymer widely investigated and used due to its antibacterial activity, mucoadhesive, analgesic, and hemostatic properties. Its biocompatibility makes chitosan a favorable candidate for different applications in tissue engineering (TE), such as skin, bone, and cartilage tissue regeneration. Despite promising results obtained with chitosan 3D scaffolds, significant challenges persist in fabricating hydrogel structures with ordered architectures and biological properties to mimic native tissues. In this work, chitosan has been investigated aiming at designing and fabricating uniaxial scaffolds which can be proposed for the regeneration of anisotropic tissues (i.e., skin, skeletal muscle, myocardium) by 3D printing technology. Chitosan was blended with gelatin to form a polyelectrolyte complex in two different ratios, to improve printability and shape retention. After the optimization of the printing process parameters, different crosslinking conditions were investigated, and the 3D printed samples were characterized. Tripolyphosphate (TPP) was used as crosslinker for chitosan-based scaffolds. For the optimization of the printing temperature, the sol-gel temperature of the chitosan-gelatin blend was determined by rheological measurements and extrusion temperature was set to 20°C (i.e., below sol-gel temperature). The shape fidelity and surface morphology of the 3D printed scaffolds after crosslinking was dependent on crosslinking conditions. Interestingly, mechanical properties of the scaffolds were also significantly affected by the crosslinking conditions, nonetheless the stability of the scaffolds was strongly determined by the content of gelatin in the blend. Lastly, in vitro cytocompatibility test was performed to evaluate the interactions between L929 cells and the 3D printed samples. 2% w/v chitosan and 4% w/v gelatin hydrogel scaffolds crosslinked with 10% TPP, 30 min at 4°C following 30 min at 37°C have shown cytocompatible and stable characteristics, compared to all other tested conditions, showing suitable properties for the regeneration of anisotropic tissues.
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Project description:It is common knowledge that pure alginate hydrogel is more likely to have weak mechanical strength, a lack of cell recognition sites, extensive swelling and uncontrolled degradation, and thus be unable to satisfy the demands of the ideal scaffold. To address these problems, we attempted to fabricate alginate/bacterial cellulose nanocrystals-chitosan-gelatin (Alg/BCNs-CS-GT) composite scaffolds using the combined method involving the incorporation of BCNs in the alginate matrix, internal gelation through the hydroxyapatite-d-glucono-δ-lactone (HAP-GDL) complex, and layer-by-layer (LBL) electrostatic assembly of polyelectrolytes. Meanwhile, the effect of various contents of BCNs on the scaffold morphology, porosity, mechanical properties, and swelling and degradation behavior was investigated. The experimental results showed that the fabricated Alg/BCNs-CS-GT composite scaffolds exhibited regular 3D morphologies and well-developed pore structures. With the increase in BCNs content, the pore size of Alg/BCNs-CS-GT composite scaffolds was gradually reduced from 200 μm to 70 μm. Furthermore, BCNs were fully embedded in the alginate matrix through the intermolecular hydrogen bond with alginate. Moreover, the addition of BCNs could effectively control the swelling and biodegradation of the Alg/BCNs-CS-GT composite scaffolds. Furthermore, the in vitro cytotoxicity studies indicated that the porous fiber network of BCNs could fully mimic the extracellular matrix structure, which promoted the adhesion and spreading of MG63 cells and MC3T3-E1 cells on the Alg/BCNs-CS-GT composite scaffolds. In addition, these cells could grow in the 3D-porous structure of composite scaffolds, which exhibited good proliferative viability. Based on the effect of BCNs on the cytocompatibility of composite scaffolds, the optimum BCNs content for the Alg/BCNs-CS-GT composite scaffolds was 0.2% (w/v). On the basis of good merits, such as regular 3D morphology, well-developed pore structure, controlled swelling and biodegradation behavior, and good cytocompatibility, the Alg/BCNs-CS-GT composite scaffolds may exhibit great potential as the ideal scaffold in the bone tissue engineering field.
| S-EPMC8400783 | biostudies-literature
Project description:Although regenerative medicine utilizing tissue scaffolds has made enormous strides in recent years, many constraints still hamper their effectiveness. A limitation of many scaffolds is that they form surface patches, which are not particularly effective for some types of "wounds" that are deep within tissues, e.g., stroke and myocardial infarction. In this study, we reported the generation of fibrous micro-scaffolds feasible for delivering cells by injection into the tissue parenchyma. The micro-scaffolds (widths<100?m) were made by dewetting of poly(lactic-co-glycolic acid) thin films containing parallel strips, and cells were seeded to form cell/polymer micro-constructs during or post the micro-scaffold fabrication process. Five types of cells including rat induced vascular progenitor cells were assessed for the formation of the micro-constructs. Critical factors in forming fibrous micro-scaffolds via dewetting of polymer thin films were found to be properties of polymers and supporting substrates, temperature, and proteins in the culture medium. Also, the ability of cells to attach to the micro-scaffolds was essential in forming cell/polymer micro-constructs. Both in vitro and in vivo assessments of injecting these micro-scaffolding constructs showed, as compared to free cells, enhanced cell retention at the injected site, which could lead to improved tissue engineering and regeneration.
| S-EPMC4292840 | biostudies-literature
Project description:Introduction:Bone marrow lesions (BMLs) are frequently identified by MRI in the subchondral bone in knee osteoarthritis (KOA). BMLs are known to be closely associated with joint pain, loss of the cartilage and structural changes in the subchondral trabecular bone (SCTB). Despite this, understanding of the nature of BMLs at the trabecular tissue level is incomplete. Thus, we used Raman microspectroscopy to examine the biochemical properties of SCTB from KOA patients with presence or absence of BMLs (OA-BML, OA No-BML; respectively), in comparison with age-matched cadaveric non-symptomatic controls (Non-OA CTL). Methods:Tibial plateau (TP) specimens were collected from 19 KOA arthroplasty patients (6-Male, 13-Female; aged 56-74 years). BMLs were identified ex-vivo by MRI, using PDFS- and T1-weighted sequences. The KOA specimens were then categorized into an OA-BML group (n = 12; containing a BML within the medial condyle only) and an OA No-BML group (n = 7; with no BMLs identified in the TP). The control (CTL) group consisted of Non-OA cadaveric TP samples with no BMLs and no macroscopic or microscopic evidence of OA-related changes (n = 8; 5-Male, 3-Female; aged 44-80 years). Confocal Raman microspectroscopy, with high spatial resolution, was used to quantify the biochemical properties of SCTB tissue of both the medial and the lateral condyle in each group. Results:The ratios of peak intensity and integrated area of bone matrix mineral (Phosphate (v1), Phosphate (v2) and Phosphate (v4)), to surrogates of the organic phase of bone matrix (Amide I, Proline and Amide III), were calculated. Within the medial compartment, the mineral:organic matrix ratios were significantly lower for OA-BML, compared to Non-OA CTL. These ratios were also significantly lower for the OA-BML medial compartment, compared to the OA-BML lateral compartment. There were no group or compartmental differences for Carbonate:Phosphate (v1, v2 and v4), Amide III (?-helix):Amide III (random-coil), Hydroxyproline:Proline, or Crystallinity. Conclusion:As measured by Raman microspectroscopy, SCTB tissue in BML zones in KOA is significantly less mineralized than the corresponding zones in individuals without OA. These data are consistent with those obtained using other methods (e.g. Fourier transform infrared spectroscopy; FTIR) and with the increased rate of bone remodeling observed in BML zones. Reduced mineralization may change the biomechanical properties of the trabecular bone in BMLs and the mechanical interaction between subchondral bone and its overlying cartilage, with potential implications for the development and progression of OA.
| S-EPMC7210419 | biostudies-literature
Project description:"Bone marrow lesion" (BML) is a common term used to describe the presence of fluid in the bone marrow. Although various pathologies can cause BMLs seen on magnetic resonance imaging, in this Technical Note we focus on treating the lesions associated with osteoarthritis in the knee joint. The role of the subchondral bone in transferring loads within the knee joint, as well as in cartilage homeostasis, is well established. In addition, cartilage and subchondral bone are increasingly considered as an osteochondral unit, rather than as 2 separate structures. Knee osteoarthritis, along with insufficiency fracture, is one of the main indications for the treatment of painful BMLs. Nowadays, there is a growing interest in this field, and new approaches are being developed. Our technique can be defined as a surgical procedure aimed directly at pathology within the subchondral bone and is named "osteo-core plasty." It consists of 2 parts: The first is decompression of bone marrow to decrease intraosseous pressure, and the second is administration of bone marrow aspirate concentrate for better healing potential and bone autograft to deliver supportive tissue. It should be noted that the cause of BMLs must be known before this kind of treatment is performed.
| S-EPMC7695615 | biostudies-literature
Project description:Injectable gelatin hydrogels formed with bioorthogonal click chemistry (ClickGel) are cell-responsive ECM mimics for in vitro and in vivo biomaterials applications. Gelatin polymers with pendant norbornene (GelN) or tetrazine (GelT) groups can quickly and spontaneously crosslink upon mixing, allowing for high viability of encapsulated cells, establishment of 3D elongated cell morphologies, and biodegradation when injected in vivo.
| S-EPMC4849477 | biostudies-literature
Project description:BackgroundThe emergence of skin substitutes provides a new approach for the treatment of wound repair and healing. The consistent and steady release of angiogenic factors is an important factor in the promotion of angiogenesis in skin substitutes, which usually lack, yet need, a vascular network.MethodsIn this study, ginsenoside Rg1, a natural compound isolated from Panax notoginseng (PNS), was incorporated into a collagen/chitosan-gelatin microsphere (CC-GMS) scaffold. The cumulative release kinetics were evaluated, and the effects of the released Rg1 on human umbilical vein endothelial cells (HUVECs) behavior, including proliferation, migration, tube formation, cell-cycle progression, cell apoptosis, and vascular endothelial growth factor (VEGF) secretion, were investigated. Additionally, HUVECs were cultured on the CC-GMS scaffold to test its biocompatibility. Standard Rg1 and VEGF were used as positive controls.ResultsThe results indicated that the CC-GMS scaffold had good release kinetics. The Rg1 released from the CC-GMS scaffold did not lose its activity and had a significant effect on HUVEC proliferation. Both Rg1 and VEGF promoted HUVEC migration and tube formation. Rg1 did not induce HUVEC apoptosis but instead promoted HUVEC progression into the S and G2/M phases of the cell cycle. Rg1 significantly increased VEGF secretion compared with that in the control group. HUVEC culture on the CC-GMS scaffold indicated that this scaffold has good biocompatibility and that CC-GMS scaffolds containing different concentrations of Rg1 promote HUVEC attachment in a dose- and time-dependent manner.ConclusionsRg1 may represent a new class of angiogenic agent that can be encapsulated in CC-GMS scaffolds to exert angiogenic effects in engineered tissue.
| S-EPMC3937171 | biostudies-literature