Intervertebral disc degeneration is a clinical disease that reduces the quality of patient's life. The degeneration usually initiates in the nucleus pulposus (NP), hence the use of hydrogels ...represents a promising therapeutic approach. However, the viscoelastic nature of hydrogel and its ability to provide biomimetic architecture and biochemical cues influence the regeneration capability. This study focused on tuning the physical nature of a glycosaminoglycan hydrogel (κ-carrageenan) as well as the release kinetics of a chondrogenic factor (kartogenin - KGN) through physical cross-linking. For this, κ-carrageenan was cross linked with 2.5 % and 5 % potassium chloride (KCl) for 15 and 30 min and loaded with KGN molecule at 50 μM and 100 μM. The tight network structure with low water retention and degradation property was seen in hydrogel cross-linked with increased KCl concentration and time. However, optimal degradation along with NP mimicking viscoelastic nature was exhibited by 5 wt% KCl treated hydrogel (H3 hydrogel). All hydrogel groups exhibited burst KGN release at 24 h followed by a sustained release for 5 days. However, hydrogel cross-linked with 5 wt% KCl enhanced chondrogenic differentiation, mainly at lower KGN dose. In summary, this study shows the potential application of biomimetic KGN laden carrageenan hydrogel in NP regeneration.
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Transplantation of synovial fluid-derived mesenchymal stem cells (SF-MSCs) is a viable therapy for cartilage degeneration of osteoarthritis (OA). But controlling chondrogenic differentiation of the ...transplanted SF-MSCs in the joints remains a challenge. Kartogenin (KGN) is a small molecule that has been discovered to induce differentiation of SF-MSCs to chondrocytes both in vitro and in vivo. The clinical application of KGN however is limited by its low water solubility. KGN forms precipitates in the cell, resulting in low effective concentration and thus limiting its chondrogesis-promoting activity. Here we report that targeted delivery of KGN to SF-MSCs by engineered exosomes leads to even dispersion of KGN in the cytosol, increases its effective concentration in the cell, and strongly promotes the chondrogenesis of SF-MSCs in vitro and in vivo. Fusing an MSC-binding peptide E7 with the exosomal membrane protein Lamp 2b yields exosomes with E7 peptide displayed on the surface (E7-Exo) that has SF-MSC targeting capability. KGN delivered by E7-Exo efficiently enters SF-MSCs and induces higher degree of cartilage differentiation than KGN alone or KGN delivered by exosomes without E7. Co-administration of SF-MSCs with E7-Exo/KGN in the knee joints via intra-articular injection also shows more pronounced therapeutic effects in a rat OA model than KGN alone or KGN delivered by exosomes without E7. Altogether, transplantation of SF-MSCs with in situ chondrogenesis enabled by E7-Exo delivered KGN holds promise towards as an advanced stem cell therapy for OA.
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Hydrogels have been widely used as the carrier material of therapeutic cell and drugs for articular cartilage repair. We previously demonstrated a unique host-guest macromer (HGM) approach to prepare ...mechanically resilient, self-healing and injectable supramolecular gelatin hydrogels free of chemical crosslinking. In this work, we show that compared with conventional hydrogels our supramolecular gelatin hydrogels mediate more sustained release of small molecular (kartogenin) and proteinaceous (TGF-β1) chondrogenic agents, leading to enhanced chondrogenesis of the encapsulated human bone marrow-derived mesenchymal stem cells (hBMSCs) in vitro and in vivo. More importantly, the supramolecular nature of our hydrogels allows injection of the pre-fabricated hydrogels containing the encapsulated hBMSCs and chondrogenic agents, and our data show that the injection process has little negative impact on the viability and chondrogenesis of the encapsulated cells and subsequent neocartilage development. Furthermore, the stem cell-laden supramolecular hydrogels administered via injection through a needle effectively promote the regeneration of both hyaline cartilage and subchondral bone in the rat osteochondral defect model. These results demonstrate that our supramolecular HGM hydrogels are promising delivery biomaterials of therapeutic agents and cells for cartilage repair via minimally invasive procedures. This unique capability of injecting cell-laden hydrogels to target sites will greatly facilitate stem cell therapies.
Cartilage lesions can progress into secondary osteoarthritis and cause severe clinical problems in numerous patients. As a prospective treatment of such lesions, human-derived induced pluripotent ...stem cells (iPSCs) were shown to be 3D bioprinted into cartilage mimics using a nanofibrillated cellulose (NFC) composite bioink when co-printed with irradiated human chondrocytes. Two bioinks were investigated: NFC with alginate (NFC/A) or hyaluronic acid (NFC/HA). Low proliferation and phenotypic changes away from pluripotency were seen in the case of NFC/HA. However, in the case of the 3D-bioprinted NFC/A (60/40, dry weight % ratio) constructs, pluripotency was initially maintained, and after five weeks, hyaline-like cartilaginous tissue with collagen type II expression and lacking tumorigenic Oct4 expression was observed in 3D -bioprinted NFC/A (60/40, dry weight % relation) constructs. Moreover, a marked increase in cell number within the cartilaginous tissue was detected by 2-photon fluorescence microscopy, indicating the importance of high cell densities in the pursuit of achieving good survival after printing. We conclude that NFC/A bioink is suitable for bioprinting iPSCs to support cartilage production in co-cultures with irradiated chondrocytes.
Abstract
Objectives
Owing to its avascular nature, cartilage tissue has a restricted capacity for regeneration. These structural features make it difficult for a fully functional tissue to regenerate ...after damage. Therefore, studies aiming at cartilage tissue regeneration are getting quite interesting. In this study, we employed a novel approach to induce chondrogenic differentiation using a collagen mimetic peptide amphihile (PA) nanofiber. The nanofiber comprised a specific peptide sequence – glycine-phenylalanine-hydroxyproline-glycine-glutamate-arginine (GFOGER), corresponding to the α1 (I) collagen chain. This sequence was selected for its ability to mimic the structure and function of natural collagen in the extracellular matrix (ECM). This specific peptide sequence is expected to enhance the chondrogenic differentiation process by providing a more efficient and effective method for tissue engineering applications.
Methods
ATDC5 cells were cultured on the synthetic scaffold of collagen-mimicking PA nanofibers, facilitating adhesion, division, and chondrogenic cell differentiation.
Results
In our study, ATDC5 cells cultured on collagen mimetic peptide nanofiber expressed chondrogenic marker proteins, namely Collagen II and Sox9, significantly high at the 5th and 10th days compared to cells cultured on TCP in the absence of insulin as inducer.
Conclusions
According to our results, the collagen mimetic peptide-based scaffold supports cell growth and differentiation by mimicking the natural cell matrix.
In indirect co-culture system, chondrocytes can induce differentiation of bone marrow mesenchymal stem cells (BMSCs) to chondrocytes without additional inducer. The participation of microRNAs ...(miRNAs) may take part in the chondrogenic differentiation. Present study aimed to investigate the effect and mechanism of chondrocytes-derived exosomal miRNA in BMSCs chondrogenic differentiation. Our data showed that miR-8485 was the exosomal miRNA derived from chondrocytes and transmitted to BMSCs. Functionally, miR-8485 silence in chondrocytes impaired exosome-induced chondrogenic differentiation of BMSCs. Mechanistically, exosomal miR-8485 targeted GSK3B to repress GSK-3β expression and targeted DACT1 to induce p-GSK-3β (Ser9), activating Wnt/β-catenin pathways. Our study firstly showed that chondrocytes-derived exosomal miR-8485 regulated the Wnt/β-catenin pathways to promote chondrogenic differentiation of BMSCs, providing innovative thoughts for cartilage reconstruction.
•miR-8485, an exosomal miRNA, could be transmit from chondrocytes to BMSCs.•miR-8485 played a role in exosome-induced chondrogenic differentiation of BMSCs.•miR-8485 targeted GSK3B by repressing GSK-3β expression and inducing GSK-3β phosphorylation to active Wnt/β-catenin pathway.
Heterotopic ossification (HO) is a pathological bone formation based on endochondral ossification distinguished by ossification within muscles, tendons, or other soft tissues. There has been growing ...studies focusing on the treatment with rapamycin to inhibit HO, but the mechanism of mTORC1 on HO remains unclear. Tendon cells (TDs) are the first cells to form during tendon heterotopic ossification. Here, we used an in vivo model of HO and an in vitro model of chondrogenesis induction to elucidate the effect and underlying mechanism of mTORC1 in HO. The current study highlights the effect of rapamycin on murine Achilles tenotomy-induced HO and the role of mTORC1 signaling pathway on TDs. Our result showed that mTORC1 was activation in the early stage of HO, whereas the mTORC1 maintained low expression in the mature ectopic cartilage tissue and the ectopic bone formation sites. The use of mTORC1-specific inhibitor (rapamycin) immediately after Achilles tendon injury could suppress the formation of HO; once ectopic cartilage and bone had formed, treatment with rapamycin could not significantly inhibit the progression of HO.
Mechanistically, mTORC1 stimulation by silencing of TSC1 promoted the expression of the chondrogenic markers in TDs. In TDs, treated with mTORC1 stimulation by silencing of TSC1, mTORC1 increased the activation of the NF-κB signaling pathway. NF-κB selective inhibitor BAY11-7082 significantly suppressed the chondrogenesis of TDs that treated with mTORC1 stimulation by silencing of TSC1. Together, our findings demonstrated that mTORC1 promoted HO by regulating TDs chondrogenesis partly through the NF-κB signaling pathway; and rapamycin could be a viable HO therapeutic regimen.
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•Mg-QCS/PF composite hydrogels mediated the sustained release of magnesium ions.•The hydrogels exhibited self-healing, adhesive properties and biocompatibility.•The hydrogels greatly ...promoted BMSCs adhesion and chondrogenic differentiation.•The hydrogels significantly increased the fibrocartilage regeneration in vivo.
The fibrocartilaginous enthesis regeneration after rotator repair is still a major challenge. Although magnesium-based alloy orthopedic implant is effective to promote fibrocartilage formation at the tendon-bone interface in the anterior cruciate ligament reconstruction model, it was limited in the rotator cuff repair for its special anatomical structure. Herein, we developed a multifunctional self-healing magnesium ions-quaternized chitosan/Pluronic® F127 (Mg-QCS/PF) hydrogels to achieve in-situ and customized release of Mg2+, and demonstrated the sustained release of Mg2+ from the hydrogel to significantly promote the rotator cuff repair in the rabbit rotator cuff tear model. The obtained hydrogels showed excellent self-healing and anti-compressive performance. Additionally, the good injectability and adhesive properties of the hydrogels make it easier and stable to deliver Mg2+ at the tendon-bone interface with irregular shapes. The release of Mg2+ from Mg-QCS/PF hydrogels improved the adhesion, proliferation and migration of bone mesenchymal stem cells (BMSCs) and MC3T3 cells in vitro compared with QCS/PF without Mg2+. Furthermore, the composite hydrogels significantly enhanced the fibrocartilaginous interface regeneration in the rabbit rotator cuff tear model in terms of repaired tendon mature scores, fibrocartilage regeneration, collagen remodeling and biomechanical properties. This is the first study to demonstrate the positive effects of Mg2+ for the rotator cuff healing in the rabbit preclinical model, and the results indicate that the acellular injectable self-healing Mg-doped hydrogels are candidates to effectively promote in situ regeneration of rotator cuff.
Cartilage regeneration by biomimetic cartilage matrix with synchronously recruited stem cells was one of ideal strategies. Inspired by catechol for proteins adhesion, dopamine modified polysaccharide ...hybrid hydrogel (HD-C) was prepared by integrating collagen I (Col I) and hyaluronic acid derivatives (HA-DN) with sulfhydryl modified polysaccharide hybrid hydrogel (HS-C) as control. Because of double-crosslinking architecture, HD-C hydrogel was endowed with a more compact pore structure, higher mechanical properties and water retention ability in comparison with those of HS-C hydrogel. Meanwhile, it significantly promoted the proliferation and spread of rabbit bone marrow stem cells (rBMSCs), and accelerated cartilaginous matrix secretion. RT-PCR results also verified higher related gene expression of chondrogenesis (Sox 9, Agg and Col II). Moreover, HD-C hydrogel could enhance the enrichment and migration of rBMSCs in vitro by potential functional protein adsorption mechanisms, and this phenomenon was further confirmed by more rBMSCs migration in short-term joint implantation experiments in vivo.
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•Dopamine modified hybrid hydrogel had higher mechanical and swelling properties.•The HD-C hydrogel promoted adhesion, spread and GAG secretion of rBMSCs in vitro.•The catechol of HD-C facilitated potential functional protein adsorption in vitro.•The HD-C hydrogel enhanced enrichment and migration of rBMSCs in vitro and in vivo.
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•Exo-AMG shows promise for treating growth plate injuries by promoting cartilage regeneration.•It fosters a conducive microenvironment of growth plate regeneration.•It suppresses ...inflammation, and enhances BMSC proliferation and chondrogenic differentiation.
The growth plate is vital for childhood development, and most of the injuries can formation of bone bridges on it, which has negative impact on limb development. Current biomaterial scaffolds often lack essential cartilage matrix components, which hurdles to get an acceptable regeneration. Additionally, stem cell transplantation faces challenges including high apoptosis rates and uncertain differentiation. Then, we designed an injectable composite hydrogel (Exo-AMG) loaded with exosomes (Exo) and composed of methacrylate-modified acellular cartilage matrix (ACMMA) and gelatin methacrylate (GelMA) to repair the growth plate for children and adolescents. The hydrogel is administered via minimally invasive injection and undergoes in-situ photocrosslinking to fill the growth plate defects. It exhibited suitable mechanical properties, biodegradation rates; it also could sustain exosome release. Moreover, it promoted M2 macrophage polarization and secretion of anti-inflammatory factors, and it enhanced bone mesenchymal stem cells (BMSCs) proliferation and chondrogenic differentiation in vitro. In vivo study, this Exo-AMG induced growth plate cartilage regeneration without bone bridge formation. Therefore, this injectable and in-situ photocrosslinkable composite hydrogel is promising on growth plate injury repair by alleviating inflammation and promoting chondrogenic differentiation.
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