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Extracellular vesicles (EVs) have been found to have the characteristics of their parent cells. Based on the characteristics of these EVs, various studies on disease treatment using mesenchymal stem cell (MSC)-derived EVs with regenerative activity have been actively conducted. The therapeutic nature of MSC-derived EVs has been shown in several studies, but in recent years, there have been many efforts to functionalize EVs to give them more potent therapeutic effects. Strategies for functionalizing EVs include endogenous and exogenous methods. In this study, human umbilical cord MSC (UCMSC)-derived EVs were selected for optimum OA treatments with expectation via bioinformatics analysis based on antibody array. And we created a novel nanovesicle system called the IGF-si-EV, which has the properties of both cartilage regeneration and long-term retention in the lesion site, attaching positively charged insulin-like growth factor-1 (IGF-1) to the surface of the UCMSC-derived Evs carrying siRNA, which inhibits MMP13. The downregulation of inflammation-related cytokine (MMP13, NF-kB, and IL-6) and the upregulation of cartilage-regeneration-related factors (Col2, Acan) were achieved with IGF-si-EV. Moreover, the ability of IGF-si-EV to remain in the lesion site for a long time has been proven through an ex vivo system. Collectively, the final constructed IGF-si-EV can be proposed as an effective OA treatment through its successful MMP13 inhibition, chondroprotective effect, and cartilage adhesion ability. We also believe that this EV-based nanoparticle-manufacturing technology can be applied as a platform technology for various diseases.
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Vesículas Extracelulares , Fator de Crescimento Insulin-Like I , Células-Tronco Mesenquimais , Osteoartrite , RNA Interferente Pequeno , Fator de Crescimento Insulin-Like I/metabolismo , Vesículas Extracelulares/metabolismo , Humanos , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/citologia , Osteoartrite/terapia , Osteoartrite/metabolismo , RNA Interferente Pequeno/genética , Animais , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 13 da Matriz/genéticaRESUMO
Human induced pluripotent stem cells (iPSCs) hold great promise for personalized medicine, as they can be differentiated into specific cell types, especially mesenchymal stem cells (MSCs). Therefore, our study sought to assess the feasibility of deriving MSCs from teratomas generated from human iPSCs. Teratomas serve as a model to mimic multilineage human development, thus enriching specific somatic progenitors and stem cells. Here, we discovered a small, condensed mass of MSCs within iPSC-generated teratomas. Afterward, we successfully isolated MSCs from this condensed mass, which was a byproduct of teratoma development. To evaluate the characteristics and cell behaviors of iPSC-derived MSCs (iPSC-MSCs), we conducted comprehensive assessments using qPCR, immunophenotype analysis, and cell proliferation-related assays. Remarkably, iPSC-MSCs exhibited an immunophenotype resembling that of conventional MSCs, and they displayed robust proliferative capabilities, similar to those of higher pluripotent stem cell-derived MSCs. Furthermore, iPSC-MSCs demonstrated the ability to differentiate into multiple lineages in vitro. Finally, we evaluated the therapeutic potential of iPSC-MSCs using an osteochondral defect model. Our findings demonstrated that teratomas are a promising source for the isolation of condensed MSCs. More importantly, our results suggest that iPSC-MSCs derived from teratomas possess the capacity for tissue regeneration, highlighting their promise for future therapeutic applications.
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Zinc oxide nanostructures (ZnO NS) were fabricated in situ within a ternary hydrogel system composed of carboxymethyl cellulose-agarose-polyvinylpyrrolidone (CAP@ZnO TNCHs) by a one-pot method employing moist-heat solution casting. The percentages of CMC and ZnO NS were varied in the CAP hydrogel films and then they were investigated by different techniques, such as ATR/FTIR, TGA, XRD, XPS, and FE-SEM analysis. Furthermore, the mechanical properties, hydrophilicity, swelling, porosity, and antibacterial activity of the CAP@ZnO TNCHs were studied. In-vitro biocompatibility assays were performed with skin fibroblast (CCD-986sk) cells. In-vitro culture of CCD-986sk fibroblasts showed that the ZnO NS facilitated cell adhesion and proliferation. Furthermore, the application of CAP@ZnO TNCHs enhanced cellular interactions and physico-chemical, antibacterial bacterial, and biological performance relative to unmodified CAP hydrogels. Also, an in vivo wound healing study verified that the CAP@ZnO TNCHs promoted wound healing significantly within 18 days, an effect superior to that of unmodified CAP hydrogels. Hence, these newly developed cellulose-based ZnO TNCHs are promising materials for wound healing applications.
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Nanoestruturas , Óxido de Zinco , Hidrogéis/farmacologia , Hidrogéis/química , Óxido de Zinco/farmacologia , Óxido de Zinco/química , Carboximetilcelulose Sódica/química , Antibacterianos/química , Nanoestruturas/química , CicatrizaçãoRESUMO
Osteoporosis is a pathological condition characterized by an accelerated bone resorption rate, resulting in decreased bone density and increased susceptibility to fractures, particularly among the elderly population. While conventional treatments for osteoporosis have shown efficacy, they are associated with certain limitations, including limited drug bioavailability, non-specific administration, and the occurrence of adverse effects. In recent years, nanoparticle-based drug delivery systems have emerged as a promising approach for managing osteoporosis. Nanoparticles possess unique physicochemical properties, such as a small size, large surface area-to-volume ratio, and tunable surface characteristics, which enable them to overcome the limitations of conventional therapies. These nanoparticles offer several advantages, including enhanced drug stability, controlled release kinetics, targeted bone tissue delivery, and improved drug bioavailability. This comprehensive review aims to provide insights into the recent advancements in nanoparticle-based therapy for osteoporosis. It elucidates the various types of nanoparticles employed in this context, including silica, polymeric, solid lipid, and metallic nanoparticles, along with their specific processing techniques and inherent properties that render them suitable as potential drug carriers for osteoporosis treatment. Furthermore, this review discusses the challenges and future suggestions associated with the development and translation of nanoparticle drug delivery systems for clinical use. These challenges encompass issues such as scalability, safety assessment, and regulatory considerations. However, despite these challenges, the utilization of nanoparticle-based drug delivery systems holds immense promise in revolutionizing the field of osteoporosis management by enabling more effective and targeted therapies, ultimately leading to improved patient outcomes.
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Spinal fusion surgery is a surgical technique that connects one or more vertebrae at the same time to prevent movement between the vertebrae. Although synthetic bone substitutes or osteogenesis-inducing recombinant proteins were introduced to promote bone union, the rate of revision surgery is still high due to pseudarthrosis. To promote successful fusion after surgery, stem cells with or without biomaterials were introduced; however, conventional 2D-culture environments have resulted in a considerable loss of the innate therapeutic properties of stem cells. Therefore, we conducted a preclinical study applying 3D-spheroids of human bone marrow-dewrived mesenchymal stem cells (MSCs) to a mouse spinal fusion model. First, we built a large-scale manufacturing platform for MSC spheroids, which is applicable to good manufacturing practice (GMP). Comprehensive biomolecular examinations, which include liquid chromatography-mass spectrometry and bioinformatics could suggest a framework of quality control (QC) standards for the MSC spheroid product regarding the identity, purity, viability, and potency. In our animal study, the mass-produced and quality-controlled MSC spheroids, either undifferentiated or osteogenically differentiated were well-integrated into decorticated bone of the lumbar spine, and efficiently improved angiogenesis, bone regeneration, and mechanical stability with statistical significance compared to 2D-cultured MSCs. This study proposes a GMP-applicable bioprocessing platform and QC directions of MSC spheroids aiming for their clinical application in spinal fusion surgery as a new bone graft substitute.
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Substitutos Ósseos , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais , Fusão Vertebral , Animais , Camundongos , Humanos , Fusão Vertebral/métodos , Transplante de Células-Tronco Mesenquimais/métodos , Medula Óssea , Osteogênese , Materiais Biocompatíveis , Proteínas RecombinantesRESUMO
Bone growth occurs in the epiphyseal growth plate (EGP) and epiphyseal growth plate cells (EGPCs) exist in EGP. EGPCs, including skeletal stem cells (SSCs), are cells that induce bone growth and development through endochondral ossification. Recently, the superiority of bone regeneration through endochondral ossification has been reported. Our study compared EGPCs with bone marrow-derived mesenchymal stem cells (BM-MSCs) and suggested the therapeutic potential of new bone regeneration. In this study, we analyzed the characteristics between EGPCs and BM-MSCs based on morphological characteristics and molecular profiles. EGPCs expressed chondrogenic and osteogenic markers higher than BM-MSCs. Additionally, in co-culture with BM-MSCs, EGPCs induced an increase in chondrogenic, osteogenic, and hypertrophic markers of BM-MSCs. Finally, EGPCs induced higher bone regeneration than BM-MSCs in the osteoporosis model. Overall, we suggest the possibility of EGPCs as cell therapy for effective bone regeneration.
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Osteoarthritis (OA) is an incurable joint disease affecting 240 million elderly population, and major unmet medical needs exist for better therapeutic options for OA. During skeletal development, Nkx3.2 has been shown to promote chondrocyte differentiation and survival, but to suppress cartilage hypertrophy and blood vessel invasion. Here we show that Nkx3.2 plays a key role in osteoarthritis (OA) pathogenesis. Marked reduction of Nkx3.2 expression was observed in three different murine OA models. Consistent with these findings, analyses of surgery-induced and age-driven OA models revealed that cartilage-specific post-natal induction of Nkx3.2 can suppress OA progression in mice. These results suggest that Nkx3.2 may serve as a promising target for OA drug development.
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Proteínas de Homeodomínio/metabolismo , Osteoartrite/metabolismo , Fatores de Transcrição/metabolismo , Animais , Modelos Animais de Doenças , Proteínas de Homeodomínio/genética , Camundongos , Osteoartrite/patologia , Osteoartrite/cirurgia , Fatores de Transcrição/genéticaRESUMO
Extracellular matrix (ECM) remodeling is necessary for the development and self-healing of tissue, and the process is tissue specific. Matrix metalloproteinases (MMPs) play a role in ECM remodeling by unwinding and cleaving ECM. We hypothesized that ECM remodeling by MMPs is involved in the differentiation of stem cells into specific lineages during self-healing. To prove the hypothesis, we investigated which MMPs are involved in the osteogenic differentiation of human mesenchymal stem cells (hMSCs) grown on a type I collagen (Col I) matrix, and we found that specifically high expression of MMP13 in hMSCs grown on a Col I matirx during osteogenic differentiation. Moreover, knocking down of MMP13 decreased the osteogenic differentiation of hMSCs grown on a Col I matrix. In addition, pre-treatment of recombinant human MMP13 lead to remodeling of Col I matrix and increased the osteogenic differentiation of hMSCs and in vivo bone formation following the upregulation of the expression of runt-related transcription factor 2 (RUNX2), integrin α3 (ITGA3), and focal adhesion kinase. Furthermore, the transcription factor RUNX2 bound to the MMP13 promoter. These results suggest that growth on a remodeled Col I matrix by MMP13 stimulates osteogenic differentiation of hMSCs and self-healing of bone tissue via an MMP13/ITGA3/RUNX2 positive feedback loop. STATEMENT OF SIGNIFICANCE: Self-healing of tissue could be the key to treating diseases that cannot be overcome by present technology. We investigated the mechanism underlying the self-healing of tissue and we found that the osteogenic differentiation was increased in hMSCs grown on a remodeled Col I matrix by the optimized concentration of MMP13 not in hMSCs grown on a Col I fragments cleaved by a high concentration of MMP13. In addition, we found the remodeled Col I matrix by MMP13 increased the osteogenic capacity through a MMP13/integrin α3/RUNX2 positive feedback loop. This result would be able to not only provide a strategy for bone tissue-specific functional materials following strong evidence about the self-healing mechanism of bone through the interaction between stem cells and the ECM matrix. As such, we strongly believe our finding will be of interest to researchers studying biomaterials, stem cell biology and matrix interaction for regenerative medicine and therapy.
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Células-Tronco Mesenquimais , Osteogênese , Regeneração Óssea , Osso e Ossos , Diferenciação Celular , Células Cultivadas , Colágeno , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Retroalimentação , Humanos , Integrina alfa3 , Ligantes , Metaloproteinase 13 da Matriz/genéticaRESUMO
Adipose-derived mesenchymal stromal cells (Ad-MSCs) are a promising tool for articular cartilage repair and regeneration. However, the terminal hypertrophic differentiation of Ad-MSC-derived cartilage is a critical barrier during hyaline cartilage regeneration. In this study, we investigated the role of matrilin-3 in preventing Ad-MSC-derived chondrocyte hypertrophy in vitro and in an osteoarthritis (OA) destabilization of the medial meniscus (DMM) model. Methacrylated hyaluron (MAHA) (1%) was used to encapsulate and make scaffolds containing Ad-MSCs and matrilin-3. Subsequently, the encapsulated cells in the scaffolds were differentiated in chondrogenic medium (TGF-ß, 1-14 days) and thyroid hormone hypertrophic medium (T3, 15-28 days). The presence of matrilin-3 with Ad-MSCs in the MAHA scaffold significantly increased the chondrogenic marker and decreased the hypertrophy marker mRNA and protein expression. Furthermore, matrilin-3 significantly modified the expression of TGF-ß2, BMP-2, and BMP-4. Next, we prepared the OA model and transplanted Ad-MSCs primed with matrilin-3, either as a single-cell suspension or in spheroid form. Safranin-O staining and the OA score suggested that the regenerated cartilage morphology in the matrilin-3-primed Ad-MSC spheroids was similar to the positive control. Furthermore, matrilin-3-primed Ad-MSC spheroids prevented subchondral bone sclerosis in the mouse model. Here, we show that matrilin-3 plays a major role in modulating Ad-MSCs' therapeutic effect on cartilage regeneration and hypertrophy suppression.
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Cartilagem Hialina/crescimento & desenvolvimento , Hipertrofia/genética , Células-Tronco Mesenquimais/citologia , Osteoartrite/genética , Animais , Proteína Morfogenética Óssea 2/genética , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Condrócitos/efeitos dos fármacos , Condrogênese/genética , Humanos , Ácido Hialurônico/farmacologia , Hipertrofia/patologia , Hipertrofia/prevenção & controle , Hipertrofia/terapia , Proteínas Matrilinas/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Camundongos , Osteoartrite/terapia , Regeneração/efeitos dos fármacos , Esferoides Celulares/efeitos dos fármacos , Alicerces Teciduais , Fator de Crescimento Transformador beta/genéticaRESUMO
A high level of reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) upregulates pro-inflammatory cytokines and inhibits the osteogenic differentiation of mesenchymal stem cells (MSCs), which are key factors in bone regeneration. Ursodeoxycholic acid (UDCA), a hydrophilic bile acid, has antioxidant and anti-inflammatory activities and also plays beneficial roles in bone regeneration by stimulating the osteogenic differentiation of MSCs while suppressing their adipogenic differentiation. Despite its remarkable capacity for bone regeneration, multiple injections of UDCA induce adverse side effects such as mechanical stress and contamination in bone defects. To fully exploit the beneficial roles of UDCA, a concept polymeric prodrug was developed based on the hypothesis that removal of overproduced H2O2 will potentiate the osteogenic functions of UDCA. In this work, we report bone regenerative nanoparticles (NPs) formulated from a polymeric prodrug of UDCA (PUDCA) with UDCA incorporated in its backbone through H2O2-responsive peroxalate linkages. The PUDCA NPs displayed potent antioxidant and anti-inflammatory activities in MSCs and induced osteogenic rather than adipogenic differentiation of the MSCs. In rat models of bone defect, the PUDCA NPs exhibited significantly better bone regeneration capacity and anti-inflammatory effects than equivalent amounts of UDCA. We anticipate that PUDCA NPs have tremendous translational potential as bone regenerative agents.
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Células-Tronco Mesenquimais , Nanopartículas , Pró-Fármacos , Animais , Ácidos e Sais Biliares , Regeneração Óssea , Diferenciação Celular , Peróxido de Hidrogênio , Osteogênese , RatosRESUMO
Human pluripotent stem cells (hPSCs) are a potent source of clinically relevant mesenchymal stem cells (MSCs) that confer functional and structural benefits in cell therapy and tissue regeneration. Obtaining sufficient numbers of MSCs in a short period of time and enhancing the differentiation potential of MSCs can be offered the potential to improve the regenerative activity of MSCs therapy. In addition, the underlying processes in the isolation and derivation of MSCs from hPSCs are still poorly understood and controlled. To overcome these clinical needs, an efficient and simplified technique on the isolation of MSCs from spontaneously differentiated human embryonic stem cells (hESCs) via integrin α5ß1 (fibronectin (FN) receptor)-to-FN interactions (hESC-FN-MSCs) is successfully developed. It is demonstrated that hESC-FN-MSCs exhibit a typical MSC surface phenotype, cellular morphology, with the whole transcriptome similar to conventional adult MSCs; but show higher proliferative capacity, more efficient trilineage differentiation, enhanced cytokine secretion, and attenuated cellular senescence. In addition, the therapeutic potential and regenerative capacity of the isolated hESC-FN-MSCs are confirmed by in vitro and in vivo multilineage differentiation. This novel method will be useful in the generation of abundant amounts of clinically relevant MSCs for stem cell therapeutics and regenerative medicine.
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BACKGROUND: Chronic low back pain is a prevalent disability, often caused by intervertebral disc (IVD) degeneration. Mesenchymal stem cell (MSC) therapy could be a safe and feasible option for repairing the degenerated disc. However, for successful translation to the clinic, various challenges need to be overcome including unwanted adverse effects due to acidic pH, hypoxia, and limited nutrition. Matrilin-3 is an essential extracellular matrix (ECM) component during cartilage development and ossification and exerts chondrocyte protective effects. METHODS: This study evaluated the effects of matrilin-3-primed adipose-derived MSCs (Ad-MSCs) on the repair of the degenerated disc in vitro and in vivo. We determined the optimal priming concentration and duration and developed an optimal protocol for Ad-MSC spheroid generation. RESULTS: Priming with 10 ng/ml matrilin-3 for 5 days resulted in the highest mRNA expression of type 2 collagen and aggrecan in vitro. Furthermore, Ad-MSC spheroids with a density of 250 cells/microwell showed the increased secretion of favorable growth factors such as transforming growth factor beta (TGF-ß1), TGF-ß2, interleukin-10 (IL-10), granulocyte colony-stimulating factor (G-CSF), and matrix metalloproteinase 1 (MMP1) and decreased secretion of hypertrophic ECM components. In addition, matrilin-3-primed Ad-MSC spheroid implantation was associated with optimal repair in a rabbit model. CONCLUSION: Our results suggest that priming MSCs with matrilin-3 and spheroid formation could be an effective strategy to overcome the challenges associated with the use of MSCs for the treatment of IVD degeneration.
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Degeneração do Disco Intervertebral , Disco Intervertebral , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais , Animais , Degeneração do Disco Intervertebral/terapia , Proteínas Matrilinas/genética , CoelhosRESUMO
Cholesterol and lipid metabolism are associated with osteoarthritis (OA) in human cartilage. High cholesterol levels in OA chondrocytes leads to decreased membrane fluidity and blocks the signaling cascade associated with the expression of chondrogenic genes. It is known that bile acid plays a role in regulating cholesterol homeostasis and the digestion of fats in the human body. Tauroursodeoxycholic acid (TUDCA), as a member of the bile acid family, also aids in the transport of cellular cholesterol. In this study, we hypothesized that TUDCA might be able to promote the restoration of OA cartilage by reducing membrane cholesterol levels in OA chondrocytes and by stimulating the chondrogenic signaling cascade. To assess this hypothesis, we investigated the effects of TUDCA on degenerated chondrocytes isolated from patients with OA. Importantly, treatment with TUDCA at sub-micellar concentrations (2500 µM) significantly increased cell proliferation and Cyclin D1 expression compared with the controls. In addition, the expression of chondrogenic marker genes (SOX9, COL2, and ACAN), proteins (SOX9 and COL2), and glycosaminoglycan (Chondroitin sulfate) was much higher in the TUDCA-treated group compared to the controls. We also found that TUDCA treatment significantly reduced the intracellular cholesterol levels in the chondrocytes and increased membrane fluidity. Furthermore, the stability of TGF receptor 1 and activity of focal adhesion proteins were also increased following TUDCA treatment. Together, these results demonstrated that TUDCA could be used as an alternative treatment for the restoration of OA cartilage.
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Colesterol/metabolismo , Condrócitos/efeitos dos fármacos , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Fluidez de Membrana/efeitos dos fármacos , Osteoartrite/tratamento farmacológico , Ácido Tauroquenodesoxicólico/farmacologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Condrócitos/metabolismo , Condrócitos/patologia , Condrogênese/efeitos dos fármacos , Relação Dose-Resposta a Droga , Adesões Focais/efeitos dos fármacos , Humanos , Osteoartrite/metabolismo , Osteoartrite/patologia , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Transdução de Sinais/efeitos dos fármacos , Ácido Tauroquenodesoxicólico/química , Ácido Tauroquenodesoxicólico/uso terapêuticoRESUMO
The directed differentiation of human adipose-derived stem cells (hASCs) into different cell types has shown great therapeutic potential in treating various diseases. To maximize the therapeutic potentials, researchers have tried manipulating master transcriptional genes that promote efficient differentiation of mesenchymal stem cells (MSCs) such as the MAPK/ERK signaling pathway. Sprouty (SPRY) is a family of proteins that are known to inhibit the MAPK/ERK signaling pathway. Although the role of some SPRY isoforms in MSC differentiation is known, the function of SPRY4 isoform has not been fully elucidated. In the present study, the role of SPRY4 in the multilineage differentiation of hASCs has been elucidated. To investigate the role of SPRY4 in hASC differentiation and tissue regeneration, we performed a transient knockdown of SPRY expression via a small interfering RNA (siSPRY4). Western blot and quantitative polymerase chain reaction results revealed that the treatment of siSPRY4 before induction of differentiation had no significant effect on adipogenic, but reduced chondrogenic, differentiation of hASCs. Interestingly, SPRY4 transient knockdown had a significant effect on the osteogenic differentiation as indicated by the increased messenger RNA (mRNA) and protein expression of osteogenic markers such as alkaline phosphatase (ALP; 2.3-fold) and osteopontin (OPN; 3.5-fold) and increased calcium deposition measured via Alizarin red staining (3.3-fold). Moreover, in vivo tissue regeneration of siSPRY4-treated hASCs in ectopic bone formation and calvarial defect mouse models showed higher bone volume (5.24-fold) and trabecular number (4.59-fold) assessed via histological and microcomputed tomography analyses. We also determined that the enhanced osteogenic differentiation in SPRY4-treated hASCs was due to the induction of ERK1/2 phosphorylation. Taken together, our results suggest that the regulation of SPRY4 through MAPK signaling is a potentially critical aspect on the osteogenic differentiation of hASCs and for bone tissue regeneration, and thus, may be utilized as a potent technique in the development of effective bone therapeutics. Impact Statement This study tried to expand our current understanding of the osteogenic differentiation of mesenchymal stem cells. The transient downregulation of the SPRY4 expression via small interfering RNA (siRNA) showed significant enhancement of the osteogenic differentiation of adipose-derived stem cells via the induction of ERK 1/2 phosphorylation. This suggests the possible mechanism to maximize the potential of stem cell as therapeutics and has a great potential in treating various bone-related diseases.
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Diferenciação Celular , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Osteogênese , Adipogenia , Animais , Proliferação de Células , Condrogênese , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Regulação da Expressão Gênica , Humanos , Sistema de Sinalização das MAP Quinases , Células-Tronco Mesenquimais/enzimologia , Camundongos Endogâmicos BALB C , Camundongos Nus , FosforilaçãoRESUMO
The most efficient method for obtaining high concentration, monodispersed quantum dots is the sol gel method. This manuscript reports room temperature photoluminescence (PL) of lead sulfide (PbS) quantum dots with the effect of a capping agent. Lead sulfide quantum dots were prepared from a waveguide sol gel material with an organic-inorganic capping agent. The quantum dots were made using silica, a photoreactive methylmethacrylate group and a zirconia sol gel solution to give various concentrations of PbS quantum dots. UV-visible absorption spectra linearly increased with increasing PbS quantum dot concentration up to 10 649 ppm, after which increase was nonlinear. At room temperature, strong photoluminescence was achieved with a weak excitation light source, and the intensity increased almost linearly. This indicated that the quantum dots were distributed uniformly in the sol gel matrix. The thermal process slightly reduced the luminescence intensity. A red shift due to band gap energy was observed from 1.55 eV to 1.49 eV.
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Chumbo/química , Pontos Quânticos/química , Sulfetos/química , Luminescência , Tamanho da Partícula , TemperaturaRESUMO
Bioengineering strategies to enhance the natural targeting function of nanocarriers would expand their therapeutic applications. Here, we designed bioengineered stem cell membrane-functionalized nanocarriers (BSMNCs) harboring C-X-C chemokine receptor type 4 (CXCR4) to achieve robust targeting and also to increase their retention time in ischemic tissue. Stem cell membrane coated nanocarrier (SMNCs) or poly (lactic-co-glycolic acid) (PLGA) nanocarriers (PNCs) and BSMNCs were prepared by functionalizing PNCs with human adipose-derived stem cells (hASCs) membranes and hASCs engineered to overexpress CXCR4-receptor, respectively. The functionalization of PNCs with stem cell membranes derived from hASCs significantly enhance the nanocarrier penetration across endothelial cell barrier compare to PNCs. In addition, stem cell membrane functionalization on PNCs also significantly decreased the nanoparticles uptake in J774 (murine) and THP (human) macrophages respectively from 84% to 76%-29% and 24%. Interestingly, BSMNCs showed much higher level of accumulation in ischemic tissue than SMNCs. Systemic retro-orbital injection of BSMNCs loaded with VEGF into mice with hindlimb ischemia resulted substantially enhancement of blood reperfusion, muscle repair, and limb salvage compared to animals treated with SMNCs loaded with similar concentration of VEGF. The reported strategy could be used to create biocompatible and custom-tailored biomimetic nanoparticles with various hybrid functionalities, which may overcome the limitations of current nanoparticle-based therapeutic and imaging platforms.
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Membrana Celular/química , Portadores de Fármacos/química , Membro Posterior/irrigação sanguínea , Isquemia/tratamento farmacológico , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Células-Tronco/química , Fator A de Crescimento do Endotélio Vascular/administração & dosagem , Animais , Bioengenharia , Linhagem Celular , Células Cultivadas , Sistemas de Liberação de Medicamentos , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Nanoestruturas/química , Receptores CXCR4/análise , Transplante de Células-Tronco/métodos , Fator A de Crescimento do Endotélio Vascular/uso terapêuticoRESUMO
Artificial scaffolds made up of various synthetic biodegradable polymers have been reported to have many advantages including cheap manufacturing, easy scale up, high mechanical strength, convenient manipulation, and molding into an unlimited variety of shapes. However, the synthetic biodegradable polymers still have the insufficiency for cartilage regeneration owing to their acidic degradation products. To reduce acidification by degradation of synthetic polymers, we incorporated magnesium hydroxide (MH) nanoparticles into porous polymer scaffold not only to effectively neutralize the acidic hydrolysate but also to minimize the structural disturbance of scaffolds. The neutralization effect of poly(D,L-lactic-co-glycolic acid; PLGA)/MH scaffold was confirmed with the maintenance of neutral pH, contrary to a PLGA scaffold with low pH. Further, the scaffolds were applied to evaluate the chondrogenic differentiation of the human bone marrow mesenchymal stem cells. In in vitro study, the PLGA/MH scaffold enhanced the chondrogenesis markers and reduced the calcification, compared to the PLGA scaffold. Additionally, the PLGA/MH scaffold reduced the release of inflammatory cytokines, compared to the PLGA scaffold, as the cell death decreased. Moreover, the addition of MH reduced necrotic cell death at the early stage of chondrogenic differentiation. Further, the necrotic cell death by the PLGA scaffold was mediated by cleavage of caspase-1, the so-called interleukin 1-converting enzyme, and MH alleviated it as well as nuclear factor kappa B expression. Furthermore, the PLGA/MH scaffold highly supported chondrogenic healing of rat osteochondral defect sites in in vivo study. Therefore, it was suggested that a synthetic polymer scaffold containing MH could be a novel healing tool to support cartilage regeneration and further treatment of orthopedic patients. STATEMENT OF SIGNIFICANCE: Synthetic polymer scaffolds have been widely utilized for tissue regeneration. However, they have a disadvantage of releasing acidic products through degradation. This paper demonstrated a novel type of synthetic polymer scaffold with pH-neutralizing ceramic nanoparticles composed of magnesium hydroxide for cartilage regeneration. This polymer showed pH-neutralization property during polymer degradation and significant enhancement of chondrogenic differentiation of mesenchymal stem cells. It reduced not only chondrogenic calcification but also release of proinflammatory cytokines. Moreover, it has an inhibitory effect on necrotic cell death, particularly caspase-1-mediated necrotic cell death (pyroptosis). In in vivo study, it showed higher healing rate of the damaged cartilage in a rat osteochondral defect model. We expected that this novel type of scaffold can be effectively applied to support cartilage regeneration and further treatment of orthopedic patients.
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Condrogênese/efeitos dos fármacos , Hidróxido de Magnésio , Células-Tronco Mesenquimais/metabolismo , Nanopartículas , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Alicerces Teciduais/química , Humanos , Hidróxido de Magnésio/química , Hidróxido de Magnésio/farmacologia , Células-Tronco Mesenquimais/citologia , Nanopartículas/química , Nanopartículas/uso terapêutico , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/farmacologiaRESUMO
BACKGROUND: Early stage osteoarthritis (OA) is clinically asymptomatic due to the avascular and the aneural nature of the cartilage tissue. Nevertheless, early detection of cartilage tissue is critical in order to impede the progression of OA. Hence, in order to develop effective preventive therapy for OA, diagnosis in the early stages is necessary. METHODS: To achieve this goal, we have developed targeted, fluorescent nanosomes conjugated with monoclonal anti-type II collagen antibodies (MabCII) for diagnosis of early OA. The MabCII-coated nanosomes (targeted-nanosomes) bind to the damaged cartilage explants in vitro and in vivo in an OA mouse model that mimics early stage OA. The OA mouse model was induced by destabilization of the medial meniscus (DMM) in 9-10 weeks old C57Bl/6 mice. RESULTS: The targeted-nanosomes enhanced the binding specificity to the cartilage tissue according to the severity of damage. CONCLUSION: We show that MabCII-nanosomes can precisely detect early stage OA in the DMM mouse model. Thus, MabCII-nanosomes have the potential to be used as a non-invasive method for diagnosing the early osteoarthritic lesions.
Assuntos
Meniscos Tibiais/diagnóstico por imagem , Nanopartículas/química , Nanopartículas/uso terapêutico , Osteoartrite/diagnóstico por imagem , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/metabolismo , Colágeno Tipo II/imunologia , Colágeno Tipo II/metabolismo , Modelos Animais de Doenças , Corantes Fluorescentes/química , Corantes Fluorescentes/uso terapêutico , Masculino , Meniscos Tibiais/patologia , Camundongos Endogâmicos C57BL , Imagem Óptica/métodos , Osteoartrite/metabolismo , Osteoartrite/patologiaRESUMO
It is very useful to evaluate the content and 3D distribution of extracellular matrix non-destructively in tissue engineering. This study evaluated the feasibility of using micro-computed tomography (µCT) with Hexabrix to measure quantitatively sulfated glycosaminoglycans (GAGs) of engineered cartilage. Rabbit chondrocytes at passage 2 were used to produce artificial cartilages in polyglycolic acid scaffolds in vitro. Engineered cartilages were incubated with Hexabrix 320 for 20 min and analyzed via µCT scanning. The number of voxels in the 2D and 3D scanning images were counted to estimate the amount of sulfated GAGs. The optimal threshold value for quantification was determined by regression analysis. The 2D µCT images of an engineered cartilage showed positive correlation with the histological image of Safranin-O staining. Quantitative data obtained with the 3D µCT images of 14 engineered cartilages showed strong correlation with sulfated GAGs contents obtained by biochemical analysis (R2 = 0.883, p < 0.001). Repeated exposure of engineered cartilages to Hexabrix 320 and µCT scanning did not significantly affect cell viability, total DNA content, or the total content of sulfated GAGs. We conclude that µCT imaging using Hexabrix 320 provides high spatial resolution and sensitivity to assess the content and 3D distribution of sulfated GAGs in engineered cartilages. It is expected to be a valuable tool to evaluate the quality of engineered cartilage for commercial development in the future.
RESUMO
Matrilin-3 is an essential extracellular matrix component present only in cartilaginous tissues. Matrilin-3 exerts chondroprotective effects by regulating an anti-inflammatory function and extracellular matrix components. We hypothesized that the codelivery of matrilin-3 with infrapatellar adipose-tissue-derived mesenchymal stem cells (Ad-MSCs) may enhance articular cartilage regeneration. Matrilin-3 treatment of Ad-MSCs in serum-free media induced collagen II and aggrecan expression, and matrilin-3 in chondrogenic media also enhanced in vitro chondrogenic differentiation. Next, the in vivo effect of matrilin-3 codelivery with Ad-MSCs on cartilage regeneration was assessed in an osteochondral defect model in Sprague Dawley rats: Ad-MSCs and hyaluronic acid were implanted at the defect site with or without matrilin-3 (140, 280, and 700 ng). Safranin O staining revealed that matrilin-3 (140 and 280 ng) treatment significantly improved cartilage regeneration and glycosaminoglycan accumulation. In the animals treated with 140-ng matrilin-3, in particular, the defect site exhibited complete integration with surrounding tissue and a smooth glistening surface. The International Cartilage Repair Society macroscopic and O'Driscoll microscopic scores for regenerated cartilage were furthermore shown to be considerably higher for this group (matrilin-3; 140 ng) compared with the other groups. Furthermore, the defects treated with 140-ng matrilin-3 revealed significant hyaline-like cartilage regeneration in the osteochondral defect model; in contrast, the defects treated with 700-ng matrilin-3 exhibited drastically reduced cartilage regeneration with mixed hyaline-fibrocartilage morphology. Codelivery of matrilin-3 with Ad-MSCs significantly influenced articular cartilage regeneration, supporting the potential use of this tissue-specific protein for a cartilage-targeted stem cell therapy.