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Purpose: Osteoarthritis (OA) is a prevalent cause of disability in older adults. Identifying diagnostic markers for OA is essential for elucidating its mechanisms and facilitating early diagnosis. Methods: We analyzed 53 synovial tissue samples (n = 30 for OA, n = 23 for the control group) from two datasets in the Gene Express Omnibus (GEO) database. We identified differentially expressed genes (DEGs) between the groups and applied dimensionality reduction using six machine learning algorithms to pinpoint characteristic genes (key genes). We classified the OA samples into subtypes based on these key genes and explored the differences in biological functions and immune characteristics among subtypes, as well as the roles of the key genes. Additionally, we constructed a protein-protein interaction network to predict small molecules that target these genes. Further, we accessed synovial tissue sample data from the single-cell RNA dataset GSE152805, categorized the cells into various types, and examined variations in gene expression and their correlation with OA progression. Validation of key gene expression was conducted in cellular experiments using the qPCR method. Results: Four genes AGMAT, MAP3K8, PER1, and XIST, were identified as characteristic genes of OA. All can independently predict the occurrence of OA. With these genes, the OA samples can be clustered into two subtypes, which showed significant differences in functional pathways and immune infiltration. Eight cell types were obtained by analyzing the single-cell RNA data, with synovial intimal fibroblasts (SIF) accounting for the highest proportion in each sample. The key genes were found over-expressed in SIF and significantly correlated with OA progression and the content of immune cells (ICs). We validated the relative levels of key genes in OA and normal cartilage tissue cells, which showed an expression trend consistency with the bioinformatics result except for XIST. Conclusion: Four genes, AGMAT, MAP3K8, PER1, and XIST are closely related to the progression of OA, and play as diagnostic and predictive markers in early OA.
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Osteogenesis is tightly coupled with angiogenesis spatiotemporally. Previous studies have demonstrated that type H blood vessel formed by endothelial cells with high expression of CD31 and Emcn (CD31hi Emcnhi ECs) play a crucial role in bone regeneration. The mechanism of the molecular communication around CD31hi Emcnhi ECs and bone mesenchymal stem cells (BMSCs) in the osteogenic microenvironment is unclear. This study indicates that exosomes from bone mesenchymal stem cells with 7 days osteogenic differentiation (7D-BMSCs-exo) may promote CD31hi Emcnhi ECs angiogenesis, which was verified by tube formation assay, qRT-PCR, Western blot, immunofluorescence staining and µCT assays etc. in vitro and in vivo. Furthermore, by exosomal miRNA microarray and WGCNA assays, we identified downregulated miR-150-5p as the most relative hub gene coupling osteogenic differentiation and type H blood vessel angiogenesis. With bioinformatics assays, dual luciferase reporter experiments, qRT-PCR and Western blot assays, SOX2(SRY-Box Transcription Factor 2) was confirmed as a novel downstream target gene of miR-150-5p in exosomes, which might be a pivotal mechanism regulating CD31hi Emcnhi ECs formation. Additionally, JC-1 immunofluorescence staining, Western blot and seahorse assay results showed that the overexpression of SOX2 could shift metabolic reprogramming from oxidative phosphorylation (OXPHOS) to glycolysis to enhance the CD31hi Emcnhi ECs formation. The PI3k/Akt signaling pathway might play a key role in this process. In summary, BMSCs in osteogenic differentiation might secrete exosomes with low miR-150-5p expression to induce type H blood vessel formation by mediating SOX2 overexpression in ECs. These findings might reveal a molecular mechanism of osteogenesis coupled with type H blood vessel angiogenesis in the osteogenic microenvironment and provide a new therapeutic target or cell-free remedy for osteogenesis impaired diseases.
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Diferenciación Celular , Células Endoteliales , Exosomas , Células Madre Mesenquimatosas , MicroARNs , Neovascularización Fisiológica , Osteogénesis , MicroARNs/genética , MicroARNs/metabolismo , Exosomas/metabolismo , Osteogénesis/genética , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Diferenciación Celular/genética , Neovascularización Fisiológica/genética , Animales , Células Endoteliales/metabolismo , Células Endoteliales/citología , Ratones , Humanos , Células Cultivadas , Transducción de Señal , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/metabolismo , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/genética , Factores de Transcripción SOXB1/metabolismo , Factores de Transcripción SOXB1/genética , Reprogramación Metabólica , AngiogénesisRESUMEN
Myoblast is a kind of activated muscle stem cell. Its biological activities, such as proliferation, migration, differentiation, and fusion, play a crucial role in maintaining the integrity of the skeletal muscle system. These activities of myoblasts can be significantly influenced by the extracellular matrix. Collagen, being a principal constituent of the extracellular matrix, substantially influences these biological activities. In skeletal muscle, collagen I and III are two kinds of primary collagen types. Their influence on myoblasts and the difference between them remain ambiguous. The purpose of this study is to discover the influence of collagen I and III on biological function of myoblasts and compare their differences. We used C2C12 cell line and primary myoblasts to discover the effect of collagen I and III on proliferation, migration and differentiation of myoblasts and then performed the transcriptome sequencing and analysis. The results showed that both collagen I and III enhanced the proliferation of myoblasts, with no statistical difference between them. Similarly, collagen I and III enhanced the migration of myoblasts, with collagen I was more pronounced in Transwell assay. On the contrary, collagen I and III inhibited myoblasts differentiation, with collagen III was more pronounced at gene expression level. The transcriptome sequencing identified DEGs and enrichment analysis elucidated different terms between Type I and III collagen. Collectively, our research preliminarily elucidated the influence of collagen I and III on myoblasts and their difference and provided the preliminary experimental foundation for subsequent research.
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Diferenciación Celular , Movimiento Celular , Proliferación Celular , Colágeno Tipo I , Mioblastos , Diferenciación Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Mioblastos/citología , Mioblastos/metabolismo , Animales , Ratones , Colágeno Tipo I/metabolismo , Colágeno Tipo III/metabolismo , Colágeno Tipo III/genética , Línea CelularRESUMEN
AIM: This study aimed to evaluate the miR-223-5p expression in patients with spinal cord injury (SCI) and to determine its role in the pathogenesis of SCI. METHODS: The serum miR-223-5p levels were analyzed using quantitative real-time polymerase chain reaction. The diagnostic accuracy of miR-223-5p was evaluated using the receiving operating characteristic curves. LPS-induced PC12 cells were established as an in vitro inflammatory cell model. Cell apoptosis, inflammation and oxidative stress were examined. The SCI rat model was constructed to evaluate the effects of miR-223-5p on inflammatory response and motor function in rats. RESULTS: MiR-223-5p expression was upregulated in SCI patients. MiR-223-5p expression in the complete SCI group was significantly higher than that in incomplete SCI group. ROC analysis showed that miR-223-5p can distinguish SCI patients from healthy volunteers. In vitro experiments demonstrated that LPS upregulated apoptosis and inflammation in PC12 cells. Treatment with miR-223-5p inhibitor alleviated the changes in LPS-induced PC12 cells . Inhibition of miR-223-5p can alleviate the activation of inflammatory response and the effects of SCI on the motor function in rats. CONCLUSIONS: MiR-223-5p is a potential diagnostic marker for SCI, and it can promote the SCI progression by regulating nerve cell survival, inflammation, and oxidative stress.
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Apoptosis , Inflamación , MicroARNs , Estrés Oxidativo , Traumatismos de la Médula Espinal , Traumatismos de la Médula Espinal/metabolismo , Traumatismos de la Médula Espinal/genética , MicroARNs/genética , Animales , Ratas , Humanos , Células PC12 , Masculino , Inflamación/genética , Apoptosis/genética , Femenino , Adulto , Persona de Mediana Edad , Modelos Animales de Enfermedad , Ratas Sprague-Dawley , Biomarcadores , LipopolisacáridosRESUMEN
The M1 polarization of microglia, followed by the production of pro-inflammatory mediators, hinders functional recovery after spinal cord injury (SCI). Our previous study has illuminated that specificity protein 1 (Sp1) expression is increased following SCI, whereas the function and regulatory mechanism of Sp1 during M1 polarization of microglia following SCI remain unknown. RNA binding protein, HuR, has been shown to be up-regulated in the injured spinal cord through analysis of the GEO database. Further investigation using Chip-Atlas data suggests a binding between Sp1 and HuR. Emerging evidence indicates that HuR plays a pivotal role in neuroinflammation after SCI. In this research, Sp1 and HuR levels in mice with SCI and BV2 cells treated with lipopolysaccharide (LPS) was determined by using quantitative real-time polymerase chain reaction and Western blotting techniques. A series of in vitro assays were performed to investigate the function of Sp1 during M1 polarization of microglia. The association between Sp1 and its target gene HuR was confirmed through gene transfection and luciferase reporter assay. Enhanced expression of HuR was observed in both SCI mice and LPS-treated BV2 cells, while Sp1 knockdown restrained M1 polarization of microglia and its associated inflammation by inhibiting the NF-κB signaling pathway. Silencing Sp1 also suppressed microglia activation and its mediated inflammatory response, which could be reversed by overexpression of HuR. In conclusion, silencing Sp1 restrains M1 polarization of microglia through the HuR/NF-κB axis, leading to neuroprotection, and thus promotes functional restoration following SCI.
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FN-kappa B , Factor de Transcripción Sp1 , Traumatismos de la Médula Espinal , Animales , Ratones , Lipopolisacáridos/farmacología , Lipopolisacáridos/metabolismo , Microglía/metabolismo , FN-kappa B/metabolismo , Transducción de Señal , Médula Espinal/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Factor de Transcripción Sp1/genética , Factor de Transcripción Sp1/metabolismoRESUMEN
The pathogenesis of osteoarthritis (OA) is still unclear, leading to the lack of targeted treatment. We aimed to probe into the effect of apolipoprotein D (APOD), the key gene from our previous study through bioinformatics analysis, on fibroblast-like synoviocyte (FLS) and chondrocytes in vitro to confirm its potential roles on the delay of OA progression. Primary FLS and chondrocytes were extracted from synovium and cartilage of OA patients and stimulated with interleukin 1ß (IL-1ß) in vitro. After APOD intervention, viability and proliferation of FLS and chondrocytes were detected. Subsequently, the inflammatory factors of the two cells were detected by quantitative reverse-transcription polymerase chain reaction, enzyme-linked immunosorbent assay, and western blot, and the apoptosis and autophagy-related substances were determined at the same time. Finally, the oxidation level in FLS and chondrocytes were detected. APOD reversed the change of gene expression stimulated by IL-1ß in FLS and chondrocytes. APOD alleviated the proliferation of FLS while promoted proliferation of chondrocytes, and reduced the expression of inflammatory factors. Moreover, APOD promoted apoptosis of FLS and autography of chondrocytes, while reduced apoptosis of chondrocytes. Finally, decrease level of reactive oxygen species (ROS) in both cells were observed after APOD intervention, as well as the increased expression of antioxidant-related genes. APOD had effects on the proliferation of FLS and chondrocytes through apoptosis and autography as well as the reduction of oxidative stress, delaying the progress of OA.
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MicroARNs , Osteoartritis , Sinoviocitos , Humanos , Sinoviocitos/metabolismo , Condrocitos/metabolismo , Apolipoproteínas D/metabolismo , Osteoartritis/metabolismo , Interleucina-1beta/metabolismo , Fibroblastos/patología , Apoptosis , MicroARNs/metabolismoRESUMEN
CONTEXT: As a common disease in the elderly, osteoporosis clearly increases the risk of fractures, leading to higher mortality, but the current markers to estimate the risk of fractures are limited. MicroRNA-21 (miR-21) may play an important role in osteoporosis, but the link of this biomarker with fractures was undetermined. OBJECTIVES: We aimed to investigate the association between miR-21 levels and the presence of fragility fractures. METHODS: A total of 200 patients were recruited and miR-21 was collected from baseline serum. The correlation between miR-21 and the fracture risk assessment tool (FRAX) score was analyzed. The incidence of fragility fractures was presented by Kaplan-Meier analysis, and Cox regression analysis was utilized to evaluate risk factors. The diagnostic value of miR-21 was conducted by the area under curve (AUC). RESULTS: The FRAX score was significantly associated with miR-21 level (p<0.001). According to the 50th percentile of miR-21 content in the overall distribution, the cumulative incidence of fragility fractures was significantly higher in patients with higher miR-21 levels than those with lower levels (75.4, 95â¯% CI: 69.0-81.8 vs. 59.2, 95â¯% CI: 42.1-76.3, p<0.001). The results of the Cox regression analysis showed that the miR-21 level was an independent risk factor linked to the incidence of fracture (p=0.005). The optimal cut-off value of the miR-21 was 6.08, and the AUC for predicting fracture was 0.718 (95â¯% CI, 0.645-0.790). CONCLUSIONS: This study showed that miR-21 has optimal diagnostic performance in the discrimination of fragility fracture, and the circulating miR-21 level in predicting the risk of fragility fracture may have a certain value.
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MicroARN Circulante , MicroARNs , Osteoporosis , Fracturas Osteoporóticas , Humanos , Anciano , Fracturas Osteoporóticas/epidemiología , Fracturas Osteoporóticas/etiología , Densidad Ósea , Medición de Riesgo/métodos , Osteoporosis/complicacionesRESUMEN
The balance among different CD4+ T cell subsets is crucial for repairing the injured spinal cord. Dendritic cell (DC)-derived small extracellular vesicles (DsEVs) effectively activate T-cell immunity. Altered peptide ligands (APLs), derived from myelin basic protein (MBP), have been shown to affect CD4+ T cell subsets and reduce neuroinflammation levels. However, the application of APLs is challenging because of their poor stability and associated side effects. Herein, it is demonstrate that DsEVs can act as carriers for APL MBP87-99 A91 (A91-DsEVs) to induce the activation of 2 helper T (Th2) and regulatory T (Treg) cells for spinal cord injury (SCI) in mice. These stimulated CD4+ T cells can efficiently "home" to the lesion area and establish a beneficial microenvironment through inducing the activation of M2 macrophages/microglia, inhibiting the expression of inflammatory cytokines, and increasing the release of neurotrophic factors. The microenvironment mediated by A91-DsEVs may enhance axon regrowth, protect neurons, and promote remyelination, which may support the recovery of motor function in the SCI model mice. In conclusion, using A91-DsEVs as a therapeutic vaccine may help induce neuroprotective immunity in the treatment of SCI.
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Vesículas Extracelulares , Traumatismos de la Médula Espinal , Vacunas , Ratas , Ratones , Animales , Ratas Sprague-Dawley , Ligandos , Traumatismos de la Médula Espinal/terapia , Traumatismos de la Médula Espinal/metabolismo , Vacunas/farmacología , Vacunas/uso terapéutico , Péptidos , Linfocitos T Reguladores , Vesículas Extracelulares/metabolismo , Células DendríticasRESUMEN
Chondrocyte ferroptosis constitutes a major cause of the development of osteoarthritis (OA). Bone marrow mesenchymal stem cell-derived exosomes (BMSC-Exos) have a protective role against ferroptosis in various diseases. Hence, we aimed to determine whether BMSC-Exos alleviated chondrocyte ferroptosis and its effect on OA, and to dissect out the possible mechanisms. An OA rat chondrocyte model was established by interleukin-1ß (IL-1ß) exposure, and treated with BMSC-Exos/ferroptosis inhibitor Ferrostatin-1. Cell viability/ferroptosis-related index levels [reactive oxygen species (ROS)/malondialdehyde (MDA)/glutathione (GSH)]/cell death/ACSL4 mRNA and protein levels and METTL3 levels were assessed by MTT/kits/immunohistochemical method and TUNEL staining/RT-qPCR and Western blot. METTL3/ACSL4 were overexpressed in rat chondrocytes to evaluate their role in BMSC-Exo-produced repression on chondrocyte ferroptosis. Bioinformatics website predicted the presence of m6A modification sites on ACSL4 mRNA, with the m6A level enriched on it assessed by MeRIP/RT-qPCR. ACSL4 mRNA stability was detected by actinomycin D assay. A surgical destabilized medial meniscus rat OA model was also established, followed by injection with BMSC-Exos to verify their function. IL-1ß stimulation in rat chondrocytes inhibited cell viability, elevated Fe2+/ROS/MDA levels, declined GSH levels and increased TUNEL positive cell number and ACSL4 level, which were neutralized by BMSC-Exos. BMSC-Exos limited chondrocyte ferroptosis by down-regulating METTL3, with the effect abrogated by METTL3 overexpression. METTL3 regulated the m6A modification of ACSL4 mRNA, increasing ACSL4 mRNA stability and ACSL4 expression. BMSC-Exos reduced chondrocyte ferroptosis and prevented OA progression via disruption of the METTL3-m6A-ACSL4 axis. BMSC-Exos might exert a chondroprotective effect by attenuating chondrocyte ferroptosis and alleviate OA progression.
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Exosomas , Células Madre Mesenquimatosas , MicroARNs , Osteoartritis , Ratas , Animales , Exosomas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Células Madre Mesenquimatosas/metabolismo , ARN Mensajero/metabolismo , MicroARNs/metabolismoRESUMEN
Background: Osteoarthritis (OA) progression involves multiple factors, including cartilage erosion as the basic pathological mechanism of degeneration, and is closely related to chondrocyte apoptosis. To analyze the correlation between apoptosis and OA development, we selected apoptosis genes from the differentially expressed genes (DEGs) between OA and normal samples from the Gene Expression Omnibus (GEO) database, used lasso regression analysis to identify characteristic genes, and performed consensus cluster analysis to further explore the pathogenesis of this disease. Methods: The Gene expression profile datasets of OA samples, GSE12021 and GSE55235, were downloaded from GEO. The datasets were combined and analyzed for DEGs. Apoptosis-related genes (ARGs) were collected from the GeneCards database and intersected with DEGs for apoptosis-related DEGs (ARDEGs). Least absolute shrinkage and selection operator (LASSO) regression analysis was performed to obtain characteristic genes, and a nomogram was constructed based on these genes. A consensus cluster analysis was performed to divide the patients into clusters. The immune characteristics, functional enrichment, and immune infiltration statuses of the clusters were compared. In addition, a protein-protein interaction network of mRNA drugs, mRNA-transcription factors (TFs), and mRNA-miRNAs was constructed. Results: A total of 95 DEGs were identified, of which 47 were upregulated and 48 were downregulated, and 31 hub genes were selected as ARDEGs. LASSO regression analysis revealed nine characteristic genes: growth differentiation factor 15 (GDF15), NAMPT, TLR7, CXCL2, KLF2, REV3L, KLF9, THBD, and MTHFD2. Clusters A and B were identified, and neutrophil activation and neutrophil activation involved in the immune response were highly enriched in Cluster B, whereas protein repair and purine salvage signal pathways were enriched in Cluster A. The number of activated natural killer cells in Cluster B was significantly higher than that in Cluster A. GDF15 and KLF9 interacted with 193 and 32 TFs, respectively, and CXCL2 and REV3L interacted with 48 and 82 miRNAs, respectively. Conclusion: ARGs could predict the occurrence of OA and may be related to different degrees of OA progression.
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Apoptosis , MicroARNs , Humanos , Consenso , Apoptosis/genética , MicroARNs/genética , Análisis por Conglomerados , ARN Mensajero , ADN Polimerasa Dirigida por ADN , Proteínas de Unión al ADN , Factores de Transcripción de Tipo KruppelRESUMEN
Tendinopathy is a disease with surging prevalence. Lacking understanding of molecular mechanisms impedes the development of therapeutic approaches and agents. Lysine lactylation (Kla) is a newly discovered post-translational modification related to glycolysis. It has long been noted that manipulation of glycolysis metabolism could affect tendon cell function, tendon homeostasis, and healing process of tendon. However, protein lactylation sites in tendinopathy remain unexplored. Here, we conducted the first proteome-wide Kla analysis in tendon samples harvested from patients with rotator cuff tendinopathy (RCT), which identified 872 Kla sites across 284 proteins. Compared with normal counterparts, 136 Kla sites on 77 proteins were identified as upregulated in the pathological tendon, while 56 sites on 32 proteins were downregulated. Function enrichment analysis demonstrated that the majority of proteins with upregulated Kla levels functioned in organization of the tendon matrix and cholesterol metabolism, accompanied by lower expression levels which meant impaired cholesterol metabolism and degeneration of the tendon matrix, indicating potential cross-talk between protein lactylation and expression levels. At last, by western blotting and immunofluorescence, we verified the correlation between high lactylation and the downregulation of matrix and cholesterol-related proteins including BGN, MYL3, TPM3, and APOC3. ProteomeXchange: PXD033146.
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Manguito de los Rotadores , Tendinopatía , Humanos , Manguito de los Rotadores/metabolismo , Manguito de los Rotadores/patología , Proteínas/metabolismo , Tendones/metabolismo , Tendones/patología , Lisina/metabolismo , Tendinopatía/genética , Tendinopatía/metabolismo , Tendinopatía/patologíaRESUMEN
Purpose: Osteoarthritis (OA) is a common degenerative disease, which still lacks specific therapeutic drugs. Synovitis is one of the most important pathological process in OA. Therefore, we aim to identify and analyze the hub genes and their related networks of OA synovium with bioinformatics tools to provide theoretical basis for potential drugs. Materials and methods: Two datasets were obtained from GEO. DEGs and hub genes of OA synovial tissue were screened through Gene Ontology (GO) annotation, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment as well as protein-protein interaction (PPI) network analysis. Subsequently, the correlation between expression of hub genes and ferroptosis or pyroptosis was analyzed. CeRNA regulatory network was constructed after predicting the upstream miRNAs and lncRNAs. The validation of hub genes was undertook through RT-qPCR and ELISA. Finally, potential drugs targeting pathways and hub genes were identified, followed by the validation of the effect of two potential drugs on OA. Results: A total of 161 commom DEGs were obtained, of which 8 genes were finally identified as hub genes through GO and KEGG enrichment analysis as well as PPI network analysis. Eight genes related to ferroptosis and pyroptosis respectively were significantly correlated to the expression of hub genes. 24 miRNAs and 69 lncRNAs were identified to construct the ceRNA regulatory network. The validation of EGR1, JUN, MYC, FOSL1, and FOSL2 met the trend of bioinformatics analysis. Etanercept and Iguratimod reduced the secretion of MMP-13 and ADAMTS5 of fibroblast-like synoviocyte. Conclusion: EGR1, JUN, MYC, FOSL1, and FOSL2 were identified as hub genes in the development of OA after series of bioinformatics analysis and validation. Etanercept and Iguratimod seemed to have opportunities to be novel drugs for OA.
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OBJECTIVE: Osteoarthritis (OA) is a common disease with complex and unclear pathogenesis. Ferroptosis is a new cell death mode, which is proved to be involved in different kinds of disease. We hypothesized that ferroptosis contributes to the progress of human OA. DESIGN: Chondrocytes were extracted from waste cartilage of total knee arthroplasty, and stimulated with interleukin-1ß (IL-1ß). Then, we detected the morphology, proliferation, and viability, and levels of Fe3+, glutathione (GSH), reactive oxygen species (ROS), malondialdehyde (MDA), and 5 proteins related to ferroptosis with or without intervention of ferrostatin-1 (Fer-1). In addition, we compared the effect of Fer-1 and liproxstatin-1 (Lip-1) on ferroptosis and the protection of chondrocytes by detecting several markers of both ferroptosis and OA. RESULTS: After stimulation of IL-1ß, there were significant changes on the shape of chondrocyte, with lower viability and proliferation. There was accumulation of intracellular Fe3+, GSH, ROS, and MDA, with the changes of expression of 5 ferroptosis-related proteins. With the contribution of Fer-1, results above were reversed. Moreover, there was no significant difference in GPX4 and ACSL4 between Fer-1 and Lip-1 group. However, the expression of COLX, ADAMTS5, and MMP-13 are lower after the treatment of Fer-1 compared with Lip-1. CONCLUSIONS: Ferroptosis plays an important role in human OA chondrocytes, which can be reversed by Fer-1, illustrating that inhibitor of ferroptosis may be a potential treatment of OA. Moreover, Lip-1 and Fer-1 can both alleviate the level of ferroptosis in OA chondrocytes, but Fer-1 had a more protective effect.
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Ferroptosis , Osteoartritis , Humanos , Condrocitos/metabolismo , Interleucina-1beta/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Especies Reactivas de Oxígeno/farmacología , Células Cultivadas , Osteoartritis/metabolismoRESUMEN
Following spinal cord injury (SCI), astrocyte activation and proliferation result in the development of glial scars, which impede axonal growth and neurological recovery. Dysregulation of microRNAs (miRNAs) during SCI results in altered expression of downstream genes. Our previous study has revealed that miR-135a-5p regulates neuronal apoptosis and axonal growth by targeting specificity protein 1 (SP1). This study attempted to investigate whether the miR-135a-5p/SP1 axis has regulatory effect on astrocytes. Herein, lipopolysaccharide (LPS) reduced miR-135a-5p expression in astrocytes. miR-135a-5p overexpression in astrocytes resulted in a decrease in CyclinD1, MMP9, GFAP, and vimentin proteins, and thus attenuated LPS-induced proliferation and migration of astrocytes. Moreover, miR-135a-5p overexpression decreased astrocyte size and the total quantity of cell protrusions, suggesting a role for miR-135a-5p in regulating astrocyte morphology. SP1 silencing also decreased astrocyte proliferation and migration by LPS. SP1 silencing could significantly reverse the promoting effect of miR-135a-5p inhibition on astrocyte proliferation and migration. In summary, the miR-135a-5p/SP1 axis regulates astrocyte proliferation and migration after SCI. This finding benefits for the development of novel ways in treating SCI effectively.
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MicroARNs , Traumatismos de la Médula Espinal , Humanos , Astrocitos/metabolismo , Lipopolisacáridos/farmacología , Lipopolisacáridos/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Apoptosis , Proliferación Celular , Traumatismos de la Médula Espinal/metabolismo , Factor de Transcripción Sp1/genética , Factor de Transcripción Sp1/metabolismoRESUMEN
BACKGROUND: Osteosarcoma is a common primary bone malignancy prevalent among adolescents and young adults. PTEN-induced kinase 1 (PINK1) regulates Parkinson's disease, but its role in cancers is unknown. OBJECTIVE: This study was designed to analyze the mechanism by which PINK1 affects osteosarcoma using bioinformatics and cell experiments. MATERIALS AND METHODS: The gene expression profiles were downloaded from the TARGET database. Several online databases were used to analyze the expression and proteinâprotein interaction networks. CCK-8 cell viability assays and cisplatin treatment were used to assess cell activity with or without cisplatin treatment. Acridine orange/ethidium bromide (AO/EB) fluorescence staining was used to calculate the percentage of apoptotic cells. RESULTS: Through bioinformatics analysis, we found that high expression of PINK1 was associated with poor prognosis in patients with osteosarcoma, and PINK1 inhibited apoptosis and promoted proliferation pathways. Next, we found that both PINK1 mRNA and protein levels were upregulated in osteosarcoma tissues. Additionally, we found that PTEN was reduced, while FOXO3a was markedly increased in osteosarcoma, suggesting that FOXO3a and not PTEN induced the overexpression of PINK1. CCK-8 and clonogenic assays showed that the knockdown of PINK1 decreased the growth of U2OS osteosarcoma cells. Ki67 immunofluorescence staining revealed that reduced cell proliferation in U2OS cells resulted in the depletion of PINK1. In addition, our AO/EB staining results indicated that the knockdown of PINK1 resulted in an increase in apoptotic cells and increased the levels of cleaved caspase-3. Furthermore, our experiments revealed that cisplatin promotes OS cell apoptosis by downregulating PINK1. CONCLUSION: Collectively, our findings demonstrate that PINK1 is crucially involved in osteosarcoma and suggests that it can promote the apoptosis of OS cells as the downstream target gene of cisplatin.
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Neoplasias Óseas , Osteosarcoma , Proteínas Quinasas , Adolescente , Humanos , Adulto Joven , Apoptosis/genética , Neoplasias Óseas/tratamiento farmacológico , Neoplasias Óseas/genética , Neoplasias Óseas/metabolismo , Línea Celular Tumoral , Proliferación Celular/genética , Cisplatino/farmacología , Regulación Neoplásica de la Expresión Génica , Osteosarcoma/tratamiento farmacológico , Osteosarcoma/genética , Osteosarcoma/metabolismo , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismoRESUMEN
OBJECTIVES: Keshin-Beck disease (KBD) is a particular type of osteoarthritis that affects many joints. However, the deformity of atlantoaxial joint has been rarely reported in KBD, and therefore its clinical and radiograph features have not been identified. METHODS: We reviewed data in 14 patients who were diagnosed with atlantoaxial dislocation (AAD) in KBD at our institution. The demographic data, clinical history, imaging data, operative data, and Japanese Orthopaedic Association score were collected for evaluation. RESULTS: The mean age at presentation was 50 ± 1.7 years old. The most common features of AAD in KBD were the osteoarthritis, characterized by hypertrophic dens and anterior arch of the atlas. The average inner anteroposterior diameter (IAPD) of C1 was 28 ± 3.5 mm and the average spinal canal diameter was 14 ± 3.3 mm, which were respectively lower than the control level. Five patients had severe C1 stenosis (IAPD < 26mm). Separated odontoid process, like os odontoideum, was seen 9 patients. The tip of dens fused to C1 was observed in 4 patients; 12 patients had high-riding vertebral artery; and 5 patients had severe C1 stenosis, and they underwent C1 laminectomy with C1-C2 interarticular fusion or occipital-cervical fusion. All the patients displayed neurologic improvement after surgery. CONCLUSIONS: The atlantoaxial level could be affected by KBD, which may lead to typical abnormalities and cause AAD. A C1 laminectomy with an C1-C2 interarticular fusion or occipital-cervical fusion is recommended for the patient with severe stenosis.
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Articulación Atlantoaxoidea , Luxaciones Articulares , Enfermedad de Kashin-Beck , Osteoartritis , Enfermedades de la Columna Vertebral , Fusión Vertebral , Espondiloartropatías , Humanos , Persona de Mediana Edad , Constricción Patológica , Luxaciones Articulares/cirugía , Radiografía , Fusión Vertebral/métodos , Articulación Atlantoaxoidea/cirugíaRESUMEN
The microenvironment in the healing process of large bone defects requires suitable conditions to promote osteogenesis and angiogenesis. Coaxial electrospinning is a mature method in bone tissue engineering (BTE) and allows functional modification. Appropriate modification methods can be used to improve the bioactivity of scaffolds for BTE. In this study, coaxial electrospinning with QK peptide (a Vascular endothelial growth factor mimetic peptide) and BMP-2 peptide-DFO (BD) was performed to produce double-modified PQBD scaffolds with vascularizing and osteogenic features. The morphology of coaxially electrospun scaffolds was verified by scanning electron microscopy (SEM) and transmission electron microscopy. Laser scanning confocal microscopy and Fourier transform infrared spectroscopy confirmed that BD covalently bound to the surface of the P and PQ scaffolds. In vitro, the PQBD scaffold promoted the adhesion and proliferation of bone marrow stromal cells (BMSCs). Both QK peptide and BD showed sustainable release and preservation of biological activity, enhancing the osteogenic differentiation of BMSCs and the migration of human umbilical vein endothelial cells and promoting angiogenesis. The combined ability of these factors to promote osteogenesis and angiogenesis is superior to that of each alone. In vivo, the PQBD scaffold was implanted into the bone defect, and after 8 weeks, the defect area was almost completely covered by new bone tissue. Histology showed more mature bone tissue and more blood vessels. PQBD scaffolds promote both angiogenesis and osteogenesis, offering a promising approach to enhance bone regeneration in the treatment of large bone defects.
Asunto(s)
Deferoxamina , Osteogénesis , Humanos , Andamios del Tejido/química , Factor A de Crecimiento Endotelial Vascular , Regeneración Ósea , Ingeniería de Tejidos/métodos , Diferenciación Celular , Péptidos/farmacología , Péptidos/química , Células Endoteliales de la Vena Umbilical HumanaRESUMEN
Osteosarcoma (OS) is the most common primary malignant bone tumor and is mainly diagnosed in children. Toll-like receptor 9 (TLR9) is expressed in various tumor cells and was correlated with cancer progression. However, the underlying mechanism of TLR9 on the OS progression remains unclear. Our previous study demonstrated that the expression of TLR9 was positively correlated with the development stage of OS. Herein, we further evaluated the actual roles and the molecular mechanism of TLR9 on regulating OS cell proliferation and metastasis. Our data showed that TLR9 was upregulated in OS cells compared to normal osteoblastic cells, and knockdown of TLR9 inhibited OS cell proliferation and induced cell cycle arrest by the decreased expression of cyclin D1, CDK2, and p-Rb, while TLR9 overexpression exerted the inverse effects. Furthermore, TLR9 overexpression could enhance the migration and invasion activities of the OS cells by the upregulation of matrix metalloproteinases 2 (MMP2) and MMP9, and the opposite result was observed in TLR9-silenced cells. Moreover, the nuclear factor kappa B (NF-κB) signaling pathway was activated by TLR9, and TLR9-induced malignant phenotype of OS cells was abrogated by the NF-κB antagonist BAY11-7082. Our study indicated that TLR9 might play a critical role in facilitating OS progression by activating the NF-κB signaling pathway, which may provide a valuable therapeutic target for OS.