Temporal transcriptome highlights the involvement of cytokine/JAK/STAT3 signaling pathway in the osteoinduction of BMSCs.

BACKGROUND: Mesenchymal stem cells (MSCs)-based therapy offers an effective strategy for bone regeneration to solve the clinical orthopedic problems. However, the transcriptional regulation of multiple transitional stages of continuous osteogenesis from MSCs has not been fully characterized. METHODS: Bone marrow mesenchymal stem cells (BMSCs) stimulated with osteogenic induction media were utilized to construct the in vitro osteogenic differentiation model. BMSCs were harvested after induction for 0, 7, 14 and 21 days, respectively, to perform the mRNA-sequencing (mRNA-Seq). The transcription factor networks and common molecules during the osteogenesis were revealed by using the temporal transcriptome. Further verification was performed by the quantitative real-time polymerase chain reaction (qRT-PCR), immunofluorescence and Western blotting. RESULTS: It showed that BMSCs could differentiate into osteogenic, and crucial regulator in the MAPK signaling pathway, the PPAR signaling pathway, the Toll-like receptor signaling and the Cytokine/JAK/STAT signaling pathway. PPI protein interaction analysis also suggested that three cytokines are involved in osteogenic differentiation as core genes, including leukemia inhibitory factor (LIF), interleukin-6 (IL6) and colony-stimulating factor 3 (CSF3). The osteogenic process was negatively affected by the inhibition of JAK/STAT3 signaling pathway. CONCLUSIONS: This work might provide new insights in the crucial features of the transcriptional regulation during the osteogenesis, as well as offer important clues about the activity and regulation of the relatively long-activated Cytokine/JAK/STAT3 signaling pathway in osteoinduction of BMSCs.

Articular chondrocyte-derived extracellular vesicles promote cartilage differentiation of human umbilical cord mesenchymal stem cells by activation of autophagy.

BACKGROUND: Umbilical cord mesenchymal stem cell (HUCMSC)-based therapies were previously utilised for cartilage regeneration because of the chondrogenic potential of MSCs. However, chondrogenic differentiation of HUCMSCs is limited by the administration of growth factors like TGF-ß that may cause cartilage hypertrophy. It has been reported that extracellular vesicles (EVs) could modulate the phenotypic expression of stem cells. However, the role of human chondrogenic-derived EVs (C-EVs) in chondrogenic differentiation of HUCMSCs has not been reported. RESULTS: We successfully isolated C-EVs from human multi-finger cartilage and found that C-EVs efficiently promoted the proliferation and chondrogenic differentiation of HUCMSCs, evidenced by highly expressed aggrecan (ACAN), COL2A, and SOX-9. Moreover, the expression of the fibrotic marker COL1A and hypertrophic marker COL10 was significantly lower than that induced by TGF-ß. In vivo, C-EVs induced HUCMSCs accelerated the repair of the rabbit model of knee cartilage defect. Furthermore, C-EVs led to an increase in autophagosomes during the process of chondrogenic differentiation, indicating that C-EVs promote cartilage regeneration through the activation of autophagy. CONCLUSIONS: C-EVs play an essential role in fostering chondrogenic differentiation and proliferation of HUCMSCs, which may be beneficial for articular cartilage repair.

Identification of potential specific biomarkers and key signaling pathways between osteogenic and adipogenic differentiation of hBMSCs for osteoporosis therapy.

BACKGROUND: The differentiation of bone mesenchymal stem cells (BMSCs) into adipogenesis (AD) rather than osteogenesis (OS) is an important pathological feature of osteoporosis. Illuminating the detailed mechanisms of the differentiation of BMSCs into OS and AD would contribute to the interpretation of osteoporosis pathology. METHODS: To identify the regulated mechanism in lineage commitment of the BMSCs into OS and AD in the early stages, the gene expression profiles with temporal series were downloaded to reveal the distinct fates when BMSCs adopt a committed lineage. For both OS and AD lineages, the profiles of days 2-4 were compared with day 0 to screen the differentially expressed genes (DEGs), respectively. Next, the functional enrichment analysis was utilized to find out the biological function, and protein-protein interaction network to predict the central genes. Finally, experiments were performed to verify our finding. RESULTS: FoxO signaling pathway with central genes like FoxO3, IL6, and CAT is the crucial mechanism of OS, while Rap1 signaling pathway of VEGFA and FGF2 enrichment is more significant for AD. Besides, PI3K-Akt signaling pathway might serve as the latent mechanism about the initiation of differentiation of BMSCs into multiple lineages. CONCLUSION: Above hub genes and early-responder signaling pathways control osteogenic and adipogenic fates of BMSCs, which maybe mechanistic models clarifying the changes of bone metabolism in the clinical progress of osteoporosis. The findings provide a crucial reference for the prevention and therapy of osteoporosis.

Electrospun poly(3-hydroxybutyrate-co-4-hydroxybutyrate) /Octacalcium phosphate Nanofibrous membranes for effective guided bone regeneration.

Membranes used in guided bone regeneration (GBR) are required to exhibit high mechanical strength, biocompatibility, biodegradation, osteogenic and osteoinductive potential. In our study, poly(3-hydroxybutyrate-co-4-hydroxybutyrate)(P(3HB-co-4HB))/octacalcium phosphate (OCP) (P(3HB-co-4HB)/OCP) nanofibrous membranes were fabricated by electrospinning with two different P(3HB-co-4HB) to OCP ratios (P(3HB-co-4HB):OCP = 95:5 wt% and 90:10 wt%, termed P(3HB-co-4HB)/OCP(5)and P(3HB-co-4HB)/OCP (10), respectively) for GBR. The developed P(3HB-co-4HB)/OCP nanofibrous membranes were analysed for their osteogenic and osteoinductive properties using mesenchymal stem cells (MSCs) in vitro and in a calvarial bone defect rat model. The composite P(3HB-co-4HB)/OCP nanofibrous membranes showed decreased fibre size and enhanced tensile strength compared with those of P(3HB-co-4HB) nanofibrous membranes. In the in vitro studies, the P(3HB-co-4HB)/OCP membranes facilitated cell growth and osteoblastic differentiation of MSCs and were superior to P(3HB-co-4HB) membranes. After covered on the calvarial bone defects, P(3HB-co-4HB)/OCP membranes facilitated greater neobone formation than P(3HB-co-4HB) membranes did, as the result of histological evaluation and micro-CT analysis with higher bone volume/total volume (BV/TV) ratio and bone mineral density (BMD). P(3HB-co-4HB)/OCP(10) membranes with higher OCP content showed greater stiffness and osteoinductivity than P(3HB-co-4HB)/OCP (5)membranes, demonstrating the role of OCP in the composite membranes. These results indicated that electrospun P(3HB-co-4HB)/OCP nanofibrous membranes hold promise for the clinical application of GBR.

BTN3A2 serves as a prognostic marker and favors immune infiltration in triple-negative breast cancer.

Immune infiltration is reported to be highly associated with tumor progress. Since butyrophilin subfamily 3 member A2 (BTN3A2) serves as a crucial mediator in immune activation, we aimed to investigate the correlation of BTN3A2 in immune infiltration and tumor prognosis via extensive-cancer analysis. The levels of BTN3A2 expression in extensive cancers were analyzed with Oncomine and TIMER databases. Univariate cox and multivariate cox regression analyses were conducted to assess the associations of BTN3A2 to prognosis of various cancers. The correlations of BTN3A2 with immune infiltration were assessed by TIMER database. It suggested that BTN3A2 was a potential prognosis signature for breast cancer (BRCA) and ovarian cancer (OV). However, immune infiltrations were highly correlated with BTN3A2 in triple-negative breast cancer (TNBC), compared with OV and other subtypes of BRCA. Multivariate cox regression analysis revealed that BTN3A2 was an independently prognostic signature of TNBC, as well as weighted correlation network analysis suggested BTN3A2 was only correlated with TNBC, rather than other subtypes of BRCA. Immune cell subtypes correlation analysis showed that BTN3A2 was highly correlated with general T, CD8+ T, T helper type 1, exhausted T cells, and dendritic cells in TNBC. And the coexpression geneset of BTN3A2 was mainly involved in T-cell receptor interaction and the nuclear factor-κB (NF-κB) signaling pathway. Collectively, BTN3A2 that was positively associated with better prognosis could be served as a special diagnostic and independently prognostic marker for TNBC by regulating the T-cell receptor interaction and NF-κB signaling pathways.

Electrospun PLGA/PCL/OCP nanofiber membranes promote osteogenic differentiation of mesenchymal stem cells (MSCs).

Nanofibers as niche-biomimetic scaffolds hold promise in guided bone regeneration (GBR). Here we fabricated poly (lactic-co-glycolic acid) (PLGA)/poly(caprolactone) (PCL)-doped octacalcium phosphate (OCP) nanofiber membranes via electrospinning and investigated the osteogenic behavior of marrow mesenchymal stem cells (MSCs) on the membranes. By adjusting different ratio of OCP to PLGA/PCL, three hybrid stents including PLGA/PCL, PLGA/PCL/2 wt%OCP, PLGA/PCL/4wt%OCP were successfully prepared. The PLGA/PCL/OCP membranes showed a decrease in fiber diameter compared with PLGA/PCL, leading to enhanced mechanical strength. In-vitro studies showed that PLGA/PCL/OCP membranes better supported cell adhesion, spreading and proliferation than PLGA/PCL. The incorporation of OCP via electrospinning also endowed the membranes with osteoinductive capacity, as evidenced by activation of ALP activity, increased gene expression of bone specific markers (such as Runx2, ALP, Col 1a1, OPN, OCN, BMP2), and mineral nodules formation compared to PLGA/PCL. Comparatively, PLGA/PCL/4wt%OCP showed better mechanical and biological performance than PLGA/PCL/2 wt%OCP, demonstrating the role of OCP in nanofiber membranes. Thus, the electrospun PLGA/PCL/OCP nanofiber membranes can be potentially developed as a promising hybrid stent for GBR.

Comparative profiling of chondrogenic differentiation of mesenchymal stem cells (MSCs) driven by two different growth factors.

This study aimed to investigate the mechanism of nerve growth factor (NGF) from cobra venom and human transforming growth factor-ß1 (TGF-ß1) on the chondrogenic induction of mesenchymal stem cells (MSCs). NGF and TGF-ß1 were used to induce chondrogenesis of MSCs from rabbits for 7 days. Total RNA was extracted for mRNA sequencing. Differentially expressed genes (DEGs), gene ontology (GO), KEGG pathway enrichment, and PPI network analysis were conducted to screen the specific signalling pathways and target genes. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to further confirm the relative target genes. The results showed that NGF could significantly promote the expression of hyaline cartilage specific genes (collagen type II alpha 1 chain, COL2A1) compared with TGF-ß1. PI3K-AKT signalling pathway is commonly involved in the chondrogenesis of MSCs induced by NGF and TGF-ß1. However, the expression levels of the genes in the PI3K-AKT signalling pathway were significantly higher in NGF group than that in the TGF-ß1 group. In the process of chondrogenesis of MSCs induced by NGF and TGF-ß1, integrin (ITGAs) were the targeted hub genes to activate the PI3K-AKT signalling pathway. NGF could activate more proliferation and differentiation genes in the process of chondrogenesis of MSCs than TGF-ß1. TGF-ß1 promoted angiogenesis by targeting the thrombospondin (THBS1) and THBS2 which might contribute to the osteophyte formation. PI3K-AKT was the crucial signalling pathway for chondrogenic differentiation. NGF could activate the PI3K-AKT signalling pathway to a higher level, and NGF had more specificity for promoting expression of specific genes of chondrocyte compared with TGF-ß1. SIGNIFICANCE OF THE STUDY: In our study, we compared two different growth factors in promoting cartilage differentiation of MSCs and found some similarities and differences. We revealed that both NGF and TGF-ß1 could activate the PI3K-AKT signalling pathway (the expression of it in NGF was higher) by targeting the ITGAs in the process of chondrogenesis from MSCs. However, NGF could activate more proliferation and differentiation genes in the process of chondrogenesis of MSCs, whereas TGF-ß1 caused osteophyte formation by activating THBS1 and THBS2. These might be the reason why NGF could promote cartilage differentiation more specifically.

Comparison of rheumatoid arthritis (RA) and osteoarthritis (OA) based on microarray profiles of human joint fibroblast-like synoviocytes.

The purpose of the present study was to investigate the underlying molecular mechanism of osteoarthritis (OA) and rheumatoid arthritis (RA) based on microarray profiles. Three human joint fibroblast-like synoviocytes (FLSs) microarray profiles including 26 OA samples, 33 RA samples, and 20 healthy control (HC) samples were downloaded from the GEO database. Differentially expressed genes (DEGs) between OA and HC (DEGsOA) and RA and HC (DEGsRA) were identified. Co-expressed and specific genes were analysed between DEGsOA and DEGsRA. Gene ontology, KEGG pathway enrichment, PPI network, and GSEA were performed to predict the function of DEGs. Two hundred seventy-six and 410 differential genes in DEGsOA and DEGsRA were observed. One hundred fifty coexpressed genes and 126 OA-specific genes (SELE, SERPINE1, and NFKBIA were the key genes) between DEGsOA and DEGsRA were enriched in the tumour necrosis factor (TNF) signalling pathway. However, 260 RA-specific genes of which the key genes were CCR5, CCR7, CXCR4, CCL5, and CCR4 were enriched in chemokine signalling pathway. Therefore, FLSs might exert an inflammatory effect by regulating TNF signalling pathway, targeting SELE, SERPINE1, and NFKBIA during the process of OA. Although TNF signalling pathway was also involved in the synovitis of RA, chemokine signalling pathway played the key role in RA FLSs mediating cell migration, invasion, and release of chemotaxis. In addition, CCR5, CCR7, CXCR4, CCL5, and CCR4 might be hub genes in RA. The different biomarkers and pathways identified in OA and RA may provide references for further study. SIGNIFICANCE OF THE STUDY: This study revealed the similar and different mechanisms of FLSs and different biomarkers that might with important regulatory effects on RA and OA. In OA, FLSs played an inflammatory role through TNF signalling pathway, targeting SELE, SERPINE1, and NFKBIA. Although TNF signalling pathway was also involved in the synovitis of RA, chemokine signalling pathway was a crucial pathway in mediating FLSs migration, invasion, and release of chemotaxis. CCR5, CCR7, CXCR4, CCL5, and CCR4 might be keys genes in RA. We expect that our results will bring more comprehensively understanding between RA and OA for researchers.