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Objective To observe the regulatory effect of exosomes from adipose-derived stem cells （ADSC-Exos） on the biological activity of scar fibroblasts. Methods Scar fibroblast were prepared and divided into the control and ADSC-Exos groups. The proliferation of scar fibroblasts between two groups was observed by CCK-8 assay. The expression levels of fibrosis-related indexes were detected by Western blot. The expression levels of fibrosis-related gene mRNA were determined by fluorescent quantitative real-time （RT） -PCR. Results Compared with the control group, the proliferation capability of scar fibroblasts was significantly weakened, and the fibrosis-related indexes were significantly decreased in ADSC-Exos group （both P < 0.05）. Meanwhile, the expression levels of collagen I, III and α-smooth muscle actin mRNA of scar fibroblasts were significantly down-regulated in the ADSC-Exos group（all P < 0.05）. Conclusion ADSC-Exos can inhibit the biological activity of scar fibroblasts.

Studies addressing the anatomical values of the supraspinatus outlet area (SOA) and the available supraspinatus outlet area (ASOA) are insufficient. This study focused on precisely measuring the SOA and ASOA values in a sample from the Chinese population using 3D CT (computed tomography) reconstruction. We analyzed CT imaging of 96 normal patients (59 males and 37 females) who underwent shoulder examinations in a hospital between 2011 and 2021. The SOA, ASOA, acromiohumeral distance (AHD), coracohumeral distance (CHD), coracoacromial arch radius (CAR), and humeral head radius (HHR) were estimated, and statistical correlation analyses were performed. There were significant sex differences observed in SOA (men: 957.62 ± 158.66 mm2; women: 735.87 ± 95.86 mm2) and ASOA (men: 661.35 ± 104.88 mm2; women: 511.49 ± 69.26 mm2), CHD (men: 11.22 ± 2.24 mm; women: 9.23 ± 1.35 mm), CAR (men: 37.18 ± 2.70 mm; women: 33.04 ± 3.15 mm), and HHR (men: 22.65 ± 1.44 mm; women: 20.53 ± 0.95 mm). Additionally, both SOA and ASOA showed positive and linear correlations with AHD, CHD, CAR, and HHR (R: 0.304–0.494, all p < 0.05). This study provides physiologic reference values of SOA and ASOA in the Chinese population, highlighting the sex differences and the correlations with shoulder anatomical parameters. [ABSTRACT FROM AUTHOR]

Studies addressing the anatomical values of the supraspinatus outlet area (SOA) and the available supraspinatus outlet area (ASOA) are insufficient. This study focused on precisely measuring the SOA and ASOA values in a sample from the Chinese population using 3D CT (computed tomography) reconstruction. We analyzed CT imaging of 96 normal patients (59 males and 37 females) who underwent shoulder examinations in a hospital between 2011 and 2021. The SOA, ASOA, acromiohumeral distance (AHD), coracohumeral distance (CHD), coracoacromial arch radius (CAR), and humeral head radius (HHR) were estimated, and statistical correlation analyses were performed. There were significant sex differences observed in SOA (men: 957.62 ± 158.66 mm 2 ; women: 735.87 ± 95.86 mm 2 ) and ASOA (men: 661.35 ± 104.88 mm 2 ; women: 511.49 ± 69.26 mm 2 ), CHD (men: 11.22 ± 2.24 mm; women: 9.23 ± 1.35 mm), CAR (men: 37.18 ± 2.70 mm; women: 33.04 ± 3.15 mm), and HHR (men: 22.65 ± 1.44 mm; women: 20.53 ± 0.95 mm). Additionally, both SOA and ASOA showed positive and linear correlations with AHD, CHD, CAR, and HHR (R: 0.304-0.494, all p < 0.05). This study provides physiologic reference values of SOA and ASOA in the Chinese population, highlighting the sex differences and the correlations with shoulder anatomical parameters.

Background: Angiogenesis is an essential step in tissue engineering. MSC exosomes play an important role in angiogenesis. Functional biomolecules in exosomes vested by the culture microenvironment can be transferred to recipient cells and affects their effect. 3D culture can improve the proliferation and activity of MSCs. However, whether exosomes derived from 3D culture of MSCs have an enhanced effect on angiogenesis is unclear., Methods: Herein, we compared the bioactivity of exosomes produced by conventional 2D culture (2D-exos) and 3D culture (3D-exos) of bone marrow stem cells (BMSCs) in angiogenesis., Results: A series of in vitro and in vivo experiments indicated that 3D-exos exhibited stronger effects on HUVEC cell proliferation, migration, tube formation, and in vivo angiogenesis compared with 2D-exos. Moreover, the superiority of 3D-exos might be attributed to the activation of HMGB1/AKT signaling., Conclusions: These results indicate that exosomes from 3D culture of MSCs may serve as a potential therapeutic approach for pro-angiogenesis., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Understanding the mechanisms that govern cell fate will lead to the development of techniques for the induction of human mesenchymal stem cell differentiation into desired cell outcomes and the production of an autologous source of tissue for regenerative medicine. Here, we demonstrate that stem cells derived from adult bone marrow grown with 3D pellets take on characteristics similar to human cartilage. The NFAT signaling pathway is primarily linked to cell differentiation and influences chondrogenic differentiation. Based on our previous results that alterations in the expression of the NFATc1 gene affect chondrogenesis, we screened a microarray and identified 29 genes with altered expression, including 13 up-regulated (fold change ≥ 2) and 16 down-regulated (fold change ≤ 2) genes, compared with the control group. We then used RT-PCR to validate the chip data. Gene ontology and pathway analyses were performed on these altered genes. We found that these altered genes function in the complement and coagulation cascades, metabolism, biosynthesis, transcriptional regulation, proteolysis, and intracellular signaling pathways, such as the cytoplasmic calcineurin-dependent signaling pathway, the cyclin-dependent kinase inhibitor 2C signaling pathway, the MAPK signaling pathway, and the insulin signaling pathway. Our study suggests that these pathways may play important roles in chondrogenesis, which could be useful in the design of biomaterials., Competing Interests: None.

Objective: To integrate the result of whole genome expression data and whole genome promoter CpG island methylation data, to screen the epigenetic modulated differentially expressed genes from transformed porcine bone marrow mesenchymal stem cells (BMSCs) after long-term cultivation., Methods: Bone marrow from 6 landrace pigs, 3-month-old about 50 kg weight, was aspirated from the medullary cavity of the proximal tibia. The BMSCs were isolated, and purified by Ficoll density gradient centrifugation combined with adherent culture method. The transfor mation of BMSCs was tested by several methods including cell morphology observation, karyotype analysis, clone forming in soft agarose, serum requirement assay, and tumor forming in mice. The Agilent Pig 4x44k Gene Expression Microarray was used to investigate the differentially expressed mRNA. The methylated genes expression profile was performed using customized pig methylation chip. The gene expression and DNA methylation profiles were integrated to find out the epigenetic modulated differentially expressed genes, and to complete the bioinformatic analysis., Results: BMSCs showed a change in appearance, from the initial spindle shape to a more flatted morphology then to small contact shape. After additional passages, BMSCs gradually acquired recovery of proliferating capacity and transformation properties such as anchorage-independent growth, chromosomal abnormality, and tumor formation in nude mice. The gene chip analysis demonstrated that 257 genes were upregulated and 315 genes were downregulated during long-term cultures as well as multiple signal pathways transduction involved, such as cell cycle, ECM-receptor interaction, focal adhesion, regulation of actin cytoskeleton, pathways in cancer, and P53. The analysis from methylation chip of coding genes suggested epigenetic regulation was involved in BMSCs spontaneous transformation and play a important role on it; 962 genes were hypermethylated and 1219 genes were hypomethylated, which were involved in the biological process of cellular metabolic, structure, and tumor generation. The combined analysis of genes regulated by methylation in the transformation process of BMSCs found that the methylation changes of the 35 genes were contrary to the direction of expression change (correlation coefficient r =-0.686, P =0.000); in which the methylation level of 21 genes promoter regions were increased while the gene expression decreased, and the methylation level of the 14 genes promoter regions decreased and the gene expression increased. At the same time, KEGG enrichment analysis revealed multiple genes regulated by methylation, involved in stem cell differentiation and multiple cell signaling pathways. Among the 14 down-regulated genes, many of them have the role of regulating the interaction of tumor and immunization, and the change of the methylation status of the CDKN3 promoter region may be closely related to the cell oncology., Conclusion: The results deepen our understanding of the crucial role of coding genes methylation modification in BMSCs transformation, and may provide new approach to establish safe criteria for BMSCs clinical applications and transformation prevention.

Chondrogenic differentiation of mesenchymal stem cells is regulated by many different pathways. Recent studies have established that hypoxia and epigenetic alterations potently affect expression of chondrogenesis marker genes. Sox9 is generally regarded as a master regulator of chondrogenesis and microRNA-124 (miRNA-124) regulates gene expression in murine bone marrow-derived mesenchymal stem cells. Therefore, in this study we investigated whether epigenetic regulation of miRNA-124 could affect the expression of Sox9 and thereby regulate chondrogenesis. A cell pellet culture model was used to induce chondrogenesis in C3H10T1/2 cells under hypoxic conditions (2% O 2 ) to determine the effects of hypoxia on miR-124 expression and DNA methylation. The expression of miR-124 was significantly downregulated under hypoxic conditions compared to normoxic conditions (21% O 2 ). The expression of chondrogenesis marker genes was significantly increased under hypoxic conditions. Bisulfite sequencing of the CpG islands in the promoter region of miR-124-3 showed that CpG methylation was significantly increased under hypoxic conditions. Treating the cells with the DNA demethylating agent 5'-AZA significantly increased miR-124 expression and decreased expression of markers of chondrogenesis. Overexpressing miR-124 under hypoxic conditions inhibited NFATc1 reporter activity. NFATc1 was shown to bind to the promoter region of Sox9. Taken together, our data provide evidence that miR-124 acts as an inhibitor of NFATc1. Under hypoxic conditions when miR-124 is downregulated by methylation of CpG islands in the promoter, NFATc1 can bind to the Sox9 promoter and induce the expression of Sox9 leading to chondrogenesis. These results support the role of epigenetic regulation in establishing and maintaining a chondrogenic phenotype., Competing Interests: None.