Osteoblasts are essential for maintaining skeletal architecture and modulating bone microenvironment homeostasis. From numerous associated investigations, the BMP-2 pathway has been well-defined as a vital positive modulator of bone homeostasis. Gremlin2 (Grem2) is a bone morphogenetic protein (BMP) antagonists. However, the effect of Grem2 on the BMP-2-induced osteogenesis of human bone marrow-derived mesenchymal stem cells (hBMSCs) remains ambiguous. This study aimed to analyze the procedure in vitro and in vivo. The differentiation of hBMSCs was assessed by determining the expression levels of several osteoblastic genes, as well as the enzymatic activity and calcification of alkaline phosphatase. We found that Grem2 expression was upregulated by BMP-2 within the range of 0-1 μg/mL, and significant increases were evident at 48, 72, and 96 h after BMP-2 treatment. Si-Grem2 increased the BMP-2-induced osteogenic differentiation of hBMSCs, whereas overexpression of Grem2 had the opposite trend. The result was confirmed using a defective femur model. We also discovered that the BMP-2/Smad/Runx2 pathway played an important role in the process. This study showed that si-Grem2 increased the BMP-2-induced osteogenic differentiation of hBMSCs via the BMP-2/Smad/Runx2 pathway. J. Cell. Biochem. 118: 286-297, 2017. © 2016 Wiley Periodicals, Inc.

Background Extrachromosomal circular deoxyribonucleic acid (eccDNA) is evolving as a valuable biomarker, while little is known about its presence in urine. Methods Here, we report the discovery and analysis of urinary cell‐free eccDNAs (ucf‐eccDNAs) in healthy controls and patients with advanced chronic kidney disease (CKD) by Circle‐Seq. Results Millions of unique ucf‐eccDNAs were identified and comprehensively characterised. The ucf‐eccDNAs are GC‐rich. Most ucf‐eccDNAs are less than 1000 bp and are enriched in four pronounced peaks at 207, 358, 553 and 732 bp. Analysis of the genomic distribution of ucf‐eccDNAs shows that eccDNAs are found on all chromosomes but enriched on chromosomes 17, 19 and 20 with a high density of protein‐coding genes, CpG islands, short interspersed transposable elements (SINEs) and simple repeat elements. Analysis of eccDNA junction sequences further suggests that microhomology and palindromic repeats might be involved in eccDNA formation. The ucf‐eccDNAs in CKD patients are significantly higher than those in healthy controls. Moreover, eccDNA with miRNA genes is highly enriched in CKD ucf‐eccDNA. Conclusions This work discovers and provides the first deep characterisation of ucf‐eccDNAs and suggests ucf‐eccDNA as a valuable noninnvasive biomarker for urogenital disorder diagnosis and monitoring.

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Developmental dysplasia of the hip (DDH) is a developmental disorder that has long-term chronic pain and limited hip joint mobility as major pathological characteristics. This study aims to access the association between the development of DDH and cartilage metabolic disorders. Cartilage tissue samples were acquired from patients with DDH, osteoarthritis (OA) and femoral neck fracture. The proteoglycan level was evaluated by safranin O-fast green, toluidine blue and hematoxylin-eosin (HE) staining. The levels of collagen-II (Col-II), collagen-X (Col-X) and metal matrix proteinase-13 (MMP-13) were evaluated by immunohistochemistry (IHC) and Western blotting analysis. The morphologic evaluation of cartilage was conducted by transmission electron microscopy (TEM). Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to detect the mRNA level of aggrecan, Col-II, Col-X and MMP-13. The aggrecan level in the cartilage matrix was significantly decreased in DDH patients by safranin O-fast green and toluidine blue staining in comparison with that in the OA and control groups. In contrast with the OA group, the Col-II expression was reduced while the MMP-13 expression increased in DDH patients, as shown by IHC and Western blotting analysis. The collagenous fibrils in cartilage of DDH patients appeared significantly sparse and disordered in the TEM analysis. In DDH patients, the mRNA expression levels of Col-II and aggrecan were markedly reduced, while the mRNA expression of Col-X was markedly increased, compared with the OA patients. There is severe articular cartilage degeneration in DDH patients. This observation provides us with new insight into cartilage metabolic regulation in DDH. © 2017 IUBMB Life, 69(3):179-187, 2017.