摘要: Objectives Mesenchymal stem cells (MSCs) could regulate the function of various immune cells. It remains unclear whether MSCs additionally possess immunostimulatory properties. We investigated the impact of human MSCs on the responsiveness of primary natural killer (NK) cells in terms of induction of anti-inflammatory purinergic signalling. Material and Methods We obtained human bone marrow mesenchymal stem cells (BMMSCs) and dental pulp stem cells (DPSCs). NK cells were isolated from peripheral blood of healthy volunteers. Activated NK cells were cultured with MSCs. Proliferation assay, apoptosis analysis, activating or inhibitory receptor expression and degranulation assay were used to explore NK cells' function. High-performance liquid chromatography was used to investigate the purinergic signalling in activated NK cells. Results Both DPSCs and BMMSCs could impair proliferation and promote apoptosis of activated NK cells. Also, activated NK cells could cause DPSCs to lyse. Furthermore, the expression of activating NK cells' receptors was decreased, but inhibitory receptors of NK cells were elevated following co-cultivation. NK cells acquired CD73 expression, while MSCs could release ATP into the extracellular space where nucleotides were converted into adenosine (ADO) following co-culture system. Under the existence of exogenous 2-chloroadenosine (CADO), the cytotoxic capacity of NK cells was remarkably depressed in a concentration-dependent manner. Conclusions DPSCs and BMMSCs could depress NK cells' function by hydrolysing ATP to ADO using CD39 and CD73 enzymatic activity. Our data suggested that DPSCs might represent a new strategy for treating immune-related diseases by regulating previously unrecognized functions in innate immune responses.

摘要: Objectives Exosomes, as important players in intercellular communication due to their ability to transfer certain molecules to target cells, are believed to take similar effects in promoting bone regeneration with their derived stem cells. Studies have suggested that umbilical cord mesenchymal stem cells (uMSCs) could promote angiogenesis. This study investigated whether exosomes derived from uMSCs (uMSC-Exos) could enhance fracture healing as primary factors by promoting angiogenesis. Materials and Methods uMSCs were obtained to isolate uMSC-Exos by ultrafiltration, with exosomes from human embryonic kidney 293 cells (HEK293) and phosphate-buffered saline (PBS) being used as control groups. NanoSight, laser light scattering spectrometer, transmission electron microscopy and Western blotting were used to identify exosomes. Next, uMSC-Exos combined with hydrogel were transplanted into the fracture site in a rat model of femoral fracture. Bone healing processes were monitored and evaluated by radiographic methods on days 7, 14, 21 and 31 after surgery; angiogenesis of the fracture sites was assessed by radiographic and histological strategies on post-operative day 14. In vitro, the expression levels of osteogenesis- or angiogenesis-related genes after being cultured with uMSC-Exos were identified by qRT-PCR. The internalization ability of exosomes was determined using the PKH67 assay. Cell cycle analysis, EdU incorporation and immunofluorescence staining, scratch wound assay and tube formation analysis were also used to determine the altered abilities of human umbilical vein endothelial cells (HUVECs) administered with uMSC-Exos in proliferation, migration and angiogenesis. Finally, to further explore the underlying molecular mechanisms, specific RNA inhibitors or siRNAs were used, and the subsequent effects were observed. Results uMSC-Exos had a diameter of approximately 100 nm, were spherical, meanwhile expressing CD9, CD63 and CD81. Transplantation of uMSC-Exos markedly enhanced angiogenesis and bone healing processes in a rat model of femoral fracture. In vitro, other than enhancing osteogenic differentiation, uMSC-Exos increased the expression of vascular endothelial growth factor (VEGF) and hypoxia inducible factor-1 alpha (HIF-1 alpha). uMSC-Exos were taken up by HUVECs and enhanced their proliferation, migration and tube formation. Finally, by using specific RNA inhibitors or siRNAs, it has been confirmed that HIF-1 alpha played an important role in the uMSC-Exos-induced VEGF expression, pro-angiogenesis and enhanced fracture repair, which may be one of the underlying mechanisms. Conclusions These results revealed a novel role of exosomes in uMSC-mediated therapy and suggested that implanted uMSC-Exos may represent a crucial clinical strategy to accelerate fracture healing via the promotion of angiogenesis. HIF-1 alpha played an important role in this process.

摘要: Objectives Circular RNAs (circRNAs) exist extensively in the eukaryotic genome. The study aimed to identify the role of hsa_circ_0008365 (Circ-SERPINE2) in gastric carcinoma (GC) cells and its downstream mechanisms. Materials and methods Gene Expression Omnibus (GEO) database was applied to screen differentially expressed circRNAs. CircInteractome, TargetScan and miRecords websites were used to predict target relationships. qRT-PCR and RNase R treatment were utilised to detect molecule expression and confirm the existence of circ-SERPINE2. RNA pull-down assay and dual-luciferase reporter assay were performed for interaction between circRNA and miRNA or mRNA. EdU assay, colony formation assay, and flow cytometry for apoptosis and cell cycle detections were utilised to assess cell function. Western blot and immunohistochemistry (IHC) assays were applied for detection of proteins in tissues or cells. Results Circ-SERPINE2 and YWHAZ were upregulated, and miR-375 was downregulated in GC tissues and cells. Circ-SERPINE2 and YWHAZ targetedly bound to miR-375. Circ-SERPINE2 promoted cell proliferation and cell cycle progress and inhibited cell apoptosis by sponging miR-375 and regulating YWHAZ expression in vitro. Circ-SERPINE2 repressed solid tumour growth through enhancing miR-375 expression and reducing YWHAZ expression in vivo. Conclusions Circ-SERPINE2 is a novel proliferative promoter through the regulation of miR-375/YWHAZ. Circ-SERPINE2/miR-375/YWHAZ axis might provide a novel therapeutic target of GC.

摘要: Objectives To reveal the role of circular RNA (circRNA) DOCK1 (circDOCK1) as a potential biomarker and therapeutic target and its competing endogenous RNA mechanism in bladder carcinoma (BC). Methods The next-generation sequencing (NGS) technology was introduced to screen the circRNA expression profiles of BC using microarray. qPCR and Western blots assay were employed to measure the gene expression in different groups. Cell counting kit-8, EdU and transwell assays were applied to detect the cell viability, proliferation and migration potential, respectively. Luciferase reporter assay was used to test the binds between hsa-miR-132-3p/Sox5. Xenografted tumour growth of nude mice was performed to test the role of circDOCK1 in vivo. Results CircDOCK1 was upregulated in BC tissues and cell lines. Repression of circDOCK1 reduced cell viability, inhibited cell proliferation and curbed the cell migration potential of BC cell. CircDOCK1 played its role via regulation of circDOCK1/hsa-miR-132-3p/Sox5 pathway in BC cells. Suppression circDOCK1 inhibited the tumour growth in vivo. Conclusion In this study, we revealed that circDOCK1 affected the progression of BC via modulation of circDOCK1/hsa-miR-132-3p/Sox5 pathway both in vitro and in vivo and providing a potential biomarker and therapeutic targets for BC.

摘要: Objectives Fluorine, an organic trace element, has been shown to unfavourably effect osteoclasts function at a low dose. Use of hydroxyapatite (HA) has been effective in exploring its roles in promoting bone repair. In this study, we used HA modified with fluorine to investigate whether it could influence osteoclastic activity in vitro and ovariectomy-induced osteoclasts hyperfunction in vivo. Materials and methods Fluorohydroxyapatite (FHA) was obtained and characterized by scanning electron microscope (SEM). Osteoclasts proliferation and apoptosis treated with FHA were assessed by MTT and TUNEL assay. SEM, F-actin, TRAP activity and bone resorption experiment were performed to determine the influence of FHA on osteoclasts differentiation and function. Moreover, HA and FHA were implanted into ovariectomized osteoporotic and sham surgery rats. Histology and Micro-CT were examined for further verification. Results Fluorine released from FHA slowly and sustainably. FHA hampered osteoclasts proliferation, promoted osteoclasts apoptosis, suppressed osteoclasts differentiation and function. Experiments in vivo validated that FHA participation brought about an inhibitory effect on osteoclasts hyperfunction and less bone absorption. Conclusion The results indicated that FHA served as an efficient regulator to attenuate osteoclasts formation and function and was proposed as a candidature for bone tissue engineering applications.