Circular RNA AFF4 modulates osteogenic differentiation in BM-MSCs by activating SMAD1/5 pathway through miR-135a-5p/FNDC5/Irisin axis.

Bone marrow-derived mesenchymal stem cells (BM-MSCs), the common progenitor cells of adipocytes and osteoblasts, have been recognized as the key mediator during bone formation. Herein, our study aim to investigate molecular mechanisms underlying circular RNA (circRNA) AFF4 (circ_AFF4)-regulated BM-MSCs osteogenesis. BM-MSCs were characterized by FACS, ARS, and ALP staining. Expression patterns of circ_AFF4, miR-135a-5p, FNDC5/Irisin, SMAD1/5, and osteogenesis markers, including ALP, BMP4, RUNX2, Spp1, and Colla1 were detected by qRT-PCR, western blot, or immunofluorescence staining, respectively. Interactions between circ_AFF4 and miR-135a-5p, FNDC5, and miR-135a-5p were analyzed using web tools including TargetScan, miRanda, and miRDB, and further confirmed by luciferase reporter assay and RNA pull-down. Complex formation between Irisin and Integrin αV was verified by Co-immunoprecipitation. To further verify the functional role of circ_AFF4 in vivo during bone formation, we conducted animal experiments harboring circ_AFF4 knockdown, and born samples were evaluated by immunohistochemistry, hematoxylin and eosin, and Masson staining. Circ_AFF4 was upregulated upon osteogenic differentiation induction in BM-MSCs, and miR-135a-5p expression declined as differentiation proceeds. Circ_AFF4 knockdown significantly inhibited osteogenesis potential in BM-MSCs. Circ_AFF4 stimulated FNDC5/Irisin expression through complementary binding to its downstream target molecule miR-135a-5p. Irisin formed an intermolecular complex with Integrin αV and activated the SMAD1/5 pathway during osteogenic differentiation. Our work revealed that circ_AFF4, acting as a sponge of miR-135a-5p, triggers the promotion of FNDC5/Irisin via activating the SMAD1/5 pathway to induce osteogenic differentiation in BM-MSCs. These findings gained a deeper insight into the circRNA-miRNA regulatory system in the bone marrow microenvironment and may improve our understanding of bone formation-related diseases at physiological and pathological levels.

Circ-ITCH sponges miR-214 to promote the osteogenic differentiation in osteoporosis via upregulating YAP1.

Osteoporosis is the most prevailing primary bone disease and a growing health care burden. The aim of this study was to clarify the functional roles and mechanisms of the circ-ITCH regulating osteogenic differentiation of osteoporosis. Circ-ITCH and yes-associated protein 1 (YAP1) levels were downregulated, but the miR-214 level was upregulated in osteoporotic mice and patients. Knockdown of circ-ITCH inhibited the alkaline phosphatase (ALP) activity, mineralized nodule formation, and expression of runt-related transcription factor 2 (RUNX2), osteopontin (OPN), and osteocalcin (OCN) during osteogenic induction. Furthermore, miR-214 was a target of circ-ITCH, knockdown of miR-214 could impede the regulatory effects of sh-circ-ITCH on osteogenic differentiation. Moreover, miR-214 suppressed hBMSCs osteogenic differentiation by downregulating YAP1. Finally, in vivo experiments indicated that overexpression of circ-ITCH could improve osteogenesis in ovariectomized mice. In conclusion, circ-ITCH upregulated YAP1 expression to promote osteogenic differentiation in osteoporosis via sponging miR-214. Circ-ITCH could act as a novel therapeutic target for osteoporosis.

LncRNA DANCR and miR-320a suppressed osteogenic differentiation in osteoporosis by directly inhibiting the Wnt/ß-catenin signaling pathway.

Our study aimed to determine how lncRNA DANCR, miR-320a, and CTNNB1 interact with each other and regulate osteogenic differentiation in osteoporosis. qRT-PCR and western blotting were performed to determine the expression of DANCR, miR-320a, CTNNB1, and the osteoporosis- or Wnt/ß-catenin pathway-related markers T-cell factor 1 (TCF-1), runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP), osteocalcin (OCN), and osteopontin (OPN). Interactions between CTNNB1, DANCR, and miR-320a were predicted by bioinformatics approaches and validated using a luciferase assay. Osteoblastic phenotypes were evaluated by ALP staining, ALP activity assay and Alizarin Red staining. The bilateral ovariectomy method was used to establish an in vivo osteoporosis model. Bone morphological changes were examined using hematoxylin and eosin (H&E) and Alcian Blue staining. The expression levels of DANCR and miR-320a in BMSCs derived from osteoporosis patients were upregulated, whereas CTNNB1 expression was downregulated compared with that in healthy controls. Importantly, we demonstrated that miR-320a and DANCR acted independently from each other and both inhibited CTNNB1 expression, whereas the inhibitory effect was additive when miR-320a and DANCR were cooverexpressed. Moreover, we found that DANCR overexpression largely abrogated the effect of the miR-320a inhibitor on CTNNB1 expression and the Wnt/ß-catenin signaling pathway in BMSCs during osteogenic differentiation. We further confirmed the results above in BMSCs derived from an osteoporosis animal model. Taken together, our findings revealed that DANCR and miR-320a regulated the Wnt/ß-catenin signaling pathway during osteogenic differentiation in osteoporosis through CTNNB1 inhibition. Our results highlight the potential value of DANCR and miR-320a as promising therapeutic targets for osteoporosis treatment.

Pseudogene PTENP1 sponges miR-214 to regulate the expression of PTEN to modulate osteoclast differentiation and attenuate osteoporosis.

BACKGROUND AIMS: Osteoporosis (OP) is a common bone metabolic disease with a high incidence. Our study aimed to explore the pseudogene PTENP1/miR-214/PTEN axis to modulate the osteoclast differentiation in osteoporosis. METHODS: Patients with osteoporosis were recruited in our study, and RANKL-induced osteoclast differentiation and ovariectomy-induced osteoporosis mouse model were established in vitro and in vivo, respectively. RESULTS: Pseudogene PTENP1 and PTEN were significantly down-regulated and miR-214 was up-regulated in osteoporosis patients. In addition, overexpression of PTENP1 or silence of miR-214 inhibited the expression levels of osteoclast specific markers and osteoclast differentiation induced by RANKL. Overexpression of PTENP1 or silence of miR-214 also inhibited the levels of phosphorylation of PI3K and AKT, p65 nuclear translocation, IκBα degradation and the expression level of NFATc1. AlsoSilence of PTENP1 or overexpression of miR-214 induced the osteoclast differentiation under normal physiological condition. Pseudogene PTENP1 sponged miR-214 to regulate the expression of PTEN. CONCLUSIONS: In an ovariectomy-induced osteoporosis mouse model, obvious pathological changes in bone tissues were found, and bone marrow mononuclear cells in this group were more likely to differentiate into osteoclasts. Therefore, pseudogene PTENP1 sponged miR-214 to regulate the expression of PTEN to inhibit osteoclast differentiation and attenuate osteoporosis by suppressing the PI3K/AKT/NF-κB signaling pathway.

A Comparative Study of Patients' Subjective Feelings Toward Total Hip Arthroplasty with Patient-Specific Instruments and Traditional Total Hip Arthroplasty.

OBJECTIVE: To determine whether differences exist in patients' subjective feelings, daily life, and surgical satisfaction between those who underwent surgery for developmental dysplasia of the hip (DDH) using patient-specific instruments (PSIs) and those who underwent traditional surgical total hip arthroplasty (THA). METHODS: We selected 30 adult patients with various types of DDH who underwent surgery during 2016-2017 at our hospital. The patients were divided into PSI surgery group and the traditional surgery group. All patients underwent follow-up, and we collected data on the Harris Hip Score, Oxford University Hip Score (OHS), Forgotten Joint Score (FJS-12), Visual Analogue Scale (VAS) score, patient satisfaction score, intraoperative surgical time, amount of bleeding and postoperative complications incidence for both groups. We then performed statistical analyses on the data. RESULTS: The Harris Hip Score, OHS, VAS score, patient satisfaction score, and mean bleeding volume did not differ statistically significantly (t-tests, P > 0.05). No statistically significant differences were found between surgical groups in the incidence of complication and sub-trochanteric osteotomy, or in the surgical side (chi-square tests, P > 0.05). For the experimental group, the FJS-12 score was 80.0 ± 12.0, and for the control group the score was 68.5 ± 16.1. The operative time of the experimental group was 138.4 ± 32.2 min, while that of the control group was 88.9 ± 26.8 min. The values of these data differed significantly (t-tests, P < 0.05). CONCLUSIONS: The novel PSI designed by our group has certain advantages for the short-term subjective feelings of patients after THA, but it may cause prolonged operative times.

Mixed Reality Combined with Three-Dimensional Printing Technology in Total Hip Arthroplasty: An Updated Review with a Preliminary Case Presentation.

Three-dimensional (3D) printing technology, virtual reality, and augmented reality technology have been used to help surgeons to complete complex total hip arthroplasty, while their respective shortcomings limit their further application. With the development of technology, mixed reality (MR) technology has been applied to improve the success rate of complicated hip arthroplasty because of its unique advantages. We presented a case of a 59-year-old man with an intertrochanteric fracture in the left femur, who had received a prior left hip fusion. After admission to our hospital, a left total hip arthroplasty was performed on the patient using a combination of MR technology and 3D printing technology. Before surgery, 3D reconstruction of a certain bony landmark exposed in the surgical area was first performed. Then a veneer part was designed according to the bony landmark and connected to a reference registration landmark outside the body through a connecting rod. After that, the series of parts were made into a holistic reference registration instrument using 3D printing technology, and the patient's data for bone and surrounding tissue, along with digital 3D information of the reference registration instrument, were imported into the head-mounted display (HMD). During the operation, the disinfected reference registration instrument was installed on the selected bony landmark, and then the automatic real-time registration was realized by HMD through recognizing the registration landmark on the reference registration instrument, whereby the patient's virtual bone and other anatomical structures were quickly and accurately superimposed on the real body of the patient. To the best of our knowledge, this is the first report to use MR combined with 3D printing technology in total hip arthroplasty.

LncRNA KCNQ1OT1 promoted BMP2 expression to regulate osteogenic differentiation by sponging miRNA-214.

BACKGROUND: Osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) is of much significance for bone formation, the imbalance of it would result in osteoporosis and other pathological bone defects. Increasing evidences showed that long non-coding RNAs (lncRNAs) and miRNAs played vital roles in the regulation of osteogenic differentiation. LncRNA KCNQ1OT1 was often regarded as an imprinted lncRNA and was related to tumor progression, while its function in osteogenic differentiation remained unclear. METHOD: qRT-PCR was performed to detect the expression of KCNQ1OT1, miR-214 and osteogenesis-related genes BMP2, Runx2, OPN, and OCN. Western blotting was carried out to detect osteogenesis-related markers. The osteoblastic phenotype was evidenced by alkaline phosphatase (ALP) activity and Alizarin Red S accumulation detection. Bioinformatics and luciferase assays were used to predict and validate the interaction between KCNQ1OT1 and miR-214 as well as BMP2 and miR-214. RESULTS: KCNQ1OT1 was significantly up-regulated during the process of osteogenic induction while miR-214 was contrarily down-regulated. Knockdown of KCNQ1OT1 inhibited osteogenic differentiation and down-regulated BMP2 and osteogenesis-related genes. It was also confirmed that KCNQ1OT1 directly interacted with miR-214. Meanwhile, miR-214 could bind to 3'UTR of BMP2 and therefore inhibited its expression. Furthermore, co-transfection of miR-214 inhibitor could rescue the down-regulation of BMP2 and osteogenesis-related genes and osteogenic differentiation suppression induced by KCNQ1OT1 knockdown. Moreover, miR-214 inhibitor significantly reversed the decreased protein levels of p-Smad1/5/8, Runx2 and Osterix induced by shKCNQ1OT1. CONCLUSIONS: KCNQ1OT1 positively regulated osteogenic differentiation of BMSCs by acting as a ceRNA to regulate BMP2 expression through sponging miR-214.

Decreased expression of SLC 39A14 is associated with tumor aggressiveness and biochemical recurrence of human prostate cancer.

OBJECTIVE: Solute carrier family 39, member 14 (SLC39A14), has been identified as a potential biomarker for various cancers. However, its roles in prostate cancer (PCa) are still unclear. The aim of this study was to investigate the clinical significance of SLC39A14 in patients with PCa and its functions in malignant phenotypes of PCa cells. PATIENTS AND METHODS: Subcellular localization and expression pattern of SLC39A14 protein were examined by immunohistochemistry. Then, the associations of SLC39A14 expression with various clinicopathological features and clinical outcome of patients with PCa were statistically evaluated. Subsequently, the effects of SLC39A14 overexpression and knockdown on PCa cell proliferation and motility were, respectively, examined by Cell Counting Kit-8, transwell, and wound-healing assays. RESULTS: The immunoreactive scores of SLC39A14 protein in human PCa tissues were significantly lower than those in normal prostate tissues. Based on the Taylor dataset, SLC39A14 downregulation occurred more frequently in patients with PCa with a higher Gleason score (P<0.001), advanced clinical stage (P=0.008), presence of metastasis (P=0.009), and prostate-specific antigen failure (P=0.006). More interestingly, the survival analysis identified SLC39A14 as an independent factor for predicting the biochemical recurrence-free survival of patients with PCa (P=0.017). Functionally, the enforced expression of SLC39A14 could suppress cell proliferation, invasion, and migration of PCa cell lines in vitro, which could be reversed by the knockdown of SLC39A14. CONCLUSION: Decreased expression of SLC39A14 may lead to malignant phenotypes of PCa cells and aggressive tumor progression in patients with PCa. Importantly, SLC39A14 may function as a tumor suppressor and a biomarker for screening patients with biochemical recurrence following radical prostatectomy.