Effect of Diabetes Mellitus on Implant Osseointegration of Titanium Screws: An Animal Experimental Study.

OBJECTIVE: To explore the effect of diabetes mellitus (DM) on implant osseointegration of titanium screws. METHODS: Sixty rats were randomly divided into a DM group and a control group (each group, n = 30). DM group rats were injected with 1% Streptozotocin solution at 65 mg/kg to establish a DM model. Titanium screws were implanted into the rats' distal femurs in both groups. The rats were sacrificed for micro-CT scanning, micro-indentation, biomechanical detection, confocal Raman microspectroscopy, and histological and histomorphometric analysis at 4, 8, and 12 weeks post-implantation, respectively. Messenger RNA (mRNA) expression and protein expression of the related growth factors around the implant were analyzed using real-time polymerase chain reaction and Western blots. RESULTS: At 4, 8 and 12 weeks, micro-CT scanning, hematoxylin-eosin (HE) staining, Gieson's acid-magenta staining, and fluorescent labeled staining showed disorder in the bone tissue arrangement, a lack of new bone tissue, poor maturity and continuity, and poor trabecular bone parameters around the implant in the DM group. At 4, 8, and 12 weeks, the interfacial bone binding rate in the DM group was significantly lower (16.2% ± 4.8%, 25.7% ± 5.7%, 42.5% ± 5.8%, respectively) than that in the control group (23.6% ± 5.2%, 40.8% ± 6.3%, 64.2% ± 7.3%, respectively; P < 0.05). At 8 and 12 weeks, the elastic modulus (17.0 ± 1.8 and 15.1 ± 1.5 GPa, respectively) and trabecular bone hardness (571 ± 39 and 401 ± 37 MPa, respectively) in the DM group were significantly lower than the elastic modulus (23.4 ± 2.3 and 23.8 ± 1.8 GPa, respectively) and trabecular bone hardness (711 ± 45 and 719 ± 46 MPa, respectively) in the control group (P < 0.05). The maximum load required for the prosthesis pull-out experiment in the DM group at 4, 8, and 12 weeks (55.14 ± 6.74 N, 73.34 ± 8.43 N, and 83.45 ± 8.32 N, respectively) was significantly lower than that in the control group (77.45 ± 7.48 N, 93.28 ± 8.29 N, and 123.62 ± 9.43 N, respectively, P < 0.05). At 8 and 12 weeks, the mineral-to-collagen ratio in the DM group (6.56 % ± 1.35% and 4.45%± 1.25%, respectively) was significantly higher than that in the control group (5.31% ± 1.42% and 3.62% ± 1.33%, respectively, P < 0.05). At 12 weeks, mRNA and protein expression levels of bone morphogenetic protein 2, transforming growth factor-ß1, vascular endothelial growth factor, osteopontin, osteocalcin, and runt-related transcription factor 2 in the DM group were significantly lower than that in the control group. CONCLUSIONS: DM can negatively affect bone osseointegration, manifesting as disorder in bone tissue arrangement around the implant, a lack of new bone tissue, poor maturity and continuity, poor trabecular bone parameters and lower expression of the related growth factors.

Long noncoding RNA H19 accelerates tenogenic differentiation by modulating miR-140-5p/VEGFA signaling.

Rotator cuff tear (RCT) is a common tendon injury, but the mechanisms of tendon healing remain incompletely understood. Elucidating the molecular mechanisms of tenogenic differentiation is essential to develop novel therapeutic strategies in clinical treatment of RCT. The long noncoding RNA H19 plays a regulatory role in tenogenic differentiation and tendon healing, but its detailed mechanism of action remains unknown. To elucidate the role of H19 in tenogenic differentiation and tendon healing, tendon-derived stem cells were harvested from the Achilles tendons of Sprague Dawley rats and a rat model of cuff tear was established for the exploration of the function of H19 in promoting tenogenic differentiation. The results showed that H19 overexpression promoted, while H19 silencing suppressed, tenogenic differentiation of tendon-derived stem cells (TDSCs). Furthermore, bioinformatic analyses and a luciferase reporter gene assay showed that H19 directly targeted and inhibited miR-140-5p to promote tenogenic differentiation. Further, inhibiting miR-140-5p directly increased VEGFA expression, revealing a novel regulatory axis between H19, miR-140-5p, and VEGFA in modulating tenogenic differentiation. In rats with RTC, implantation of H19-overexpressing TDSCs at the lesion promoted tendon healing and functional recovery. In general, the data suggest that H19 promotes tenogenic differentiation and tendon-bone healing by targeting miR-140-5p and increasing VEGFA levels. Modulation of the H19/miR-140-5p/VEGFA axis in TDSCs is a new potential strategy for clinical treatment of tendon injury.

Effect of Conditioned Medium from Human Umbilical Cord-Derived Mesenchymal Stromal Cells on Rejuvenation of Nucleus Pulposus Derived Stem/Progenitor Cells from Degenerated Intervertebral Disc.

BACKGROUND AND OBJECTIVES: Mesenchymal stromal cells (MSCs)-based treatment for degeneration of intervertebral disc (IVD) has been proposed recently. We here addressed whether MSC secreted factors can rejuvenate nucleus pulposus- derived stem/progenitor cells from degenerated disc (D-NPSCs) in vitro. METHODS AND RESULTS: We analyzed the expression of MSCs and NP cell specific surface markers, pluripotency related genes, multilineage potential and cell proliferative capacity of D-NPSCs upon incubation with the conditioned medium which was collected from the umbilical cord derived MSCs (UCMSCs). Our results indicated that the conditioned medium restore the stemness of D-NPSCs by up-regulating the expression level of CD29 and CD105, pluripotency related genes OCT4 and Nanog, and NP progenitor marker Tie2. The increased stemness was accompanied by promoted cell proliferative capacity and improved osteogenic and chondrogenic differentiation potential. CONCLUSIONS: Our findings suggested that the UCMSCs derived conditioned medium might be used to rejuvenate the degenerated NP stem/progenitor cells.

HSP90 regulates osteosarcoma cell apoptosis by targeting the p53/TCF-1-mediated transcriptional network.

Osteosarcoma (OS) is the most common bone tumor that occurs predominantly in children and teenagers. Although many genes, such as p53 and Rb1, have been shown to be mutated, deregulation of the canonical Wnt/ß-catenin signaling pathway is frequently observed in OS. We recently demonstrated that heat shock protein 90 (HSP90) is involved in the regulation of runt-related transcription factor 2 via the AKT/GSK-3ß/ß-catenin signaling pathway in OS. However, the precise role of T cell factors/lymphoid enhancer-binding factor (TCFs/LEF) family members, which are the major binding complex of ß-catenin, in OS is poorly understood. In the present study, we first demonstrated that TCF-1 is overexpressed in OS compared with other bone tumors. Knockdown of TCF-1 significantly induced cell cycle arrest, severe DNA damage, and subsequent caspase-3-dependent apoptosis. Interestingly, coexpression of HSP90 and TCF-1 was observed in OS, and mechanistically, we demonstrated that TCF-1 expression is regulated by HSP90 either through a ß-catenin-dependent mechanism or a direct degradation of the proteasome. We also found that overexpression of TCF-1 partially abolishes the apoptosis induced by HSP90 inhibition. Furthermore, we provided evidence that p53, but not miR-34a, plays a crucial role in the HSP90-regulated TCF-1 expression and subsequent apoptosis. Given the diverse combination regimens of HSP90 inhibition with some other treatments, we propose that the p53 status and the expression level of TCF-1 should be taken into consideration to enhance the therapeutic efficacy of HSP90 inhibition.

Regenerative potential of human nucleus pulposus resident stem/progenitor cells declines with ageing and intervertebral disc degeneration.

Numerous studies have demonstrated the presence of resident nucleus pulposus stem/progenitor cells (NPSCs) in the tissue of the intervertebral disc (IVD). However, the cellular identity of NPSCs during IVD degeneration and ageing are poorly defined at present, despite significant progress in the understanding of NPSC biology. In the present study, NPSCs were isolated from human degenerated IVD and were characterized by flow cytometry, gene expression assays and proliferation and multipotency analysis. The results of the present study demonstrated that NPSCs isolated from human degenerated IVD may be divided into two groups according to the expression of mesenchymal stem cell (MSC) surface markers: The high expression of MSC surface markers group (H­NPSCs) was highly positive for CD29, CD44, CD73, CD90 and CD105 at rates >95%, and the low expression of MSC markers surface markers group (L­NPSCs), with the expression of CD29 and CD105 exhibiting individual variability, however, all at rates <95%. The donors for H­NPSCs were aged <20 years, while the majority of donors for L­NPSCs were aged >25 years, with one exception aged <20 years. The results highlighted that the low expression of MSC surface markers in NPSCs from aged and degenerated NP tissues were associated with a low rate of proliferation and reduced differentiation potential, as well as downregulation of the NP progenitor marker Tie2 and higher expression of NP cell­specific markers. These findings demonstrated that the regenerative potential of human NPSCs declines with ageing and degeneration of the IVD.

Comparison of nucleus pulposus stem/progenitor cells isolated from degenerated intervertebral discs with umbilical cord derived mesenchymal stem cells.

Mesenchymal stem-cell based therapies have been proposed as novel treatments for intervertebral disc (IVD) degeneration. The development of these treatment strategies, however, has been hindered by the incomplete understanding of the origin, biological properties of nucleus pulposus (NP) derived stem/progenitor cells and their effects on the IVD degeneration. The goal of this study is to explore the biological properties of NP stem/progenitor cells isolated from degenerated IVD (D-NPMSCs) regarding immunotype, proliferative capacity, multi-lineage differentiation abilities, and the expression of NP specific cell surface markers compared to human umbilical cord mesenchymal stem cells (UCMSCs). Our results indicate that although D-NPMSCs shared the mesenchymal stromal cells (MSCs) characteristics with UCMSCs, significant differences exist in phenotype signatures and biological capacities between D-NPMSCs and UCMSCs. D-NPMSCs expressed lower expression levels of CD29 and CD105, the phenotype markers of MSCs, and exhibited reduced proliferation capability and differentiation potentials, which might account for the distinct NP microenvironment and the poor capacity for disc regeneration. This study will lay a foundation for further understanding the mechanism of stem cell-based therapy for IVD degeneration.

The effects of liquid crystal-based composite substrates on cell functional responses of human umbilical cord-derived mesenchymal stem cells by mechano-regulatory process.

Physical properties of extracellular matrix, including elasticity and microstructure, have been considered as important factors inducing stem cell differentiation. This study developed a novel type of liquid crystal-based matrix by combining the elastic property of polyurethane with viscoelastic liquid crystal to generate a soft elastic response resembling physical microenvironment of stem cell niche, and explored the mechano-driving cell behaviors. Addition of varying liquid crystal concentration (10 wt%, 30 wt% and 50 wt%) had great effects on surface morphology and elastic modulus of liquid crystal/ polyurethane composite substrates. Changes in microstructure and elastic modulus of the substrates could cause intense cell responses that influenced cell properties, including proliferation, adhesion, and osteogenic differentiation. Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) cultured on both liquid crystal-10/polyurethane and liquid crystal-30/polyurethane substrates exhibited higher viability, more actin filament, and larger spreading area while liquid crystal-50/polyurethane substrates seemed not to favor cell attachment and spreading. Alkaline phosphatase activity and calcium deposition were significantly improved with hUC-MSCs on both liquid crystal-10/ polyurethane and liquid crystal-30/ polyurethane substrates, and the maximal alkaline phosphatase activity was observed in liquid crystal-10/ polyurethane while the lowest in liquid crystal-50/ polyurethane. Osteopontin was upregulated to a high level in both liquid crystal-10/ polyurethane and liquid crystal-30/ polyurethane groups after 14 days culturing; the maximal expression of osteocalcin and related transcription factor 2 were found in liquid crystal-10/ polyurethane group on day 21. Our findings revealed that hUC-MSCs could intensely sense the bioactive patterns and soft-matter feature of liquid crystal domains and subsequently modulated cell behaviors, which may prove useful in the development of new class of biomaterials for applications in tissue engineering and regenerative medicine.