Lianfu Deng

Upregulating Hif-1α by Hydrogel Nanofibrous Scaffolds for Rapidly Recruiting Angiogenesis Relative Cells in Diabetic Wound.

Nonhealing chronic wounds on foot are one of the most dreaded complications of diabetes, and biomedical scaffolds remain an attractive option for repairing or regenerating tissues. Accelerating angiogenesis in the early stage after injury is critical to wound healing process; however, the scaffolds accelerate the angiogenesis in the beginning but with the acceleration of vessel network formation the scaffold network hinders the process. In this study, the water soluble drugs-loaded hydrogel nanofibrous scaffolds are designed for rapidly recruiting angiogenesis relative cells and promoting wound healing. The sustained release profile of desferrioxamine (DFO), which continues for about 72 h, leads to significantly increase of neovascularization. The majority of the scaffold is degraded in 14 d, leaving enough space for cell proliferation and vessel formation. The in vitro results show that the scaffolds upregulate the expression of Hif-1α and vascular endothelial growth factor, and enhance the interaction between fibroblasts and endothelial cells. The in vivo studies show a higher expression of angiogenesis related cytokines. This study demonstrates that the DFO released from hydrogel nanofibrous scaffolds of quick degradation can interfere with the required prolyl-hydroxylases cofactors by acting as Fe(2+) chelator and upregulate the expression of Hif-1α, leading to a significant increase of the neovascularization.

Heme oxygenase-1 attenuates IL-1β induced alteration of anabolic and catabolic activities in intervertebral disc degeneration

Intervertebral disc degeneration (IDD) is characterized by disordered extracellular matrix (ECM) metabolism, implicating subdued anabolism and enhanced catabolic activities in the nucleus pulposus (NP) of discs. Pro-inflammatory cytokines such as interleukin-1β (IL-1β) are considered to be potent mediators of ECM breakdown. Hemeoxygenase-1 (HO-1) has been reported to participate in cellular anti-inflammatory processes. The purpose of this study was to investigate HO-1 modulation of ECM metabolism in human NP cells under IL-1β stimulation. Our results revealed that expression of HO-1 decreased considerably during IDD progression. Induction of HO-1 by cobalt protoporphyrin IX attenuated the inhibition of sulfate glycosaminoglycan and collagen type II (COL-II) synthesis and ameliorated the reduced expressions of aggrecan, COL-II, SOX-6 and SOX-9 mediated by IL-1β. Induction of HO-1 also reversed the effect of IL-1β on expression of the catabolic markers matrix metalloproteinases-1, 3, 9 and 13. This was combined with inhibition of the activation of mitogen-activated protein kinase signaling. These findings suggest that HO-1 might play a pivotal role in IDD, and that manipulating HO-1 expression might mitigate the impairment of ECM metabolism in NP, thus potentially offering a novel therapeutic approach to the treatment of IDD.

Increased EZH2 and decreased osteoblastogenesis during local irradiation-induced bone loss in rats

Radiation therapy is commonly used to treat cancer patients but exhibits adverse effects, including insufficiency fractures and bone loss. Epigenetic regulation plays an important role in osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). Here, we reported local bone changes after single-dose exposure to 137CS irradiation in rats. Femur bone mineral density (BMD) and trabecular bone volume in the tibia were significantly decreased at 12 weeks after irradiation. Micro-CT results showed that tBMD, Tb.h and Tb.N were also significantly reduced at 12 weeks after irradiation exposure. ALP-positive OB.S/BS was decreased by 42.3% at 2 weeks after irradiation and was decreased by 50.8% at 12 weeks after exposure. In contrast to the decreased expression of Runx2 and BMP2, we found EZH2 expression was significantly increased at 2 weeks after single-dose 137CS irradiation in BMSCs. Together, our results demonstrated that single-dose 137CS irradiation induces BMD loss and the deterioration of bone microarchitecture in the rat skeleton. Furthermore, EZH2 expression increased and osteoblastogenesis decreased after irradiation. The underlying mechanisms warrant further investigation.

Desferrioxamine reduces ultrahigh-molecular-weight polyethylene-induced osteolysis by restraining inflammatory osteoclastogenesis via heme oxygenase-1

As wear particles-induced osteolysis still remains the leading cause of early implant loosening in endoprosthetic surgery, and promotion of osteoclastogenesis by wear particles has been confirmed to be responsible for osteolysis. Therapeutic agents targeting osteoclasts formation are considered for the treatment of wear particles-induced osteolysis. In the present study, we demonstrated for the first time that desferrioxamine (DFO), a powerful iron chelator, could significantly alleviate osteolysis in an ultrahigh-molecular-weight polyethylene (UHMWPE) particles-induced mice calvaria osteolysis model. Furthermore, DFO attenuated calvaria osteolysis by restraining enhanced inflammatory osteoclastogenesis induced by UHMWPE particles. Consistent with the in vivo results, we found DFO was also able to inhibit osteoclastogenesis in a dose-dependent manner in vitro, as evidenced by reduction of osteoclasts formation and suppression of osteoclast specific genes expression. In addition, DFO dampened osteoclasts differentiation and formation at early stage but not at late stage. Mechanistically, the reduction of osteoclastogenesis by DFO was due to increased heme oxygenase-1 (HO-1) expression, as decreased osteoclasts formation induced by DFO was significantly restored after HO-1 was silenced by siRNA, while HO-1 agonist COPP treatment enhanced DFO-induced osteoclastogenesis inhibition. In addition, blocking of p38 mitogen-activated protein kinase (p38MAPK) signaling pathway promoted DFO-induced HO-1 expression, implicating that p38 signaling pathway was involved in DFO-mediated HO-1 expression. Taken together, our results suggested that DFO inhibited UHMWPE particles-induced osteolysis by restraining inflammatory osteoclastogenesis through upregulation of HO-1 via p38MAPK pathway. Thus, DFO might be used as an innovative and safe therapeutic alternative for treating wear particles-induced aseptic loosening.

Activation of HIFa Pathway in Mature Osteoblasts Disrupts the Integrity of the Osteocyte/Canalicular Network

The hypoxia-inducible factors (HIFs), HIF-1α and HIF-2α, are the central mediators of the homeostatic response that enables cells to survive and differentiate in low-oxygen conditions. Previous studies indicated that disruption of the von Hippel-Lindau gene (Vhl) coincides with the activation of HIFα signaling. Here we show that inactivation of Vhl in mature osteoblasts/osteocytes induces their apoptosis and disrupts the cell/canalicular network. VHL-deficient (ΔVHL) mice exhibited a significantly increased cortical bone area resulting from enhanced proliferation and osteogenic differentiation of the bone marrow stromal cells (BMSCs) by inducing the expression of β-catenin in the BMSC. Our data suggest that the VHL/HIFα pathway in mature osteoblasts/osteocytes plays a critical role in the bone cell/canalicular network and that the changes of osteocyte morphology/function and cell/canalicular network may unleash the bone formation, The underlying mechanism of which was the accumulation of β-catenin in the osteoblasts/osteoprogenitors of the bone marrow.

Inflammatory microRNA-194 and -515 attenuate the biosynthesis of chondroitin sulfate during human intervertebral disc degeneration

Intervertebral disc degeneration (IDD) is characterized by dehydration and loss of extracellular matrixes in the nucleus pulposus region. Chondroitin sulfate has been found to be the water-binding molecule that played a key role in IDD. Although investigators have reported that inflammatory cytokines are involved in the reduction of chondroitin sulfate in IDD, but the underlying mechanism is unrevealed. Since chondroitin sulfate synthesis is controlled by chondroitin sulfate glycosyltransferases CHSY-1/2/3 and CSGALNACT-1/2, their functional role and regulatory mechanism in IDD is not fully studied. Here, we set out to investigate the function and regulatory roles of these factors during IDD development. We found that among these chondroitin sulfate glycosyltransferases, CHSY-1/2/3 are significantly down-regulated in severe IDD samples than mild IDD samples. In vitro experiments revealed that Interleukin-1β and Tumor Necrosis Factor-α stimulation led to significant reduction of CHSY-1/2/3 at protein level than mRNA level in NP cells, indicating a post-transcriptional regulatory mechanisms are involved. By computational prediction and analysis, we found that inflammatory cytokines stimulated microRNA-194 and -515 target CHSY-1/2/3 mRNA and significantly interrupt their translation and downstream chondroitin sulfate deposition. Inhibition of microRNA-194 and -515 however, significantly rescued CHSY-1/2/3 expressions and chondroitin sulfate deposition. These findings together demonstrated a vital role of inflammatory stimulated microRNAs in promoting intervertebral disc degeneration by interrupt chondroitin sulfate synthesis, which may provide new insights into the mechanism and therapeutic approaches in IDD.

Strontium Ranelate Reduces the Fracture Incidence in a Growing Mouse Model of Osteogenesis Imperfecta.

Osteogenesis imperfecta (OI) is a genetic bone dysplasia characterized by brittle bones with increased fracture risk. Although current treatment options to improve bone strength in OI focus on antiresorptive bisphosphonates, controlled clinical trials suggest they have an equivocal effect on reducing fracture risk. Strontium ranelate (SrR) is a promising therapy with a dual mode of action that is capable of simultaneously maintaining bone formation and reducing bone resorption, and may be beneficial for the treatment of OI. In this study, SrR therapy was investigated to assess its effects on fracture frequency and bone mass and strength in an animal model of OI, the oim/oim mouse. Three‐week‐old oim/oim and wt/wt mice were treated with either SrR or vehicle (Veh) for 11 weeks. After treatment, the average number of fractures sustained by SrR‐treated oim/oim mice was significantly reduced compared to Veh‐treated oim/oim mice. Micro–computed tomographic (μCT) analyses of femurs showed that both trabecular and cortical bone mass were significantly improved with SrR treatment in both genotypes. SrR significantly inhibited bone resorption, whereas bone formation indices were maintained. Biomechanical testing revealed improved bone structural properties in both oim/oim and wild‐type (wt/wt) mice under the treatment, whereas no significant effects on bone brittleness and material quality were observed. In conclusion, SrR was able to effectively reduce fractures in oim/oim mice by improving bone mass and strength and thus represents a potential therapy for the treatment of pediatric OI.

Double-stranded RNA released from damaged articular chondrocytes promotes cartilage degeneration via Toll-like receptor 3-interleukin-33 pathway.

Pattern recognition receptors (PRRs), including Toll-like receptor 3 (TLR3), are involved in arthritic responses; however, whether interleukin-33 (IL-33) is involved in TLR3-mediated cartilage degeneration is unknown. Here, we found that IL-33 was abundantly increased in chondrocytes of osteoarthritis, especially the chondrocytes of weight-bearing cartilage. Furthermore, double-stranded RNA (dsRNA) released from damaged articular chondrocytes induced by mechanical stretching upregulated IL-33 expression to a greater degree than IL-1β and tumor necrosis factor-α. dsRNA induced IL-33 expression via the TLR3-p38 mitogen-activated protein kinase-nuclear factor-κB (NF-κB) pathway. In addition, formation of the p65 and peroxisome proliferator-activated receptor-γ transcriptional complex was required for dsRNA-induced IL-33 expression. IL-33, in turn, acted on chondrocytes to induce matrix metalloproteinase-1/13 and inhibit type II collagen expression. These findings reveal that dsRNA released from damaged articular chondrocytes promotes cartilage degeneration via the TLR3-IL-33 pathway.

Increased 15-lipoxygenase-1 expression in chondrocytes contributes to the pathogenesis of osteoarthritis

15-Lipoxygenase-1 (15-LO-1) is involved in many pathological processes. The purpose of this study was to determine the potential role of 15-LO-1 in osteoarthritis (OA). The levels of 15-LO-1 expression were measured by western blotting and quantitative real-time PCR in articular cartilage from the OA rat models and OA patients. To further investigate the effects of 15-LO-1 on chondrocyte functions, such as extracellular matrix (ECM) secretion, the release of matrix-degrading enzymes, the production of reactive oxygen species (ROS), cell proliferation and apoptosis, we decreased or increased 15-LO-1 expression in chondrocytes by means of transfecting with siRNA targeting 15-LO-1 and plasmid encoding 15-LO-1, respectively. The results showed that 15-LO-1 expression was obviously increased in articular cartilage from OA rats and OA patients. It was also found that many factor-related OA, such as mechanical loading, ROS, SNP and inflammatory factor, significantly promoted 15-LO-1 expression and activity in chondrocytes. Silencing 15-LO-1 was able to markedly alleviate mechanical loading-induced cartilage ECM secretion, cartilage-degrading enzyme secretion and ROS production. Overexpression of 15-LO-1 could inhibit chondrocyte proliferation and induce chondrocyte apoptosis. In addition, reduction of 15-LO-1 in vivo significantly alleviated OA. Taken together, these results indicate that 15-LO-1 has an important role in the disease progression of OA. Thus 15-LO-1 may be a good target for developing drugs in the treatment of OA.

Estrogen inhibits osteoclasts formation and bone resorption via microRNA-27a targeting PPARγ and APC: GUO et al.

Inhibition of osteoclasts formation and bone resorption by estrogen is very important in the etiology of postmenopausal osteoporosis. The mechanisms of this process are still not fully understood. Recent studies implicated an important role of microRNAs in estrogen‐mediated responses in various cellular processes, including cell differentiation and proliferation. Thus, we hypothesized that these regulatory molecules might be implicated in the process of estrogen‐decreased osteoclasts formation and bone resorption. Western blot, quantitative real‐time polymerase chain reaction, tartrate‐resistant acid phosphatase staining, pit formation assay and luciferase assay were used to investigate the role of microRNAs in estrogen‐inhibited osteoclast differentiation and bone resorption. We found that estrogen could directly suppress receptor activator of nuclear factor B ligand/macrophage colony‐stimulating factor‐induced differentiation of bone marrow‐derived macrophages into osteoclasts in the absence of stromal cell. MicroRNA‐27a was significantly increased during the process of estrogen‐decreased osteoclast differentiation. Overexpressing of microRNA‐27a remarkably enhanced the inhibitory effect of estrogen on osteoclast differentiation and bone resorption, whereas which were alleviated by microRNA‐27a depletion. Mechanistic studies showed that microRNA‐27a inhibited peroxisome proliferator‐activated receptor gamma (PPARγ) and adenomatous polyposis coli (APC) expression in osteoclasts through a microRNA‐27a binding site within the 3′‐untranslational region of PPARγ and APC. PPARγ and APC respectively contributed to microRNA‐27a‐decreased osteoclast differentiation and bone resorption. Taken together, these results showed that microRNA‐27a may play a significant role in the process of estrogen‐inhibited osteoclast differentiation and function.