Zhenheng Wang

Particle-induced osteolysis mediated by endoplasmic reticulum stress in prosthesis loosening

We hypothesized that endoplasmic reticulum (ER) stress in macrophages induced by wear particles was one of the reasons for particle-induced osteolysis (PIO) in total hip arthroplasty (THA) failure. In the present study, the expression of ER stress markers was examined by Western blot in macrophages treated with particles from materials used in prosthetics, specimens from PIO animal models and patients suffering from aseptic loosening. To address whether ER stress triggers these inflammatory responses, the effect of an ER stress blocker on the expression of inflammatory cytokines in particle-treated macrophages and PIO animal models was tested. The results demonstrated that ER stress markers were significantly upregulated in particle-treated macrophages, periosteum tissues from PIO animal models and clinical specimens of prosthesis loosening. Blocking ER stress with a specific inhibitor dramatically reduced the particle-induced expression of inflammatory cytokines in vitro and in vivo. Furthermore, in PIO animal models, this ER stress blocker dramatically suppressed the differentiation of osteoclasts and reduced the severity of osteolysis. Thus, the results of the present study suggest that ER stress plays a key role in particle-induced osteolysis and that targeting the ER stress pathway may lead to novel therapeutic approaches for the treatment of aseptic prosthesis loosening.

Specifically Formed Corona on Silica Nanoparticles Enhances Transforming Growth Factor β1 Activity in Triggering Lung Fibrosis

A corona is a layer of macromolecules formed on a nanoparticle surface in vivo. It can substantially change the biological identity of nanomaterials and possibly trigger adverse responses from the body tissues. Dissecting the role of the corona in the development of a particular disease may provide profound insights for understanding toxicity of nanomaterials in general. In our present study, we explored the capability of different silica nanoparticles (SiNPs) to induce silicosis in the mouse lung and analyzed the composition of coronas formed on these particles. We found that SiNPs of certain size and surface chemistry could specifically recruit transforming growth factor β1 (TGF-β1) into their corona, which subsequently induces the development of lung fibrosis. Once embedded into the corona on SiNPs, TGF-β1 was remarkably more stable than in its free form, and its fibrosis-triggering activity was significantly prolonged. Our study meaningfully demonstrates that a specific corona component on a certain nanoparticle could initiate a particular pathogenic process in a clinically relevant disease model. Our findings may shed light on the understanding of molecular mechanisms of human health risks correlated with exposure to small-scale substances.

SIRT1 protects osteoblasts against particle-induced inflammatory responses and apoptosis in aseptic prosthesis loosening

We hypothesized that SIRT1 downregulation in osteoblasts induced by wear particles was one of the reasons for particle-induced osteolysis (PIO) in total joint arthroplasty failure. In the present study, the expression of SIRT1 was examined in osteoblasts treated with TiAl6V4 particles (TiPs) and CoCrMo particles (CoPs) from materials used in prosthetics and specimens from PIO animal models. To address whether SIRT1 downregulation triggers inflammatory responses and apoptosis in osteoblasts, the effect of a SIRT1 activator, resveratrol on the expression of inflammatory cytokines and apoptosis in particle-treated osteoblasts was tested. The results demonstrated that SIRT1 expression was significantly downregulated in particle-treated osteoblasts and PIO animal models. Both pharmacological activation and overexpression of SIRT1 dramatically reduced the particle-induced expression of inflammatory cytokines and osteoblast apoptosis through NF-κB and p53 signaling, respectively. Furthermore, in PIO animal models, resveratrol significantly reduced the severity of osteolysis. Collectively, the results of the present study indicated that SIRT1 plays a vital role in the pathogenesis of aseptic loosening, and further treatment targeted at SIRT1 possibly lead to novel approaches for prevention of aseptic prosthesis loosening. STATEMENT OF SIGNIFICANCE Aseptic loosening is the most common cause of total hip arthroplasty (THA) and total knee arthroplasty (TKA) failure and revision surgery. However, there is still no effective therapeutic target in the clinical treatment. Besides, the underlying mechanism of aseptic loosening is largely unknown. The result of our study indicated that SIRT1 has the ability to effectively regulate the wear particle-induced inflammatory responses, apoptosis, osteolysis in particle-stimulated osteoblasts and particle-induced osteolysis animal models. Our study provides a potential target for the prevention and treatment of aseptic loosening and further investigated the underlying mechanism of aseptic loosening, which may make contribution to decrease the incidence of THA and TKA failure in the clinical practice.

ER Stress Mediates TiAl6V4 Particle-Induced Peri-Implant Osteolysis by Promoting RANKL Expression in Fibroblasts.

Wear particle-induced osteolysis is a major cause of aseptic loosening, which is one of the most common reasons for total hip arthroplasty (THA) failure. Previous studies have shown that the synovial fibroblasts present in the periprosthetic membrane are important targets of wear debris during osteolysis. However, the interaction mechanisms between the wear debris and fibroblasts remain largely unknown. In the present study, we investigated the effect of ER (endoplasmic reticulum) stress induced by TiAl6V4 particles (TiPs) in human synovial fibroblasts and calvarial resorption animal models. The expression of ER stress markers, including IRE1-α, GRP78/Bip and CHOP, were determined by western blot in fibroblasts that had been treated with TiPs for various times and concentration. To address whether ER stress was involved in the expression of RANKL, the effects of ER stress blockers (including 4-PBA and TUDCA) on the expression of RANKL in TiPs-treated fibroblasts were examined by real-time PCR, western blot and ELISA. Osteoclastogenesis was assessed by tartrate resistant acid phosphatase (TRAP) staining. Our study demonstrated that ER stress markers were markedly upregulated in TiPs-treated fibroblasts. Blocking ER stress significantly reduced the TiPs-induced expression of RANKL both in vitro and in vivo. Moreover, the inhibition of ER stress ameliorated wear particle-induced osteolysis in animal models. Taken together, these results suggested that the expression of RANKL induced by TiPs was mediated by ER stress in fibroblasts. Therefore, down regulating the ER stress of fibroblasts represents a potential therapeutic approach for wear particle-induced periprosthetic osteolysis.

The metal nanoparticle-induced inflammatory response is regulated by SIRT1 through NF-κB deacetylation in aseptic loosening

Aseptic loosening is the most common cause of total hip arthroplasty (THA) failure, and osteolysis induced by wear particles plays a major role in aseptic loosening. Various pathways in multiple cell types contribute to the pathogenesis of osteolysis, but the role of Sirtuin 1 (SIRT1), which can regulate inflammatory responses through its deacetylation, has never been investigated. We hypothesized that the downregulation of SIRT1 in macrophages induced by metal nanoparticles was one of the reasons for osteolysis in THA failure. In this study, the expression of SIRT1 was examined in macrophages stimulated with metal nanoparticles from materials used in prosthetics and in specimens from patients suffering from aseptic loosening. To address whether SIRT1 downregulation triggers these inflammatory responses, the effects of the SIRT1 activator resveratrol on the expression of inflammatory cytokines in metal nanoparticle-stimulated macrophages were tested. The results demonstrated that SIRT1 expression was significantly downregulated in metal nanoparticle-stimulated macrophages and clinical specimens of prosthesis loosening. Pharmacological activation of SIRT1 dramatically reduced the particle-induced expression of inflammatory cytokines in vitro and osteolysis in vivo. Furthermore, SIRT1 regulated particle-induced inflammatory responses through nuclear factor kappa B (NF-κB) acetylation. Thus, the results of this study suggest that SIRT1 plays a key role in metal nanoparticle-induced inflammatory responses and that targeting the SIRT1 pathway may lead to novel therapeutic approaches for the treatment of aseptic prosthesis loosening.

Probiotics protect mice from CoCrMo particles-induced osteolysis

Wear particle-induced inflammatory osteolysis is the primary cause of aseptic loosening, which is the most common reason for total hip arthroplasty (THA) failure in the med- and long term. Recent studies have suggested an important role of gut microbiota (GM) in modulating the host metabolism and immune system, leading to alterations in bone mass. Probiotic bacteria administered in adequate amounts can alter the composition of GM and confer health benefits to the host. Given the inflammatory osteolysis that occurs in wear debris-induced prosthesis loosening, we examined whether the probiotic Lactobacillus casei could reduce osteolysis in a mouse calvarial resorption model. In this study, L. casei markedly protected mice from CoCrMo particles (CoPs)-induced osteolysis. Osteoclast gene markers and the number of osteoclasts were significantly decreased in L. casei-treated mice. Probiotic treatment decreased the M1-like macrophage phenotype indicated by downregulation of tumor necrosis factor α (TNF-α), interleukin (IL)-6 and inducible nitric oxide synthase (iNOS) and increased the M2-like macrophage phenotype indicated by upregulation of IL-4, IL-10 and arginase. Collectively, these results indicated that the L. casei treatment modulated the immune status and suppressed wear particle-induced osteolysis in vivo. Thus, probiotic treatment may represent a potential preventive and therapeutic approach to reduced wear debris-induced osteolysis.

Autophagy mediated TiAl 6 V 4 particle-induced peri-implant osteolysis by promoting expression of TNF-α

Peri-prosthetic osteolysis and the consequent aseptic loosening constitute the most common reason for total joint arthroplasty failure and surgical revision. Although numerous studies suggest that pro-inflammatory cytokines induced by wear particles is involved in the pathological process of aseptic loosening, the underlying mechanism linking wear particles to pro-inflammatory cytokines remains to be illustrated. In the present study, we investigated the effect of autophagy on TNF-α secretion induced by TiAl6V4 particles (TiPs) in macrophages and in a calvarial resorption animal model. Our study demonstrated that TiPs activated autophage in macrophages and particle-induced osteolysis animal models as well as periprosthetic membranes of patients with aseptic loosening. The autophagy inhibitor 3-MA (3-methyladenine) could dramatically reduce TiPs-induced TNF-α expression both in macrophages and in membranes from animal models. Furthermore, inhibition of autophagy with 3-MA ameliorated the severity of osteolysis in PIO animal models. Collectively, these results suggest that autophagy plays a key role in TiPs-induced osteolysis by promoting TNF-α expression and that blocking autophagy may represent a potential therapeutic approach for treating particle-induced peri-implant osteolysis.

Enhanced Uptake of Fe3O4 Nanoparticles by Intestinal Epithelial Cells in a State of Inflammation.

Fe3O4 nanoparticles (Fe3O4 NPs) have been used for medical and drug applications, although the mechanisms of cellular uptake and transport need to be further evaluated under inflammatory conditions. In the present study, we investigated the uptake of Fe3O4 NPs (20, 50, 100, and 200 nm) by intestinal epithelial cells under inflammatory conditions via the light scattering of flow cytometry and inductively coupled plasma mass spectrometry (ICP-MS) techniques. The results of the correlation analysis indicated that the uptake ratios of Fe3O4 NPs by intestinal epithelial cells under inflammatory conditions were higher than those under the control conditions. The transportation ratios of NPs by inflammatory Caco-2 cells increased almost 0.8–1.2 fold compared to the control. The internalization of the Fe3O4 NPs in Caco-2 cells was mediated by clathrin-related routes in both the control and an interleukin-1β (IL-1β)-induced inflammatory condition. The level of mRNA of clathrin expressed in Caco-2 cells that were stimulated by IL-1β was almost three times more than the control. Consistently with the mRNA expression, the level of protein in the clathrin was upregulated. Additionally, it was verified for the first time that the expression of clathrin was upregulated in IL-1β-stimulated Caco-2 cells. Collectively, these results provided a further potential understanding about the mechanism of Fe3O4 NPs’ uptake by intestinal epithelial cells under inflammatory conditions.

Proanthocyanidins Attenuation of H2O2-Induced Oxidative Damage in Tendon-Derived Stem Cells via Upregulating Nrf-2 Signaling Pathway

Proanthocyanidins (PCs) have shown inhibition of oxidative damage by improving Nrf-2 expression in many tissues. However, the cytoprotective effects of PCs on H2O2-induced tendon damage have not been verified. The current study was aimed at assessing the cytoprotection of PCs on the oxidative cellular toxicity of tendon-derived stem cells (TDSCs) induced by H2O2. The TDSCs were isolated from patellar tendons of Sprague Dawley (SD) rats, and the cells after third passage were used for subsequent experiments. The isolated cells were identified by flow cytometry assay and multidifferentiation potential assay. Cell Counting Kit-8 assay was performed to examine cell viability. Real-Time PCR and Western Blot were employed to, respectively, assess the mRNA and protein expressions of Nrf-2, GCLM, NQO-1, and HO-1. PCs significantly improved the cell viability of TDSCs. Furthermore, H2O2 upregulated Nrf-2, GCLM, NQO-1, and HO-1 without significant difference, while the proteins expressions were increased with significant difference in PCs group and PCs + H2O2 cotreated group. All the findings indicated that PCs could protect against the oxidative damage induced by H2O2 in TDSCs, and the cytoprotective effects might be due to the ability of PCs to activate the expressions of GCLM, HO-1, and NQO-1 via upregulating Nrf-2 signaling pathway.

Expression of XBP1s in fibroblasts is critical for TiAl6 V4 particle-induced RANKL expression and osteolysis.

Wear particle‐induced osteolysis is a major cause of aseptic loosening, which is one of the most common reasons for total hip arthroplasty (THA) failure. Previous studies have shown that the expression of Receptor activation of nuclear factor (NF)‐kB (RANKL) by fibroblasts in periprosthetic membrane played a crucial role in wear particle‐induced osteolysis. However, the underlying mechanism of RANKL expression remains largely unknown. In the present study, we investigated the effect of TiAl6V4 particle (TiPs)‐induced XBP1s (spliced form of X‐box binding protein 1) on RANKL expression and osteoclastogenesis both in vitro and in vivo. The levels of XBP1s in peri‐implant membrane, animal models, and TiPs‐stimulated fibroblasts were determined by western blots. To assess the effect of XBP1s on RANKL expression, fibroblasts were treated with both a small interfering RNA (siRNA) and an inhibitor of XBP1 prior to exposure to TiPs. The effect of XBP1s on osteoclasts formation was determined by tartrate‐resistant acid phosphatase (TRAP) staining in vitro osteoclastogenesis assay and in animal models. The resorption of bone was assessed by micro‐computed tomography (micro‐CT) with three‐dimensional reconstruction. Our results demonstrated that XBP1s was activated in periprosthetic membrane, mouse calvaria models, and TiPs‐stimulated human synovial fibroblasts. Further, inhibition of XBP1s decreased the expression of RANKL and osteoclasts formation in vitro. In mouse calvaria models, both of the osteoclastogenesis and osteolysis were inhibited XBP1s inhibitor. Our results suggested that XBP1s mediated TiPs‐induced of RANKL expression in fibroblasts, and down regulating XBP1s may represent a potential therapy for wear particle‐induced osteolysis.