Rspo2 exacerbates rheumatoid arthritis by targeting aggressive phenotype of fibroblast-like synoviocytes and disrupting chondrocyte homeostasis via Wnt/ß-catenin pathway.

BACKGROUND: The aggressive phenotype of fibroblast-like synoviocytes (FLS) has been identified as a contributing factor to the exacerbation of rheumatoid arthritis (RA) through the promotion of synovitis and cartilage damage. Regrettably, there is currently no effective therapeutic intervention available to address this issue. Recent research has shed light on the crucial regulatory role of R-spondin-2 (Rspo2) in cellular proliferation, cartilage degradation, and tumorigenesis. However, the specific impact of Rspo2 on RA remains poorly understood. We aim to investigate the function and mechanism of Rspo2 in regulating the aggressive phenotype of FLS and maintaining chondrocyte homeostasis in the context of RA. METHODS: The expression of Rspo2 in knee joint synovium and cartilage were detected in RA mice with antigen-induced arthritis (AIA) and RA patients. Recombinant mouse Rspo2 (rmRspo2), Rspo2 neutralizing antibody (Rspo2-NAb), and recombinant mouse DKK1 (rmDKK1, a potent inhibitor of Wnt signaling pathway) were used to explore the role and mechanism of Rspo2 in the progression of RA, specifically in relation to the aggressive phenotype of FLS and chondrocyte homeostasis, both in vivo and in vitro. RESULTS: We indicated that Rspo2 expression was upregulated both in synovium and articular cartilage as RA progressed in RA mice and RA patients. Increased Rspo2 upregulated the expression of leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5), as the ligand for Rspo2, and ß-catenin in FLS and chondrocytes. Subsequent investigations revealed that intra-articular administration of rmRspo2 caused striking progressive synovitis and articular cartilage destruction to exacerbate RA progress in mice. Conversely, neutralization of Rspo2 or inhibition of the Wnt/ß-catenin pathway effectively alleviated experimental RA development. Moreover, Rspo2 facilitated FLS aggressive phenotype and disrupted chondrocyte homeostasis primarily through activating Wnt/ß-catenin pathway, which were effectively alleviated by Rspo2-NAb or rmDKK1. CONCLUSIONS: Our data confirmed a critical role of Rspo2 in enhancing the aggressive phenotype of FLS and disrupting chondrocyte homeostasis through the Wnt/ß-catenin pathway in the context of RA. Furthermore, the results indicated that intra-articular administration of Rspo2 neutralizing antibody or recombinant DKK1 might represent a promising therapeutic strategy for the treatment of RA.

LncRNA Neat1 promotes the macrophage inflammatory response and acts as a therapeutic target in titanium particle-induced osteolysis.

Aseptic loosening (AL), secondary to particle-caused periprosthetic osteolysis, is one of the main reasons of artificial joint failure. Suppressing the macrophage inflammatory response caused by wear particles extends the life of prosthesis, and the long noncoding RNAs (lncRNAs) may play a predominant part in it. Here, titanium particles' (TiPs') stimulation increases both the cytoplasmic and nuclear levels of lncRNA Neat1 in bone marrow derived macrophages (BMDMs), which further induces the inflammatory response. Mechanically, Neat1 facilitates Bruton's tyrosine kinase (BTK) transcription by reducing the transcriptional factor KLF4, which further activates the NF-κB pathway, NLRP3 inflammation, and M1 polarization in BMDMs. Cytoplasmic Neat1 also works as an miRNA sponge in miR-188-5p-regulated BTK expression in the post-transcriptional stage. In vivo, Neat1 downregulation can reduce the TiP-induced pro-inflammatory factors and reverse the osteolysis induced by BTK overexpression. In addition, the PLGA-based microparticles loaded with si-Neat1 are developed for the treatment of the mouse calvarial osteolysis model via local injection, presenting satisfactory anti-osteolysis efficacy. These findings indicate that Neat1 is a key regulator of AL. STATEMENT OF SIGNIFICANCE: Due to released particles, aseptic loosening (AL) is the most common reason for prosthesis failure and surgical revision and represents a substantial economic burden worldwide. Herein, we reported that lncRNA Neat1 is a key regulator in regulating wear particles-induced osteolysis by activating NF-κB pathway, NLRP3 inflammation and M1 polarization via BTK, and the underlying mechanisms of Neat1-BTK interaction were further portrayed. For potential clinical application, the microparticles are developed for effective si-Neat1 delivery, leading to a dramatically enhanced effect for the treatment of osteolysis, which might be a novel strategy to extend the life of the implant.

Increased expression of osteopontin in subchondral bone promotes bone turnover and remodeling, and accelerates the progression of OA in a mouse model.

Osteopontin (OPN) has been proved to be closely related to the pathogenesis of osteoarthritis (OA), but the role of OPN in the pathogenesis of OA has not been fully clarified. Current studies on OPN in OA mostly focus on articular cartilage, synovial membrane and articular fluid, while ignoring its role in OA subchondral bone turnover and remodeling. In this study, we used a destabilization OA mouse model to investigate the role of OPN in OA subchondral bone changes. Our results indicate that increased expression of OPN accelerates the turnover and remodeling of OA subchondral bone, promotes the formation of h-type vessels in subchondral bone, and mediates articular cartilage degeneration induced by subchondral bone metabolism. In addition, our results confirmed that inhibition of PI3K/AKT signaling pathway inhibits OPN-mediated OA subchondral bone remodeling and cartilage degeneration. This study revealed the role and mechanism of OPN in OA subchondral bone, which is of great significance for exploring specific biological indicators for early diagnosis of OA and monitoring disease progression, as well as for developing drugs to regulate the metabolism and turnover of subchondral bone and alleviate the subchondral bone sclerosis of OA.

Spermidine activates RIP1 deubiquitination to inhibit TNF-α-induced NF-κB/p65 signaling pathway in osteoarthritis.

Spermidine has been known to inhibit the production of pro-inflammatory cytokines. However, there are no reports about anti-inflammatory effects of spermidine on osteoarthritis (OA). Herein, we examined whether OA progression could be delayed by intraperitoneal injection (i.p.) of spermidine in the anterior cruciate ligament transection (ACLT) and TNF-α induced arthritis (TIA) mouse models. During the process, human FLS cells (H-FLS) were used to investigate the potential ubiquitination mechanism of spermidine-mediated RIP1 in TNF-α-induced NF-κB/p65 signaling. We found that spermidine attenuated synovitis, cartilage degeneration and osteophyte formation, resulting in substantially lower OARSI scores and TNF-α scores in spermidine-treated ACLT and TIA mice. In terms of the mechanism, 9 µM spermidine did not affect the viability, proliferation, cell cycle and apoptosis of H-FLS, and exerted inhibitory effects by activating CYLD-mediated RIP1 deubiquitination on TNF-α-induced NF-κB/p65 signaling in H-FLS. From these data, we can conclude that spermidine attenuates OA progression by the inhibition of TNF-α-induced NF-κB pathway via the deubiquitination of RIP1 in FLS. Therefore, intake of spermidine could be a potential therapy for preventing OA.

Hesperidin inhibits synovial cell inflammation and macrophage polarization through suppression of the PI3K/AKT pathway in complete Freund's adjuvant-induced arthritis in mice.

Rheumatoid arthritis (RA) is an autoimmune disease characterized by synovitis. Synovitis can cause joint injury by releasing inflammatory factors and metalloproteinases (MMPs). Therefore, it is necessary to find drugs that can control synovitis in the process of RA. Herein, we investigate the anti-inflammatory effect of Hesperidin (HSN) on fibroblast-like synovial (FLS) cells induced by lipopolysaccharide (LPS) and the protective action of M1 polarization level of synovial macrophages on antigen-induced arthritis (AIA) in order to elucidate the reduction of inflammatory cytokines and MMPs and the inhibition of macrophage activation. The functional effect of HSN on LPS-induced mRNA and protein expressions of inflammatory cytokines and MMPs in FLS cells as well as on LPS-induced macrophage M1 and M2 polarization markers was determined by quantitative real-time PCR (qPCR) or Western blot analyses, respectively. AIA in 2-month-old mice was generated using intraperitoneal injection with HSN (20 mg/kg/day) or LY294002 (20 mg/kg/day). The results show HSN significantly inhibited the LPS-induced gene expression of the inflammatory mediators. Furthermore, treatment with HSN relieved the antigen-induced arthritis and reduced the protein levels of MMP3, MMP9, and MMP13 in FLS and inhibited the polarization of macrophages to M1. Based on the results of our analyses, we concluded that HSN has significant anti-inflammatory activities and reduces the potential of MMPs in rheumatoid arthritis and the degree of polarization of macrophages to M1. Through the study of signaling pathways, we established that the inhibition of the PI3K/AKT signaling pathway by HSN may show therapeutic effects in the progression of rheumatoid arthritis.

Synovial macrophage M1 polarisation exacerbates experimental osteoarthritis partially through R-spondin-2.

OBJECTIVES: To investigate the roles and regulatory mechanisms of synovial macrophages and their polarisation in the development of osteoarthritis (OA). METHODS: Synovial tissues from normal patients and patients with OA were collected. M1 or M2-polarised macrophages in synovial tissues of patients with OA and OA mice were analysed by immunofluorescence and immunohistochemical staining. Mice with tuberous sclerosis complex 1 (TSC1) or Rheb deletion specifically in the myeloid lineage were generated and subjected to intra-articular injection of collagenase (collagenase-induced osteoarthritis, CIOA) and destabilisation of the medial meniscus (DMM) surgery to induce OA. Cartilage damage and osteophyte size were measured by Osteoarthritis Research Society International score and micro-CT, respectively. mRNA sequencing was performed in M1 and control macrophages. Mice and ATDC5 cells were treated with R-spondin-2 (Rspo2) or anti-Rspo2 to investigate the role of Rspo2 in OA. RESULTS: M1 but not M2-polarised macrophages accumulated in human and mouse OA synovial tissue. TSC1 deletion in the myeloid lineage constitutively activated mechanistic target of rapamycin complex 1 (mTORC1), increased M1 polarisation in synovial macrophages and exacerbated experimental OA in both CIOA and DMM models, while Rheb deletion inhibited mTORC1, enhanced M2 polarisation and alleviated CIOA in mice. The results show that promoting the macrophage M1 polarisation leads to exacerbation of experimental OA partially through secretion of Rspo2 and activation of ß-catenin signalling in chondrocytes. CONCLUSIONS: Synovial macrophage M1 polarisation exacerbates experimental CIOA partially through Rspo2. M1 macrophages and Rspo2 are potential therapeutic targets for OA treatment.

Blocking PI3K/AKT signaling inhibits bone sclerosis in subchondral bone and attenuates post-traumatic osteoarthritis.

PI3K/AKT signaling is essential in regulating pathophysiology of osteoarthritis (OA). However, its potential modulatory role in early OA progression has not been investigated yet. Here, a mouse destabilization OA model in the tibia was used to investigate roles of PI3K/AKT signaling in the early subchondral bone changes and OA pathological process. We revealed a significant increase in PI3K/AKT signaling activation which was associated with aberrant bone formation in tibial subchondral bone following destabilizing the medial meniscus (DMM), which was effectively prevented by treatment with PI3K/AKT signaling inhibitor LY294002. PI3K/AKT signaling inhibition attenuated articular cartilage degeneration. Serum and bone biochemical analyses revealed increased levels of MMP-13, which was found expressed mainly by osteoblastic cells in subchondral bone. However, this MMP-13 induction was attenuated by LY294002 treatment. Furthermore, PI3K/AKT signaling was found to enhance preosteoblast proliferation, differentiation, and expression of MMP-13 by activating NF-κB pathway. In conclusion, inhibition of PI3K/AKT/NF-κB axis was able to prevent aberrant bone formation and attenuate cartilage degeneration in OA mice.

Anterograde Fixation Module for Posterior Acetabular Column Fracture: Computer-Assisted Determination of Optimal Entry Point, Angle, and Length for Screw Insertion.

BACKGROUND The aim of this study was to provide valid data for a plate-screw fixation model for fractured posterior-anterior columns of the acetabulum. MATERIAL AND METHODS Nineteen cadaveric bony hemi-pelvis specimens were obtained and 50 healthy adults were enrolled. The modified Stoppa approach and computed tomography (CT) imaging were used to collect the measured parameter data of the module. RESULTS The measured parameter data were as follows: OP, 0.96±0.32 cm in females and 0.92±0.16 cm in males (P>0.05); PI, 0.98±0.28 cm in females, and 0.75±0.23 cm in males (P>0.05); ÐÏ´, 59.68°±6.28° in females and 56.75°±3.22° in males (P>0.05); and Ðφ, 41.27°±2.76° in females and 34.31°±2.78° in males (P<0.05). The corresponding CT image data were as follows: PI, 1.08±0.22 cm in females and 0.85±0.27 cm in males (P>0.05); OP, 1.06±0.29 cm in females and 1.12±0.24 cm in males (P>0.05); ÐÏ´, 55.33°±4.00° in females and 55. 50°±3.43° in males (P>0.05); and Ðφ was 39.21°±2.45°in females and 35.58°±2.31°in males (P<0.05). No significant difference with respect to sex and side existed between specimens and healthy adults (P>0.05). CONCLUSIONS The measured parameter data obtained in healthy adults and cadaveric specimens provided an anatomic basis for the designation of the guide module, and thus confirmed the accuracy and safety of screw placement in fractured columns of the acetabulum.

Kinematics of anterior cruciate ligament-deficient knees in a Chinese population during stair ascent.

BACKGROUND: The purpose of this study was to measure the tibiofemoral kinematics of anterior cruciate ligament (ACL) deficiency in a Chinese population and compare the kinematics with published data about a Caucasian population. METHODS: Unilateral knees of 18 Chinese ACL-deficient (ACL-D) subjects were studied while subjects ascended stairs. Kinematic alteration was compared between ACL-D knees and contralateral ACL-intact (ACL-I) knees. The kinematic alteration of ACL deficiency was also compared between the Chinese population and published data about a Caucasian population. RESULTS: A statistical difference was found in the three-dimensional rotations between ACL-D and ACL-I knees. In the sagittal plane, ACL-I knees had a larger flexion angle than ACL-D knees during 40 to 50 % of the activity during stair ascent (P < 0.027) and throughout the gait cycle. A significant difference in rotational motion between ACL-D and ACL-I knees was also observed in the frontal plane during 40 to 60 % (P < 0.017) of the activity and in the transverse plane during 70 to 80 % (P < 0.028) of the activity. A greater tibial varus was demonstrated in the Chinese population while the published data revealed external tibial rotation and a statistical difference in translation in the Caucasian population. CONCLUSIONS: ACL-D knees show different kinematics than ACL-I knees in the Chinese population. ACL-I knees had a larger flexion angle than ACL-D knees in the middle stage of the activity during stair ascent. A significant difference in rotational motion between ACL-D and ACL-I knees was also observed in the frontal plane during the middle phase and in the transverse plane during the terminal phase of the activity. A greater tibial varus was demonstrated in the Caucasian population while the published data revealed external tibial rotation and a statistical difference in translation in the Caucasian population.

Percutaneous fixation of traumatic pubic symphysis diastasis using a TightRope and external fixator versus using a cannulated screw.

BACKGROUND: The aim of the study was to introduce a new percutaneous technique for the treatment of traumatic pubic symphysis diastasis using a TightRope and external fixator. A comparison between this technique and percutaneous fixation using a cannulated screw was performed. METHODS: From January 2009 to December 2013, 26 patients with type II traumatic pubic symphysis diastasis were treated at two level 1 regional trauma centers. Among them, 10 patients were treated with a percutaneous TightRope and external fixator and 16 patients were treated with percutaneous cannulated screw fixation. Functional outcomes were evaluated using the Majeed scoring system. Patient satisfaction was evaluated using the modified visual analog scale. Radiological results were assessed based on the width of pubic symphysis preoperatively, immediately postoperatively, and at the final follow-up. Postoperative complications were also recorded. RESULTS: There were no significant differences between the groups in Majeed scores and patient satisfaction (p > 0.05). There were no significant differences in the width of pubic symphysis preoperatively, immediately postoperatively, and at the final follow-up (p > 0.05). No significant differences were found regarding infection, fixation failure, or the need for revision surgery (p > 0.05). CONCLUSIONS: The new percutaneous technique using a TightRope and external fixator is a successful alternative for the treatment of type II traumatic pubic symphysis diastasis, which results in similar outcomes comparing to percutaneous cannulated screw fixation.

Enhancement of osteogenesis post-splenectomy does not attenuate bone loss in ovariectomized rats.

The roles of different immune cell populations and cytokines in bone metabolism have been extensively investigated. However, the influence of whole immune organ removal on osteopathology remains unknown. In the current study, we investigated the effects of splenectomy on bone metabolism and microarchitecture in rats with or without concurrent ovariectomy. Ovariectomized (OVX) rats were used as osteoporosis model. Sixty 12-week-old female rats were randomized into 4 groups (n = 15): sham, splenectomized (SP), ovariectomized, as well as ovariectomized and splenectomized (OVX + SP). Bone microarchitecture was assessed by micro CT analysis at 4 week and 12 week post-operation, respectively. Bone pathology and metabolism were evaluated via immunohistochemical staining. The serum levels of alkaline phosphatase (ALP), tumor necrosis factor-alpha (TNF-α), tartrate-resistant acid phosphatase 5b (Tracp5b), and C-terminal telopeptide (CTx) were analyzed at 4 and 12 weeks post-operation. Removal of the spleen led to alterations in the homeostasis of bone metabolism and increased bone formation in rats. In this study, our findings indicate that the spleen is involved in skeletal metabolism.