Manipulating PP2Acα-ASK-JNK signaling to favor apoptotic over necroptotic hepatocyte fate reduces the extent of necrosis and fibrosis upon acute liver injury.

In the widely used Carbon tetrachloride (CCl4)-induced acute liver injury (ALI) mouse model, hepatocytes are known to die from programmed cell death (PCD) processes including apoptosis and necroptosis. Both in vivo and in vitro experiments showed that CCl4 treatment could induce both apoptosis and necroptosis. Treatment of mice with the apoptosis inducer SMAC mimetic reduced necroptosis, led to less pronounced liver damage, and improved overall liver function. By LC-MS/MS, we found that PP2Acα expression was increased in ALI mice liver, and we confirmed its high expression in subacute hepatitis patients. We observed that ALI severity (including aggravated fibrogenesis) was significantly alleviated in hepatocyte-specific PP2Acα conditional knockout (PP2Acα cKO) mice. Furthermore, the relative extent of apoptosis over necroptosis was increased in the PP2Acα cKO ALI mice. Pursuing the idea that biasing the type of PCD towards apoptosis may reduce liver damage, we found that treatment of PP2Acα cKO ALI mice with the apoptosis inhibitor z-Vad-fmk increased the extent of necroptosis and caused severer damage. Mechanistically, disruption of PP2Acα prevents the dephosphorylation of pASK1(Ser967), thereby preventing the sustained activation of JNK. Inhibition of PP2Acα prevents CCl4-induced liver injury and fibrogenesis by disrupting ASK/JNK pathway mediated PCD signaling, ultimately improving liver function by biasing hepatocytes towards an apoptotic rather than necroptotic cell fate. Thus, targeting PP2A and/or ASK1 to favor apoptotic over necroptotic hepatocyte fate may represent an attractive therapeutic strategy for treating ALI.

Heterogeneity analysis of astrocytes following spinal cord injury at single-cell resolution.

Astrocytes play many important functions in response to spinal cord injury (SCI) in an activated manner, including clearance of necrotic tissue, formation of protective barrier, maintenance of microenvironment balance, interaction with immune cells, and formation of the glial scar. More and more studies have shown that the astrocytes are heterogeneous, such as inflammatory astrocyte 1 (A1) and neuroprotective astrocyte 2 (A2) types. However, the subtypes of astrocyte resulting from SCI have not been clearly defined. In this study, using single-cell RNA sequencing, we constructed the transcriptomic profile of astrocytes from uninjured spinal cord tissue and injured tissue nearby the lesion epicenter at 0.5, 1, 3, 7, 14, 60, and 90 days after mouse hemisection spinal cord surgery. Our analysis uncovered six transcriptionally distinct astrocyte states, including Atp1b2+ , S100a4+ , Gpr84+ , C3+ /G0s2+ , GFAP+ /Tm4sf1+ , and Gss+ /Cryab+ astrocytes. We used these new signatures combined with canonical astrocyte markers to determine the distribution of morphologically and physiologically distinct astrocyte population at injured sites by immunofluorescence staining. Then we identified the dynamic evolution process of each astrocyte subtype following SCI. Finally, we also revealed the evolution of highly expressed genes in these astrocyte subtypes at different phases of SCI. Together, we provided six astrocyte subtypes at single-cell resolution following SCI. These data not only contribute to understand the heterogeneity of astrocytes during SCI but also help to find new astrocyte subtypes as a target for SCI repair.

Effects of Mechanical Compression on Chondrogenesis of Human Synovium-Derived Mesenchymal Stem Cells in Agarose Hydrogel.

Mechanical compression is a double-edged sword for cartilage remodeling, and the effect of mechanical compression on chondrogenic differentiation still remains elusive to date. Herein, we investigate the effect of mechanical dynamic compression on the chondrogenic differentiation of human synovium-derived mesenchymal stem cells (SMSCs). To this aim, SMSCs encapsulated in agarose hydrogels were cultured in chondrogenic-induced medium with or without dynamic compression. Dynamic compression was applied at either early time-point (day 1) or late time-point (day 21) during chondrogenic induction period. We found that dynamic compression initiated at early time-point downregulated the expression level of chondrocyte-specific markers as well as hypertrophy-specific markers compared with unloaded control. On the contrary, dynamic compression applied at late time-point not only enhanced the levels of cartilage matrix gene expression, but also suppressed the hypertrophic development of SMSCs compared with unloaded controls. Taken together, our findings suggest that dynamic mechanical compression loading not only promotes chondrogenic differentiation of SMSCs, but also plays a vital role in the maintenance of cartilage phenotype, and our findings also provide an experimental guide for stem cell-based cartilage repair and regeneration.

Load-induced regulation of tendon homeostasis by SPARC, a genetic predisposition factor for tendon and ligament injuries.

Tendons and tendon interfaces have a very limited regenerative capacity, rendering their injuries clinically challenging to resolve. Tendons sense muscle-mediated load; however, our knowledge on how loading affects tendon structure and functional adaption remains fragmentary. Here, we provide evidence that the matricellular protein secreted protein acidic and rich in cysteine (SPARC) is critically involved in the mechanobiology of tendons and is required for tissue maturation, homeostasis, and enthesis development. We show that tendon loading at the early postnatal stage leads to tissue hypotrophy and impaired maturation of Achilles tendon enthesis in Sparc -/- mice. Treadmill training revealed a higher prevalence of spontaneous tendon ruptures and a net catabolic adaptation in Sparc -/- mice. Tendon hypoplasia was attenuated in Sparc -/- mice in response to muscle unloading with botulinum toxin A. In vitro culture of Sparc -/- three-dimensional tendon constructs showed load-dependent impairment of ribosomal S6 kinase activation, resulting in reduced type I collagen synthesis. Further, functional calcium imaging revealed that lower stresses were required to trigger mechanically induced responses in Sparc -/- tendon fascicles. To underscore the clinical relevance of the findings, we further demonstrate that a missense mutation (p.Cys130Gln) in the follistatin-like domain of SPARC, which causes impaired protein secretion and type I collagen fibrillogenesis, is associated with tendon and ligament injuries in patients. Together, our results demonstrate that SPARC is a key extracellular matrix protein essential for load-induced tendon tissue maturation and homeostasis.

Serum levels of leptin, osteopontin, and sclerostin in patients with and without knee osteoarthritis.

OBJECTIVE: To investigate the relationship between leptin, osteopontin (OPN), sclerostin (SOST) and severity of knee osteoarthritis (KOA). METHODS: The study included 148 consecutive patients with knee OA and 101 non-KOA subjects enrolled in this cross-sectional study. All patients fulfilled the American College of Rheumatology criteria for primary knee OA. Severity of the disease was assessed using plain radiography of the affected knee, according to the Kellgren and Lawrence classification. Fasting blood samples were obtained from all patients and controls; the serum samples were kept at - 80 °C before assessment of leptin, OPN, and SOST using a multiplex particle-based flow cytometric assay. RESULTS: KOA patients group compared with the control group, serum leptin (KOA, 26581.7 ± 2011.5 pg/ml, vs control,6936.4 ± 702.2 pg/ml),OPN (KOA, 4908.3 ± 769.4 pg/ml, vs control, 2182.5 ± 217.8 pg/ml), and SOST (KOA, 2481.9 ± 543.5 pg/ml, vs control, 1288.9 ± 267.7 pg/ml) in the KOA group were higher than control group; there were also differences in three bone metabolic factors between male and female in the KOA group; meanwhile, there was correlation between each factor and the incidence of KOA. CONCLUSION: Our study of 249 serum samples was conducted. Serum leptin, OPN, and SOST were significantly increased in KOA patients, and there was an internal correlation; these findings could, at best, contribute to the identification of novel targets for medical interventions. Key Points • The aim of this study was to assess the relationships of radiographic knee OA with altered serum levels of leptin, OPN, and SOST. Our study of 249 serum samples was conducted. Serum leptin, OPN, and SOST were significantly increased in KOA patients compared with control group. There were gender differences in the concentration of three serum bone turnover factors in KOA group and control group. Serum SOST concentration increased with Kellgren-Lawrence (K-L) grading. We found that serum leptin, OPN, and SOST were significantly increased in KOA patients, and there was an internal correlation. Leptin had a remarkable diagnostic value in the incidence of KOA.

Application of robotic-assisted in situ 3D printing in cartilage regeneration with HAMA hydrogel: An in vivo study.

The concept of in situ 3D bio-printing was previously reported, while its realization has still encountered with several difficulties. The present study aimed to report robotic-assisted in situ 3D bio-printing technology for cartilage regeneration, and explore its potential in clinical application. A six-degree-of-freedom (6-DOF) robot was introduced in this study, and a fast tool center point (TCP) calibration method was developed to improve printing accuracy. The bio-ink consisted of hyaluronic acid methacrylate and acrylate-terminated 4-armed polyethylene glycol was employed as well. The in vitro experiment was performed on a resin model to verify the printing accuracy. The in vivo experiment was conducted on rabbits to evaluate the cartilage treatment capability. According to our results, the accuracy of the robot could be notably improved, and the error of printed surface was less than 30 µm. The osteochondral defect could be repaired during about 60 s, and the regenerated cartilage in hydrogel implantation and in situ 3D bio-printing groups demonstrated the same biomechanical and biochemical performance. We found that the cartilage injury could be treated by using this method. The robotic-assisted in situ 3D bio-printing is highly appropriate for improving surgical procedure, as well as promoting cartilage regeneration.

Estrogen prevents articular cartilage destruction in a mouse model of AMPK deficiency via ERK-mTOR pathway.

BACKGROUND: To investigate the mechanism underlying the chondroprotective effect of estrogen in AMP-activated protein kinase (AMPK) deficiency mice. METHODS: Female cartilage-specific AMPKα double knockout (AMPKα cDKO) mice were generated and subjected to ovariectomy (OVX). The model of osteoarthritis (OA) was induced by destabilization of medial meniscus (DMM). Histopathological changes were evaluated by using OARSI scoring systems. Autophagy changes were analyzed by immunofluorescence staining. Human chondrocytes were subjected to mechanical stress to mimic OA development. and incubated in presence of or absence of 17ß-estradiol or/and compound C (AMPK inhibitor) or/and U0126 (ERK inhibitor). The expression levels of ERK1/2 phosphorylation, p70S6K phosphorylation and light chain 3 (LC3) were detected by Western blot. RESULTS: Compared with in OVX-sham AMPKα cDKO and OVX-sham WT mice, DMM-induced OA is more severe, and significantly low level of LC3 was observed in articular cartilage in OVX AMPK cDKO mice. Both mechanical stress and compound C were shown to induce an increase in phosphorylation of p70S6K, respectively. 17ß-estradiol stimulation led to a reduction in the basal level of p70S6K phosphorylation as well as in the compound C or mechanical stress-induced level of p70S6K phosphorylation. 17ß-estradiol stimulation not only led to an increase in LC3 conversion but also overrode the inhibitory effect of compound C on LC3 conversion. The effects of 17ß-estradiol were abrogated by blocking ERK signaling pathway. CONCLUSIONS: Our findings suggest that estrogen can protect articular cartilage from damage during OA development by promoting chondrocyte autophagy via ERK-mammalian target of rapamycin (mTOR) signaling, and give new insight into the mechanism of the chondroprotective effect of estrogen.

Inhibition of pyroptosis attenuates Staphylococcus aureus-induced bone injury in traumatic osteomyelitis.

BACKGROUND: Osteomyelitis is a severe bone infection and typically leads to progressive bone resorption, destruction and dysfunction. Pyroptosis is a form of programmed cell death involved in various infectious diseases. However, the identification of pyroptosis and the role it plays in osteomyelitis remains to be clarified. In this study, we investigated the expression of pyroptosis-associated proteins in osteomyelitis and the effects of inhibiting pyroptosis on S. aureus-induced osteomyelitis both in vitro and in vivo. METHODS: The expression of pyroptosis-associated protein-NLRP3 (NLR Family Pyrin Domain Containing 3), Caspase1 and GSDMD (GasderminD) were examined in murine and human infectious bone fragments by western blot. Bone destruction was evaluated by microcomputed tomography (µCT). The concentration of inflammatory factors was tested by Enzyme linked Immunosorbent Assay (ELISA). The expression of pyroptosis-associated gene was detected by real-time quantitative polymerase chain reaction (RT-qPCR). RESULTS: The expression of pyroptosis-associated proteins in infectious bone fragments from patients with osteomyelitis was significantly higher than uninfected bone. Additionally, in S. aureus-induced murine osteomyelitis model, higher expression of pyroptosis-associated proteins was noticed. Furthermore, the inhibitors of pyroptosis-associated proteins alleviated S. aureus-induced pyroptosis both in vivo and in vitro. More importantly, the inhibition of pyroptosis restored the bone formative property, attenuated the aberrant activation of osteoclast in vitro and reversed bone injury in vivo. CONCLUSIONS: Our study identified pyroptosis as a key pathway in osteomyelitis and elaborated that the inhibition of pyroptosis could attenuate S. aureus-induced bone destruction in osteomyelitis, providing a potential treatment target to osteomyelitis.

Inactivation of Lkb1 in postnatal chondrocytes leads to epiphyseal growth-plate abnormalities and promotes enchondroma-like formation.

Liver serine-threonine kinase B1 (LKB1) is a tumor suppressor that has been linked to many types of tumors. However, the role of LKB1 in cartilaginous tumorigenesis is still poorly understood. In this study, we find that cartilage-specific, tamoxifen-inducible Lkb1 knockout results in multiple enchondroma-like lesions adjacent to the disorganized growth plates. We showed that chondrocytes retain an immature status caused by loss of Lkb1, which may lead to the dramatic expansion of growth-plate cartilage and the formation of enchondroma-like lesions. Additionally, increased mammalian target of rapamycin complex 1 (mTORC1) activity is observed in the Lkb1 conditional knockout (cKO) chondrocytes, and rapamycin (mTORC1 inhibitor) treatment significantly alleviates the expansion of growth-plate cartilage and eliminates the enchondroma-like lesions in Lkb1 cKO mice. Thus, our findings indicate that loss of Lkb1 leads to the expansion of chondrocytes and the formation of enchondroma-like lesions during postnatal cartilage development, and that the up-regulated mTORC1-signaling pathway is implicated in this process. Our findings suggest that modulation of LKB1 and related signaling is a potential therapy in cartilaginous tumorigenesis.-Zhou, S., Li, Y., Qiao, L., Ge, Y., Huang, X., Gao, X., Ju, H., Wang, W., Zhang, J., Yan, J., Teng, H., Jiang, Q. Inactivation of Lkb1 in postnatal chondrocytes leads to epiphyseal growth-plate abnormalities and promotes enchondroma-like formation.

Recruitment of Brd3 and Brd4 to acetylated chromatin is essential for proinflammatory cytokine-induced matrix-degrading enzyme expression.

BACKGROUND: Proinflammatory cytokines, which can upregulate the expression of matrix-degrading enzymes in chondrocytes, play important roles in the development of osteoarthritis. BET family proteins, acting as the "readers" of acetylated modifications on histones, have been linked to transcriptional regulation. And a BET protein inhibitor, I-BET151, has been shown to inhibit the induction of matrix-degrading enzymes by proinflammatory cytokines in chondrocytes. Our objective is to clarify the role and mechanism of BET proteins on matrix-degrading enzyme gene expression by using a human chondrosarcoma cell line (SW1353). METHODS: We pretreated SW1353 cells with I-BET151 prior to treatment with IL-1ß or TNF-α and then checked the expression of four matrix-degrading enzyme genes (MMP1, MMP3, MMP13, and ADAMTS4). We performed knockdown of BET protein family members (BRD2, BRD3, and BRD4) with corresponding siRNAs in SW1353 cells prior to treatment with IL-1ß or TNF-α and checked the expression of the matrix-degrading enzyme genes. We evaluated Brd-mediated transcriptional regulation on the matrix-degrading enzyme genes by ChIP assay. RESULTS: We confirmed that I-BET151 could suppress the IL-1ß- or TNF-α-induced expression of MMP1, MMP3, MMP13, and ADAMTS4 in SW1353 cells. Brd3 and Brd4 were required for the IL-1ß- or TNF-α-induced expression of matrix-degrading enzyme genes in SW1353 cells. We revealed that inducible acetylation of H4k5/8/12 and the recruitment of Brd3, Brd4, and p-TEFb to chromatin were involved in IL-1ß- or TNF-α-induced transcription. CONCLUSIONS: Our findings suggested that Brd3 and Brd4 were essential for the IL-1ß- or TNF-α-induced transcription of matrix-degrading enzyme genes, and recruitment of Brd3 and Brd4 to chromatin of these genes played the main role in this process.

A genome-wide association study identifies new genes associated with developmental dysplasia of the hip.

Developmental dysplasia of the hip (DDH) is one of the most common congenital malformations and covers a spectrum of hip disorders from mild dysplasia to irreducible dislocation. The pathological mechanisms of DDH are poorly understood, which hampers the development of diagnostic tools and treatments. To gain insight into its disease mechanism, we explored the potential biological processes that underlie DDH by integrating pathway analysis tools and performing a genome-wide association study (GWAS). A total of 406 DDH-associated genes (P < 0.001) were identified by our GWAS using a Chinese Han cohort consisting of 386 DDH cases and 500 healthy controls (Set A). We verified the significant loci (P < 10-5 ) in another Chinese Han cohort consisting of 574 DDH patients and 569 healthy controls (Set B). An intronic Single Nucleotide Polymorphism (SNP) (rs61930502) showed significant association in Set A and Set B (P = 2.65 × 10-7 and 2.0 × 10-4 , respectively). The minor allele, rs61930502-A, which tended to prevent DDH showed a dominant effect. Heat shock 70 kDa protein 8 (HSPA8) showed the most direct interactions with other proteins which were coded by DDH-associated genes in the protein-protein interaction analysis. Interestingly, Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis suggested a relation between DDH and the genes involved in type II diabetes mellitus pathway (P = 0.0067). Our genetic and protein interaction evidence could open avenues for future studies of DDH.

The effectiveness of allogeneic mesenchymal stem cells therapy for knee osteoarthritis in pigs.

BACKGROUND: Intraarticular injection of the mesenchymal stem cells (MSCs) has shown to be successful for treating osteoarthritis (OA). Nevertheless, many studies have been focusing on autologous MSCs. The following study investigates the safety and effectiveness of intraarticular injection of allogenic MSCs in a pig OA model. METHODS: Superparamagnetic iron oxide (SPIO) nanoparticles were labelled with bone marrow-derived mesenchymal stem cells (BM-MSCs) to allow cells tracking using magnetic resonance imaging (MRI). A pig OA model was established by bilateral medial meniscectomy. Next, SPIO-BM-MSCs were injected into the right knee, while the left knee was left untreated. MRI and radiography were used to assess the degree of OA and to evaluate the effectiveness of allogenic MSCs. Hematoxylin and eosin (H&E), safranin-o fast green staining, toluidine blue, and immunohistochemical staining were used to evaluate the therapeutic effect of the injections. RESULTS: At concentration of ≤20 µg/mL, SPIO caused no toxicity to BM-MSCs. Four weeks after surgery, OA changes were observed on MRI scan. The SPIO labeled BM-MSCs were found moving towards the impaired part of the cartilage 8 to 24 h after injections. In addition, no significant differences between the right side (therapeutic side) and the left side (untreated side) were observed following histological and immunohistochemistry analysis. CONCLUSIONS: The suitable concentration of SPIO for labelling BMSCs was 20 µg/mL, while the allogenic MSCs could move towards and accumulate around the impaired cartilage. No significant difference was found between treatment and control group.

Evaluation of a polyvinyl alcohol-alginate based hydrogel for precise 3D bioprinting.

In this study, we designed a polyvinyl alcohol (PVA)-alginate based hydrogel and evaluated its cytocompatibility and printability. The samples were fabricated by 3D printing using a freeze-thaw process. The scanning electron microscope, material testing machine, rheometer, and cell counting kit-8 assay were used to examine the morphology, mechanical properties, rheological properties, and cytocompatiblity of the scaffolds, respectively. The mechanical strength, cytocompatiblity, crosslinking time, and printability were remarkably improved with the use of PVA. To sum up, our data suggest that hybrid bio-ink is more appropriate for precise 3D bioprinting due to its rapid prototyping capability and better cytocompatibility. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2944-2954, 2018.

Egr1 deficiency disrupts dynamic equilibrium of chondrocyte extracellular matrix through PPARγ/RUNX2 signaling pathways.

BACKGROUND: This study is to investigate the effect of Egr1 on the mineralization and accumulation of chondrocyte extracellular matrix. METHODS: The femoral heads of patients of various heights were collected. Egr1 knockout mice were used. Their limb lengtha nd body weight were assessed. The bone characteristics were detected by micro-CT scan and histological staining. Immature murine articular chondrocytes (iMACs) were isolated. Gross morphology was observed by histological staining. Relevant mRNA and protein expression were detected by qRT-PCR and Western blot, respectively. the related proteins were observed by immunohistochemical staining and immunofluorescence assay. Chromatin immunoprecipitation and reporter gene assay were also used. TUNEL was used to detect apoptosis. RESULTS: It was found that shorter patients had reduced Egr1 expression levels in the hypertrophic cartilage zone of the femoral head. In addition, Egr1 knockout mice exhibited reduced body size. Micro-CT analysis showed that these mice also had reduced bone volume. Safranin-O staining showed that the extracellular matrix of these mice exhibited a relatively limited degree of mineralization, and TUNEL staining showed reduced cell apoptosis levels. After transfecting the iMACs with dominant-negative Egr1 adenoviruses to inhibit Egr1, the enzymes of Adamst4, Adamst5, Mmp3 and Mmp13 were significantly upregulated. ChIP and luciferase assays revealed that Egr1 might regulate the chondrocyte extracellular matrix by the PPARγ/RUNX2 signaling pathways. CONCLUSION: Egr1 has an important regulatory effect on the dynamic equilibrium of the chondrocyte extracellular matrix, which may be achieved through the PPARγ/RUNX2 signaling pathways.

3D printing individualized heel cup for improving the self-reported pain of plantar fasciitis.

BACKGROUND: To explore the therapeutic effect and the biomechanical mechanism of 3D printing individualized heel cup in treating of plantar heel pain. METHODS: The clinical effect was evaluated by plantar pressure analysis and pain assessment in participants. Its biomechanical mechanism of protecting the plantar heel was explored using finite element simulation. RESULTS: The individualized heel cup could support and protect the osseous structure and soft tissue of plantar heel while walking and jogging, as well as significantly reduce the self-reported pain after being worn for 4 weeks. The nylon heel cup could alter the load concentration of the heel as well as decrease the load affected on plantar fascia and calcaneus bone. It also provided an obvious support for heel pad. CONCLUSION: To summarize, the 3D printed individualized heel cup can be used as an effective method for the treatment of plantar heel pain.

High preoperative serum leptin level is an independent risk factor for deep vein thrombosis after total knee arthroplasty in osteoarthritis patients: A prospective and cross-sectional study.

It suggests that a high leptin level may increase the risk of venous thromboembolism (VTE) in animal studies. However, clinical studies in this field are still largely unexplored. Our objective was to evaluate the relationship between the preoperative serum leptin levels and postoperative VTE incidence in osteoarthritis (OA) patients who underwent total knee arthroplasty (TKA) at our institute.We conducted a prospective and cross-sectional study in these OA patients from March 2014 to March 2016. Preoperative leptin levels were analyzed by Luminex assays. VTE was assessed preoperatively and on postoperative day 5 and 7. The potential risk factors for VTE were also documented.We enrolled 203 OA patients. No PE was detected and DVT was diagnosed in 34 patients postoperatively. There were significant differences between the median leptin levels in DVT group and non-DVT group [25.13 ng/mL (interquartile range, 14.51-44.31) vs 18.71 ng/mL (8.26-28.99), P = .007]. The relative risk of DVT significantly increased with natural logarithm (ln) leptin (per SD increase) (OR 2.37, 95% confidence interval (95% CI), 1.29-4.33, P = .005). Multivariate analyses adjusted for potential confounders showed ln leptin (per SD increase) was significantly associated with the relative risk of DVT (OR 2.17, 95% CI, 1.01-4.64, P = .046). When patients were subdivided into tertiles according to their leptin values, the OR for DVT increased with increasing tertiles of serum leptin (OR 1.03, 95% CI, 1.01-1.06, P for trend = .023).In the present study, our results indicate that a high preoperative leptin level may be an independent risk factor for postoperative DVT.

Bi-directional regulation of cartilage metabolism by inhibiting BET proteins-analysis of the effect of I-BET151 on human chondrocytes and murine joints.

BACKGROUND: Proinflammatory cytokines, which can upregulate the expression of matrix-degrading enzymes in chondrocytes, play important roles in the development of osteoarthritis. And a BET protein inhibitor, I-BET151, has been shown to exert an anti-inflammatory effect by repressing the BET protein-mediated expression of inflammatory genes. Our objective is to investigate the effect of I-BET151 on a surgical mouse model of osteoarthritis (OA) and human chondrocytes. METHODS: We first treated a surgical mouse model of OA with I-BET151 once per day and evaluated the knee joints at 6 and 8 weeks after treatment. We then pretreated the human chondrocytes with I-BET151 prior to treatment with IL-1ß or TNF-α and checked the expression and activity of the matrix-degrading enzyme genes. We also checked the expression of ACAN, COL2A1, and SOX9. RESULTS: We demonstrated that I-BET151 could prevent articular cartilage damage in the surgical mouse model of OA at an earlier time after treatment, but not at a later time after treatment. I-BET151 could robustly suppress the IL-1ß- and TNF-α-induced expression and activity of several matrix-degrading enzymes in human chondrocytes. I-BET151 could also suppress the expression of ACAN, COL2A1, and SOX9. CONCLUSIONS: Our findings suggested that inhibiting BET proteins could exert a repression effect on both of chondrocyte anabolism and catabolism, and the effect of BET protein inhibitor on surgical mouse model of OA needs further evaluation.

FTO variant is not associated with osteoarthritis in the Chinese Han population: replication study for a genome-wide association study identified risk loci.

BACKGROUND: Osteoarthritis is the most prevalent form of arthritis worldwide and is the major cause of pain and loss of function in elderly people. A signal of the fat mass and obesity-associated (FTO) gene had been reported in a genome-wide association study of osteoarthritis. The FTO polymorphism (rs8044769) might exert its effect on osteoarthritis through obesity, because it was reported as a body mass index-associated single-nucleotide polymorphism. And replication studies showed inconsistent results for this association. Our present study is to check the association of rs8044769 with osteoarthritis and body mass index in Chinese Han population. METHODS: A case-control association study was conducted by using 890 osteoarthritis cases and 844 controls in Chinese Han population. rs8044769 was genotyped in all subjects. Allelic and genotypic frequencies were compared between osteoarthritis cases and control subjects. Associations between rs8044769 and body mass index, and body mass index and osteoarthritis were also assessed. RESULTS: No significant difference was detected in genotype or allele distribution between osteoarthritis cases and controls (P > 0.05). Stratification by gender and body mass index revealed negative association between rs8044769 and osteoarthritis. We did not find any solid association between rs8044769 and higher body mass index. Meanwhile, we demonstrated that higher body mass index (body mass index ≥ 25) was associated with osteoarthritis. CONCLUSION: Our present study suggested that rs8044769 was not associated with osteoarthritis susceptibility or higher body mass index, and higher body mass index was a risk factor for osteoarthritis in the Chinese Han population. We also proposed that stratification by clinical parameters was crucial to reduce false-positive result in OA association studies.

Zhuangguguanjie formulation protects articular cartilage from degeneration in joint instability-induced murine knee osteoarthritis.

Zhuangguguanjie formulation (ZG) can provide noticeable relief from joint pain in patients suffering from knee osteoarthritis (OA). However, the underlying mechanism has not been fully described. Male C57BL/6 mice were administered either ZG or normal saline (NS) following surgical destabilization of the medial meniscus (DMM). At weeks 4, 6 and 8 (post-surgery), knee joints were harvested and assessed with Safranin-O staining. Blood serum was collected and tested. In vitro analysis was carried out to evaluate the effects of ZG on the expression of the OA-related genes. DMM mice indicated reduced cartilage destruction and lower blood serum biomarkers of OA (COMP1 and CTX-1) following ZG treatment. Moreover, the femoral condyle and tibial plateau histological scores were significantly reduced following ZG treatment of the DMM mice. ZG could markedly downregulate the expression of OA-related genes namely, ADAMTS5, MMP3 and MMP13, while it simultaneously upregulated collagen II as demonstrated by in vitro assays. Moreover, chondrocyte apoptosis was significantly decreased following ZG treatment. These results may be caused by the up-regulation of p-AKT expression levels, since the anti-apoptotic effects of ZG can be blocked by treatment with an AKT inhibitor. ZG is capable of preventing and/or reducing the progression of OA by inhibiting chondrocyte apoptosis via the p-AKT/Caspase 3 pathway.

Age-dependent variations of cancellous bone in response to ovariectomy in C57BL/6J mice.

The ovariectomized (OVX) mouse model has been widely accepted to be suitable for the study of postmenopausal osteoporosis. However, whether C57BL/6J mice, a commonly used genetic background mouse strain, is an appropriate model for postmenopausal osteoporosis remains controversial. The present study investigated the effect of the OVX model on alterations in bone density and microarchitecture in C57BL/6J female mice of different ages. C57BL/6J mice were divided into 8-, 12- and 16-week-old groups (OVX8, OVX12 and OVX16) from the beginning of OVX. At 8 weeks post-surgery, the mice were anesthetized and micro-computed tomography was used to analyze the bone density and microarchitecture. The results revealed that OVX-induced loss of cancellous bone was greatest in OVX8, moderate in OVX12, and only a weak bone loss was observed in the OVX16 group when compared with the SHAM16 control group. In addition, the effect of genetic backgrounds in response to the OVX model were examined. Several other strains of mice, including inbred (BALB/c) and outbred (ICR and Kunming), were used in the present study, all of which were subjected to OVX at 8 weeks of age. The present findings revealed that the highest rate of bone loss was detected in C57BL/6J female mice. In addition, treatment with estrogen (17ß-estradiol, 30 µg/kg five times per week) led to a significant increase in bone density in C57BL/6J mice compared with the other strains of mice. Therefore, these results may provide novel insights into the age- and strain-associated effect of OVX on regulating turnover of bone in female mice. The present findings also suggest 8-week-old C57BL/6J mice as an animal model for postmenopausal osteoporosis and preclinical testing of potential therapies for this disease.