METTL3 promotes the progression of osteosarcoma through the N6-methyladenosine modification of MCAM via IGF2BP1.

BACKGROUND: The molecular mechanisms of osteosarcoma (OS) are complex. In this study, we focused on the functions of melanoma cell adhesion molecule (MCAM), methyltransferase 3 (METTL3) and insulin like growth factor 2 mRNA binding protein 1 (IGF2BP1) in OS development. METHODS: qRT-PCR assay and western blot assay were performed to determine mRNA and protein expression of MCAM, METTL3, IGF2BP1 and YY1. MTT assay and colony formation assay were conducted to assess cell proliferation. Cell apoptosis, invasion and migration were evaluated by flow cytometry analysis, transwell assay and wound-healing assay, respectively. Methylated RNA Immunoprecipitation (MeRIP), dual-luciferase reporter, Co-IP, RIP and ChIP assays were performed to analyze the relationships of MCAM, METTL3, IGF2BP1 and YY1. The functions of METTL3 and MCAM in tumor growth were explored through in vivo experiments. RESULTS: MCAM was upregulated in OS, and MCAM overexpression promoted OS cell growth, invasion and migration and inhibited apoptosis. METTL3 and IGF2BP1 were demonstrated to mediate the m6A methylation of MCAM. Functionally, METTL3 or IGF2BP1 silencing inhibited OS cell progression, while MCAM overexpression ameliorated the effects. Transcription factor YY1 promoted the transcription level of METTL3 and regulated METTL3 expression in OS cells. Additionally, METTL3 deficiency suppressed tumor growth in vivo, while MCAM overexpression abated the effect. CONCLUSION: YY1/METTL3/IGF2BP1/MCAM axis aggravated OS development, which might provide novel therapy targets for OS.

Inflammation-related pathways involved in damaged articular cartilage of rats exposed to T-2 toxin based on RNA-sequencing analysis.

Many studies have shown that ingestion of the T-2 toxin is harmful to articular cartilage. However, the mechanisms underlying damaged articular cartilage induced by T-2 toxin have not been elucidated. Twenty-four SD rats were randomly divided into T-2 toxin and control groups. In the control group, the 12 rats were administered 4% absolute ethanol by gavage, and in the T-2 toxin group, the 12 rats were administered T-2 toxin (100 ng/g, BW/day) by gavage. After the rats were sacrificed, the knee joints were collected, and RNA was extracted using TRIzol reagent for RNA sequencing (RNA-seq). Differentially expressed mRNA was identified based on p < 0.05 and | log2 (fold change) | > 1. The T-2 toxin-related genes were obtained from the GeneCards database. An online tool (https://www.bioinformatics.com.cn) was used for enrichment analysis. Hematoxylin and eosin (H&E) staining was used to observe damaged articular cartilage, and immunohistochemical (IHC) staining was used to validate differentially expressed proteins. The H&E staining shows the number of cells decreased significantly, and the arrangement of chondrocytes became disordered in the T-2 toxin group. RNA-seq analysis identified 195 upregulated and 89 downregulated mRNAs in the T-2 toxin group. The top immune-related biological processes (Gene Ontology) were regulation of hormone secretion, regulation of peptide hormone secretion, and regulation of transcription involved in cell fate commitment. KEGG pathway enrichment analysis revealed that the IL-17 and tumor necrosis factor signaling pathways were significantly expressed, and the IL-17 signaling pathway was also identified in the enrichment analysis of T-2 toxin-related genes. Also, Mmp3, Tnf, Mapk10, Ccl11, Creb5, Cxcl2, and Cebpb were significantly enriched in the two pathways. The immunohistochemical staining showed that the levels of Mmp3 and Tnf proteins were significantly increased in the T-2 toxin group, which was consistent with the RNA-seq results. This study revealed the critical roles of IL-17 and TNF signaling pathways in damaged cartilage induced by T-2 toxin.

Calcium-binding protein 39 overexpression promotes macrophages from 'M1' into 'M2' phenotype and improves chondrocyte damage in osteoarthritis by activating the AMP-activated protein kinase/sirtuin 1 axis.

Osteoarthritis (OA) is a degenerative joint disease that affects cartilage and its peripheral tissues. Up-regulation of Calcium-binding protein 39 (CAB39) has a significant protective effect on osteoblasts, but the role and related molecular mechanisms of CAB39 in OA have not yet been reported. CAB39 overexpression and knockdown models were set up in chondrocytes (ATDC5) and macrophages (RAW264.7). The OA cell model was induced in ATDC5 cells with IL-1ß (10 ng/mL). Cell viability was tested by the cell counting kit-8 assay, apoptosis was checked by flow cytometry. Western blot was applied for checking the expression of MMP3, MMP13, Aggrecan, the AMPK/Sirt-1 pathway, apoptosis-related proteins (Bax, Bcl-2, and Caspase-3), and macrophage phenotypic markers (CD86, iNOS, CD206, and Arg1). An OA model was constructed in mice, and CAB39 overexpression plasmids were administered to the knee cavity of the OA model mice. As a result, CAB39 was down-regulated in IL-1ß-treated chondrocytes and OA mice. Overexpressing CAB39 enhanced ATDC5 cell viability and choked IL-1ß-mediated apoptosis. Overexpression of CAB39 boosted the polarization of macrophages from M1-phenotype into M2 phenotype. In addition, overexpressing CAB39 facilitated the AMPK/Sirt-1 pathway activation, and AMPK inhibitors reversed the protective effect of CAB39 overexpression on chondrocytes. Moreover, CAB39 exhibited anti-inflammatory effects in OA mice by activating the AMPK/Sirt-1 pathway. Collectively, overexpressing CAB39 heightened macrophages' M2 polarization and declined chondrocyte injury in OA by activating the AMPK/Sirt-1 pathway.Abbreviations AMPK: AMP-activated protein kinaseArg1: arginase 1Bax: Bcl-2-associated X proteinBcl-2: B-cell lymphoma-2CAB39: Calcium-binding protein 39CM: Conditioned mediumDMM: destabilization of the medial meniscusECM: extracellular matrixELISA: enzyme-linked immunosorbent assayFCM: Flow cytometryIL-1ß: interleukin-1ßIL-4: interleukin-4IL-6: interleukin-6IL-10: interleukin-10IFN - γ: Interferon-gammaIHC: ImmunohistochemistryiNOS: Inducible nitric oxide synthaseLKB1: liver kinase B1MMP3: Matrix metalloproteinase3MMP13:Matrix metalloproteinase13NF-κB: NF-kappaBOA: OsteoarthritisqRT-PCR: Quantitative reverse transcription-polymerase chain reactionRT: room temperatureSirt-1: sirtuin 1STRAD: STE20-related adaptor alphaWB: Western blot.

The influence of prior arthroscopy on outcomes of primary total lower extremity arthroplasty: A systematic review and meta-analysis.

PURPOSE: The primary purpose of this systematic review and meta-analysis was to investigate the impact of prior arthroscopy on postoperative revisions, complications, and other clinical outcomes after conversion total lower extremity arthroplasty. METHODS: Two individual researchers conducted the platform searches on the Embase, PubMed, Cochrane Central, and Google Scholar electronic databases from inception to June 02, 2021. We identified cohort trials that compared the outcomes of patients who underwent primary THA or TKA in the prior arthroscopy or control groups. The primary outcome was revision, and secondary outcomes included reoperation, patient-reported outcomes, and postoperative complications. A modified version of the Downs and Black tool was used to assess the methodological quality of the non-randomized cohort studies. RESULTS: Of the 23 included studies with 319946 cases, 18 were matched retrospectively and five were non-matched retrospectively. Methodological quality was high in ten studies and moderate in thirteen studies. Our analysis demonstrated that TKA or THA patients with prior arthroscopy were associated with an increased risk of revision, reoperation, infection, and aseptic loosening. THA patients with prior arthroscopy were also associated with an increased risk of dislocation. Furthermore, there were no significant intergroup differences in periprosthetic fracture, range of motion, Harris Hip Score, or Knee Society Score. CONCLUSION: Arthroscopy performed before total lower extremity arthroplasty substantially increased the revision, reoperation, infection, and aseptic loosening rates. THA patients with prior arthroscopy were also associated with an increased risk of dislocation. Patients should be counseled on the potential increased risks associated with conversion total lower extremity arthroplasty after prior arthroscopy. Further research is needed to better characterize these findings.

Cross-species transmission of feline herpesvirus 1 (FHV-1) to chinchillas.

BACKGROUND: Herpesviruses are a class of double-stranded DNA viruses found in both vertebrates and invertebrates. They are usually highly host-specific and do not easily spread across species. Chinchillas have gradually entered the Chinese pet market in recent years, but references to viral infections in chinchillas are extremely scarce, and only two reports about the herpesvirus in chinchillas are available at present. OBJECTIVES: The aim of this study was to present the first report of FHV-1 infection in chinchillas. METHODS: A total of 130 nasopharyngeal swab samples of chinchillas and three nasopharyngeal swabs of domestic cats collected from a chinchillas farm were investigated by nested PCR for FHV-1. RESULTS: Four chinchillas were infected with FHV-1, the positive rate was 3.08% (4/130), and two domestic cats were FHV-1 positive (2/3). The 253 bp fragments of FHV-1 gD gene from four chinchillas and two domestic cats were 100% identical, respectively, and the homology between chinchillas and domestic cat was 99.21%, but they all shared nearly 98.81% homology with the reference strain sequences. Phylogenetic tree analysis showed that these four chinchillas strains were clustered together with FHV-1. CONCLUSIONS: This is the first time that FHV-1 was detected in chinchillas and suggested chinchillas are susceptible to FHV-1 and may play a role as a temporary reservoir for FHV-1.

miR-29 promotes osteosarcoma cell proliferation and migration by targeting PTEN.

Osteosarcoma (OS) is an aggressive malignant neoplasm that arises from primitively transformed cells of mesenchymal origin, and that exhibits osteoblastic differentiation and produces malignant osteoid. MicroRNAs (miRNAs) have been widely reported to have important regulatory roles in various human tumors, including OS. However, the potential mechanism of miR-29 in OS remains largely unknown. miR-29 was highly expressed in OS and overexpression of miR-29 promoted OS cell proliferation, as well as proliferating cell nuclear antigen (PCNA) expression and migration, whereas lower expression of miR-29 inhibited OS cell proliferation, PCNA expression and migration. In the present study, a dual-luciferase reporter system supporting phosphatase and tensin homolog (PTEN) was a target of miR-29 and its expression was inhibited by miR-29 mimic, but increased by miR-29 inhibitor. Overexpression of PTEN inhibited OS cell proliferation and migration and it could attenuate miR-29 promotion effect on OS progression. Overall, the results revealed that miR-29, as a tumor promoter, is involved in OS progression and metastasis by targeting PTEN, indicating that the miR-29/PTEN pathway is a potential therapeutic target for the treatment of OS.

Deep eutectic solvents-derived carbon dots for detection of mercury (II), photocatalytic antifungal activity and fluorescent labeling for C. albicans.

A novel nitrogen and chloride co-doped carbon dots (N/Cl-CDs) based choline chloride-urea deep eutectic solvents (DESs) were synthesized by one-step hydrothermal method with high quantum yield of 37% and excellent photoluminescent properties. This N/Cl-CDs fluorescent probe had been successfully applied to sensitively and selectively detect the concentration of Hg2+ with a linear range of 0.2-40 µM and a detection limit of 0.05 µM. Moreover, the N/Cl-CDs displayed a strong photocatalytic antifungal activity against C. albicans and their photoinduced antifungal functions were evaluated under conditions of varying other experimental parameters. The antifungal efficiency of N/Cl-CDs (7 mg/mL) against C. albicans is upon 100% when extending the visible light irradiation time to 80 min. In addition, the excellent luminescence properties of N/Cl-CDs can also label C. albicans and displayed multicolour fluorescence imaging at different excitation wavelengths. Based on their functions of fluorescence probe, antifungal and fluorescence imaging, N/Cl-CDs would provide potentials for a wide range of applications in the detection and microbe in the near future.

Tranexamic acid versus aminocaproic acid for blood management after total knee and total hip arthroplasty: A systematic review and meta-analysis.

OBJECTIVE: To compare the efficacy and safety of tranexamic acid and aminocaproic acid for reducing blood loss and transfusion requirements after total knee and total hip arthroplasty. METHODS: We conduct electronic searches of Medline (1966-2017.11), PubMed (1966-2017.11), Embase (1980-2017.11), ScienceDirect (1985-2017.11) and the Cochrane Library (1900-2017.11). The primary outcomes, including total blood loss, hemoglobin decline and transfusion requirements. Secondary outcomes include length of hospital stay and postoperative complications such as the incidence of deep vein thrombosis and pulmonary embolism. Each outcome is combined and calculated using the statistical software STATA 12.0. Fixed/random effect model is adopted based on the heterogeneity tested by I2 statistic. RESULTS: A total of 1714 patients are analyzed across three randomized controlled trials (RCTs) and one non-RCT. The present meta-analysis reveals that TXA is associated with a significantly reduction of total blood loss and postoperative hemoglobin drop compared with EACA. No significant differences are identified in terms of transfusion rates, length of hospital stay, and the incidence of postoperative complications. CONCLUSION: Although total blood loss and postoperative hemoglobin drop are significant greater in EACA groups, there is no significant difference between TXA and EACA groups in terms of transfusion rates. Based on the current evidence available, higher quality RCTs are still required for further research.

Honokiol triggers receptor­interacting protein kinase 3­mediated cell death of neuroblastoma cells by upregulating reactive oxygen species.

Neuroblastoma is the most common form of childhood extracranial tumor and almost half of neuroblastoma cases occur in infants under two years old. Neuroblastoma accounts for ~6­10% of childhood cancers and 15% of cancer­associated childhood mortality. However, an effective treatment remains to be developed. Honokiol exhibits long­lasting central muscle relaxation, anti­inflammatory, antibacterial, antimicrobial, antiulcer, antioxidation, antiaging and antitumor effects. Honokiol has been previously demonstrated to kill neuroblastoma cells, however, the underlying mechanism of action remains unclear. The present study reports that honokiol inhibits the growth of neuroblastoma cells via upregulation of reactive oxygen species (ROS). MTT assays demonstrated that treatment of Neuro­2a neuroblastoma cells with honokiol resulted in time­ and dose­dependent inhibition of cell proliferation, which was associated with upregulation of the protein expression of receptor­interacting protein kinase 3 (RIP3), as demonstrated by western blot analysis. Furthermore, knockdown of RIP3 by small interfering RNA, or pharmacological inhibition of RIP3 by the RIP3 specific inhibitor necrosulfonamide, reversed honokiol­induced loss of cell viability in Neuro­2a cells. Importantly, honokiol significantly increased the intracellular ROS levels as determined by a 2',7'­dichlorofluorescin diacetate assay, while ROS scavenger N­acetyl cysteine significantly prevented the induction of ROS and RIP3 by honokiol. The results of the present study indicate that honokiol may suppress the growth of neuroblastoma Neuro­2a cells, at least partially, through ROS­mediated upregulation of RIP3.