miR-219a-5p inhibits the pyroptosis in knee osteoarthritis by inactivating the NLRP3 signaling via targeting FBXO3.

PURPOSE: This work was to identify the function and mechanism of miR-219a-5p in regulating knee osteoarthritis (KOA). METHODS: Rat fibroblast-like synoviocytes (FLSs) were isolated to construct KOA cell model by lipopolysaccharide and adenosine triphosphate treatment. miR-219a-5p and FBXO3 expression in FLSs was modulated by transfection. Flow cytometry was executed to research FLSs apoptosis. Caspase-1 and IL-1ß expression in FLSs was researched by immunofluorescence. The binding between miR-219a-5p and FBXO3 was identified by dual luciferase reporter gene assay. KOA rat model and miR-219a-5p up-modulation KOA rat model were constructed. Step size of rats was analyzed. Knee joints of rats were experienced Safranin O-fast green staining to evaluate the knee joint injury. FBXO3, pyroptosis-associated proteins, and IL-1ß and IL-18 expression in FLSs and articular cartilage tissues of rats were assessed by Western blot, qRT-PCR and Enzyme-linked immunosorbent assay. RESULTS: KOA cell model had higher apoptosis percentage, expression of pyroptosis-associated proteins, and IL-1ß and IL-18 level. miR-219a-5p up-modulation decreased the above indicators, whereas miR-219a-5p down-modulation increased the above indicators. FBXO3 expression was directly repressed by miR-219a-5p. Loss of FBXO3 suppressed the above indicators. FBXO3 counteracted the suppression of miR-219a-5p on the above indicators. miR-219a-5p agomir attenuated knee joint injury, increased step size of KOA rats, and reduced FBXO3, pyroptosis-associated proteins and level of IL-1ß and IL-18 in the articular cartilage tissues of KOA rats. CONCLUSION: miR-219a-5p suppressed the pyroptosis in KOA by inactivating the NLRP3 signaling via targeting FBXO3, which might be a promising target for ameliorating KOA in the clinic.

Fatigue Performance of RC Beams Strengthened with CFRP under Overloads with a Ladder Spectrum.

Vehicle overload is detrimental to bridges and traffic safety. This paper presents a study on the fatigue performance of typical reinforced concrete (RC) beams of highway bridges under vehicle overload. A definition method of vehicle overload and a construction method of overload ladder spectrum were first proposed based on traffic data acquisition, statistical analysis and structural calculation of the highway bridges in Guangzhou. A fatigue experimental method was also proposed with the three-ladder vehicle overload spectrum, and the fatigue tests of 15 RC beams strengthened with carbon fiber reinforced polymer (CFRP) under three loading levels were then carried out. The fatigue performance and the failure mechanism of the strengthened beams were presented and discussed, and two fatigue life prediction methods were proposed with the established modified Palmgren-Miner rule and the loading level equivalent method respectively. The results showed that the fatigue performance of the strengthened RC beams was severely degraded under overload ladder spectrum compared with that under constant amplitude cyclic load, and the life prediction methods were proved effective.

[Effect of acupuncture on HIF-1α/NLRP3 inflammatory signaling pathway in the rats with cerebral ischemia-reperfusion injury].

OBJECTIVE: To observe the effects of Xingnao Kaiqiao (regaining consciousness and opening orifices) acupuncture therapy on the expression of hypoxia-inducible factor 1α (HIF-1α) and Nod-like receptor protein 3 (NLRP3) in cerebral ischemia-reperfusion rats, and to explore the mechanism of acupuncture against cerebral ischemia-reperfusion injury. METHODS: Seventy-two male SD rats were randomly divided into a sham-operation group, a model group, an acupuncture group and a non-point acupuncture group, with 18 rats in each one. Using modified Longa thread embolization method, the rat model of acute focal cerebral ischemia was prepared; and after 2 h ischemia, the reperfusion was performed to prepared the model of cerebral ischemia-reperfusion. Immediately after reperfusion, Xingnao Kaiqiao acupuncture method was applied to bilateral "Neiguan" (PC 6) and "Shuigou" (GV 26) in the acupuncture group, while in the non-point acupuncture group, acupuncture was delivered at non-points and all of the needles were retained for 30 min in these two groups. The samples were collected 24 h after reperfusion in the rats of each group. Zea-Longa neurological deficit score was used to evaluate the degree of cerebral neurological impairment, TTC staining was adopted to observe the volume percentage of cerebral infarction, HE staining was provided to observe the morphological changes of brain, and Western blot was applied for detecting the expression of HIF-1α and NLRP3 proteins in the cerebral cortex on the right side. RESULTS: Compared with the sham-operation group, neurological deficit score and volume percentage of cerebral infarction were increased in the model group (P<0.01), and HIF-1α and NLRP3 protein expression was elevated (P<0.01). Compared with the model group, neurological deficit score and volume percentage of cerebral infarction were decreased (P<0.01), and HIF-1α and NLRP3 protein expression was lower (P<0.01) in the acupuncture group. There was no significant difference in above indexes in the non-point acupuncture group compared with the model group (P>0.05). Compared with the sham-operation group, the brain tissue of the rats in the model group and the non-point acupuncture group was loose and edema, and the nuclei were shriveled. The brain tissue morphology in the acupuncture group was similar to that of the sham-operation group. CONCLUSION: Acupuncture can alleviate cerebral ischemia-reperfusion injury, and its mechanism may be related to the regulation of HIF-1α/NLRP3 signaling pathway to attenuate inflammatory response.

Bioinformatics analysis of miRNA and mRNA expression profiles to reveal the key miRNAs and genes in osteoarthritis.

BACKGROUND: Osteoarthritis (OA) is a chronic degenerative joint disease and the most frequent type of arthritis. This study aimed to identify the key miRNAs and genes associated with OA progression. METHODS: The GSE105027 (microRNA [miRNA/miR] expression profile; 12 OA samples and 12 normal samples) and GSE48556 (messenger RNA [mRNA] expression profile; 106 OA samples and 33 normal samples) datasets were selected from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) and miRNAs (DEMs) were analyzed using the limma and ROCR packages in R, respectively. The target genes that negatively correlated with the DEMs were predicted, followed by functional enrichment analysis and construction of the miRNA-gene and miRNA-transcription factor (TF)-gene regulatory networks. Additionally, key miRNAs and genes were screened, and their expression levels were verified by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). RESULTS: A total of 1696 DEGs (739 upregulated and 957 downregulated) and 108 DEMs (56 upregulated and 52 downregulated) were identified in the OA samples. Furthermore, 56 target genes that negatively correlated with the DEMs were predicted and found to be enriched in three functional terms (e.g., positive regulation of intracellular protein transport) and three pathways (e.g., human cytomegalovirus infection). In addition, three key miRNAs (miR-98-5p, miR-7-5p, and miR-182-5p) and six key genes (murine double minute 2, MDM2; glycogen synthase kinase 3-beta, GSK3B; transmembrane P24-trafficking protein 10, TMED10; DDB1 and CUL4-associated factor 12, DCAF12; caspase 3, CASP3; and ring finger protein 44, RNF44) were screened, among which the miR-7-5p → TMED10/DCAF12, miR-98-5p → CASP3/RNF44, and miR-182-5p → GSK3B pairs were observed in the regulatory network. Moreover, the expression levels of TMED10, miR-7-5p, CASP3, miR-98-5p, GSK3B, and miR-182-5p showed a negative correlation with qRT-PCR verification. CONCLUSION: MiR-98-5p, miR-7-5p, miR-182-5p, MDM2, GSK3B, TMED10, DCAF12, CASP3, and RNF44 may play critical roles in OA progression.

Fatigue Life Prediction of Reinforced Concrete Beams Strengthened with CFRP: Study Based on an Accumulative Damage Model.

With the prestressed carbon fiber reinforced polymer (CFRP) strengthening technique widely used in reinforced concrete (RC) structures, it is more and more important to study the fatigue performance of RC structures. Since the fracture of a tensile steel bar at the main cracked section is the leading reason for the failure of RC beams reinforced by prestressed CFRP, a fatigue life prediction model of RC beams reinforced by prestressed CFRP was developed based on an accumulative damage model. Moreover, gradual degradation of the performance of the concrete was considered in the fatigue life prediction model. An experimental study was also conducted to research the fatigue behavior of RC beams reinforced by prestressed or non-prestressed carbon fiber laminate (CFL). During the tests, fatigue crack patterns were captured using a digital image correlation (DIC) technique, and the fatigue lives of a total of 30 beams were recorded. The results showed that the predicted main crack propagation curves and the fatigue lives were close to the experimental data. This study also exhibited that the prestressed CFRP could reduce the stress of main steel bars in RC beams and effectively improve the fatigue performance of the RC beams.

Specialization of mitochondrial and vascular oxidant modulated VEGFR in the denervated skeletal muscle.

Denervation of skeletal muscles results in timely muscular inflammation and muscle-T cell interaction, the cellular events might orchestrate a local circuit involved with IL-1ß and IL-15. In the present study, by a combination assay of nerve-muscle preparation, western blot, immuno-precipitation, and radioactive of enzyme activity, we confirmed that mitochondrial and vascular oxidants were considerably up-regulated following gastrocnemius denervation, which was due to gradual decay in mitochondrial biogenesis and XO pathway and accompanied by strengthened IL-1ß-VEGFR-2 and IL-15-VEGFR-1 signaling. Intriguingly, these alterations could be triggered by the early established muscular inflammation. In contrast, with prolonged muscle denervation, settings of organelle interconnection were ultimately conveyed by ER bound PTP1B, which promoted VEGFR-1 signaling and contributed to VEGFR-2 activation, and the process could be modulated by mitochondrial and vascular oxidant. Importantly, VEGFR-2 could rescue the disruption of MuSK activity and AchR cluster exerted by IL-1ß and IL-15, with PGC-1α and XO involvement. Altogether, extensive network centered on VEGFR-2 signaling was essentially contributed to early recovery processes regarding muscle denervation. Increasing knowledge of this mechanism might open up a conduit for functional response to muscle atrophy, and enable the development of better agents to combat the related disorders.