Long non-coding RNA XIST protects chondrocytes ATDC5 and CHON-001 from IL-1ß-induced injury via regulating miR-653-5p/SIRT1 axis.

Chondrocyte apoptosis is linked to cartilage degeneration, and considered as a crucial event during the development of osteoarthritis (OA). X inactive specific transcript (XIST) is an oncogenic long non-coding RNA (lncRNA). However, its role in the pathophysiological process of OA remains largely unknown. In this work, quantitative real-time reverse transcriptase PCR (qRT-PCR) was employed to measure the expression of XIST, miR-653-5p and sirtuin1 (SIRT1) mRNA in OA and normal cartilage tissues. Chondrocyte cell lines, CHON-001 and ATDC5, were treated with different doses of interleukin- 1ß (IL-1ß) to mimic the inflammatory environment of OA in vitro. Overexpression plasmids, microRNA (miRNA) mimics, miRNA inhibitors and small interfering RNAs (siRNAs) were constructed and transfected into CHON-001 and ATDC5 cells. 3-(4,5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) assay was adopted to determine the cell viability. Western blot was used to detect the expression of apoptosis-related proteins. Enzyme-linked immunosorbent assay (ELISA) was employed to probe the expression levels of inflammatory factors. Flow cytometry was used to analyze the cell apoptosis. StarBase and TargetScan databases were used to predict the binding sites between XIST and miR-653-5p, miR-653-5p and 3'UTR of SIRT1, respectively, which were then verified by dual luciferase reporter assay. The data in the present study demonstrated that XIST and SIRT1 were down-regulated while miR-653-5p was up-regulated in OA tissues and cell models. The up-regulation of XIST increased the viability of CHON-001 and ATDC5 cells, while it impeded their apoptosis and inflammatory response induced by IL-1ß. Conversely, miR-653-5p had opposite effects. It was proved that miR-653-5p could be sponged and suppressed by XIST. Additionally, SIRT1 was identified as a target of miR-653-5p, and SIRT1 could be suppressed by XIST indirectly. In conclusion, down-regulated XIST was involved in the injury of chondrocytes during the pathophysiological process of OA, and XIST up-regulation protected chondrocytes from inflammatory injury via regulating miR-653-5p/SIRT1 axis.

[Regulation of m

N^(6)-methyladenosine (m^(6)A) has been identified as a conserved epitranscriptomic modification of eukaryotic mRNAs, and plays important biological roles in the regulation of cellular metabolic processes. However, its role in myogenic differentiation is unclear. Here, we altered the m^(6)A RNA methylation level by overexpression of METTL3, and explored the effect of m^(6)A RNA methylation on myogenic differentiation of murine myoblasts in vitro. The m6A RNA methylation level is regulated by exogenous methylation inhibitor cycloleucine (Cyc) and methyl donor betaine (Bet). Therefore, chemical reagents of Cyc and Bet were used to test the regulatory effect of m^(6)A RNA methylation on myogenic differentiation. Results showed that METTL3 and Bet positively regulated the m^(6)A RNA methylation levels, and Cyc negatively regulated m^(6)A RNA methylation levels. In addition, m^(6)A methylation positively regulated myogenic differentiation in murine myoblasts. These findings provide insight in the mechanisms underlying the effect of m^(6)A RNA methylation on myogenesis.

Effect analysis of early bedside hemo-filtration in treatment of severe pneumonia with acute renal failure of children.

OBJECTIVE: To investigate the best opportunity for bedside continuous blood purification (CBP) to treat severe pneumonia with acute renal failure (ARF) of children and look for the sensitive marker to evaluate the clinical effects and prognosis. PATIENTS AND METHODS: 54 children patients that were diagnosed as severe pneumonia with ARF by Pediatric Intensive Care Unit (PICU) were enrolled in our study as experimental group. In the meanwhile, 46 children patients that were diagnosed as severe pneumonia with ARF by PICU were enrolled as a normal control group. Patients in the experimental group started CBP treatment within 24 h after onset while patients in the control group started CBP treatment 24h after onset. The differences of clinical effects between two groups were compared for statistical significance. RESULTS: The survival rates of the observation group in day 7, day 28 and 6 months were significantly higher than those in the control group. After treatment for 7 days, IL-6 and TNF-α, YKL-40 and Annexin A1 levels of the experimental group were significantly lower than those of the control group. 7-day infection-related organ failure score (SOFA) of the experimental group was significantly lower than that of the control group. CONCLUSIONS: CBP therapy for treating severe pneumonia with acute renal failure of children within 24 hours could significantly improve the survival rate and reduce the inflammatory reactions.

Nitrogen supply after removing the shoot apex increases the nicotine concentration and nitrogen content of tobacco plants.

BACKGROUND: and Aims High nicotine concentrations in leaves, especially in the upper leaves, offer a serious problem for the cultivation of tobacco (Nicotiana tabacum). Preliminary field experiments showed that rapid mineralization of soil N during late stages of growth may contribute to high nicotine concentrations in leaves. METHODS: A sand-culture experiment was carried out in the greenhouse. The N supply was controlled during the experiment, and different amounts of 15N were supplied during late stages of growth (after removal of the shoot apex), to investigate the contribution of the N taken up at this time to the N content of and nicotine concentration in tobacco plants. KEY RESULTS: Addition of 1.6 g or 4 g 15N-labelled NH4NO3 after removing the shoot apex and flushing out the 14N did not increase leaf dry weights; however, it did result in delayed leaf senescence, more lateral bud formation, and an increase in 15N as a proportion of total N, and nicotine-15N as a proportion of total nicotine-N in each organ. The nicotine concentration, 15N and nicotine-15N abundances were increased from the bottom to the top leaves. When more 15N-labelled NH4NO3 was supplied, the nicotine concentration in leaves increased, and so did the 15N abundance in nicotine-N. CONCLUSION: Enhanced N supply in the later growth stages (after removing the apex) increased N content and nicotine concentration in tobacco plants. Nicotine was synthesized de novo during the late growth stages.