WTAP affects intracranial aneurysm progression by regulating m6A methylation modification.

Intracranial aneurysm (IA) is a type of cerebrovascular disease that mainly occurs in the circle of Willis. Abnormalities in RNA methylation at the N6-methyladenosine (m6A) site have been associated with numerous types of human diseases. WTAP recruits the m6A methyltransferase complexes to the mRNA targets, and its expression is positively correlated with m6A methylation levels. This research aimed to explore the potential mechanisms of m6A methylation in IA. A selective arterial ligation method was used to establish an IA rat model; thereafter, the m6A methylation level and m6A methylation-related genes were determined in blood and circle of Willis samples using a commercial kit and real-time quantitative PCR, respectively. Subsequently, rat brain microvascular endothelial cells (rBMVECs) were treated with TNF-α, and the expression of m6A methylation-related genes within the cells were assessed. Lastly, the effects of WTAP on TNF-α-induced rBMVECs were further investigated through in vitro experiments. In result, the m6A RNA methylation level evidently declined in the blood and circle of Willis' samples of the IA rats, as compared to the corresponding samples from the control rats (P < 0.05). Compared to the results in the control rats/cells, WTAP expression was significantly downregulated, whereas ALKBH1 expression was evidently upregulated in the blood and circle of Willis samples of the TNF-α-induced rBMVECs of IA rats. Consequently, TNF-α-induced rBMVECs and rBMVECs with WTAP overexpression were successfully established. TNF-α inhibited the viability of the rBMVECs, promoted apoptosis, and significantly upregulated cleaved-caspase3 and downregulated WTAP expression. In contrast, WTAP overexpression significantly reversed these changes caused by TNF-α (P < 0.05). In conclusion, WTAP overexpression may modulate the growth of TNF-α-induced rBMVECs by enhancing WTAP expression and its m6A methylation.

An ultra-sensitive fluorescent "Turn On" biosensor for glutathione and its application in living cells.

In this work, an effective controlled-release biosensor based on Au nanocages (AuNCs) capped with disulfide-containing DNA molecular gates was developed for ultra-sensitive and highly selective detection of glutathione (GSH). Oligonucleotides containing the S-S bonds were assembled on the surface of the AuNCs by means of electrostatic interactions in order to inhibit the release of fluorescent molecules such as Rhodamine B (RhB) loaded by AuNCs. In the presence of GSH, due to the specific cleavage of S-S bonds in disulfide-containing single-stranded DNAs (ssDNAs) as well as their subsequent departure from the surface of AuNCs, the pores could be opened, and then the dye molecules would be released from AuNCs. The concentration of GSH ranged from 1.0 × 10-12 to 6.0 × 10-10 M could be detected. The developed amplification strategy based on the controlled-release of fluorescent molecules reached an extraordinary sensitivity of GSH. A detection limit of as low as 4.8 × 10-13 M with an excellent selectivity toward GSH could be achieved. The results of fluorescence microscopy imaging of GSH in living cells indicate that the fabricated system is an efficient controlled-release biosensor in response to intracellular target molecules and predict its potential use for in situ molecular imaging in living systems.

Tert-butylhydroquinone protects the spinal cord against inflammatory response produced by spinal cord injury.

Antioxidant transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) has been shown in our previous studies to play an important role in protection against spinal cord injury (SCI) induced inflammatory response. The objective of this study was to test whether tert-butylhydroquinone (tBHQ), a novel Nrf2 activator, can protect the spinal cord against SCI-induced inflammatory damage. Adult male Sprague-Dawley rats were subjected to laminectomy at T8-T9 and compression with a vascular clip. Three groups were analyzed: a sham group, a SCI group, and a SCI+rhEPO group (n=16 per group). We measured Nrf2 and nuclear factor kappa B (NF-κB) binding activities by an electrophoretic mobility shift assay (EMSA). We also measured the concentrations of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6) by an enzyme-linked immunosorbent assay (ELISA); we also measured hindlimb locomotion function by the Basso, Beattie, and Bresnahan (BBB) rating, spinal cord edema by wet/dry weight method, and apoptosis by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) analysis. The results showed that the induction of the Nrf2 activity by tBHQ markedly decreased NF-κB activation and inflammatory cytokines production in the injured spinal cord. Administration of tBHQ also significantly attenuated SCI induced hindlimb locomotion deficits, spinal cord edema, and apoptosis. To conclude, pre-treatment with tBHQ could attenuate the spinal cord inflammatory response after SCI.

Protective effects of erythropoietin in traumatic spinal cord injury by inducing the Nrf2 signaling pathway activation.

BACKGROUND: Erythropoietin has demonstrated neuroprotective effects against traumatic spinal cord injury (SCI), but the underlying mechanisms remain unclear. The signaling pathway of an antioxidant transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2), has been shown to play an important role in protecting SCI-induced secondary spinal cord damage. This study was undertaken to explore the effect of recombinant human erythropoietin (rhEPO) on the activation of Nrf2 signaling pathway and secondary spinal cord damage in rats after SCI. METHODS: Adult male Sprague-Dawley rats were subjected to laminectomy at T8-T9 and compression with a vascular clip. Three groups were analyzed: (1) sham group, (2) SCI group, and (3) SCI + rhEPO group (n = 16 per group). In the SCI + rhEPO group, rhEPO was administered at a dose of 5,000 IU/kg at 30 minutes after SCI. Spinal cord samples were extracted at 72 hours after the trauma. RESULTS: As a result, we found that the treatment with rhEPO markedly up-regulated the messenger RNA expressions and activities of Nrf2 signaling pathway-related agents, including Nrf2, NAD(P)H:quinone oxidoreductase 1(NQO1), and glutathione S-transferase. The administration of rhEPO also significantly ameliorated the secondary spinal cord damage, as shown by a decreased severity of locomotion deficit, spinal cord edema, and apoptosis. CONCLUSION: Post-SCI rhEPO administration induces Nrf2-mediated cytoprotective response in the injured spinal cord, and this may be a mechanism whereby rhEPO improves the outcome following SCI.

Induction of rat facial nerve regeneration by functional collagen scaffolds.

Nerve conduit provides a promising strategy for nerve regeneration, and the proper microenvironment in the lumen could improve the regeneration. Our previous work had demonstrated that linear ordered collagen scaffold (LOCS) could effectively guide the oriented growth of axons. Laminin is known as an important nerve growth promoting factor and can facilitate the growth cone formation. In addition, ciliary neurotrophic factor (CNTF) and brain-derived neurotrophic factor (BDNF) can effectively improve the nerve regeneration after nerve injuries. However, in practice, diffusion caused by the body fluids is the major obstacle in their applications. To retain CNTF or BDNF on the scaffolds, we produced collagen binding CNTF (CBD-CNTF), collagen binding BDNF (CBD-BDNF) and laminin binding CNTF (LBD-CNTF), laminin binding BDNF (LBD-BDNF) respectively. In this work, we developed laminin modified LOCS fibers (L × LOCS) by chemical cross-linking LOCS fibers with laminin. Collagen binding or laminin binding neurotrophic factors were combined with LOCS or L × LOCS, and then filled them into the collagen nerve conduit. They were found to guide the ordered growth of axons, and improve the nerve functional recovery in the rat facial nerve transection model. The combination of CNTF and BDNF greatly enhanced the facial nerve regeneration and functional recovery.