MicroRNA-210 Promotes Accumulation of Neural Precursor Cells Around Ischemic Foci After Cerebral Ischemia by Regulating the SOCS1-STAT3-VEGF-C Pathway.

BACKGROUND: Neural precursor cell (NPC) migration toward lesions is key for neurological functional recovery. The neovasculature plays an important role in guiding NPC migration. MicroRNA-210 (miR-210) promotes angiogenesis and neurogenesis in the subventricular zone and hippocampus after cerebral ischemia; however, whether miR-210 regulates NPC migration and the underlying mechanism is still unclear. This study investigated the role of miR-210 in NPC migration. METHODS AND RESULTS: Neovascularization and NPC accumulation was detected around ischemic foci in a mouse model of middle cerebral artery occlusion (MCAO) and reperfusion. Bone marrow-derived endothelial progenitor cells (EPCs) were found to participate in neovascularization. miR-210 was markedly upregulated after focal cerebral ischemia/reperfusion. Overexpressed miR-210 enhanced neovascularization and NPC accumulation around the ischemic lesion and vice versa, strongly suggesting that miR-210 might be involved in neovascularization and NPC accumulation after focal cerebral ischemia/reperfusion. In vitro experiments were conducted to explore the underlying mechanism. The transwell assay showed that EPCs facilitated NPC migration, which was further promoted by miR-210 overexpression in EPCs. In addition, miR-210 facilitated VEGF-C (vascular endothelial growth factor C) expression both in vitro and in vivo. Moreover, the luciferase reporter assay demonstrated that miR-210 directly targeted the 3' untranslated region of SOCS1 (suppressor of cytokine signaling 1), and miR-210 overexpression in HEK293 cells or EPCs decreased SOCS1 and increased STAT3 (signal transducer and activator of transcription 3) and VEGF-C expression. When EPCs were simultaneously transfected with miR-210 mimics and SOCS1, the expression of STAT3 and VEGF-C was reversed. CONCLUSIONS: miR-210 promoted neovascularization and NPC migration via the SOCS1-STAT3-VEGF-C pathway.

[Effect of low serum culture on the synchrony of cell cycle of mesenchymal stem cells].

AIM: To investigate a suitable method of inducing cell cycle synchronization at G0/G1 stage of mesenchymal stem cells (MSCs). METHODS: MSCs were cultured and identified with CD44, CD90, CD71 and CD11b by flow cytometer. Cell cycle and apotosis under normal and low serum culture were detected by flow cytometer. RESULTS: MSCs were positive for CD44, CD90 and CD71 and negative for CD11b. The cells at G0/G1 cell stage decreased, while the cells at S and G2 cell stage increased in 1 day of 50 mL/L fetal bovine serum (FBS) and 1 to 3 days of 5 mL/L FBS. However, prolonged culture in serum-starvation medium induced cell cycle arrest at G0/G1 stage. The ratio of apoptotic cells did not increase in 50 mL/L FBS. In 5 mL/L FBS, the ratio of apoptotic cells increased in 3 days and then decreased in 4 to 5 days. The proportion of cells at G0/G1 phase was significantly increased from 75.9% to 89.4% while the ratio of apoptotic cells was only 0.162% in 5 days of 5 mL/L FBS. CONCLUSION: It is a good method to induce the synchrony of cell cycle of MSCs at G0/G1 stage in five days of 5 mL/L FBS.

[Triplex-forming oligonucleotide inhibits the expression of tissue factor gene in endothelial cells induced by the blood flow shear stress in rats].

AIM: To study the effect of antiparallel phosphorothioate triplex-forming oligonucleotide (apsTFO) matching with the shear stress response element (SSRE) of tissue factor (TF) gene promoter region on the expression of TF in endothelial cells (ECs) of rat common carotid artery stenosis. METHODS: The model of common carotid artery middle segment stenosis was established by silica gel pipe loop ligation in SD rats. The mRNA expression and protein synthesis of TF, early growth response-1 (Egr-1) and specificity protein 1 (Sp1) were measured by in situ hybridization (ISH) and immunohistochemistry (IHC) technique. GT21-apsTFO, GT20-apsTFO, GT20-psTFO and FITC-labeled apsTFO, matching with the SSRE of TF gene promoter region, were designed, and intravenously injected into rats at 0.5 h before operation. TFO was detected 4 h after the operation, and the mRNA expression and protein synthesis of TF, Egr-1 and Sp1 were detected 6 h after the operation. RESULTS: There were much fluorescence in vascular tissue, especially in the nuclear of ECs 4.5 h after the injection of apsTFO. The mRNA expression and protein synthesis of TF reduced by 22% - 23% with injection of GT20-apsTFO 6.5 h after stenosis (P < 0.01) and by 10% - 11% with GT21-apsTFO at the same time (P < 0.05). The inhibition by GT20-apsTFO was stronger than that of the GT21-apsTFO (P < 0.05). The expression of TF was not inhibited by the GT20-psTFO (P > 0.05). The mRNA expression and protein synthesis of Egr-1 and Sp1 did not change in the rat treated with GT20-apsTFO, GT20-psTFO and GT21-apsTFO (P > 0.05). CONCLUSION: apsTFO could mero-inhibit the expression of TF gene but could not change the expression of Egr-1 and Sp1 protein.

[The study of the effect of the administration of mannitol on flash visual evoked potentials].

OBJECTIVE: To study the relationship between the change of N2 wave in flash visual evoked potentials (fVEP) and the intracranial pressure after the administration of mannitol. METHODS: Fifty-two patients with elevated intracranial pressure were chosen and are divided into two groups: the former (n = 32) were treated with mannitol, the latter (n = 20) were treated with mannitol and glycerol and sodium chloride injection. The latency and amplitude of N2 wave of fVEP was measured by NIP-200 noninvasive intracranial pressure apparatus before and after the 1st, 4th, 10th administration of mannitol. RESULTS: The latency of N2 wave began to reduce at 30 min after the injection of mannitol and was lowest at 2 h. At 4 h after the injection of mannitol, the latency of N2 wave increased but was still shorter than that before the injection of mannitol. The amplitude of N2 wave did not change significantly. The reduction of the latency of N2 wave after the administration of mannitol decreased with the multiple mannitol injection. The latency of N2 wave did not change significantly after the administration of mannitol when combined with glycerol and sodium chloride injection. CONCLUSION: The latency of N2 wave of fVEP changes after the administration of mannitol, which suggests that the change of the latency of N2 wave of fVEP can reflect the change of intracranial pressure.

[Approach to the relationship between the changes of the content of free zinc in hippocampus and ischemic neuronal damage].

AIM: To make approach to the relationship between the changes of free zinc and ischemic neuronal damage in hippocampus after forebrain ischemia/reperfusion. METHODS: The models of forebrain ischemia/reperfusion were established in rats. The contents of free Zn2+ were measured by TSQ fluorescence method. The Zn2+ chelator (CaEDTA) was injected into lateral ventricles in order to evaluate the effect of free Zn2+ on ischemic neuronal damage. RESULTS: (1) Zn2+ fluorescence in the hilus of dentate gyrus, CA3 region and the stratum radiatum and stratum oriens of CA1 decreased slightly at forty-eight hours after reperfusion. From seventy-two hours to ninety-six hour after reperfusion, the decreased fluorescence gradually returned to the normal level, but some fluorescence dots were found in pyramidal neurons of CA1 and the hilus of dentate gyrus. Seven days after reperfusion, all the changes of the fluorescence almost recovered. (2) The cell membrane-impermeable Zn2+ chelator CaEDTA could reduce the intracellular concentration of free Zn2+ and reduced neuronal damage after forebrain ischemia/reperfusion. CONCLUSION: (1) The synaptic vesicle Zn2+ released and then translocated into postsynaptic neurons after forebrain ischemia/reperfusion and played a role in ischemic neuronal damage. (2) The cell membrane-impermeable chelator CaEDTA could provide neuroprotection.