microRNA-451 protects neurons against ischemia/reperfusion injury-induced cell death by targeting CELF2

Figures & data

Table 1 The sequence of the primers for qRT-PCR

hsa-miR-451	Forward	5′-ACACTCCAGCTGGGAAACCGTTACCATTACT-3′
	Reverse	5′-CTGGTGTCGTGGAGTCGGCAA-3′
hsa-CELF2	Forward	5′-CTGGCGGGAAACAAACTCTG-3′
	Reverse	5′-TCTAAGCCCTTGGCCTCCTC-3′
GAPDH	Forward	5′-GCACCGTCAAGGCTGAGAAC-3′
	Reverse	5′-TGGTGAAGACGCCAGTGGA-3′

Abbreviation: qRT-PCR, quantitative reverse transcription PCR.

Figure 1 miR-451 expression was decreased in the OGD/R cells.

Notes: The relative expression of miR-451 was measured using qRT-PCR upon hypoxia for 12 or 24 hours. **P<0.01, ***P<0.001.
Abbreviations: OGD/R, oxygen and glucose deprivation/reoxygenation; qRT-PCR, quantitative reverse transcription PCR.

Figure 2 Overexpression of miR-451 inhibited apoptosis and oxidative stress in the OGD/R cells.

Notes: The OGD/R cells were transfected with miR-451 or NC. (A) The relative expression of miR-451, (B) cell viability, (C) LDH level, (D) SOD activity, (E) MDA level, (F) relative ROS level, (G) apoptosis and (H) the expression of cleaved-caspase-3 were assessed using qRT-PCR, CCK-8, LDH, SOD, MDA and ROS assays, flow cytometry and Western blot analysis, respectively. ***P<0.001. ^###P<0.001.
Abbreviations: CCK-8, Cell Counting Kit-8; FITC, fluorescein isothiocyanate; MDA, malondialdehyde; NC, negative control; OGD/R, oxygen and glucose deprivation/reoxygenation; PI, propidium iodide; qRT-PCR, quantitative reverse transcription PCR.

Figure 3 CELF2 was a target of miR-451 in OGD/R cells. The OGD/R cells were transfected with miR-451 mimic or NC mimic.

Notes: (A) The potential binding sequences between CELF2 and miR-451, and the relative luciferase activity were examined by bioinformatics analysis and the dual-luciferase reporter assay. (B) The transfection efficiency of CELF2 was examined using the Western blot analysis. (C, D) The relative expression of CELF2 was measured using qRT-PCR and Western blot analysis. **P<0.01, ***P<0.001. ^###P<0.001. ^#Represents the comparison between the OGD/R+miR-451 group and the OGD/R+NC group.
Abbreviations: NC, negative control; OGD/R, oxygen and glucose deprivation/reoxygenation; qRT-PCR, quantitative reverse transcription PCR.

Figure 4 Knockdown of CELF2 inhibited apoptosis and oxidative stress in the OGD/R cells. The OGD/R cells were transfected with siCELF2 or NC.

Notes: (A) The relative expression of CELF2, (B) cell viability, (C) LDH level, (D) SOD activity, (E) MDA level, (F) relative ROS level, (G) apoptosis and (H) the expression of cleaved-caspase-3 were assessed using qRT-PCR, CCK-8, LDH, SOD, MDA and ROS assays, flow cytometry and Western blot analysis, respectively. ***P<0.001. ^###P<0.001.
Abbreviations: CCK-8, Cell Counting Kit-8; FITC, fluorescein isothiocyanate; MDA, malondialdehyde; NC, negative control; OGD/R, oxygen and glucose deprivation/reoxygenation; PI, propidium iodide; qRT-PCR, quantitative reverse transcription PCR.

Figure 5 CELF2 overexpression alleviated the inhibitory effect of miR-451 on apoptosis and oxidative stress of the OGD/R cells.

Notes: The OGD/R cells were transfected with CELF2 or miR-451. (A) Cell viability, (B) LDH level, (C) SOD activity, (D) MDA level, (E) relative ROS level, (F) apoptosis and (G) the expression of cleaved-caspase-3 were assessed using the CCK-8, LDH, SOD, MDA and ROS assays, flow cytometry and Western blot analysis, respectively. ***P<0.001. ^###P<0.001.
Abbreviations: CCK-8, Cell Counting Kit-8; FITC, fluorescein isothiocyanate; MDA, malondialdehyde; NC, negative control; OGD/R, oxygen and glucose deprivation/reoxygenation; PI, propidium iodide.