Retracted Article: Berberine alleviates amyloid beta-induced injury in Alzheimer's disease by miR-107/ZNF217

Abstract

Berberine plays a neuroprotective role in neurodegenerative disorders, including Alzheimer's disease (AD). However, the underlying mechanism by which berberine inhibits AD progression remains largely unclear. The AD model was established using PC12 cells after treatment of amyloid beta (Aβ)_25-35. Cells were transfected with microRNA (miRNA)-107 mimic, inhibitor, zinc finger protein 217 (ZNF217) overexpression or corresponding negative controls. Cell viability, apoptosis and inflammatory cytokine secretion were measured by MTT, flow cytometry or enzyme linked immunosorbent assay, respectively. The expressions of miR-107, ZNF217 and phosphorylated tau (p-Tau) were detected by quantitative real-time polymerase chain reaction or Western blot. The association between miR-107 and ZNF217 was explored by luciferase reporter assay and RNA immunoprecipitation. Berberine attenuated Aβ_25-35-induced viability suppression in PC12 cells. Moreover, berberine inhibited the Aβ_25-35-induced increase of inflammatory cytokine expression, apoptosis and p-Tau level in PC12 cells. miR-107 expression was reduced in Aβ_25-35-treated PC12 cells and its overexpression alleviated Aβ_25-35-induced injury, which was further weakened by combination with berberine. ZNF217 was a target of miR-107 and its addition reversed miR-107-mediated inhibition of inflammatory injury, apoptosis and phosphorylation of tau. Besides, ZNF217 protein level was decreased by berberine via regulating miR-107 in Aβ_25-35-treated PC12 cells. Berberine protected against Aβ_25-35-induced inflammatory injury, apoptosis and phosphorylation of tau by regulating miR-107 and ZNF217, indicating berberine as a promising neuroprotective agent for therapeutics of AD.

Fig. 1. Berberine regulates cell viability of PC12. (A) The chemical structure of berberine. (B) Cell viability was measured in PC12 cells after treatment of different concentrations of berberine for 24 h by MTT. (C) Cell viability was detected in PC12 cells after treatment of different concentrations of berberine and 10 μM Aβ for 24 h by MTT. *P < 0.05, **P < 0.01.

Fig. 2. Berberine inhibits Aβ_25-35-induced injury in PC12 cells. PC12 cells were exposed to 1 μM berberine and 10 μM Aβ for 24 h. The expressions of IL-1β, IL-6 and TNF-α in cell medium (A–C), cell apoptosis (D) and p-Tau level (E) were measured by ELISA, flow cytometry or Western blot assays, respectively. **P < 0.01, ***P < 0.001.

Fig. 3. Berberine suppresses Aβ_25-35-induced injury in PC12 cells by up-regulating miR-107. PC12 cells were transfected with miR-107 mimic or miR-NC for 24 h and then treated with 1 μM berberine and 10 μM Aβ for 24 h. The miR-107 level (A), expressions of IL-1β, IL-6 and TNF-α in cell medium (B–D), cell apoptosis (E) and p-Tau level (F) were measured by qRT-PCR, ELISA, flow cytometry or Western blot assays, respectively. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig. 4. ZNF217 is a target of miR-107. (A) The potential binding sites of miR-107 and ZNF-217. (B) Luciferase activity was measured in PC12 cells after co-transfection of ZNF217-WT or ZNF217-MUT and miR-107 mimic or miR-NC. (C) The enrichment of ZNF217 was detected in PC12 cells transfected with miR-107 mimic or miR-NC after RIP assay. (D) The protein level of ZNF217 was measured in PC12 cells transfected with miR-NC, miR-107 mimic, anti-miR-107 or anti-miR-NC by Western blot. **P < 0.01, ***P < 0.001, ns: no significance, P > 0.05.

Fig. 5. miR-107 suppresses Aβ_25-35-induced injury in PC12 cells by regulating ZNF217. PC12 cells were transfected with miR-NC, miR-107 mimic, miR-107 mimic and ZNF217 or pcDNA for 24 h and then treated with 1 μM berberine and 10 μM Aβ for 24 h. The ZNF217 protein level (A), expressions of IL-1β, IL-6 and TNF-α in cell medium (B–D), cell apoptosis (E) and p-Tau level (F) were detected by Western blot, ELISA or flow cytometry assays, respectively. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig. 6. Berberine decreases ZNF217 protein expression by regulating miR-107 in Aβ_25-35-treated PC12 cells. PC12 cells were transfected with miR-NC, miR-107 mimic, anti-miR-NC or anti-miR-107 for 24 h and treated with 1 μM berberine and 10 μM Aβ for 24 h. (A and B) The abundance of ZNF217 protein was measured in treated PC12 cells by Western blot. *P < 0.05, ***P < 0.001.