Blockade on Lin28a Prevents Cognitive Impairment and Disruption of the Blood-Brain Barrier Induced by Chronic Cerebral Hypoperfusion

Abstract

Lin28a is an RNA-binding protein involved in the translation and regulation of multiple mRNAs. Lin28a is overexpressed in animal models of brain injury. Similarly, our preliminary study found increased Lin28a expression levels in the animal models four to seven days after chronic cerebral hypoperfusion. Therefore, this current study aimed to evaluate the effects of modulating Lin28a on cognition and brain functions. Vascular dementia (VaD) was induced in 12-week-old male Wistar rats using permanent bilateral common carotid artery occlusion (BCCAO), and these rats were treated with Lin28a siRNA on the fourth and seventh day after BCCAO. From the 42nd day after BCCAO, cognitive behavioral experiments were performed for two weeks. VaD induced by BCCAO resulted in cognitive impairment and microglial activation. Lin28a expression was upregulated after BCCAO. Lin28a siRNA treatment alleviated cognitive impairment and overexpression of GFAP and Iba-1 in the brain. Furthermore, the treatment ameliorated the VaD-induced damage to the blood-brain barrier (BBB) components, including PECAM-1, PDGFRβ, occludin, claudin-9, and ZO-1. CCR6 activation after VaD, associated with BBB disruption, was diminished by treatment with Lin28a siRNA. The treatment inhibited VaD-induced microglial activity and alleviated BBB damage. Thus, blocking Lin28a may alleviate cognitive impairment caused by VaD.

Keywords: Lin28a; blood-brain barrier; chronic cerebral hypoperfusion; cognitive impairment; vascular dementia.

Figure 1

Expression of Lin28a in the hippocampal dentate gyrus after VaD. Representative images of Lin28a expression in the hippocampal dentate gyrus at 4, 7, 14, 28, and 42 days after BCCAO surgery or sham operation. Data are presented as mean ± S.E.M. * p < 0.05, compared to the sham group. A scale bar represents 50 μm.

Figure 2

Expression of Iba-1 and GFAP in the hippocampal dentate gyrus after VaD. Representative images of Iba-1 and GFAP expression in the hippocampal dentate gyrus at 4, 7, 14, 28, and 42 days after BCCAO surgery or sham operation. Data are presented as mean ± S.E.M. * p < 0.05, compared to the sham group. A scale bar represents 50 μm.

Figure 3

Blocking Lin28a ameliorates the loss of tight junction proteins in the brain. (A) Representative photographs of immunofluorescent staining of occludin (green), PDGFRβ (green), and RECA-1 (red) in the brain. (B) Representative bands of PECAM-1, PDGFRβ, occludin, claudin-9 and ZO-1, eNOS, and Laminin proteins in the brain were detected by western blotting. The data are presented as the mean ± S.E.M. * p < 0.05, compared to the sham group. # p < 0.05, compared to the VaD group. A scale bar represents 20 μm.

Figure 4

Blocking Lin28a inhibits the activation of GFAP and CCR6 in the brain after VaD. Representative photographs of immunofluorescent staining of GFAP (green), Lin28a (red), CCR6 (red), and DAPI (blue) in the brain. The data are presented as the mean ± S.E.M. * p < 0.05, compared to the sham group. # p < 0.05, compared to the VaD group. A scale bar represents 20 μm.

Figure 5

Effect of blocking Lin28a on VaD-induced cognitive impairment using passive avoidance and radial 8-arm maze tests. (A) In the passive avoidance test, VaD+Lin28a siRNA group show significantly improved latency times compared to the VaD group (B) In the radial 8-arm maze test, VaD+Lin28a siRNA group show significantly improved number of correct and errors choices compared to the VaD group. The data are presented as the mean ± S.E.M. * p < 0.05, compared to the sham group. # p < 0.05, compared to the VaD group.