Fig 2: miR-145-5p targets PXN. (A) The potential targets of miR-145-5p were predicted by three bioinformatics analysis tools. (B) The lncRNA-miRNA-mRNAs interaction network on the basis of predicted results. (C) The binding sites between candidate genes and miR-145-5p and the minimum free energy were predicted by RNAhybrid. (D) The binding sites between miR-145-5p and PXN were predicted by Targetscan database. (E) The binding relation between miR-145-5p and PXN was validated through dual luciferase reporter assays (n = 3). (F) The enrichment level of Fendrr and miR-145-5p was detected by RIP assay (n = 6). (G) The miR-145-5p and PXN co-localization in hippocampal tissue was detected by FISH. Scale bar, 20 µm; Blue: DAPI; red: PXN; green: miR-145-5p. (H) The PXN expression was examined by RT-qPCR after the up-regulation of miR-145-5p (n = 6). (I) The PXN expression was detected by Western Blot after the up-regulation of miR-145-5p. (J) The bar chart shows quantification for PXN expression (n = 6). Datasets reflect the mean ± SD. ** P < 0.01 versus the agomir-NC group or 2VO+agomir-NC group; *** P < 0.001 versus the Ago2 group.

Fig 3: The mechanism of action of TMAO on neuroinflammation after VaD. TMAO can promote neuroinflammation and aggravate neural damage in vascular dementia by regulating the lncRNA Fendrr/miR-145-5p/PXN axis.

Fig 4: Inhibition of miR-145-5p reversed the Fendrr knockdown-mediated influence on 2VO rats. (A) The PXN expression was examined by RT-qPCR after the down-regulation of Fendrr (n = 6). (B) The PXN expression was examined by Western Blot after the down-regulation of Fendrr. (C) The bar chart shows quantification for PXN expression (n = 6). (D) Representational movement tracks, (E) total distance traveled and (F) time spent in the center in each group were measured using the OFT (n = 24). (G) The escape latency and (H) swimming speeds in each group were recorded by MWM (n = 24). (I) Neuronal damage in the CA1 region of the hippocampus was assessed by Nissl staining in each group of rats. Scale bar, 20 µm (n = 6). (J) The bar chart shows the quantification of the Nissl-positive cells (n = 6). (K) The PXN expression was examined by RT-qPCR after the down-regulation of Fendrr and down-regulation of miR-145-5p (n = 6). (L) The PXN expression was examined by Western Blot after the down-regulation of Fendrr and down-regulation of miR-145-5p (n = 6). (M) The bar chart shows quantification for PXN expression (n = 6). The inflammation-related indexes (N) TNF-α, (O) IL-1β, (P) IL-6 in the hippocampus of 2VO rats were detected by ELISA (n = 6). Datasets reflect the mean ± SD. * P < 0.05 versus the 2VO+sh-NC+antagomir-NC group; ** P < 0.01 versus the 2VO+sh-NC group or 2VO+sh-NC+antagomir-NC group; # P < 0.05 and ## P < 0.01 versus the 2VO+sh-Fendrr+antagomir-NC group.

Fig 5: Knockdown of PXN can reduce neurological function injury in 2VO rats. (A) The expression of PXN in hippocampus of rats was detected by RT-qPCR (n = 6). (B) The expression of PXN in hippocampus of rats was tested by Western Blot. (C) The bar chart shows quantification for PXN expression (n = 6). (D) The knockdown efficiency of PXN expression was detected by RT-qPCR (n = 6). (E) The efficiency of knockdown of PXN expression was detected by Western Blot. (F) The bar chart shows quantification for PXN expression (n = 6). (G) Representational movement tracks, (H) total distance traveled and (I) time spent in the center in each group were measured using the OFT (n = 24). (J) The escape latency and (K) swimming speeds in each group were recorded by MWM (n = 24). (L) Neuronal damage in the CA1 region of the hippocampus was assessed by Nissl staining in each group of rats. Scale bar, 20 µm (n = 6). (M) The bar chart shows the quantification of the Nissl-positive cells (n = 6). The inflammation-related indexes (N) TNF-α, (O) IL-1β, (P) IL-6 in the hippocampus of 2VO rats were detected by ELISA (n = 6). Datasets reflect the mean ± SD. * P < 0.05 versus the 2VO+sh-NC group; ** P < 0.01 versus the Sham group or 2VO+sh-NC group.