Tongxinluo Exerts Inhibitory Effects on Pyroptosis and Amyloid-ß Peptide Accumulation after Cerebral Ischemia/Reperfusion in Rats.

Amyloid-ß peptide (Aß) accumulation is a detrimental factor in cerebral ischemia/reperfusion (I/R) injuries accounting for dementia induced by ischemic stroke. In addition to blood brain barrier (BBB), the glymphatic system mediated by aquaporin-4 (AQP-4) on astrocytic endfeet functions as an important pathway for the clearance of Aß in the brain. Cerebral I/R induced astrocytic pyroptosis potentially causes the AQP-4 polarization loss and dysfunctional BBB-glymphatic system exacerbating the accumulation of Aß. Furthermore, Aß toxicity has been identified as a trigger of pyroptosis and BBB damage, suggesting an amplified effect of Aß accumulation after cerebral I/R. Therefore, based on our previous work, this study was designed to explore the intervention effects of Tongxinluo (TXL) on astrocytic pyroptosis and Aß accumulation after cerebral I/R in rats. The results showed that TXL intervention obviously alleviated the degree of pyroptosis by downregulating expression levels of cleaved caspase-11/1, N-terminal gasdermin D, nucleotide-binding oligomerization domain-like receptors pyrin domain containing 3 (NLRP3), interleukin-6 (IL-6), and cleaved IL-1ß and abated astrocytic pyroptosis after cerebral I/R. Moreover, TXL intervention facilitated to restore AQP-4 polarization and accordingly relieve Aß accumulation around astrocytes in ischemic cortex and hippocampus as well as the formation of toxic Aß (Aß 1-42 oligomer). Our study indicated that TXL intervention could exert protective effects on ischemic brain tissues against pyroptotic cell death, inhibit astrocytic pyroptosis, and reduce toxic Aß accumulation around astrocytes in cerebral I/R injuries. Furthermore, our study provides biological evidence for the potential possibility of preventing and treating poststroke dementia with TXL in clinical practice.

Yi-Zhi-Fang-Dai Formula Exerts a Protective Effect on the Injury of Tight Junction Scaffold Proteins in Vitro and in Vivo by Mediating Autophagy through Regulation of the RAGE/CaMKKß/AMPK/mTOR Pathway.

Alzheimer's disease (AD) is a chronic neurodegeneration disease that is closely related to the abnormal tight junction scaffold proteins (TJ) proteins of the blood-brain barrier (BBB). Recently, Yi-Zhi-Fang-Dai Formula (YZFDF) had exerted a neuronal protective effect against amyloid peptide (Aß) toxicity. Still, the therapeutic mechanism of YZFDF in restoring Aß-induced injury of TJ proteins (ZO-1, Occludin, and Claudin-5) remains unclear. This study aimed to explore the underlying mechanism of YZFDF in alleviating the injury of TJ proteins. We examined the impacts of YZFDF on autophagy-related proteins and the histopathology of Aß in the APP/PS1 double-transgenic male mice. We then performed the free intracellular calcium levels [Ca2+]i analysis and the cognitive behavior test of the AD model. Our results showed that YZFDF ameliorated the injury of TJ proteins by reducing the mRNA transcription and expression of the receptor for advanced glycation end-products (RAGE), the levels of [Ca2+]i, calmodulin-dependent protein kinase ß (CaMKKß), phosphorylated AMP-activated protein kinase (AMPK). Accordingly, YZFDF increased the expression of the phosphorylated mammalian targets of rapamycin (mTOR), leading to inhibition of autophagy (downregulated LC3 and upregulated P62). Moreover, the Aß1-42 oligomers-induced alterations of autophagy in murine mouse brain capillary (bEnd.3) cells were blocked by RAGE small interfering RNA (siRNA). These results suggest that YZFDF restored TJ proteins' injury by suppressing autophagy via RAGE signaling. Furthermore, YZFDF reduced the pathological precipitation of Aß in the hippocampus, and improved cognitive behavior impairment of the AD model suggested that YZFDF might be a potential therapeutic candidate for treating AD through RAGE/CaMKKß/AMPK/mTOR-regulated autophagy pathway.

Yi-Zhi-Fang-Dai Formula Protects against Aß1-42 Oligomer Induced Cell Damage via Increasing Hsp70 and Grp78 Expression in SH-SY5Y Cells.

Yi-Zhi-Fang-Dai formula (YZFDF) is an experiential prescription used to cure dementia cases like Alzheimer's disease (AD). In this study, the main effective compounds of YZFDF have been identified from this formula, and the neuroprotective effect against Aß1-42 oligomer of YZFDF has been tested in SH-SY5Y cells. Our results showed that YZFDF could increase cell viability and could attenuate endothelial reticula- (ER-) mediated apoptosis. Evidence indicated that protein folding and endothelial reticula stress (ERS) played an important role in the AD pathological mechanism. We further explored the expression of Hsp70, an important molecular chaperon facilitating the folding of other proteins, and Grp78, the marker protein of ERS in SH-SY5Y cells. Data told us that YZFDF pretreatment could influence the mRNA and protein expression of these two proteins. At last, we also found that YZFDF pretreatment could activate Akt in SH-SY5Y cells. All these above indicate that YZFDF could be a potent therapeutic candidate for AD treatment.

EGb761 improves cognitive function and regulates inflammatory responses in the APP/PS1 mouse.

There is accumulating evidence that the Ginkgo biloba extract EGb761 may help to prevent Alzheimer's disease (AD). However, the underlying mechanism of its action remains to be elaborated. In this study, we examined the effects of EGb761 using the APP/PS1 transgenic mouse model of AD. Two-month-old APP/PS1 mice were supplemented with EGb761 daily for 6months. We found that this chronic treatment with EGb761 improved the cognitive function of these mice and also significantly alleviated amyloid plaque deposition. Although the level of insoluble amyloid beta (Aß) was decreased, the soluble content of Aß was not changed after administration of EGb761. We then determined the changes in central inflammation and observed that the activated microglia around amyloid plaque was increased in these treated mice. We also found that chronic EGb761 treatment downregulated pro-inflammatory cytokines and inducible nitric oxide synthase (iNOS), and upregulated anti-inflammatory cytokines and Arginase-1 (Arg-1), suggesting that EGb761 regulated the phenotype of activated microglia in the APP/PS1 mouse brain. In support of this, pretreatment of the BV2 microglial cell line with EGb761 inhibited the inflammatory reaction to Aß. Furthermore, the addition of conditioned media derived from BV2 cells that were co-treated with Aß and EGb761, protected neurons against treatment of Aß and inhibited apoptotic damage. Taken together, our results demonstrated that EGb761 provided a protective effect in APP/PS1 mouse. This protection was correlated with an inhibition of the pro-inflammatory effects of microglia and an induction of anti-inflammatory effects. These results strongly suggest that EGb761 provides a protective effect in APP/PS1 mouse via regulation of inflammation in the brain.