Dajianzhong decoction ameliorated D-gal-induced cognitive aging by triggering mitophagy in vivo and in vitro

Dajianzhong decoction (DJZ) is a classical famous formula for treating yang-deficiency-syndrome in traditional Chinese medicine and recorded in Jin-Kui-Yao-Lue in Dynasty of Dong Han. Cognitive aging can present similar features of mitochondrial energy deficits to the clinical features of Yang deficiency. However, there is poor understanding of the effects of DJZ treatment on mitophagy in cognitive aging. The aims of this work were to decipher the effectiveness and mechanism of DJZ against cognitive aging, focusing on mitophagy. YFP-Parkin HeLa cells, D-galactose (D-gal) -induced mice (500 mg/kg for 35 d, s. c.) and SH-SY5Y cells (80 mg/ml for 6 h) were established. Firstly, the formation of YFP-Parkin puncta (a well-known mitophagy marker) in YFP-Parkin HeLa cells was employed to discover the mitophagy induction of DJZ. Moreover, the genes and proteins related to PINK1/Parkin pathway and mitochondrial functions were evaluated after treatment with DJZ in vivo (3.5 g/kg or 1.75 g/kg, i. g, 35 d) and in vitro (0.2, 2 and 20 μg/ml, 12 h). Furthermore, the effectiveness of DJZ (3.5 g/kg or 1.75 g/kg, i. g) for alleviating cognitive aging and nerve damage was measured in D-gal mice. Finally, siPINK1 was applied to reverse validation of DJZ in vitro. The formation of YFP-Parkin puncta in YFP-Parkin HeLa cells was markedly induced by DJZ in a dose-dependent manner. The immunofluorescence intensity of Parkin and the protein expression of Parkin in mitochondrial membrane in D-gal mice were significantly increased after treatment of DJZ. The inhibition of PINK1/Parkin pathway in D-gal-induced mice and SH-SY5Y cells was significantly activated by DJZ. Simultaneously, the impairment of mitochondrial functions induced by D-gal were markedly reversed by DJZ. In addition, DJZ significantly ameliorated the neuropathological injury and cognitive declines in D-gal mice. Finally, after PINK1 was knocked down by siPINK1 in vitro, the neuroprotective effects of DJZ and the Parkin enhancement effect of DJZ were markedly reversed. Our findings firstly showed DJZ could relieve cognitive aging through facilitating PINK1/Parkin-mediated mitophagy to protect against mitochondrial functions, indicating DJZ may be regarded as a promising intervention in cognitive aging.