A metabolic exposure-oriented network regulation strategy for the identification of effective combination in the extract of Ginkgo biloba L.

Nowadays, network pharmacology-based methods were increasing proposed to screen synergistic or combinatorial compounds from herbal medicines (HMs), while these researches mainly focused on structural prediction or experiment-based interaction between single compound and target protein. The proportion of each chemical in the nature and their metabolic process was ignored, which might decide an optimized composition for their synergistic effect. To exact the effective combination of HMs, a metabolic distribution-oriented network regulation strategy was developed for the identification of effective combination. Firstly, comprehensive chemical profiling and metabolic exposure of HMs in a pathological state were conducted. Then the effective combination for HMs was screened by combining network regulation and the metabolic exposure level of HMs. Finally, with the extract of Ginkgo biloba (EGB) as a case, a combination of 12 active compounds was found for treating ischemia stroke, showing bioactivity equivalence with original herb. The results also indicated that beside the well-known ginkgolides and flavonoids, trace compounds might also play an important role of the holistic effect of EGB. This method can be used as an alternative for effective combination screening.

Ginkgolide K attenuates neuronal injury after ischemic stroke by inhibiting mitochondrial fission and GSK-3ß-dependent increases in mitochondrial membrane permeability.

Ginkgolide K (GK) belongs to the ginkgolide family of natural compounds found in Ginkgo biloba leaves, which have been used for centuries to treat cerebrovascular and cardiovascular diseases. We evaluated the protective effects of GK against neuronal apoptosis by assessing its ability to sustain mitochondrial integrity and function. Co-immunoprecipitation showed that Drp1 binding to GSK-3ß was increased after an oxygen-glucose deprivation/reperfusion (OGD/R) insult in cultured neuroblastoma cells. This induced Drp1 and GSK-3ß translocation to mitochondria and mitochondrial dysfunction, which was attenuated by GK. GK also reduced mitochondrial fission by increasing Drp1 phosphorylation at Ser637 and inhibiting mitochondrial Drp1 recruitment. In addition, GK exposure induced GSK-3ß phosphorylation at Ser9 and enhanced the interaction between adenine nucleotide translocator (ANT) and p-GSK-3ß. This interaction suppressed the interaction between ANT and cyclophilin D (CypD), which inhibited mitochondrial permeability transition pore (mPTP) opening. Similarly, suppression of mitochondrial fission by Mdivi-1 also inhibited GSK-3ß-induced mPTP opening. Treating mice with GK prevented GSK-3ß and Drp1 translocation to mitochondria and attenuated mitochondrial dysfunction after middle cerebral artery occlusion. We therefore propose that by inhibiting mitochondrial fission and attenuating mPTP opening, GK exerts neuroprotective effects that mitigate or prevent neuronal damage secondary to ischemic stroke.

Succinate-induced neuronal mitochondrial fission and hexokinase II malfunction in ischemic stroke: Therapeutical effects of kaempferol.

Mitochondrial dysfunction is known as one of causative factors in ischemic stroke, leading to neuronal cell death. The present work was undertaken to investigate whether succinate induces neuron apoptosis by regulating mitochondrial morphology and function. In neurons, oxygen-glucose deprivation induced succinate accumulation due to the reversal of succinate dehydrogenase (SDH) activation, leading to mitochondrial fission. Kaempferol inhibited mitochondrial fission and maintained mitochondrial HK-II through activation of Akt, and thereby protected neurons from succinate-mediated ischemi injury. Knockdown of Akt2 with siRNA diminished the effect of kaempferol, indicating that kaempferol suppressed dynamin-related protein 1 (Drp1) activation and promoted HK-II mitochondrial binding dependently on Akt. Moreover, we demonstrated that kaempferol potentiated autophagy during oxygen and glucose deprivation, contributing to protecting neuron survival against succinate insult. In vivo, oral administration of kaempferol in mice attenuated the infract volume after ischemic and reperfusion (I/R) injury and reproduced the similar mitochondrial protective effect in the brain infract area. This study indicates that succinate accumulation plays a pivotal role in I/R injury-induced neuronal mitochondrial dysfunction, and suggests that modulation of Drp1 phosphorylation might be potential therapeutic strategy to protect neuron mitochondrial integrity and treat ischemic stroke.