# Piperine improves ischemic brain injury by promoting the regulation of the AMPK/PGC-1α pathway by Apelin 13

## TL;DR

Piperine, a compound from traditional Chinese medicine, helps reduce brain damage from stroke by boosting mitochondrial function through the Apelin 13 and AMPK/PGC-1α pathways.

## Contribution

This study reveals a novel mechanism where piperine enhances Apelin 13 to activate mitochondrial biogenesis via the AMPK/PGC-1α pathway in ischemic stroke.

## Key findings

- Piperine combined with Apelin 13 significantly improved mitochondrial function and reduced oxidative stress in stroke models.
- Piperine increased Apelin 13 expression, which was essential for activating the AMPK/PGC-1α pathway and promoting mitochondrial biogenesis.
- Knocking down Apelin 13 blocked the neuroprotective effects of piperine and Apelin 13, confirming its critical role in the observed mechanism.

## Abstract

Ischemic stroke (IS) persists as the second foremost cause of mortality and the primary cause of long-term disability globally, a burden largely attributable to a paucity of effective therapeutic strategies. Piperine (PIP) is a bioactive component of traditional Chinese medicine that has shown potential to reduce cell inflammation and pyroptosis. Recent studies indicate that mitochondrial biogenesis can improve ischemic stroke.

In this study, we aimed to investigate the effect of PIP combined with Apelin 13 on mitochondrial biosynthesis in IS and determine its mechanism and whether PIP promotes Apelin 13.

We used network pharmacology to screen chemical drugs for combination therapy for IS. Male Sprague–Dawley rats were utilized to induce a model of pMCAO, and primary cortical neuron cells were extracted to establish an oxygen–sugar deprivation–reperfusion model. To evaluate the changes in mitochondrial function of neuronal cells, we observed mitochondrial membrane potential via fluorescence microscopy, detected ROS levels by flow cytometry, and determined the ATP concentration by using a chemiluminescence multifunctional microplate reader. Western blot and qRT-PCR were used to detect the protein expression and mRNA content of Apelin 13 and the AMPK/PGC-1α pathway. In addition, the underlying mechanism of action of PIP promoting Apelin 13 in the regulation of the AMPK/PGC-1α pathway by using siRNA to reduce the content of Apelin 13 in primary cortical neurons was investigated.

The results of network pharmacology research indicated that Apelin 13 affects IS. PIP combined with Apelin 13 exerts neuroprotective effects against IS. The OGD/R group showed obvious mitochondrial functional damage, reduced mitochondrial membrane potential, increased reactive oxygen species level, and decreased ATP content compared with the Con group. Compared with the OGD/R group, the mitochondrial function detection and expression level of mitochondrial biogenesis-related factors in the PIP and Apelin 13 groups significantly improved, and the neuroprotective effect was more significant when the two were combined. Our in vitro and in vivo experiments revealed that, compared with the normal group, the mRNA and protein expression of Apelin 13 in the model group significantly decreased. Furthermore, the abundance of Apelin 13 in the PIP group substantially rose compared with that in the model group. When the expression of Apelin 13 was knocked down by si-Apelin 13, si-Apelin 13 effectively blocked the individual or even combined effects of PIP and Apelin 13.

This study showed that PIP could promote Apelin 13 to activate mitochondrial biogenesis and decreased mitochondrial functional damage. The potential mechanism of activating mitochondrial biogenesis lies in the regulation of the AMPK/PGC-1α pathway. This study not only expands the understanding of the clinical application of PIP in the treatment of IS but also provides new insights into its internal mechanism.

A detailed scientific diagram with three main sections: the pMCAO model, mitochondrial function, and molecular experiments. The pMCAO model shows experiments on rats, including initial conditions and stroke effects. Mitochondrial function features assessments like JC-1 assay, ATP, and ROS levels illustrated with colored graphs and fluorescence images. The molecular experiment section displays Western Blot and RT-PCR results with bar graphs showing variations in protein and mRNA levels. Each part is interconnected with arrows, indicating the experimental progression.