Buyang Huanwu Decoction restores the balance of mitochondrial dynamics after cerebral ischemia-reperfusion through calcium overload reduction by the PKCε-Nampt-Sirt5 axis

Ethnopharmacological relevance: Stroke is a common condition that poses a significant threat to human health. Buyang Huanwu Decoction (BYHWD) is a traditional treatment used for stroke management. However, the exact mechanism by which BYHWD mitigates cerebral ischemia-reperfusion by regulating calcium overload and restoring mitochondrial function is not yet fully understood. Aim: The objective of this research was to examine the neuroprotective properties of BYHWD in reducing the damage produced by cerebral ischemia/reperfusion (I/R) injury via the modulation of calcium overload and mitochondrial dynamics (MD). Methods: MCAO/R model success was evaluated via PSI laser scatter flowmetry. The neurological function scores were assessed. The cerebral infarct (CI) volume was detected via TTC staining. NeuN expression was detected via immunohistochemistry, and degenerated neurons were observed via FJC staining. The mitochondrial permeability transition pore (mPTP), the mitochondrial membrane potential (MMP), and ATP were detected. The reactive oxygen species (ROS) content and the NAD+/NADH ratio were determined. The glutamate (Glu) and glutamine (Gln) contents as well as the Ca2+ concentration were determined. The expression of PKCa, p-PKCa, namely, Sirt5, GLS, Drp1, p-Drp1 616, Fis1, Opa1, and Mfn2 was determined via Western blotting. Immunohistochemistry was used to detect p-PKCa, which is expressed at high levels. Immunofluorescence was used to detect p-Drp1 616, Opa1 and Sirt5 fluorescence intensity. Results: BYHWD treatment enhanced neurological function, decreased the amount of CI, mitigated neuronal damage, decreased mPTP opening, restored the MMP, increased ATP synthesis, and decreased the ROS content after brain I/R. It also increased PKCa, p-PKCa, Sirt5, GLS, Opa1 and Mfn2 expression; downregulated p-Drp1 616, Drp1 and Fis1 expression; elevated the NAD+/NADH ratio and Gln content; and decreased the Glu content and Ca2+ concentration. The effects of BYHWD were reversed by the administration of the PKCa inhibitor aV1-2. BYHWD administration led to increased PKCa mRNA expression. Conclusions: BYHWD modulates MD by diminishing calcium overload through the PKCa-Nampt-Sirt5 axis, which restores mitochondrial function and mitigates brain I/R damage.