Clinical Efficacy and Mechanistic Insights of Anshen Dingzhi Prescription on Breast Cancer-Related PTSD Through Network Pharmacology and Molecular Docking.

Anshen Dingzhi prescription (ADP) is a classic prescription of traditional Chinese medicine, which has been used in the treatment of neuropsychiatric diseases. However, its treatment of breast cancer-related post-traumatic stress disorder (BC-PTSD) lacks clinical research evidence and its mechanism is not clear. The present study investigated the efficacy and action mechanism of ADP against BC-PTSD. The results of the clinical trial showed that after 4 weeks of treatment, both groups showed reduced post-traumatic stress disorder checklist-civilian version (PCL-C), Pittsburgh sleep quality index (PSQI), self-rating depression scale (SDS) and self-rating anxiety scale (SAS) scores, and increased functional assessment of cancer therapy-breast (FACT-B) scores. The serum cortisol (CORT), tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1ß) levels were decreased and brain-derived neurotrophic factor (BDNF) level were increased, and the improvement of serum TNF-α, IL-1ß, and BDNF in treatment group was better than that of the control group. The overall treatment efficacy in the treatment group (43.90%) was superior to that in the control group (23.81%), and the overall incidence of adverse effects was lower than that in the control group. The results of network analysis and molecular docking showed that ADP blood components could act on IL1B, TNF, and BDNF. ADP contributes to the treatment of BC-PTSD symptoms, with a mechanism possibly related to its regulatory effect on TNF-α, IL-1ß, and BDNF levels.Trial registration: Chinese Clinical Trial Registry, http://www.chictr.org.cn,ChiCTR2300077801.

Novel mechanisms of Anshen Dingzhi prescription against PTSD: Inhibiting DCC to modulate synaptic function and inflammatory responses.

ETHNOPHARMACOLOGICAL RELEVANCE: Anshen Dingzhi prescription (ADP), documented in "Yi Xue Xin Wu", is a famous prescription for treating panic-related mental disorders such as post-traumatic stress disorder (PTSD). However, the underlying mechanism remains unclear. AIM OF THE STUDY: This study aimed to investigate the mechanisms by which ADP intervened in PTSD-like behaviors. METHODS: A mouse model of single prolonged stress (SPS) was established to evaluate the ameliorative effects and mechanisms of ADP on PTSD. Behavioral tests were used to assess PTSD-like behaviors in mice; transmission electron microscopy was used to observe changes in the ultrastructure of hippocampal synapses, and western blot, immunofluorescence, and ELISA were used to detect the expression of hippocampal deleted in colorectal cancer (DCC) and downstream Ras-related C3 botulinum toxin substrate 1 (Rac1) - P21-activated kinase 1 (PAK1) signal, as well as levels of synaptic proteins and inflammatory factors. Molecular docking technology simulated the binding of potential brain-penetrating components of ADP to DCC. RESULTS: SPS induced PTSD-like behaviors in mice and increased expression of hippocampal netrin-1 (NT-1) and DCC on the 14th day post-modeling, with concurrent elevation in serum NT-1 levels. Simultaneously, SPS also decreased p-Rac1 level and increased p-PAK1 level, the down-stream molecules of DCC. Lentiviral overexpression of DCC induced or exacerbated PTSD-like behaviors in control and SPS mice, respectively, whereas neutralization antibody against NT-1 reduced DCC activation and ameliorated PTSD-like behaviors in SPS mice. Interestingly, downstream Rac1-PAK1 signal was altered according to DCC expression. Moreover, DCC overexpression down-regulated N-methyl-d-aspartate (NMDA) receptor 2A (GluN2A) and postsynaptic density 95 (PSD95), up-regulated NMDA receptor 2B (GluN2B) and increased neuroinflammatory responses. Administration of ADP (36.8 mg/kg) improved PTSD-like behaviors in the SPS mice, suppressed hippocampal DCC, and downstream Rac1-PAK1 signal, upregulated GluN2A and PSD95, downregulated GluN2B, and reduced levels of inflammatory factors NOD-like receptor protein 3 (NLRP3), nuclear factor kappa-B (NF-κB) and interleukin-6 (IL-6). Importantly, DCC overexpression could also reduce the ameliorative effect of ADP on PTSD. Additionally, DCC demonstrated a favorable molecular docking pattern with the potential brain-penetrating components of ADP, further suggesting DCC as a potential target of ADP. CONCLUSION: Our data indicate that DCC is a key target for the regulation of synaptic function and inflammatory response in the onset of PTSD, and ADP likely reduces DCC to prevent PTSD via modulating downstream Rac1-PAK1 pathway. This study provides a novel mechanism for the onset of PTSD and warrants the clinical application of ADP.

Exploring the effect of Anshen Dingzhi prescription on hippocampal mitochondrial signals in single prolonged stress mouse model.

HEADINGS ETHNOPHARMACOLOGICAL RELEVANCE: Anshen Dingzhi prescription (ADP), which was first published in the masterpiece of traditional Chinese Medicine in the Qing Dynasty, "Yi Xue Xin Wu" (1732 CE), is documented to interrupt panic-related disorders. However, the mechanism of its action is still not clear. AIM OF THE STUDY: This study aims to investigate the effects of ADP on post-traumatic stress disorder (PTSD)-like behaviors and explore the mechanism from perspective of sirtuin1 (SIRT1)-peroxisome proliferator-activated receptor gamma co-activator 1 alpha (PGC-1α)-dependent mitochondrial function. MATERIALS AND METHODS: The changes of SIRT1-PGC-1α signal and mitochondrial function were evaluated in the hippocampus of mice receiving single prolonged stress (SPS). Later, the roles of this signaling pathway played in fear memory generalization and anxiety-like behavior in SPS mice was investigated using two agonists of this signaling pathway. On this basis, the effects of ADP (36.8 mg/kg) with definite therapeutic effects, on mitochondrial function were investigated and further confirmed by a SIRT1 inhibitor. Finally, the possible components of ADP targeting PGC-1α were monitored through bioinformatics. RESULTS: Compared with control mice, SIRT1-PGC-1α signal in the hippocampus was impaired in SPS mice, accompanied with dysfunction of mitochondria and abnormal expression of synaptic proteins. The agonists of SIRT1-PGC-1α signal, ZLN005, as well as resveratrol improved the behavioral changes of mice caused by SPS, reversed the decline of proteins in SIRT1-PGC-1α signal, mitochondrial dysfunction, and the abnormal expression of synaptic proteins. The fingerprint was established for the quality control of ADP. At a dose of 36.8 mg/kg, ADP could prevent fear memory generalization and anxiety-like behavior in SPS mice. Mechanically, ADP promoted SIRT1-PGC-1α signal and repaired mitochondrial function. Importantly, SIRT1 inhibitor, selisistat eliminated the ameliorative effects of ADP on behavioral and mitochondrial function. Through molecular docking simulation, the brain-entering components of ADP, including malkangunin, Rg5, fumarine, frutinone A, celabenzine, and inermin had high binding energy with PGC-1α. CONCLUSION: Dysfunction of SIRT1-PGC-1α-dependent mitochondrial function is attributed to SPS-triggered fear generalization and anxiety-like behavior, and ADP could improve PTSD-like behaviors likely through activating this signaling pathway.

The improving effect and mechanisms of Anshen Dingzhi Prescription on Alzheimer's disease-like behavior induced by D-galactose combined with AβO in mice / 药学学报

The purpose of this study was to explore the improving effect of Anshen Dingzhi Prescription (ADP) on Alzheimer's disease (AD)-like behavior in mice and its mechanisms. The main chemical components of ADP were identified by ultra performance liquid chromatography-time-of-flight mass spectrometry (UPLC-Q-TOF-MS). The AD-like mouse model was induced by D-galactose (D-gal) combined with Aβ1-42 oligomer (AβO). The effect of ADP on AD-like behavior in mice was assessed using various behavioral experiments; pathomorphological changes in mouse hippocampal tissue were observed by Nissl staining and transmission electron microscopy; ELISA was used in the assessment of oxidative stress factors and inflammation-related factor levels; Western blot was performed to detect the expression of Aβ, Tau and glial fibrillary acidic protein (GFAP) proteins. The active components of ADP were screened according to TCMSP and HERB database, and the action targets of active components were predicted by Swiss Target Prediction platform. In addition, the targets of AD were predicted through DisGeNET database. Further, GO and KEGG enrichment analysis of common targets was carried out by Metascape database. Combined with the results of GO and KEGG analysis, in vivo experiments were carried out to explore the potential mechanism of ADP improving AD-like behavior in mice from the PI3K/Akt, calcium signal pathway and synaptic function. Finally, the core components of ADP were molecularly docked to the validated targets using Autodock Vina. Animal experiments were approved by the Animal Ethics Committee of Anhui University of Chinese Medicine (approval number: AHUCM-mouse-2021080). The results showed that the five chemical components, including ginsenoside Rg1, ginsenoside Rb1, tenuifolin, poricoic acid B and α-asarone were found in the ADP. ADP significantly improved the anxiety-like behavior and memory impairment, protected hippocampal neurons, decreased the levels of oxidative stress and inflammation, and inhibited the expression of Aβ and p-Tau induced by D-galactose combined with AβO in mice. The results of network pharmacology suggested that PI3K/Akt, calcium signal pathway and cell components related to postsynaptic membrane might be the key factors for ADP to improve AD. Animal experiments revealed that ADP up-regulated N-methyl-D-aspartate receptor 2A (GluN2A), postsynaptic density protein 95 (PSD95), calpain-1, phosphorylated protein kinase B (p-Akt), phosphorylated cAMP response element binding protein (p-CREB), brain-derived neurotrophic factor (BDNF) expression and inhibited p-GluN2B and calpain-2 expression in the hippocampus of AD-like mice. The molecular docking results demonstrated that the core components of ADP, such as panaxacol, dehydroeburicoic acid, deoxyharringtonine, etc. had a high binding ability with the validated targets GRIN2A, GRIN2B, PSD95, etc. In summary, our results indicate ADP improves AD-like pathological and behavioral changes induced by D-galactose combined with AβO in mice, and the mechanism might be related to the NMDAR/calpain axis and Akt/CREB/BDNF pathway.

Anshen Dingzhi prescription in the treatment of PTSD in mice: Investigation of the underlying mechanism from the perspective of hippocampal synaptic function.

BACKGROUND: Anshen Dingzhi prescription (ADP) is an important prescription for the treatment of mental diseases in traditional Chinese medicine and is widely used to treat neuropsychiatric disorders. PURPOSE: To explore the ameliorative effect of ADP on post-traumatic stress disorder (PTSD)-like behaviors in mice and determine the underlying mechanism. METHODS: The constituents of ADP were analyzed by UPLC-Q-TOF/MS. The PTSD-like behaviors of mice subjected to single prolonged stress (SPS) were evaluated using behavioral tests. Potential pathological changes in the hippocampus were assessed by hematoxylin and eosin (H&E) staining. Western blotting and immunohistochemistry (IHC) were employed to detect the expression of proteins involved in relevant signaling pathways. RESULTS: Five quality control markers (ginsenoside Rg1, ginsenoside Rb1, tenuifolin, poricoic acid B, and α-asarone) were detected in the ADP solution. The ginsenoside Rg1 content in ADP was found to be 0.114 mg/g. Mice subjected to SPS showed obvious fear generalization and anxiety-like behaviors. ADP treatment prevented the behavioral changes caused by exposure to SPS. Compared with control animals, the number of normal pyramidal cells in the hippocampal CA1 region of mice exposed to SPS was decreased and the number of degenerating pyramidal cells was increased; however, ADP administration could counteract these effects. Furthermore, the protein expression of BDNF, p-TrkB, µ-calpain, PSD95, GluN2A, GluA1, p-AKT, p-mTOR, and ARC was decreased, while that of PTEN and GluN2B was increased in the hippocampus of mice subjected to SPS compared with that in control animals; however, these changes in protein expression were reversed following ADP treatment. Importantly, the ameliorative effect of ADP on PTSD-like behaviors and synaptic protein expression were inhibited by rapamycin administration. CONCLUSIONS: ADP administration improves PTSD-like behaviors in mice and this effect may be mediated through an mTOR-dependent improvement in synaptic function in the hippocampus.