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.

Shenqi Yizhi prescription prevents AßO-induced memory impairment in mice by regulating the contractility of brain pericytes.

BACKGROUND: Cerebral microcirculation disturbance manifested by decrease of cerebral blood flow (CBF) is one of early features of Alzheimer's disease (AD). Shenqi Yizhi prescription (SQYZ) is widely used in the treatment of AD. However, the effect of SQYZ on the early feature of AD is not clarified. PURPOSE: To explore the effect and mechanism of SQYZ on AD-like behavior from the perspective of early pathological features of AD. METHODS: The fingerprint of SQYZ was established by ultra-high-performance liquid chromatograph. The improvement effect of SQYZ on Aß1-42 Oligomer (AßO)-induced AD-like behavior of mice was evaluated by behavioral test. The changes of CBF were detected by laser doppler meter and laser speckle imaging. The pathological changes of the hippocampus were observed by HE staining and transmission electron microscope. The expressions of intercellular communication molecules were detected by western blotting or immunofluorescence staining. The content of platelet-derived growth factor-BB (PDGF-BB) was detected by ELISA. Finally, the core components of SQYZ were docked with platelet-derived growth factor receptor beta (PDGFRß) using AutoDock Vina software. RESULTS: The similarity of the components in SQYZ extracted from different batches of medicinal materials was higher than 0.9. SQYZ administration could improve AßO-induced memory impairment and CBF reduction. Compared with the sham group, the number of neurons in the hippocampi of AßO group was significantly reduced, and the microvessels were shrunken and deformed. By contrary, SQYZ administration mitigated those pathological changes. Compared with the sham mice, the expressions of CD31, N-cadherin, PDGFRß, glial fibrillary acidic protein, phosphorylation of focal adhesion kinase, integrin ß1, and integrin α5 in the hippocampi of AßO mice were significantly increased. However, SQYZ administration significantly reduced AßO-induced expression of those proteins. Interestingly, the effect of PDGFRß inhibitor, sunitinib demonstrated a consistent modulating effect as SQYZ. Finally, the brain-entering components of SQYZ, including ginsenoside Rg5, coptisine, cryptotanshinone, dihydrotanshinone IIA, stigmasterol, and tanshinone IIA had high binding force with PDGFRß, implicating PDGFRß as a potential target for SQYZ. CONCLUSIONS: Our data indicate that SQYZ improves CBF in AßO-triggered AD-like mice through inhibiting brain pericyte contractility, indicating the treatment potential of SQYZ for AD at the early stage.

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.

Effects of electroacupuncture on the microvascular structure and related protein expression in the hippocampus of vascular dementia rats. / ç”µé’ˆå¯¹è¡€ç®¡æ€§ç—´å‘†å¤§é¼ æµ·é©¬å¾®è¡€ç®¡ç»“æž„å’Œç›¸å ³è›‹ç™½è¡¨è¾¾çš„å½±å“.

OBJECTIVES: To explore the effect of electroacupuncture (EA) at "Baihui" (GV20) and "Shenting" (GV24) on the microvascular structure and related protein expression in the hippocampus of vascular dementia (VD) rat model, and to investigate the mechanism of EA in the treatment of VD. METHODS: A total of 24 SD rats were randomly divided into sham operation, model, EA, and oxiracetam groups, with 6 rats in each group. Multiple cerebral infarction method was used to establish VD model. In the EA group, EA was applied to GV20 and GV24 for 30 min, once daily for 14 days. Rats in the oxiracetam group were treated with oxiracetam (50 mg/kg) by intraperitoneal injection, and the course of treatment was the same as that in the EA group. Learning and memory ability were evaluated by using Morris water maze test and new object recognition experiment. The cerebral blood flow was detected by laser doppler. The microvascular structure in the hippocampus was observed by transmission electron microscopy. The expression of vascular structure related proteins of platelet-derived growth factor receptor (PDGFR)-ß, platelet endothelial cell adhesion molecule-1(CD31), neural cadherin N-Cadherin, Zonula occludens protein-1(ZO-1) in the hippocampus were measured by Western blot. RESULTS: Compared with the sham operation group, the rats in the model group had a significant increase in time of first crossing the platform, a significant decrease in the number of crossing platform and the new object preference index (P<0.05), a significant decrease in cerebral blood flow (P<0.05), and a significant increase in the brain weight (P<0.05). The structure boundary of pericyte and endothelial cells in the microvessels of the hippocampal CA1 area of model group was blurred, accompanied by obvious edema around the vessels and the reduction of tight junctions. The protein expression levels of PDGFR-ß, CD31, N-Cadherin, ZO-1 were significantly decreased in the model group compared with those in the sham operation group (P<0.05). Compared with the model group, the time of first crossing the platform of rats in the EA and oxiracetam group was shortened, the number of crossing platform were increased (P<0.05), the cerebral blood flow was increased (P<0.05), the brain weight was decreased (P<0.05), the morphology and structure of pericyte and endothelial cells in the microvessels of hippocampal CA1 area were intact, accompanied by the increase of tight junctions. Additionally, Compared with the model group, the EA group had a significant increase in the new object preference index (P<0.05), the protein expression levels of PDGFR-ß, CD31, ZO-1 in the EA group were increased (P<0.05), and the expression of PDGFR-ß, N-Cadherin, ZO-1 in the oxiracetam group were increased (P<0.05). CONCLUSIONS: EA at GV20 and GV24 can improve the learning and memory ability of VD rats, and the mechanism may be related to the repair of microvascular structures and improvement of cerebral blood flow.

Neurovascular glial unit: A target of phytotherapy for cognitive impairments.

BACKGROUND: Neurovascular glial unit (NVGU) dysfunction has been reported to be an early and critical event in the pathophysiology of Alzheimer's disease (AD) and vascular dementia (VD). Although herbal medicines, with their favorable safety profiles and low adverse effects, have been suggested to be useful for the treatment of cognitive impairment, the potential role of the NVGU as the target of the effects of herbal medicines is still unclear. PURPOSE: This review aimed to retrieve evidence from experimental studies of phytopharmaceuticals targeting the NVGU for the treatment of cognitive impairment in AD and VD, and discussed the potential of phytopharmaceuticals to improve cognitive impairment from the perspective of the NVGU. STUDY DESIGN AND METHODS: We systematically searched PubMed, Google Scholar, Web of Science, and CNKI. The keywords used for searching information on the NVGU in the treatment of cognitive impairments included "Alzheimer's disease," "Vascular dementia," "Herbal medicines," "Natural products," "Neurovascular," "Adverse reaction," and "Toxicity, etc." We selected studies on the basis of predefined eligibility criteria. RESULTS: NVGU mainly consists of endothelial cells, pericytes, astrocytes, microglia, oligodendrocytes, and neurons, and damage to these cells can induce cognitive impairment by impairing the blood-brain barrier (BBB) and cerebral blood flow (CBF) as well as neuronal function. The active components of herbal medicines, including Ginkgo biloba L., Ginseng Radix et Rhizoma, Epimedium Folium, Chuanxiong Rhizoma, Carthami flos, and Acorus tatarinowii Schott, as well as traditional Chinese medicine prescriptions have shown the potential to improve BBB function and increase CBF to prevent cognitive impairment by inhibiting astrocyte and microglia activation, protecting oligodendrocyte myelin function, reducing neuronal apoptosis, and promoting angiogenesis. CONCLUSIONS: Herbal medicines demonstrate great potential to prevent cognitive impairment. Multiple components from herbal medicines may function through different signaling pathways to target the NVGU. Future studies using novel drug-carrier or delivery systems targeting the NVGU will certainly facilitate the development of phytopharmaceuticals for AD and VD.

Tenuifolin in the prevention of Alzheimer's disease-like phenotypes: Investigation of the mechanisms from the perspectives of calpain system, ferroptosis, and apoptosis.

Polygala tenuifolia was documented to calm the mind and promote wisdom. However, its underlying mechanisms are still unclear. This study aimed to investigate the mechanisms underlying the effects of tenuifolin (Ten) on Alzheimer's disease (AD)-like phenotypes. We first applied bioinformatics methods to screen the mechanisms of P. tenuifolia in the treatment of AD. Thereafter, the d-galactose combined with Aß1-42 (GCA) was applied to model AD-like behaviors and investigate the action mechanisms of Ten, one active component of P. tenuifolia. The data showed that P. tenuifolia actioned through multi-targets and multi-pathways, including regulation of synaptic plasticity, apoptosis, and calcium signaling, and so forth. Furthermore, in vitro experiments demonstrated that Ten prevented intracellular calcium overload, abnormal calpain system, and down-regulation of BDNF/TrkB signaling induced by GCA. Moreover, Ten suppressed oxidative stress and ferroptosis in HT-22 cells induced by GCA. Calpeptin and ferroptosis inhibitor prevented the decrease of cell viability induced by GCA. Interestingly, calpeptin did not interrupt GCA-induced ferroptosis in HT-22 cells but blocked the apoptosis. Animal experiments further demonstrated that Ten prevented GCA-induced memory impairment in mice and increased synaptic protein expression while reducing m-calpain expression. Ten prevents AD-like phenotypes through multiple signaling by inhibiting oxidative stress and ferroptosis, maintaining the stability of calpain system, and suppressing neuronal apoptosis.

Curculigoside rescues hippocampal synaptic deficits elicited by PTSD through activating cAMP-PKA signaling.

Chronic traumatic stress results in various psychiatric disorders, especially posttraumatic stress disorder (PTSD). Previous study demonstrated that curculigoside (CUR) a component of Rhizoma Curculiginis prevented fear extinction and stress-induced depression-like behaviors. However, its effects on PTSD and the mechanisms are still not completely clear. In this study, we observed typical PTSD-like phenotypes, synaptic deficit, and reduction of BDNF/TrkB signaling pathway in mice receiving modified single prolonged stress and electrical stimulation (SPS&S). By contrast, systemic administration of CUR blocked PTSD-like phenotypes and synaptic deficits, including reduction of BDNF/TrkB signaling pathway, GluA1 and Arc expression. Importantly, CUR reversed the impairment of PKA signaling pathway elicited by PTSD. We further confirmed that the effects of CUR on synaptic function were through PKA signaling pathway, as H-89, an inhibitor of PKA blocked the effect of CUR on behavioral changes and BDNF/TrkB signaling pathway. Thereafter, we verified that CUR on synaptic function were through PKA pathway using direct intracerebral injection of CUR and H-89. Direct intracerebral injection of CUR activated PKA/CREB/BDNF/TrkB, which was blocked by H-89. Additionally, the docking results showed high binding energies of CUR with A2AR, AC, PRKACA, and PRKAR1A, which might indicate that CUR functions through regulating PKA signaling pathway. In conclusion, CUR prevented the behavioral changes and hippocampal synaptic deficits elicited by PTSD through activating cAMP-PKA signaling.

Use of Steaming Process to Improve Biochemical Activity of Polygonatum sibiricum Polysaccharides against D-Galactose-Induced Memory Impairment in Mice.

Polysaccharide from Polygonatum sibiricum (PSP) possesses antioxidant, antiaging, and neuroprotective activities. However, whether and how the steaming process influences the biological activities of PSP, especially against aging-related memory impairment, is not yet known. In this study, Polygonatum sibiricum rhizome was subjected to a "nine steaming and nine drying" process, then PSPs with different steaming times were abstracted. Thereafter, the physicochemical properties were qualified; the antioxidant activities of PSPs were evaluated in a D-gal-induced HT-22 cell model, and the effects of PSPs (PSP0, PSP5 and PSP9) on memory was evaluated using D-gal-injured mice. Our results showed that while the steamed PSPs had a low pH value and a large negative charge, they shared similar main chains and substituents. Cellular experiments showed that the antioxidant activity of steamed PSPs increased. PSP0, PSP5, and PSP9 could significantly ameliorate the memory impairment of D-gal-injured mice, with PSP5 showing the optimal effect. Meanwhile, PSP5 demonstrated the best effect in terms of preventing cell death and synaptic injury in D-gal-injured mice. Additionally, the steamed PSPs increased anti-oxidative stress-related protein expression and decreased inflammation-related protein expression in D-gal-injured mice. Collectively, the steaming process improves the effects of PSPs against D-gal-induced memory impairment in mice, likely by increasing the antioxidant activity of PSPs.

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.

Polysaccharides from Polygonatum cyrtonema Hua Reduce Depression-Like Behavior in Mice by Inhibiting Oxidative Stress-Calpain-1-NLRP3 Signaling Axis.

Polysaccharides from Polygonatum cyrtonema Hua (PSP) exert antioxidant, anti-inflammatory, and antidepressant effects. Production of reactive oxygen species (ROS) and activation of the calpain system and the NOD-like receptor protein 3 (NLRP3) inflammasome are closely related to the pathogenesis of depression. However, the relationships among those pathways and the protective effects of PSP have not been characterized. In this study, lipopolysaccharide (LPS) and chronic unpredictable mild stress- (CUMS-) induced depression models were used to evaluate the protective mechanisms of PSP against depression. ROS levels were measured in HT-22 cells using flow cytometry. Brain tissues were collected to determine the levels of oxidation-related indicators and inflammatory cytokines. The protein levels of calpain-1, calpain-2, calpastatin, phosphatase and Tensin Homolog deleted on Chromosome 10 (PTEN), suprachiasmatic nucleus circadian oscillatory protein (SCOP), nuclear factor-erythroid factor 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), NLRP3, apoptosis-associated speck-like protein (ASC), caspase-1, cleaved-caspase-1, ionized calcium binding adapter molecule 1 (Iba1), phosphorylation of extracellular signal-regulated kinase (p-ERK), nuclear factor-kappa B (NF-κB), interleukin-1ß (IL-1ß), and glial fibrillary acidic protein (GFAP) were measured using western blotting or immunofluorescence. In cellular experiments, we showed that PSP attenuated LPS-induced production of ROS in HT-22 cells. In animal experiments, we found that LPS increased the expression of calpain-1, NLRP3, ASC, caspase-1, cleaved-caspase-1, Iba1, p-ERK, NF-κB, and GFAP and reduced the expression of calpastatin, PTEN, SCOP, and Nrf2. Administration of PSP reversed these changes. N-Acetyl-L-cysteine (NAC) administration also inhibited oxidative stress and activation of the calpain system and the NLRP3 inflammasome. Furthermore, PSP, calpeptin, MCC950 (a selective NLRP3 inflammasome inhibitor), and NAC reduced LPS-induced proinflammatory cytokine release. We also showed that PSP prevented CUMS-induced changes in the calpain system and the Nrf2 and NLRP3 signaling pathways and reduced depression-like behavior. These results indicate that PSP exerts antidepressant effects through regulation of the oxidative stress-calpain-1-NLRP3 signaling axis.

Utility of 7,8-dihydroxyflavone in preventing astrocytic and synaptic deficits in the hippocampus elicited by PTSD.

Astrocytic functions and brain-derived neurotrophic factor (BDNF)-tyrosine kinase receptor B (TrkB) signaling pathways are impaired in stress-related neuropsychiatric diseases. Previous studies have reported neuroprotective effects of 7,8-dihydroxyflavone (7,8-DHF), a TrkB activator. Here, we investigated the molecular mechanisms underlying pathogenesis of post-traumatic stress disorder (PTSD) using a modified single-prolonged stress (SPS&S) model and the potential beneficial effects of 7,8-DHF. SPS&S reduced the hippocampal expression of glial fibrillary acidic protein (GFAP), a marker of astrocytes, and induced morphological changes in astrocytes. From the perspective of synaptic function, the SPS&S model displayed reduced expression of BDNF, p-TrkB, postsynaptic density protein 95 (PSD95), AMPA receptor subunit GluR1 (GluA1), NMDA receptor subunit N2A/N2B ratio, calpain-1, phosphorylated protein kinase B (Akt) and phosphorylated mammalian target of rapamycin (mTOR) and conversely, higher phosphatase and tension homolog (PTEN) expression in the hippocampus. Acute or continuous intraperitoneal administration of 7,8-DHF (5 mg/kg) after SPS&S procedures prevented SPS&S-induced fear memory generalization and anxiety-like behaviors as well as abnormalities of hippocampal oscillations. Most importantly, 7,8-DHF attenuated SPS&S-induced abnormal BDNF-TrkB signaling and calpain-1-dependent cascade of synaptic deficits. Furthermore, treatment with a TrkB inhibitor completely blocked while an mTOR inhibitor partially blocked the effects of 7,8-DHF on behavioral changes of SPS&S model mice. Our collective findings suggest that 7,8-DHF effectively alleviates PTSD-like symptoms, including fear generalization and anxiety-like behavior, potentially by preventing astrocytic and synaptic deficits in the hippocampus through targeting of TrkB.

Depichering the Effects of Astragaloside IV on AD-Like Phenotypes: A Systematic and Experimental Investigation.

Astragaloside IV (AS-IV) is an active component in Astragalus membranaceus with the potential to treat neurodegenerative diseases, especially Alzheimer's diseases (ADs). However, its mechanisms are still not known. Herein, we aimed to explore the systematic pharmacological mechanism of AS-IV for treating AD. Drug prediction, network pharmacology, and functional bioinformatics analyses were conducted. Molecular docking was applied to validate reliability of the interactions and binding affinities between AS-IV and related targets. Finally, experimental verification was carried out in AßO infusion produced AD-like phenotypes to investigate the molecular mechanisms. We found that AS-IV works through a multitarget synergistic mechanism, including inflammation, nervous system, cell proliferation, apoptosis, pyroptosis, calcium ion, and steroid. AS-IV highly interacted with PPARγ, caspase-1, GSK3Β, PSEN1, and TRPV1 after docking simulations. Meanwhile, PPARγ interacts with caspase-1, GSK3Β, PSEN1, and TRPV1. In vivo experiments showed that AßO infusion produced AD-like phenotypes in mice, including impairment of fear memory, neuronal loss, tau hyperphosphorylation, neuroinflammation, and synaptic deficits in the hippocampus. Especially, the expression of PPARγ, as well as BDNF, was also reduced in the hippocampus of AD-like mice. Conversely, AS-IV improved AßO infusion-induced memory impairment, inhibited neuronal loss and the phosphorylation of tau, and prevented the synaptic deficits. AS-IV prevented AßO infusion-induced reduction of PPARγ and BDNF. Moreover, the inhibition of PPARγ attenuated the effects of AS-IV on BDNF, neuroflammation, and pyroptosis in AD-like mice. Taken together, AS-IV could prevent AD-like phenotypes and reduce tau hyperphosphorylation, synaptic deficits, neuroinflammation, and pyroptosis, possibly via regulating PPARγ.

Astrocytic Kir4.1 regulates NMDAR/calpain signaling axis in lipopolysaccharide-induced depression-like behaviors in mice.

The activation of Nod-like receptor protein 3 (NLRP3) inflammasome propagates pro-inflammatory signaling cascades linking to depression-like behaviors. However, the signaling pathway contributing to NLRP3 inflammasome activation and depression-like behaviors is still not clear. In this study, we evidenced that lipopolysaccharide (LPS) injection (i.p.) triggered depression-like behaviors, promoted the expression of Kir4.1, p-GluN2B and calpain-1, and activated NLRP3 inflammasome. The blockage of N-methyl-d-aspartate receptors (NMDAR) by memantine reduced LPS-induced depression-like behaviors, NLRP3 inflammasome and astrocyte activation, and calpain-1 expression. Additionally, memantine also inhibited LPS-induced reduction of postsynaptic density protein 95 (PSD-95) and Arc expression. Specific reduction of Kir4.1 in astrocytes attenuated LPS-induced expression of NLRP3 and calpain-1, and phosphorylation of GluN2B. Interestingly, LPS-induced expression of calpain-1 largely co-localized with GFAP, indicating the specific function of calpain-1 in astrocytes. Together, these data indicate that astrocytic Kir4.1 could regulate NMDAR/calpain-1 signaling axis, contributing to depression-like behaviors, likely through regulating NLRP3 inflammasome activation.

p53-mediated ferroptosis is required for 1-methyl-4-phenylpyridinium-induced senescence of PC12 cells.

Parkinson's disease (PD) is characterized by the loss of dopaminergic neurons in the substantia nigra and striatum. Aging is the most important risk factor of PD. Ferroptosis is an iron-dependent form of cell death associated with PD. However, it is not clear whether ferroptosis accelerates PD by promoting cellular senescence. This study investigated the mechanism of 1-methyl-4-phenylpyridinium (MPP+) -induced PC12 cells injury. We found that MPP+ induced cell senescence with increased ß-galactosidase activity and the expression of p53, p21 and p16 activation in cells. In addition, MPP+ treatment showed smaller mitochondria and increased membrane density, downregulation of ferritin heavy chain 1 expression and upregulation of acyl-CoA synthetase long chain family member 4 expression, and enhanced levels of oxidative stress, which were important characteristics of ferroptosis. Ferrostatin-1 (Fer-1), a ferroptosis inhibitor, was tested to eliminate MPP+-induced cell senescence. Fer-1 downregulated the expression of p53 and upregulated the expression of solute carrier family 7 member 11 (SLC7A11) and glutathione peroxidase-4 (GPX4) in MPP+-induced ferroptosis. Inhibition of p53 eliminated cell senescence by upregulation the expression of of SLC7A11 and GPX4. Thus, these results suggest that MPP+ induces senescence in PC12 cells via the p53/ SLC7A11/ GPX4 signaling pathway in the ferroptosis regulation mechanism.

Brusatol reverses lipopolysaccharide-induced epithelial-mesenchymal transformation and induces apoptosis through PI3K/Akt/NF-кB pathway in human gastric cancer SGC-7901 cells.

Brusatol is a butyrolactone compound isolated from traditional Chinese medicine Brucea javanica. It has been reported to possess strong cytotoxicity against various cancer cells, thus showing its potential as an anticancer drug. Besides, lipopolysaccharide (LPS) plays a central role in the tumor microenvironment, while epithelial-mesenchymal transformation (EMT), a biological process by which epithelial cells are transformed into mesenchymal phenotypic cells through specific procedures, participates in chronic inflammation and tumor metastasis. This study aimed to investigate the inhibition of LPS-induced tumor cell invasion and metastasis and the molecular mechanism of apoptosis induced by brusatol in human gastric cancer SGC-7901 cells. Cell viability, cell migration and invasion ability, inflammatory factor release, and protein expression were detected using methyl thiazolyl tetrazolium assays, transwell assays, ELISA kit, and Western blot analysis, respectively. The change of EMT marker protein vimentin was assessed using immunofluorescence, while the apoptosis rate was measured using flow cytometry. In summary, brusatol inhibited LPS-induced EMT via the deactivation of the PI3K/Akt/NF-кB signaling pathway. This provides a useful new theoretical basis for the treatment of gastric cancer in the future.

Astragaloside IV prevents Aß1-42 oligomers-induced memory impairment and hippocampal cell apoptosis by promoting PPARγ/BDNF signaling pathway.

Astragaloside IV (AS-IV), a natural product derived from Radix Astragali (Astragalus membranaceus), is beneficial for the treatment of Alzheimer's disease (AD), but the mechanisms underlying this benefit are not completely understood. Peroxisome proliferator-activated receptor gamma (PPARγ) and brain-derived neurotrophic factor (BDNF) are potential therapeutic targets for AD. In this study, we found that amyloid ß protein fragment 1-42 oligomers (AßO) suppressed BDNF and PPARγ expression, and inhibited tyrosine receptor kinase B (TrkB) phosphorylation in cultured hippocampal neurons; these changes were ameliorated by treatment with AS-IV. Inhibition of PPARγ by genetic and pharmacological methods also blocked the effect of AS-IV on BDNF expression in AßO-treated cells. Importantly, exogenous BDNF protected against neurotoxicity and apoptosis induced by AßO, whereas inhibition of PPARγ reversed protective effects of AS-IV against these outcomes. In vivo data further revealed that AS-IV improved AßO-induced memory impairment and reduced apoptosis of hippocampal neurons. Moreover, AS-IV suppressed the AßO-induced reduction in BDNF by promoting PPARγ activation in the hippocampus. Taken together, these results indicate that AS-IV prevents AßO-induced memory impairment and hippocampal neuronal apoptosis, probably by promoting the PPARγ/BDNF signaling pathway.

Gastrodin Inhibits H2O2-Induced Ferroptosis through Its Antioxidative Effect in Rat Glioma Cell Line C6.

Ferroptosis is a form of necrosis caused by iron-induced accumulation of lipid hydroperoxide, involving several molecular events, and has been implicated in Parkinson's disease. Gastrodin is a component of Gastrodia elata Blume with strong antioxidant activity. We examined whether gastrodin can prevent H2O2-induced cytotoxicity in rat glioma cell line C6. For this purpose, C6 cells were pretreated with gastrodin (1, 5, 25 µM) and then exposed to 100 µM H2O2. Results showed that pretreatment of C6 cells with gastrodin decreased H2O2-induced lactate dehydrogenase (LDH) release and cell death. Moreover, gastrodin decreased intracellular malondialdehyde (MDA) level, whereas increased glutathione peroxidase (GPX) activity and glutathione (GSH) level after H2O2 treatment. In addition, treatment of deferoxamine (DFO), ferrostatin-1, and liproxstatin-1 abolished ferroptosis induced by H2O2 or erastin pretreatment. Treatment with gastrodin attenuated H2O2-induced ferroptosis and decreased lipid reactive oxygen species (ROS) (C11-BODIPY) production in C6 cells. Moreover, gastrodin increased the protein expression of nuclear factor erythroid 2-related factor 2 (Nrf2), GPX4, ferroportin-1 (FPN1), and heme oxygenase-1 (HO-1) in C6 cells treated with H2O2. RSL3, a GPX4 inhibitor, inhibited GPX4 protein level in cells co-treated with gastrodin and 100 µM H2O2. These findings indicate that gastrodin can inhibit H2O2-induced ferroptosis through its antioxidative effect in C6 cells.

Gastrodin protects against glutamate-induced ferroptosis in HT-22 cells through Nrf2/HO-1 signaling pathway.

Gastrodin (GAS) is a component of Gastrodia elata Blume, with strong antioxidant activity in neurodegenerative diseases. Ferroptosis is similar to glutamate-induced cell death. This study was designed to explore the protective effects of GAS against glutamate-induced neurotoxicity in mice hippocampal neurons (HT-22) cells. HT-22 cells were cultured with glutamate in the presence or absence of GAS (1, 5, 25 µM). Results showed that GAS inhibited glutamate-induced ferroptosis via Nrf2/HO-1 signaling pathway. Pretreatment of HT-22 cells with GAS significantly decreased glutamate-induced cell death and release of LDH. Ferrostatin-1, liproxstatin-1, and DFO treatments canceled these effect. GAS decreased glutamate-treatment ROS production in HT-22 cells. The concentration of iron ion was analyzed using ICP-MS. Metal analysis showed that GAS pretreatment normalized iron ion concentration in HT-22 cells. We found that GAS increased the nuclear translocation of Nrf2, up-regulated the downstream HO-1 protein expression in HT-22 cells following treatment with glutamate. Nrf2 knockdown greatly decreased glutamate-induced ferroptosis through HO-1. In conclusion, these results show that GAS protects HT-22 cells from the ferroptosis induced by glutamate through a new mechanism of Nrf2/HO-1 signaling pathway.

Curculigoside facilitates fear extinction and prevents depression-like behaviors in a mouse learned helplessness model through increasing hippocampal BDNF.

Curculigoside (CUR) is the main active component of traditional Chinese medicine Curculigoorchioides Gaertn (Xianmao in Chinese), which exhibits a variety of pharmacological activities. In this study we investigated the effects of CUR on fear extinction and related depression-like behaviors in mice. In fear conditioning task, we found that administration of CUR (1.6, 8, 40 mg·kg-1·d-1, ip, for 7 days) did not affect memory consolidation, but CUR at higher doses (8, 40 mg·kg-1·d-1) significantly facilitated fear extinction, especially on D3 and D4. Moreover, CUR administration significantly ameliorated the fear conditioning-induced depression-like behaviors, likely through promoting fear extinction. We showed that CUR increased the expression of brain-derived neurotrophic factor (BDNF) and phosphorylation of tropomyosin receptor kinase B (TrkB) in the hippocampus, and activated protein kinase B (Akt)-mammalian target of the rapamycin (mTOR) signaling pathway. Administration of the selective TrkB agonist 7,8-dihydroxyflavone (7,8-DHF, 5 mg·kg-1·d-1, ip) also facilitated fear extinction, ameliorated depression-like behaviors. We established a mouse learned helplessness (LH) model to evaluate the antidepressant activity of CUR. The spatial memory was assessed in Morris water maze. We showed that LH-induced depression-like behaviors, including prolonged immobility times in forced swim and tail suspension tests as well as spatial memory impairments; LH also downregulated BDNF expression and the Akt-mTOR signaling pathway in the hippocampus. Administration of CUR (1.6, 8, 40 mg·kg-1·d-1, ip, for 14 days) or 7,8-DHF (5 mg·kg-1·d-1, ip, for 3 days) prevented LH-induced depression-like behaviors and promoted BDNF expression and the Akt-mTOR signaling pathway. In conclusion, CUR can accelerate the fear memory extinction and ameliorate depression-like behaviors in mice via promoting BDNF expression and activating the Akt-mTOR signaling pathway in the hippocampus.

MPTP-induced changes in hippocampal synaptic plasticity and memory are prevented by memantine through the BDNF-TrkB pathway.

BACKGROUND AND PURPOSE: Mild cognitive deficit in early Parkinson's disease (PD) has been widely studied. Here we have examined the effects of memantine in preventing memory deficit in experimental PD models and elucidated some of the underlying mechanisms. EXPERIMENTAL APPROACHES: I.p. injection of 1-methyl-4- phenyl-1,2,3,6-tetrahydro pyridine (MPTP) in C57BL/6 mice was used to produce models of PD. We used behavioural tasks to test memory. In vitro, we used slices of hippocampus, with electrophysiological, Western blotting, real time PCR, elisa and immunochemical techniques. KEY RESULTS: Following MPTP injection, long-term memory was impaired and these changes were prevented by pre-treatment with memantine. In hippocampal slices from MPTP treated mice, long-term potentiation (LTP) -induced by θ burst stimulation (10 bursts, 4 pulses) was decreased, while long-term depression (LTD) induced by low-frequency stimulation (1 Hz, 900 pulses) was enhanced, compared with control values. A single dose of memantine (i.p., 10 mg·kg(-1) ) reversed the decreased LTP and the increased LTD in this PD model. Activity-dependent changes in tyrosine kinase receptor B (TrkB), ERK and brain-derived neurotrophic factor (BDNF) expression were decreased in slices from mice after MPTP treatment. These effects were reversed by pretreatment with memantine. Incubation of slices in vitro with 1-methyl-4-phenylpyridinium (MPP(+) ) decreased depolarization-induced expression of BDNF. This effect was prevented by pretreatment of slices with memantine or with calpain inhibitor III, suggesting the involvement of an overactivated calcium signalling pathway. CONCLUSIONS AND IMPLICATIONS: Memantine should be useful in preventing loss of memory and hippocampal synaptic plasticity in PD models.