[Content analysis and quality evaluation of main active components and mineral elements of Cynomorium songaricum in different habitats].

To clarify the content characteristics of the main active components and mineral elements of Cynomorium songaricum under different habitat conditions, and further explore the relationship between the quality of C. songaricum and habitats, this study took C. songaricum from 25 different habitats in China as the research object, and measured the contents of 8 main active components and 12 mineral elements separately. Diversity analysis, correlation analysis, principal component analysis and cluster analysis were carried out. The results showed that the genetic diversity of total flavonoids, ursolic acid, ether extract, potassium(K), phosphorus(P) and zinc(Zn) in C. songaricum was high. The coefficient of variation of crude polysaccharide, ether extract, gallic acid, protocatechuic aldehyde, catechin, epicatechin, calcium(Ca), sodium(Na), magnesium(Mg), sulfur(S), iron(Fe), manganese(Mn), selenium(Se) and nickel(Ni) were all over 36%, indicating that the quality of C. songaricum was significantly affected by habitats. There were strong synergistic and weak antagonistic effects among the contents of the 8 active components, and complex antagonistic and synergistic effects among the contents of the 12 mineral elements. Principal component analysis revealed that crude polysaccharide, ursolic acid, catechin, epicatechin and total flavonoids could be used as the characteristic components to evaluate the quality of C. songaricum, and Na, copper(Cu), Mn and Ni were the characteristic elements to evaluate the quality of C. songaricum. In cluster ana-lysis, the second group with the main active components as cluster center had better quality in terms of the content of active substances, and the second group with the mineral elements as cluster center had higher utilization potential in the exploitation of mineral elements. This study could provide a basis for resource evaluation and breeding of excellent varieties of C. songaricum in different habitats, and provide a reference for cultivation and identification of C. songaricum.

Astragaloside IV promotes cerebral tissue restoration through activating AMPK- mediated microglia polarization in ischemic stroke rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Astragaloside IV (AS), a key active ingredient obtained from Chinese herb Astragalus mongholicus Bunge, exerts potent neuroprotective and anti-inflammatory effects for treating neurodegenerative diseases. However, mechanisms of AS on improvement of ischemic brain tissue repair remain unclear. AIM OF THE STUDY: This research aims at using magnetic resonance imaging (MRI) to noninvasively determine whether AS facilitates brain tissue repair, and investigating whether AS exerts brain remodeling through adenosine monophosphate-activated protein kinase (AMPK) metabolic signaling regulating key glycolytic enzymes and energy transporters, thereby impacting microglia polarization. MATERIALS AND METHODS: Ischemic stroke model in male Sprague-Dawley rats were induced through permanent occlusion of the middle cerebral artery (MCAO). Infarct volume, the alterations of brain microstructure and nerve fibers reorganization were examined by multi-parametric MRI. The pathological damages of myelinated axons and microglia polarization surrounding infarct tissue were detected using pathological techniques. Furthermore, M1/M2 microglia polarization associated protein, glycolytic rate-limiting enzymes, energy transporters and AMPK/mammalian target of rapamycin (mTOR)/hypoxia inducible factor-1α (HIF-1α) signal were examined both in ischemic stroke rats and BV2 microglia treated with lipopolysaccharide (LPS) + interferon-γ (IFN-γ) by western blotting. RESULTS: MRI revealed that AS obviously decreased infarct volume, relieved brain microstructure damage and improved nerve fibers reorganization in ischemic stroke rats. Histological tests supported MRI findings. Notably, AS promoted microglia M2 and reduced M1 polarization, induced the AMPK activation accompanied with decreased levels of phosphorylated mTOR and HIF-1α. Moreover, AS suppressed the expression of glycolytic rate-limiting enzymes and energy transporters in ischemic stroke rats and BV2 microglia. In contrast, these beneficial effects were greatly blocked by AMPK inhibitor compound C. CONCLUSION: Overall, these results collectively suggested that AS facilitated tissue remodeling that may be partially through modulating polarization of microglia in AMPK- dependent metabolic pathways after ischemic stroke.

Buyang Huanwu Decoction promotes neurovascular remodeling by modulating astrocyte and microglia polarization in ischemic stroke rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Buyang Huanwu Decoction (BYHWD), one of the most commonly utilized traditional Chinese medicine prescription for treatment of cerebral ischemic stroke. However, the understanding of BYHWD on neurovascular repair following cerebral ischemia is so far limited. AIM OF THE STUDY: This research investigated the influence of BYHWD on neurovascular remodeling by magnetic resonance imaging (MRI) technology and revealed the potential neurovascular repair mechanism underlying post-treatment with BYHWD after ischemic stroke. MATERIALS AND METHODS: Male Sprague-Dawley rats were utilized as an ischemic stroke model by permanent occlusion of the middle cerebral artery (MCAO). BYHWD was intragastrically administrated once daily for 30 days straight. Multimodal MRI was performed to detect brain tissue injuries, axonal microstructural damages, cerebral blood flow and intracranial vessels on the 30th day after BYHWD treatment. Proangiogenic factors, axonal/synaptic plasticity-related factors, energy transporters and adenosine monophosphate-activated protein kinase (AMPK) signal pathway were evaluated using western blot. Double immunofluorescent staining and western blot were applied to evaluate astrocytes and microglia polarization. RESULTS: Administration of BYHWD significantly alleviated infarct volume and brain tissue injuries and ameliorated microstructural damages, accompanied with improved axonal/synaptic plasticity-related factors, axonal growth guidance factors and decreased axonal growth inhibitors. Meanwhile, BYHWD remarkably improved cerebral blood flow, cerebral vascular signal and promoted the expression of proangiogenic factors. Particularly, treatment with BYHWD obviously suppressed astrocytes A1 and microglia M1 polarization accompanied with promoted astrocyte A2 and microglia M2 polarization. Furthermore, BYHWD effectively improved energy transporters. Especially, BYHWD markedly increased expression of phosphorylated AMPK, cyclic AMP-response element binding protein (CREB) and brain-derived neurotrophic factor (BDNF) accompanied by inactivation of the NF-κB. CONCLUSION: Taken together, these findings identified that the beneficial roles of BYHWD on neurovascular remodeling were related to AMPK pathways -mediated energy transporters and NFκB/CREB pathways.

Proposing a novel molecular subtyping scheme for predicting distant recurrence-free survival in breast cancer post-neoadjuvant chemotherapy with close correlation to metabolism and senescence.

Background: High relapse rates remain a clinical challenge in the management of breast cancer (BC), with distant recurrence being a major driver of patient deterioration. To optimize the surveillance regimen for distant recurrence after neoadjuvant chemotherapy (NAC), we conducted a comprehensive analysis using bioinformatics and machine learning approaches. Materials and methods: Microarray data were retrieved from the GEO database, and differential expression analysis was performed with the R package 'Limma'. We used the Metascape tool for enrichment analyses, and 'WGCNA' was utilized to establish co-expression networks, selecting the soft threshold power with the 'pickSoftThreshold' algorithm. We integrated ten machine learning algorithms and 101 algorithm combinations to identify key genes associated with distant recurrence in BC. Unsupervised clustering was performed with the R package 'ConsensusCluster Plus'. To further screen the key gene signature of residual cancer burden (RCB), multiple knockdown studies were analyzed with the Genetic Perturbation Similarity Analysis (GPSA) database. Single-cell RNA sequencing (scRNA-seq) analysis was conducted through the Tumour Immune Single-cell Hub (TISCH) database, and the XSum algorithm was used to screen candidate small molecule drugs based on the Connectivity Map (CMAP) database. Molecular docking processes were conducted using Schrodinger software. GMT files containing gene sets associated with metabolism and senescence were obtained from GSEA MutSigDB database. The GSVA score for each gene set across diverse samples was computed using the ssGSEA function implemented in the GSVA package. Results: Our analysis, which combined Limma, WGCNA, and machine learning approaches, identified 16 RCB-relevant gene signatures influencing distant recurrence-free survival (DRFS) in BC patients following NAC. We then screened GATA3 as the key gene signature of high RCB index using GPSA analysis. A novel molecular subtyping scheme was developed to divide patients into two clusters (C1 and C2) with different distant recurrence risks. This molecular subtyping scheme was found to be closely associated with tumor metabolism and cellular senescence. Patients in cluster C2 had a poorer DRFS than those in cluster C1 (HR: 4.04; 95% CI: 2.60-6.29; log-rank test p < 0.0001). High GATA3 expression, high levels of resting mast cell infiltration, and a high proportion of estrogen receptor (ER)-positive patients contributed to better DRFS in cluster C1. We established a nomogram based on the N stage, RCB class, and molecular subtyping. The ROC curve for 5-year DRFS showed excellent predictive value (AUC=0.91, 95% CI: 0.95-0.86), with a C-index of 0.85 (95% CI: 0.81-0.90). Entinostat was identified as a potential small molecule compound to reverse high RCB after NAC. We also provided a comprehensive review of the EDCs exposures that potentially impact the effectiveness of NAC among BC patients. Conclusion: This study established a molecular classification scheme associated with tumor metabolism and cancer cell senescence to predict RCB and DRFS in BC patients after NAC. Furthermore, GATA3 was identified and validated as a key gene associated with BC recurrence.

Tetramethylpyrazine promotes stroke recovery by inducing the restoration of neurovascular unit and transformation of A1/A2 reactive astrocytes.

2,3,5,6-Tetramethylpyrazine (TMP) as an active ingredient extracted from a traditional Chinese herbal medicine Ligusticum chuanxiong Hort. has been proved to penetrate blood-brain barrier (BBB) and show neuroprotective effects on cerebral ischemia. However, whether TMP could regulate astrocytic reactivity to facilitate neurovascular restoration in the subacute ischemic stroke needs to be urgently verified. In this research, permanent occlusion of the middle cerebral artery (MCAO) model was conducted and TMP (10, 20, 40 mg/kg) was intraperitoneally administrated to rats once daily for 2 weeks. Neurological function was evaluated by motor deficit score (MDS). Magnetic resonance imaging (MRI) was implemented to analyze tissue injury and cerebral blood flow (CBF). Magnetic resonance angiography (MRA) was applied to exhibit vascular signals. Transmission electron microscopy (TEM) was performed to detect the neurovascular unit (NVU) ultrastructure. Haematoxylin and eosin (HE) staining was utilized to evaluate cerebral histopathological lesions. The neurogenesis, angiogenesis, A1/A2 reactivity, aquaporin 4 (AQP4) and connexin 43 (Cx43) of astrocytes were observed with immunofluorescent staining. Then FGF2/PI3K/AKT signals were measured by western blot. Findings revealed TMP ameliorated neurological functional recovery, preserved NVU integrity, and enhanced endogenous neurogenesis and angiogenesis of rats with subacute ischemia. Shifting A1 to A2 reactivity, suppressing excessive AQP4 and Cx43 expression of astrocytes, and activating FGF2/PI3K/AKT pathway might be potential mechanisms of promoting neurovascular restoration with TMP after ischemic stroke.

Magnetic Resonance Imaging Investigation of Neuroplasticity After Ischemic Stroke in Tetramethylpyrazine-Treated Rats.

Ischemic stroke elicits white matter injury typically signed by axonal disintegration and demyelination; thus, the development of white matter reorganization is needed. 2,3,5,6-Tetramethylpyrazine (TMP) is widely used to treat ischemic stroke. This study was aimed to investigate whether TMP could protect the white matter and promote axonal repair after cerebral ischemia. Male Sprague-Dawley rats were subjected to permanent middle cerebral artery occlusion (MCAO) and treated with TMP (10, 20, 40 mg/kg) intraperitoneally for 14 days. The motor function related to gait was evaluated by the gait analysis system. Multiparametric magnetic resonance imaging (MRI) was conducted to noninvasively identify gray-white matter structural integrity, axonal reorganization, and cerebral blood flow (CBF), followed by histological analysis. The expressions of axonal growth-associated protein 43 (GAP-43), synaptophysin (SYN), axonal growth-inhibitory signals, and guidance factors were measured by Western blot. Our results showed TMP reduced infarct volume, relieved gray-white matter damage, promoted axonal remodeling, and restored CBF along the peri-infarct cortex, external capsule, and internal capsule. These MRI findings were confirmed by histopathological data. Moreover, motor function, especially gait impairment, was improved by TMP treatment. Notably, TMP upregulated GAP-43 and SYN and enhanced axonal guidance cues such as Netrin-1/DCC and Slit-2/Robo-1 but downregulated intrinsic growth-inhibitory signals NogoA/NgR/RhoA/ROCK-2. Taken together, our data indicated that TMP facilitated poststroke axonal remodeling and motor functional recovery. Moreover, our findings suggested that TMP restored local CBF, augmented guidance cues, and restrained intrinsic growth-inhibitory signals, all of which might improve the intracerebral microenvironment of ischemic areas and then benefit white matter remodeling.

Trillium tschonoskii rhizomes' saponins induces oligodendrogenesis and axonal reorganization for ischemic stroke recovery in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Trillium tschonoskii Maxim. is one of traditional Chinese medical herbs that has been utilized to treat brain damages and cephalalgia. The neuroprotective effect of total saponins from Trillium tschonoskii rhizome (TSTT) has been demonstrated efficacy in rats following ischemia. However, the axonal remodeling effect of TSTT and the detailed mechanisms after ischemic stroke have not been investigated. AIM OF THE STUDY: We aimed to estimate therapeutic role of TSTT in axonal remodeling using magnetic resonance imaging (MRI) technique, and explored possible mechanisms underlying this process followed by histological assays in ischemic rats. METHODS: Male Sprague-Dawley (SD) rats underwent permanently focal cerebral ischemia induced by occluding right permanent middle cerebral artery. TSTT was intragastrically administrated 6 h after surgery and once daily for consecutive 15 days. Neurological function was assessed by the motor deficit score and beam walking test. T2 relaxation mapping and diffusion tensor imaging (DTI) were applied for detecting cerebral tissues damages and microstructural integrity of axons. Luxol fast blue (LFB) and transmission electron microscope (TEM) were performed to evaluate histopathology in myelinated axons. Double immunofluorescent staining was conducted to assess oligodendrogenesis. Furthermore, the protein expressions regarding to axonal remodeling related signaling pathways were detected by Western blot assays. RESULTS: TSTT treatment (65, 33 mg/kg) markedly improved motor function after ischemic stroke. T2 mapping MRI demonstrated that TSTT decreased lesion volumes, and DTI further confirmed that TSTT preserved axonal microstructure of the sensorimotor cortex and internal capsule. Meanwhile, diffusion tensor tractography (DTT) showed that TSTT elevated correspondent density and length of fiber in the internal capsule. These MRI measurements were confirmed by histological examinations. Notably, TSTT significantly increased Ki67/NG2, Ki67/CNPase double-labeled cells along the boundary zone of ischemic cortex and striatum. Meanwhile, TSTT treatment up-regulated the phosphorylation level of Ser 9 in GSK-3ß, and down-regulated phosphorylated ß-catenin and CRMP-2 expression. CONCLUSION: Taken together, our findings indicated that TSTT (65, 33 mg/kg) enhanced post-stroke functional recovery, amplified endogenous oligodendrogenesis and promoted axonal regeneration. The beneficial role of TSTT might be correlated with GSK-3/ß-catenin/CRMP-2 modulating axonal reorganization after ischemic stroke.

Effect of Neurorepair for Motor Functional Recovery Enhanced by Total Saponins From Trillium tschonoskii Maxim. Treatment in a Rat Model of Focal Ischemia.

Trillium tschonoskii Maxim. (TTM), is a perennial herb from Liliaceae, that has been widely used as a traditional Chinese medicine treating cephalgia and traumatic hemorrhage. The present work was designed to investigate whether the total saponins from Trillium tschonoskii Maxim. (TSTT) would promote brain remodeling and improve gait impairment in the chronic phase of ischemic stroke. A focal ischemic model of male Sprague-Dawley (SD) rats was established by permanent middle cerebral artery occlusion (MCAO). Six hours later, rats were intragastrically treated with TSTT (120, 60, and 30 mg/kg) and once daily up to day 30. The gait changes were assessed by the CatWalk-automated gait analysis system. The brain tissues injuries, cerebral perfusion and changes of axonal microstructures were detected by multimodal magnetic resonance imaging (MRI), followed by histological examinations. The axonal regeneration related signaling pathways including phosphatidylinositol 3-kinases (PI3K)/protein kinase B (AKT)/glycogen synthase kinase-3 (GSK-3)/collapsin response mediator protein-2 (CRMP-2) were measured by western blotting. TSTT treatment significantly improved gait impairment of rats. MRI analysis revealed that TSTT alleviated tissues injuries, significantly improved cerebral blood flow (CBF), enhanced microstructural integrity of axon and myelin sheath in the ipsilesional sensorimotor cortex and internal capsule. In parallel to MRI findings, TSTT preserved myelinated axons and promoted oligodendrogenesis. Specifically, TSTT interventions markedly up-regulated expression of phosphorylated GSK-3, accompanied by increased expression of phosphorylated PI3K, AKT, but reduced phosphorylated CRMP-2 expression. Taken together, our results suggested that TSTT facilitated brain remodeling. This correlated with improving CBF, encouraging reorganization of axonal microstructure, promoting oligodendrogenesis and activating PI3K/AKT/GSK-3/CRMP-2 signaling, thereby improving poststroke gait impairments.

The performance of an anaerobic ammonium oxidation upflow anaerobic sludge blanket reactor during natural periodic temperature variations.

An anaerobic ammonium oxidation-upflow anaerobic sludge blanket (anammox-UASB) reactor was operated without temperature control during the four seasons and was therefore subjected to natural periodic temperature variations between 9 and 28 ℃. The anammox reactor had a high nitrogen removal ability at intermediate and low temperatures. The total nitrogen (TN) concentration of the influent increased from 200 to 1200 mg/L, the nitrogen removal efficiency was maintained at 90%, and the nitrogen removal rate (NRR) increased to 9.15 ±â€¯0.35 kg N/m3/d. The enrichment of anammox bacteria in the UASB granular sludge reached 53.8%, and the dominant bacteria changed from Candidatus Brocadia to Candidatus Kuenenia after several seasons of cultivation. Dynamics analysis revealed that the maximum reaction rate of the anammox-UASB sludge was 62.5 kg N/m3/d, reflecting the high potential nitrogen removal ability of the reactor.

Estrogenic properties of six compounds derived from Eucommia ulmoides Oliv. and their differing biological activity through estrogen receptors α and ß.

Eucommia ulmoides Oliv. (Du-zhong), a well known Chinese herbal medicine, has been increasingly used as a nutraceutical supplement. The aim of this study was to identify and characterise novel estrogenic compounds in E. ulmoides. We report that six compounds, namely pinoresinol 4'-O-ß-d-glucopyranoside, pinoresinol di-O-ß-d-glucopyranoside, aucubin, wogonin, baicalein, and α-O-ß-d-glucopyranosyl-4,2',4'-trihydroxydihydrochalcone, activated estrogen receptor (ER)-dependent transcription of both transfected and endogenous target genes. Strikingly, these compounds exhibited significant difference in ER subtype (α vs. ß) selectivity. Cell proliferation and ER subcellular localisation analyses also demonstrated that they mediated the estrogenic effects by the genomic action of ERα. This study suggests that E. ulmoides is a rich source of new selective estrogen receptor modulators and evaluations of its health benefit and safety as a food additive should take into account the diverse phytoestrogen activities of the individual components.