The Effects of Crataegus pinnatifida and Wolfiporia extensa Combination on Diet-Induced Obesity and Gut Microbiota.

Obesity is a multifactorial chronic metabolic disease with multiple complications. Crataegus pinnatifida (CP) and Wolfiporia extensa (WE) are traditional functional foods with improving metabolic health properties. This study demonstrated the effect of CP and WE combination on ameliorating obesity induced by a high-fat diet (HFD). Moreover, the CP-WE food pair ameliorated HFD-induced metabolic disorders, including glucose intolerance, insulin resistance, hyperlipidemia, and hepatic steatosis. 16S rRNA gene amplicon sequencing and analysis revealed that CP combined with WE reshaped the composition of gut microbiota in HFD-fed mice. Furthermore, correlation analysis revealed a substantial association between the obesity-related parameters and the shifts in predominant bacterial genera influenced by the food pair intervention. In conclusion, this study demonstrated that the CP-WE food pair ameliorated HFD-induced obesity and reshaped gut microbiota composition, providing a promising approach to combat obesity through specific food combinations.

Mechanism of Astragalus mongholicus Bunge ameliorating cerebral ischemia-reperfusion injury: Based on network pharmacology analysis and experimental verification.

ETHNOPHARMACOLOGICAL RELEVANCE: Astragalus mongholicus Bunge (AMB) is a herb with wide application in traditional Chinese medicine, exerting a wealth of pharmacological effects. AMB has been proven to have an evident therapeutic effect on ischemic cerebrovascular diseases, including cerebral ischemia-reperfusion injury (CIRI). However, the specific mechanism underlying AMB in CIRI remains unclear. AIM OF THE STUDY: This study aimed to investigate the potential role of AMB in CIRI through a comprehensive approach of network pharmacology and in vivo experimental research. METHODS: The intersection genes of drugs and diseases were obtained through analysis of the Traditional Chinese Medicine Systems Pharmacology (TCMSP) database and Gene Expression Omnibus (GEO) database. The protein-protein interaction (PPI) network was created through the string website. Meanwhile, the gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis was carried out using R studio, and thereafter the key genes were screened. Then, the molecular docking prediction was made between the main active ingredients and target genes, and hub genes with high binding energy were obtained. In addition, molecular dynamic (MD) simulation was used to validate the result of molecular docking. Based on the results of network pharmacology, we used animal experiments to verify the predicted hub genes. First, the rat middle cerebral artery occlusion and reperfusion (MACO/R) model was established and the effective dose of AMB in CIRI was determined by behavioral detection and 2,3,5-Triphenyltetrazolium chloride (TTC) staining. Then the target proteins corresponding to the hub genes were measured by Western blot. Moreover, the level of neuronal death was measured using hematoxylin and eosin (HE) and Nissl staining. RESULTS: Based on the analysis of the TCMSP database and GEO database, a total of 62 intersection target genes of diseases and drugs were obtained. The KEGG enrichment analysis showed that the therapeutic effect of AMB on CIRI might be realized through the advanced glycation endproduct-the receptor of advanced glycation endproduct (AGE-RAGE) signaling pathway in diabetic complications, nuclear factor kappa-B (NF-κB) signaling pathway and other pathways. Molecular docking results showed that the active ingredients of AMB had good binding potential with hub genes that included Prkcb, Ikbkb, Gsk3b, Fos and Rela. Animal experiments showed that AWE (60 g/kg) could alleviate CIRI by regulating the phosphorylation of PKCß, IKKß, GSK3ß, c-Fos and NF-κB p65 proteins. CONCLUSION: AMB exerts multi-target and multi-pathway effects against CIRI, and the underlying mechanism may be related to anti-apoptosis, anti-inflammation, anti-oxidative stress and inhibiting calcium overload.

Arenobufagin regulates the p62-Keap1-Nrf2 pathway to induce autophagy-dependent ferroptosis in HepG2 cells.

Hepatocellular carcinoma (HCC) is the most prevalent type of primary liver cancer, accounting for the overwhelming majority of malignant liver tumors. Therefore, how to effectively prevent and cure HCC has become a research hotspot. Many studies have shown that arenobufagin can induce apoptosis, ferroptosis, and autophagy of tumor cells. An increasing number of studies have shown that autophagy is closely linked to ferroptosis. In this study, HepG2 cells and BALB/c nude mice were used as research objects to explore the effect and preliminary mechanism of hepatoma cell autophagy and ferroptosis induced by arenobufagin. We found that arenobufagin can significantly inhibit tumor growth in vivo, and interestingly, we found that arenobufagin inhibited ferroptosis-related proteins Nrf2 and COX-2 in a dose-dependent manner and decreased the levels of reduced glutathione (GSH) and superoxide dismutase (T-SOD) in tissues, while increased the level of reduced malondialdehyde (MDA). In addition, we found that arenobufagin increased the levels of COX-2 and MDA in cells, decreased the levels of Nrf2, GSH, and T-SOD, increased the levels of tissue reactive oxygen species (ROS) and lipid ROS in a dose-dependent manner, and promoted ferroptosis in HepG2 cells. HepG2 cells were preprotected by autophagy inhibitor chloroquine (CQ) and ferroptosis inhibitor deferoxamine (DFO), and then treated with arenobufagin. It was found that CQ partially reversed the changes of COX-2 and Nrf2 expression and lipid peroxidation induced by arenobufagin-induced autophagy and HepG2 cells. Interestingly, CQ partially reversed the inhibition of arenobufagin on cytoplasmic junction protein (Keap1) and heme oxygenase-1 (HO-1) in p62-Keap1-Nrf2 pathway. At the same time, we found that the effect of arenobufagin on oxidative stress of HepG2 cells overexpressed by Nrf2 was significantly less than that of the control group. To sum up, arenobufagin promotes autophagy-dependent ferroptosis of HepG2 cells by inducing autophagy and regulating p62-Keap1-Nrf2 pathway. It is suggested that arenobufagin can be used as a potential intervention therapy.

Cinobufacini injection delays hepatocellular carcinoma progression by regulating lipid metabolism via SREBP1 signaling pathway and affecting macrophage polarization.

ETHNOPHARMACOLOGICAL RELEVANCE: Cinobufacini injection, an aqueous extract of the toad, is a commonly used anti-tumor animal herbal medicine in clinical practice. It has the effects of detoxifying, reducing swelling, and relieving pain. AIMS OF THE STUDY: To investigate the effects of Cinobufacini injection on hepatocellular carcinoma progression by regulating lipid metabolism and macrophage polarization in the tumor microenvironment and to identify the potential molecular mechanisms. MATERIALS AND METHODS: To establish the axillary transplantation tumor model of hepatocellular carcinoma Hepa1-6 in C57BL/6 mice, and to evaluate the inhibitory effect of Cinobufacini injection on hepatocellular carcinoma in vivo as well as drug delivery security. Combined metabolomics and transcriptomics analysis of the effect of Cinobufagin Injection on tumor microenvironment. An in vitro mouse co-culture model of peritoneal macrophages and Hepa1-6 cells was established to research the effects of Cinobufacini injection on macrophage polarization, hepatocellular carcinoma cell growth, migration, and changes in lipid metabolism. Cinobufacini injection inhibition of the AMPK/SREBP1/FASN signaling pathway regulating cholesterol metabolism and affecting macrophage polarization was examined using qRT-PCR, lentiviral transfection, immunofluorescence, and Western blot. RESULT: In vivo experiments demonstrated that Cinobufacini injection treatment significantly inhibited the growth of Hepa1-6 hepatomas, along with a reduction in cholesterol content and a decrease in the percentage of M2 macrophages in tumor tissue. In vitro, we found that Cinobufacini injection inhibits IL-4-induced M2 macrophage polarization, reduces the cholesterol content of Hepa1-6 cells in a co-culture system, and inhibits the promotion of hepatocellular carcinoma cells by M2 macrophages. In addition, successful overexpression of SREBP1 in Hepa1-6 cells showed more pronounced cellular activity whereas Cinobufacini injection inhibited this change and reduced intracellular lipid levels. CONCLUSION: Cinobufacini injection inhibits cholesterol synthesis within the tumor microenvironment via the AMPK/SERBP1/FASN signaling pathway, which in turn blocks the M2 polarization of macrophages, leading to the weakening of hepatocellular carcinoma growth and migration, and the promotion of its apoptosis. Our findings provide an important Introduction to understanding the molecular mechanism of Cinobufacini injection's anticancer activity and provide reliable theoretical and experimental support for its clinical application.

Basic research and clinical exploration of cold atmospheric plasma for skin wounds.

Skin wounds, such as burns, diabetic foot ulcers, pressure sores, and wounds formed after laser or surgical treatment, comprise a very high proportion of dermatological disorders. Wounds are treated in a variety of ways; however, some wounds are greatly resistant, resulting in delayed healing and an urgent need to introduce new alternatives. Our previous studies have shown that cold atmospheric plasma (CAP) has antibacterial activity and promotes cell proliferation, differentiation, and migration in vitro. To further advance the role of CAP in wound healing, we evaluated the safety and efficacy of CAP in vitro by irradiation of common refractory bacteria on the skin, irradiation of normal skin of rats and observing reactions, treatment of scald wounds in rats, and treating clinically common acute wounds. Our findings revealed that CAP can eliminate refractory skin bacteria in vitro; CAP positively affected wound healing in a rat scalding wound model; and direct CAP irradiation of low intensity and short duration did not lead to skin erythema or edema. CAP promises to be a new, economical, and safe means of wound treatment.

Loureirin B protects against cerebral ischemia/reperfusion injury through modulating M1/M2 microglial polarization via STAT6 / NF-kappaB signaling pathway.

The latest research indicates that modulating microglial polarization from M1 to M2 phenotype may be a coping therapy for ischemic stroke. The present study thereby evaluated the effects of loureirin B (LB), a monomer compound extracted from Sanguis Draconis flavones (SDF), on cerebral ischemic injury and the potential mechanisms. The middle cerebral artery occlusion (MCAO) model was established in male Sprague-Dawley rats to induce cerebral ischemia/reperfusion (I/R) injury in vivo, and BV2 cells were exposed to oxygen-glucose deprivation and reintroduction (OGD/R) to mimic cerebral I/R injury in vitro. The results showed that LB significantly reduced infarct volume, neurological deficits and neurobehavioral deficits, apparently improved histopathological changes and neuronal loss in cortex and hippocampus of MCAO/R rats, markedly decreased the proportion of M1 microglia cells and the level of pro-inflammatory cytokines, and increased the proportion of M2 microglia and the level of anti-inflammatory cytokines both in vivo and in vitro. In addition, LB evidently improved the p-STAT6 expression and reduced the NF-κB (p-p65) expression after cerebral I/R injury in vivo and in vitro. IL-4 (a STAT6 agonist) exhibited a similar impact to that of LB, while AS1517499 (a STAT6 inhibitor) significantly reversed the effect of LB on BV-2 cells after OGD/R. These findings point to the protection of LB against cerebral I/R injury by modulating M1/M2 polarization of microglia via the STAT6/NF-κB signaling pathway, hence LB may be a viable treatment option for ischemic stroke.

Effects of urban particulate matter on gut microbiome and partial schizophrenia-like symptoms in mice: Evidence from shotgun metagenomic and metabolomic profiling.

BACKGROUND: Epidemiological evidence reported that particulate matter (PM) was associated with increased schizophrenia (SCZ) risk. Disturbance of gut microbiome was involved in SCZ. However, it remains unclear whether PM induces SCZ-like symptoms and how gut microbiome regulates them. Therefore, a multi-omics animal experiment was conducted to verify how urban PM induces SCZ-like behavior and altered gut microbiota and metabolic pathways. METHODS: Using a completely random design, mice were divided into three groups: PM group, control group and MK801 group, which received daily tracheal instillation of PM solution, sterile PBS solution and intraperitoneal injection of MK801 (establish SCZ model), respectively. After a 14-day intervention, feces were collected for multi-omics testing (shotgun metagenomic sequencing and untargeted metabolomic profiling), followed by open field test, tail suspension test, and passive avoidance test. Besides, fecal microbiome of PM group and control group were transplanted into "pseudo-sterile" mice, then behavioral tests were conducted. RESULTS: Similar to MK801 group, mice in PM group showed SCZ-like symptoms, including increased spontaneous activity, excitability, anxiety and decreased learning and spatial memory. PM exposure significantly increased the relative abundance of Verrucomicrobia and decreased that of Fibrobacteres et al. The metabolism pathways of estrogen signaling (estriol, 16-glucuronide-estriol and 21-desoxycortisol) and choline metabolism (phosphocholine) were significantly altered by PM exposure. Verrucomicrobia was negatively correlated with the level of estriol, which was correlated with decreased learning and spatial memory. Fibrobacteres and Deinococcus-Thermus were positively correlated with the level of phosphocholine, which was correlated with increased spontaneous activity, excitability and anxiety. Fecal microbiome transplantation from PM group mice reproduced excitability and anxiety symptoms. CONCLUSIONS: Exposure to PM may affect composition of gut microbiome and alterations of estrogen signaling pathway and choline metabolism pathway, which were associated with partial SCZ-like behaviors. But whether gut microbiome regulates these metabolic pathways and behaviors remains to be determined.

Gambogenic acid antagonizes the expression and effects of long non-coding RNA NEAT1 and triggers autophagy and ferroptosis in melanoma.

In this study, we investigated the molecular mechanism underlying melanoma proliferation, with the aim to discover effective interventions which may markedly improve clinical prognosis. The results showed that gambogenic acid (GNA) could inhibit the proliferation of melanoma cells in vivo (C57BL/6 mice) and in vitro. Long non-coding RNA sequencing was used to identify the most significant long non-coding RNA, i.e., nuclear enriched abundant transcript 1 (NEAT1). NEAT1 was is up-regulated in melanoma, which was found to closely relate to cell proliferation. Melanoma cell lines either over-expressing NEAT1 or with NEAT1 knockdown was established through cloning experiments. A model of transplanted tumors was established to verify the inhibitory effect of GNA on the proliferation of melanoma cells in vitro and in vivo by downregulating NEAT1. Downregulation of NEAT1-induced ferroptosis and autophagy was demonstrated by detecting the effects of NEAT1 overexpressed and downregulated melanoma cell lines and melanoma transplantation model mice. Mechanistically, downregulation of NEAT1 can weaken the direct binding of Slc7a11, indirectly leading to inhibiting GPX-4 activity and subsequent ferroptosis, while, mediating the AMPK/mTOR signal axis-induced autophagy. The levels of Furthermore, NEAT1 decrease under the treatment of Gambogenic acid (GNA), a promising natural anticancer compound, while NEAT1 overexpression suppresses GNA inhibition on cell vitality and eliminates GNA-induced melanoma cell ferroptosis and autophagy.

Anfibatide alleviates inflammation and apoptosis via inhibiting NF-kappaB/NLRP3 axis in ischemic stroke.

Recent evidence suggests that Nod-like receptor protein-3 (NLRP3) inflammasome is a key mediator of inflammatory response and can induce the activation of apoptosis signaling pathways in ischemic stroke. In this research, we assessed the effects of anfibatide (ANF) on inflammatory and apoptosis in cerebral ischemic injury and the potential mechanisms. Middle cerebral artery occlusion (MCAO) model was established on male Sprague-Dawley rats to induce cerebral ischemia/reperfusion (I/R) injury in vivo. Primary cortical neurons (PCN) cells were exposed to oxygen-glucose deprivation and reintroduction (OGD/R) to mimic cerebral I/R injury in vitro. The results showed that ANF markedly alleviated infarct volume, neurological deficit and neurobehavioral impairment in MCAO/R rats, enhanced cell viability and decreased LDH release in PCN after OGD/R. The number of TUNEL-positive cells, Bax, cleaved-caspase-3, p-IκBα, p-p65, NLRP3, ASC, cleaved caspase-1, IL-ß and IL-18 proteins expression were significantly upregulated in the cortex of MCAO/R rats and PCN exposed to OGD/R, NLRP3 and caspase-1 mRNA levels were also evidently elevated. Bcl-2 protein expression significantly decreased in the cortex of MCAO/R rats. Treatment with ANF obviously inhibited the expression of p-IκBα, p-p65, NLRP3, ASC, cleaved caspase-1, Bax and cleaved-caspase-3, promoted the expression of Bcl-2, then decreased the TUNEL-positive cell number and the level of inflammatory cytokines (IL-ß and IL-18) in cerebral ischemia reperfusion in vito and in vitro. Our findings suggest that ANF exerts effects of alleviating inflammation and apoptosis through inhibiting NF-kappaB/NLRP3 axis. ANF is a potential candidate for treating cerebral I/R injury.

Assessment of the toxicity and carcinogenicity of double-walled carbon nanotubes in the rat lung after intratracheal instillation: a two-year study.

BACKGROUND: Considering the expanding industrial applications of carbon nanotubes (CNTs), safety assessment of these materials is far less than needed. Very few long-term in vivo studies have been carried out. This is the first 2-year in vivo study to assess the effects of double walled carbon nanotubes (DWCNTs) in the lung and pleura of rats after pulmonary exposure. METHODS: Rats were divided into six groups: untreated, Vehicle, 3 DWCNT groups (0.12 mg/rat, 0.25 mg/rat and 0.5 mg/rat), and MWCNT-7 (0.5 mg/rat). The test materials were administrated by intratracheal-intrapulmonary spraying (TIPS) every other day for 15 days. Rats were observed without further treatment until sacrifice. RESULTS: DWCNT were biopersistent in the rat lung and induced marked pulmonary inflammation with a significant increase in macrophage count and levels of the chemotactic cytokines CCL2 and CCL3. In addition, the 0.5 mg DWCNT treated rats had significantly higher pulmonary collagen deposition compared to the vehicle controls. The development of carcinomas in the lungs of rats treated with 0.5 mg DWCNT (4/24) was not quite statistically higher (p = 0.0502) than the vehicle control group (0/25), however, the overall incidence of lung tumor development, bronchiolo-alveolar adenoma and bronchiolo-alveolar carcinoma combined, in the lungs of rats treated with 0.5 mg DWCNT (7/24) was statistically higher (p < 0.05) than the vehicle control group (1/25). Notably, two of the rats treated with DWCNT, one in the 0.25 mg group and one in the 0.5 mg group, developed pleural mesotheliomas. However, both of these lesions developed in the visceral pleura, and unlike the rats administered MWCNT-7, rats administered DWCNT did not have elevated levels of HMGB1 in their pleural lavage fluids. This indicates that the mechanism by which the mesotheliomas that developed in the DWCNT treated rats is not relevant to humans. CONCLUSIONS: Our results demonstrate that the DWCNT fibers we tested are biopersistent in the rat lung and induce chronic inflammation. Rats treated with 0.5 mg DWCNT developed pleural fibrosis and lung tumors. These findings demonstrate that the possibility that at least some types of DWCNTs are fibrogenic and tumorigenic cannot be ignored.

Use of cellular metabolomics and lipidomics to decipher the mechanism of Huachansu injection-based intervention against human hepatocellular carcinoma cells.

Huachansu (HCS) injection has been used for the clinical treatment of hepatocellular carcinoma (HCC) patients in China for more than thirty years. However, the underlying mechanisms of HCS injection in inhibiting HCC remains unclear. In this study, we aimed to evaluate the effects of HCS injection on HCC both in vitro and in vivo and to systematically explore the underlying mechanisms of HCS's actions against HCC using an ultrahigh-performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS)-based cellular metabolomics and lipidomics approach. MTT and DAPI staining were performed to evaluate the effects of HCS injection on cell proliferation and apoptosis, respectively, in human HepG2 HCC cells with and without HCS treatment. An H22 hepatoma-bearing mouse model was employed to investigate the anti-HCC action of HCS injection in vivo. A UPLC-HRMS-based cellular metabolomics and lipidomics method was employed to characterize cellular metabolite fingerprinting, to identify potential biomarkers associated with the effect of HCS injection on the inhibition of HCC and to explore the anti-HCC mechanisms of HCS injection from the perspective of cellular metabolism. As a result, HCS injection inhibited the proliferation of HepG2 cells and induced their apoptosis. Body weight loss, sarcoma weights and volumes of H22 hepatoma-bearing mice in the HCS treatment group were significantly attenuated compared to those in the model group. Twenty-five biomarkers responsible for the intervention effect of HCS were screened and annotated using UPLC-HRMS-based cellular metabolomics and lipidomics. Among these biomarkers, two cellular metabolites, uridine diphosphate-N-acetylglucosamine (UDP-GlcNAc) and glutathione, were confirmed unambiguously using the reference substances. Metabolic pathway analysis revealed that the anti-HCC effects of HCS injection primarily involved the metabolism of glutathione, cysteine and methionine, sphingolipid, fatty acid biosynthesis, amino sugar and nucleotide sugars. As evidenced by western blotting experiments, the changes in UDP-GlcNAc content in response to HCS injection were linked to the inhibition of UDP-GlcNAc pyrophosphorylase 1 (UAP1) expression in HepG2 cells. In conclusion, HCS injection has a curative anti-HCC effect, and a UPLC-HRMS-based cellular metabolomics and lipidomics approach combined with further confirmatory experiments is a promising means of discovering its underlying mechanisms.

Qingluoyin granules protect against adjuvant-induced arthritis in rats via downregulating the CXCL12/CXCR4-NF-κB signalling pathway.

CONTEXT: Qingluoyin (QLY) is a traditional Chinese medicine (TCM) formula which has been used in treating human rheumatoid arthritis (RA) for years in China. OBJECTIVE: This study investigates the effect of QLY granules on adjuvant arthritis (AA) and the possible mechanism. MATERIALS AND METHODS: Sprague-Dawley (SD) rats were injected with Complete Freund's adjuvant (CFA) to induce the AA model. After the onset of arthritis, rats received intragastric administrations of the QLY granules (1.35, 2.70, and 5.40 g/kg) or Tripterygium glycosides (TG) tablets (positive drug, 10 mg/kg) for 14 d. After 28 d immunization, the symptoms, inflammatory parameters and molecular mechanisms were investigated. RESULTS: In the QLY granule (1.35, 2.70, and 5.40 g/kg) therapy groups, the arthritis index decreased to 6.30 ± 2.06, 5.80 ± 1.55, 5.30 ± 1.16 compared with the model (9.00 ± 3.01), paw swelling decreased to 1.56 ± 0.40, 1.28 ± 0.38, 1.12 ± 0.41 mL compared with the model (2.22 ± 0.73 mL). QLY granules (1.35, 2.70 and 5.40 g/kg) significantly reduced the thymus and the spleen indexes, inhibited the production of pro-inflammatory cytokines, and alleviated the pathological changes of joints compared with the model group. Furthermore, the treatment of QLY granules (2.70 and 5.40 g/kg) markedly inhibited CXCL12, CXCR4 (in spleen and synovium) and p-NF-κB p65 (in synovium) protein expression of AA rats. CONCLUSIONS: QLY granules have obvious therapeutic effects on AA rats, which may be associated with downregulating the CXCL12/CXCR4-NF-κB signalling pathway. QLY granules can be used as a candidate for the treatment of RA, which deserves further study.

Anfibatide Preserves Blood-Brain Barrier Integrity by Inhibiting TLR4/RhoA/ROCK Pathway After Cerebral Ischemia/Reperfusion Injury in Rat.

The disruption of the blood-brain barrier (BBB) and the consequent brain edema are major contributors to the pathogenesis of cerebral ischemia/reperfusion injury. RhoA is generally thought to play a crucial role in the process of BBB disruption and participate in the signaling pathways emanating from TLR4. However, it remains unverified the regulatory role of TLR4 in the RhoA/ROCK pathway in cerebral I/R injury and its effects on the BBB as well. The present study probes into the protective effect of ANF on the BBB after cerebral I/R injury and the possible mechanisms. Focal cerebral ischemia was induced by 120 min of transient middle cerebral artery occlusion (MCAO). ANF (1, 2, 4 µg/kg) was achieved by intravenous injection after 120 min of MCAO followed by 1, 24, 48, and 72 h reperfusion. Evans blue extravasation, brain water content, RhoA activity, and the expressions of TLR4, ROCK1/2, p-MLC2, MMP-2/9, ZO-1, occludin, and claudin-5 protein in rat brain were evaluated 72 h after reperfusion. ANF could significantly reduce the Evans blue extravasation and water content in the ipsilateral hemisphere and obviously increase the occludin, claudin-5, and ZO-1 expression after cerebral I/R injury. Furthermore, cerebral I/R injury induced apparently increased expression of TLR4, RhoA-GTP, ROCK1/2, p-MLC2, and MMMP-2/9, which, however, could be remarkably alleviated by ANF intervention. Taken together, the TLR4/RhoA/ROCK signaling pathway is implicated in BBB breakdown after cerebral I/R injury, and ANF preserves BBB integrity, probably via inhibiting the TLR4/RhoA/ROCK signaling pathway.

[Efficacy of Kangshuailing Gao on benign prostatic hyperplasia in rats].

OBJECTIVE: To investigate the effect of Kangshuailing Gao (KG) on benign prostatic hyperplasia (BPH) in rats and its action mechanisms. METHODS: Fifty BPH model rats were randomized into five groups of equal number, BPH model control, finasteride control, and high-, medium- and low-dose KG, to be treated intragastrically with distilled water, finasteride solution at 0.52 mg/kg, and KG solution at 4.16, 2.08 and 1.04 g/kg respectively once a day for 30 days consecutively. Another 10 normal healthy rats were taken as blank controls. The rats were weighed once a week during the treatment. The wet weight and index of the prostate were obtained after treatment, followed by measurement of the contents of serum estradiol (E2) and dihydrotestosterone (DHT), testosterone (T) and hypoxia-inducible factor-1α (HIF-1α) in the prostatic tissue, and observation of histomorphological changes in the prostate under the light microscope. RESULTS: Compared with the BPH model control group, high- and medium-dose KG significantly reduced the prostate wet weight (ï¼»0.84 ± 0.08ï¼½ vs ï¼»0.69 ± 0.04ï¼½ and ï¼»0.71 ± 0.07ï¼½ g, P < 0.01), the prostatic index (ï¼»0.28 ± 0.03ï¼½% vs ï¼»0.20 ± 0.02ï¼½% and ï¼»0.22 ± 0.03ï¼½%, P < 0.01), and the levels of T (ï¼»4.63 ± 1.25ï¼½ vs ï¼»2.44 ± 0.47ï¼½ and ï¼»2.91 ± 0.69ï¼½ ng/L, P < 0.01) and DHT (ï¼»154.44 ± 20.25ï¼½ vs ï¼»88.23 ± 13.63ï¼½ and ï¼»90.52 ± 16.44ï¼½ nmol/L, P < 0.01), but increased the level of E2 (ï¼»0.95 ± 0.24ï¼½ vs ï¼»1.19 ± 0.14ï¼½ and ï¼»1.20 ± 0.22ï¼½ nmol/L, P < 0.01) in the serum. High-dose KG remarkably reduced the overexpression of HIF-1α in the prostate tissue of the BPH model rats (P < 0.01) and alleviated such BPH-related symptoms as epithelium thinning, intraglandular secretion reduction, and interstitial substance decrease. CONCLUSIONS: Kangshuailing Gao acted effectively on BPH in the model rats by reducing the androgen level, balancing the estrogen/androgen ratio, and downregulating the expression of HIF-1α in the prostate tissue.

The Protective Effect of Total Flavones from Rhododendron simsii Planch. on Myocardial Ischemia/Reperfusion Injury and Its Underlying Mechanism.

OBJECTIVES: Total flavones from Rhododendron simsii Planch. (TFR) are the effective part extracted from the flowers of Rhododendron simsii Planch. and have obvious protective effects against cerebral ischemic or myocardial injuries in rabbits and rats. However, their mechanism of cardioprotection is still unrevealed. Therefore, the present study was designed to investigate the effect of TFR on myocardial I/R injury and the underlying mechanism. METHODS: TFR groups were treated by gavage once a day for 3 days at a dose of 20, 40, and 80 mg/kg, respectively, and then the model of myocardial I/R injury was established. Myocardial infarction, ST-segment elevation, and the expression of UTR, ROCK1, ROCK2, and p-MLC protein in rat myocardium were determined at 90 min after reperfusion. UTR siRNA in vivo transfection and competition binding assay method were used to study the relationship between the protective effect of TFR and UTR. RESULTS: The expression of UTR protein markedly decreased in myocardium of UTR siRNA transfection group rats. TFR could significantly reduce the infarct size and inhibit the increase of RhoA activity and ROCK1, ROCK2, and p-MLC protein expressions both in WT and UTR knockdown rats. The reducing rate of TFR in myocardial infarction area, RhoA activity, and ROCK1, ROCK2, and p-MLC protein expressions in UTR knockdown rats decreased markedly compared with that in WT rats. In addition, TFR had no obvious effect on the increase of ΣST in UTR knockdown rats in comparison with that in model group. In particular, TFR could significantly inhibit the combination of [125I]-hu-II and UTR, and IC50 was 0.854 mg/l. CONCLUSIONS: The results indicate that the protective effect of TFR on I/R injury may be correlated with its blocking UTR and the subsequent inhibition of RhoA/ROCK signaling pathway.

Anfibatide protects against rat cerebral ischemia/reperfusion injury via TLR4/JNK/caspase-3 pathway.

Anfibatide (ANF) is a GPIb antagonist derived from the protein complex agglucetin. Previous studies have showed that it has protective effect on cerebral ischemia/reperfusion injury, the mechanism of which is still unclear, however. The present study was designed to investigate the protective effect of ANF on cerebral I/R injury in rats and the possible mechanisms. Focal cerebral ischemia was induced by 90min of transient middle cerebral artery occlusion (MCAO). ANF (1, 2, 4µg/kg) was achieved by intravenous injection after 120min of MCAO followed by 1h, 24h ,48h and 72h reperfusion. Neurological deficit, infarct volume, histopathology, neuronal apoptosis, NeuN and the expression of TLR4, total and phosphorylated c-Jun NH2-terminal kinase (JNK/p-JNK), Bcl-2, Bax, caspase-3, NF-κB protein in rat brain, the levels of IL-1ß, IL-6 and TNF-α in serum were evaluated 72h after reperfusion. ANF could significantly decrease neurological score, reduce the infarct volumes, ameliorate the histopathological alteration, attenuate the neuronal apoptosis and increase the fluorescence density of NeuN in the rat brain. Furthermore, ANF could obviously decrease the expression of TLR4, p-JNK, caspase-3, NF-κB , relative ratio of Bax/Bcl-2 in brain and the levels of IL-1ß, IL-6 and TNF-α in serum. The results indicate that ANF has protective effect against cerebral I/R injury in rats and the underlying mechanism may be associated with the suppression of apoptosis through inhibiting TLR4/JNK/caspase-3 signaling pathway.

The effect of ethyl acetate extract from persimmon leaves on Alzheimer's disease and its underlying mechanism.

BACKGROUND: Alzheimer's disease (AD) is one of the most prevalent neurodegenerative disorders characterized by neuronal loss in the brain and cognitive impairment. AD is now considered to be the third major cause of death in developed countries, after cardiovascular disease and cancer. Persimmon leaves are used as a popular folk medicine to treat hypertension, angina and internal haemorrhage in Cyangbhina, and it has been reported that ethyl acetate extract of persimmon leaves (EAPL) displays a potential therapeutic effect on neurodegenerative diseases. HYPOTHESIS/PURPOSE: This study was designed to investigate the effects of EAPL on AD, to clarify the possible mechanism by which EAPL exerts its beneficial effects and prevents AD, and to determine the major constituents involved. STUDY DESIGN: AD model was established by bilateral injection of Aß1-42 into the hippocampus of rats. The cognitive performance was determined by the Morris water maze and step-down tests. Superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), malondialdehyde (MDA), apoptosis, total and phosphorylated c-Jun NH2-terminal kinase (JNK/p-JNK), caspase-3, Bax and Bcl-2 were determined. In addition, a sensitive and reliable LC-QTOF-MS method was applied to identify the major compounds present in EAPL. RESULTS: EAPL at doses of 200mg/kg, 400mg/kg could markedly reduce the latency, significantly increase the time in the first quadrant and number of the target crossing times in Morris water maze test, markedly increase the latency and reduce the number of errors in the step-down test, significantly inhibit the reductions in SOD and GSH-Px activities, and increase the level of MDA. In addition, EAPL treatment attenuated neuronal apoptosis in the hippocampus, reduced the expression of p-JNK, caspase-3, and the relative ratio of Bax/Bcl-2. Meanwhile, 32 constituents were identified by LC-QTOF-MS/MS assays. CONCLUSION: The results indicate that EAPL has a potent protective effect on cognitive deficits induced by Aß in rats and this effect appears to be associated with the regulation of the antioxidative defense system and the mechanism of mitochondrial-mediated apoptosis. Furthermore, analysis of the LC-MS data suggests that flavonoids and triterpenoids may be responsible for the potential biological effects of EAPL.

Urotensin-â ¡Receptor Antagonist SB-710411 Protects Rat Heart against Ischemia-Reperfusion Injury via RhoA/ROCK Pathway.

AIM: SB-710411 is a rat selective urotensin-II (U-II) receptor antagonist, which can block U-II-induced contraction of the aorta and inhibit U-II-induced myocardial fibrosis in rats. However, the effect of SB-710411 on myocardial ischemia-reperfusion (I/R) injury is unclear. The present study was designed to investigate whether SB-710411 has a protective effect on myocardial I/R injury in rats and the possible mechanisms. METHODS AND RESULTS: Myocardial I/R injury was induced by occluding the left anterior descending coronary artery in adult male Sprague-Dawley rats. Hemodynamic parameters, electrocardiogram (ECG), infarct size, histological alteration, lactate dehydrogenase (LDH), creatine phosphokinase-MB (CK-MB), cardiac troponin I (cTnI), RhoA, and the protein expressions of U-II receptor (UTR), ROCK1 and ROCK2 were evaluated. Cardiac I/R injury significantly up-regulated the expressions of UTR, ROCK1 and ROCK2 proteins in rat myocardium. SB-710411 1.0 and 2.0 µg/kg significantly reduced cardiac I/R-induced the infarct size and histological damage in rat myocardium, markedly inhibited the changes of hemodynamic parameters and the increases of ST-segment in ECG, the serum LDH and CK-MB activities and cTnI level in rats subjected to myocardial I/R injury. Furthermore, SB-710411 obviously prevented myocardial I/R-increased RhoA activity and UTR, ROCK1 and ROCK2 protein expressions. CONCLUSIONS: Our results indicate that cardiac I/R injury increases myocardial UTR expression, and SB-710411 has a potent protective effect on myocardial I/R injury in rats. The cardioprotection may be associated with the inhibition of UTR-RhoA/ROCK pathway.

A novel snake venom-derived GPIb antagonist, anfibatide, protects mice from acute experimental ischaemic stroke and reperfusion injury.

BACKGROUND AND PURPOSE: Ischaemic stroke is a serious disease with limited therapy options. Glycoprotein (GP)Ib binding to von Willebrand factor (vWF) exposed at vascular injury initiates platelet adhesion and contributes to platelet aggregation. GPIb has been suggested as an effective target for antithrombotic therapy in stroke. Anfibatide is a GPIb antagonist derived from snake venom and we investigated its protective effect on experimental brain ischaemia in mice. EXPERIMENTAL APPROACH: Focal cerebral ischaemia was induced by 90 min of transient middle cerebral artery occlusion (MCAO). These mice were then treated with anfibatide (4, 2, 1 µg·kg(-1) ), injected i.v., after 90 min of MCAO, followed by 1 h of reperfusion. Tirofiban, a GPIIb/IIIα antagonist, was used as a positive control. KEY RESULTS: Twenty-four hours after MCAO, anfibatide-treated mice showed significantly improved ischaemic lesions in a dose-dependent manner. The mice had smaller infarct volumes, less severe neurological deficits and histopathology of cerebrum tissues compared with the untreated MCAO mice. Moreover, anfibatide decreased the amount of GPIbα, vWF and accumulation of fibrin(ogen) in the vasculature of the ischaemic hemisphere. Tirofiban had similar effects on infarct size and fibrin(ogen) deposition compared with the MCAO group. Importantly, the anfibatide-treated mice showed a lower incidence of intracerebral haemorrhage and shorter tail bleeding time compared with the tirofiban-treated mice. CONCLUSIONS AND IMPLICATIONS: Our data indicate anfibatide is a safe GPIb antagonist that exerts a protective effect on cerebral ischaemia and reperfusion injury. Anfibatide is a promising candidate that could be beneficial for the treatment of ischaemic stroke.