Rearranged Homoadamantane-Type Polycyclic Polyprenylated Acylphloroglucinols from Hypericum pseudohenryi.

Hypseudohenones A-C (1-3), the first rearranged homoadamantane-type polycyclic polyprenylated acylphloroglucinols, were isolated from Hypericum pseudohenryi. Their structures with an unprecedented tricyclo[4.3.1.13,8]undecane-2,4,10-trione core were determined by spectroscopic analysis, quantum-chemical calculations, and X-ray crystallography. A method for determining the relative configuration at C-3 was established by the peak shape of H-28 or J-value of H-3/H-28. Moreover, 2-3 exhibited significant AChE inhibitory activity, and the interactions of 2-3 with AChE were evaluated by molecular docking.

A time-course study of microglial activation and dopaminergic neuron loss in the substantia nigra of mice with paraquat-induced Parkinson's disease.

Activated microglia play an active role in the pathogenesis of PD and paraquat (PQ) induces PD. The study was to understand the time relationship between microglial activation and dopaminergic neuron loss in the substantia nigra (SN) of PQ-induced PD mice. Male C57BL/6 mice were injected intraperitoneally with PQ, twice a week for six weeks. Some mice underwent behavioral assessments each week and were sacrificed for SN tissues, in which histopathological analysis, dopaminergic neuron loss, microglial activation and phenotypic characteristics were evaluated. The results showed that motor retardation, coordination disorders and limb stiffness occurred four weeks after PQ exposure, as well as the degeneration and loss of dopaminergic neurons in the SN. Activated microglia and increased CD68 expression appeared two weeks after PQ exposure in time-dependent manners. Increased CD86 and decreased CD206 expression were observed four weeks after PQ exposure, accompanied by increased TNF-α and IL-6 levels and decreased IL-10 and TGF-ß levels. These results indicate that PQ can activate microglia in vivo, and microglial activation precedes neuronal loss in the SN. Activated microglia are characterized by mixed M1/M2 polarization in the early stage and M1 polarization in the late stage of PQ-induced PD development.

Cardioprotective effects of Amentoflavone by suppression of apoptosis and inflammation on an in vitro and vivo model of myocardial ischemia-reperfusion injury.

Inflammation modulation is currently considered a promising therapeutic strategy to counteract the burden of cardiovascular disease. Amentoflavone (AME) is a natural biflavone with two apigenin molecules that, possess promising anti-inflammatory, anti-oxidative, and anti-cancer properties. In the present study, we aimed to investigate the effects of AME on myocardial ischemia-reperfusion injury in vivo and in vitro, and to elucidate the underlying mechanism. Our results showed that AME significantly reduced the levels of LDH, CK-MB, IL-6, IL-1ß, and TNF-α after hypoxia (H) 12 h/reoxygenation (R) 4 h treatment, and significantly increased the cell survival rate of H9c2 cardiomyocytes induced by H/R and inhibited their apoptosis rate. AME (25, 50, 100 mg·kg-1·d-1, i.g.) or a positive control drug diltiazem (DIZ) (16 mg·kg-1·d-1, i.g.) was used as pretreatment for 7 days; the myocardial ischemia-reperfusion(I/R) model was established. TTC staining results showed that the infarct volume was significantly reduced after AME and DIZ treatment. Oral administration of AME dose-dependently ameliorated I/R injury-induced increase in pro-inflammatory factors (IL-6, IL-1ß, and TNF-α) and levels of LDH and CK-MB. Results of TUNEL and HE staining showed that the I/R model had more induced apoptosis, but could be effectively reduced by pretreatment with AME. After surgery, the heart of the rat was examined via western blotting to detect inflammation-related proteins. Compared with the sham group, the p-AKT in the I/R group was significantly reduced and the content of p-NF-κBp65 was significantly increased. However, these changes could be reversed by AME treatment. DIZ treatment exerted similar beneficial effects in I/R rats as the high dose of AME did. This study highlights the excellent therapeutic potential of AME for managing myocardial ischemia-reperfusion injury.

Ultrasound Combined With Microbubbles Loading BDNF Retrovirus to Open BloodBrain Barrier for Treatment of Alzheimer's Disease.

Background: Brain-derived nerve growth factor (BDNF) is a promising effective target for the treatment of Alzheimer's disease (AD). BDNF, which has a high molecular weight, has difficulty in crossing the blood-brain barrier (BBB). The study aimed to prepare microbubbles loading brain-derived nerve growth factor (BDNF) retrovirus (MpLXSN-BDNF), to verify the characteristics of the microbubbles, and to study the therapeutic effect of the microbubbles combined with ultrasound on the opening of the blood-brain barrier in an AD rat model. Methods: 32 adult male SD rats were randomly divided into four groups: control group, ultrasound + pLXSN-EGFP microbubble group (U + MpLXSN-BDNF), ultrasound + pLXSN-BDNF microbubble group, and ultrasound + microbubble + pLXSN-BDNF virus group (U + MpLXSN-BDNF), with eight rats in each group. At the same time, the left hippocampus of rats was irradiated with low-frequency focused ultrasound guided by MRI to open the blood-brain barrier (BBB). The effects of BDNF overexpression on AD rats were evaluated behaviorally before and 1 month after the treatment. The number of acetylcholinesterase (ChAT)-positive cells and the content of acetylcholine (ACh) in brain tissues were determined by immunohistochemistry and high-performance liquid chromatography (HPLC), respectively. IF staining of synaptic spines and Western blot of synaptophysin presented herein detected synaptic density recovery. Results: Signal intensity enhancement at the BBB disruption sites could be observed on the MR images. The behavioral evaluation showed that the times of crossing the original platform in the U + MpLXSN-BDNF group increased significantly after treatment. Immunohistochemistry and HPLC revealed that the number of ChAT-positive neurons and the contents of ACh in the brain were significantly decreased in the treated groups compared with the controls. IF staining of synaptic spines and Western blot data of synaptophysin showed that the U + MpLXSN-BDNF group can recover the synaptic loss better by BDNF supplementation than the other treatment groups. Conclusion: Ultrasound combined with viral microbubbles carrying BDNF can increase the transfection efficiency of brain neurons, promote the high expression of exogenous gene BDNF, and play a therapeutic role in the AD model rats.

Attenuation of paraquat-induced inflammation by inhibitors of phosphorylation of mitogen-activated protein kinases in BV2 microglial cells.

Paraquat has dopaminergic neurotoxicity and potentially contributes to Parkinson's disease (PD) as a risk factor. However, the cellular and molecular mechanisms of PQ-induced neurodegeneration have not been clearly elucidated. Studies have shown that PQ induces microglial neuroinflammation through toll-like receptor 4 (TLR4)-nuclear factor-κB pathway, resulting in neuronal cell loss. Mitogen-activated protein kinases (MAPKs) are involved in the production of pro-inflammatory cytokines in microglia, and in this study, the role of MAPKs in PQ-activated microglial inflammation was investigated. Murine BV2 microglial cells were treated with 40 µM of PQ following pretreatment of the cells with selective inhibitor of MAPKs phosphorylation for blockage of the phosphorylation of ERK, JNK and P38, or a specific TLR4 inhibitor for blocking the activation of TLR4. The protein expression of phosphorylated ERK, JNK and p38, and the transcription expression of pro-inflammatory mediators were assessed with Western blotting and qRT-PCR technique, respectively. The results indicated that PQ significantly induced the phosphorylation of ERK, JNK and P38 in microglia, while MAPKs inhibitors suppressed PQ-induced phosphorylation of ERK, JNK and P38, and reduced the transcription level of pro-inflammatory cytokines. PQ-stimulated phosphorylation of ERK, JNK and P38 was also reduced by TLR4 inhibitor. The inhibited intensity in the level of pro-inflammatory cytokine transcription was obviously greater in TLR4 inhibitor + PQ group than in each MAPK inhibitor + PQ group. Taken together, inhibition of MAPKs phosphorylation partially attenuates PQ-induced microglial inflammation, which may become a potential intervention strategy for PQ neurotoxicity.

Garsubelone A, the First Dimeric Polycyclic Polyprenylated Acylphloroglucinols with Complicated Heptacyclic Architecture from Garcinia subelliptica.

Garsubelone A (1), the first dimeric polycyclic polyprenylated acylphloroglucinols type metabolite featuring a complicated 6/6/6/6/6/6/6 heptacyclic architecture containing 10 stereogenic centers, was isolated from Garcinia subelliptica. Biogenetically, this compound was constructed by the plausible monomeric precursor, garsubelone B (2) and secohyperforin, via a key Diels-Alder cycloaddition to form an unique 2-oxabicyclo[3.3.1]nonane core. Their structures and absolute configurations were determined by comprehensive spectroscopic and X-ray diffraction techniques. The cytotoxic activities of these isolates were also evaluated.

Inhibiting expression of HSP60 and TLR4 attenuates paraquat-induced microglial inflammation.

Accumulating evidences suggest that heat shock protein 60 (HSP60) and toll-like receptor 4 (TLR4) are involved in triggering inflammatory response in microglia. Paraquat (PQ) evokes microglial inflammation by up-regulating expression of HSP60-TLR4-myeloid differentiation factor 88 (Myd88)-nuclear factor-kappa B (NF-κB) in vitro. The aim of this study is to investigate the potential modulatory roles of HSP60 and TLR4 in PQ-induced inflammation. Before treated with PQ, microglia BV2 cells were pretreated using siRNA to knockdown HSP60 or with specific inhibitor to inhibit TLR4 expression. Expression of TLR4 and MyD88, and nuclear translocation of NF-κB subunit p65 were studied with immunoblotting and immunofluorescence, respectively. Expression of pro-inflammatory factors was assessed with quantitative real-time PCR. Knockdown of HSP60 or inhibition of TLR4 significantly reduced the expression of TLR4 and MyD88 and decreased the accumulation of NF-κB p65 in the nucleus. Gene expression of tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1ß), interleukin-6 (IL-6) and inducible nitric oxide synthase (iNOS) were also significantly decreased in response to PQ. These results suggest that HSP60 and TLR4 can modulate intracellular signaling of PQ-induced inflammation. Inhibiting HSP60 or TLR4 reduces significantly the intensity of inflammation in PQ-activated microglia.

Efficacy of a Chinese Herbal Medicine Compound Zhangpi Ointment against Hydroxyurea-Induced Leg Ulcers: A Prospective, Randomized, Open-Label, Controlled Clinical Trial.

Objective. The randomized controlled trial was to evaluate the efficacy of topical Chinese herbal Zhangpi Ointment for hydroxyurea-induced leg ulcers in patients with myeloproliferative neoplasms. Patients and Methods. This single-center, prospective, randomized, open-label, controlled clinical trial conducted at Shanghai Ninth People's Hospital enrolled 54 patients with hydroxyurea-induced leg ulcers. Patients were randomly assigned to the control group (n = 27) treated with chlorhexidine dressing or the intervention group (n = 27) treated with the Zhangpi Ointment. Finally, 26 patients in the control group and 23 patients in the intervention group completed 8 weeks of observation. Results. The rate of complete healing was 100% for the intervention group, which was significantly higher than that of the control group (96.15%) (P<0.05). Furthermore, the intervention group achieved a significantly higher rate of wound healing (95.56%) than the control group (69.02%) at week 4 (P<0.01). The intervention group took 34 ± 5 days to achieve complete healing while the control group took 41 ± 7 days (P < 0.01). Moreover, grade 3/4 side effects were observed in neither group. Conclusion. The Zhangpi Ointment is effective in promoting the healing of hydroxyurea-induced leg ulcers in patients with myeloproliferative neoplasms, providing a therapeutic option for a condition that is recalcitrant to conventional therapy.

[Effect of adiponectin postconditioning against myocardial ischemia/reperfusion injury in rats and role of ADP/PI3K/Akt pathway in adiponectin postconditioning].

OBJECTIVE: To investigate the effects of adiponectin(ADP) postconditioning against myocardial ischemia/reperfusion injury(MIRI) in rats and role of ADP/PI3K/Akt pathway in ADP postconditioning. METHODS: SD rat was connected to ventilator by tracheal intubation under anesthesia, then left anterior descending coronary artery (LAD) was threaded between left auricle and pulmonary artery cone after exposing heart by surgery. MIRI model was induced by ligation of LAD for 30 min and the following reperfusion for 120 min. Rats were divided randomly into 5 groups (n=12):① Sham group:LAD was threaded without ligation; ② MIRI group; ③ADP group (ADP postconditioning) were subjected to intravenous injection of ADP when LAD ligation for 10 min and the ligation held for 20 min after that, then reperfusion for 120 min; ④ ADP+LY294002 group were subjected to injection of ADP and LY294002 when LAD ligation for 10 min, the other steps were the same as ADP group; ⑤ LY294002 group were subjected to injection of LY294002 when LAD ligation for 10 min, the other steps were the same as ADP group. Titers of lactate dehydrogenase(LDH) and cardiac troponin I(cTnI) in plasma were observed, expressions of PI3K, Akt, phosphorylated-Akt(p-Akt), ADP mRNA, ADPR1 mRNA and PI3k mRNA in myocardial tissue were measured. RESULTS: Compared with sham group, the levels of LDH and cTnI in MIRI group were increased (P<0.05); Compared with MIRI group, the levels of LDH and cTnI in ADP group were decreased (P<0.05); Compared with ADP group, the levels of LDH and cTnI were increased in LY294002 applying groups(P<0.05). Compared with MIRI group, the expressions of PI3K, p-Akt, ADP mRNA, ADPR1 mRNA and PI3K mRNA were increased in ADP group (P<0.05), the above mentioned 5 parameters in LY294002 applying groups were decreased(P<0.05). CONCLUSIONS: ADP postconditioning could reduce MIRI in rats, the protective effect might have relation to ADP/PI3k/Akt pathway.

Synergistic effects of curcumin and bevacizumab on cell signaling pathways in hepatocellular carcinoma.

The aim of the present study was to explore the effects of curcumin in combination with bevacizumab on the vascular endothelial growth factor (VEGF)/VEGF receptor (VEGFR)/K-ras pathway in hepatocellular carcinoma. A total of 30 Sprague Dawley (SD) rats were randomly divided into five groups: Control, model, curcumin, VEGF blocker, and curcumin + VEGF blocker groups. The mRNA levels of VEGF and VEGFR in all groups were subsequently measured by quantitative reverse transcriptase-polymerase chain reaction and the protein expression of K-ras was detected by western blot analysis. Compared with the control group, the mRNA levels of VEGF and VEGFR were revealed to be significantly increased in the model, curcumin and VEGF blocker groups. The VEGF mRNA levels in the curcumin, VEGF blocker and curcumin + VEGF blocker groups were all decreased when compared with the model group. In addition, the VEGF mRNA levels in the curcumin + VEGF blocker group were significantly lower compared with the curcumin group (P<0.05). The VEGF mRNA levels in the curcumin, VEGF blocker and curcumin + VEGF blocker groups were decreased when compared with the model group (P=0.0001). No significant differences in VEGF mRNA levels were identified between the VEGF blocker and curcumin groups (P=0.863), whereas the VEGF mRNA levels in the curcumin + VEGF blocker group were significantly lower than that of the curcumin group (P=0.025). Curcumin and the VEGF blocker are each capable of inhibiting hepatocellular carcinoma progression by regulating the VEGF/VEGFR/K-ras pathway. The combination of the two compounds has a synergistic effect on the inhibition of the effects of the VEGF signaling pathways in hepatocellular carcinoma progression.

Oxygen consumption and ammonia excretion of black carp (Mylopharyngdon piceus Richardson) and allogynogenetic crucian carp (Carassius auratus gibelio female x Cyprinus carpio male) fed different carbohydrate diets.

Oxygen consumption and ammonia excretion of black carp (Mylopharyngdon piceus Richardson) (4.6±0.3 g) and allogynogenetic crucian carp (Carassius auratus gibelio ♀×Cyprinus carpio ♂) (5.7±0.5 g) were examined when fish fed two types of carbohydrate (dextrin and glucose) at two levels (20 and 40%) each. The diets were isonitrogenous (40% dry matter) and isocaloric at 18.5 kJ g(−1) (dry matter) by adjusting the oil content to 10.1 and 1.5%, respectively. In black carp, the interactions between the carbohydrate type and level were found in oxygen consumption at 3 and 6 h and in ammonia excretion at 6 h after feeding. At 20% carbohydrate, no significant difference was observed between dextrin and glucose in oxygen consumption. However, at 40% carbohydrate, oxygen consumption in fish fed glucose was significantly higher than that in fish fed dextrin at 3 and 6 h after feeding. Within the dextrin diets, no significant differences in both oxygen consumption and ammonia excretion were detected between the two carbohydrate levels. Within the glucose diets, however, fish fed 40% glucose showed significantly higher oxygen consumption than those fed 20% glucose at 3 and 6 h after feeding. Ammonia excretion in black carp fed 40% glucose was higher than that in black carp fed 40% dextrin at 6 h and also found higher than those in the other three treatments at 24 h after feeding. The postprandial oxygen consumption and the ammonia excretion in crucian carp fed 40% glucose were the highest, but no significant differences were observed. Our data indicate that the escalation of glucose to 40% in a fish diet results in high oxygen consumption and ammonia excretion in black carp, suggesting that the efficiency of glucose as an energy source for this fish is compromised by the high metabolic expenditure after feeding. Crucian carp, on the other hand, have a better ability to cope with dietary carbohydrates.

Down-regulation of the expression of angiotensin II type 1 receptor in neonatal rat cardiac fibroblast by activation of PPARgamma signal pathway.

Peroxisome proliferator-activated receptor-gamma (PPARgamma) is one of the hormone nuclear receptors. Recent data have shown that activation of PPARgamma signal pathway has many positive effects on cardiovascular system. The goals of this study were to determine whether PPARgamma activator affects cardiac fibrosis and the possible mechanisms. Cardiac fibroblasts (CFs) of SD neonate rats were used in the study. Cells were divided into 4 groups: I--control group; II--pioglitazone group (Piog--PPARgamma agonist); III--angiotensin II (Ang II) group; and IV--Piog + Ang II group (Piog plus angiotensin II). mRNA and protein expression of collagen type I, III and angiotensin II type 1 receptor (AT1-R) were tested by reverse transcription--polymerase chain reaction and Western blotting. With the inhibition of actinomycin D, we investigated the impacts of Piog on the stability of AT1-RmRNA. Compared with group I, the mRNA and protein expression of collagen type I, III and AT1-R were up-regulated in group III (P < 0.05). However with the effects of Piog in group IV, the expressions mentioned above were attenuated significantly (P < 0.05). With the effects of actinomycin D, AT1-RmRNA was reduced at the same degree in control and Piog groups at the same time points. These results indicated that treatment with Piog can attenuate Ang II-induced collagen synthesis in CFs through down-regulation of the AT1-R expression. With the intervention of actinomycin D, we suggested that PPARgamma agonist didn't affect the stability of AT1-RmRNA.