Yuan-Fu Lu

Neuroprotective effect of resveratrol on 6-OHDA-induced Parkinson's disease in rats.

The present study was undertaken to investigate the neuroprotective effects of resveratrol on 6-hydroxydopamine (6-OHDA)-induced Parkinsons disease in rats. 6-OHDA-induced Parkinsons disease rat model involves chronic inflammation, mitochondrial dysfunction, and oxidative stress, and the loss of the dopaminergic neurons in the substantia nigra is the predominant lesion. Resveratrol has been shown to have anti-inflammatory actions, and thus was tested for its beneficial effects using 6-OHDA-induced Parkinsons disease rat model. Adult Sprague-Dawley (SD) rats were unilaterally injected with 6-OHDA (5 microg/2 microl) into the right striatum, and the striatum damage was assessed by rotational test, ultrahistopathology, and molecular alterations. Resveratrol (10, 20 and 40 mg/kg) was then given orally to Parkinsons disease rats, daily for 10 weeks to examine the protective effects. Rotational test (turns of rats) showed that resveratrol significantly attenuated apomorphine-induced turns of rats in 6-OHDA-injuried Parkinsons disease rat model as early as two weeks of administration. Ultrastructural analysis showed that resveratrol alleviated 6-OHDA-induced chromatin condensation, mitochondrial tumefaction and vacuolization of dopaminergic neurons in rat substantia nigra. Furthermore, resveratrol treatment also significantly decreased the levels of COX-2 and TNF-alpha mRNA in the substantia nigra as detected by real-time RT-PCR. COX-2 protein expression in the substantia nigra was also decreased as evidenced by Western blotting. These results demonstrate that resveratrol exerts a neuroprotective effect on 6-OHDA-induced Parkinsons disease rat model, and this protection is related to the reduced inflammatory reaction.

Icariin isolated from Epimedium brevicornum Maxim attenuates learning and memory deficits induced by d-galactose in rats.

The effects of icariin (ICA), a major constituent of flavonoids from the Chinese medical herb Epimedium brevicornum Maxim, on spatial memory performances and expressions of hippocampus brain-derived neurotrophic factor (BDNF) and tyrosine kinase TrkB (tropomyosin receptor kinase B) were investigated in d-galactose (d-gal)-treated rats. Subcutaneous injection of d-gal (500mg/kg/d) for four months caused memory loss as detected by the Morris water maze, morphologic abnormalities of neurons in hippocampus region and the reduced expression of BDNF and TrkB were observed. ICA (60mg/kg/d) given orally 1h after subcutaneous injection of d-gal daily for 4months markedly attenuated d-gal-induced rats behavioral dysfunction and neurodegeneration, as evidenced by shortened escape latency and searching distance and rescued morphologic abnormalities, and also elevated the mRNA levels and the protein expressions of BDNF and TrkB in hippocampus, as evidenced by quantitative real-time RT-PCR and Western blotting analysis. But ICA had no significant influence on normal rats which were not injected d-gal. These results clearly demonstrated that d-gal produced learning and memory deficits after chronic administration, and ICA can protect neuron from d-gal insults and improve the memory loss.

Protective effects of icariin against learning and memory deficits induced by aluminium in rats.

1 The present study examined the protective effects of icariin against the learning and memory deficits in aluminium‐treated rats and its potential mechanisms of action. 2 Qualified rats were treated with 1600 p.p.m. AlCl3 in drinking water for 8 months and the ability of spatial learning and memory was tested by the Morris water maze. In the place navigation test, aluminium administration significantly increased the mean escape latency and searching distance. In space probing test, aluminium markedly decreased the searching time and searching distance in the quadrant where the platform was originally located. All tests indicated deficits in rat spatial learning and memory induced by aluminium. Icariin treatment (60 and 120 mg/kg, by gavage for 3 months) dose‐dependently protected against the development of aluminium‐induced spatial learning and memory deficits. 3 To examine the mechanisms responsible for the protection afforded by icariin, the superoxide dismutase (SOD) activity and malondialdehyde (MDA) content in the hippocampus were assayed biochemically and the level of Aβ1−40 in the hippocampus was determined immunohistochemically. Icariin treatment significantly increased SOD activity and decreased MDA and Aβ1−40 content in the hippocampus of aluminium‐intoxicated rats. 4 In conclusion, the present study demonstrates that icariin is effective in improving the spatial learning and memory of aluminium‐intoxicated rats. The mechanisms responsible appear to be due, at least in part, to an increased anti‐oxidant capacity and decreased lipid peroxidation and Aβ1−40 levels in the rat hippocampus.

Protective effects of icariin on brain dysfunction induced by lipopolysaccharide in rats

In this study we examined the protective effects of icariin, a flavonol isolated from Herba epimedii, on learning and memory in a rat model with brain inflammation induced by lipopolysaccharide (LPS). Injecting LPS into the lateral ventricle caused rat brain dysfunction, as evidenced by deficits of spatial learning and memory in the Morris water maze. With administration of icariin (30, 60, 120mg/kg body wt./day) for 17 consecutive days, spatial learning and memory abilities were markedly altered. Escape latency and searching distance decreased, and the expressions of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and cyclooxygenase-2 (COX-2) of brain were significantly reduced as observed by real-time RT-PCR and immunohistochemistry. This study used ibuprofen (40mg/kg body wt./day) as positive control. In conclusion, this study suggested that icariin can improve spatial learning and memory abilities in rats with brain dysfunction induced by LPS, an effect which may be due to decreased expressions of TNF-α, IL-1β and COX-2 in the hippocampus.

Resveratrol Protects Cortical Neurons against Microglia-mediated Neuroinflammation

Neuroinflammation is closely associated with the pathogenesis of neurological disorders. The hallmark of neuroinflammation is considered to be microglial activation. Therefore, inhibition of microglial activation might hold a promising therapy for neurological disorders. Resveratrol, a natural non‐flavonoid polyphenol found in grapes and red wine, has been recognized as a bioactive agent with potential benefit for health. Several lines of evidence show that resveratrol could exert neuroprotection against ischemia, seizure, and neurodegenerative diseases. However, the mechanisms underlying its beneficial neuroprotective effects are poorly defined. Here, by using rat primary cortical neuron‐glia cultures, results showed that resveratrol attenuated lipopolysaccharide (LPS)‐induced cortical neurotoxicity. Further studies revealed that microglia were responsible for resveratrol‐mediated neuroprotection. Resveratrol significantly inhibited LPS‐induced microglial activation and subsequent production of multiple pro‐inflammatory and cytotoxic factors such as tumor necrosis factor‐α, nitric oxide, and interleukin‐1β. Collectively, resveratrol produced neuroprotection against microglia‐induced neurotoxicity. Thus, resveratrol might represent a potential benefit for the treatment of inflammation‐related neurological disorders. Copyright

Sex Differences in the Circadian Variation of Cytochrome P450 Genes and Corresponding Nuclear Receptors in Mouse Liver

Sex differences and circadian variation are two major factors that affect the expression of drug-processing genes. This study aimed to examine sex differences in the circadian variation of hepatic cytochrome P450 (Cyp) genes and corresponding nuclear receptors. Adult mice were acclimated to environmentally controlled facilities for 2 wks, and livers were collected every 4 h during a 24-h period. Total RNA and protein were isolated and subjected to real-time reverse transcriptase–polymerase chain reaction (RT-PCR) and Western blot analysis. The mRNA expression of the aryl hydrocarbon receptor (AhR) and AhR-regulated Cyp1a1 and Cyp1a2 were higher in females and higher during the light phase. The mRNA expression of constitutive and rostane receptor (CAR) and CYP2B10 protein was female-predominant and higher in the dark phase. Pregnane X receptor (PXR) peaked around 18:00 h, but PXR-regulated Cyp3a11 and Cyp3a25 were higher at 10:00 h, without apparent sex dimorphism at protein levels. Peroxisome proliferator-activated receptor-α (PPARα), Cyp4a10, and Cyp4a14 were higher in females and peaked between 14:00 and 18:00 h. The mRNA levels of farnesoid X receptor (FXR), Cyp7a1, and Cyp27a1 peaked around 18:00 h and CYP7A1 protein was higher during the dark phase and higher in females. Cyp7b1(male-predominant) and Cyp2a4 (female-predominant) both showed circadian variation. Circadian variation of hepatic clock genes such as nuclear receptor Rev-erbα, cryptochrome 1 (Cry1), and brain muscle ARNT-like protein 1 (Bmal1) showed distinct patterns. Sex differences and circadian rhythmicity of Cyp genes and corresponding nuclear receptors exist in mouse liver that could impact xenobiotic metabolism and toxicity at different times of the day.

Resveratrol promotes neurotrophic factor release from astroglia

Neurotrophic factors such as glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF) are considered to contribute to the development, maintenance and survival of neurons, glia and oligodendrocytes. Astroglia are a major source of various neurotrophic factors. Thus, enhancement of astroglia-mediated neurotrophic factor release might hold promising potential for neurological diseases. Resveratrol, a natural non-flavonoid polyphenol found in grapes and red wine, has been recognized to be beneficial for health. Here, rat primary astroglia-enriched cultures were used to investigate the effects of resveratrol-mediated neurotrophic factor release and the related mechanisms. The cultures were treated with 25–100 μmmol/L resveratrol for 12–48 h. Results showed resveratrol increased BDNF and GDNF production in the culture medium. In addition, the production of BDNF in the supernatant of cultures was increased five-fold over control cultures 24 h after resveratrol treatment and then remained high 36 h later. Meanwhile, the production of GDNF was initially increased by up to four-fold 24 h after resveratrol treatment and continued to increase to six-fold at 36 h and remained at a high level till 48 h. Western blot analysis of BDNF and GDNF protein in astroglia at different time points after resveratrol treatment indicated similar increases. Furthermore, resveratrol significantly induced the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and cAMP responsive element-binding protein (CREB) in astroglia. Overall, resveratrol is effective in promoting astroglia-derived neurotrophic factor release, and this effect is mediated, at least in part, by the activation of ERK1/2 and CREB.

Realgar and realgar-containing Liu-Shen-Wan are less acutely toxic than arsenite and arsenate.

ETHNOPHARMACOLOGICAL RELEVANCE Liu-Shen-Wan (LSW) is a widely-used traditional Chinese medicine containing realgar (As(4)S(4)). AIM OF THE STUDY Realgar has been included in many traditional medicines, and is often taken as arsenite for risk assessment in realgar-containing traditional remedies. Is realgar toxicologically similar to arsenite? MATERIALS AND METHOD Mice were orally given LSW (60 and 200mg/kg; 200mg LSW contains 27 mg realgar), realgar (30 mg/kg, equivalent to 21 mg As/kg), and the equivalent As dose as sodium arsenite (NaAsO(2)), or as arsenate (Na(2)HAsO(4)). Acute toxicity and tissue As accumulation were determined 8h later. RESULTS Arsenite and arsenate increased serum alanine aminotransferase (ALT) levels, indicative of liver injury; blood urea nitrogen (BUN) was also increased by arsenite and arsenate, indicative of nephrotoxicity. No elevations of ALT and BUN were observed in LSW and realgar groups. Histopathology showed more damage in arsenite- and arsenate-treated liver and kidneys, while in realgar- and LSW- treated animals, only mild alterations were seen. Hepatic and renal As contents were dramatically increased to 6200 and 3350ng/g, respectively, after arsenite, but only increased to 260 and 180 ng/g after LSW. The expressions of arsenic-sensitive stress genes, namely metallothionein-1 and heme oxygenase-1, were increased after arsenite or arsenate by 3-10-folds, but were unaltered after LWS and realgar. CONCLUSIONS Realgar and LSW are much less toxic than arsenite and arenate. The use of total As content to evaluate the safety of realgar-containing traditional medicines is not scientifically sound.

Realgar, cinnabar and An-Gong-Niu-Huang Wan are much less chronically nephrotoxic than common arsenicals and mercurials

Realgar (As4S4) and cinnabar (HgS) are frequently included in traditional Chinese medicines and Indian Ayurvedic medicines. Both As and Hg are well known for toxic effects, and their safety is of concern. The aim of this study was to compare chronic nephrotoxicity of An-Gong-Niu-Huang Wan (AGNH), realgar and cinnabar with common arsenicals and mercurials. Mice were orally administrated with AGNH (3 g/kg, 6-fold of clinical dose), cinnabar (0.3 g/kg, amount in AGNH) and realgar (0.3 g/kg, amount in AGNH), HgCl2 (0.118 mmol/kg, 1/10 of cinnabar), MeHg (0.012 mmol/kg, 1/100 of cinnabar), NaAsO2 (As3+ 0.028 mmol/kg, 1/100 of realgar) or Na2HAsO4 (As5+ 0.056 mmol/kg, 1/50 of realgar), daily for six weeks, and nephrotoxicity was examined. Animal body weights were decreased by MeHg and HgCl2. Blood urea nitrogen and creatinine levels were elevated by MeHg. Renal pathology was severe in the MeHg and HgCl2 groups, moderate in the arsenite, arsenate and realgar groups and mild in the cinnabar and AGNH groups. Renal Hg accumulation in the MeHg and HgCl2 groups was 50–200 folds higher than the cinnabar group. Expressions of metallothionein-1 and heme oxygenase-1, biomarkers for metal toxicity, were increased 2–5 folds by arsenite, arsenate, MeHg and HgCl2, but not by realgar, cinnabar and AGNH. The chemokine and glutathione-S transferase-α4, markers for inflammation, were also increased by MeHg and HgCl2. Expressions of cell adhesion gene S100a9 and E-cadherin were altered by HgCl2, arsenite and realgar. Taken together, chemical forms of mercury and arsenic are major determinants in their disposition and toxicity.

Realgar- and cinnabar-containing An-Gong-Niu-Huang Wan (AGNH) is much less acutely toxic than sodium arsenite and mercuric chloride

An-gong-niu-huang wan (AGNH) is a famous traditional Chinese medicine used for brain trauma, hemorrhage, and coma. AGNH contains 10% realgar (As₄S₄) and 10% cinnabar (HgS). Both As and Hg are well-known for their toxic effects, and the safety of AGNH is of concern. To address this question, the acute toxicity of AGNH, realgar and cinnabar were compared to sodium arsenite (NaAsO₂) and mercuric chloride (HgCl₂). Mice were administrated orally AGNH at 1, 3 and 6g/kg. AGNH at 3g/kg contains 2.8mmol As/kg as realgar and 1.18mmol Hg/kg as cinnabar. Realgar, cinnabar, arsenite (0.28 mmol/kg, 10% of realgar) and HgCl₂ (0.256 mmol/kg, 20% of cinnabar) were orally given to mice for comparison. Blood and tissues were collected 8h later for toxicity evaluation. Serum alanine aminotransferase was increased by arsenite and blood urea nitrogen was increased by HgCl₂. Total As accumulation after arsenite in liver (100-fold) and kidney (13-fold) was much higher than that after realgar. The accumulation of Hg after HgCl₂ in liver was 400-fold higher and kidney 30-fold higher than after cinnabar. Histopathology showed moderate liver and kidney injuries after arsenite and HgCl₂, but injuries were mild or absent after AGNH, realgar, and cinnabar. The expression of metallothionein-1, a biomarker of metal exposure, was increased 4-10-fold by arsenite and HgCl₂, but was unchanged by AGNH, realgar and cinnabar. Thus, AGNH, realgar and cinnabar are much less toxic acutely than arsenite and HgCl₂. The chemical forms of As and Hg are extremely important factors in determining their disposition and toxicity.