Yi-Fang Li

Anti-Stress Effects of Carnosine on Restraint-Evoked Immunocompromise in Mice through Spleen Lymphocyte Number Maintenance

Carnosine (β-alanyl-L-histidine), a naturally occurring dipeptide, has been characterized as a putative neurotransmitter and serves as a reservoir for brain histamine, which could act on histaminergic neurons system to relieve stress-induced damages. However, understanding of the role of carnosine in stress-evoked immunocompromise is limited. In this study, results showed that when mice were subjected to restraint stress, spleen index and the number of spleen lymphocytes including Natural Killer (NK) cells were obviously decreased. Results also demonstrated that restraint stress decreased the cytotoxic activity of NK cells per spleen (LU10/spleen) while the activity of a single NK cell (LU10/106 cells) was not changed. However, oral administration of carnosine (150 and 300 mg/kg) increased spleen index and number of spleen lymphocytes (including NK cells), and elevated the cytotoxic activity of NK cells per spleen in restraint-stressed mice. These results indicated that carnosine ameliorated stress-evoked immunocompromise through spleen lymphocyte number maintenance. Carnosine was further found to reduce stress-induced elevation of plasma corticosterone level. On the other hand, results showed that carnosine and RU486 (a glucocorticoids receptor antagonist) treatment prevented the reduction in mitochondrion membrane potential and the release of mitochondrial cytochrome c into cytoplasm, increased Bcl-2/Bax mRNA ratio, as well as decreased terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL)-positive cells in spleen lymphocytes of stressed mice. The results above suggested that the maintenance of spleen lymphocyte number by carnosine was related with the inhibition of lymphocytes apoptosis caused by glucocorticoids overflow. The stimulation of lymphocyte proliferation by carnosine also contributed to the maintenance of spleen lymphocyte number in stressed mice. In view of the elevated histamine level, the anti-stress effects of carnosine on restraint-evoked immunocompromise might be via carnosine-histamine metabolic pathway. Taken together, carnosine maintained spleen lymphocyte number by inhibiting lymphocyte apoptosis and stimulating lymphocyte proliferation, thus prevented immunocompromise in restraint-stressed mice.

Pharmacological studies on the anxiolytic effect of standardized Schisandra lignans extract on restraint-stressed mice.

Fruits of Fructus Schisandrae were used as sedatives and hypnotics in traditional Chinese medicine for a long history. In this study, we investigated the effects of schisandra lignans extract (SLE) on anxiety disorder in restraint-stressed mice using light-dark (L-D) test. The influences of restraint stress on the levels of monoamines: noradrenaline (NE), dopamine (DA) and serotonin (5-HT) in cerebral cortex, as well as plasma corticosterone (CORT) were studied in mice. The HPLC fingerprint of SLE was recorded and the percentage composition of Schisandra lignans was determined as 82.63%. In L-D test, it was found out that 18h of restraint stress significantly decreased the anxiolytic parameters (explorative behaviors, e.g. number of entries, time spent) in light area indicating high state of anxiety in stressed mice. In addition, restraint stress elevated NE, DA, and 5-HT levels in cerebral cortex of anxiety mice. Plasma CORT level was also increased. Oral administration of SLE (100 and 200mg/kg/day, 8 days) emolliating the level of stress-induced anxiety by significantly increasing the anxiolytic parameters mentioned above. We also observed decreases in cerebral cortex monoamines levels, as well as plasma CORT level in stressed mice. These results suggested that SLE reversed stress-induced anxiety level, changes of cortex monoamine transmitters and plasma CORT. The anxiolytic effects of SLE might be related to its anti-stress activity by modulation of hyperactive HPA axis.

Theacrine, a Purine Alkaloid Obtained from Camellia assamica var. kucha, Attenuates Restraint Stress-Provoked Liver Damage in Mice

Theacrine (1,3,7,9-tetramethyluric acid), a purine alkaloid, has proven to be beneficial in maintaining several brain functions and is being studied for potential medicinal uses in recent years. In this study, we isolated theacrine from Camellia assamica var. kucha and investigated its protective effects on liver damage induced by restraint stress in mice. Results showed that 18 h of restraint stress could induce liver damage, with an obvious increase in levels of plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST). This finding was further confirmed by hepatic pathological examination, which showed inflammatory cell infiltration and focal necrosis of hepatocytes. However, oral administration of theacrine (10, 20, 30 mg/kg for 7 consecutive days) was found to decrease plasma ALT and AST levels, reduce hepatic mRNA levels of inflammatory mediators (IL-1β, TNF-α, IL-6, and IFN-γ), and reverse the histologic damages in stressed mice. Simultaneously, theacrine also significantly decreased the content of malondialdehyde and increased oxygen radical absorbance capacity (ORAC) level in the plasma and liver of stressed mice. These results suggested that the protective effects of theacrine on stress-induced liver damage might be correlated with its antioxidative activity. The antioxidative capacity of theacrine was further evaluated by in vitro ORAC and cellular antioxidant activity assay. The results suggested that the antioxidative capacity of theacrine was not due to the direct action on free radical clearance. Moreover, the elevated activities and gene expressions of superoxide dismutase, catalase, and glutathione peroxidase, as well as the reduced activity of xanthine oxidase by theacrine treatment in stressed mice suggested that the antioxidative activity might be due to the strengthening of the antioxidant system in vivo. On the basis of the above results, theacrine is possibly a good candidate for protecting against or treating lifestyle diseases and might contribute to the study of natural products.

A SIRT3/AMPK/autophagy network orchestrates the protective effects of trans-resveratrol in stressed peritoneal macrophages and RAW 264.7 macrophages

Resveratrol gains a great interest for its strong antioxidant properties, while the molecular mechanisms underlie the beneficial effects on psychosocial stress remain controversial. In this study, we demonstrated that resveratrol protected peritoneal macrophages and RAW 264.7 cells from stress-induced decrease in the total cell count, phagocytic capability, reactive oxygen species generation, monodansylcadaverine and mitochondrial membrane potential in stressed mice. Resveratrol promoted stress-induced autophagy in both models. Modulation of autophagy by rapamycin or 3-methyladenine regulated the protective effect of resveratrol, suggesting a role of autophagy in the protective mechanisms of resveratrol. The comparison studies revealed that distinct mechanisms were implicated in the protective effect of resveratrol and other antioxidants (vitamin C and edaravone). Resveratrol promoted autophagy via upregulating SIRT3 expression and phosphorylation of AMP-activated protein kinase (AMPK). Knockdown of SIRT3 resulted in decreased autophagy and abolished protective effect of resveratrol. SIRT1 was also involved in the protective mechanism of resveratrol, although its effect on autophagy was unnoticeable. Pharmacological manipulation of autophagy modulated the effects of resveratrol on SIRT3 and AMPK, revealing the engagement of a positive feedback loop. In sharp contrast, vitamin C and edaravone effectively protected macrophages from stress-induced cytotoxicity, accompanied by downregulated SIRT3 expression and AMPK phosphorylation, and decreased level of autophagy response. Taken together, we conclude that a SIRT3/AMPK/autophagy network orchestrates in the protective effect of resveratrol in macrophages.

Anti-inflammatory effects of anthocyanins-rich extract from bilberry (Vaccinium myrtillus L.) on croton oil-induced ear edema and Propionibacterium acnes plus LPS-induced liver damage in mice

Abstract Bilberry (Vaccinium myrtillus L.) has been known to play a protective role in human health due to its high anthocyanin content. This study investigated the anti-inflammatory effects of bilberry extract (BE, containing 42.04% anthocyanin) on Propionibacterium acnes (P. acnes) plus lipopolysaccharide (LPS) induced liver injury and croton oil-induced ear edema in mice. Results showed that BE could effectively inhibit croton oil-induced ear edema and liver inflammation provoked by P. acnes plus LPS, as reflected by the reduced plasma alanine aminotransferase and aspartate aminotransferase activities. These findings were confirmed by hepatic pathological examination. Moreover, BE administration markedly suppressed the increase of liver mRNA levels of iNOS, TNF-α, IL-1β and IL-6, and the protein levels of iNOS, TNF-α and NF-κB. In addition, liver malondialdehyde and NO contents were significantly reduced by BE treatment. These results indicated that BE has potent protective effects on acute and immunological inflammation, which might contribute to the study of the anti-inflammatory effects of natural products and healthy food.

Anti-Inflammatory Effects of a Polyphenols-Rich Extract from Tea (Camellia sinensis) Flowers in Acute and Chronic Mice Models

While beneficial health properties of tea leaves have been extensively studied, less attention is paid to the flowers of tea. In this study, the anti-inflammatory effects of hot water extract of tea (Camellia sinensis) flowers were investigated. Pharmacological studies found that administration of tea flowers extract (TFE) could effectively inhibit croton oil-induced ear edema and carrageenin-induced paw edema. Furthermore, administration of TFE also protected against Propionibacterium acnes (P. ances) plus lipopolysaccharide-(LPS-) induced liver inflammation by reversing the histologic damage and plasma alanine aminotransferase (ALT) increase. Moreover, the levels of nitric oxide (NO), tumor necrosis factor-(TNF)-α and interleukin-(IL-) 1β mRNA in mouse liver were markedly suppressed after treatment with TFE in mice with immunological liver inflammation. These results indicated that tea flowers had potent anti-inflammatory effects on acute and immunological inflammation in vivo, and may be used as a functional natural food.

Autophagy is involved in the effects of resveratrol on prevention of splenocyte apoptosis caused by oxidative stress in restrained mice

SCOPE Resveratrol, a powerful natural compound for human health, is widely reported for its immunity-related beneficial properties. However, few works have studied its effect mechanism on immunity. The present study was conducted to investigate the effects of resveratrol on splenic immunity in restraint stressed mice and the mechanism was further studied as autophagy induction. METHODS AND RESULTS Mice were administered with resveratrol for 7 days consecutively, fixed in restraint cages for 18 h, and recovered for 12 h after the last administration. Data showed that restraint led to spleen damages, including declined spleen index, decreased CD4(+) T-cell number, increased mitochondrial oxidative damage, and apoptosis of splenocytes. Resveratrol, vitamin C (antioxidant), and rapamycin (autophagy agonist) protected spleen functions. Meanwhile, rapamycin augmented the effects of resveratrol that were abolished by chloroquine (autophagy antagonists). Further studies showed that expressions of Beclin 1 and LC3β required in autophagy development were significantly upregulated by resveratrol but not by vitamin C. CONCLUSION This study demonstrated that resveratrol preserved splenic immunity of restraint stressed mice. It is meaningful to find that autophagy, apart from reactive oxygen species clearance, is included as a potential mechanism via which resveratrol ameliorated the state of oxidative stress and thus protected splenocytes in mice.

Comparing antioxidant capacity of purine alkaloids: a new, efficient trio for screening and discovering potential antioxidants in vitro and in vivo.

The most commonly applied strategies for the evaluation of antioxidant capacity are the chemical- or cell-based approaches. However, the results obtained from these methods might not reflect the antioxidant ability of test samples within organisms. In this study, we propose a combination of experiments, including oxygen radical absorbance capacity (ORAC), cellular antioxidant activity assay (CAA), and the chick embryo model, as an efficient trio to evaluate antioxidant capacity of food components. Taking purine alkaloids as example, results demonstrate that chemical and cellular method might misinterpret their true ability on antioxidation. In chick embryo model, caffeine and theacrine can significantly improve vessel density on chorioallantoic membrane and myocardial apoptosis. The mechanism can be involving multiple targets within the organism. We believe that the trio proposed can be widely utilized in screening massive number of antioxidant in a cost-effective way. It will also help discovering new antioxidants that are easily being omitted due to their relatively poor in vitro activities.

Caffeine interferes embryonic development through over-stimulating serotonergic system in chicken embryo.

The potential harmful effects of caffeine in pregnant women aroused public interests due to its possibility to jeopardize fetal development. Monoamine neurotransmitters are thought to regulate neural development processes through maternal-fetal interactions, which may have long term impact on mental and behavioral effects. The current study focuses on investigating the effects of caffeine on the monoamine neurotransmitter system using developmental chicken embryos. The ED(50) value of caffeine toxicity was 27.3 μmol/egg in chicken embryo. Administration of caffeine, with lower dosage than ED(50) (2.5, 5.0 and 10.0 μmol/egg), caused failure of neural tube closure. In addition, contents of 5-HT and its metabolite 5-HIAA were increased under dosage of 10.0 μmol/egg caffeine administration. Gene expression of TPH2 was also increased by caffeine treatment. Caffeine could result in defect of neural tube closure and induce disorder of serotonergic system development, which may increase teratogenic rate of embryos. Meanwhile, it is probably an underlying factor for inducing psychological and behavioral disorders in adult. Moreover, caffeine was found to be accumulated in the embryonic brain and not being metabolized, which may incur a magnification of adverse effects. This study may provide valuable data for further investigations on toxicology of caffeine during different stages of pregnancy.

Anti-inflammatory effects of Reduning Injection (热毒宁注射液) on lipopolysaccharide-induced acute lung injury of rats

ObjectiveTo evaluate the protective effects of Reduning Injection (热毒宁注射液, RDN), a patent Chinese medicine, on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in rats and its underlying mechanisms of action.MethodsSixty male Sprague-Dawley rats were randomly divided into 6 groups, including normal control, model, dexamethasone (DEX, 5 mg/kg), RDN-H (720 mg/kg), RDN-M (360 mg/kg) and RDN-L (180 mg/kg) groups, with 10 rats in each group. Rats were challenged with intravenous injection of LPS 1 h after intraperitoneal treatment with RDN or DEX. At 6 h after LPS challenge, lung tissues and bronchoalveolar lavage fluid (BALF) were collected, and the number of inflammatory cells was determined. The right lungs were collected for histopathologic examination, measurement of gene and protein expressions, superoxide dismutase (SOD) and myeloperoxidase (MPO) activities.ResultsIn vivo pretreatment of RDN (360, 720 mg/kg) significantly reduced the weight of wet to dry (W/D) ratio of lung, protein content in BALF, and led to remarkable attenuation of LPS-induced histopathological changes in the lungs. Meanwhile, RDN enormously decreased BALF total inflammatory cells, especially neutrophil and macrophage cell numbers. Moreover, RDN increased SOD activity, inhibited MPO activity, alleviated LPS-induced tumor neurosis factor-α (TNF-α) and inducible nitric oxide synthase (iNOS) expression in lung tissues. Furthermore, RDN (720 mg/kg) efficiently weakened nuclear factorkappa B (NF-κB) gene and protein expression.ConclusionAnti-inflammatory effects of RDN was demonstrated to be preventing pulmonary neutrophil infiltration, lowering MPO activity, TNF-α and iNOS gene expression by inhibiting NF-κB activity in LPS-induced ALI.