Specnuezhenide ameliorates ultraviolet-induced skin photoaging in mice by regulating the Sirtuin 3/8-Oxoguanine DNA glycosylase signal.

PURPOSE: Ultraviolet-induced skin photoaging was involved in DNA oxidative damage. Specnuezhenide, one of the secoiridoids extracted from Ligustri Lucidi Fructus, possesses antioxidant and anti-inflammatory effects. Whether specnuezhenide ameliorates skin photoaging remains unclear. This study aimed to investigate the effect of specnuezhenide on skin photoaging induced by ultraviolet and explore the underlying mechanism. METHODS: Mice were employed to treat with ultraviolet to induce skin photoaging, then administrated 10 and 20 mg/kg of specnuezhenide. Histological analysis, protein expression, network pharmacology, and autodock analysis were conducted. RESULTS: Specnuezhenide ameliorated ultraviolet-induced skin photoaging in mice via the increase in collagen contents, and decrease in epidermal thickness, malondialdehyde content, and ß-galactosidase expression in the skin. Specnuezhenide reduced cutaneous apoptosis and inflammation in mice with skin photoaging. In addition, network pharmacology data indicated that specnuezhenide possessed potential targets on the NOD-like receptor signaling pathway. Validation experiment found that specnuezhenide inhibited the expression of NOD-like receptor family pyrin domain-containing 3, gasdermin D-C1, and Caspase 1. Furthermore, the expression of 8-Oxoguanine DNA glycosylase (OGG1), sirtuin 3 (SIRT3), and superoxide dismutase 2 was increased in specnuezhenide-treated mice with photoaging. CONCLUSION: Specnuezhenide protected against ultraviolet-induced skin photoaging in mice via a probable activation of SIRT3/OGG1 signal.

Restraint stress promotes nonalcoholic steatohepatitis by regulating the farnesoid X receptor/NLRP3 signaling pathway.

Psychological stress promotes nonalcoholic steatohepatitis (NASH) development. However, the pathogenesis of psychological stress-induced NASH remains unclear. This study aims to explore the underlying mechanism of restraint stress-induced NASH, which mimics psychological stress, and to discover potential NASH candidates. Methionine choline deficient diet- and high fat diet-induced hepatosteatotic mice are subjected to restraint stress to induce NASH. The mice are administrated with Xiaoyaosan granules, NOD-like receptor family pyrin domain containing 3 (NLRP3) inhibitors, farnesoid X receptor (FXR) agonists, or macrophage scavengers. Pathological changes and NLRP3 signaling in the liver are determined. These results demonstrate that restraint stress promotes hepatic inflammation and fibrosis in hepatosteatotic mice. Restraint stress increases the expressions of NLRP3, Caspase-1, Gasdermin D, interleukin-1ß, cholesterol 7α-hydroxylase, and sterol 12α-hydroxylase and decreases the expression of FXR in NASH mice. Xiaoyaosan granules reverse hepatic inflammation and fibrosis and target FXR and NLRP3 signals. In addition, inhibition of NLRP3 reduces the NLRP3 inflammasome and liver damage in mice with restraint stress-induced NASH. Elimination of macrophages and activation of FXR also attenuate inflammation and fibrosis by inhibiting NLRP3 signaling. However, NLRP3 inhibitors or macrophage scavengers fail to affect the expression of FXR. In conclusion, restraint stress promotes NASH-related inflammation and fibrosis by regulating the FXR/NLRP3 signaling pathway. Xiaoyaosan granules, NLRP3 inhibitors, FXR agonists, and macrophage scavengers are potential candidates for the treatment of psychological stress-related NASH.

A Novel Polysaccharide Separated from Panax Notoginseng Residue Ameliorates Restraint Stress- and Lipopolysaccharide-induced Enteritis in Mice.

Polysaccharides are rich in Panax notoginseng residue after extraction. This study aims to explore the structural characteristics of PNP-20, which is a homogeneous polysaccharide, separated from P. notoginseng residue by fractional precipitation and evaluate the anti-enteritis effect of PNP-20. The structure of PNP-20 was determined by spectroscopic analyses. A mouse model with enteritis induced by restraint stress (RS) and lipopolysaccharide (LPS) was used to evaluate the pharmacological effect of PNP-20. The results indicated that PNP-20 consisted of glucose (Glc), galactose (Gal), Mannose (Man) and Rhamnose (Rha). PNP-20 was composed of Glcp-(1→, →4)-α-Glcp-(1→, →4)-α-Galp-(1→, →4,6)-α-Glcp-(1→, →4)-Manp-(1→ and →3)-Rhap-(1→, and contained two backbone fragments of →4)-α-Glcp-(1→4)- α-Glcp-(1→ and →4)-α-Galp-(1→4)-α-Glcp-(1→. PNP-20 reduced intestinal injury and inflammatory cell infiltration in RS- and LPS-induced enteritis in mice. PNP-20 decreased the expression of intestinal tumor necrosis factor-α, NOD-like receptor family pyrin domain containing 3, and nuclear factor-κB and increased the expression of intestinal superoxide dismutase 2. In conclusion, PNP-20 may be a promising material basis of P. Notoginseng for the treatment of inflammatory bowel disease.

New insights into the effects of caffeine on adult hippocampal neurogenesis in stressed mice: Inhibition of CORT-induced microglia activation.

Chronic stress-evoked depression has been implied to associate with the decline of adult hippocampal neurogenesis. Caffeine has been known to combat stress-evoked depression. Herein, we aim to investigate whether the protective effect of caffeine on depression is related with improving adult hippocampus neurogenesis and explore the mechanisms. Mouse chronic water immersion restraint stress (CWIRS) model, corticosterone (CORT)-established cell stress model, a coculture system containing CORT-treated BV-2 cells and hippocampal neural stem cells (NSCs) were utilized. Results showed that CWIRS caused obvious depressive-like disorders, abnormal 5-HT signaling, and elevated-plasma CORT levels. Notably, microglia activation-evoked brain inflammation and inhibited neurogenesis were also observed in the hippocampus of stressed mice. In comparison, intragastric administration of caffeine (10 and 20 mg/kg, 28 days) significantly reverted CWIRS-induced depressive behaviors, neurogenesis recession and microglia activation in the hippocampus. Further evidences from both in vivo and in vitro mechanistic experiments demonstrated that caffeine treatment significantly suppressed microglia activation via the A2AR/MEK/ERK/NF-κB signaling pathway. The results suggested that CORT-induced microglia activation contributes to stress-mediated neurogenesis recession. The antidepression effect of caffeine was associated with unlocking microglia activation-induced neurogenesis inhibition.

[Two new sucrose cinnamates from Polygonum lapathifolium var. salicifolium].

Two new sucrose cinnamates(1 and 2) along with nine known compounds(3-11) were isolated from ethanol extract of Polygonum lapathifolium var. salicifolium by silica gel column chromatography, ODS column chromatography and semi-preparative HPLC. Their structures were elucidated by extensive spectroscopic methods including 1 D-and 2 D-NMR experiments, as well as HR-ESI-MS analysis. Eleven compounds(7 sucrose cinnamates, 3 phenylpropanoids and 1 lactone) were obtained and their structures were identified as(1,3-O-di-p-coumaroyl)-ß-D-fructofuranosyl-(2→1)-α-D-glucopyranoside(1),(1,3-O-di-p-coumaroyl)-ß-D-fructofuranosyl-(2→1)-(6-O-acetyl)-α-D-glucopyranoside(2),(3-O-feruloyl)-ß-D-fructofuranosyl-(2→1)-(6-O-p-coumaroyl)-α-D-glucopyranoside(3), hydropiperoside(4), vanicoside C(5),(1,3-O-di-p-coumaroyl)-ß-D-fructofuranosyl-(2→1)-(6-O-feruloyl)-α-D-glucopyranoside(6), vanicoside B(7),trans-p-hydroxycinnamic acid methyl ester(8), trans-p-hydroxycinnamic acid ethyl ester(9), methyl ferulate(10) and dimethoxydimethylphthalide(11), respectively. Compounds 1 and 2 were two new sucrose cinnamates, and compounds 1-11 were isolated from this plant for the first time. The antioxidant activities of the isolated compounds 1-9 were investigated by an oxygen radical absorbance capacity(ORAC) assay, and all nine compounds were found to show strong antioxidant activities. Among them, compound 6(10 µmol·L~(-1)) was the supreme one in antioxidant activities, with its ORAC value equivalent to(1.60±0.05) times of 50 µmol·L~(-1) Trolox.

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.

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-1ß 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.

Phloridzin improves lipoprotein lipase activity in stress-loaded mice via AMPK phosphorylation.

Long-term stress exposure can lead to disturbed homeostasis and cause many life-style diseases. Phloridzin possesses various bioactivities, but the understanding of the effects of phloridzin on stress-related lipid metabolism disorder is limited. Our results demonstrate that phloridzin improved plasma lipoprotein lipase (LPL) activity and triglyceride metabolism in restrained mice. A decrease of angiopoietin-like protein 4 (ANGPTL4) mRNA expression and an increase of AMP-activated protein kinase (AMPK) phosphorylation were observed after phloridzin treatment. After inhibiting AMPK phosphorylation, the effects of phloridzin on the amelioration of plasma LPL activity and suppression of ANGPTL4 expression were blocked. In addition, cardiac AMPK phosphorylation, plasma LPL activity and ANGPTL4 expression were also affected by phloridzin, even if the glucocorticoid receptor was blocked. Taken together, the down-regulation of ANGPTL4 expression by phloridzin was probably via a direct activation of AMPK pathway. This discovery can provide a biochemical and nutritional basis for the use of phloridzin-containing food and beverage in daily life.

Erzhiwan Ameliorates Restraint Stress- and Monobenzone-Induced Depigmentation in Mice by Inhibiting Macrophage Migration Inhibitory Factor and 8-Hydroxy-2-Deoxyguanosine.

Purpose: Vitiligo is an autoimmune disease that results in the loss of epidermal melanocytes. The treatments for patients with vitiligo remain lacking. Erzhiwan (EZW), a traditional Chinese Medicine composed of Ligustri Lucidi Fructus and Ecliptae Herba, was used to ameliorate depigmentation since ancient China. This study aims to investigate the effect of EZW on vitiligo-related depigmentation. Methods: A vitiligo-related depigmentation mouse model was induced by monobenzone and restraint stress. The experimental depigmentation mice were treated with EZW. Histological observation of skin was conducted. Cutaneous oxidative damage and inflammation were determined. A network pharmacology analysis was carried out. Results: EZW reduced depigmentation score (p<0.01), cutaneous inflammatory infiltration (p<0.01), and CD8α-positive expression (p<0.01), and increased cutaneous melanin content in experimental depigmentation mice. EZW reduced stress reaction in experimental depigmentation mice (p<0.01). EZW inhibited 8-hydroxy-2-deoxyguanosine (8-OHdG)-related DNA oxidative damage in the skin (p<0.05, p<0.01). In addition, EZW reduced cutaneous macrophage migration inhibitory factor (MIF)-CD74-NF-κB signaling (p<0.01). The network pharmacology analysis demonstrated that EZW regulated necroptosis, apoptosis, and FoxO signaling pathways in vitiligo. An in vitro experiment showed that the main ingredient of EZW, specnuezhenide, protected against monobenzone and MIF-induced cell death in HaCaT cells (p<0.01). Conclusion: EZW ameliorates restraint stress- and monobenzone-induced depigmentation via the inhibition of MIF and 8-OHdG signaling. The findings provide a data basis of an utilization of EZW in vitiligo.

Restraint stress promotes monobenzone-induced depigmentation in mice via the activation of glucocorticoid receptor/macrophage migration inhibitory factor signaling pathway.

Psychological stress triggers onset and development of vitiligo in humans. However, the mechanism of psychological stress on vitiligo remains unclear. The study aims to investigate whether psychological stress promotes vitiligo and explore the underlying mechanism. A depigmentation mouse model induced by applying a skin-bleaching reagent monobenzone to dorsal skin and an in vitro HaCaT keratinocyte death model induced by monobenzone were employed to explore the effect of restraint stress, which mimics psychological stress, on depigmentation. The results indicated that restraint stress promoted vitiligo-related depigmentation, vacuolisation, spongiosis, CD8+ T lymphocyte infiltration, and loss of melanocytes in the skin. Restraint stress activated cutaneous NLR family containing pyrin domain protein 3 (NLRP3) inflammasome. In addition, restraint stress aggravated anxiety-like behaviors and increased levels of macrophage migration inhibitory factor (MIF) and corticosterone in the circulation, accompanied with decreasing the expression of cutaneous 8-oxoguanine DNA glycosylase (OGG1) in depigmentation mice. In vitro experiments demonstrated that activation of glucocorticoid receptor (GR) by cortisol upregulated NLRP3 expression dependent on MIF, and directly decreased the transcription of OGG1. Blockade of MIF reversed the NLRP3 signal in restraint stress-induced depigmentation mice. In conclusion, restraint stress promotes vitiligo-related depigmentation in mice via the activation of GR/MIF signaling pathway. The findings provide a theoretical basis for prevention and treatments of vitiligo with therapies of targeting GR, MIF, and OGG1.

Theacrine ameliorates experimental liver fibrosis in rats by lowering cholesterol storage via activation of the Sirtuin 3-farnesoid X receptor signaling pathway.

Formulations against liver fibrosis (LF) mitigate the progression of hepatitis to cirrhosis. However, notable toxicity of the currently available anti-LF drugs limits their long-term use. In the study, we aimed to investigate the anti-LF effects of theacrine, a purine alkaloid without obvious toxicity, on high-fat diet-, alcohol-, and carbon tetrachloride-induced LF in rats. The results indicated that 10 and 20 mg/kg of theacrine ameliorated hepatic fibrosis, steatosis, and inflammation in LF rats. Mechanistically, theacrine reduced hepatic stellate cell (HSC)-related α-smooth muscle actin expression, and decreased cholesterol accumulation, followed by decreased expression of transforming growth factor-ß1, interleukin (IL)-1ß, and tumor necrosis factor (TNF)-α. In addition, theacrine upregulated the phosphorylation of AMP-activated protein kinase, accompanied by decreased expression of ß-catenin and stearoyl-CoA desaturase 1, and increased the expression of sirtuin 3 (SIRT3). Further investigation revealed that the theacrine-mediated decrease in cholesterol was independent of cholesterol synthesis or low-density lipoprotein (LDL) uptake in hyperlipidemia mice. However, theacrine activated farnesoid X receptor (FXR), a ß-catenin conjugated protein, accompanied with decreased expression of cholesterol 7α-hydroxylase and sterol 12α-hydroxylase. In conclusion, theacrine alleviated experimental LF in rats by lowering cholesterol storage and decreasing cholesterol-related HSC activation. A plausible mechanism of theacrine on cholesterol metabolism may involve activation of SIRT3-FXR signaling pathway followed by decreased intestinal cholesterol absorption.

Biochanin A ameliorated oleate-induced steatosis in HepG2 cells by activating the SIRT3/AMPK/ULK-1 signaling pathway.

Biochanin A (Bio-A), an isoflavone abundant in chickpeas, possesses hypoglycemic, hypolipidemic, and anti-inflammatory effects. However, whether Bio-A has antihepatosteatosis effect remains unclear. This study aimed to evaluate the antihepatosteatosis effect of Bio-A on oleate (OA)-treated hepatocytes, and explore the underlying mechanism. When incubated with OA for 24 h, HepG2 cells were treated with various concentrations of Bio-A for 24 h to obtain an optimal antihepatosteatosis dose. HepG2 cells were treated with the AMP-activated protein kinase (AMPK) inhibitor Compound C, or the sirtuin-3 (SIRT3) inhibitor 3-TYP, and incubated with 50 µM Bio-A. The results indicated that 12.6% of lipid content, particularly 11.0% of triglyceride content, and the expression of adipocyte differentiation-related protein were significantly decreased in Bio-A-treated hepatosteatosis cells, followed by an increase in the expression of Beclin 1, phosphorylation of Unc-51-like kinase 1 (ULK-1), the microtubule-associated protein 1 light chain 3 (LC3)-II/LC3-I ratio, and a decrease in expression of p62. The results indicated that Bio-A upregulated autophagosome formation and autophagy flux. In addition, Bio-A increased SIRT3 expression and AMPK phosphorylation in OA-treated HepG2 cells. Blockade of AMPK and SIRT3 blocked the antihepatosteatosis effect and ULK-1 activation by Bio-A. AMPK inhibition did not eliminate the activation of SIRT3 by Bio-A. AutoDock analysis demonstrated that interaction might exist between Bio-A and SIRT3. In conclusion, Bio-A reduced fat accumulation in OA-treated HepG2 cells by activating SIRT3/AMPK/ULK-1-mediated autophagy. The findings provide a theoretical basis for the effect of Bio-A on hepatic steatosis-related diseases. PRACTICAL APPLICATIONS: This study highlights the antihepatosteatosis effects of biochanin A (Bio-A) on oleate (OA)-treated hepatocytes. Bio-A, one of the isoflavones in Cicer arietinum Linn., possesses multiple bioactivities such as antiobesity, anti-inflammation, and hypoglycemic and hypolipidemic effects. This study provides a new application of Bio-A to treat hepatic steatosis, and revealed the underlying mechanism of Bio-A involved in the activation of the SIRT3/AMPK/ULK-1-mediated autophagy. The findings provide a theoretical basis for the application of Bio-A to hepatic steatosis-related diseases.

Theacrine, a Potent Antidepressant Purine Alkaloid from a Special Chinese Tea, Promotes Adult Hippocampal Neurogenesis in Stressed Mice.

Daily intake of tea has been known to relate to a low risk of depression. In this study, we report that a special variety of tea in China, Camellia assamica var. kucha (kucha), possesses antidepressant effects but with less adverse effects as compared to traditional tea Camellia sinensis. This action of kucha is related to its high amount of theacrine, a purine alkaloid structurally similar to caffeine. We investigated the antidepressant-like effects and mechanisms of theacrine in chronic water immersion restraint stress and chronic unpredictable mild stress mice models. PC12 cells and primary hippocampal neural stem cells were treated with stress hormone corticosterone (CORT) to reveal the potential antidepression mechanism of theacrine from the perspective of adult hippocampus neurogenesis. Results of behavioral and neurotransmitter analysis showed that intragastric administration of theacrine significantly counteracted chronic stress-induced depression-like disorders and abnormal 5-hydroxytryptamine (5-HT) metabolism with less central excitability. Further investigation from both in vivo and in vitro experiments indicated that the antidepressant mechanism of theacrine was associated with promoting adult hippocampal neurogenesis, via the modulation of the phosphodiesterase-4 (PDE4)/cyclic adenosine monophosphate (cAMP)/cAMP response-element binding (CREB)/brain-derived neurotrophic factor (BDNF)/tropomyosin-related kinase B (TrkB) pathway. Collectively, our findings could promote the prevalence of kucha as a common beverage with uses for health care and contribute to the development of theacrine as a potential novel antidepressant medicine.

Theacrine protects against nonalcoholic fatty liver disease by regulating acylcarnitine metabolism.

OBJECTIVE: Acylcarnitine metabolism disorder contributes significantly to the pathogenesis of nonalcoholic fatty liver disease (NAFLD). There are, however, few ideal medications for NAFLD, which work by targeting acylcarnitine metabolism. The aim of this study was to investigate the protective effects of theacrine, a rare purine alkaloid isolated from Camellia assamica var. kucha, against acylcarnitine metabolism disorder in NAFLD. METHODS: The pharmacological activities of theacrine were studied using high-fat diet (HFD)-fed ApoE-/- and C57BL/6J mice models. Oleate-treated HepG2 and L-02 cells were used to investigate the molecular mechanism of theacrine on acylcarnitine metabolism. The target of theacrine was confirmed in vitro as the blockade of sirtuin 3 (SIRT3) and protein kinase A. RESULTS: Theacrine inhibits hepatic steatosis and liver inflammation and improves energy expenditure in HFD-fed mice. Theacrine ameliorates acylcarnitine metabolism disorder in HFD-fed mice and oleate-treated hepatocytes by improving fatty acid oxidation. The underlying mechanism involves theacrine's activation of the mitochondrial deacetylase SIRT3 and consequently, the increased activity of long-chain acyl coenzyme A dehydrogenase (LCAD) through deacetylation. CONCLUSION: Theacrine promotes acylcarnitine metabolism in NAFLD through the SIRT3/LCAD signaling pathway. The target of theacrine's activities on NAFLD is identified as SIRT3.

Immune regulation effect of lienal polypeptides extract in Lewis lung carcinoma-bearing mice treated with cyclophosphamide.

Polypeptides extracted from animal immune organs have been proved to exert immunomodulatory activities in previous reports. However, relative experimental data regarding the influence of a polypeptide mixture extracted from healthy calf spleen (lienal polypeptide [LP]) on the immune function in tumor therapy are limited, and the components in LP remain unclear. In the present study, the immune regulatory effect of LP was investigated in normal mice and Lewis lung carcinoma (LLC)-bearing mice treated with cyclophosphamide (CTX). The components of LP were identified by liquid chromatography-electrospray ionization-coupled with tandem mass spectrometry (LC-MS/MS) analysis and bioinformatic analysis. In LLC-bearing mice, LP showed a synergic antitumor effect with CTX, whereas LP alone did not present direct antitumor activity. Further, LP was found to enhance immune organ indexes, splenocyte number, and T lymphocyte subsets in normal mice and LLC-bearing mice treated with CTX. The decline of white blood cell and platelet counts, splenocyte proliferation activity, and peritoneal macrophage phagocytic function caused by CTX were also significantly suppressed by LP treatment in LLC-bearing mice. Notably, LP treatment significantly decreased the expression of phagocytosis-related proteins including CD47/signal regulatory protein α/Src homology phosphatase-1 in the tumor tissue of LLC-bearing mice treated with CTX. LC-MS/MS-based peptidomics unraveled the main polypeptides in LP with a length from 8 to 25 amino acids. Bioinformatics analysis further confirmed the possibility of LP to regulate immunity, especially in phagocytosis-related pathway. Our above findings indicated that LP can relieve the immunosuppression induced by chemotherapy and is a beneficial supplement in cancer therapy. Impact statement The immunomodulatory activities of polypeptides extracted from animal immune organs have incurred people's interests since a long time ago. In this study, we investigated the immune regulation effects of a polypeptide mixture extracted from health calf spleen (lienal polypeptide [LP]) in Lewis lung carcinoma-bearing mice treated with cyclophosphamide (CTX). Liquid chromatography-electrospray ionization-coupled with tandem mass spectrometry-based peptidomics and bioinformatics analysis unraveled the main polypeptides in LP and further confirmed that LP is mainly associated with immune regulating pathway, especially in tumor cell phagocytosis-related pathway. Our study for the first time revealed that polypeptides from spleen can relieve the immunosuppression induced by CTX and is a beneficial supplement in cancer therapy.

Carnosine attenuates cyclophosphamide-induced bone marrow suppression by reducing oxidative DNA damage.

Oxidative DNA damage in bone marrow cells is the main side effect of chemotherapy drugs including cyclophosphamide (CTX). However, not all antioxidants are effective in inhibiting oxidative DNA damage. In this study, we report the beneficial effect of carnosine (ß-alanyl-l-histidine), a special antioxidant with acrolein-sequestering ability, on CTX-induced bone marrow cell suppression. Our results show that carnosine treatment (100 and 200mg/kg, i.p.) significantly inhibited the generation of reactive oxygen species (ROS) and 8-hydroxy-2'-deoxyguanosine (8-oxo-dG), and decreased chromosomal abnormalities in the bone marrow cells of mice treated with CTX (20mg/kg, i.v., 24h). Furthermore, carnosine evidently mitigated CTX-induced G2/M arrest in murine bone marrow cells, accompanied by reduced ratios of p-Chk1/Chk1 and p-p53/p53 as well as decreased p21 expression. In addition, cell apoptosis caused by CTX was also suppressed by carnosine treatment, as assessed by decreased TUNEL-positive cell counts, down-regulated expressions of Bax and Cyt c, and reduced ratios of cleaved Caspase-3/Caspase-3. These results together suggest that carnosine can protect murine bone marrow cells from CTX-induced DNA damage via its antioxidant activity.

Caffeine Protects Skin from Oxidative Stress-Induced Senescence through the Activation of Autophagy.

Skin cells are vulnerable to oxidative stress-induced senescence, which may lead to abnormal aging or aging-related disorders. Therefore, strategies that can ameliorate oxidative stress-induced senescence are expected to protect skin from damage, holding the promise of treating skin diseases in the clinic. This study aims to investigate whether caffeine, a well-known purine alkaloid, is able to prevent skin from oxidative stress-induced senescence, and to explore the underlying molecular mechanisms. Methods: A free radical inducer 2,2'-Azobis (2-amidinopropane) dihydrochloride (AAPH) was used to induce oxidative stress and cellular senescence in both transformed skin cells and in normal human epidermal keratinocytes (NHEKs). Ultraviolet (UV) irradiation was established as the in vivo oxidative stress model in mouse skin tissues. Cellular senescence was determined by SA ß-galactosidase staining, immunofluorescence and western blotting. Activation of autophagy was confirmed by western blotting, immunofluorescence, and transmission electron microscopy. Reactive oxygen species (ROS) detection by commercial kits, gene knockdown by RNA interference (RNAi) and receptor activation/inactivation by agonist/antagonist treatment were applied in mechanistic experiments. Results: We report that AAPH induced senescence in both transformed skin cells and in NHEKs. Similarly, UV irradiation induced senescence in mouse skin tissues. Remarkably, low dose of caffeine (<10 µM) suppressed cellular senescence and skin damage induced by AAPH or UV. Mechanistically, caffeine facilitated the elimination of ROS by activating autophagy. Using a combination of RNAi and chemical treatment, we demonstrate that caffeine activates autophagy through a series of sequential events, starting from the inhibition of its primary cellular target adenosine A2a receptor (A2AR) to an increase in the protein level of Sirtuin 3 (SIRT3) and to the activation of 5' adenosine monophosphate-activated protein kinase (AMPK). Oral administration of caffeine increased the protein level of SIRT3, induced autophagy, and reduced senescence and tissue damage in UV-irradiated mouse skin. On the other hand, co-administration with autophagy inhibitors attenuated the protective effect of caffeine on UV-induced skin damage in mice. Conclusion: The results reveal that caffeine protects skin from oxidative stress-induced senescence through activating the A2AR/SIRT3/AMPK-mediated autophagy. Our study not only demonstrated the beneficial effect of caffeine using both in vitro and in vivo models, but also systematically investigated the underlying molecular mechanisms. These discoveries implicate the potential of caffeine in the protection of skin disease.

Isolation and identification of polyphenols from Marsilea quadrifolia with antioxidant properties in vitro and in vivo.

Marsilea quadrifolia is an edible aquatic medicinal plant used as a traditional health food in Asia. Four new polyphenols including kaempferol 3-O-(2″-O-E-caffeoyl)-ß-d-glucopyranoside (1), kaempferol 3-O-(3″-O-E-caffeoyl)-α-l-arabinopyranoside (3), 4-methy-3'-hydroxypsilotinin (4) and (±)-(E)-4b-methoxy-3b,5b-dihydroxyscirpusin A (18) together with 14 known ones (2, 5-17) were isolated from the ethanol extract of M. quadrifolia. Structures of the new compounds were elucidated by extensive spectroscopic analyses. In DPPH and oxygen radical absorbance capacity antioxidant assays, some compounds showed stronger antioxidant activities and quercetin (9) was the most potent antioxidant in both assays. In a restraint-induced oxidative stress model in mice, quercetin significantly attenuated the increase in plasma ALT and AST levels as well as liver MDA content of restrained mice. Liver SOD activity was also significantly increased by quercetin, indicating a significant in vivo antioxidant activity. As a rich source of polyphenols with strong antioxidant activities, M. quadrifolia may be developed to a product for relieving oxidative stress.

Caffeine ameliorates high energy diet-induced hepatic steatosis: sirtuin 3 acts as a bridge in the lipid metabolism pathway.

The beneficial effect of caffeine-containing food on non-alcoholic fatty liver disease (NAFLD) has been widely reported. The aim of this study was to explore the effect of caffeine on hepatic steatosis. C57BL/6 mice were randomly assigned to a normal diet or a high energy diet (HED). Caffeine was given to HED mice by oral gavage. Body weights, lipids in the liver and liver damage were measured. Meanwhile, cAMP, SIRT3 or AMPK inhibitors were treated respectively before incubation with caffeine in oleate-treated HepG2 cells. SIRT3 was further silenced by siRNA to confirm the results. Caffeine significantly decreased the mass of fat tissues, lipids, ALT and AST levels in the liver of HED-treated mice. Caffeine increased the transformation of ADP to ATP and activated the cAMP/CREB/SIRT3/AMPK/ACC pathway in the liver. Nile red staining demonstrated that suppression of cAMP, SIRT3 or AMPK in oleate-treated HepG2 cells counteracted the effect of caffeine. Moreover, knocking down SIRT3 could down-regulate AMPK and ACC phosphorylation by caffeine. These results demonstrate that caffeine could improve HED-induced hepatic steatosis by promoting lipid metabolism via the cAMP/CREB/SIRT3/AMPK/ACC pathway. SIRT3 functioned as a molecular bridge connecting caffeine and lipid metabolism.

Prognostic indicators of patients with acute kidney injury in intensive care unit.

BACKGROUND: Acute kidney injury (AKI) is associated with a high mortality. This study was undertaken to detect the factors associated with the prognosis of AKI. METHODS: We retrospectively reviewed 98 patients with AKI treated from March 2008 to August 2009 at this hospital. In these patients, 60 were male and 38 female. Their age ranged from 19 to 89 years (mean 52.4±16.1 years). The excluded patients were those who died within 24 hours after admission to ICU or those who had a history of chronic kidney disease or incomplete data. After 60 days of treatment, the patients were divided into a survival group and a death group. Clinical data including gender, age, history of chronic diseases, the worst laboratory values within 24 hours after diagnosis (values of routine blood tests, blood gas analysis, liver and renal function, levels of serum cystatin C, and blood electrolytes) were analyzed. Acute physiology, chronic health evaluation (APACHE) II scores and 60-day mortality were calculated. Univariate analysis was performed to find variables relevant to prognosis, odds ratio (OR) and 95% confidence interval (CI). Multiple-factor analysis with logistic regression analysis was made to analyze the correlation between risk factors and mortality. RESULTS: The 60-day mortality was 34.7% (34/98). The APACHE II score of the death group was higher than that of the survival group (17.4±4.3 vs. 14.2±4.8, P<0.05). The mortality of the patients with a high level of cystatin C>1.3 mg/L was higher than that of the patients with a low level of cystatin C (<1.3 mg/L) (50% vs. 20%, P<0.05). The univariate analysis indicated that organ failures≥2, oliguria, APACHE II>15 scores, cystatin C>1.3 mg/L, cystatin C>1.3 mg/L+APACHE II>15 scores were the risk factors of AKI. Logistic regression analysis, however, showed that organ failures≥2, oliguria, cystatin C>1.3 mg/L +APACHE II>15 scores were the independent risk factors of AKI. CONCLUSION: Cystatin C>1.3 mg/L+APACHE II>15 scores is useful in predicting adverse clinical outcomes in patients with AKI.