Aryl hydrocarbon receptor engagement during redox alteration determines the fate of CD4+ T cells in C57BL/6 mice.

The preservation of the redox homeostasis is critical for cell survival and functionality. Redox imbalance is an essential inducer of several pathological states. CD4+ /helper T cells are highly dependent on the redox state of their surrounding milieu. The potential of the aryl hydrocarbon receptor (AhR) engagement in controlling CD4+ T-cell fate during redox alteration is still challenging. C57BL/6 mice were treated with AhR agonist 6-formylindolo[3,2-b]carbazole (FICZ), AhR antagonist CH223191, an inhibitor of glutathione biosynthesis buthionine sulfoximine (BSO), and the antioxidant N-acetylcysteine (NAC) alone or in combination. Six days later, splenocytes were evaluated for the expression of the redox-related genes and the possible changes in T-cell subsets. FICZ like BSO significantly elevated the expression of HMOX1, GCLC, and GCLM genes but it failed to increase the expression of the Nrf2 gene. Moreover, FICZ + BSO increased while FICZ + CH223191 or NAC decreased the expression of these genes. FICZ also significantly increased Th1 cell numbers but decreased Tregs in a dose-dependent manner. Furthermore, a high dose of FICZ + CH223191 + NAC significantly enhanced Th1, Th17, and Treg cells but its low dose in such a situation increased Th2 and Th17 while decreased Treg cells. AhR engagement during redox alteration can determine the fate of CD4 + T cells, so, AhR agonists or antagonists might be useful in assessing immune responses. However, these results need further verifications in vitro and in animal models of various diseases.

Palmitate reduces starvation-induced ER stress by inhibiting ER-phagy in hypothalamic cells.

Palmitate is a saturated fatty acid that is well known to induce endoplasmic reticulum (ER) stress and autophagy. A high-fat diet increases the palmitate level in the hypothalamus, the main region of the brain regulating energy metabolism. Interestingly, hypothalamic palmitate level is also increased under starvation, urging the study to distinguish the effects of elevated hypothalamic palmitate level under different nutrient conditions. Herein, we show that ER-phagy (ER-targeted selective autophagy) is required for progress of ER stress and that palmitate decreases ER stress by inhibiting ER-phagy in hypothalamic cells under starvation. Palmitate inhibited starvation-induced ER-phagy by increasing the level of B-cell lymphoma 2 (Bcl-2) protein, which inhibits autophagy initiation. These findings suggest that, unlike the induction of ER stress under nutrient-rich conditions, palmitate protects hypothalamic cells from starvation-induced stress by inhibiting ER-phagy.

Assessment of the anti-rheumatoid arthritis activity of Gastrodia elata (tian-ma) and Radix aconitic lateralis preparata (fu-zi) via network pharmacology and untargeted metabolomics analyses.

AIM: Gastrodia elata and Radix aconiti lateralis preparrata are respectively named as Tian-Ma and Fu-Zi (TF) in Chinese. We explored the active components against rheumatoid arthritis (RA) from an extensively used couplet of Chinese herbs, Gastrodia elata and Radix aconiti lateralis preparata (TF) via untargeted metabolomics and network pharmacological approaches. METHODS: Water extracts of TF were mixed at ratios 1:1, 3:2 and 2:3 (w/w). Ultra-performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) was then utilized as metabolomics screening. Human Metabolome (http://www.hmdb.ca/) and Lipidmaps (http://www.lipidmaps.org/) databases were used to annotate detected compounds. Further identification of vital genes and important pathways associated with the anti-RA properties of the TF preparations was done via network pharmacology, and verified by real-time quantitative polymerase chain reaction (RT-qPCR). RESULTS: Four key compounds involved in unsaturated fatty acid biosynthesis and isoflavonoid biosynthesis were identified through metabolomics analyses. Three key components of TF associated with anti-RA activity were linoleic acid, daidzein, and daidzin. Results of RT-qPCR revealed that all 3 tested TF couplets (1:1, 3:2, and 2:3) markedly suppressed the transcription of PTGS2. These results were consistent with our network pharmacological predictions. CONCLUSIONS: The anti-RA properties of Tian-Ma and Fu-Zi are associated with the inhibition of arachidonic acid metabolism pathway.

SNP rs17079281 decreases lung cancer risk through creating an YY1-binding site to suppress DCBLD1 expression.

Genome-wide association studies (GWAS) have identified numerous genetic variants that are associated with lung cancer risk, but the biological mechanisms underlying these associations remain largely unknown. Here we investigated the functional relevance of a genetic region in 6q22.2 which was identified to be associated with lung cancer risk in our previous GWAS. We performed linkage disequilibrium (LD) analysis and bioinformatic prediction to screen functional SNPs linked to a tagSNP in 6q22.2 loci, followed by two case-control studies and a meta-analysis with 4403 cases and 5336 controls to identify if these functional SNPs were associated with lung cancer risk. A novel SNP rs17079281 in the DCBLD1 promoter was identified to be associated with lung cancer risk in Chinese populations. Compared with those with C allele, patients with T allele had lower risk of adenocarcinoma (adjusted OR = 0.86; 95% CI: 0.80-0.92), but not squamous cell carcinoma (adjusted OR = 0.99; 95% CI: 0.91-1.10), and patients with the C/T or T/T genotype had lower levels of DCBLD1 expression than those with C/C genotype in lung adenocarcinoma tissues. We performed functional assays to characterize its biological relevance. The results showed that the T allele of rs17079281 had higher binding affinity to transcription factor YY1 than the C allele, which suppressed DCBLD1 expression. DCBLD1 behaved like an oncogene, promoting tumor growth by influencing cell cycle progression. These findings suggest that the functional variant rs17079281C>T decreased lung adenocarcinoma risk by creating an YY1-binding site to suppress DCBLD1 expression, which may serve as a biomarker for assessing lung cancer susceptibility.

Antrodin A from mycelium of Antrodia camphorata alleviates acute alcoholic liver injury and modulates intestinal flora dysbiosis in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Antrodia camphorata (A. camphorata) is a rare functional fungus in Taiwan and contains a variety of biologically active ingredients. Antrodin A (AdA) is one of the main active ingredients in the solid-state fermented A. camphorata mycelium. It protects the liver from alcohol damage by improving the antioxidant and anti-inflammatory capacity of the liver and maintaining the stability of the intestinal flora. AIM OF THE STUDY: The aim of this study was to evaluate the hepatoprotective activities of ethyl acetate layer extract (EALE), AdA, and Antroquinonol (Aq) from mycelium of A. camphorata on alcoholic liver injury. MATERIALS AND METHODS: Mice were given with intragastrically vehicle (NC, 2% CMC-Na), alcohol (AL, 12 mL/kg bw), or different A. camphorata samples (EALE, AdA, Aq) at low (100 mg/kg bw) or high (200 mg/kg bw) dosages. The positive control (PC) group was given with silymarin (200 mg/kg bw). Except the NC group, each group of mice was fasted for 4 h after the last treatment and was intragastrically administrated with 50% alcohol (12 mL/kg bw). At the end of experiment, mouse serum was collected and the liver was excised. A portion of the liver was fixed in formalin and used for histopathological analysis, whereas the rest was used for biochemical analysis and real-time PCR analysis. The intestinal flora structure of feces was analyzed by determining the v3-v4 region sequence in 16S rDNA. RESULTS: The high-dose groups of the three samples (EALEH, AdAH, and AqH) significantly alleviated the alcohol-induced increases in liver index, serum ALT, AST, and AKP activities. Serum TG level was significantly reduced in all treatment groups. The increase of HDL-C content indicated that active ingredients of A. camphorata could reduce the lipid content in serum. Furthermore, MDA contents of the AdAH and AqH groups in liver were significantly reduced, accompanying with the levels of SOD, CAT, and GSH elevated to various extents. Antioxidant and anti-inflammatory capabilities in the liver were increased in the AdAH group, as evidenced by the mRNA expression levels of Nrf-2 and HO-1 were significantly increased; while those of CYP2e1, TNF-α, and TLR-4 were significantly decreased. Analysis of intestinal flora of feces showed that alcohol treatment significantly changed the composition of intestinal flora. Supplementation with AdA could mitigate dysbiosis of intestinal flora induced by alcohol. Flora of Faecalibaculum, Lactobacillus, and Coriobacteriaceae_UCG-002 showed significantly negative correlations with ALT, AST, AKP, and MDA levels. CONCLUSION: Antrodin A could improve the antioxidant and anti-inflammatory capacities of the liver and maintain the stability of intestinal flora. It is potentially a good candidate compound against acute alcoholic liver injury.

Mitochondrial fission protein 1 up-regulation ameliorates senescence-related endothelial dysfunction of human endothelial progenitor cells.

BACKGROUND: We investigated the contribution of mitochondrial dysfunction to the senescence of human endothelial progenitor cells (EPCs) expanded in vitro and the underlying molecular mechanism. METHODS AND RESULTS: Serial passage increased cell doubling time and those cells reaching the doubling time for more than 100% were defined as senescent EPCs, of which the activity of therapeutic angiogenesis was attenuated in mouse ischemic hindlimbs. The senescent cells, in medium free of glucose and bicarbonate, showed impaired activity in migration and tube formation. Flow cytometry indicated increased content of reactive oxygen species, mitochondria, and calcium, while bioenergetic analysis showed increased oxygen consumption and reduced ATP content. Examination of mitochondrial network showed that senescence increased the length of the network and ultrastructure analysis exhibited elongated mitochondria. Immunoblotting of the senescent EPCs demonstrated decreased expression level of fission protein1 (Fis1). In rat EPCs, the Fis1 level was decreased in the animals aged 24 months or older, compared to those of 3 months. Silencing of Fis1 in the young EPCs using Fis1-specific siRNA leads to appearance of phenotype resembling those of senescent cells, including elevated oxidative stress, disturbed mitochondrial network, reduced mitochondria membrane potential, decreasing ATP content, lower proliferation activity, and loss of therapeutic potential in ischemic hindlimbs. Fis1 over-expression in senescent EPCs reduced the oxidative stress, increased the proliferation, and restored the cobble stone-like morphology, senescence, bioenergetics, angiogenic potential, and therapeutic activity. CONCLUSION: In human EPCs, down-regulation of Fis1 is involved in mitochondrial dysfunction and contributes to the impaired activity of EPCs during the senescence process. Enhanced expression of Fis1 in senescent EPCs restores the youthful phenotype.

Induction of COX-1, suppression of COX-2 and pro-inflammatory cytokines gene expression by moringa leaves and its aqueous extract in aspirin-induced gastric ulcer rats.

The Moringa plant (Moringa oleifera) is known for its potential medicinal properties and health benefits in addition to its high nutritional value. The current study aimed to investigate the antiulcer effect of moringa leaves and its aqueous extract on pro-inflammatory cytokines and inflammatory mediators in ulcerative rats. Rats were treated with either moringa leaves (10%) or moringa extract (300 mg/kg body weight) for 4 weeks then treated with a single dose of aspirin to induce gastric ulcer. Moringa leaves and its extract markedly reduced ulcer index, gastric volume and total acidity. Both treatments induced a significant increase in gastric mucosal mucin content and plasma NO level associated with significant decrease in plasma TNFα. Moringa leaves and its extract prompted down-regulation of TNFα, TGFß1 and COX2 genes expression by 2.7, 3.5, and 8.4 fold-change for moringa leaves and 2.7, and 2.3, 4.1 fold-change for moringa extract, respectively. Moringa leaves and extract treatments altered the COX-1 gene expression levels to near normal values. This study confirms the gastro-protective influence of moringa leaves and its extract on aspirin-induced ulcer in rats as manifested by its significant reduction in inflammatory cytokines and normalization of gastric mucosal mucin and NO level. Overall, moringa leaves powder is more efficient as antiulcer agent than moringa extract.

Effect of Omega-3 Fatty Acid on STAMP2 Expression in the Heart and Kidney of 5/6 Nephrectomy Rat Model.

Six transmembrane protein of prostate 2 (STAMP2) is a critical modulator of inflammation and metabolism in adipose tissue. There are no data on the expression of STAMP2 in chronic kidney disease, which is an inflammatory disease related to metabolic disorders. This study aimed to investigate STAMP2 expression in the kidney and heart in 5/6 nephrectomy (Nx) rats, and the effect of omega-3 fatty acid (FA) on STAMP2 expression. Male Sprague Dawley rats were divided into three groups: sham control (0.9% saline), 5/6 Nx (0.9% saline), and 5/6 Nx treated with omega-3 FA (300 mg per kg per day by gastric gavage). The expression of STAMP2 in the kidney and heart were examined by western blotting. Serum creatinine levels were higher in 5/6 Nx rats than in controls. Compared with sham controls, the expression of IκB, NF-κB, NOX4, SREBP-1, and LXR were upregulated and STAMP2 and phosphorylated-AMPK expression were downregulated in the kidney and heart of 5/6 Nx rats. Omega-3 FA supplementation prevented these changes in biomarkers related to inflammation and metabolic lipid disorders. Omega 3-FA supplementation induced the upregulation of STAMP2 protein in 5/6 Nx rats, which was associated with an attenuation of inflammation- and metabolic disease-related markers.

Endocrine sensitivity of estrogen receptor-positive breast cancer is negatively correlated with aspartate-ß-hydroxylase expression.

Although prognostic markers for early estrogen receptor (ER)-positive breast cancer have been extensively developed, predictive markers for adjuvant endocrine therapy are still lacking. Focusing on the mechanisms underlying endocrine resistance, we investigated whether the endocrine sensitivity of ER-positive breast cancer cells was correlated with the expression of aspartate-ß-hydroxylase (ASPH), which is involved in the development of hepatocellular carcinoma. ASPH expression in ER-positive and tamoxifen-resistant breast cancer cells was upregulated by the MAPK and phosphoinositide-3 kinase (PI3K) pathways, which both play pivotal roles in endocrine resistance. In the clinical setting, ASPH expression was negatively correlated with recurrence-free survival of luminal B breast cancer patients that received adjuvant endocrine therapy, but not in patients that did not receive adjuvant endocrine therapy. Luminal B breast cancer is one of the intrinsic molecular subtypes identified by the Prediction Analysis of Microarray 50 (PAM50) multiple gene classifier, and because of its poor response to endocrine therapy, chemotherapy in addition to endocrine therapy is generally required after surgical resection. Our results suggest that the endocrine sensitivity of luminal B breast cancer can be assessed by examining ASPH expression, which promotes the consideration of a prospective study on the association between ASPH expression at the mRNA and protein levels in luminal B breast cancer and subsequent response to endocrine therapy.

Subcritical water-hydrolyzed fish collagen ameliorates survival of endotoxemic mice by inhibiting HMGB1 release in a HO-1-dependent manner.

To investigate potential mechanisms underlying the bioactivity of hydrolyzed fish collagen, we examined the anti-inflammatory actions of subcritical water-hydrolyzed fish collagen (SWFC) in lipopolysaccharide (LPS)-triggered inflammation and endotoxemia. SWFC markedly inhibited LPS-stimulated release of high mobility group box 1 (HMGB1) in murine RAW264.7 macrophages, along with decreased cytosolic translocation of HMGB1. Both the protein and mRNA levels of heme oxygenase-1 (HO-1) were significantly upregulated in SWFC-treated RAW 264.7 cells in an Nrf2-dependent manner. In line with these effects of SWFC, both HO-1 siRNA and ZnPPIX (zinc protoporphyrin IX) actually attenuated the effects of SWFC on HMGB1 release stimulated by LPS, indicating a possible mechanism by which SWFC modulates HMGB1 release through HO-1 signaling. Notably, administration of SWFC improved the survival rates of LPS-injected endotoxemic mice, in which the serum level of HMGB1 was significantly reduced. Taken together, these results indicate that the anti-inflammatory activities of SWFC are achieved by inhibiting HMGB1 release induced by LPS in a HO-1-sensitive manner.

Heme Oxygenase-1 Induction and Anti-inflammatory Actions of Atractylodes macrocephala and Taraxacum herba Extracts Prevented Colitis and Was More Effective than Sulfasalazine in Preventing Relapse.

BACKGROUND/AIMS: In inflammatory bowel disease (IBD), repeated bouts of remission and relapse occur in patients and can impose a risk of colitis-associated cancer. We hypothesized that plant extracts of Atractylodes macrocephala (AM) or Taraxacum herba (TH) may be better than sulfasalazine for treating this disease because these extracts can promote additional regeneration. METHODS: Murine intestinal epithelial IEC-6 cells were pretreated with AM or TH before a lipopolysaccharide (LPS)-induced challenge. Acute colitis was induced with 7 days of dextran sulfate sodium (DSS) in male C57BL/6 mice, and extracts of AM and TH were administered for 2 weeks before DSS administration. RESULTS: In vitro studies demonstrated that AM or TH treatment reduced LPS-induced COX-2 and tumor necrosis factor-α mRNA levels but increased heme oxygenase-1 (HO-1). Oral preadministration of AM and TH rescued mice from DSS-induced colitis by inhibiting inflammatory mediators via inactivated extracellular signal regulated kinase and repressed nuclear factor κB and signal transducer and activator of transcription 3, but the effect was weaker for sulfasalazine than that for the extracts. Anti-inflammatory activities occurred via the inhibition of macrophage and T lymphocyte infiltrations. Unlike sulfasalazine, which did not induce HO-1, TH extracts afforded significant HO-1 induction. CONCLUSIONS: Because the AM or TH extracts were far superior in preventing DSS-induced colitis than sulfasalazine, AM or TH extracts can be considered natural agents that can prevent IBD relapse.

Curcumin prevents mitochondrial dynamics disturbances in early 5/6 nephrectomy: Relation to oxidative stress and mitochondrial bioenergetics.

Five-sixths nephrectomy (5/6NX) is a widely used model to study the mechanisms leading to renal damage in chronic kidney disease (CKD). However, early alterations on renal function, mitochondrial dynamics, and oxidative stress have not been explored yet. Curcumin is an antioxidant that has shown nephroprotection in 5/6NX-induced renal damage. The aim of this study was to explore the effect of curcumin on early mitochondrial alterations induced by 5/6NX in rats. In isolated mitochondria, 5/6NX-induced hydrogen peroxide production was associated with decreased activity of complexes I and V, decreased activity of antioxidant enzymes, alterations in oxygen consumption and increased MDA-protein adducts. In addition, it was found that 5/6NX shifted mitochondrial dynamics to fusion, which was evidenced by increased optic atrophy 1 and mitofusin 1 (Mfn1) and decreased fission 1 and dynamin-related protein 1 expressions. These data were confirmed by morphological analysis and immunoelectron microscopy of Mfn-1. All the above-described mechanisms were prevented by curcumin. Also, it was found that curcumin prevented renal dysfunction by improving renal blood flow and the total antioxidant capacity induced by 5/6NX. Moreover, in glomeruli and proximal tubules 5/6NX-induced superoxide anion production by uncoupled nitric oxide synthase (NOS) and nicotinamide adenine dinucleotide phosphate oxidase (NOX) dependent way, this latter was associated with increased phosphorylation of serine 304 of p47phox subunit of NOX. In conclusion, this study shows that curcumin pretreatment decreases early 5/6NX-induced altered mitochondrial dynamics, bioenergetics, and oxidative stress, which may be associated with the preservation of renal function. © 2016 BioFactors, 43(2):293-310, 2017.

Neurohormetic responses of quercetin and rutin in a cell line over-expressing the amyloid precursor protein (APPswe cells).

BACKGROUND: Plant secondary metabolites may induce adaptive cellular stress-responses in a variety of cells including neurons at the sub-toxic doses ingested by humans. Such 'neurohormesis' phenomenon, activated by flavonoids such as quercetin or rutin, may involve cell responses driven by modulation of signaling pathways which are responsible for its neuroprotective effects. PURPOSE: We attempt to explore the molecular mechanisms involved in the neurohormetic responses to quercetin and rutin exposure, in a SH-SY5Y cell line which stably overexpresses the amyloid precursor protein (APP) Swedish mutation, based on a biphasic concentration-response relationship for cell viability. METHODS: We examined the impact of both natural compounds, at concentrations in its hormetic range on the following cell parameters: chymotrypsin-like activity of the proteasome system; PARP-1 protein levels and expression and caspase activation; APP processing; and the main endogenous antioxidant enzymes. RESULTS: Proteasome activities following quercetin or rutin treatment were significantly augmented in comparison with non-treated cells. Activity of caspase-3 was significantly attenuated by treatment with quercetin or rutin. Modest increased levels of PARP-1 protein and mRNA transcripts were observed in relation to the mild increase of proteasome activity. Significant reductions of the full-length APP and sAPP protein and APP mRNA levels were related to significant enhancements of α-secretase ADAM-10 protein and mRNA transcripts and significant increases of BACE processing in cells exposed to rutin. Furthermore, quercetin or rutin treatment displayed an overall increase of the four antioxidant enzymes. CONCLUSIONS: The upregulation of the proteasome activity observed upon quercetin or rutin treatment could be afforded by a mild increased of PARP-1. Consequently, targeting the proteasome by quercetin or rutin to enhance its activity in a mild manner could avoid caspase activation. Moreover, it is likely that APP processing of cells upon rutin treatment is mostly driven by the non-amyloidogenic pathway leading to a putative reduction of ßA production. Overall induction of endogenous antioxidant enzymes under quercetin or rutin treatments of APPswe cells might modulate its proteasome activity. We might conclude that quercetin and rutin might exert a neurohormetic cell response affecting various signaling pathways and molecular networks associated with modulation of proteasome function.

Valproate Induces the Unfolded Protein Response by Increasing Ceramide Levels.

Bipolar disorder (BD), which is characterized by depression and mania, affects 1-2% of the world population. Current treatments are effective in only 40-60% of cases and cause severe side effects. Valproate (VPA) is one of the most widely used drugs for the treatment of BD, but the therapeutic mechanism of action of this drug is not understood. This knowledge gap has hampered the development of effective treatments. To identify candidate pathways affected by VPA, we performed a genome-wide expression analysis in yeast cells grown in the presence or absence of the drug. VPA caused up-regulation of FEN1 and SUR4, encoding fatty acid elongases that catalyze the synthesis of very long chain fatty acids (C24 to C26) required for ceramide synthesis. Interestingly, fen1Δ and sur4Δ mutants exhibited VPA sensitivity. In agreement with increased fatty acid elongase gene expression, VPA increased levels of phytoceramide, especially those containing C24-C26 fatty acids. Consistent with an increase in ceramide, VPA decreased the expression of amino acid transporters, increased the expression of ER chaperones, and activated the unfolded protein response element (UPRE), suggesting that VPA induces the UPR pathway. These effects were rescued by supplementation of inositol and similarly observed in inositol-starved ino1Δ cells. Starvation of ino1Δ cells increased expression of FEN1 and SUR4, increased ceramide levels, decreased expression of nutrient transporters, and induced the UPR. These findings suggest that VPA-mediated inositol depletion induces the UPR by increasing the de novo synthesis of ceramide.

Receptor-Like Kinase RUPO Interacts with Potassium Transporters to Regulate Pollen Tube Growth and Integrity in Rice.

During sexual reproduction of flowering plants, the pollen tube grows fast and over a long distance within the pistil to deliver two sperms for double fertilization. Growing plant cells need to communicate constantly with external stimuli as well as monitor changes in surface tension of the cell wall and plasma membrane to coordinate these signals and internal growth machinery; however, the underlying mechanisms remain largely unknown. Here we show that the rice member of plant-specific receptor-like kinase CrRLK1Ls subfamily, Ruptured Pollen tube (RUPO), is specifically expressed in rice pollen. RUPO localizes to the apical plasma membrane and vesicle of pollen tubes and is required for male gamete transmission. K+ levels were greater in pollen of homozygous CRISPR-knockout lines than wild-type plants, and pollen tubes burst shortly after germination. We reveal the interaction of RUPO with high-affinity potassium transporters. Phosphorylation of RUPO established and dephosphorylation abolished the interaction. These results have revealed the receptor-like kinase as a regulator of high-affinity potassium transporters via phosphorylation-dependent interaction, and demonstrated a novel receptor-like kinase signaling pathway that mediates K+ homeostasis required for pollen tube growth and integrity.

Anti­inflammatory and antioxidant activity of the traditional herbal formula Gwakhyangjeonggi­san via enhancement of heme oxygenase­1 expression in RAW264.7 macrophages.

Gwakhyangjeonggi­san (GHJGS) is a mixture of herbal plants, including Agastache rugosa, Perilla frutescens, Angelica dahurica, Areca catechu, Poria cocos, Magnolia officinalis, Atractylodes macrocephala, Citrus reticulata, Pinellia ternata, Platycodon grandiflorum, Glycyrrhiza uralensis, Ziziphus jujuba and Zingiber officinale. GHJGS has been used for treating diarrhea­predominant irritable bowel syndrome in traditional Korean medicine. In the present study, the anti­inflammatory and antioxidant effects of GHJGS were investigated using the RAW 264.7 murine macrophage cell line. GHJGS significantly reduced production of the proinflammatory cytokines, tumor necrosis factor­α, interleukin­6 and prostaglandin E2 in lipopolysaccharide (LPS)­stimulated macrophages. GHJGS markedly suppressed LPS­induced phosphorylation of mitogen­activated protein kinases, whereas it had no effect on nuclear factor­κB activation. Furthermore, GHJGS enhanced expression of heme oxygenase­1 and prevented the generation of reactive oxygen species in RAW 264.7 cells. These results indicate that GHJGS is a viable therapeutic agent against inflammation and oxidative stress­associated disorders.

Biological therapy induces expression changes in Notch pathway in psoriasis.

Psoriasis is a chronic inflammatory skin disease, characterized by hyperproliferation of keratinocytes and by skin infiltration of activated T cells. To date, the pathophysiology of psoriasis has not yet been fully elucidated. The Notch pathway plays a determinant role in cell fate determination, proliferation, differentiation, immune cell development and function. Many biological agents, used in the treatment of psoriasis, include TFN-α inhibitors, such as etanercept, adalimumab, and anti IL-12/IL-23 p40 antibody, such as ustekinumab. This study aimed to determine mRNA expression levels by real-time RT-PCR, and protein expression levels, analysed by Western blot and immunohistochemistry, of some components of the Notch pathway, such as NOTCH1, NOTCH2, JAGGED1, and HES1 after biological treatments in psoriatic patients. mRNA and protein levels of NOTCH1, NOTCH2, JAGGED1 and HES1 were upregulated in skin samples from untreated psoriatic patients compared with normal controls. Biological therapy showed to downregulate differently the protein expression levels of the molecules under study. Our study suggests that Notch pathway components might be a potential therapeutic target against psoriasis.

Neonatal exposure to mild hyperoxia causes persistent increases in oxidative stress and immune cells in the lungs of mice without altering lung structure.

Preterm infants often require supplemental oxygen due to lung immaturity, but hyperoxia can contribute to an increased risk of respiratory illness later in life. Our aim was to compare the effects of mild and moderate levels of neonatal hyperoxia on markers of pulmonary oxidative stress and inflammation and on lung architecture; both immediate and persistent effects were assessed. Neonatal mice (C57BL6/J) were raised in either room air (21% O2), mild (40% O2), or moderate (65% O2) hyperoxia from birth until postnatal day 7 (P7d). The mice were killed at either P7d (immediate effects) or lived in air until adulthood (P56d, persistent effects). We enumerated macrophages in lung tissue at P7d and immune cells in bronchoalveolar lavage fluid (BALF) at P56d. At P7d and P56d, we assessed pulmonary oxidative stress [heme oxygenase-1 (HO-1) and nitrotyrosine staining] and lung architecture. The data were interrogated for sex differences. At P7d, HO-1 gene expression was greater in the 65% O2 group than in the 21% O2 group. At P56d, the area of nitrotyrosine staining and number of immune cells were greater in the 40% O2 and 65% O2 groups relative to the 21% O2 group. Exposure to 65% O2, but not 40% O2, led to larger alveoli and lower tissue fraction in the short term and to persistently fewer bronchiolar-alveolar attachments. Exposure to 40% O2 or 65% O2 causes persistent increases in pulmonary oxidative stress and immune cells, suggesting chronic inflammation within the adult lung. Unlike 65% O2, 40% O2 does not affect lung architecture.

Medicinal effect and its JP2/RyR2-based mechanism of Smilax glabra flavonoids on angiotensin II-induced hypertrophy model of cardiomyocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Rhizome and root of Smilax glabra Roxb (Liliaceae family) is a widely used traditional Chinese medicine (TCM) named Tu-fu-ling (TFL) for cardiac disease therapy. The TFL flavonoids (TFLF) has been extracted and proven to possess the anti-cardiac hypertrophy effect in our previous reports. Such effect could be mediated by the modulation of intracellular Ca(2+) flux in myocardial cells, in which junctophilin-2 (JP2) and ryanodine receptor 2 (RyR2) play an important role. However, its mechanism of the anti-cardiac hypertrophy effect remains unclarified. MATERIALS AND METHODS: 2µmol/L Ang II was applied to induce hypertrophy model of rat primary cardiomyocytes. After treatment of TFLF at 0.25, 0.5 and 1.0mg/ml, the cell size was microscopic measured, and the protein and mRNA expressions of JP2 and RyR2 in cardiomyocytes were estimated by immunofluorescence imaging, ELISA and real-time PCR assay. RESULTS: Obvious hypertrophy of cardiomyocytes was induced by Ang II but reversed by TFLF from 0.5 to 1.0mg/ml. The protein and mRNA expressions of JP2 and RyR2 in cardiomyocytes were also inhibited by Ang II but restored by TFLF at its dose range. Such effect of TFLF was exerted at a dose dependent manner, which was even better than that of verapamil. CONCLUSIONS: Our findings may evidence the correlation between JP2/RyR2 and myocardiac hypertrophy, and indicate the JP2/RyR2-mediated anti-hypertrophy mechanism of TFLF for the first time. It deserves to be developed as a promising TCM candidate of new drug for myocardial hypertrophy treatment.

The impact of FGFR1 and FRS2α expression on sorafenib treatment in metastatic renal cell carcinoma.

BACKGROUND: Angiogenesis plays a role in tumor growth and is partly mediated by factors in both the fibroblast growth factor (FGF) and vascular endothelial growth factor (VEGF) pathways. Durable clinical responses with VEGF tyrosine kinase inhibitors (TKIs) may be limited by intrinsic tumor resistance. We hypothesized that FGF signaling may impact clinical responses to sorafenib. METHODS: Nephrectomy material was available from 40 patients with metastatic renal cell carcinoma (RCC) enrolled in a phase II clinical trial of sorafenib ± interferon (ClinicalTrials.gov Identifier NCT00126594). Fibroblast growth factor receptor 1 (FGFR1) and fibroblast growth factor receptor substrate 2 alpha (FRS2α) expression was assessed by in situ hybridization and immunofluorescence, respectively. The relationship between fibroblast growth factor pathway marker levels and progression-free survival (PFS) was analyzed using Kaplan-Meier and Cox proportional hazards regression methods. RESULTS: Univariate analysis indicated that more intense FGFR1 staining was associated with shorter PFS (log-rank P = 0.0452), but FRS2α staining was not significantly associated with PFS (log-rank P = 0.2610). Multivariate Cox proportional hazards regression models were constructed for FGFR1 and FRS2α individually, adjusting for baseline Eastern Cooperative Oncology Group performance status, treatment arm and anemia status. When adjusted for each of these variables, the highest intensity level of FGFR1 (level 3 or 4) had increased progression risk relative to the lowest intensity level of FGFR1 (level 1) (P = 0.0115). The highest intensity level of FRS2α (level 3 or 4) had increased progression risk relative to the lowest intensity level of FRS2α (level 1) (P = 0.0126). CONCLUSIONS: Increased expression of FGFR1 and FRS2α was associated with decreased PFS among patients with metastatic RCC treated with sorafenib. The results suggest that FGF pathway activation may impact intrinsic resistance to VEGF receptor inhibition.