Endurance running training can lead to gradual accumulation of inflammation and soreness ultimately resulting in overuse injuries. Management of soreness and inflammation with pharmaceuticals (i.e. non-prescription pain relievers) during long-term training is not a suitable solution due to known side effects (e.g. gastrointestinal complications, etc.). Dietary polyphenols (i.e. curcumin, pomegranate, etc.) have been purported to reduce inflammation and muscle soreness, without these negative side effects making them ideal for use in an exercise model. The purpose of the present feasibility study was to explore the combined effect of optimized curcumin and pomegranate extract supplementation prior to (PRE) and after (4H and 24H) an organized half-marathon race on blood inflammatory proteins and inflammation-associated RNA. Daily supplementation (1000 mg/d) started 26 days before a half-marathon which doubled on days 27-31. Data were analyzed with R software and Welch t-test, significance set at p < 0.05. At both 4H and 24H, supplementation was associated with alterations in protein (IL-10, IL-13, IL-4, ITAC, MIP-1alpha, MIP-3alpha, BDNF, sIL-2Ralpha, and TNF-alpha; p < 0.05) and RNA (CCL22, GUSB, IL-6, LINC00305, NKILA, PTGES, THRIL, TRAF6, ARG2, CD1A, CD55, CFI, CSF2, CXC3CL1, CX3CR1, EDNRB, GATA3, LILRB5, THY1, CD3D, MRC1, GPR183, HAMP, MBL2, CASP3, B2M, KLRF2, PDCD1LG2, IL-10, PTGS2, TLR2, IL-6R, IL-8, IL-7R, MASP1, MYD88, TNFRSF1B, TNFRSF1A, and TIRAP; p < 0.05) biomarkers compared to control. Pathway classification of these biomarkers indicated supplementation may be associated with a more favorable muscle recovery profile. Our findings support the notion that combined curcumin and pomegranate supplementation may represent a useful addition to a comprehensive exercise training plan.

Keywords: inflammation; muscle injury; polyphenols; running.

We investigated the effect of Yi Gong San (YGS) decoction on iron homeostasis and the possible underlying mechanisms in a mouse model of acute inflammation in this study. Our findings suggest that YGS regulates iron homeostasis by downregulating the level of HAMP mRNA, which may depend on regulation of the IL-6/STAT3 or BMP/HJV/SMAD pathway during acute inflammation.

In this study, Spirulina maxima derived pectin nanoparticles (SmPNPs) were synthesized and multiple biological effects were investigated using in vitro and in vivo models. SmPNPs were not toxic to Raw 264.7 cells and zebrafish embryos up to 1 mg/mL and 200 µg/mL, respectively. SmPNPs upregulated Il 10, Cat, Sod 2, Def 1, Def 2, and Muc 1 in Raw 264.7 cells and tlr2, tlr4b, tlr5b, il1β, tnfα, cxcl8a, cxcl18b, ccl34a.4, ccl34b.4, muc5.1, muc5.2, muc5.3, hamp, cstd, hsp70, cat, and sod1 in the larvae and adult zebrafish, suggesting immunomodulatory activity. Exposure of larvae to SmPNPs followed by challenge with pathogenic bacterium Aeromonas hydrophila resulted a two-fold reduction of reactive oxygen species, indicating reduced oxidative stress compared to that in the control group. The cumulative percent survival of larvae exposed to SmPNPs (50 µg/mL) and adults fed diet supplemented with SmPNPs (4%) was 53.3% and 76.7%, respectively. Topical application of SmPNPs on adult zebrafish showed a higher wound healing percentage (48.9%) compared to that in the vehicle treated group (38.8%). Upregulated wound healing markers (tgfβ1, timp2b, mmp9, tnfα, il1β,ccl34a.4, and ccl34b.4), enhanced wound closure, and restored pigmentation indicated wound healing properties of SmPNPs. Overall, results uncover the multiple bioactivities of SmPNPs, which could be a promising biocompatible candidate for broad range of aquatic and human therapies.

Keywords: Aeromonas hydrophila; Spirulina maxima; immunomodulation; pectin nanoparticles; stress tolerance; wound healing; zebrafish.

Thiazolidinediones are well known anti-diabetic drugs. However, they are not widely used due to their cardiotoxic effects. Therefore, in this study we aimed to determine the molecular toxicological alterations induced in the mouse hearts after thiazolidinedione administration. Balb/c mice received doses clinically equivalent to those given to humans of the most commonly used thiazolidinediones, pioglitazone, and rosiglitazone for 30 days. After that, RNA samples were isolated from the hearts. The mRNA expression of cytochrome (cyp) p450 genes that synthesize the cardiotoxic 20-hydroxyecostarionic acid (20-HETE) in addition to 92 cardio-toxicity biomarker genes were analyzed using quantitative polymerase chain reaction array technique. The analysis demonstrated that thiazolidinediones caused a significant upregulation (p < 0.5) of the mRNA expression of cyp1a1,cyp4a12,itpr1,ccl7,ccr1, and b2m genes. In addition, thiazolidinediones caused a significant (p < 0.05) downregulation of the mRNA expression of adra2a,bsn,col15a1,fosl1,Il6,bpifa1,plau, and reg3b genes. The most affected gene was was itpr1 gene, which was upregulated by pioglitazone and rosiglitazone by 7- and 3.5-fold, respectively. In addition, pioglitazone caused significant upregulation of (p < 0.05) hamp,ppbp,psma2,sik1,timp1, and ucp1 genes, which were not affected significantly (p > 0.05) by rosiglitazone administration. In conclusion, this study showed that thiazolidinediones induces toxicological molecular alterations in the mouse hearts, such as the induction of cyp450s that synthesize 20-HETE, chemokine activation, inflammatory responses, blood clotting, and oxidative stress. These findings may help us understand the mechanism of cardiotoxicity involved in thiazolidinedione administration.

Keywords: 20-HETE; Cardiotoxicity; PCR array; molecular alteration; thiazolidinediones.

Bacteria closely control gene expression to ensure optimal physiological responses to their environment. Such careful gene expression can minimize the fitness cost associated with antibiotic resistance. We previously described a novel regulatory logic in Bacillus subtilis enabling the cell to directly monitor its need for detoxification. This cost-effective strategy is achieved via a two-component regulatory system (BceRS) working in a sensory complex with an ABC-transporter (BceAB), together acting as a flux-sensor where signaling is proportional to transport activity. How this is realized at the molecular level has remained unknown. Using experimentation and computation we here show that the histidine kinase is activated by piston-like displacements in the membrane, which are converted to helical rotations in the catalytic core via an intervening HAMP-like domain. Intriguingly, the transporter was not only required for kinase activation, but also to actively maintain the kinase in its inactive state in the absence of antibiotics. Such coupling of kinase activity to that of the transporter ensures the complete control required for transport flux-dependent signaling. Moreover, we show that the transporter likely conserves energy by signaling with sub-maximal sensitivity. These results provide the first mechanistic insights into transport flux-dependent signaling, a unique strategy for energy-efficient decision making.

Keywords: Signal transduction; antimicrobial peptide; cell envelope stress; flux-sensing.

Under anaerobic conditions, bacteria may utilize nitrates and nitrites as electron acceptors. Sensitivity to nitrous compounds is achieved via several mechanisms, some of which rely on sensor histidine kinases (HKs). The best studied nitrate- and nitrite-sensing HKs (NSHKs) are NarQ and NarX from Escherichia coli. Here, we review the function of NSHKs, analyze their natural diversity, and describe the available structural information. In particular, we show that around 6000 different NSHK sequences forming several distinct clusters may now be found in genomic databases, comprising mostly the genes from Beta- and Gammaproteobacteria as well as from Bacteroidetes and Chloroflexi, including those from anaerobic ammonia oxidation (annamox) communities. We show that the architecture of NSHKs is mostly conserved, although proteins from Bacteroidetes lack the HAMP and GAF-like domains yet sometimes have PAS. We reconcile the variation of NSHK sequences with atomistic models and pinpoint the structural elements important for signal transduction from the sensor domain to the catalytic module over the transmembrane and cytoplasmic regions spanning more than 200 Å.

Keywords: allostery; cell signaling; histidine kinases; nitrate regulation; nitrate respiration; signal transduction; two-component systems.

Hepcidin is a peptide hormone that targets the iron exporter ferroportin, thereby limiting iron entry into the bloodstream. It is generated in hepatocytes mainly in response to increased body iron stores or inflammatory cues. Iron stimulates expression of bone morphogenetic protein 6 (BMP6) from liver sinusoidal endothelial cells, which in turn binds to BMP receptors on hepatocytes and induces the SMAD signaling cascade for transcriptional activation of the hepcidin-encoding HAMP mRNA. SMAD signaling is also essential for inflammatory HAMP mRNA induction by the IL-6/STAT3 pathway. Herein, we utilized human Huh7 hepatoma cells and primary murine hepatocytes to assess the effects of iron perturbations on signaling to hepcidin. Iron chelation appeared to slightly impair signaling to hepcidin. Subsequent iron supplementation not only failed to reverse these effects, but drastically reduced basal HAMP mRNA and inhibited HAMP mRNA induction by BMP6 and/or IL-6. Thus, treatment of cells with excess iron inhibited basal and BMP6-mediated SMAD5 phosphorylation and induction of HAMP, ID1 and SMAD7 mRNAs in a dose-dependent manner. Iron also inhibited IL-6-mediated STAT3 phosphorylation and induction of HAMP and SOCS3 mRNAs. These responses were accompanied by induction of GCLC and HMOX1 mRNAs, known markers of oxidative stress. We conclude that hepatocellular iron overload suppresses hepcidin by inhibiting the SMAD and STAT3 signaling pathways downstream of their respective ligands.

We herein describe a novel technology, termed self-priming phosphorothioated hairpin-mediated isothermal amplification (SP-HAMP), enabling target nucleic acid detection. Isothermal amplification strategies are a simple process that efficiently raises the amount of nucleic acid at a constant temperature, but still has lots of problems such as the requirement of multiple exogenous primers and enzymes, which trigger non-specific background signal and increase the complexity of procedures. The key component for overcoming the above-mentioned limitations is the designed hairpin probe (HP) consisting of self-priming region along the 3' stem and the 3' overhang and phosphorothioate modifications at the 5' overhang and the specific loop part. The HP was designed to open through binding to target nucleic acid. Upon opening of HP, its self-priming (SP) region is rearranged to form a smaller hairpin whose 3' end could serve as a primer. The following extension produces the extended HP and displaces the bound target nucleic acid, which is then recycled to open another HP. Due to the reduced stability caused by the specific two phosphorothioate (PS) modifications, the 3' end of EP1 is readily rearranged to form the foldback hairpin structure, which would promote the foldback extension to produce once more extended HP. Since the two PS modifications are always located at the same positions along the 5' stem within the further extended HPs, the foldback reaction followed by the extension would be continuously repeated, consequently producing a large number of the long hairpin concatamers. Based on this unique design principle, we successfully detected even a single copy of target DNA with outstanding discrimination capability under an isothermal condition by employing only a single HP without the requirement for the complicated multiple primers. In conclusion, the sophisticated design principle employed in this work would provide great insight for the development of self-operative isothermal amplifying system enabling short target nucleic acid detection such as microRNAs or any target which is less than 200 mer.

Keywords: Loop-mediated isothermal amplification (LAMP); Nucleic acid detection; Phosphorothioate (PS) DNA; Self-priming hairpin.

Hereditary hemochromatosis (HH) is an inherited iron overload disorder characterized by normal iron-driven erythropoiesis and abnormal iron metabolism, leading to excess iron deposited in parenchymal cells of liver, heart, and endocrine glands. Iron hormone, hepcidin, plays a critical role in iron homeostasis through interaction with ferroportin (FPN), a major cellular iron exporter. Hepcidin is encoded by hepcidin antimicrobial peptide (HAMP). Mutations in hepcidin and any genes that regulate the biology of hepcidin, including hemochromatosis genes (HFE), Hemojuvelin (HJV), transferring receptor 2 (TFR2) and FPN, result in hemochromatosis. The identification of hepcidin and its role will provide a better understanding for pathogenesis of HH.

Plants fight-off pathogens and pests by manifesting an array of defense responses using their innate immunity mechanisms. Here we report the identification of a novel soybean gene encoding a plasma membrane protein, transcription of which is suppressed following infection with the fungal pathogen, Furatium virguliforme. Overexpression of the protein led to enhanced resistance against not only against F. virguliforme, but also against spider mites (Tetranychus urticae, Koch), soybean aphids (Aphis glycines, Matsumura) and soybean cyst nematode (Heterodera glycines). We, therefore, name this protein as Glycine max disease resistance 1 (GmDR1; Glyma.10g094800). The homologues of GmDR1 have been detected only in legumes, cocoa, jute and cotton. The deduced GmDR1 protein contains 73 amino acids. GmDR1 is predicted to contain an ecto- and two transmembrane domains. Transient expression of the green fluorescent protein fused GmDR1 protein in soybean leaves showed that it is a plasma membrane protein. We investigated if chitin, a pathogen associated molecular pattern (PAMP), common to all pathogen and pests considered in this study, can significantly enhance defense pathways among the GmDR1-overexpressed transgenic soybean lines. Chitin induces marker genes of the salicylic- and jasmonic acid-mediated defense pathways; but suppresses the defense pathway regulated by ethylene. Chitin induced SA- and JA-regulated defense pathways may be one of the mechanisms involved in generating broad-spectrum resistance among the GmDR1-overexpressed transgenic soybean lines against two serious pathogens and two pests including spider-mites, against which no known resistance genes have been identified in soybean and among the most other crop species.

Keywords: HAMP; MAMP; PAMP; PTI; chitin; disease resistance; pattern-recognition receptors (PRR); plasma membrane; soybean aphids; soybean cyst nematode; soybean sudden death syndrome; spider mites.

A growing emphasis in anticancer drug discovery efforts has been on targeting histone acetylation modulators. Here we comprehensively analyze the genomic alterations of the genes encoding histone acetylation modulator proteins (HAMPs) in the Cancer Genome Atlas cohort and observe that HAMPs have a high frequency of focal copy number alterations and recurrent mutations, whereas transcript fusions of HAMPs are relatively rare genomic events in common adult cancers. Collectively, 86.3% (63/73) of HAMPs have recurrent alterations in at least 1 cancer type and 16 HAMPs, including 9 understudied HAMPs, are identified as putative therapeutic targets across multiple cancer types. For example, the recurrent focal amplification of BRD9 is observed in 9 cancer types and genetic depletion of BRD9 inhibits tumor growth. Our systematic genomic analysis of HAMPs across a large-scale cancer specimen cohort may facilitate the identification and prioritization of potential drug targets and selection of suitable patients for precision treatment.

Glutaricacidemia type 1(GA1) is an autosomal recessive disease caused by reduced or missing glutaryl-CoA dehydrogenase activity which hamps metabolism of lysine, hydroxylysine and tryptophan. The catabolic products of glutarylcarnitine and glutaric acid are abnormally accumulated in the body, resulting in metabolic disorders which primarily lead to damage to the nervous system. Clinical manifestations of patients include macrocephaly, dystonia, dyskinesia, and developmental retardation. Acute encephalopathy may be induced in infants and young children due to infection, vaccination and surgery. For GA1 is a rare disease and its clinical manifestations are similar to other neurological diseases, it may be easily missed or misdiagnosed. To facilitate early diagnosis and treatment and improve the prognosis, this consensus was formulated by pediatric experts from the fields of endocrinology and genetic metabolism through full discussion and reference to the latest literature and guidelines home and abroad.

A simple, extractive heptafluoro-n-butyrylation with Extrelut columns was devised to simultaneously measure methamphetamine (MAMP), amphetamine (AMP), 4-hydroxymethamphetamine (HMAMP), and 4-hydroxyamphetamine (HAMP) in biological materials by gas chromatography-mass spectrometry (GC-MS) using 4-methoxymethamphetamine-d5 as the internal standard. Human urine, human whole blood, and porcine skeletal muscle spiked with the stimulant standards were used for evaluating the method. After deproteinization and adjustment of the pH to 12.6, the sample was applied to an Extrelut column. Using the present method, AMP, MAMP, and HMAMP could be determined in an actual forensic case study.

Plant pollens are airborne allergens, as their inhalation causes immune activation, leading to rhinitis, conjunctivitis, sinusitis and oral allergy syndrome. A myriad of pollen proteins belonging to profilin, expansin, polygalacturonase, glucan endoglucosidase, pectin esterase, and lipid transfer protein class have been identified. In the present in silico study, the protein domains of fifteen pollen sequences were extracted from the UniProt database and submitted to the interactive web tool SMART (Simple Modular Architecture Research Tool), for finding the protein domain profiles. Analysis of the data based on custom-made scripts revealed the conservation of pathogenic domains such as OmpH, PROF, PreSET, Bet_v_1, Cpl-7 and GAS2. Further, the retention of critical domains like CHASE2, Galanin, Dak2, DALR_1, HAMP, PWI, EFh, Excalibur, CT, PbH1, HELICc, and Kelch in pollen proteins, much like cockroach allergens and lethal viruses (such as HIV, HCV, Ebola, Dengue and Zika) was observed. Based on the shared motifs in proteins of taxonomicall-ydispersed organisms, it can be hypothesized that allergens and pathogens manipulate the human immune system in a similar manner. Allergens, being inanimate, cannot replicate in human body, and are neutralized by immune system. But, when the allergens are unremitting, the immune system becomes persistently hyper-sensitized, creating an inflammatory milieu. This study is expected to contribute to the understanding of pollen allergenicity and pathogenicity.

BACKGROUND

The ultimate goal of periodontal therapy is predictable regeneration of a functional attachment apparatus destroyed as a result of periodontitis. Reconstructive procedures have been used with varying success during the past decades to accomplish this goal.

OBJECTIVE

To evaluate whether the use of porous hydroxyapatite alone or a bioresorbable membrane alone would enhance the clinical results in the treatment of class II furcation defects in human lower molars.

METHODS

Fifteen patients with chronic periodontitis, aged between 39 and 49 years, with a pair of similar bilateral class II furcation defects (classification of Hamp et al.) in mandibular first molars were selected. A split-mouth design was incorporated and the selected 30 furcation defects were assigned to one of the two treatment groups, i.e., Group I treated with a bioresorbable membrane from bovine-derived collagen guided tissue regeneration membrane and Group II treated using porous hydroxyapatite bone graft material on the contralateral sides. Evaluation of clinical parameters, probing depths and attachment levels, and radiographs was done preoperatively and 6 months postoperatively.

RESULTS

Both the groups showed statistically significant mean reduction in probing depths and gain in clinical attachment levels and linear bone fill. Comparison between Group I and Group II showed insignificant difference.

CONCLUSIONS

Within the limits of this study, both the treatment modalities are beneficial for the treatment of human mandibular class II furcation defects.

The classification of hereditary abnormalities of iron metabolism was recently expanded and diversified. Genetic hemochromatosis now corresponds to six diseases, namely classical hemochromatosis HFE 1; juvenile hemochromatosis HFE 2 due to mutations in an unidentified gene on chromosome 1; hemochromatosis HFE 3 due to mutations in the transferrin receptor 2 (TfR2); hemochromatosis HFE 4 caused by a mutation in the H subunit of ferritin; and hemochromatosis HFE 6 whose gene is hepcidine (HAMP). Systemic iron overload is also associated with aceruloplasminemia, atransferrinemia and the "Gracile" syndrome caused by mutations in BCS1L. The genes responsible for neonatal and African forms of iron overload are unknown. Other genetic diseases are due to localized iron overload: Friedreich's ataxia results from the expansion of triple nucleotide repeats within the frataxin (FRDA) gene; two forms of X-linked sideroblastic anemia are due to mutations within the delta aminolevulinate synthetase (ALAS 2) or ABC-7 genes; Hallervorden-Spatz syndrome is caused by a pantothenate kinase 2 gene (PANK-2) defect; neuroferritinopathies; and hyperferritinemia--cataract syndrome due to a mutation within the L-ferritin gene. In addition to this wide range of genetic abnormalities, two other features characterize these iron disorders: 1) most are transmitted by an autosomal recessive mechanism, but some, including hemochromatosis type 4, have dominant transmission; and 2) most correspond to cytosolic iron accumulation while some, like Friedreich's ataxia, are disorders of mitochondrial metabolism.

The review summarizes the results of the state-of-the-art studies of hemochromatosis (HC): iron-regulatory genes (HFE, HJV, HAMP, TFR2, SLC40A1) have been discovered; the HC types caused by mutations in these genes (types 1, 2, 3, and 4 in the OMIM register) have been identified; the inflammation anemia (IA) mediator - the polypeptide hepatic hormone hepcidin that is an important constituent of the natural immunity system - has been found. This gives an idea of hypersiderosis and dissiderosis as types of iron microelementosis. Types 1, 2, and 3 HC in which iron absorption and its total reserves are increased in the body serve as examples of hypersiderosis. Dissiderosis is characterized by the redistribution of iron between the functional and spare funds inherent in type 4 HC and IA. By taking into account their findings, the authors briefly discuss the leading clinical and morphological manifestations of HC and IA, difficulties in differential diagnosis, and treatment ways and prospects.

The effects of an adequate supply of vitamin A and iron, in comparison with diets low or absent in vitamin A and low in iron, on the mRNA expression of some biomarkers of iron homeostasis as hepcidin (Hamp), transferrin receptor-1 (Tfrc), iron regulatory protein-2 (Ireb2) and ferritin (Fth1) in rats were investigated. 35 male Wistar rats were randomly divided into 5 dietary groups: control, sufficient in iron and insufficient in vitamin A (FesvAi), sufficient in iron and depleted in vitamin A (FesvAd), insufficient in iron and sufficient in vitamin A (FeivAs) and insufficient in both iron and vitamin A (FeivAi). After 6 weeks rats showed no significant effects of variations in vitamin A on the expression of Hamp relative to the control group (FesvAi: 1.37-fold; FesvAd: 1.22-fold); however, iron deficiency showed significant reduction on it relative to the control group (FeivAs: 71.4-fold, P = 0.0004; FeivAi: 16.1-fold, P = 0.0008). Vitamin A deficiency (FesvAd) affects expression of Fth1 independent of low dietary iron in spleen (0.29-fold, P = 0.002) and duodenum (5.15-fold, P = 0.02). Variations of dietary iron and vitamin A showed significant effects relative to the control group for expression of Tfrc in spleen (FesvAd: 0.18-fold, P = 0.01; FeivAs: 0.24-fold, P < 0.0001; FeivAi: 0.42-fold, P = 0.014), Ireb2 in spleen (FeivAs: 3.7-fold, P < 0.0001; FeivAi: 2.9-fold, P < 0.0001) and Ireb2 in duodenum (FeivAs: 2.68-fold, P = 0.012; FeivAi: 2.60-fold, P = 0.014). These results show that vitamin A and iron must be supplied together to regulate some of the main biomarkers of iron metabolism as a strategy to reduce prevalence of iron deficiency anemia.

Keywords: anemia; hepcidin; retinol; vitamin A deficiency.

This study was designed to identify early-response genes of chemical-induced renal carcinogenicity for the prediction of chemical carcinogenicity in rats. We conducted a 28-day repeated-dose test in male Crl:CD (SD) rats with 12 carcinogens and 10 non-carcinogens as the training dataset, and five carcinogens and five non-carcinogens as the validation dataset. Renal gene expression profiles were analyzed by using a microarray. Fifteen candidate genes were selected from the gene expression profiles of the training dataset as genes that showed specific expression in response to carcinogens. To assess the prediction performance of the candidate genes for renal carcinogenicity, a prediction formula was developed on the basis of the gene expression data. When this formula was applied to the training dataset to check its predictive performance, all of the carcinogens and non-carcinogens were predicted correctly; the prediction formula was then applied to the validation dataset, and five carcinogens and four non-carcinogens were correctly predicted. However, 4-Hydroxy-m-phenylenediammonium dichloride (AMIDOL), a known non-renal carcinogen, was judged as positive. Therefore, the accuracy of the prediction formula for renal carcinogenicity was 100% for the training dataset and 90% for the validation dataset. Among the predictive genes, Hamp and Ranbp1 are known to be important for cell growth and cell cycle regulation, which are important events in carcinogenesis. Given our current limited knowledge of the genes responsible for renal carcinogenesis, the identification of candidate genes of chemical-induced renal carcinogenicity by use of this gene expression-based prediction method represents a promising advance in renal carcinogen identification.

OBJECTIVE

To determine the regulation of human hepcidin (HAMP) and mouse hepcidin (hepcidin-1 and hepcidin-2) gene expression in the liver by apoptosis using in vivo and in vitro experimental models.

METHODS

For the induction of the extrinsic apoptotic pathway, HepG2 cells were treated with various concentrations of CH11, an activating antibody for human Fas receptor, for 12 h. Male C57BL/6NCR and C57BL/6J strains of mice were injected intraperitoneally with sublethal doses of an activating antibody for mouse Fas receptor, Jo2. The mice were anesthetized and sacrificed 1 or 6 h after the injection. The level of apoptosis was quantified by caspase-3 activity assay. Liver injury was assessed by measuring the levels of ALT/AST enzymes in the serum. The acute phase reaction in the liver was examined by determining the expression levels of IL-6 and SAA3 genes by SYBR green quantitative real-time PCR (qPCR). The phosphorylation of transcription factors, Stat3, Smad4 and NF-κB was determined by western blotting. Hepcidin gene expression was determined by Taqman qPCR. The binding of transcription factors to hepcidin-1 promoter was studied using chromatin immunoprecipitation (ChIP) assays.

RESULTS

The treatment of HepG2 cells with CH11 induced apoptosis, as shown by the significant activation of caspase-3 (P < 0.001), but did not cause any significant changes in HAMP expression. Short-term (1 h) Jo2 treatment (0.2 μg/g b.w.) neither induced apoptosis and acute phase reaction nor altered mRNA expression of mouse hepcidin-1 in the livers of C57BL/6NCR mice. In contrast, 6 h after Jo2 injection, the livers of C57BL/6NCR mice exhibited a significant level of apoptosis (P < 0.001) and an increase in SAA3 (P < 0.023) and IL-6 (P < 0.005) expression in the liver. However, mRNA expression of hepcidin-1 in the liver was not significantly altered. Despite the Jo2-induced phosphorylation of Stat3, no occupancy of hepcidin-1 promoter by Stat3 was observed, as shown by ChIP assays. Compared to C57BL/6NCR mice, Jo2 treatment (0.2 μg/g b.w.) of C57BL/6J strain mice for 6 h induced a more prominent activation of apoptosis, liver injury and acute phase reaction. Similar to C57BL/6NCR mice, the level of liver hepcidin-1 mRNA expression in the livers of C57BL/6J mice injected with a sublethal dose of Jo2 (0.2 μg/g b.w.) remained unchanged. The injection of C57BL/6J mice with a higher dose of Jo2 (0.32 μg/g b.w.) did not also alter hepatic hepcidin expression.

CONCLUSIONS

Our findings suggest that human or mouse hepcidin gene expression is not regulated by apoptosis induced via Fas receptor activation in the liver.

Hereditary type 1 HFE hemochromatosis is associated with homozygosity for the p.Cys282Tyr mutation of the HFE gene (C282Y mutation). The p.Cys282Tyr mutation of the HFE gene leads to an abnormal reduction in hepatic expression of hepcidin, a protein that appears to control the release of iron from enterocytes and macrophages towards plasma. Abnormally low hepcidin levels promote an increase in the bioavailability of plasma iron, characterized by elevated transferrin saturation and the appearance of non transferrin bound iron. This nontransferrin-bound iron is avidly taken up by the liver, heart, and pancreas, the principal target organs for systemic iron overload. The variable penetrance of this disease is related to environmental and genetic factors. Among the genetic factors, mutations of some newly identified genes may aggravate the phenotype of iron overload associated with homozygosity for the p.Cys282Tyr mutation of the HFE gene; these new genes include those of hemojuvelin (HJV), transferrin receptor 2 (TfR2), and hepcidin (HAMP).

Chemotaxis allows bacteria to sense gradients in their environment and respond by directing their swimming. Aer is a receptor that, instead of responding to a specific chemoattractant, allows bacteria to sense cellular energy levels and move towards favourable environments. In Pseudomonas, the number of apparent Aer homologues differs between the only two species it has been characterized in, Pseudomonas aeruginosa and Pseudomonas putida. Here we combined bioinformatic approaches with deletional mutagenesis in Pseudomonas pseudoalcaligenes KF707 to further characterize Aer. It was determined that the number of Aer homologues varies between zero and four throughout the genus Pseudomonas, and they were phylogenetically classified into five subgroups. We also used sequence analysis to show that these homologous receptors differ in their HAMP signal transduction domains. Genetic analysis also indicated that some Aer homologues have likely been subject to horizontal transfer. P. pseudoalcaligenes KF707 was unique among strains for having three Aer homologues as well as the receptors CttP and McpB. Phenotypic characterization in this strain showed that the most prevalent homologue of Aer was key, but not essential, for energy taxis. This study demonstrates that energy taxis in Pseudomonas varies between species and provides a new naming convention and associated phylogenetic details for Aer chemoreceptors.