The effects of early life Pb exposure on the expression of IL1-β, TNF-α and Aβ in cerebral cortex of mouse pups

ObjectiveTo investigate the effects of maternal lead (Pb) exposure on the learning and memory ability and expression of interleukin1-β (IL1-β), tumor necrosis factor (TNF-α) and beta amyloid protein (Aβ) in cerebral cortex of mice offspring.MethodsPb exposure initiated from beginning of gestation to weaning. Pb acetate administered in drinking solutions was dissolved in distilled deionized water at the concentrations of 0.1%, 0.5% and 1% groups, respectively. On the PND21, the learning and memory ability were tested by water maze test and the Pb levels were also determined by graphite furnace atomic absorption spectrometry. The expression of IL1-β, TNF-α and Aβ in cerebral cortex was measured by immunohistochemistry and western blotting.ResultsThe Pb levels in blood and cerebral cortex of all exposure groups were significantly higher than that of the control group (P < 0.05). In water maze test, the performances of 0.5% and 1% groups were worse than that of the control group (P < 0.05). The expression of IL1-β, TNF-α and Aβ was increased in Pb exposed groups than that of the control group (P < 0.05).ConclusionsThe high expression of IL1-β, TNF-α and Aβ in the cerebral cortex of pups may contribute to the impairment of learning and memory associated with maternal Pb exposure.     

Altered clearance of beta-amyloid from the cerebrospinal fluid following subchronic lead exposure in rats: Roles of RAGE and LRP1 in the choroid plexus

Formation of amyloid plaques is the hallmark of Alzheimer’s disease. Our early studies show that lead (Pb) exposure in PDAPP transgenic mice increases β-amyloid (Aβ) levels in the cerebrospinal fluid (CSF) and hippocampus, leading to the formation of amyloid plaques in mouse brain. Aβ in the CSF is regulated by the blood-CSF barrier (BCB) in the choroid plexus. However, the questions as to whether and how Pb exposure affected the influx and efflux of Aβ in BCB remained unknown. This study was conducted to investigate whether Pb exposure altered the Aβ efflux in the choroid plexus from the CSF to blood, and how Pb may affect the expression and subcellular translocation of two major Aβ transporters, i.e., the receptor for advanced glycation end-products (RAGE) and the low density lipoprotein receptor protein-1 (LRP1) in the choroid plexus. Sprague-Dawley rats received daily oral gavage at doses of 0, 14 (low-dose), and 27 (high-dose) mg Pb/kg as Pb acetate, 5 d/wk, for 4 or 8 wks. At the end of Pb exposure, a solution containing Aβ₄₀ (2.5 μg/mL) was infused to rat brain via a cannulated internal carotid artery. Subchronic Pb exposure at both dose levels significantly increased Aβ levels in the CSF and choroid plexus (p < 0.05) by ELISA. Confocal data showed that 4-wk Pb exposures prompted subcellular translocation of RAGE from the choroidal cytoplasm toward apical microvilli. Furthermore, it increased the RAGE expression in the choroid plexus by 34.1 % and 25.1 % over the controls (p < 0.05) in the low- and high- dose groups, respectfully. Subchronic Pb exposure did not significantly affect the expression of LRP1; yet the high-dose group showed LRP1 concentrated along the basal lamina. The data from the ventriculo-cisternal perfusion revealed a significantly decreased efflux of Aβ₄₀ from the CSF to blood via the blood-CSF barrier. Incubation of freshly dissected plexus tissues with Pb in artificial CSF supported a Pb effect on increased RAGE expression. Taken together, these data suggest that Pb accumulation in the choroid plexus after subchronic exposure reduces the clearance of Aβ from the CSF to blood by the choroid plexus, which, in turn, leads to an increase of Aβ in the CSF. Interaction of Pb with RAGE and LRP1 in choroidal epithelial cells may contribute to the altered Aβ transport by the blood-CSF barrier in brain ventricles.     

Altered cellular redox status,sirtuin abundance and clock gene expression in a mouse model of developmentally primed NASH

BackgroundWe have previously shown that high fat (HF) feeding during pregnancy primes the development of non-alcoholic steatohepatits (NASH) in the adult offspring. However, the underlying mechanisms are unclear.AimsSince the endogenous molecular clock can regulate hepatic lipid metabolism, we investigated whether exposure to a HF diet during development could alter hepatic clock gene expression and contribute to NASH onset in later life.MethodsFemale mice were fed either a control (C, 7% kcal fat) or HF (45% kcal fat) diet. Offspring were fed either a C or HF diet resulting in four offspring groups: C/C, C/HF, HF/C and HF/HF. NAFLD progression, cellular redox status, sirtuin expression (Sirt1, Sirt3), and the expression of core clock genes (Clock, Bmal1, Per2, Cry2) and clock-controlled genes involved in lipid metabolism (Rev-Erbα, Rev-Erbβ, RORα, and Srebp1c) were measured in offspring livers.ResultsOffspring fed a HF diet developed NAFLD. However HF fed offspring of mothers fed a HF diet developed NASH, coupled with significantly reduced NAD⁺/NADH (p < 0.05, HF/HF vs C/C), Sirt1 (p < 0.001, HF/HF vs C/C), Sirt3 (p < 0.01, HF/HF vs C/C), perturbed clock gene expression, and elevated expression of genes involved lipid metabolism, such as Srebp1c (p < 0.05, C/HF and HF/HF vs C/C).ConclusionOur results suggest that exposure to excess dietary fat during early and post-natal life increases the susceptibility to develop NASH in adulthood, involving altered cellular redox status, reduced sirtuin abundance, and desynchronized clock gene expression.     

The effects of early life lead exposure on the expression of P2X7 receptor and synaptophysin in the hippocampus of mouse pups

The present study was undertaken to investigate the effects of maternal lead exposure on expression of P2X7 receptor and synaptophysin in the hippocampus of mice offspring. Lead exposure initiated from beginning of gestation to weaning. Lead acetate administered in drinking solutions was dissolved in distilled deionized water at the concentrations of 0.1%, 0.5% and 1% groups, respectively. On the 21st postnatal day, the Pb levels were also determined by graphite furnace atomic absorption spectrometry. The expression of P2X7 receptor and synaptophysin in hippocampus was examined by immunohistochemistry and Western blotting. The lead levels in blood and hippocampus of all lead exposure groups were significantly higher than that of the control group (P < 0.05). Compared with the control group, the expression of P2X7 receptor was increased in lead exposed groups (P < 0.05), but the expression of synaptophysin was decreased (P < 0.05). The high expression of P2X7 receptor and low expression of synaptophysin in the hippocampus of pups may contribute to the neurotoxicity associated with maternal Pb exposure.     

Urolithin A suppresses glucolipotoxicity-induced ER stress and TXNIP/NLRP3/IL-1β inflammation signal in pancreatic β cells by regulating AMPK and autophagy

BackgroundPancreatic inflammation plays a key role in diabetes pathogenesis and progression. Urolithin A (UA), an intestinal flora metabolite of pomegranate, has anti-diabetic, anti-inflammatory and kidney protection effects among others. However, its effects on pancreatic inflammation and the potential mechanisms have not been clearly established.PurposeThis study aimed at investigating the molecular mechanisms of UA anti-pancreatic inflammation under a diabetic environment.MethodsDiabetes induction in male C57BL/6 mice was achieved by a high fat diet and intraperitoneal streptozotocin injections. Then, diabetic mice were orally administered with UA for 8 weeks. In vitro, endoplasmic reticulum stress and MIN6 pancreatic β cell inflammation were induced using 25 mM glucose and 0.5 mM palmitic acid. The effects of UA were evaluated by immunohistochemistry, Western blot, and enzyme linked immunosorbent assays. Finally, the underlying mechanisms were elucidated using an autophagy inhibitor (chloroquine, CQ) and an AMPK inhibitor (dorsomorphin dihydrochloride).ResultsUA significantly inhibited IL-1β secretion and TXNIP/NLRP3 expression in the pancreas of diabetic mice and in MIN6 pancreatic cells. UA downregulated the ER stress protein, p-PERK, and promoted AMPK phosphorylation. UA activated autophagy to inhibit TXNIP/NLRP3 IL-1β inflammatory signal, an effect that was reversed by CQ. Dorsomorphin 2HCL, reversed the autophagy-activation and anti-inflammatory effects of UA. Verapamil, clinically applied as an antiarrhythmic drug, is a TXNIP inhibitor for prevention of beta cell loss and diabetes development, but limited by its cardiac toxicity. In this study, verapamil (as positive control) inhibited NLRP3 /IL-1β signaling in MIN6 cells. Inhibitory effects of UA on TXNIP and IL-1β were weaker than those of verapamil (both at 50 μM, p < 0.05, p < 0.01). Conversely, inhibitory effects of UA on p62 were stronger, relative to those of verapamil (p < 0.05), and there were no differences in AMPK activation and LC3 enhancement effects between UA and verapamil.ConclusionUA is a potential anti-pancreatic inflammation agent that activates AMPK and autophagy to inhibit endoplasmic reticulum stress associated TXNIP/NLRP3/IL-1β signal pathway.     

Dioscin and methylprotodioscin isolated from the root of Asparagus cochinchinensis suppressed the gene expression and production of airway MUC5AC mucin induced by phorbol ester and growth factor

BackgroundThe root of Asparagus cochinchinensis (Lour.) Merr. has been utilized as mucoregulators and expectorants for controlling the airway inflammatory diseases in folk medicine.Hypothesis/purposeWe investigated whether dioscin and methylprotodioscin isolated from the root of Asparagus cochinchinensis (Lour.) Merr. suppress the gene expression and production of airway MUC5AC mucin induced by phorbol ester and growth factor.Study designConfluent NCI-H292 cells were pretreated with dioscin or methylprotodioscin for 30 min and then stimulated with EGF or PMA for 24 h. The MUC5AC mucin gene expression was measured by RT-PCR. Production of MUC5AC mucin protein was measured by ELISA.Results(1) Dioscin and methylprotodioscin suppressed the expression of MUC5AC mucin gene induced by EGF or PMA; (2) dioscin suppressed the production of MUC5AC mucin induced by either EGF at 10⁻⁵ M (p < 0.05) and 10⁻⁶ M (p < 0.05) or PMA at 10⁻⁴ M (p < 0.05), 10⁻⁵ M (p < 0.05) and 10⁻⁶ M (p < 0.05); (3) methylprotodioscin also suppressed the production of MUC5AC mucin induced by either EGF at 10⁻⁴ M (p < 0.05) or PMA at 10⁻⁴ M (p < 0.05).ConclusionThese results suggest that dioscin and methylprotodioscin isolated from the root of Asparagus cochinchinensis suppress the gene expression and production of MUC5AC mucin, by directly acting on airway epithelial cells, and the results are consistent with the traditional use of Asparagus cochinchinensis as remedy for diverse inflammatory pulmonary diseases.     

Health of children born to childhood cancer survivors: Participant characteristics and methods of the Multicenter Offspring Study

IntroductionResearch on childhood cancer survivor offspring has been limited to genetic disease occurrence, malformations or non-hereditary cancers. However, previous surveys indicated that survivors harbor fears about their (prospective) children's overall health. Our Multicenter Offspring Study examined extensive health aspects in children born to survivors and their siblings providing comprehensive information to be used in patient counseling to elucidate and alleviate existing concerns.MethodsUsing a specifically designed questionnaire, childhood cancer survivors and their siblings were surveyed on their offspring’s health (Supplementary material). Recruitment strategies depended on local infrastructures and standards of participating centers, including registry-based and direct approaches. Group differences were tested non-parametrically and effect sizes were calculated.ResultsIn total, 1126 survivors reported on 1780 offspring and 271 siblings reported on 441 offspring. Response rates ranged from 32.1% (Czech Republic) to 85.0% (Austria). Respondents were more likely to be female (p = .007), older at time of survey (p < .001), diagnosed 1980–1999 (p < .001) and treated with chemotherapy (p < .001). Compared to siblings, survivors were younger at time of survey (35 years vs. 39 years, p < .001) and at first birth (29 years vs. 30 years, p < .001). Survivor and sibling offspring only differed in terms of age at survey (6.3 years vs. 8.9 years, p < .001).ConclusionThe Multicenter Offspring Study investigates a wide variety of health aspects in offspring born to survivors and their siblings in five European countries. Our study cohorts form a solid basis for future analyses; yet, certain limitations, due to differences in approach among participating centers, must be considered when interpreting findings.     

High SLC7A11 expression in normal skin of melanoma patients

BackgroundMelanoma is one of the highest metastatic cancers and its incidence is rapidly increasing. A great effort has been devoted to determine gene mutations and expression profiles in melanoma cells, but less attention has been given to the possible influence of melanin synthesis in melanocytes on melanomagenesis. SLC7A11 encodes the cystine/glutamate antiporter xCT and its expression increases the antioxidant capacity of cells by providing cysteine that may be used for glutathione (GSH) synthesis. Melanocytes, however, can also use cysteine for pheomelanin synthesis and pigmentation. Therefore, pheomelanin synthesis may lead to chronic oxidative stress. Possible consequences of this for melanomagenesis have never been explored.MethodsWe quantified the expression of SLC7A11 and other genes that are involved in the synthesis of pheomelanin but do not regulate the transport of cysteine from the extracellular medium to the cytosol (CTNS, MC1R, ASIP and SLC45A2) in non-tumorous skin of 45 patients of cutaneous melanoma and 50 healthy individuals. We controlled for the effects of Fitzpatrick skin type, age, gender, body mass, frequency of sun exposure and sunburns and number of melanocytic nevi, as well as for the intrinsic antioxidant capacity as given by the expression of the gene NFE2L2.ResultsThe expression of SLC7A11, but not of the other genes, was significantly higher in melanoma patients than in healthy individuals. This was independent of phenotypic factors and antioxidant capacity, thus supporting an effect of pheomelanin-induced oxidative stress on melanomagenesis.ConclusionOur findings indicate that SLC7A11 downregulation in normal epidermal melanocytes may represent a preventive treatment against melanoma.     

Relationship of lead and essential elements in whole blood from school-age children in Nanning,China

ObjectivesThe aim of this study was to investigate blood lead level and its relationship to essential elements (zinc, copper, iron, calcium and magnesium) in school-age children from Nanning, China.MethodsA total of 2457 children aged from 6 to 14 years were enrolled in Nanning, China. The levels of lead (Pb), zinc (Zn), copper (Cu), iron (Fe), calcium (Ca) and magnesium (Mg) were determined by an atomic absorption spectrometer.ResultsThe mean blood lead level (BLL) was 57.21 ± 35.00 μg/L. 188 (7.65%) asymptomatic children had toxic lead level higher than 100 μg/L. The school-age boys had similar lead level among different age groups, while the elder girls had less BLL. The blood Zn and Fe were found to be increased in the boys with elevated BLL, but similar trends were not observed in the girls. Positive correlations between Pb and Fe or Mg (r = 0.112, 0.062, respectively, p < 0.01) and a negative correlation between Pb and Ca (r = −0.047, p < 0.05) were further established in the studied children.ConclusionsLead exposure in school-age children was still prevalent in Nanning. The boys and girls differed in blood levels of lead and other metallic elements. Lead exposure may induce metabolic disorder of other metallic elements in body.     

Increasing vitamin A in post-weaning diets reduces food intake and body weight and modifies gene expression in brains of male rats born to dams fed a high multivitamin diet

High multivitamin gestational diets (HV, 10-fold AIN-93G levels) increase body weight (BW) and food intake (FI) in rat offspring weaned to a recommended multivitamin (RV), but not to a HV diet. We hypothesized that high vitamin A (HA) alone, similar to HV, in post-weaning diets would prevent these effects of the HV maternal diet consistent with gene expression in FI and reward pathways. Male offspring from dams fed HV diets were weaned to a high vitamin A (HA, 10-fold AIN-93G levels), HV or RV diet for 29 weeks. BW, FI, expression of genes involved in regulation of FI and reward and global and gene-specific DNA methylation of pro-opiomelanocortin (POMC) in the hypothalamus were measured. Both HV and HA diets slowed post-weaning weight gain and modified gene expression in offspring compared to offspring fed an RV post-weaning diet. Hypothalamic POMC expression in HA offspring was not different from either HV or RV, and dopamine receptor 1 was 30% (P<.05) higher in HA vs. HV, but not different from RV group. Hippocampal expression of serotonin receptor 1A (40%, P<.01), dopamine receptor 2 (40%, P<.05) and dopamine receptor 5 (70%, P<.0001) was greater in HA vs. RV fed pups and is 40% (P<.01), 50% (P<.05) and 40% (P<.0001) in HA vs. HV pups, respectively. POMC DNA methylation was lower in HA vs. RV offspring (P<.05). We conclude that high vitamin A in post-weaning diets reduces post-weaning weight gain and FI and modifies gene expression in FI and reward pathways.     

Folate deprivation induces cell cycle arrest at G0/G1 phase and apoptosis in hippocampal neuron cells through down-regulation of IGF-1 signaling pathway

Folate deficiency contributes to impaired adult hippocampal neurogenesis, yet the mechanisms remain unclear. Here we use HT-22 hippocampal neuron cells as model to investigate the effect of folate deprivation (FD) on cell proliferation and apoptosis, and to elucidate the underlying mechanism. FD caused cell cycle arrest at G0/G1 phase and increased the rate of apoptosis, which was associated with disrupted expression of folate transport and methyl transfer genes. FOLR1 and SLC46A1 were (P < 0.01) down-regulated, while SLC19A1 was up-regulated (P < 0.01) in FD group. FD cells exhibited significantly (P < 0.05) higher protein content of BHMT, MAT2b and DNMT3a, as well as increased SAM/SAH concentrations and global DNA hypermethylation. The expression of the total and all the 3 classes of IGF-1 mRNA variants was significantly (P < 0.01) down-regulated and IGF-1 concentration was decreased (P < 0.05) in the culture media. IGF-1 signaling pathway was also compromised with diminished activation (P < 0.05) of STAT3, AKT and mTOR. CpG hypermethylation was detected in the promoter regions of IGF-1 and FOLR1 genes, while higher SLC19A1 mRNA corresponded to hypomethylation of its promoter. IGF-1 supplementation in FD media significantly abolished FD-induced decrease in cell viability. However, IGF-1 had limited effect in rescuing the cell phenotype when added 24 h after FD. Taken together, down-regulation of IGF-1 expression and signaling is involved in FD-induced cell cycle arrest and apoptosis in HT-22 hippocampal neuron cells, which is associated with an abnormal activation of methyl transfer pathway and hypermethylation of IGF-1 gene promoter.     

The shedding-derived soluble receptor for advanced glycation endproducts sustains inflammation during acute Pseudomonas aeruginosa lung infection

BackgroundThe membrane-bound isoform of the receptor for advanced glycation end products (FL-RAGE) is primarily expressed by alveolar epithelial cells and undergoes shedding by the protease ADAM10, giving rise to soluble cleaved RAGE (cRAGE). RAGE has been associated with the pathogenesis of several acute and chronic lung disorders. Whether the proteolysis of FL-RAGE is altered by a given inflammatory stimulus is unknown.Pseudomonas aeruginosa causes nosocomial infections in hospitalized patients and is the major pathogen associated with chronic lung diseases.MethodsP. aeruginosa was injected in Rage −/− and wild-type mice and the impact on RAGE expression and shedding, levels of inflammation and bacterial growth was determined.ResultsAcute P. aeruginosa lung infection in mice induces a reduction of the active form of ADAM10, which determines an increase of FL-RAGE expression on alveolar cells and a concomitant decrease of pulmonary cRAGE levels. This was associated with massive recruitment of leukocytes and release of pro-inflammatory factors, tissue damage and relocation of cRAGE in the alveolar and bronchial cavities. The administration of sRAGE worsened bacterial burden and neutrophils infiltration. RAGE genetic deficiency reduced the susceptibility to P. aeruginosa infection, mitigating leukocyte recruitment, inflammatory molecules production, and bacterial growth.ConclusionsThese data are the first to suggest that inhibition of FL-RAGE shedding, by affecting the FL-RAGE/cRAGE levels, is a novel mechanism for controlling inflammation to acute P. aeruginosa pneumonia. sRAGE in the alveolar space sustains inflammation in this setting.General significanceRAGE shedding may determine the progression of inflammatory lung diseases.     

The role of activated NLRP3 inflammatory body in acute kidney injury in rats caused by sepsis and NLRP3-TXNIP signaling pathway

ObjectiveThe model of acute renal injury (AKI) induced by sepsis in rats was established by abdominal resection through surgical suture. The activation mechanism of nod-like receptor with pyrin domain containing 3 (NLRP3) inflammatory corpuscle in AKI induced by sepsis was analyzed.MethodsHere, 60 male rats were selected and divided into two groups, including sham-operated group (NO-OPs group, n = 15) and sepsis group (CELP group, n = 45). In order to examine each index of CELP group, four time points (10, 20, 30, and 40 h) were set as control. In NO-OPs group, only abdominal resection through surgical suture was carried out. The expression levels of NLRP3, apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain (ASC), caspase-1, and the expression level of NLRP3-TXNIP signaling pathway were measured by immunohistochemistry, Western blotting, immunoprecipitation, and mito-TEMPO (a mitochondria-targeted antioxidant) 40 h after operation and 10, 20, 30, and 40 h post-operation in CELP group. Herein, 40 h post-operation in NO-OPs group and 10, 20, 30, and 40 h post-operation in CELP group, peripheral blood samples were collected.ResultsCompared with NO-OPs group, the levels of serum creatinine (Scr) and blood urea nitrogen (BUN) in CELP group were increased (P < 0.05). Compared with NO-OPs group, the expression levels of interleukin-1β (IL-1β), NLRP3, ASC, and caspase-1 in CELP group were increased (P < 0.05). The expression level of TXNIP in renal tubular epithelial cells in rats was up-regulated. There was a positive correlation between TXNIP and NLRP3. The binding of NLRP3-TXNIP signaling pathway could be inhibited by siRNA transfection or mito-TMPO, and the activity of NLRP3 inflammatory bodies could be inhibited as well.ConclusionActivation of NLRP3 inflammatory corpuscles could promote AKI induced by sepsis. Simultaneously, renal injury may lead to the production of mitochondrial reactive oxygen species (mROS), which may induce the binding of TXNIP to NLRP3.     

Th1/Th2/Th17/Treg cytokine imbalance in systemic lupus erythematosus (SLE) patients: Correlation with disease activity

AimImbalance of T-helper-cell (TH) subsets (TH1/TH2/TH17) and regulatory T-cells (Tregs) is suggested to contribute to the pathogenesis of Systemic lupus erythematosus (SLE). Therefore, we evaluated their cytokine secretion profile in SLE patients and their possible association with disease activity.MethodsSixty SLE patients, 24 rheumatoid arthritis (RA) patients and 24 healthy volunteers were included in this study. Demographic, clinical, disease activity and serological data were prospectively assessed. Plasma cytokines levels of TH1 (IL-12, IFN-γ), TH2 (IL-4, IL-6, IL-10), TH17 (IL-17, IL-23) and Treg (IL-10 and TGF-β) were measured by enzyme linked immunosorbent assays (ELISA).ResultsSLE patients were found to have significantly higher levels of IL-17 (p < 0.001), IL-6 (p < 0.01), IL-12 (p < 0.001) and IL-10 (p < 0.05) but comparable levels of IL-23 and IL-4 and slight reduction (but statistically insignificant) of TGF-β levels compared to controls. IL-6, IL-10 and IL-17 were significantly increased (p < 0.05) with disease activity. The RA group exhibited significantly higher levels of plasma IL-4 (p < 0.01), IL-6 (p < 0.05), IL-17 (p < 0.001), IL-23 (p < 0.01) and TGF-β (p < 0.5) and lower IFN-γ (p < 0.001) and IL-10 (p < 0.01) than those of healthy subjects.ConclusionOur study showed a distinct profile of cytokine imbalance in SLE patients. Reduction in IFN-γ (TH1) and TGF-β1 (Treg) with the elevation in IL-6 and IL-17 (TH17) could imply skewing of T-cells toward TH17 cells. Breaking TH17/Treg balance in peripheral blood may play an important role in the development of SLE and could be responsible for an increased pro-inflammatory response especially in the active form of the disease.     

Upregulation of NKG2D ligands in acute lymphoblastic leukemia and non-Hodgkin lymphoma cells by romidepsin and enhanced in vitro and in vivo natural killer cell cytotoxicity

Background aimsThere is a critical need to prevent and/or treat hematological relapse after allogeneic hematopoietic stem cell transplantation. The activating NKG2D receptor expressed on natural killer (NK) cells, when engaged by its corresponding ligands (MIC A/B), activates NK cells to become cytotoxic against malignant cells.MethodsWe incubated acute lymphoblastic leukemia and non-Hodgkin lymphoma cells for 24 h with 10 ng/mL of romidepsin. Flow cytometry was performed to demonstrate changes in surface expression of NKG2D ligands MIC A/B. In vitro and in vivo cytotoxicity was measured by means of modified Europium assay, and non-obese diabetic/severe combined immunodeficiency mice were xenografted with RS 4:11 cells.ResultsWe demonstrated an approximately 50, 200, 1300 and 180-fold increase in the number of cells positive for the surface expression of MIC A/B in RS 4:11 (P < 0.001), REH (P < 0.001), Ramos (P < 0.001) and Jurkat cells (P < 0.001), respectively. We further demonstrated a significant increase in NK cell–mediated in vitro cytotoxicity against RS 4:11 (P < 0.004), Ramos (P < 0.05), Jurkat (P < 0.001) and REH cells (P < 0.01), respectively. Romidepsin-mediated NK cytotoxicity was blocked by pre-incubating NK cells with anti-NKG2D-Fc in RS 4:11 (P < 0.03) and Ramos cells (P < 0.01), respectively. Finally, non-obese diabetic/severe combined immunodeficiency mice xenografted with RS 4:11 cells had a significant increase in survival (P < 0.02) in mice treated with romidepsin and interleukin-2–activated NK cells compared with each of these other treatment groups.ConclusionsRomidepsin significantly enhanced in vitro and in vivo NK cell cytotoxicity mediated in part by increased MIC A/B expression on malignant cells. This translational approach of the use of romidepsin and interleukin-2–activated NK cells should be considered in patients with relapsed/refractory leukemia or lymphoma.     

Increased Receptor for Advanced Glycation Endproducts Immunocontent in the Cerebral Cortex of Vitamin A-Treated Rats

Vitamin A, beyond its biological role, is an alternative choice in treating some life threatening pathologies, for instance leukemia and immunodeficiency. On the other hand, vitamin A therapy at moderate to high doses has caused concern among public health researchers due to the toxicological aspect resulting from such habit. It has been described hepatotoxicity, cognitive disturbances and increased mortality rates among subjects ingesting increased levels of vitamin A daily. Then, based on the previously reported data, we investigated here receptor for advanced glycation endproducts (RAGE) immunocontent and oxidative damage levels in cerebral cortex of vitamin A-treated rats at clinical doses (1,000–9,000 IU/kg day⁻¹). RAGE immunocontent, as well as oxidative damage levels, were observed increased in cerebral cortex of vitamin A-treated rats. Whether increased RAGE levels exert negative effects during vitamin A supplementation it remains to be investigated, but it is very likely that deleterious consequences may arise from such alteration.     

Vocal fold fibroblasts immunoregulate activated macrophage phenotype

Recent evidence suggests that fibroblasts play a critical role in regulating inflammation during wound healing because they express several inflammatory mediators in response to bacteria. The objective of this study was to analyze the effects of lipopolysaccharide (LPS) on the immunomodulatory properties of vocal fold fibroblasts (VFFs) derived from polyps, scar and normal tissue co-cultured with macrophages, to provide insight into their interactions during the inflammatory process. Fibroblasts were co-cultured with CD14+ monocytes and after 7 days, wells were treated with LPS for 24 and 72 h. Culture supernatants were collected and concentrations of TNF-α, IL-6, IL-8, IL-10, IL-12, IL-1β and MCP-1 were quantified by ELISA. Normal VFF and CD14+ monocultures were used as controls. Twenty-four hours after LPS activation, macrophages co-cultured with polyp VFF had significantly increased expression of TNF-α, IL-1β, IL-12 and IL-10 compared to controls (p < 0.0001). In contrast, macrophages co-cultured with scar VFF had significantly lower expression of TNF-α, IL-1β and IL-12 with significantly higher IL-10 compared to control (p < 0.0001). After 72 h, macrophages co-cultured with polyp VFF increased expression of TNF-α, IL-1β, IL-10, IL-6, IL-8, MCP-1 and TGF-β (p < 0.01) and macrophages co-cultured with scar VFF significantly decreased their expression of IL-1β and IL-12 compared to control (p < 0.0001). Scar VFF at both time points produced significantly lower levels of IL-8, MCP-1, IL-6 and TGF-β compared to controls (p < 0.05). Based on our findings, VFF and macrophages secrete several inflammatory mediators that modify their diverse functions. Polyp and scar VFF may play a role in regulating abnormal inflammatory responses, which could result in excessive ECM deposition that disrupts the function of the vocal folds.     

Genetic variation in ESR2 and estrogen receptor-beta expression in lung tumors

Objective: To investigate the association between inherited variation in the estrogen receptor beta (ERβ) gene (ESR2) and ERβ lung tumor expression, a phenotype that possibly affects survival differently in men and women. Methods: We genotyped 135 lung cancer patients for 22 ESR2 single nucleotide polymorphisms (SNPs) and measured nuclear and cytoplasmic ERβ expression by immunohistochemistry (IHC) in their primary lung tumor. Distributing Allred ERβ IHC scores according to ESR2 genotype classified under a dominant genetic model, we used rank sum tests to identify ESR2 SNPs significantly associated (p < 0.05) with ERβ expression. Results: 35%, 35%, and 29% of lung tumors showed no/low (Allred < 6), intermediate (Allred 6–7), and maximal (Allred 8) cytoplasmic ERβ expression, whereas 13%, 27%, and 60% showed no/low, intermediate, and maximal nuclear ERβ expression. For SNPs rs8021944, rs1256061 and rs10146204, ERβ expression was higher according to the rank sum test in lung tumors from patients with at least one minor allele. For each of these three SNPs, the odds of maximal (Allred 8) relative to no/low (Allred < 6) ERβ expression was 3-fold higher in tumors from patients with at least one minor allele than in tumors from patients homozygous for the common allele. Conclusion: Inherited variability in ESR2 may determine ERβ lung tumor expression.