Resveratrol differentially modulates immune responses in human THP-1 monocytes and macrophages.

Resveratrol (Res), a natural polyphenol compound found in grapes and red wine, has been shown to exhibit anti-inflammatory, antioxidant, and anticarcinogenic effects. However, proinflammatory/tumor-promoting properties of Res have also been reported, rendering the polyphenol's reported therapeutic benefits less convincing and controversial. To evaluate the underlying plausible factors contributing to the differential immunomodulatory effects imparted by Res, herein, we investigated, at both physiological and pharmacological doses, the in vitro effects of Res on cell survival/proliferation, inflammatory genes, and cytokine production in human monocytic cell line (THP-1) and phorbol 12-myristate 13-acetate differentiated human THP-1-derived macrophages. We hypothesized that the differential effects observed in monocytes and macrophages may largely depend on dietary vs pharmacological doses of Res, duration of treatment, and the target cells it acts upon. Our data showed that Res, at physiological concentrations, inhibited proliferation of THP-1 monocytes with S phase arrest. On the other hand, at pharmacological concentrations, Res induced cell apoptosis and caused G0/G1 phase arrest. Additionally, Res showed differential effects on proinflammatory cytokine expression and production measured by reverse-transcription polymerase chain reaction and enzyme-linked immunosorbent assay, respectively, in THP-1 monocytes vs macrophages: promoting inflammation in monocytes while exhibiting anti-inflammatory effects in macrophages. Comparative analysis on Res and 2 other phytochemicals, pterostilbene and genistein, revealed that the immunomodulatory effects of Res were consistent with those observed in pterostilbene and not genistein. Our results reveal a pleiotropic immunomodulatory property of Res that is dose-time-target cell-dependent and thus serve as a caution for the use of Res in the treatment of inflammatory diseases.

Bone metabolism markers are associated with neck circumference in adult Arab women.

The study aimed to determine whether neck circumference is associated with bone metabolism markers among adult Arab women and found modest but significant associations with bone resorption markers, suggesting that neck circumference, a surrogate measure of upper subcutaneous fat, influences bone turnover expression among adult females. INTRODUCTION: Body fat distribution is associated with decreased bone resorption and neck circumference (NC), a surrogate measure for upper body fat, has never been tested as a marker that can reflect bone turnover. This is the first study aimed to analyze the associations between NC and several bone biomarkers among adult Saudi women. METHODS: This cross-sectional study included a total of 265 middle-aged Saudi women [86 non-obese (mean age 52.7 ± 8.1; mean BMI 26.9 ± 2.3) and 179 obese (mean age 50.6 ± 7.5; mean BMI 35.7 ± 4.5)] recruited from primary care centers in Riyadh, Saudi Arabia. Anthropometrics included BMI, NC, waist and hip circumferences, total body fat percentage (%), and blood pressure. Biochemical parameters included glucose and lipid profile which were measured routinely. Serum levels of 25(OH) D, parathyroid hormone, RANKl, sclerostin, C-terminal telopeptide of collagen I (CTX-I), Dkk1, IL1ß, osteoprotegerin, osteopontin, and osteocalcin were measured using commercially available assays. RESULTS: In all groups, NC was inversely associated with PTH (R = - 0.22; p < 0.05) and positively associated with osteoprotegerin (R = 0.20; p < 0.05) even after adjustments for age and BMI. Using all anthropometric indices as independent variables showed that only NC explained the variance perceived in CTX-I (p = 0.049). In the non-obese, waist-hip ratio (WHR) was significantly associated with sclerostin (R = 0.40; p < 0.05) and body fat was significantly associated with osteopontin (R = 0.42; p < 0.05). CONCLUSION: NC is modestly but significantly associated with bone biomarkers, particularly the bone resorption markers, among adult Arab women. The present findings highlight the importance of NC as measure of upper body subcutaneous fat in influencing bone biomarker expression in adult females.

Correction to: Effects of obesity and metabolic syndrome on cardiovascular outcomes in pediatric kidney transplant recipients: a longitudinal study.

As originally published, this article contained errors owing to oversights in typesetting. The article has now been amended accordingly.

Increased carotid intima-media thickness in African American pediatric kidney transplant recipients.

Early signs of subclinical CV dysfunction can be detected by ultrasound for CIMT. Although A-A are at high risk for CV disease, CIMT of A-A kidney transplant recipients has not been previously investigated. The aim of this prospective, controlled, longitudinal study was to investigate determinants of CIMT in a multiracial pediatric kidney transplant population, with a focus on A-A. Transplant recipients (n = 42) had BMI, waist-to-height ratio, fasting glucose, lipid panel, HbA1c%, and CIMT measured at 1, 18, and 30 months post-transplant. Twenty-four healthy children (14 A-A) served as controls. CIMT of A-A transplant (0.49, 0.49, and 0.48 mm) was higher than non-AA transplant (0.43, 0.44, and 0.44 mm) at 1, 18, and 30 months and higher than A-A controls (0.47 mm). Hyperparathyroidism prior to transplant predicted high CIMT-for-race. A-A race was associated with 10% higher CIMT vs non-A-A transplant. Metabolic syndrome was associated with 0.03 ± 0.01 mm increase in CIMT among A-A transplant recipients only. In conclusion, A-A kidney transplant recipients have increased CIMT. Metabolic syndrome disproportionately affects CIMT of A-A children post-transplant. Identification of subclinical CV damage, detected by CIMT, may provide an opportunity for early detection of CV risk in this vulnerable population.

Effects of obesity and metabolic syndrome on cardiovascular outcomes in pediatric kidney transplant recipients: a longitudinal study.

BACKGROUND: Obesity and metabolic syndrome (MS) are common after kidney transplantation, but their contribution to adverse cardiovascular (CV) outcomes in children are not well known. A prospective, controlled, longitudinal cohort study was conducted to investigate the effects of obesity and MS on left ventricular hypertrophy (LVH) and myocardial strain in pediatric kidney transplant recipients. METHODS: Transplant recipients (n = 42) had anthropometrics [body mass index (BMI), waist circumference, waist-to-height ratio], biochemical parameters (fasting glucose, lipid panel, HbA1c%), and echocardiogram with speckle tracking analysis for strain measured at 1, 18, and 30 months post-transplant. Additionally, 35 pre-transplant echocardiograms were analyzed retrospectively. Healthy children (n = 24) served as controls. RESULTS: Waist-to-height ratio detected abdominal obesity in 46% of transplant patients, whereas only 8.1% were identified as obese by waist circumference. Ejection fraction and fractional shortening of the transplant group were normal. Prevalence of LVH was 35.2%, 17.1%, and 35.5% at 1, 18, and 30 months respectively. The longitudinal strain of transplant group was worse than controls at all time points (p < 0.001). Hemodialysis was independently associated with 21% worse longitudinal strain during the pre-transplant period (p = 0.04). After transplantation, obesity, MS, and systolic hypertension predicted increased odds of LVH (p < 0.04). Worse longitudinal strain was independently associated with obesity, MS, hypertension, and the combination of MS with elevated low density lipoprotein (LDL) cholesterol (p < 0.04), whereas higher estimated glomerular filtration rate (eGFR) conferred a protective effect (p < 0.001). CONCLUSION: Obesity and MS adversely affect CV outcomes after transplantation. Further studies are needed to investigate speckle tracking echocardiography as a tool for early detection of subclinical myocardial dysfunction in this population.

Methylation-sensitive amplified polymorphism analysis of Verticillium wilt-stressed cotton (Gossypium).

In this study, a methylation-sensitive amplification polymorphism analysis system was used to analyze DNA methylation level in three cotton accessions. Two disease-sensitive near-isogenic lines, PD94042 and IL41, and one disease-resistant Gossypium mustelinum accession were exposed to Verticillium wilt, to investigate molecular disease resistance mechanisms in cotton. We observed multiple different DNA methylation types across the three accessions following Verticillium wilt exposure. These included hypomethylation, hypermethylation, and other patterns. In general, the global DNA methylation level was significantly increased in the disease-resistant accession G. mustelinum following disease exposure. In contrast, there was no significant difference in the disease-sensitive accession PD94042, and a significant decrease was observed in IL41. Our results suggest that disease-resistant cotton might employ a mechanism to increase methylation level in response to disease stress. The differing methylation patterns, together with the increase in global DNA methylation level, might play important roles in tolerance to Verticillium wilt in cotton. Through cloning and analysis of differently methylated DNA sequences, we were also able to identify several genes that may contribute to disease resistance in cotton. Our results revealed the effect of DNA methylation on cotton disease resistance, and also identified genes that played important roles, which may shed light on the future cotton disease-resistant molecular breeding.

Nutritional and supranutritional levels of selenate differentially suppress prostate tumor growth in adult but not young nude mice.

The inhibitory effect of oral methylseleninic acid or methylselenocysteine administration on cancer cell xenograft development in nude mice is well characterized; however, less is known about the efficacy of selenate and age on selenium chemoprevention. In this study, we tested whether selenate and duration on diets would regulate prostate cancer xenograft in nude mice. Thirty-nine homozygous NU/J nude mice were fed a selenium-deficient, Torula yeast basal diet alone (Se-) or supplemented with 0.15 (Se) or 1.0 (Se+) mg selenium/kg (as Na2SeO4) for 6 months in Experiment 1 and for 4 weeks in Experiment 2, followed by a 47-day PC-3 prostate cancer cell xenograft on the designated diet. In Experiment 1, the Se- diet enhanced the initial tumor development on days 11-17, whereas the Se+ diet suppressed tumor growth on days 35-47 in adult nude mice. Tumors grown in Se- mice were loosely packed and showed increased necrosis and inflammation as compared to those in Se and Se+ mice. In Experiment 2, dietary selenium did not affect tumor development or histopathology throughout the time course. In both experiments, postmortem plasma selenium concentrations in Se and Se+ mice were comparable and were twofold greater than those in Se- mice. Taken together, dietary selenate at nutritional and supranutritional levels differentially inhibit tumor development in adult, but not young, nude mice engrafted with PC-3 prostate cancer cells.

Apoptosis-mediated inhibition of human breast cancer cell proliferation by lemon citrus extract.

Dietary phytochemicals have a variety of antitumor properties. In the present study, the antitumor activity of methanolic extract of lemon fruit (lemon extract; LE) (LE) on the MCF-7 breast cancer cell line was investigated in vitro. Apoptotic cell death was analyzed using the TUNEL assay. In addition, the apoptosis mediated by LE extract in the MCF-7 cells was associated with the increased expression of the tumor suppressor p53 and caspase-3. Additionally, the expression of a pro-apoptotic gene, bax, was increased, and the expression of an anti-apoptotic gene, bcl-2, was decreased by LE extract treatment, resulting in a shift in the Bax:Bcl-2 ratio to one that favored apoptosis. The expression of a major apoptotic gene, caspase-3, was increased by LE extract treatment. In light of the above results, we concluded that LE extract can induce the apoptosis of MCF-7 breast cancer cells via Bax-related caspase-3 activation. This study provides experimental data that are relevant to the possible future clinical use of LE to treat breast cancer.

Zinc depletion reduced Egr-1 and HNF-3beta expression and apolipoprotein A-I promoter activity in Hep G2 cells.

We examined the influence of zinc status on expression of certain transcription factors involved in regulation of apolipoprotein A-I (apoAI) expression in human hepatoblastoma Hep G2 cells. A low zinc basal medium (zinc deficient, ZD) consisting of DMEM and 10% Chelex100-treated fetal bovine serum was used to deplete cellular zinc over one passage. Cells were also cultured for one passage in medium supplemented with 0.4 (ZD0.4), 4.0 (zinc normal, ZN), 16.0 (zinc adequate, ZA), or 32.0 microM zinc (zinc supplemented, ZS). Compared with ZN cells, cellular zinc levels were 43 and 31% lower in ZD and ZD0.4 cells but 70 and 146% higher in ZA and ZS cells, respectively. Supplementation of 0.4 microM zinc significantly increased DNA contents per plate, from 65% in ZD cells to 83% in ZD0.4 cells compared with ZN cells. Addition of >4 microM zinc in medium did not further increase DNA contents. The proportion of cells in G(1)/S and S phase was about fourfold higher and threefold lower, respectively, in ZD cells compared with ZN and other groups. Nuclear Egr-1 protein was markedly decreased in ZD and ZD0.4 cells. Moreover, hepatocyte nuclear factor (HNF)-3beta was severely degraded in ZD and ZD0.4 cells. In contrast, HNF-4alpha remained stable in all groups and was not significantly lower in ZD and ZD0.4 cells. Furthermore, downregulation of trans-acting factor Egr-1 and cleavage of HNF-3beta were associated with reduction of apoAI promoter activity in zinc-deficient Hep G2 cells. Thus zinc is critical in transcriptional regulation of apoAI gene expression in hepatocytes.

Zinc status affects p53, gadd45, and c-fos expression and caspase-3 activity in human bronchial epithelial cells.

This study was designed to examine the influence of zinc depletion and supplementation on the expression of p53 gene, target genes of p53, and caspase-3 activity in normal human bronchial epithelial (NHBE) cells. A serum-free, low-zinc medium containing 0.4 micromol/l of zinc [zinc deficient (ZD)] was used to deplete cellular zinc over one passage. In addition, cells were cultured for one passage in media containing 4.0 micromol/l of zinc [zinc normal (ZN)], which represents normal culture concentrations (Clonetics); 16 micromol/l of zinc [zinc adequate (ZA)], which represents normal human plasma zinc levels; or 32 micromol/l of zinc [zinc supplemented (ZS)], which represents the high end of plasma zinc levels attainable by oral supplementation in humans. Compared with ZN cells, cellular zinc levels were 76% lower in ZD cells but 3.5-fold and 6-fold higher in ZA and ZS cells, respectively. Abundances of p53 mRNA and nuclear p53 protein were elevated in treatment groups compared with controls (ZN). For p53 mRNA abundance, the highest increase (3-fold) was observed in ZD cells. In contrast, the highest increase (17-fold) in p53 nuclear protein levels was detected in ZS cells. Moreover, gadd45 mRNA abundance was moderately elevated in ZD and ZA cells and was not altered in ZS cells compared with ZN cells. Furthermore, the only alteration in c-fos mRNA and caspase-3 activity was the twofold increase and the 25% reduction, respectively, detected in ZS compared with ZN cells. Thus p53, gadd45, and c-fos and caspase-3 activity appeared to be modulated by cellular zinc status in NHBE cells.

Regulation of intestinal apolipoprotein B mRNA editing levels by a zinc-deficient diet and cDNA cloning of editing protein in hamsters.

This study was conducted to investigate the influence of dietary zinc on intestinal apoB mRNA editing in hamsters. Apolipoprotein B-48 (apoB-48) is synthesized from the same gene as apoB-100 by a post-transcriptional, site-specific cytidine deamination, a process known as apoB mRNA editing. A cDNA encoding the hamster apoB mRNA editing enzyme was obtained by reverse transcriptase-polymerase chain reaction (RT-PCR) and the deduced amino acid sequence was found to possess high amino acid sequence identity to apoB mRNA editing enzymes from several other species. Editing activity was detected in the small intestine and colon but, like humans, none was detected in the liver. Analysis by RT-PCR indicated that the small intestine possessed the highest expression of editing enzyme mRNA abundance, whereas both liver and small intestine expressed relatively high levels of apoB mRNA. The influence of dietary zinc on intestinal apoB mRNA editing levels was examined in Golden Syrian hamsters (7 wk old) assigned to one of the following three dietary treatments: Zn-adequate (ZA, 30 mg Zn/kg diet), Zn-deficient (ZD, <0. 5 mg Zn/kg diet), or Zn-replenished (ZDA, ZD hamsters receiving ZA diet for last 2 d) for 7 wk. Hamsters consuming the ZD diet had modestly but significantly lower intestinal editing activity than ZA hamsters. Intestinal editing activity in the ZDA group was not different from that of ZA hamsters. Data derived from these studies contribute to the understanding of lipoprotein metabolism in hamsters, a suitable model for the study of atherosclerosis.

Expression of the p53 tumor suppressor gene is up-regulated by depletion of intracellular zinc in HepG2 cells.

Expression and activation of the p53 tumor suppressor protein are modulated by various cellular stimuli. The objective of this work was to examine the influence of zinc depletion on the expression of p53 in HepG2 cells. Two different low Zn (ZD) media, Zn-free Opti-MEM and a ZD medium containing Chelex-100 treated serum, were used to deplete cellular zinc over one passage. Cellular zinc levels of ZD cells were significantly lower than in their controls in both the Opti-MEM and Chelex studies. p53 mRNA abundance was 187% higher in ZD Opti-MEM cells and >100% higher in ZD Chelex cells compared with their respective controls. To examine whether the effects were specific to zinc depletion, a third, zinc-replenished group (ZDA) was included in the Opti-MEM study in which cells were cultured in ZD media for nearly one passage before a change was made to zinc-adequate (ZA) medium for the last 24 h. Zinc levels in the ZDA cells were significantly higher than in ZD cells, and p53 mRNA abundance was normalized to control levels. Nuclear p53 protein levels were >100% higher in the ZD Opti-MEM cells than in ZA cells. Interestingly, the ZDA Opti-MEM cells had significantly lower levels of nuclear p53 protein than both the ZA and ZD cells. These data suggest that expression of p53, a critical component in the maintenance of genomic stability, may be affected by reductions in cellular zinc.

Apolipoprotein A-I gene expression is regulated by cellular zinc status in hep G2 cells.

The influence of Zn on the expression of the apolipoprotein A-I (apoA-I) gene in Hep G2 cells was examined. Zn depletion was achieved with a low-Zn (ZD) medium prepared from Zn-free growth medium (Opti), a ZD medium containing Chelex 100-extracted fetal bovine serum (CHE), and a medium containing chelator 1, 10-phenanthroline (OP). Compared with those for their respective controls, cellular Zn levels were reduced by 55, 48, and 46% and apoA-I mRNA abundances were reduced by 20, 29, and 28% in Opti, CHE, and OP systems, respectively, after one passage in ZD media or 24 h in OP medium. To establish the specificity of Zn treatment, groups of ZD cells were treated with their respective control media for the last 24 h (ZDA) or normal cells were cultured with OP medium supplemented with Zn (OP-Zn). ZDA treatments partially normalized cellular Zn levels in the Opti system and restored or elevated apoA-I mRNA levels in the Opti or CHE system, respectively. Similarly, the OP-Zn treatment restored the cellular Zn and apoA-I mRNA levels. Furthermore, one passage of culture with Zn-supplemented media in both the Opti and CHE systems resulted in higher cellular Zn and apoA-I mRNA levels than those for controls. Most significantly, short-term high-Zn induction to normal cells markedly elevated the cellular Zn (3-fold) and apoA-I mRNA (5-fold) levels. Data derived from this study strongly suggest that the expression of apoA-I is regulated by cellular Zn status.

Plasma apolipoprotein B-48, hepatic apolipoprotein B mRNA editing and apolipoprotein B mRNA editing catalytic subunit-1 mRNA levels are altered in zinc-deficient rats.

Apolipoprotein B (apoB) exists as two major isoforms and serves as an obligatory component of lipid-rich plasma lipoprotein particles. Apolipoprotein B mRNA editing is a zinc-dependent, site-specific cytidine deamination that determines whether the apoB-100 or apoB-48 isoform is synthesized. The objective of this work was to examine whether dietary zinc levels affect apoB mRNA editing in vivo. Adult male Sprague-Dawley rats were randomly assigned to zinc-deficient (ZD, <0.5 mg Zn/kg diet), zinc-adequate (ZA, 30 mg Zn/kg diet) or zinc-replenished (ZDA, ZD rats fed the ZA diet for last 2 d) dietary groups for 18 d. The ratio of plasma apolipoprotein B-48 (apoB-48) to total apoB was significantly lower in zinc-deficient compared with zinc-adequate rats. Primer extension analysis indicated a modest but significant reduction in hepatic apoB mRNA editing in ZD rats compared with that of the ZA group. In ZDA rats, hepatic apoB mRNA editing and the percentage of plasma apoB-48 to total apoB were not different from ZA rats. The mRNA abundance of hepatic apobec-1 (apoB mRNA editing catalytic subunit 1) was significantly lower in ZD and ZDA rats than in ZA rats. In summary, the plasma ratio of apoB-48 to total apoB protein as well as hepatic apoB mRNA editing and hepatic apobec-1 mRNA levels were reduced in rats consuming a zinc-deficient diet. These data suggest that one or more components of apoB metabolism may be influenced by dietary zinc status.

Zinc deficiency decreases plasma level and hepatic mRNA abundance of apolipoprotein A-I in rats and hamsters.

The influence of Zn deficiency on the plasma level as well as the hepatic and intestinal gene expression of apolipoprotein (apo) A-I was examined in rats and hamsters. Male Sprague-Dawley rats (8 wk old) and Golden Syrian hamsters (7 wk old) were assigned to three dietary treatments: Zn adequate (ZA, 30 mg Zn/kg diet), Zn deficient (ZD, <0.5 mg Zn/kg diet), and Zn replete (ZDA, ZD animals fed the ZA diet for the last 2 days). The dietary treatments lasted for 18 days for rats or 6 wk for hamsters. For the measurement of apoA-I mRNA abundance, hamster apoA-I cDNA was cloned from the small intestine. The full-length 905-base pair cDNA shared approximately 80% similarity with the human, rat, and mouse apoA-I cDNAs. Hepatic and plasma Zn levels were reduced in ZD animals but normalized in ZDA rats and increased in ZDA hamsters compared with ZA animals. Zn deficiency reduced plasma apoA-I and hepatic apoA-I mRNA levels 13 and 38%, respectively, in ZD rats. The 2 days of Zn replenishment raised plasma apoA-I and hepatic apoA-I mRNA levels in ZDA rats by 34 and 28%, respectively, higher than ZA rats. Similarly, these levels were decreased by 18 and 25%, respectively, in ZD hamsters but normalized in ZDA hamsters compared with ZA hamsters. In contrast to the alterations of hepatic apoA-I mRNA levels, neither Zn deficiency nor subsequent Zn repletion produced alterations in the intestinal apoA-I mRNA abundance. Data from this study demonstrated that Zn deficiency specifically decreases hepatic apoA-I gene expression, which may at least be partly responsible for the reduction of plasma apoA-I levels.

Dexamethasone and interleukin-1 potently synergize to stimulate the production of granulocyte colony-stimulating factor in differentiated THP-1 cells.

The human monocytic leukemic cell line, THP-1, which differentiates toward macrophages in response to phorbol 12-myristate 13-acetate (PMA) was investigated for its ability to produce granulocyte colony-stimulating factor (G-CSF). G-CSF protein was neither produced during PMA-induced differentiation nor in response to dexamethasone (Dex) alone. However, when combined, PMA and Dex synergistically stimulated THP-1 cells to produce G-CSF. The synergistic interaction between PMA and Dex on G-CSF production appeared to be mediated through the production of interleukin-1 (IL-1) since neutralization of IL-1 activity completely inhibited G-CSF production. Further experiments demonstrated that in THP-1 cells pretreated with PMA, Dex potently synergized with IL-1 to stimulate G-CSF production.

Regulation of apolipoprotein A-I gene expression in Hep G2 cells depleted of Cu by cupruretic tetramine.

Studies were designed to examine the regulation of apolipoprotein (apo) A-I gene expression in Cu-depleted Hep G2 cells. The cupruretic chelator N,N'-bis(2-aminoethyl)-1,3-propanediamine 4 HCl (2,3,2-tetramine or TETA) was used to maintain a 77% reduction in cellular Cu in Hep G2 cells. After two passages of TETA treatment, the relative abundance of apoA-I mRNA was elevated 52%. In TETA-treated cells, the rate of apoA-I mRNA decay measured by an actinomycin D chase study was accelerated 108%, and the synthesis of apoA-I mRNA determined by a nuclear runoff assay was enhanced 2.5-fold in TETA-treated cells. All of those changes could be reverted toward the control values with Cu supplementation for only 2 days. In transient transfection assays, a 26.7% increase in chloramphenicol O-acetyltransferase (CAT) activity for the reporter construct -256AI-CAT was observed in the treated cells. However, the ability of apoA-I regulatory protein 1 (ARP-1) to repress the CAT activity was not affected by the depressed Cu status. In addition, gel retardation experiments demonstrated that Cu depletion enhanced the binding of hepatocyte nuclear factor 4 (HNF-4) and other undefined nuclear factors to oligonucleotides containing site A, one of three regulatory sites of the apoA-I gene promoter. Moreover, the relative abundance of HNF-4 mRNA was increased 58% in the Cu-depleted cells. Thus the observed increase in apoA-I gene transcription may be mediated mostly by an elevated level of the regulatory factor, HNF-4. In summary, the present findings established the mechanism by which a depressed cellular Cu status can enhance apoA-I mRNA production and subsequently increase apoA-I synthesis.

Dexamethasone potently enhances phorbol ester-induced IL-1beta gene expression and nuclear factor NF-kappaB activation.

The synthetic glucocorticoid dexamethasone, an immunosuppressive and anti-inflammatory agent, was investigated for its effect on PMA-mediated expression of the inflammatory cytokine IL-1beta in the human monocytic leukemic cell line THP-1. PMA alone induced the production of low levels of IL-1beta in THP-1 cells, whereas dexamethasone alone had no effect. However, dexamethasone potently enhanced PMA-mediated IL-1beta production. Using a selective and potent inhibitor of protein kinase C, we found that synergistic interaction between PMA and dexamethasone requires protein kinase C activation. PMA has been known to activate nuclear factor NF-kappaB in THP-1 cells. Using an oligonucleotide probe corresponding to an NF-kappaB DNA-binding motif of the IL-1beta gene promoter in gel electrophoresis mobility shift assays, we demonstrated that PMA-induced NF-kappaB activation was greatly potentiated by dexamethasone. Our results indicate that glucocorticoids can be positive regulators of inflammatory cytokine gene expression during monocytic cell differentiation.

Copper deficiency increases total protein and apolipoprotein A-I synthesis in the rat small intestine.

This study was designed to determine whether an enhanced intestinal synthesis of apolipoprotein (apo) A-I is associated with the hyperapolipoproteinemia observed in copper-deficient rats. Male weanling Sprague-Dawley rats were assigned to two dietary treatments, Cu deficient (0.6 ppm Cu) and Cu adequate (6.0 ppm Cu) for 6 weeks. In vivo studies were then performed after rats were injected with a flooding dose of 150 microM [3H]phenylalanine (PHE, 50 microCi/ml/100 g body wt). Three rats from each treatment were sacrificed at 5, 10, 15, 30, and 60 min postinjection. The small intestine was rapidly rinsed and frozen in liquid N2. In vitro studies were performed by labeling freshly isolated 6-cm segments from duodenum, jejunum and ileum with [3H]PHE (33 microCi/ml, 49.7 Cl/mmol) in PHE-free minimum essential medium for 7 and 14 min. In vivo and in vitro intestinal samples were sonicated, solubilized in 1% Triton X-100, and centrifuged to provide the detergent soluble fraction for the isolation of nascent apo A-I and total protein. Radioactivities associated with nascent apo A-I isolated by immunoprecipitation and SDS-PAGE, and with total protein precipitated by trichloroacetic acid, were measured to determine the influence of Cu deficiency on nascent apo A-I and total protein synthesis. In the Cu-deficient small intestine, the synthesis of total protein was measured only in the duodenum and was enhanced after 1 hr for the in vivo studies. Moreover, total protein synthesis was enhanced at both 7 and 14 min of the in vitro studies for all three small intestinal segments of the Cu-deficient rats. Apo A-I synthesis was measured only at the jejunum and was also enhanced by Cu deficiency in the in vitro studies. Thus, an increase in intestinal apo A-I synthesis may contribute to the elevated plasma apo A-I level in Cu-deficient rats.

Copper deficiency increases hepatic apolipoprotein B secretion and mRNA editing in rats.

Male weanling Sprague-Dawley rats were assigned to copper-deficient (9.0 mumol Cu/kg diet) or copper-adequate (102 mumol Cu/kg diet) dietary treatments for 6 wk. Pulse-chase studies using freshly isolated rat liver parenchymal cells demonstrated that apolipoprotein B (apoB)-48 and apoB-100 syntheses were not altered, but secretion was increased twofold in hepatocytes derived from copper-deficient rats. Both plasma apoB-48 and apoB-100 levels were increased by copper deficiency, but only the apoB-48 increase was significant. Hepatic apoB mRNA editing, expressed as a ratio of apoB-48 mRNA to apoB-48 plus apoB-100 mRNA, was significantly increased from 60.8% in copper-adequate to 70.2% in copper-deficient rats. Moreover, hepatic apoB mRNA abundance was not significantly altered by copper deficiency. Thus the increased amount of nascent apoB-48 secreted into the medium as well as the enhanced apoB mRNA editing may have contributed to the differential increase in plasma apoB-48 over apoB-100 level in copper-deficient rats.