RHS6 coordinately regulates the Th2 cytokine genes by recruiting GATA3, SATB1, and IRF4.

BACKGROUND: Asthma is a Th2 cell-driven inflammatory disease and a major public health concern. The cis-acting element Rad50 hypersensitive site 6 (RHS6) in the Th2 locus control region is essential for regulation of the Th2 cytokine genes; however, its role in allergic airway inflammation and underlying molecular mechanisms of the regulation by RHS6 are poorly understood. OBJECTIVE: We sought to understand the role of RHS6 in the development of allergic airway inflammation and its molecular mechanism for Th2 cytokine expression. METHODS: We used an ovalbumin-induced allergic inflammation model with RHS6-deficient mice to examine the role of RHS6 in this process. To examine molecular mechanism of RHS6 for Th2 cytokine expression, we used DNA affinity chromatography and mass spectrometry, quantitative RT-PCR, ELISA, intracellular cytokine staining, chromatin immunoprecipitation, and co-immunoprecipitation. RESULTS: Deletion of RHS6 caused a dramatic resistance to allergic airway inflammation. RHS6 recruited transcription factors GATA3, SATB1, and IRF4, which play important roles in expression of all three Th2 cytokine genes. RHS6 deficiency caused inhibition of transcription factor-induced Th2 cytokine gene expression. CONCLUSION: RHS6 is a critical regulatory element for allergic airway inflammation and for coordinate regulation of Th2 cytokine genes by recruiting GATA3, SATB1, and IRF4.

Weight loss outcomes are generally worse for dogs and cats with class II obesity, defined as > 40% overweight.

In pet dogs and cats, adiposity is most-often estimated clinically using a 9-category body condition score (BCS), with BCS 9 equating to ~ 40% overweight. Animals that are more overweight (> 40%) are seen in clinical practice but are not appropriately depicted by descriptions in the existing categories. To determine whether being > 40% overweight has clinical relevance, this study aimed to compare the outcomes of weight management in animals that were > 40% overweight with those < 40% overweight. Records of dogs and cats attending a specialist obesity care clinic, where adiposity is determined using dual-energy X-ray absorptiometry (DXA), were reviewed. Animals were assigned to two classes (class I ≤ 40% overweight: 118/398 [40%] dogs and 68/116 [59%] cats; class II, > 40% overweight: 180/398 [60%] dogs and 48/116 [41%] cats) based on DXA results, and weight loss outcomes were compared. Fewer class II dogs obesity completed weight management than class I dogs (P < 0.001), rate of weight loss was also slower (P = 0.012) and lean tissue loss greater (P < 0.001). Compared with class I, cats with class II obesity lost more weight (P = 0.048) albeit over a longer period (P = 0.043) leading to greater lean tissue loss (P = 0.004). Approximately half the pets presenting to a specialist clinic were have class II obesity (> 40% overweight), and some weight loss outcomes are worse for these animals.

An inherited enzymatic defect in porphyria cutanea tarda: decreased uroporphyrinogen decarboxylase activity.

Uroporphyrinogen decarboxylase activity was measured in liver and erythrocytes of normal subjects and in patients with porphyria cutanea tarda and their relatives. In patients with porphyria cutanea tarda, hepatic uroporphyrinogen decarboxylase activity was significantly reduced (mean 0.43 U/mg protein; range 0.25-0.99) as compared to normal subjects (mean 1.61 U/mg protein; range 1.27-2.42). Erythrocyte uroporphyrinogen decarboxylase was also decreased in patients with porphyria cutanea tarda. The mean erythrocyte enzymatic activity in male patients was 0.23 U/mg Hb (range 0.16-0.30) and in female patients was 0.17 U/mg Hb (range 0.15-0.18) as compared with mean values in normal subjects of 0.38 U/mg Hb (range 0.33-0.45) in men and 0.26 U/mg Hb (range 0.18-0.36) in women. With the erythrocyte assay, multiple examples of decreased uroporphyrinogen decarboxylase activity were detected in members of three families of patients with porphyria cutanea tarda. In two of these families subclinical porphyria was also recognized. The inheritance pattern was consistant with an autosomal dominant trait. The difference in erythrocyte enzymatic activity between men and women was not explained but could have been due to estrogens. This possibility was supported by the observation that men under therapy with estrogens for carcinoma of the prostate had values in the normal female range. It is proposed that porphyria cutanea tarda results from the combination of an inherited defect in uroporphyrinogen decarboxylase and an acquired factor, usually siderosis associated with alcoholic liver disease.

The role of superoxide anion radical in the reduction of ferritin iron by xanthine oxidase.

Superoxide dismutase exerted a pronounced inhibitory effect upon xanthine oxidase-mediated reduction of iron in ferritin, ferric chloride, or ferric ADP. Maximal inhibition was observed when the superoxide dismutase concentration was only about 1% of that found in normal porcine liver. These observations indicate that superoxide anion radical is an intermediate in the reduction of iron by xanthine oxidase in vitro but not in vivo.

The role of iron in the pathogenesis of porphyria cutanea tarda. II. Inhibition of uroporphyrinogen decarboxylase.

Porphria cutanea tarda is characterized biochemically by excessive hepatic synthesis and urinary excretion of uroporphyrin I and 7-carboxylporphyrins. This pattern of excretion suggest an impaired ability to decarboxylate uroporphyrinogen to the paired ability to decarboxylate uroporphyringen to the 4-carboxyl porphyrinogen, coproporphyrinogen, a reaction catalyzed by the enzyme uroporphyringen decarboxylase. Because clinical evidence has implicated iron in the pathogenesis of porphyria cutanea tarda, these experiments were designed to study the effect of iron on uroporphyrinogen decarboxylase in procine crude liver extracts. Mitochondria-free crude liver extracts were preincubated with ferrous ion and aliquots were assayed for uroporphyrinogen decarboxylase activity. Uroporphyrinogens I and III, the substrates for the decarboxylase assay, were prepared enzymatically from (3H)porphobilinogen. The products of the decarboxylase reaction were identified and quantitated by three methods: (a) extraction into 1.5 N HCl and spectrophotometric quantitation; (b) adsorption onto talc, esterification, paper chromatographic identification, and quantitation by liquid scintillation counting; and (c) adsorption onto talc, esterification, thin-layer chromatographic identification on silica gel, and quantitation by liquid scintillation counting. The thin-layer scinllation method proved most sensitive as it was the only method which accurately identified and quantitated the 7-carboxyl porphyrin reaction product. Uroporphyrinogens I and III were decarboxylated at the same rate by porcine hepatic uroporphyrinogen decarboxylase, and the addition of iron induced marked inhibition of the decarboxylase activity. Ortholpehanthroline blocked the inhibitory effect of iron. The inhibition of uroporphyrinogen decarboxylase by ferrous ion, coupled with its previously reported inhibitory effect on uroporphyrinogen III cosynthetase, provides a possible biochemical explanation for the pattern of urinary porphyrin excretion observed in patients with porphyria cutanea tarda and the clinical association with disordered iron metabolism.

The role of iron in the pathogenesis of porphyria cutanea tarda. An in vitro model.

Porphyria cutanea tarda (PCT) is characterized biochemically by excessive hepatic synthesis and urinary excretion of uroporphyrin I. Clinical evidence has implicated iron in the pathogenesis of PCT. The synthesis of the normally occurring isomer of uroporphyrin, namely uroporphyrin III, from porphobilinogen (PBG) requires two enzymes; uroporphyrinogen I synthetase and uroporphyrinogen III cosynthetase (COSYN). In the absence of COSYN only uroporphyrinogen I is formed. These experiments were designed to study the effect of iron on porphyrin biosynthesis in porcine and human crude liver extracts and to measure COSYN activity in the presence of iron.Mitochondria-free crude liver extracts were prepared in 0.25 m sucrose at pH 7.4 by centrifugation at 37,000 g. Preparations were incubated with either 0.2 mm amino-levulinic acid (ALA) or 0.1 mm PBG. The addition of ferrous ion (either from ferritin iron [4 mug/ml] and cysteine [6.7 mm] or ferrous ammonium sulfate [0.3 mm Fe] and cysteine) significantly increased the rate of uroporphyrin synthesis from either ALA or PBG. The predominant porphyrin synthesized in the presence of ferrous ion was uroporphyrin I whereas coproporphyrin III predominated in its absence. Orthophenanthroline blocked these effects of ferrous ion.To investigate the effect of ferrous ion on COSYN, crude liver extracts were incubated with ferrous ammonium sulfate (0.3 mm Fe) and cysteine (6.7 mm) and the COSYN activity of the incubates was assayed directly. In both porcine and human extracts ferrous ion caused marked inhibition of COSYN activity. Orthophenanthroline blocked the inhibitory effect.Inactivation of COSYN by heating resulted in marked enhancement of porphyrin synthesis from PBG. The sole product was uroporphyrin I.Thus, inactivation of COSYN results in accelerated synthesis of uroporphyrin I. This effect of ferrous ion provides a possible biochemical explanation for the excess production and excretion of uroporphyrin I in patients with PCT and the reversal of this defect by phlebotomy.

Iron metabolism in copper-deficient swine.

The way in which iron is handled by the duodenal mucosa, the reticuloendothelial system, the hepatic parenchymal cell, and the normoblast was investigated in copper-deficient swine.Copper-deficient swine failed to absorb dietary iron at the normal rate. Increased amounts of stainable iron were observed in fixed sections of duodenum from such animals. When (59)iron was administered orally, the mucosa of copper-deficient animals extracted iron from the duodenal lumen at the normal rate, but the subsequent transfer to plasma was impaired.When intramuscular iron supplements were given to copper-deficient pigs, increased amounts of iron were found in the reticuloendothelial system, the hepatic parenchymal cells, and in normoblasts (sideroblasts). Hypoferremia was observed in the early stages of copper deficiency, even though iron stores were normal or increased. When red cells that were damaged by prolonged storage were administered, the reticuloendothelial system failed to extract and transfer the erythrocyte iron to the plasma at the normal rate. Administration of copper to copper-deficient animals with normal iron stores resulted in a prompt increase in the plasma iron. The observed abnormalities in iron metabolism are best explained by an impaired ability of the duodenal mucosa, the reticuloendothelial system, and the hepatic parenchymal cell to release iron to the plasma. It is suggested that copper is essential to the normal release of iron from these tissues. This concept is compatible with the suggestion made by others that the transfer of iron from tissues to plasma requires the enzymatic oxidation of ferrous iron, and that the plasma copper protein, ceruloplasmin, is the enzyme (ferroxidase) which catalyzes the reaction. Because excessive amounts of iron were found in normoblasts, it is suggested that an additional defect in iron metabolism affects these cells and plays a major role in the development of anemia. As a result of the proposed defect, iron cannot be incorporated into hemoglobin and, instead, accumulates as nonhemoglobin iron.

The role of ceruloplasmin in iron metabolism.

The importance of ceruloplasmin in iron metabolism was studied in swine made hypoceruloplasminemic by copper deprivation. When the plasma ceruloplasmin level fell below 1% of normal, cell-to-plasma iron flow became sufficiently impaired to cause hypoferremia, even though total body iron stores were normal. When ceruloplasmin was administered to such animals, plasma iron increased immediately and continued to rise at a rate proportional to the logarithm of the ceruloplasmin dose. The administration of inorganic copper induced increases in plasma iron only after ceruloplasmin appeared in the circulation. Thus, ceruloplasmin appeared to be essential to the normal movement of iron from cells to plasma. Studies designed to define the mechanism of action of ceruloplasmin were based on the in vitro observation that ceruloplasmin behaves as an enzyme (ferroxidase) that catalyzes oxidation of ferrous iron. Retention of injected ferrous iron in the plasma of ceruloplasmin-deficient swine was significantly less than that of ferric iron, reflecting impaired transferrin iron binding. Rat ceruloplasmin, which has little ferroxidase activity, was much less effective than porcine or human ceruloplasmin in inducing increases in plasma iron. These observations suggest that ceruloplasmin acts by virtue of its ferroxidase activity. Eight patients with Wilson's disease were evaluated in order to investigate iron metabolism in a disorder characterized by reduced ceruloplasmin levels. Evidence of iron deficiency was found in six of these, and in five of the six, plasma ceruloplasmin was less than 5% of normal. In comparison, the two patients without evidence of iron deficiency had ceruloplasmin levels of 11 and 18% of normal. It is suggested that iron deficiency tends to occur in those patients with Wilson's disease who have the severest degrees of hypoceruloplasminemia, possibly because of defective transfer of iron from intestinal mucosal cells to plasma.

Porphyria cutanea tarda associated with disinfectant misuse.

Porphyria cutanea tarda was detected in a 44-year-old janitress. The illness was probably caused by the unintentional synthesis of a polychlorinated phenol as a result of mixing commonly available household ingredients in toilet bowls and shower stalls. Although the evidence for this hypothesis is circumstantial, its likelihood and the wide-spread household use of these reagents justify calling attention to the innovative misuse of disinfectants as a potential source of toxic exposure.

The anemia of chronic disease.

The anemia of chronic disease (ACD) is defined as a mild anemia associated with a chronic inflammatory, infectious or neoplastic illness and with a characteristic disturbance of iron metabolism. Many of the findings in ACD can be accounted for by release of a monokine called leukocyte endogenous mediator (LEM), endogenous pyrogen, or interleukin-1. This substance is released from "activated" monocytes. Bacterial endotoxins, certain lymphokines and phagocytic challenges are among the factors stimulating its biosynthesis. LEM induces fever, leukocytosis, biosynthesis. LEM induces fever, leukocytosis, and a variety of biochemical changes, including hypoferremia and alterations in plasma protein synthesis, collectively known as the "acute phase response." It is proposed that ACD results from the long-term elaboration of LEM and that release of this material is the common pathogenetic factor found in the illnesses that are associated with ACD. Some suggestions are made for testing the hypothesis. The hypoferremia associated with ACD is probably caused by defective release of iron from cells--particularly from macrophages, but also from hepatocytes and intestinal epithelium. Two possible mechanisms for this abnormality have been proposed: liberation of lactoferrin from neutrophilic leukocytes and induction of apoferritin synthesis. Neither mechanism has been established. Erythrokinetic studies in ACD have detected a modest reduction of erythrocyte survival without an adequate compensatory increase in the rate of red cell production. The reduced erythrocyte survival is probably related to an increase in phagocytic activity by activated macrophages. Impaired bone marrow response is partly related to the restricted iron supply, but there is substantial evidence for an additional defect in erythropoietin secretion. In some malignant diseases, there is evidence of an additional abnormality: impaired marrow response to a normal amount of erythropoietin. The nature of the erythropoietic defects and the relation of LEM to them remain to be established.

Antisense DNA-targeting protein kinase A-RIA subunit: a novel approach to cancer treatment.

Enhanced expression of the RIa subunit of cAMP-dependent protein kinase type I (PKA-I) has been shown during carcinogenesis, in human cancer cell lines and in primary tumors. We demonstrate that the sequence-specific inhibition of RIa gene expression by antisense oligonucleotides results in the differentiation of leukemia cells and growth arrest of cancer cells of epithelial origin and tumors in mice. The loss of RI by the antisense results in rapid increase in the half-life of the competitor molecule, RII protein, via its stabilization in a holoenzyme complex (PKA-II) that insures depletion of PKA-I and sustained inhibition of tumor growth. RI antisense, which restrains tumor cell growth by turning on the signals for blockade of tumor cell survival, namely blockade of the tyrosine kinase signaling, cell cycle deregulation and apoptosis, provides a single gene-targeting approach to treatment of cancer.

The influence of community context on the preferences of older adults for entering a nursing home.

Previous research has established that rural elders are more likely to enter a nursing home than elders living in suburban and urban areas. This research examined preferences for long-term care alternatives using a telephone survey of community-dwelling elders (age 65+). In contrast to admission patterns, urban elders were more likely to prefer nursing home care if unable to live independently. This difference persisted in multinomial logit models that included other predictors of nursing home use. Thus, rural elders may be more likely to experience discrepancies between their preferred mode of long-term care and the actual outcomes that they may ultimately experience.

Gender differences in the depressive effect of widowhood in later life.

OBJECTIVES: This study documented the stronger adverse effect of widowhood on the psychological well-being of men than that of women and explained why this gender difference in the effect of widowhood exists. METHODS: Data came from Wave 1 of the National Survey of Families and Households. Married and widowed people aged 65 and older were selected (n = 1,686). The dependent variable was the Center for Epidemiologic Studies--Depression scale (CES-D). RESULTS: Widowhood was indeed more depressing for men than women. However, this was due primarily to the fact that married men were much less depressed than married women; widowed men and women were comparably depressed. Other contributors to the stronger effect of widowhood for men included men's shorter average time since widowhood, lower frequency of church attendance, stronger dislike of domestic labor, and lessened ability to assist their children. DISCUSSION: Although widowhood has a strong depressive effect for older men, its effect for women is nonsignificant, and it explains a small proportion of the variation in depressive symptomatology. This suggests that most people, particularly women, adapt relatively well in the long run.

Are expectations for care related to the receipt of care? An analysis of parent care among disabled elders.

This investigation explores the relationship between the degree to which older parents expect assistance from their children and the actual amount of care they receive from their children. Combining the theories of reasoned action and intergenerational solidarity, we hypothesize that global expectations (social norms about receiving care) influence specific expectations (behavioral intentions to seek care), but that it is specific expectations that influence the amount of care received from children. Data were collected at three points over a 12-month period among a sample of impaired older parents (65+) who lived independently in community settings (n = 334). Using structural equation models, the results were consistent with the hypothesis. We conclude that the theory of reasoned action is a useful adjunct to the theory of intergenerational solidarity by proposing that parents adjust their global expectations to reflect the specific realities of the lives of their children.

The health status, health services utilization, and support networks of the rural elderly: a decade review.

Research from the 1980s on several dimensions of health and health care among the rural elderly is reviewed. Following a brief discussion of the demographic patterns and life conditions of the rural elderly, the current state of knowledge regarding health status, health services utilization, and the potential for informal and familial care of the elderly is examined. The review concentrates on studies that include comparisons between rural and urban populations and/or control for additional variables that are known to covary with residence. Such analyses permit the documentation of the effects of residential location on health indicators net of other factors. The review concludes that the rural elderly are relatively disadvantaged in terms of both health status and access to health care services, and have little if any advantage over the urban elderly in their access to informal sources of care. Following the review, an agenda for future research is identified.

Success rates for functional MR imaging in children.

BACKGROUND AND PURPOSE: Functional MR imaging is widely used for research in functional brain development in healthy children. However, obtaining high-quality brain imaging data from pediatric research participants requires cooperation that is challenging for young children. In this study, we examined success rates for fMRI in typically developing children in both longitudinal and cross-sectional research study designs to inform the recruitment needs of future pediatric brain imaging studies. MATERIALS AND METHODS: In the cross-sectional study, 459 healthy children (5-18 years of age, 215 girls) were recruited. A subset of 30 healthy children 5-7 years of age from the cross-sectional cohort were selected and scanned for 10 consecutive years in the longitudinal arm of the study. Following anatomic scans, each participant attempted 4 functional MR imaging tasks. Success rate was defined as the proportion of fMRI tasks completed. Differences in success rates across sexes and in cross-sectional-versus-longitudinal cohorts were evaluated by using the Fischer exact test. RESULTS: In the cross-sectional study, 74% of the children completed all tasks. Success rates for individual tasks ranged from 34% to 67% for children 5-7 years of age and 76%-100% for those 8-18 years of age. In the longitudinal study, 89% of children completed all tasks in all 10 years. We established significance (P < .0001) between the cross-sectional and longitudinal cohorts for both 0% and 100% task completion rates. There was no significance between sexes. CONCLUSIONS: When designing pediatric fMRI studies in children, the sample sizes indicated by power analysis should be scaled up according to age (ie, 33% for ages 8-18 years, 50% for ages 5-7 years).