Comparison of urine albumin creatinine ratio with the pediatric index of mortality 2 score for prediction of pediatric intensive care unit outcomes.

BACKGROUND: Predicting morbidity and mortality in a pediatric intensive care unit (PICU) is of extreme importance to make precise decisions for better outcomes. AIM: We compared the urine albumin creatinine ratio (ACR) with the established PICU score, pediatric index of mortality 2 (PIM 2) for predicting PICU outcomes. METHODS: This cross-sectional study enrolled 67 patients admitted to PICU with systemic inflammatory response syndrome. Urine ACR was estimated on admission, and PIM 2 score was calculated. ACR was compared with PIM 2 for PICU outcome measures: the need for inotropes, development of multiple organ dysfunction syndrome (MODS), duration of PICU stay, and survival. RESULTS: Microalbuminuria was found in 77.6% of patients with a median ACR of 80 mg/g. ACR showed a significant association with the need for inotropes (p < 0.001), MODS (p = 0.001), and significant correlation to PICU stay (p 0.001, rho = 0.361). The area under the receiver operating characteristic curve for ACR (0.798) was comparable to that of PIM 2 (0.896). The cutoff value of ACR derived to predict mortality was 110 mg/g. The study subjects were divided into 2 groups: below cutoff and above the cutoff. Outcome variables, inotrope use, MODS, mortality, and PICU stay compared between these subgroups, were statistically significant. CONCLUSION: ACR is a good predictor of PICU outcomes and is comparable to PIM 2 for mortality prediction.

Combined effects of arginine and hydroxyurea on BFU-E derived colony growth and HbF synthesis in erythroid progenitors isolated from sickle cell blood.

Hydroxyurea (HU) increases HbF synthesis in sickle cell disease (SCD). Recent studies suggest HU-induced HbF synthesis is mediated through a NO-cGMP pathway. Since arginine is the main precursor of NO, we investigated the effects of arginine and HU mixtures on HbF synthesis and burst forming unit erythroid (BFU-E) proliferation. Mixtures of HU (0, 15, 25, 100µM) and arginine (0, 25, 50, and 100µM) resulting in optimal HbF synthesis and minimal HU-induced cytotoxicity in erythroid progenitors were determined. HU dose-dependently attenuated growth of BFU-E colonies and stimulated HbF synthesis. In contrast, arginine dose-dependently increased BFU-E colonies without affecting HbF synthesis. Furthermore, arginine at concentrations >100µM in combination with varying concentrations of HU, decreased HbF synthesis compared to HU controls. HU, 15-25µM, in combination with 25-50µM arginine not only minimized cytotoxicity, but also increased HbF synthesis when compared with HU controls. NG-nitro-L-arginine-methyl ester (L-NAME; 100µM), a nitric oxide synthase inhibitor, attenuated the effects of HU+arginine on HbF synthesis compared to HU and HU+arginine controls. These results suggest HU+arginine-induced HbF synthesis in human erythroid progenitors is NO dependent. The synergistic effect on HbF synthesis seen with combinations HU+arginine is an important observation in understanding potential therapeutic uses of HU and arginine.

Comparative validity of microalbuminuria versus clinical mortality scores to predict pediatric intensive care unit outcomes.

BACKGROUND: Predicting the prognosis of patients admitted to the pediatric intensive care unit (PICU) is very important in determining further management and resource allocation. The prognostication of critically ill children can be challenging; hence, accurate methods for predicting outcomes are needed. PURPOSE: To evaluate the role of microalbuminuria at admission as a prognostic marker in comparison to standard Pediatric Risk of Mortality (PRISM) and Pediatric Logistic Organ Dysfunction (PELOD) mortality scores in children admitted to the PICU. METHODS: This cross-sectional study was conducted from January 2015 to October 2016. Eighty-four patients aged 1 month to 18 years admitted to the PICU of teaching hospitals for more than 24 hours were enrolled by convenience sampling method. Microalbuminuria was estimated by spot urinary albumin-creatinine ratio. PRISM and PELOD scores were calculated using an online calculator. Outcome measures were PICU length of stay, inotrope usage, multiorgan dysfunction, and survival. ACR was compared with mortality scores for predicting survival. RESULTS: Microalbuminuria was present in 79.8% with a median value of 85 mg/g (interquartile range, 41.5-254 mg/g). A positive correlation was found between albumin-creatinine ratio and PICU length of stay (P=0.013, r=0.271). Albumin-creatinine ratio was significantly associated with organ dysfunction (P=0.004) and need for inotropes (P=0.006). Eight deaths were observed in the PICU. The area under the curve for mortality for albumin-creatinine ratio (0.822) was comparable to that for PRISM (0.928) and PELOD (0.877). Albumin-creatinine ratio >109 mg/g predicted mortality with a sensitivity of 87.5% and specificity of 63.2%. CONCLUSION: Microalbuminuria is a good predictor of PICU outcomes comparable with mortality scores.

Dihydroxyphenylalanine in rat food containing wheat and oats.

Dopa has been identified in rat food by three different fluorimetric assays and paper chromatography. Incubation of the rat food with proteolytic enzymes dramatically increased the measurable free dopa. Analysis of samples of six individual protein-containing constituents of rat food revealed that both wheat and oats contain dopa.

High turnover rate of transfer RNA in tumor tissue.

Cancer patients and tumor-bearing animals excrete high levels of modified purines and pyrimidines some of which, e.g., N2,N2-dimethylguanosine, can originate only from transfer RNA (tRNA). Until recently, it could not be ascertained whether the high level of excretion of such compounds is due to cell death or specific tRNA turnover. However, an approach to this problem became feasible, with beta-aminoisobutyric acid as a probe. This compound is a terminal degradation product of thymine which is present in both DNA and tRNA. Since the pathway of synthesis of thymine is different in the two macromolecules, it and its end product, beta-aminoisobutyric acid can be differentially labeled with [14C]formate and [3H3]methylmethionine as precursors. Therefore the ratio of the two labels in the excreted beta-aminoisobutyric acid is a measure of the macromolecular origin of the degradation product. We have found from such analysis that tRNA's are not homogeneous in their turnover rate. There is a subpopulation that turns over much faster than the rest. The turnover rate of a subpopulation of tRNA's in tumor tissue exceeds the turnover rate of tRNA's in normal tissue. Such rapid degradation of tRNA's must be the source of the massive excretion of modified nucleosides by cancer patients which can be 10-fold higher than in normal subjects.

Mechanism of D-amphetamine inhibition of protein synthesis.

At 1 h after intraperitoneal administration of D-amphetamine sulphate (15 mg/kg), rat brain polyribosomes show disaggregation accompanied by reduced capacity for in vitro peptide chain elongation. The direct action of amphetamine on cell-fine protein-synthesizing systems was therefore explored. When brain or liver polyribosomes from untreated rats were incubated with pH 5 enzyme, peptide chain elongation was not inhibited by the addition 4 mM amphetamine to the medium. On the other hand, an initiation-dependent system consisting of rat liver of brain mRNA and wheat germ S-30 fraction showed inhibition of [3H]leucine incorporation by 50% when 4 mM amphetamine were added. The metabolites of amphetamine, p-hydroxyamphetamine and p-hydroxynorephedrine, had no inhibitory action in either system, but the potent neurotoxin p-chloroamphetamine was a more powerful inhibitor of initiation than amphetamine. By using [3H]amphetamine, it was shown that amphetamine binds to the 80-S ribosomes of the wheat germ system. This binding depended on the presence in the system of natural liver or brain mRNA or several synthetic mRNAs, but was not promoted by polyuridylic acid as the messenger. Significantly, polyuridylic acid-dependent polyphenylalanine synthesis by the wheat germ system was not inhibited by amphetamine or p-chloroamphetamine. Therefore, it was concluded that amphetamine inhibits protein synthesis by interfering with initiation through a step related to formation of the mRNA ribosome complex.

Erk pathway inhibitor U0126 induces gamma-globin expression in erythroid cells.

Fetal hemoglobin (HbF) induction is an effective approach to improve clinical symptoms in sickle cell disease. Understanding molecular mechanisms for gamma-gene re-activation will aid efforts to design lead compounds. A potential inhibitory role for the extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase (MAPK) pathway in gamma-gene expression has been suggested recently. Therefore, we determined the ability of U0126, a selective inhibitor of MEK1/2 the upstream activators of ERK, to re-activate gamma-globin expression. K562 stable lines over-expressing constitutively active MEK1 were established. A significant increase in ERK phosphorylation was observed and gamma-gene expression was silenced concomitantly, however U0126 attenuated this effect. Studies in human erythroid progenitors confirmed the ability of U0126 to induce HbF. Cellular mechanisms for the inhibitory role of ERK signaling in drug-mediated HbF induction will be discussed.

Fetal hemoglobin induction by the histone deacetylase inhibitor, scriptaid.

Many different classes of drugs induce fetal hemoglobin (HbF) including histone deacetylase (HDAC) inhibitors such as butyrate and trichostatin A. Although these agents induce gamma-globin expression in culture many are ineffective in vivo, therefore research efforts continue to identify clinically useful fetal globin inducers. We and others demonstrated a role for p38 mitogen activated protein kinase (MAPK) in gamma-globin promoter activation by HDAC inhibitors. In this study we determined the ability of scriptaid, a novel HDAC inhibitor, to induce gamma-globin expression via p38 MAPK signaling. Scriptaid induced gamma-globin in K562 cells and human erythroid progenitors. Furthermore the p38-selective inhibitor SB203580 completely reversed the ability of scriptaid to induce HbF. To test the potential efficacy of scriptaid in humans, in vivo studies were completed in beta-YAC transgenic mice where the gamma-gene is completely silenced. Scriptaid induced reticulocytosis and human gamma-globin mRNA synthesis. At a concentration of 1 mg/kg/day given by intraperitoneal injections twice weekly we observed a significant 1.8-fold increase in gamma-globin mRNA transcripts. The potential for scriptaid as a treatment option for sickle cell disease will be discussed.

Mechanism of differentiation of human erythroleukaemic cell line K562 by hemin.

The human erythroleukaemic cell line K562, in response to various chemical agents, undergoes differentiation and exhibits exclusive production of fetal and embryonic haemoglobins. In this study we have compared the efficiency of natural growth factors interleukin-3 and erythropoietin and three chemical inducers such as dimethyl sulfoxide (DMSO, 1.9%), phorbol-12-myristate-13-acetate (PMA, 50 ng/ml) and hemin (25 microM) on growth and differentiation of these cells. Erythropoietin significantly stimulated the growth of K562 cells (P<0.0001), while interleukin-3 did not (P = 0.2783). However, neither of these growth factors individually or together induced differentiation of K562 cells. Hemin appears to be more efficient than DMSO or PMA in differentiation of K562 cells as measured by benzidine positive cells (70% or more). The differentiation of K562 cells by hemin occurs independently of protein kinase-C activation and the arrest of DNA synthesis. In contrast, hemin significantly stimulated RNA and protein synthesis (P<0.0001) as measured by [3H]-uridine and [3H]-leucine incorporation respectively. Analysis of hemin-treated K562 nuclear extract on sodium dodecylsulphate gel electrophoresis showed that one protein band of molecular weight 70 kDa decreased after 48 h of incubation in the presence of 25 microM hemin. The disappearance of this protein can be prevented by cycloheximide (100 microg/ml) and actinomycin D (0.1 microg/ml) and thus indicating that the removal of 70 kDa protein seems to be dependent on RNA and protein synthesis. The regulatory role of 70 kDa protein in hemin-induced differentiation of K562 cells is discussed.

Impaired protein kinase C activation as one of the possible mechanisms of reduced lymphocyte proliferation in iron deficiency in mice.

We investigated the effects of iron deficiency in mice on protein kinase C (PKC) activation, an enzyme required for optimal lymphocyte proliferation. C57BL/6 mice were fed either an iron-deficient diet (ID; 10 mg Fe/kg diet), a control diet (C; 50 mg/kg diet), or were pair fed (PF) to ID mice for 34 d. PKC activity was studied in spleen cells by histone phosphorylation. Iron deficiency significantly reduced cytosol activity in unstimulated cells and membrane-bound activity in cells stimulated by concanavalin A (Con A) or phorbol-12-myristate-13-acetate (PMA), and the ratio of membrane-bound over cytosol activity in mitogen-stimulated cells. In PF mice the ratio of membrane-bound activity to cytosol activity was greater than normal in Con A-treated cells and only slightly decreased in PMA-treated cells. PKC activity positively correlated with iron status. We conclude that reduced PKC activity and poor translocation results in aberrant signal transduction, which in turn might be responsible for the impaired lymphocyte proliferation associated with iron deficiency.

Effects of iron deficiency anemia on delayed cutaneous hypersensitivity in mice.

In the present study, the effect of iron-deficiency anemia on delayed cutaneous hypersensitivity was measured using weanling C57BL/6 female mice which were fed either an ad libitum control diet supplemented with 25 to 30 mg Fe/kg diet (FePO4), an iron-deficient test diet (5 to 6 mg Fe/kg diet), or pairfed control diet (25 to 30 mg Fe/kg diet). When skin sensitizing agent (dinitrofluorobenzene) was applied to these animals and skin responses were measured 3 to 5 days later, anemic mice showed a significantly decreased inflammatory skin response than either control or pairfed mice. Five days after sensitization, the animals were challenged with dinitrofluorobenzene painted on the right ear and an equal dose of only the solvent on the left ear followed by 125I-deoxyuridine injected intraperitoneally. The ratio of either total or DNA associated radioactivity incorporated into the right over the left ears was significantly lower in anemic mice than either control or pairfed mice. A single dose of Imferon injected 24 h before the recall dose of dinitrofluorobenzene restored the ratio of 125I-dUR incorporated in anemic mice without having any significant effect on either the control or pairfed groups. The results suggest that iron is not required for sensitization but is required for an effective inflammatory response.

The effect of iron-deficiency anemia on cytolytic activity of mice spleen and peritoneal cells against allogenic tumor cells.

The capacity of spleen and peritoneal cells from iron deficient mice, ad libitum fed control mice, and pair-fed mice to kill allogenic tumor cells (mastocytoma tumor P815) has been investigated. In the first study, mice were sensitized in vivo with 10(7) viable tumor cells 51 and 56 days after weaning. The capacity of splenic cells and peritoneal cells from sensitized and nonsensitized mice to kill tumor cells was evaluated 5 days after the second dose of tumor cells. At ratios of 2.5:1 to 100:1 of attacker to target cells, the percentage 51Cr release after 4 h of incubation was significantly less in iron-deficient mice than control and/or pair-fed mice (p less than 0.05). Protein-energy undernutrition in pair-fed mice had no significant effect. In the second study, spleen cells and enriched T cell fractions were incubated in vitro for 5 days with uv irradiated Balb/C spleen cells in a 2:1 ratio. The cytotoxic capacity against the same allogenic tumor cells was again evaluated. The percentage chromium release at different attacker to target cells was less than 30% in the iron-deficient group compared to either control or pair-fed supporting the results of in vivo sensitized cells. The possible mode of impairment of the cytotoxic capacity is discussed.

Impairment of blastogenic response of splenic lymphocytes from iron-deficient mice: in vivo repletion.

Iron-deficiency anemia impaired the blastogenic response of splenic lymphocytes and partially purified T cells to Concanavalin A and phytohemagglutinin. The response of splenic lymphocytes and partially B cells to bacterial lipopolysaccharide was also significantly impaired. Caloric restriction in pair-fed mice did not have any significant effect. Blastogenic response to the three mitogens was restored to normal after anemic mice were fed the regular diet containing 25 to 30 mg Fe/kg (FeSO4) for approximately 10 days. We also found that in the anemic mice the mean wet weights per 100 g of body of spleen, heart, brain, and kidney increased, while those of the thymus and liver decreased. In the pair-fed mice only the mean wet weight of the liver significantly decreased. There was a small but significant decrease in the white blood count and peripheral lymphocyte count in the anemic but not the pair-fed mice. The mechanism by which iron deficiency impairs the cell-mediated immune response is discussed.

Control of RNA content of developing human placenta.

Human placentae obtained early in pregnancy or at full term were examined for RNA content per cell, RNA polymerase types and activities, and chromatin template availability. The RNA:DNA ratio fell from 0.7 at 15 to 20 weeks to 0.4 at 40 weeks of pregnancy. Since RNase activities were similar at both times, the reduction in RNA content was attributed not to increased degradation, but to reduced synthesis. At both stages of pregnancy, about 55 to 60 per cent of the RNA polymerase activity in isolated placental nuclei was accounted for by RNA polymerase II, as judged by suppression of activity with alpha-amanitin and by separation of the extracted polymerases on DEAE-Sephadex. The relative roles of changes in polymerase activity and template availability were measured in nuclei from 20- and 40-week placentae. Nuclei showed 20 per cent greater polymerase activity in full-term than in early placentae, but the template availability of isolated chromatin for transcription by RNA polymerase II was 70 per cent less at full term. We conclude that the reduced amount of RNA per cell in the full-term placenta is due to reduced template availability that more than offsets the slight increase in polymerase activity.

Synthesis and processing of RNA by isolated human placental nuclei.

Conditions for isolating intact and active nuclei from human term pacenta and for studying their transcription products are described. The isolated nuclei can synthesize cell-free RNA for a prolonged period at 29 degree C in a medium containing 100 mM KCl and 5 mM MgCl2. Actinomycin D inhibited 92 per cent of RNA synthesis, whereas approximately 60 per cent of RNA synthesis was sensitive to alpha-amanitin. When nuclei were incubated at 29 degrees C for 1 h, about 27 per cent of the newly synthesized RNA was released into the medium outside the nucleus. Analysis of this released material by affinity chromatography on an oligo(dT)-cellulose column revealed that 2.4 per cent of the total released RNA was adsorbed at high salt concentration. Most of this fraction was eluted with a low-salt buffer at 45 degrees C and the remainder by 50 per cent formamide, conditions that are necessary for elution of poly(A)-containing mRNP particles from oligo(dT)-cellulose. These results show that placental nuclei incubated in vitro synthesize poly(A)-containing RNA, which is released as a protein-bound complex. This procedure allows exploration of changes in mRNA release during placental development.

Hyperhomocysteinemia in type 2 diabetes mellitus: cardiovascular risk factors and effect of treatment with folic acid and pyridoxine.

OBJECTIVE: To determine whether hyperhomocysteinemia (HH) exacerbates other cardiovascular risk factors and markers of coagulation and hemostasis in patients with type 2 diabetes mellitus (DM) and whether treatment of HH with vitamins will alter these risk factors. METHODS: We measured several cardiovascular risk factors and markers of coagulation and hemostasis in patients with type 2 DM with and without HH. We also treated patients with type 2 DM and coexistent HH with high doses of folic acid and pyridoxine to determine whether this treatment would lower plasma total homocysteine concentrations as well as correct other associated cardiovascular risk factors in this population. RESULTS: Plasma levels of plasminogen activator inhibitor type 1 and fibrinogen were significantly higher in all patients with DM in comparison with control subjects (P<0.01), whether they had HH or not. No significant difference was noted between the two groups of patients with DM. The presence of hypertension and microalbuminuria did not lead to a higher plasma total homocysteine. After treatment with folic acid, 15 mg daily, and pyridoxine, 600 mg daily, fasting (basal) plasma total homocysteine declined significantly in patients with DM from 12.3 +/- 2.9 micromol/L to 9.1 +/- 1.1 micromol/L (P<0.01). The peak post-methionine load plasma total homocysteine in the patients with DM decreased from 39.9 +/- 11.4 micromol/L to 30.4 +/- 6.5 micromol/L (P<0.05). Neither fasting nor peak plasma total homocysteine changed in normal subjects. None of the cardiovascular risk factors measured changed significantly with the vitamin treatment. CONCLUSION: The coexistence of type 2 DM and HH does not lead to an exacerbation of abnormalities in the measured variables of coagulation and hemostasis. Treatment with high doses of folic acid and pyridoxine lowers the plasma total homocysteine significantly but does not improve any of the associated cardiovascular risk factors that we measured. Long-term clinical trials should be conducted to determine whether high-dose vitamin treatment will diminish the increased morbidity and mortality associated with cardiovascular disease in patients with type 2 DM.

A study of lupus anticoagulant in unexplained fetal wastage.

Lupus Anticoagulant, first discovered in patients of SLE, is now known to be associated with a wide spectrum of diseases. The presence of LA is associated with adverse fetal outcome in an obstetric population. In the present series the incidence of LA was found to be 16.6% and 80% of LA positive subgroup had an unsuccessful outcome.

Evaluation of adnexal masses by ultrasound and fine needle aspiration cytology.

Forty Eight cases of adnexal masses were subjected to ultrasound and FNAC. In this study, FNAC could differentiate benign and malignant adnexal masses in 98% of cases where as ultrasound was successful in 85% of cases. FNAC is a safe simple, rapid & reliable investigation. The complex adnexal masses present a diagnostic challenge with particular reference to the findings predictive of malignancy.

Multiple primary visceral malignancy--a review with report of two cases.

Multiple primary cancers are being reported with increasing frequency in recent years, the frequency varying from 0.3 to 4.3% in different studies. A combination of primary cancer of larynx and lung is the most common followed by malignant neoplasm involving lip-larynx, skin-larynx, skin-lung, breast-ovary and breast-endometrium. Two interesting cases of rare combinations of primary cancers are being presented. In the first case primary adenocarcinoma of the gall bladder was associated with appendiceal adenocarcinoma. In the second case primary malignant papillary serous cystadenocarcinoma of right ovary and squamous cell carcinoma of cervix uteri were found.