A multi-level analysis showing associations between school neighborhood and child body mass index.

OBJECTIVE: The objective of this study is to examine associations between aspects of the environment in school neighborhoods and childhood body mass index percentile (BMIp). METHODS: Trained medical students visited 46 elementary schools in the Kansas City metropolitan area to conduct medical screenings that included the height and weight measurements of 12 118 boys and girls 4-12 years of age in the academic year 2008-2009. For the same time period, aspects of the built environment in a 2-mile radius around each school was obtained from the Walkscore database. Other environmental characteristics (for example, population change) of these areas were also obtained from various sources. Hierarchical linear modeling was used to estimate the associations between neighborhood- and individual-level factors and BMIp. RESULTS: Population size along with the number of fast-food restaurants and grocery stores were positively associated with BMIp, whereas population change along with the number of parks and fitness centers were inversely associated with BMIp. CONCLUSIONS: After considering individual-level factors and the random effects of schools, environmental elements of school neighborhoods predict childhood BMIp. This study offers evidence of the health influence of school neighborhoods in a way that can inform neighborhood redevelopment efforts.

Triplex formation inhibits HER-2/neu transcription in vitro.

Triplex-forming oligonucleotides (TFOs) have been shown to bind to target DNA sequences in several human gene promoters such as the c-myc oncogene, the epidermal growth factor receptor, and the dihydrofolate reductase genes. TFOs have been shown to inhibit transcription in vitro and gene expression in cell culture of the c-myc and other genes. The HER-2/neu oncogene, which is overexpressed in breast cancer and other human malignancies, contains a purine-rich sequence in its promoter, which is favorable for purine:purine:pyrimidine (R:R:Y) triplex formation. Although its function in the HER-2/neu promoter is unknown, this purine-rich site is homologous to a protein-binding sequence in the promoter of the epidermal growth factor receptor that is necessary for efficient transcription of this gene. We have shown that this sequence is a site for nuclear protein binding by incubation with a crude nuclear extract. We describe the formation of an interstrand triplex using a purine-rich oligonucleotide antiparallel to this purine-rich target sequence of the HER-2/neu promoter. Triplex formation by the oligonucleotide prevents protein binding to the target site in the HER-2/neu promoter in vitro. We have shown that this oligonucleotide is a potent and specific inhibitor of HER-2/neu transcription in an in vitro assay. The triplex target site contains a single pyrimidine base that does not conform to the R:R:Y triplex motif. In an attempt to abrogate the potentially destabilizing effects of this pyrimidine base on triplex formation, we have substituted an abasic linker for the pyrimidine residue in the triplex forming oligonucleotide. Triplex formation with the modified oligonucleotide appears to occur with approximately equivalent binding affinity. Triplex formation in the HER-2/neu oncogene promoter prevents transcription in vitro and may represent a future modality for specific inhibition of this gene in vivo.

Genotypic characterization of Ureaplasma species by pulsed field gel electrophoresis.

We describe the first use of pulsed field gel electrophoresis to genotype human Ureaplasma species. This technique can distinguish between U. urealyticum and U. parvum, differentiate most of the 14 serovars from one another, and identify differences among clinical isolates of the same serovar.

Psoralen-modified clamp-forming antisense oligonucleotides reduce cellular c-Myc protein expression and B16-F0 proliferation.

The c-myc protooncogene plays an important role in the abnormal growth pattern of melanoma cells. In an attempt to inhibit c-Myc expression and the growth of an established murine melanoma cell line, we targeted homopurine sequences within the mouse myc mRNA with modified antisense oligonucleotides (AS ODNs). Psoralen was conjugated to the 5'-end of these clamp-forming oligonucleotides (clamp ODNs). Gel mobility shift analysis demonstrated a sequence-specific interaction between the active clamp ODNs (Myc-E2C and Myc-E3C) and the 1.4 kb c-myc mRNA, but no interaction with the control clamp ODN (SCR**). This association was further confirmed by thermal denaturation studies. In vitro translation assays demonstrated that both Myc-E2C and Myc-E3C at 5 microM inhibited c-Myc expression >99% after UV activation at 366 nm. Immunostaining of B16-F0 cells with a c-Myc monoclonal antibody revealed a significant reduction in c-Myc after clamp ODN treatment compared with the untreated or SCR** control-treated cells. This result was corroborated by western blot analysis. Utilizing the MTT assay to determine the effects of ODN-mediated c-Myc reduction on B16-F0 growth, we observed 60 and 64% reductions in growth after treatment with 5 microM Myc-E3C and Myc-E2C, respectively. We attribute the enhanced effectiveness of the clamp ODNs to psoralen activation. Our preliminary data suggest that inhibiting c-Myc overexpression results in a significant reduction in abnormal proliferation of B16-F0 melanoma cells and that the increased efficiency of clamp ODNs may provide an important advantage for their use in antisense therapies.

Human neonatal keratinocytes have very high levels of cellular vitamin A-binding proteins.

Since cellular retinol- and retinoic acid-binding proteins (CRBP and CRABP) mediate the effects of vitamin A on epidermal differentiation, the levels of these binding proteins were measured in the epidermal and dermal layers of newborn, human foreskin as well as in primary cultures of keratinocytes and fibroblasts from these layers. Ligand binding assays with saturating concentrations of all trans-[3H]retinol or of all trans-[11-3H]retinoic acid were used to quantitate amounts of binding proteins in cytosols prepared from these skin layers or cultured cells. The epidermal levels of CRABP and CRBP (60.9 +/- 14.4 and 7.3 +/- 1.7 pmol per mg cytosol protein, respectively) were markedly higher than that reported for adult epidermis but were comparable to levels in keratinocytes cultured from neonatal foreskin epidermis (61.8 +/- 7.8 and 10.7 +/- 2.5, respectively). The levels of CRABP were much lower in the foreskin dermis than in the epidermis and the levels measured in the fibroblasts cultured from this dermis were consistent with the dermal levels. However, CRBP levels in cultured dermal fibroblasts were very low and could not account for the dermal CRBP levels, suggesting that another dermal cell type has high levels of CRBP.

Novel inhibitors of rat lens aldose reductase: N-[[(substituted amino)phenyl]sulfonyl]glycines.

A number of N-[[(substituted amino)phenyl]sulfonyl]glycines 3a-n were synthesized as analogues of the simple (phenylsulfonyl)glycines 1a-c with increased lipophilic character and therefore greater aldose reductase inhibitory potential. The 2-benzoylamino derivative 3c was found to be less potent than the corresponding amine 1c as an inhibitor of rat lens aldose reductase, but both the 3- and 4-benzoylamino analogues, 3b and 3a, are substantially more potent than their amines 1b and 1a; compound 3a is the most effective inhibitor of this series, with an IC50 of 0.41 microM. The 4-benzoylamino derivative 3a is also significantly more active than the 4-acetylamino analogue 3d and the 4-benzylamino (3e) and 4-dimethylamino (3f) derivatives, suggesting that both the additional carbonyl moiety and aromatic ring present in this compound may bind to complementary sites present on the enzyme. Furthermore, structure-activity studies reveal that increasing the number of atoms between the carbonyl and aromatic moieties of 3a results in a decrease in inhibitory activity. Kinetic studies demonstrate that 3a, like other known inhibitors of aldose reductase, functions as an uncompetitive inhibitor with respect to the substrate and therefore may interact at the proposed common inhibitor binding site of this enzyme.

N- and 2-substituted N-(phenylsulfonyl)glycines as inhibitors of rat lens aldose reductase.

A variety of N-(phenylsulfonyl)-N-phenylglycines 5, N-(phenylsulfonyl)-2-phenylglycines 6, and N-(phenylsulfonyl)anthranilic acids 7 were prepared as analogues of the N-(phenylsulfonyl)glycine 1 aldose reductase inhibitors. In the rat lens assay, several derivatives of 5 display greater inhibitory activity than the corresponding glycines 1, suggesting that N-phenyl substitution enhances affinity for aldose reductase. Enzyme kinetic evaluations of the 4-benzoylamino analogues of 5 and 1 demonstrate that these compounds produce inhibition by the same mechanism. However, the significant differences in relative inhibitory potencies between compounds of series 5 and 1 may indicate that these compounds do not interact with the inhibitor binding site in precisely the same manner. Evaluation of the individual enantiomers of series 6 reveals that the S isomers are substantially more active than the corresponding R isomers. Also, with the exception of the naphthalene analogue 6n, the S stereoisomers of this series display greater inhibitory potencies than the glycines 1. The anthranilates 7 generally are less active than the glycines 1, demonstrating that direct incorporation of an aromatic ring in the glycine side chain may result in a decrease in affinity for aldose reductase.

Relative structure-inhibition analyses of the N-benzoyl and N-(phenylsulfonyl) amino acid aldose reductase inhibitors.

A number of N-benzoyl amino acids were synthesized and tested to compare structure-inhibition relationships with the isosteric N-(phenylsulfonyl) amino acid (PS-amino acid) aldose reductase inhibitors. Inhibition analyses with these series reveals that their kinetic mechanisms of inhibition are similar, but that significant differences in structure-inhibition relationships exist. For example, while the PS-alanines and PS-2-phenylglycines produce enantioselective inhibition (S greater than R), no consistent pattern of enantioselectivity is observed with the isosteric N-benzoylalanines and 2-phenylglycines. Also, N-methyl and N-phenyl substitution in the PS-amino acid series does not substantially alter inhibitory activity, while similar substitutions in the N-benzoyl series (particularly N-phenyl) results in a significant increase in inhibitory activity. Proton NMR analysis of the N-benzoylsarcosines reveals that these compounds exist as a mixture of rotamers in solutions including the enzyme assay buffer and that the preferred conformer is one in which the carboxymethyl moiety is trans to the aromatic ring. Similar analyses with the N-benzoyl-N-phenylglycines demonstrate that these derivatives exist exclusively in the trans rotameric conformation in solution. No such N-substituent effects on conformation were observed in the PS-amino acid series. These results suggest that the differences in structure-inhibition trends between these structurally related series may result from the effect of substituents on preferred conformation.

Synthesis and in vitro aldose reductase inhibitory activity of compounds containing an N-acylglycine moiety.

A number of N-benzoylglycines (6), N-acetyl-N-phenylglycines (7), N-benzoyl-N-phenylglycines (8), and tricyclic N-acetic acids (9-12) were synthesized as analogues of the N-acylglycine-containing aldose reductase inhibitors alrestatin and 2-oxoquinoline-1-acetic acid. Derivatives of 6, which represent ring-simplified analogues of alrestatin, are very weak inhibitors of aldose reductase obtained from rat lens, producing 50% inhibition only at concentrations exceeding 100 microM. Compounds of series 7 were designed as ring-opened analogues of the 2-oxoquinolines. While these derivatives are more potent than compounds of series 6 (IC50S of 6-80 microM), they are less active than the corresponding 2-oxoquinolines. Analogues of series 8 were designed as hybrid structures of both alrestatin and the 2-oxoquinoline-1-acetic acids. These compounds are substantially more potent than compounds of series 6 and 7 and display inhibitory activities comparable to or greater than alrestatin or the 2-oxoquinolines (IC50S of 0.1-10 microM). Of the rigid analogues of 8, the most potent derivative is benzoxindole (12) with an IC50 of 0.67 microM, suggesting that fusion of the two aromatic rings of 8 in a coplanar conformation may optimize affinity for aldose reductase in this series.

Inhibitory activity and mechanism of inhibition of the N-[[(4-benzoylamino)phenyl]sulfonyl]amino acid aldose reductase inhibitors.

A series of substituted N-[[(4-benzoylamino)phenyl]sulfonyl]amino acids (BAPS-amino acids) were synthesized by established methods, and the stereochemistry of the products was confirmed by HPLC analysis after chiral derivatization. When tested against aldose reductase (alditol:NADP+ oxidoreductase; EC 1.1.1.21; ALR2) isolated from rat lens, all of the BAPS-amino acids were determined to be significantly more inhibitory than the corresponding N-(phenylsulfonyl)amino acids. Structure-inhibition and enzyme kinetic analyses suggest that the BAPS-amino acids inhibit ALR2 by a mechanism similar to the N-(phenylsulfonyl)amino acids. However, multiple inhibition analyses indicate that the increased inhibitory activity of the BAPS-amino acids is a result of interaction with multiple sites present on ALR2. Enzyme specificity studies with several of the BAPS-amino acids demonstrated that these compounds do not produce significant inhibition of other nucleotide-requiring enzymes including aldehyde reductase (alcohol: NADP+ oxidoreductase; EC 1.1.1.2; ALR1).

HDL cholesterol quantitation by phosphotungstate-Mg2+ and by dextran sulfate-Mn2+-polyethylene glycol precipitation, both with enzymic cholesterol assay compared with the lipid research method.

Two methods using commercial kits for high density lipoprotein (HDL) cholesterol quantitation were compared with the Lipid Research Clinics (LRC) procedures. HDL cholesterol quantitations on 50 patient specimens by the Lancer HDL cholesterol Rapid Stat Kit (Lancer) with phosphotungstate-Mg2+ precipitation and enzymic cholesterol assay averaged 424 mg/L, and by a method with dextran sulfate-Mn2+-polyethylene glycol (dextran sulfate) precipitation and enzymic cholesterol assay averaged 474 mg/L. By comparison, the LRC method (heparin-Mn2+ precipitation combined with a Liebermann-Burchard reagent cholesterol assay) averaged 478 mg/L. Supernates obtained by the three precipitation methods had similar cholesterol values when analyzed by the LRC assay, suggesting that the observed differences were primarily due to differences between the cholesterol assays. Results were consistent with underestimation by the enzymic assay of cholesterol in the supernates, offset by a positive interference of Mn2+ in the dextran sulfate-produced supernates. Among-day CVs of 4-5% were observed for the Lancer method, and 6-7% for the dextran sulfate method. Sedimentation of precipitates in hypertriglyceridemic specimens was excellent by both methods.

Creaming and plasma clearance rate of intravenous fat emulsion in critically ill patients.

Intravenous fat emulsion incubated with serum or plasma in vitro may result in the aggregation of fat (creaming). Twenty critically ill patients were tested for in vitro creaming of the fat emulsion Intralipid. An intravenous fat tolerance test was used to determine the plasma clearance rate of Intralipid in each patient. Eleven patients (55%) were found to be creamers. These patients had a higher mean plasma clearance rate of Intralipid than non-creamers (5.73 +/- 0.56 vs. 2.77 +/- 0.37% per min; p < 0.001); however, the rates of both groups were within the range reported in normal healthy subjects. Mean C-reactive protein concentration was significantly higher (p < 0.01), and albumin levels were lower (p < 0.01) in creamers compared to non-creamers. Ionized calcium levels did not differ between the two groups. The results of this study indicate that in vitro creaming is common in acutely ill patients. The clinical significance of creaming is probably minimal since creamers tolerated 50 to 100 g/day of intravenous fat emulsion while receiving total parenteral nutrition. Creaming was uncommon when the fat was mixed with blood in vitro.

A treatment algorithm for pneumothoraces complicating central venous catheter insertion.

BACKGROUND: We investigated the role of observation or insertion of a small French pigtail catheter with Heimlich valve as alternative management to a tube thoracostomy for iatrogenic pneumothorax complicating central venous catheter (CVC) insertion. METHODS: A retrospective review of 9,637 consecutive patients who had had subclavian CVCs inserted on an outpatient basis identified 100 patients with pneumothoraces. Treatment consisted of (1) observation, (2) outpatient insertion of a Heimlich valve, or (3) inpatient tube thoracostomy. RESULTS: The median pneumothorax size was 10% (range 1% to 100%). Fifty-eight patients had observation as initial treatment, and this strategy was successful in 35 (60%). Thirty-four patients were treated initially with Heimlich valves, and this strategy was successful in 29 (85%). Tube thoracostomy as initial therapy was successful in 7 (88%) of 8 patients. Patients in who initial treatment failed were treated with insertion of a Heimlich valve or tube thoracostomy. CONCLUSION: In appropriately selected patients, pneumothorax after insertion of a subclavian CVC can be successfully managed in the outpatient setting with observation. Patients in whom observation fails can be treated with insertion of a Heimlich valve. Tube thoracostomy can be reserved for refractory PTX or emergent situations.

In vitro aldose reductase inhibitory activity of substituted N-benzenesulfonylglycine derivatives.

A number of N-benzenesulfonylglycines, alanines, sarcosine, and prolines, which contain the minimum pharmacophore moieties necessary for aldose reductase inhibitory activity, were prepared and tested in the rat lens assay. In this assay, the benzenesulfonylglycines are considerably more potent than the corresponding alanine and sarcosine derivatives which, in turn, are more active than the proline analogues. Of the monosubstituted benzenesulfonylglycines, the 2-nitro and 4-amino derivatives were most active with 50% inhibitory concentration (IC50) values of 13 and 16 microM, respectively. The most potent derivatives evaluated were the beta- and alpha-naphthylenesulfonylglycines with IC50 values of 0.4 and 1.3 microM, respectively. The structure-activity data obtained from evaluation of the benzenesulfonylamino acids suggests that the aromatic ring and ring substituents, as well as the sulfonamide group and carboxylate moiety, all contribute to the inhibitory potency through direct interaction with complimentary binding sites present on aldose reductase.

The G-C specific DNA binding drug, mithramycin, selectively inhibits transcription of the C-MYC and C-HA-RAS genes in regenerating liver.

Expression of the c-myc and c-Ha-ras protooncogenes is dramatically increased in regenerating rat liver as an early response to partial hepatectomy. Nuclear runon transcription studies confirm that the increased c-myc and c-Ha-ras mRNA levels in regenerating livers reflect transcriptional activation of these genes. Mithramycin, a G-C specific DNA binding drug, prevents the increased transcriptional activity of c-myc and c-Ha-ras genes after hepatectomy but does not alter the transcriptional activity of the beta-actin gene. Continuous exposure of rats to mithramycin after hepatectomy prevents the increase in both c-myc and c-Ha-ras expression and blocks the increased cellular proliferation characteristic of regeneration. The delayed increase in c-myc and c-Ha-ras gene expression is associated with a delay in cellular proliferation. The inhibition of c-myc and c-Ha-ras transcription by mithramycin, the delay in cellular proliferation, and the ability of mithramycin to prevent protein binding to the c-myc promoter, suggest that the increased expression of these genes is a necessary component of liver regeneration.

Assessing the nicotine content of smokeless tobacco products.

The nicotine content of 11 popular brands of smokeless tobacco--including moist snuff, plug and loose-leaf chewing tobacco--was analyzed. In general, moist snuff has the highest nicotine content and loose-leaf chewing tobacco has the lowest, with plug tobacco falling in the middle. Variability in nicotine content may affect smokeless tobacco use and should be considered when studying usage as a variable for adverse effects of ST use.

Diastereoisomeric derivatization and liquid chromatographic analysis of N-(phenylsulfonyl)-2-phenylglycine aldose reductase inhibitors.

A series of (S)- and (R)-N-(phenylsulfonyl)-2-phenylglycines are synthesized as potential inhibitors of the enzyme aldose reductase. In vitro analysis of these compounds reveals that the S-enantiomers are more potent than the corresponding R-enantiomers and that the difference in potencies between enantiomeric pairs is dependent on the nature of the ring substituent. To ensure that the enantioselectivity observed does not reflect varying degrees of racemization during the synthesis of the N-(phenylsulfonyl)-2-phenylglycines, the enantiomeric purity of these products is determined by HPLC after chiral derivatization. Each 2-phenylglycine inhibitor is derivatized with R-alpha-methylbenzylamine, and the resulting diastereomers are analyzed using reversed and normal achiral stationary phases. Reversed-phase methods with C18 or phenyl stationary phases and solvent mixtures of acetonitrile or methanol in water do not provide satisfactory resolution of the diastereomers. However, normal-phase analyses with a silica stationary phase and mixtures of methanol, ethanol, or acetonitrile in chloroform provide good separations with relatively short analysis times. The normal-phase analyses demonstrate that a single diastereomeric amide forms from each N-(phenylsulfonyl)-2-phenylglycine product, establishing that these compounds do not racemize during synthesis.

Liquid chromatographic measurement of hydrophobicity constants for N-arylsulfonylglycine aldose reductase inhibitors.

The hydrophobicity constants for a series of aldose reductase inhibitors (ARIs) are determined by reversed-phase liquid chromatography. A series of reference compounds consisting of 23 barbituric acid derivatives are separated on two phenylsilica stationary phases over a range of methanol concentrations (30-80%) in 0.05 M phosphate buffer. Linear regression analysis of the measured log k' data is used to estimate the capacity factor in 100% water (log k'w) for each compound. The log k'w values are regressed against the shake-flask-measured 1-octanol-water partition coefficients, producing a correlation of 0.953. The same procedure is then used to estimate the log k'w values for a large group of ARIs and their log P values, calculated from the established relationship between log k'w and log P from the reference compounds. An initial analysis of the aldose reductase inhibitory activity of these compounds as a function of hydrophobicity alone fails to reveal a clear relationship, demonstrating the need for a multivariant approach for quantitative structure-activity analysis in this series of compounds.

Effect of music tempo on task performance.

Two studies were conducted to evaluate the effect of music tempo on task performance. In Study 1, 44 undergraduate business students were asked to be "workers" in a stock market project by collecting closing stock prices and calculating the percentage of change in the price from week to week. Subjects were randomly divided into groups such that they either listened to fast-paced music while they worked, to slow-paced music, or to no music. Analyses of variance and covariance were conducted on both the quantity and quality of the subjects' work, using music listening habits as a covariate. There were no differences in either the quantity or quality of the work produced by the groups. There were some methodological concerns regarding Study 1, so a second study was conducted. The 70 undergraduate business students in Study 2 completed the same task under the same music conditions as in Study 1. Analyses of variance indicated women performed significantly better than men, performance was significantly higher in the rock condition than in the heartbeat condition, and subjects in the rock condition had a significantly higher perceived level of distraction by the music.