Pyropheophorbide 2-deoxyglucosamide: a new photosensitizer targeting glucose transporters

To prepare near-infrared fluorescence imaging and photodynamic therapy agents targeted at glucose transporters, pyropheophorbide 2-deoxyglucosamide (Pyro-2DG) was synthesized and evaluated in a 9L glioma rat model. Fluorescence imaging studies demonstrate that Pyro-2DG is selectively accumulated in the tumor. Upon its photoactivation, we demonstrate that this agent efficiently causes selective mitochondrial damage to the region of a tumor that was photoirradiated after administration of this agent, but does not affect tissues photoirradiated in the absence of the agent or tissues treated with the agent that are not photoirradiated. Preliminary confocal microscopy studies suggest that Pyro-2DG is delivered and trapped in tumor cells via the GLUT/hexokinase pathway and therefore is useful both as a tumor-targeted NIR fluorescence imaging probe and as a PDT agent for the destruction of cancer.     

Structural effects of carbohydrate-containing polycations on gene delivery. 2. Charge center type

Recent polycation structure-gene delivery studies reveal that subtle changes in the molecular structure of polycations have substantial influences on DNA-binding and condensation and on in vitro toxicity and gene delivery efficiency. In Part 1 of this structure-property study using carbohydrate-containing polycations (1), it is demonstrated that as the amidine charge center is removed further from the carbohydrate unit within the polycation structure, the toxicity increases. Inclusion of larger carbohydrate species within the polycation backbone also reduces the toxicity. Here, the effect that polycation charge center type has on toxicity and gene delivery efficiency is investigated. A series of quaternary ammonium polycations containing N,N,N',N'-tetramethyl-1,6-hexanediamine, d-trehalose, and beta-cyclodextrin are synthesized in order to elucidate the effects of charge center type (by comparison to the data given in Part 1) on gene delivery. In all cases, it is found that the quaternary ammonium analogues exhibit lower gene expression values and similar toxicities to their amidine analogues. Additionally, transfection experiments conducted in the presence of chloroquine reveal increased gene expression from quaternary ammonium containing polycations and not from their amidine analogues.     

Weight lifting belt use patterns among a population of health club members

The purpose of this study was to identify patterns of weight belt use in a population of recreational weight trainers. We hypothesized that the majority of weight belt users utilized weight belts to reduce injury risk and/or improve performance while lifting submaximal loads. Three hundred fifty-two strength training health center members (189 men, 156 women, 7 gender not indicated) aged 20-72 years (36.5 +/- 10.5 years) completed a voluntary weight belt use survey. Overall, 27% (94/352) were weight belt users and 73% (258/352) were nonusers. Ninety percent (85/94) of belt users and 63% (55/88) of former belt users utilized belts to prevent injury, whereas 22% (21/94) of belt users and 28% (25/88) of former belt users utilized belts to improve performance. Many belt users were found to utilize belts during inappropriate situations such as lifting light loads or during exercises that do not typically stress the trunk musculature. Based on these findings we suggest that specific educational interventions be developed at health and fitness facilities to help provide a foundation for more informed decision-making regarding weight belt use.     

End-of-life issues as perceived by Lebanese judges

Objectives: to assess the attitudes of judges in Beirut, Lebanon, regarding end-of-life issues such as assisted suicide and withholding or withdrawing life-sustaining treatment.
Subjects and Methods: 85% of all currently acting and in-training judges and public prosecutors in Beirut (N =135) were surveyed using a mailed questionnaire that assessed attitudes toward intervention in five hypothetical cases. The associations of attitudes, on a scale from least to most 'sympathetic' toward assisting those who desire to end their lives, were measured by a variety of personal, social and professional variables.
Results: younger individuals, and those who have not yet been formally appointed as judges, were significantly more sympathetic to withdrawal or withholding of life-sustaining devices when patients or their proxies requested it, and more in support of assisted suicide. Gender, religious denomination, religious practice, and personal experience with prolonged illness leading to death among close friends or family, were generally not significant predictors of respondents' attitudes. Years of experience as a judge correlated strongly with age and may have contributed to its predictive effect.
Discussion and conclusions: a relatively more sympathetic attitude among younger judges, many of them women, and among trainees, may reflect a historical evolution in younger age-groups in Lebanon today. A survey of opinions in the public may help reach a more conclusive understanding in this regard. In any case, judges in Lebanon will remain important partners in the debate, as they will continue to be the final interpreters of the letter of the law in end-of-life issues.     

Ribosomal protein L11 negatively regulates oncoprotein MDM2 and mediates a p53-dependent ribosomal-stress checkpoint pathway

The gene encoding p53 mediates a major tumor suppression pathway that is frequently altered in human cancers. p53 function is kept at a low level during normal cell growth and is activated in response to various cellular stresses. The MDM2 oncoprotein plays a key role in negatively regulating p53 activity by either direct repression of p53 transactivation activity in the nucleus or promotion of p53 degradation in the cytoplasm. DNA damage and oncogenic insults, the two best-characterized p53-dependent checkpoint pathways, both activate p53 through inhibition of MDM2. Here we report that the human homologue of MDM2, HDM2, binds to ribosomal protein L11. L11 binds a central region in HDM2 that is distinct from the ARF binding site. We show that the functional consequence of L11-HDM2 association, like that with ARF, results in the prevention of HDM2-mediated p53 ubiquitination and degradation, subsequently restoring p53-mediated transactivation, accumulating p21 protein levels, and inducing a p53-dependent cell cycle arrest by canceling the inhibitory function of HDM2. Interference with ribosomal biogenesis by a low concentration of actinomycin D is associated with an increased L11-HDM2 interaction and subsequent p53 stabilization. We suggest that L11 functions as a negative regulator of HDM2 and that there might exist in vivo an L11-HDM2-p53 pathway for monitoring ribosomal integrity.     

The comparative amino acid sequences, substrate specificities and gene or cDNA nucleotide sequences of some prokaryote and eukaryote amidinotransferases: implications for evolution

The amino acid sequences of the amidinotransferases and the nucleotide sequences of their genes or cDNA from four Streptomyces species (seven genes) and from the kidneys of rat, pig, human and human pancreas were compared. The overall amino acid and nucleotide sequences of the prokaryotes and eukaryotes were very similar and further, three regions were identified that were highly identical. Evidence is presented that there is virtually zero chance that the overall and high identity regions of the amino acid sequence similarities and the overall nucleotide sequence similarities between Streptomyces and mammals represent random match. Both rat and lamprey amidinotransferases were able to use inosamine phosphate, the amidine group acceptor of Streptomyces. We have concluded that the structure and function of the amidinotransferases and their genes has been highly conserved through evolution from prokaryotes to eukaryotes. The evolution has occurred with: (1) a high degree of retention of nucleotide and amino acid sequences; (2) a high degree of retention of the primitive Streptomyces guanine+cytosine (G+C) third codon position composition in certain high identity regions of the eukaryote cDNA; (3) a decrease in the specificities for the amidine group acceptors; and (4) most of the mutations silent in the regions suggested to code for active sites in the enzymes.     

Characterization of a Helicobacter pylori vaccine candidate by proteome techniques

In a previous two-dimensional (2D) gel electrophoretic study of protein antigens of the gastric pathogen, Helicobacter pylori recognized by human sera, one of the highly and consistently reactive antigens, a protein with M_r of approximately 30?000 (Spot 15) seemed to be of special interest because of low yields on N-terminal protein sequencing. This suggested possible N-terminal modification, as the N-terminal sequence analysis of this 30?000 protein (Spot 15) did not provide a definitive match within the H. pylori genomic database. This protein was isolated by 2D polyacrylamide gel electrophoresis, evaluated by liquid chromatography–mass spectrometry, and found to consist of two related species of approximately 28?100 and 26?500. In parallel, the proteins within this spot were digested in situ with the endoprotease Lys-C. Analysis of the Lys-C digest by matrix-assisted laser desorption time-of-flight mass spectrometry, peptide mapping, and sequence analysis was conducted. Comparison of the mass and sequence of the Lys-C peptides with those derived from a H. pylori genomic library identified an open reading frame of approximately 300 base pairs as the source of the Spot 15 protein. This corresponded to HP0175 in the recently reported H. pylori genome sequence, an open reading frame with some homology to Campylobacter jejeuni cell binding protein 2. Mass spectral and sequence analysis indicated that Spot 15 was a processed product generated by proteolytic cleavage at both the carboxy and amino termini of the 34 open reading frame precursor.     

The Tryptophan Fluorescence of Tetracarbidium conophorum Agglutinin II and a Solution-Based Assay for the Binding of a Biantennary Glycopeptide

The plant lectin Tetracarbidium conophorum agglutinin II binds to glycoproteins and glycopeptides in a structurally specific manner [Animashaun et al., (1994) Glycoconjugate J. 11, 299–303]. We have characterized the steady-state and time-resolved fluorescence of the tryptophan residues of this lectin. The fluorescence (λ_ex = 295 nm, λ_em = 350 nm) decay is complex and can be described by four decay times with the following values: τ₁ = 7.4nsec, α₁ = 0.22; τ₂ = 2.9 nsec, α₂ = 0.25; τ₃ = l.0 nsec, α₃ = 0.34; τ₄ = 0.2 nsec, α₄ = 0.18. The addition of a biantennary glycopeptide $\begin{array}{*{20}c} {Gal\beta (1 \to 4)GlcNAc\beta (1 \to 2)Man\alpha (1 \to 6)\neg } \\ {Man\beta (1 \to 4)GlcNAC\beta (1 \to 4)GlcAc\beta (1 \to )\begin{array}{*{20}c} {Glu - Nh_2 } \\ | \\ {Asn} \\ | \\ {COOH} \\ \end{array} } \\ {Gal\beta (1 \to 4)GlcNAc\beta (1 \to 2)Man\alpha (1 \to 3)} \\ \end{array} $ to the lectin results in a quench and an 8 nm blue shift of the emission spectrum. The effect is saturable, and is described by an association constant of 1.8×10⁵ M^?1. The tryptophan fluorescence of Tetracarbidium conophorum agglutinin II may therefore be utilized to characterize thermodynamically the binding interactions between this lectin and complex glycoprotein.     

The Tryptophan Fluorescence of Tetracarbidium conophorum Agglutinin II and a Solution-Based Assay for the Binding of a Biantennary Glycopeptide

The plant lectin Tetracarbidium conophorum agglutinin II binds to glycoproteins and glycopeptides in a structurally specific manner [Animashaun et al., (1994) Glycoconjugate J. 11, 299–303]. We have characterized the steady-state and time-resolved fluorescence of the tryptophan residues of this lectin. The fluorescence (_ex = 295 nm, _em = 350 nm) decay is complex and can be described by four decay times with the following values: ₁ = 7.4nsec, ₁ = 0.22; ₂ = 2.9 nsec, ₂ = 0.25; ₃ = l.0 nsec, ₃ = 0.34; ₄ = 0.2 nsec, ₄ = 0.18. The addition of a biantennary glycopeptide to the lectin results in a quench and an 8 nm blue shift of the emission spectrum. The effect is saturable, and is described by an association constant of 1.8×10⁵ M^–1. The tryptophan fluorescence of Tetracarbidium conophorum agglutinin II may therefore be utilized to characterize thermodynamically the binding interactions between this lectin and complex glycoprotein.     

Where We Used to Live: Validating Retrospective Measures of Childhood Neighborhood Context for Life Course Epidemiologic Studies

Early life exposures influence numerous social determinants of health, as distal causes or confounders of later health outcomes. Although a growing literature is documenting how early life socioeconomic position affects later life health, few epidemiologic studies have tested measures for operationalizing early life neighborhood context, or examined their effects on later life health. In the Life-course Influences on Fetal Environments (LIFE) Study, a retrospective cohort study among Black women in Southfield, Michigan (71% response rate), we tested the validity and reliability of retrospectively-reported survey-based subjective measures of early life neighborhood context(N=693). We compared 3 subjective childhood neighborhood measures (disorder, informal social control, victimization), with 3 objective childhood neighborhood measures derived from 4 decades of historical census tract data 1970-2000, linked through geocoded residential histories (tract % poverty, tract % black, tract deprivation score derived from principal components analysis), as well as with 2 subjective neighborhood measures in adulthood. Our results documented that internal consistency reliability was high for the subjective childhood neighborhood scales (Cronbach’s α =0.89, 0.93). Comparison of subjective with objective childhood neighborhood measures found moderate associations in hypothesized directions. Associations with objective variables were strongest for neighborhood disorder (rhos=.40), as opposed to with social control or victimization. Associations between subjective neighborhood context in childhood versus adulthood were moderate and stronger for residentially-stable populations. We lastly formally tested for, but found little evidence of, recall bias of the retrospective subjective reports of childhood context. These results provide evidence that retrospective reports of subjective neighborhood context may be a cost-effective, valid, and reliable method to operationalize early life context for health studies.