JAK-mediated autophagy regulates stemness and cell survival in cisplatin resistant bladder cancer cells.

BACKGROUND: Autophagy is a critical process in acquiring drug resistance in solid tumors. However, the mechanisms by which autophagy modulate resistance to chemotherapy in bladder cancer remains poorly understood. MATERIAL AND METHODS: We have established cisplatin resistant patient derived primary cultured cells as well as T24 bladder cancer cells. The autophagy flux as well as the effect of chemotherapeutic agents, gemcitabine (GC) and mitomycin (MM) were evaluated in these cells. Side population analysis, tumorisphere assay and expression of stemness genes were also studied to evaluate the role of autophagy during chemoresistance. The role of cytokines in autophagy induced chemoresistance was demonstrated. RESULTS: Our results show that resistant cells have higher basal autophagic flux. GC and MM led to increase in autophagy and combination of autophagy inhibitors synergistically inhibited bladder cancer cell growth. When autophagy was inhibited, only IFN-γ level was decreased in cytokine array. Blocking IFN-γ decreased JAK2 expression and reduced the autophagy. Inhibition of JAK2 decreased the proportion of cancer stem cell enriched side population, tumor sphere forming ability and led to decrease in expression of stemness genes. Combined treatment with inhibitors of JAK2 and autophagy led to inhibition in cell growth, reduced the levels of inflammatory cytokines and decreased the resistance gene expression. CONCLUSION: GC and MM treatment increased autophagy process via IFN-γ mediated JAK2 and STAT3 pathway. GENERAL SIGNIFICANCE: These findings support a key role of the autophagy in drug resistance. Thus, autophagy-based, "customized" combinatorial approaches may effectively manage resistance and relapse in bladder cancer.

Bone mineral density among long-term survivors of childhood acute lymphoblastic leukemia: results from the St. Jude Lifetime Cohort Study.

BACKGROUND: The prevalence of low bone mineral density (BMD) in adult survivors of childhood acute lymphoblastic leukemia (ALL), and the degree of recovery or decline, are not well elucidated. PROCEDURE: Study subjects (age ≥ 18 years and ≥10 years post-diagnosis) participated in an institutional follow-up protocol and risk-based clinical evaluation based on Children's Oncology Group guidelines. Trabecular volumetric BMD was ascertained using quantitative computed tomography, reported as age- and sex-specific Z-scores. RESULTS: At median age 31 years, 5.7% of 845 subjects had a BMD Z-score of ≤-2 and 23.8% had a Z-score of -1 to -2. Cranial radiation dose of ≥24 Gy, but not cumulative methotrexate or prednisone equivalence doses, was associated with a twofold elevated risk of a BMD Z-score of ≤-1. The cranial radiation effect was stronger in females than in males. In a subset of 400 subjects, 67% of those who previously had a BMD Z-score of ≤-2 improved by one or more categories a median of 8.5 years later. CONCLUSIONS: Very low BMD was relatively uncommon in this sample of adult survivors of childhood ALL, and BMD Z-scores tended to improve from adolescence to young adulthood. High-dose cranial or craniospinal radiation exposure was the primary predictor of suboptimal BMD in our study. Given that cranial radiation treatment for childhood ALL is used far more sparingly now than in earlier treatment eras, concerns about persistently low BMD among most current childhood ALL patients may be unwarranted.

Parental risk perception and influenza vaccination of children in daycare centres.

Little information is available about perceptions of influenza vaccination of parents with healthy children in daycare. Therefore, we systematically explored the relationship between parental risk perception and influenza vaccination in children attending daycare. We distributed a self-administered paper survey to parents of children aged 6-59 months attending licensed daycare centres in Tarrant County, Texas. We used conditional logistic regression with penalized conditional likelihood to estimate odds ratios (ORs) and 95% profile likelihood confidence limits (PL) for parental risk-perception factors and influenza vaccination. A high level of parental prevention behaviours (OR 9.1, 95% PL 3.2, 31) and physician recommendation (OR 8.2, 95% PL 2.7, 30) had the highest magnitudes of association with influenza vaccination of healthy children in daycare. Our results provide evidence about critical determinants of influenza vaccination of healthy children in daycare, which could help inform public health interventions aimed at increasing influenza vaccination coverage in this population.

Platelet glycoprotein IaC807T polymorphisms and ischemic stroke in young Chinese Han population.

OBJECTIVES: The objective of this study was to investigate the association between platelet glycoprotein (GP) Ia C807T polymorphisms and ischemic stroke in young Chinese Han Population. MATERIALS AND METHODS: We conducted a case-control study in 92 consecutive young ( 50 years), and 160 age- and sex-matched healthy control. Genotyping of platelet GP Ia C807Tpolymorphisms was performed by polymerase chain reaction followed by sequencing nucleic acid with dideoxy chain-termination method and an ABI PRISM3100 (Perkin-Elmer Co) genetic analyzer. Student's t-test, chi-square test, and logistic regression modeling were used for data significance analyses. RESULTS: Hypertension and smoking were found to be the independent risk factors for ischemic stroke patients (aged 50 years). There was no significant difference observed in the T allele frequency of GPIa C807T polymorphisms between young stroke patients and corresponding controls. CONCLUSIONS: These findings suggest that there is no role of GPIa C807T polymorphisms in the development of young first-ever ischemic stroke in Chinese Han Population.

Swing phase considerations in prosthetic knee design: Case series to validate simulations.

BACKGROUND: Previously reported simulations comparing a new polycentric knee (called IPK) and a single-axis knee suggested that polycentricity could lead to improved performance during swing phase and negate the need for an extension assist. They also showed that an anteriorly translated socket adapter for enhanced stance stability compromises foot clearance during swing. OBJECTIVES: The objectives of this study are to validate those findings using gait trials to enable further improvement in the IPK design before mass production. METHODS: Three subjects regularly using three different passive knees, single-axis knee without extension assist, single-axis knee with extension assist (SAK-EA), and polycentric knee with extension assist (Mobility India-sourced polycentric knee), participated in this study. Their gait with their regular prosthesis and with IPK (having no extension assist) were analyzed, compared, and broadly correlated with simulation results. RESULTS: Extension assist in single-axis knee with extension assist improved swing performance, affected foot clearance in Mobility India-sourced polycentric knee, and was found to be unnecessary in the IPK. With an anteriorly placed socket adapter in the IPK, compensatory strategies were necessary for foot clearance. The IPK was found to provide better knee extension characteristics with lower hip effort (up to 42% reduction) than other knees. CONCLUSIONS: This case series confirmed previously reported simulation results on the swing phase behavior of passive prosthetic knees. The performance of the IPK during swing obviated the need for an extension assist, thereby simplifying the design. Appropriate design changes in the IPK's socket adapter location are required to achieve both stance stability and reduce gait compensations for foot clearance.

Quantum chemical study of gas-phase reactions of isoprene with OH radicals producing highly oxidised second-generation products.

The formation of secondary organic aerosols caused by atmospheric oxidation of isoprene is harmful to human health and the climate; thus, isoprene oxidation is further mandatory to obtain less harmful or harmless highly oxidised products. In this numerical investigation, 2-hydroperoxy-2-methylbut-3-en-1-ol (ISOPOOH) was considered the model compound to investigate the formation of three RO2 radicals (C5H11O4, C5H11O6 and C5H11O5) and two saturated highly oxidised products (C5H12O6 and C5H10O6). The complete reaction network and its thermodynamics and kinetics were analysed to obtain the most probable and feasible reaction pathways. Four different levels of theories (HF, B3LYP, M06-2X and ωB97XD with basis set of 6-31+g(d,p)) were employed to explore a global minimum of ISOPOOH. All theories provided approximately close energetics; however, because of the novelty of the functional and parameterisation of the basis set, the ωB97XD functional was selected to examine the reaction mechanism. C5H12O6 was formed as the second-generation highly oxidised product during ISOPOOH oxidation.

Statistical mechanics of a gas-fluidized particle.

Characterization of the microscopic fluctuations in systems that are far from equilibrium is crucial for understanding the macroscopic response. One approach is to use an 'effective temperature'--such a quantity has been invoked for chaotic fluids, spin glasses, glasses and colloids, as well as non-thermal systems such as flowing granular materials and foams. We therefore ask to what extent the concept of effective temperature is valid. Here we investigate this question experimentally in a simple system consisting of a sphere placed on a fine screen in an upward flow of gas; the sphere rolls because of the turbulence it generates in the gas stream. In contrast to many-particle systems, in which it is difficult to measure and predict fluctuations, our system has no particle-particle interactions and its dynamics can be captured fully by video imaging. Surprisingly, we find that the sphere behaves exactly like a harmonically bound brownian particle. The random driving force and frequency-dependent drag satisfy the fluctuation-dissipation relation, a cornerstone of statistical mechanics. The statistical mechanics of near-equilibrium systems is therefore unexpectedly useful for studying at least some classes of systems that are driven far from equilibrium.

Tobacco seedlings: damage by excessive nitrogen lessened by added phosphorus.

Concentrations of nitrogen (as ammonium nitrate) of 50 kilo-grams per hectare and greater are toxic to tobacco seedlings; symptoms are poor growth and white leaves. The leaves have markedly less chlorophylls (especially chlorophyll a) and more carotene and xanthophyll than seedlings grown in media low in nitrogen. These ill eflects are aggravated by short supply of phosphorus; they can be counteracted by more liberal supply of phosphorus.

Statistical characterization of the forces on spheres in an upflow of air.

The dynamics of a sphere fluidized in a nearly levitating upflow of air were previously found to be identical to those of a Brownian particle in a two-dimensional harmonic trap, consistent with a Langevin equation [Ojha et al., Nature (London) 427, 521 (2004)]. The random forcing, the drag, and the trapping potential represent different aspects of the interaction of the sphere with the air flow. In this paper we vary the experimental conditions for a single sphere, and report on how the force terms in the Langevin equation scale with air flow speed, sphere radius, sphere density, and system size. We also report on the effective interaction potential between two spheres in an upflow of air.

Effect of dyslipidemia on intima-media thickness of intra- and extracranial atherosclerosis by regulating the expression of hsp70 in rabbits.

The aim of this study was to explore the effect of dyslipidemia on intima-media thickness (IMT) of Intra- and extracranial atherosclerosis by regulating the expression of heat shock protein 70 (HSP70) in rabbits. Twenty-seven male white rabbits were randomly divided into normal control group A, high fat group B and high fat + endothelial injury operation group C (each group was 9), we measured lipids and obtained tissues from different cerebral arteries including Bilateral common carotid artery (CCA), Internal carotid artery (ICA), middle cerebral artery (MCA) and vertebral artery (VA). Pathological analysis were done, western blot analysis was used to detect the expression of HSP70 in CCA and MCA. The Serum lipid levels were overall significantly increased at 12(th) week in Group B and Group C compared to normal control (P < 0.05); at 12(th) week, the IMT of CCA and MCA in group B and C were showed significant increment compared with Group A; the correlation between HDL/CHOL/LDL and IMT of different cerebral arteries are as follows: MCA > ICA > CCA > VA; between TG and IMT of different cerebral arteries: VA > ICA > MCA > CCA; the expression of HSP70 from MCA were increased compared with CCA in group B and group C (P < 0.05). Significant positive correlations were observed between hyperlipidemia and different cerebral arteries. Hyperlipidemia has more impact on IMT of intracranial cerebral arteries. The expression of HSP70 from intracranial cerebral arteries is significantly increased. The mechanisms underlied was speculated that might be involved in inhibiting the inflammatory via HSP70.

Molecular dynamics simulation study of DNA triplex formed by mixed sequences in solution.

The unrestrained molecular dynamics simulation of the triple helical DNA with mix sequences d(GACTGGTGAC).d(CTGACCACTG)*d (GACTGGTGAC), using the particle mesh Ewald sum, is presented here. The Ewald summation method effectively eliminates the usualcut-of of the long range interactions and allowed us to evaluate the full effect of the electrostatic forces. The AMBER5.0 force field has been used during the simulation in solvent. The MD results support a dynamically stable model of DNA triplex over the entire length of the trajectory. The duplex structure assumes the conformation, which is very close to B-DNA. In mixed sequences the purine bases occurs in both strand of DNA duplex. The bases of third strand do not favor the Hoogsteen or/and reverse Hoogsteen type of Hydrogen bonding but they form hydrogen bonds with the bases of both the strand of DNA duplex. The orientation of the third strand is parallel to one of the strand of duplex and all nucleotides (C, A, G & T) show isomorphic behavior with respect to the DNA duplex. The conformation of all the three strands is almost same except few exceptions. Due to interaction of third strand the conformational change in the duplex structure and a finite amount of displacement in the W-C base pairs have been observed. The conformational variation of the back bone torsion angles and helicoidal parameters, groove widths have been discussed. The sequence dependent effects on local conformation, helicoidal and morphological structure, width of the grooves of DNA helix may have important implication for understanding the functional energetics and specificity of interactions of DNA and its triplexes with proteins, pharmaceutical agents and other ligands.

Structure of an anti-HIV-1 hammerhead ribozyme complex with a 17-mer DNA substrate analog of HIV-1 gag RNA and a mechanism for the cleavage reaction: 750 MHz NMR and computer experiments.

The structure of an anti-HIV-1 ribozyme-DNA abortive substrate complex was investigated by 750 MHz NMR and computer modeling experiments. The ribozyme was a chimeric molecule with 30 residues-18 DNA nucleotides, and 12 RNA residues in the conserved core. The DNA substrate analog had 17 residues. The chimeric ribozyme and the DNA substrate formed a shortened ribozyme-abortive substrate complex of 47 nucleotides with two DNA stems (stems I and III) and a loop consisting of the conserved core residues. Circular dichroism spectra showed that the DNA stems assume A-family conformation at the NMR concentration and a temperature of 15 degrees C, contrary to the conventional wisdom that DNA duplexes in aqueous solution populate entirely in the B-form. It is proposed that the A-family RNA residues at the core expand the A-family initiated at the core into the DNA stems because of the large free energy requirement for the formation of A/B junctions. Assignments of the base H8/H6 protons and H1' of the 47 residues were made by a NOESY walk. In addition to the methyl groups of all T's, the imino resonances of stems I and III and AH2's were assigned from appropriate NOESY walks. The extracted NMR data along with available crystallographic data, were used to derive a structural model of the complex. Stems I and III of the final model displayed a remarkable similarity to the A form of DNA; in stem III, a GC base pair was found to be moving into the floor of the minor groove defined by flanking AT pairs; data suggest the formation of a buckled rhombic structure with the adjacent pair; in addition, the base pair at the interface of stem III and the loop region displayed deformed geometry. The loop with the catalytic core, and the immediate region of the stems displayed conformational multiplicity within the NMR time scale. A catalytic mechanism for ribozyme action based on the derived structure, and consistent with biochemical data in the literature, is proposed. The complex between the anti HIV-1 gag ribozyme and its abortive DNA substrate manifests in the detection of a continuous track of A.T base pairs; this suggests that the interaction between the ribozyme and its DNA substrate is stronger than the one observed in the case of the free ribozyme where the bases in stem I and stem III regions interact strongly with the ribozyme core region (Sarma, R. H., et al. FEBS Letters 375, 317-23, 1995). The complex formation provides certain guidelines in the design of suitable therapeutic ribozymes. If the residues in the ribozyme stem regions interact with the conserved core, it may either prevent or interfere with the formation of a catalytically active tertiary structure.

Biological activity of 3-deazauridine nucleoside analogue: a theoretical study.

The incorporation model of Sanyal et al. has been used to understand the biological activity of the cytostatic compound 3-deazauridine. The interaction energies of various types of binding pattern of the enterant molecule with nucleic acid fragments have been computed. The energy values and the sites of association of the analogous base, obtained by optimization of energy values as well as the sites of association of nucleic acid bases during the transcription process have been compared. The specificity of the binding of the interacting molecule has been discussed, along with the inhibitory effect of 3-deazauridine. They are in agreement with the experimentally observed evidence.

Myopia and plasma cortisol.

Estimation of plasma cortisol by fluorometric technique desorbed by Mallingly's was carried out in 56 cases, including 38 cases of myopia (19 cases of simple myopia and 19 cases of degenerative myopia) and 18 normal individuals. Urinary 17-keto steroids/24 hours was also estimated by Zimmermann technique in 12 out of the 56 cases, which include 8 cases of myopia (4 cases of simple myopia and 4 cases of degenerative myopia) and 4 normal individuals as control. Plasma cortisol level is lower in simple and degenerative myopia than in normal subjects, but on statistical analysis the difference was not of much significance. Patients with simple myopia with positive family history showed lower value of plasma cortisol than patients with simple myopia with negative family history, the difference was also statistically insignificant (P-0.1). In degenerative myopia patients, with and without family history, there was very little difference in plasma cortisol level and statistically highly insignificant (P-0.8). Urinary 17 keto steroid/24 hour values are lower in simple and degenerative myopia than in normal subjects.

Amsacrine binds in a cooperative manner to deoxyribonucleic acid.

The intercalative binding of the acridine antitumour drug 4'-(9-acridinylamino) methane-sulphonate-m-anisidine, a known inhibitor of nucleic acid synthesis, to native calf thymus DNA has been studied using optical titration method. Amsacrine (AMSA) exhibits positive cooperativity in their equilibrium binding to DNA as indicated by the positive slope in the initial region of the binding isotherms (Scatchard plots) under conditions simulating physiological ionic strengths. m-AMSA binds with a higher degree of cooperativity than o-AMSA. Although this correlates with the effectiveness of the drugs as antitumour agents, the exact relationship between the observation of cooperative binding and pharmacological activity is yet to be determined.

Mode of action of intercalators: a theoretical study.

Mode of action of some intercalators has been theoretically investigated on the basis of quantum mechanical perturbation method. Energies of H-bond interaction between drug chromophore and base pairs have been calculated in all possible orientations. The stacking energy has also been calculated with the base pairs. The effect of these interactions on specific recognition has also been discussed. On the basis of these studies, a model for the interaction of these drugs has been proposed. This explains the relative activities of acridine intercalators and satisfies the experimental observations.

An investigation of the conformation of peptide-T and its D-Ala analog by NMR and molecular dynamics simulations.

Peptide-T (ASTTTNYT) and its D-Ala analog (D-ASTTTNYT-NH2) have been designed to block the adsorption of HIV to CD4 receptors on T-cell lymphocytes, thus inhibiting viral infectivity. The conformation of these important peptides has been investigated by 2D-NMR and molecular dynamics simulations. The NMR studies in DMSO show that the peptides exist in solution as a mixture of conformations. beta-Turns and non-specific folded conformations are present in a small proportion in the ensemble of conformations, which is largely dominated by more or less extended structures. This result is in line with molecular dynamics simulations where beta-turns were found to occur with a low frequency and with energies 10 to 17 kcal/mole higher than the global minimum structure. Our findings differ from previous reports on the conformation of peptide-T determined by NMR.

Neonatal presentation of incontinentia pigmenti with a family history extending over four generations--a case report.

We report a case of Incontinentia pigmenti (IP). A day 2 female presented to the special care nursery with seizures. EEG showed multifocal epileptiform discharges and cranial MRI revealed extensive cerebral infarction. A rash appeared shortly after birth. Eye examination revealed changes of IP. There is a strong family history of IP. Genetic testing excluded the most common mutation. Follow-up reveals significant development delay.