The role of spin in biological processes: O2, NO,nucleobases, nucleosides,nucleotides and Watson–Crick base pairs

The experimental electron affinities of adenine, guanine, cytosine, thymine and uracil have been determined from reduction potentials and negative ion photoelectron spectra. Updated values for purine, pyrimidine and other nitrogen heterocyclics, which have not been measured in the gas phase, are presented. The electron affinity of Watson–Crick guanine–cytosine is estimated empirically. The experimental values are consistent with quantum mechanical semi-empirical multiconfiguration configuration interaction calculations. The bulk hydration energies of the nucleobase anions, 2.34 eV, determined from the experimental data and sequential anion hydration energy difference of about 0.20(5) eV suggest that 10–15 water molecules complete the hydration shell. The electron affinities for the formation of doublet and quartet anions of the nucleobases, nucleosides, nucleotides and Watson–Crick base pairs are calculated. We postulate that low-lying quartet anion states and their spin distribution can and will participate in electron conduction, radiation damage, oxidation damage and repair, strand breakage and protein synthesis.     

Aerosol generation using nanometer liposome suspensions for pulmonary drug delivery applications

Abstract

Pulmonary lung targeting finds applications in drug delivery to the lung itself and to other body organs, via blood circulation following transfer across alveolar membranes. Understanding pulmonary drug delivery systems towards improving their efficacy needs identification of particle sizes of relevance and elucidation of links between suspension properties, techniques of atomisation and properties of the generated aerosols. This review article is focussed on understanding the elements of pulmonary drug delivery, specifically related to suspensions of small liposomes. Specific objectives of this review include (a) understanding aerosol particle deposition and absorption on pulmonary surface, (b) links between properties of aerosol generation and colloidal drug carriers used for drug encapsulation, and (c) investigation on the controlled properties of liposome aerosols generated using different atomisation techniques for efficacious aerosol therapy.     

Physicochemical study of the protein–liposome interactions: influence of liposome composition and concentration on protein binding

Abstract

The aim of the present study is to investigate the interactions between liposomes and proteins and to evaluate the role of liposomal lipid composition and concentration in the formation of protein corona. Liposomes composed of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) or hydrogenated soybean phosphatidylcholine (HSPC) with 1,2-dipalmitoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (sodium salt) (DPPG), 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-3000] (DPPE-PEG 3000), cholesterol (CH) or mixtures of these lipids, were prepared at different concentrations by the thin-film hydration method. After liposomes were dispersed in HPLC-grade water and foetal bovine serum (FBS), their physicochemical characteristics, such as size, size distribution, and ζ-potential, were determined using dynamic and electrophoretic light scattering. Aggregation of DPPC, HSPC, DPPC:CH (9:1 molar ratio), and HSPC:CH (9:1 molar ratio) in FBS was observed. On the contrary, liposomes incorporating DPPG lipids and CH both in a molar ratio of 11% were found to be stable over time, while their size did not alter dramatically in biological medium. Liposomes containing CH and PEGylated lipids retain their size in the presence of serum as well as their physical stability. In addition, our results indicate that the protein binding depends on the presence of polyethylene glycol (PEG), CH, concentration and surface charge. In this paper, we introduce a new parameter, fraction of stealthiness (F_s), for investigating the extent of protein binding to liposomes. This parameter depends on the changes in size of liposomes after serum incubation, while liposomes have stealth properties when F_s is close to 1. Thus, we conclude that lipid composition and concentration affect the adsorption of proteins and the liposomal stabilization.     

Flow cytometry and live confocal analysis for the evaluation of the uptake and intracellular distribution of FITC-ODN into HaCaT cells

In this study, the mechanism of the internalization and the cellular distribution of 59 fluorescein conjugated PS-ODN (FITC-ODN) after transfection with different mixed lipidic vesicles/oligo complexes (lipoplexes) have been investigated. Mixed lipidic vesicles were prepared with one of the most used cationic lipid (DOTAP) and different amounts of a cholic acid (UDCA) to release the oligo into HaCaT cells. Using flow cytometry, the cellular uptake of the oligo was studied with and without different inhibitors able to block selectively the different pathways involved in the internalization mechanism. The intracellular distribution of the oligo was analyzed by confocal laser scanning microscopy (CLSM), treating the cells with the lipoplexes and directly observing without any fixing procedure. To better carry out the colocalization studies, fluorescent-labeled markers, specific for the different cellular compartments, were coincubated with 59 fluorescein-conjugated 29-mer phosphorotioate oligonucleotide (FITC-ODN). The different lipidic vesicles affect the internalization mechanism of FITC-ODN. After using the inhibitors, the uptake of complexes involved a different internalization mechanism. The live CLSM analysis demonstrated that, after 1 hour from the complex incubation, the oligo was transferred into cells and localized into the endosomes; after 24 hours, the oligo was intracellularly localized close to the nuclear structure in a punctuate pattern. However, the results from fusion experiments showed also a binding of a quite low amount of oligo with the cell membranes.     

Multispectroscopic studies of the interaction of folic acid with glycated human serum albumin

Abstract

The interaction between glycated human serum albumin (gHSA) and folic acid (FA) was investigated by various spectroscopic techniques, such as fluorescence, circular dichroism, UV–vis absorption spectroscopy and electrophoretic light scattering technique. These methods characterize the binding properties of an albumin–folic acid system. The binding constants values (K_a) at 300 and 310 K are about 10⁴ M^?1. The standard enthalpy change (ΔH) and the standard entropy change (ΔS) were calculated to be ～?20?kJ mol^?1 and ～16 J mol^?1 K^?1, respectively, which indicate characteristic electrostatic interactions between gHSA and folic acid. The CD studies showed that there are no significant conformational changes in the secondary structure of the protein. Moreover, the zeta potential measurements proved that under physiological conditions the gHSA–folic acid complex shows instability. No significant changes in the secondary structure of the protein and reversible drug binding are the desirable effect from pharmacological point of view.

Communicated by Ramaswamy H. Sarma     

FAILURE OF EXTRAOCULAR LIGHT TO FACILITATE CIRCADIAN RHYTHM REENTRAINMENT IN HUMANS

Although extraocular light can entrain the circadian rhythms of invertebrates and nonmammalian vertebrates, almost all studies show that the mammalian circadian system can only be affected by light to the eyes. The exception is a recent study by Campbell and Murphy that reported phase shifts in humans to bright light applied with fiber-optic pads behind the knees (popliteal region). We tested whether this extraocular light stimulus could accelerate the entrainment of circadian rhythms to a shift of the sleep schedule, as occurs in shift work or jet lag. In experiment 1, the sleep/dark episodes were delayed 8h from baseline for 2 days, and 3h light exposures were timed to occur before the temperature minimum to help delay circadian rhythms. There were three groups: (1) bright (about 13,000 lux) extraocular light from fiber-optic pads, (2) control (dim light, 10–20 lux), and (3) medium-intensity (about 1000 lux) ocular light from light boxes. In experiment 2, the sleep/dark episodes were inverted, and extraocular light was applied either before the temperature minimum to help delay circadian rhythms or after the temperature minimum to help advance rhythms. Circadian phase markers were the salivary dim light melatonin onset (DLMO) and the rectal temperature minimum. There was no evidence that the popliteal extraocular light had a phase-shifting effect in either experiment. Possible reasons for phase shifts in the Campbell and Murphy study and not the current study include the many differences between the protocols. In the current study, there was substantial sleep deprivation before the extraocular light was applied. There was a large shift in the sleep/dark schedule, rather than allowing subjects to sleep each day from midnight to noon, as in the Campbell and Murphy study. Also, when extraocular light was applied in the current protocol, subjects did not experience a change from sleeping to awake, a change in posture (from lying in bed to sitting in a chair), or a change in ocular light (from dark to dim light). Further research is necessary to determine the conditions under which extraocular light might produce phase shifts in human circadian rhythms. (Chronobiology International, 17(6), 807–826, 2000).     

Shining light on nanotechnology to help repair and regeneration

Phototherapy can be used in two completely different but complementary therapeutic applications. While low level laser (or light) therapy (LLLT) uses red or near-infrared light alone to reduce inflammation, pain and stimulate tissue repair and regeneration, photodynamic therapy (PDT) uses the combination of light plus non-toxic dyes (called photosensitizers) to produce reactive oxygen species that can kill infectious microorganisms and cancer cells or destroy unwanted tissue (neo-vascularization in the choroid, atherosclerotic plaques in the arteries). The recent development of nanotechnology applied to medicine (nanomedicine) has opened a new front of advancement in the field of phototherapy and has provided hope for the development of nanoscale drug delivery platforms for effective killing of pathological cells and to promote repair and regeneration. Despite the well-known beneficial effects of phototherapy and nanomaterials in producing the killing of unwanted cells and promoting repair and regeneration, there are few reports that combine all three elements i.e. phototherapy, nanotechnology and, tissue repair and regeneration. However, these areas in all possible binary combinations have been addressed by many workers. The present review aims at highlighting the combined multi-model applications of phototherapy, nanotechnology and, reparative and regeneration medicine and outlines current strategies, future applications and limitations of nanoscale-assisted phototherapy for the management of cancers, microbial infections and other diseases, and to promote tissue repair and regeneration.     

A Personal Light-Treatment Device for Improving Sleep Quality in the Elderly: Dynamics of Nocturnal Melatonin Suppression at Two Exposure Levels

Light treatment has been used as a non-pharmacological tool to help mitigate poor sleep quality frequently found in older people. In order to increase compliance to non-pharmacological light treatments, new, more efficacious light-delivery systems need to be developed. A prototype personal light-treatment device equipped with low brightness blue light-emitting diodes (LEDs) (peak wavelength near 470 nm) was tested for its effectiveness in suppressing nocturnal melatonin, a measure of circadian stimulation. Two levels of corneal irradiance were set to deliver two prescribed doses of circadian light exposure. Eleven older subjects, between 51 and 80 yrs of age who met the selection criteria, were exposed to a high and a low level of light for 90 min on separate nights from the personal light-treatment device. Blood and saliva samples were collected at prescribed times for subsequent melatonin assay. After 1 h of light exposure, the light-induced nocturnal melatonin suppression level was about 35% for the low-light level and about 60% for the high-light level. The higher level of blue light suppressed melatonin more quickly, to a greater extent over the course of the 90 min exposure period, and maintained suppression after 60 min. The constant exposure of the low-light level resulted in a decrease in nocturnal melatonin suppression for the last sampling time, whereas for the high-light level, suppression continued throughout the entire exposure period. The present study performed with healthy adults suggests that the tested personal light-treatment device might be a practical, comfortable, and effective way to deliver light treatment to those suffering from circadian sleep disorders; however, the acceptance and effectiveness of personal light-treatment devices by older people and by other segments of the population suffering from sleep disorders in a real-life situation need to be directly tested. (Author correspondence: figuem@rpi.edu)