Spectroscopic Characterization of Mitochondrial G-Quadruplexes.

Guanine quadruplexes (G4s) are highly polymorphic four-stranded structures formed within guanine-rich DNA and RNA sequences that play a crucial role in biological processes. The recent discovery of the first G4 structures within mitochondrial DNA has led to a small revolution in the field. In particular, the G-rich conserved sequence block II (CSB II) can form different types of G4s that are thought to play a crucial role in replication. In this study, we decipher the most relevant G4 structures that can be formed within CSB II: RNA G4 at the RNA transcript, DNA G4 within the non-transcribed strand and DNA:RNA hybrid between the RNA transcript and the non-transcribed strand. We show that the more abundant, but unexplored, G6AG7 (37%) and G6AG8 (35%) sequences in CSB II yield more stable G4s than the less profuse G5AG7 sequence. Moreover, the existence of a guanine located 1 bp upstream promotes G4 formation. In all cases, parallel G4s are formed, but their topology changes from a less ordered to a highly ordered G4 when adding small amounts of potassium or sodium cations. Circular dichroism was used due to discriminate different conformations and topologies of nucleic acids and was complemented with gel electrophoresis and fluorescence spectroscopy studies.

A Surfactant Concentration Model for the Systematic Determination of the Critical Micellar Concentration and the Transition Width.

The critical micellar concentration (cmc) is a fundamental property of surfactant solutions. Many proposed methods for the definition and determination of the cmc from property-concentration plots yield values, which depend on the studied property, on the specific technique used for its analysis and in many cases on the subjective choice of the chosen type of plot and concentration interval. In this focus review, we revise the application of a surfactant concentration model we proposed earlier that defines the cmc directly based on the surfactant concentration. Known equations for the concentration-dependence of different surfactant properties can then be combined with this concentration model and fitted to experimental data. This modular concept makes it possible to determine the cmc and the transition width in a systematic and unambiguous way. We revise its use in the literature in different contexts: the determination of the cmc of surfactants and their mixtures from different properties (electrical conductivity, NMR chemical shift, self-diffusion, surface tension, UV-Vis absorption, fluorescence intensity and fluorescence correlation). We also revise the dependence of the width of the transition region on composition, detailed studies of the properties of fluorescent probes and the aggregation of non-surfactant systems, namely amyloid peptides.

Fluorescence-labeled bis-benzamidines as fluorogenic DNA minor-groove binders: photophysics and binding dynamics.

In recent decades there has been great interest in the design of highly sensitive sequence-specific DNA binders. The eligibility of the binder depends on the magnitude of the fluorescence increase upon binding, related to its photophysics, and on its affinity and specificity, which is, in turn, determined by the dynamics of the binding process. Therefore, progress in the design of DNA binders requires both thorough photophysical studies and precise determination of the association and dissociation rate constants involved. We have studied two bis-benzamidine (BBA) derivatives labeled by linkers of various lengths with the dye Oregon Green (OG). These fluorogenic binders show a dramatic fluorescence enhancement upon binding to the minor groove of double-stranded (ds) DNA, as well as significant improvement in their sequence specificity versus the parent BBA, although with decreased affinity constants. Detailed photophysical analysis shows that static and dynamic quenching of the OG fluorescence by BBA through photoinduced electron transfer is suppressed upon insertion of BBA into the minor groove of DNA. Fluorescence correlation spectroscopy yields precise dynamic rate constants that prove that the association process of these fluorogenic binders to dsDNA is very similar to that of BBA alone and that their lower affinity is mainly a consequence of their weaker attachment to the minor groove and the resultant faster dissociation process. The conclusions of this study will allow us to go one step further in the design of new DNA binders with tunable fluorescence and binding properties.

Towards ratiometric sensing of amyloid fibrils in vitro.

The aggregation of amyloid-ß peptide and its accumulation in the human brain has an important role in the etiology of Alzheimer's disease. Thioflavin T has been widely used as a fluorescent marker for these amyloid aggregates. Nevertheless, its complex photophysical behavior, with strong wavelength dependencies of all its fluorescence properties, requires searching for new fluorescent probes. The use of 2-(2'-hydroxyphenyl)imidazo[4,5-b]pyridine (HPIP), which shows two emission bands and a rich excited-state behavior due to the existence of excited-state intramolecular processes of proton transfer and charge transfer, is proposed. These properties result in a high sensitivity of HPIP fluorescence to its microenvironment and cause a large differential fluorescence enhancement of the two bands upon binding to aggregates of the amyloid-ß peptide. Based on this behavior, a very sensitive ratiometric method is established for the detection and quantification of amyloid fibrils, which can be combined with the monitoring of fluorescence anisotropy. The binding selectivity of HPIP is discussed on the basis of the apparent binding equilibrium constants of this probe to amyloid-ß (1-42) fibrils and to the nonfibrillar protein bovine serum albumin. Finally, an exhaustive comparison between HPIP and thioflavin T is presented to discuss the sensitivity and specificity of these probes to amyloid aggregates and the significant advantages of the HPIP dye for quantitative determinations.

Quick adjustment of imaging tracer payload, for in vivo applications of theranostic nanostructures in the brain.

In order to provide sufficient sensibility for detection, selection of an adequate payload of imaging probe is critical, during the design of MRI theranostic nanoplatforms. This fact is particularly crucial for in vivo applications in the brain, where delivery of macromolecules is limited by the blood-brain barrier. Here we report a simple and quick process for the estimation of adequate payloads of gadolinium in liposomes with potential to act as theranostic agents, for in vivo MRI applications in the brain. Our studies show that an excessive payload of gadolinium in liposomes may actually have a negative influence on in vivo T1 contrast. By preparing and characterizing 4 different liposomal compositions of increasing Gadolinium loads, we show that a superior sensitivity for in vivo detection of MRI theranostic molecules can be quickly improved by adjusting the payload of imaging probe in the molecules. FROM THE CLINICAL EDITOR: This team of authors report the development of a simple and quick process for the estimation of adequate payloads of gadolinium in liposomes as theranostic agents for in vivo brain MRI studies, using a rodent model.

Validating the Enright Forgiveness Inventory in Ecuador.

Research on forgiveness is limited in Ecuador. This study validated the Enright Forgiveness Inventory-30 (EFI-30) among 960 participants in Ecuador, resulting in robust reliability and validity values. Our findings provide avenues for future research and practices.

Early Aggregation of Amyloid-ß(1-42) Studied by Fluorescence Correlation Spectroscopy.

Alzheimer's disease (AD) is a progressive neurodegenerative disease affecting cognitive and memory abilities and is believed to be linked to the formation and accumulation of neurotoxic aggregates of the Amyloid-ß peptide (Aß). In particular, it is the formation of soluble pre-fibrillar oligomers within the early stage of Aß aggregation which is thought to represent a key step in the development of AD, thus underlining the interest in characterizing the aggregation process and the nature of these aggregates. In this context, fluorescence correlation spectroscopy (FCS) has emerged as a valuable alternative for the study of these systems in solution. Indeed, the use of FCS to study terminally labelled Aß provides a means to detect changes in the size and concentration of initially monomeric Aß samples by monitoring these fluorescently labelled species freely diffusing in solution with single-molecule resolution. Herein, we show how to employ FCS to study the early aggregation process of Aß(1-42) and how this can be used to estimate the critical concentration for oligomer formation and to characterize the aggregates formed.

Supramolecular association studied by Fluorescence correlation spectroscopy.

A comprehensive description of a supramolecular system involves a full understanding of its thermodynamic and dynamic properties, as well as detailed knowledge of its structure. Fluorescence Correlation Spectroscopy (FCS) constitutes a powerful technique to acquire this information. Fluorescence correlation curves show a characteristic diffusion term that is related to the binding equilibrium constant or other thermodynamic properties of the supramolecular system. The association and dissociation rate constants of the binding process can be determined in FCS when the relaxation time of the binding is faster than the observation time-a regime called fast-exchange dynamics - in opposition to the slow-exchange regime. In all cases, structural information can be inferred from the diffusional properties of the supramolecular complexes. A short overview of the use of FCS for the study of supramolecular systems is given with examples which belong to the fast and slow regime.

Study of informal reasoning in judicial agents in sexual aggression cases.

Background/Objective: Judicial decisions must rest on formal reasoning. Nevertheless, informal reasoning sources (cognitive and motivational biases) were observed in judicial judgment making. Literature has identified sexual aggression cases as the most favorable for informal reasoning. Thus, a field study was designed with the aim of assessing the incidence and effects of cognitive and motivational biases in judicial agents in a case to rape to a woman. Methods: As for this, Chilean judicial agents (N = 217) assessed an allegation (weak evidence) of sexual assault in a case where the perpetrator was known or unknown to the victim. The judicial agents answered to a measure of the myths about sexual aggression, the attribution of responsibility to complainant, the attribution of responsibility to accused, the attribution of credibility to the complainant testimony, the attribution of a nature of a rape to the alleged facts and an estimation of the probability of false/unfounded accusations. Results: The results revealed an estimation of false/unfounded accusations of sexual aggression significantly higher than the mean of the best estimates, but into the upper limit of the best estimates; that the studied population did not share, in general, the myths about sexual aggression; and that the sources of attributional biases were driven in favor and against the complainant. Nevertheless, the case study showed that a large number of judicial agents participated of an overestimation of the probabilities of false or unfounded allegations, and of the myths about sexual aggressions and of attributional biases against the complainant. Conclusion: In conclusion, informal reasoning sources were observed in judicial agents when only formal reasoning should prevail. Thus, judicial agents should be trained to control these sources of bias substituting them by formal reasoning (evidence).

Estimating the Psychological Harm Consequence of Bullying Victimization: A Meta-Analytic Review for Forensic Evaluation.

The prevalence of traditional bullying victimization has been estimated at around 36%, while that of cyberbullying has been estimated at 15%. The victimization of bullying brings with it harm to mental health that must be compensated for, after a forensic evaluation, by the aggressor or legal guardian. Thus, a meta-analytic review was undertaken with the aim of knowing the effect of bullying victimization on psychological harm, as well as quantifying the magnitude of the harm and estimating the probability that no harm associated with bullying victimization is generated. METHOD: A random-effects correlational meta-analysis correcting effect size by sampling error and criterion and predictor unreliability was performed. RESULTS: The results exhibited a positive (i.e., more victimization and more psychological harm) and significant mean true effect size, implying an average psychological harm associated to bullying victimization of 29.7%. Nevertheless, 26.7% of students victimized by bullying did not develop psychological harm. CONCLUSIONS: Bullying victimization causes psychological harm, with an average increase in psychological harm associated with bullying victimization of 29.7%.

Role of electrostatic and hydrophobic forces in the interaction of ionic dyes with charged micelles.

The nature and strength of the interactions between a cationic fluorophore, Rhodamine 123 (R123), and surfactants of different head charge are investigated. Series of absorption and fluorescence emission spectra and of fluorescence decays are measured. R123 does not interact with the monomers of both nonionic and cationic surfactants but it presents affinity to their micelles. A partition equilibrium model was proposed and the corresponding equilibrium constants were obtained, as well as the photophysical properties of the dye bound to the micelles. In the case of the cationic surfactants, changes of the fluorescence properties were already observed below the critical micellar concentration (CMC) due to dynamic quenching caused by the free counterions. In the presence of anionic surfactants R123 shows a very different behaviour with dramatic spectral changes below the CMC. The observed variations are attributed to a first, strong interaction between R123 and the surfactant monomers, which yields ionic pairs of singular photophysical properties and dominates at low surfactant concentrations, followed by the association of R123 with the surfactant premicellar and micellar aggregates at higher surfactant concentrations near the CMC. This behaviour results from the competition between the strong electrostatic interactions of the cationic dye with the anionic surfactant head groups and the hydrophobic forces stabilizing the dye inside the micelles. The results of this work illustrate the complex physicochemical and photophysical behaviour of a charged dye in micellar systems, which resembles the expected situation in similar systems such as biological membranes.

Host-guest complexation studied by fluorescence correlation spectroscopy: adamantane-cyclodextrin inclusion.

The host-guest complexation between an Alexa 488 labelled adamantane derivative and beta-cyclodextrin is studied by Fluorescence Correlation Spectroscopy (FCS). A 1:1 complex stoichiometry and a high association equilibrium constant of K = 5.2 x 10(4) M(-1) are obtained in aqueous solution at 25 degrees C and pH = 6. The necessary experimental conditions are discussed. FCS proves to be an excellent method for the determination of stoichiometry and association equilibrium constant of this type of complexes, where both host and guest are nonfluorescent and which are therefore not easily amenable to standard fluorescence spectroscopic methods.

Adolescent-to-Parent Violence: Psychological and Family Adjustment.

Adolescent-to-Parent Violence (APV) or Child-to-Parent Violence (CPV) is a specific form of violence that has remained inconspicuous until recently, but is becoming a mounting social issue and is increasingly the focus of scientific research. Of the variables related to APV, the study assessed the characteristics of the family system and its relationship to the psychosocial adjustment of adolescents, an aspect scarcely examined in the literature. Thus, a field study was performed on a community sample of 210 adolescents aged 12-17 years (51.4% girls) who were assessed on measurements of APV, parenting (parental socialization), victimization, and psychological adjustment (personal, family, and school). The results revealed higher rates of psychological APV, and no gender effects in violence exercised against either parent. The adolescents involved in APV exhibited a greater psychological maladjustment in the different areas under analysis. Moreover, adolescents engaging in psychological APV reported a parental socialization style characterized by severe strictness and supervision in comparison to non-aggressors not implicated in psychological APV. Finally, adolescents exercising APV who were victimized by their parents showed more psychological, personal, and school maladjustment. These results have implications for needs analysis and the planning of community prevention strategies.

Host-assisted guest self-assembly: enhancement of the dimerization of pyronines Y and B by gamma-cyclodextrin.

Buckle up! The dimerization of small fluorescent guests is strongly enhanced in presence of a cyclodextrin host. The host cavity acts like a belt to assist the self-assembly of guests (see picture). Small variations in the guest structure have significant influence on the stability and geometry of the aggregates.The role of small variations in the structural properties of host and guest molecules on the stoichiometry and strength of supramolecular associations is analyzed. Earlier we found that a change in substituents from pyronine B to pyronine Y has a dramatic effect on both the stability and the dynamics of the association of these guests with beta-cyclodextrin as host. Now we study the association between these two pyronines and a cyclodextrin with a bigger cavity (gamma-cyclodextrin) using UV/Vis absorption and fluorescence spectroscopy. The absorption spectra of the pyronines show complex variations with cyclodextrin concentration indicating that pyronine dimerization is strongly enhanced inside the cavity of the cyclodextrin. A full model is proposed and the equilibrium constants of the involved processes and the absorption and emission spectra of the different species are estimated. The equilibrium constants of the formation of complexed dimers are much higher than those for free dimerization or for the inclusion of a single guest. The gamma-cyclodextrin host acts like a belt to assist the guest self-assembly. The differences in the stability of pyronine B and pyronine Y dimers are explained on the basis of their structure and geometry.

[Creation and validation of a forensic protocol to assess psychological harm in battered women]. / Creación y validación de un protocolo de evaluación forense de las secuelas psicológicas de la violencia de género.

Psychological harm is one of the sequela of the victimization of a crime. Nevertheless, a diagnosis of psychological harm, that is, posttraumatic stress disorder (PTSD), does not constitute sufficient evidence alone given that, in legal terms, faking or false testimony must be detected and eliminated before an expert testimony can be admitted. To assess the aptitudes and strategies for faking psychological harm associated with domestic violence, 101 women were asked to fake psychological harm using the MMPI-2 and a forensic clinical interview. The results showed that most women (84.2%) were capable of faking in MMPI-2. The original validity scales of the MMPI-2 and their configurations were effective for the detection of faking though with a wide margin of error was observed. In the forensic clinical interview, it was much more difficult to simulate since only 3 malingerers, in which protocols were not observed any feigning strategy, were capable of simulating the psychological injury linked to domestic violence. Consequently, a multimethod approach (MMPI and interview) was contrasted and observed to classify correctly all subjects. Thus, a protocol for forensic assessment of the psychological harm consequence of domestic violence against women was formulated.

Assessing the effects of an education program on mental health problems in separated parents.

BACKGROUND: Parental separation is a stressful experience that can lead to parents suffering mental health problems (MHPs). Parental separation education programs for coping with post-separation adjustment have proven to be effective in reducing conflict and improving co-parenting. However, the effects of these programs on MHPs have not been assessed. A field study was carried out to assess the impact of a parental separation education program on parental MHPs. METHOD: A total of 116 separated parents who completed the program "Parental separation, not family breakdown" completed the Brief Symptom Inventory (BSI) pre- and post-intervention. RESULTS: Separated parents had significantly higher pre-intervention scores on the nine symptom dimensions and the global indexes of distress in comparison to the normative population. The intervention yielded a significant improvement (i.e., reduction of clinical symptoms) in all MHPs, ranging from 19% in phobic anxiety to 36% in depression and general anxiety; and in the global indexes of distress (36% in the global severity index; 28% in the positive symptom distress index; and 33% in the positive symptom total). Approximately 45% of parents significantly improved through the intervention. CONCLUSIONS: The implications of the outcomes of the separation and intervention in parents' MHPs and children wellbeing are discussed.

Unveiling the multi-step solubilization mechanism of sub-micron size vesicles by detergents.

The solubilization of membranes by detergents is critical for many technological applications and has become widely used in biochemistry research to induce cell rupture, extract cell constituents, and to purify, reconstitute and crystallize membrane proteins. The thermodynamic details of solubilization have been extensively investigated, but the kinetic aspects remain poorly understood. Here we used a combination of single-vesicle Förster resonance energy transfer (svFRET), fluorescence correlation spectroscopy and quartz-crystal microbalance with dissipation monitoring to access the real-time kinetics and elementary solubilization steps of sub-micron sized vesicles, which are inaccessible by conventional diffraction-limited optical methods. Real-time injection of a non-ionic detergent, Triton X, induced biphasic solubilization kinetics of surface-immobilized vesicles labelled with the Dil/DiD FRET pair. The nanoscale sensitivity accessible by svFRET allowed us to unambiguously assign each kinetic step to distortions of the vesicle structure comprising an initial fast vesicle-swelling event followed by slow lipid loss and micellization. We expect the svFRET platform to be applicable beyond the sub-micron sizes studied here and become a unique tool to unravel the complex kinetics of detergent-lipid interactions.

Dynamics of supramolecular association monitored by fluorescence correlation spectroscopy.

Supramolecular binding is a key process in many biological systems and in newly developed supramolecular assemblies. Most of the scientific work on these systems is focused on their structural properties and on the thermodynamics of the association process. However, the underlying dynamics are usually much less known, in spite of the great importance they have during the binding process in these highly dynamic systems. Understanding supramolecular binding in biological systems and controlling the functionality of new synthetic supramolecular systems can only be achieved through knowledge of the structure-dynamics relationship. There is a strong need for suitable techniques which cover the typically wide time interval of the association dynamics and which do not need a perturbation of the system. We briefly review high-resolution fluorescence correlation spectroscopy (FCS) as a technique to monitor supramolecular dynamics and to give information on how structure determines the dynamics of host-guest association. The comparison of hosts and guests with different structures shows that geometrical and orientational requirements determine the association rate constant, whereas the dissociation is defined by the strength of specific interactions. As model hosts cyclodextrins and micelles are studied.

Neighborhood and community factors: effects on deviant behavior and social competence.

Socialization in a neighborhood and community at risk, defined in terms of violence, social alienation, school failure, and disruptive behavior, is a risk factor for the acquisition of antisocial and delinquent behavior. In order to test this hypothesis and examine the underlying mechanisms involved, 346 participants, 155 high-risk and 191 low-risk, aged 11 to 13, that is, under the age of criminal responsibility as established by the Spanish Law 5/2000 were selected. The results reveal that high-risk youngsters had higher rates of antisocial behavior and lower levels of social skills (i.e., greater tendency to externalize attribution of responsibility, fewer conflict resolution strategies, lower self esteem, and a lower degrees of emotional intelligence) in comparison to the lower-risk group. Finally, the results and implications of the study are discussed in the light of designing prevention programs.

Critical aggregation concentration for the formation of early Amyloid-ß (1-42) oligomers.

The oligomers formed during the early steps of amyloid aggregation are thought to be responsible for the neurotoxic damage associated with Alzheimer's disease. It is therefore of great interest to characterize this early aggregation process and the aggregates formed, especially for the most significant peptide in amyloid fibrils, Amyloid-ß(1-42) (Aß42). For this purpose, we directly monitored the changes in size and concentration of initially monomeric Aß42 samples, using Fluorescence Correlation Spectroscopy. We found that Aß42 undergoes aggregation only when the amount of amyloid monomers exceeds the critical aggregation concentration (cac) of about 90 nM. This spontaneous, cooperative process resembles surfactants self-assembly and yields stable micelle-like oligomers whose size (≈50 monomers, R h ≈ 7-11 nm) and elongated shape are independent of incubation time and peptide concentration. These findings reveal essential features of in vitro amyloid aggregation, which may illuminate the complex in vivo process.