Development of stable cell lines for production or regulated expression using matrix attachment regions.

One of the major hurdles of isolating stable, inducible or constitutive high-level producer cell lines is the time-consuming selection procedure. Given the variation in the expression levels of the same construct in individual clones, hundreds of clones must be isolated and tested to identify one or more with the desired characteristics. Various boundary elements (BEs), matrix attachment regions, and locus control regions (LCRs) were screened for their ability to augment the expression of heterologous genes in Chinese hamster ovary (CHO) cells. Of the chromatin elements assayed, the chicken lysozyme matrix-attachment region (MAR) was the only element to significantly increase stable reporter expression. We found that the use of the MAR increases the proportion of high-producing clones, thus reducing the number of clones that need to be screened. These benefits are observed both for constructs with MARs flanking the transgene expression cassette, as well as when constructs are co-transfected with the MAR on a separate plasmid. Moreover, the MAR was co-transfected with a multicomponent regulatable beta-galactosidase expression system in C2C12 cells and several clones exhibiting regulated expression were identified. Hence, MARs are useful in the development of stable cell lines for production or regulated expression.

Another way of thinking about ADHD: the predictive role of early attachment deprivation in adolescents' level of symptoms.

PURPOSE: Attention deficit and hyperactivity disorder (ADHD) is one of the most frequent disorders in childhood and adolescence. Both neurocognitive and environmental factors have been related to ADHD. The current study contributes to the documentation of the predictive relation between early attachment deprivation and ADHD. METHOD: Data were collected from 641 adopted adolescents (53.2 % girls) aged 11-16 years in five countries, using the DSM oriented scale for ADHD of the Child Behavior Checklist (CBCL) (Achenbach and Rescorla, Manual for the ASEBA school-age forms and profiles. University of Vermont, Research Center for Children, Youth and Families, Burlington, 2001). The influence of attachment deprivation on ADHD symptoms was initially tested taking into consideration several key variables that have been reported as influencing ADHD at the adoptee level (age, gender, length of time in the adoptive family, parents' educational level and marital status), and at the level of the country of origin and country of adoption (poverty, quality of health services and values). The analyses were computed using the multilevel modeling technique. RESULTS: The results showed that an increase in the level of ADHD symptoms was predicted by the duration of exposure to early attachment deprivation, estimated from the age of adoption, after controlling for the influence of adoptee and country variables. The effect of the age of adoption was also demonstrated to be specific to the level of ADHD symptoms in comparison to both the externalizing and internalizing behavior scales of the CBCL. CONCLUSION: Deprivation of stable and sensitive care in infancy may have long-lasting consequences for children's development.

The Effect of Sexual Abuse on Children's Attachment Representations in Chile.

Child sexual abuse is associated with problems in children's emotional development, particularly increased insecurity of attachment. However, few studies have examined its effect on the organization of attachment representations in preschoolers, and the findings of those that have been conducted have not been entirely consistent. Therefore, this study aims to analyze the effect of child sexual abuse on attachment representation quality in a sample of children 3 to 7 years old in Chile. The results indicate child sexual abuse does affect children's attachment representation quality. The attachment narratives of child sexual abuse victims scored significantly higher than nonvictims on the hyperactivity and disorganization dimensions of attachment. These results are discussed in terms of attachment theory, clinical findings on child sexual abuse, and clinical implications.

Attachment of Combination Rails to Concrete Parapets Utilizing Epoxy Adhesive Anchors, RPFP-IOWA-1, 2015

The Iowa Department of Transportation (IaDOT) was interested in investigating the use of epoxy adhesive anchorages for the attachment of posts used in the BR27C combination bridge rail system. Alternative anchorage concepts were developed using a modified version of the ACI 318-11 procedures for embedded anchor design. Four design concepts were developed for review by IaDOT, including: (1) a four-bolt square anchorage, (2) a four-bolt spread anchorage, (3) a twobolt centered anchorage, and (4) a two-bolt offset anchorage. IaDOT representatives selected the four-bolt spread anchorage and the two-bolt offset anchorage as the preferred designs for evaluation. In addition to these two proposed configurations, IaDOT also requested that the researchers evaluate a third option that had been previously installed on the US-20 bridge near Hardin, IA. The proposed alternative anchorages and the original cast-in-place anchorage for the BR27C combination bridge rail were evaluated through dynamic component testing. The test of the original cast-in-place anchorage was used a baseline for comparison with the alternative designs. Test no. IBP-1 of the original cast-in-place anchorage developed a peak load of 22.9 kips (101.9 kN) at a deflection of 1.5 in. (38 mm). All three of the tested alternative anchorages provided greater load capacity than the original cast-in-place design and were deemed acceptable surrogates. Of the three alternative designs, the two-bolt offset design was deemed the best option.

Models of social networks based on social distance attachment

We propose a class of models of social network formation based on a mathematical abstraction of the concept of social distance. Social distance attachment is represented by the tendency of peers to establish acquaintances via a decreasing function of the relative distance in a representative social space. We derive analytical results (corroborated by extensive numerical simulations), showing that the model reproduces the main statistical characteristics of real social networks: large clustering coefficient, positive degree correlations, and the emergence of a hierarchy of communities. The model is confronted with the social network formed by people that shares confidential information using the Pretty Good Privacy (PGP) encryption algorithm, the so-called web of trust of PGP.

Molecular characterization of a human matrix attachment region epigenetic regulator.

Matrix attachment regions (MAR) generally act as epigenetic regulatory sequences that increase gene expression, and they were proposed to partition chromosomes into loop-forming domains. However, their molecular mode of action remains poorly understood. Here, we assessed the possible contribution of the AT-rich core and adjacent transcription factor binding motifs to the transcription augmenting and anti-silencing effects of human MAR 1-68. Either flanking sequences together with the AT-rich core were required to obtain the full MAR effects. Shortened MAR derivatives retaining full MAR activity were constructed from combinations of the AT-rich sequence and multimerized transcription factor binding motifs, implying that both transcription factors and the AT-rich microsatellite sequence are required to mediate the MAR effect. Genomic analysis indicated that MAR AT-rich cores may be depleted of histones and enriched in RNA polymerase II, providing a molecular interpretation of their chromatin domain insulator and transcriptional augmentation activities.

Genome-wide prediction of matrix attachment regions that increase gene expression in mammalian cells.

Gene transfer in eukaryotic cells and organisms suffers from epigenetic effects that result in low or unstable transgene expression and high clonal variability. Use of epigenetic regulators such as matrix attachment regions (MARs) is a promising approach to alleviate such unwanted effects. Dissection of a known MAR allowed the identification of sequence motifs that mediate elevated transgene expression. Bioinformatics analysis implied that these motifs adopt a curved DNA structure that positions nucleosomes and binds specific transcription factors. From these observations, we computed putative MARs from the human genome. Cloning of several predicted MARs indicated that they are much more potent than the previously known element, boosting the expression of recombinant proteins from cultured cells as well as mediating high and sustained expression in mice. Thus we computationally identified potent epigenetic regulators, opening new strategies toward high and stable transgene expression for research, therapeutic production or gene-based therapies.

Attachment, psychopathology, and religion: Introduction to this special section of Mental Health, Religion, & Culture

An international conference of psychology of religion, organised at the University of Lausanne (Switzerland) on 16 May 2012, took up the theme: "Attachment, psychopathology, and religion". Four speakers were invited: Pehr Granvist, Andrew Gumley, Isabelle Rieben, and Pascal Roman. Their reworked contributions are gathered in this special section of Mental Health, Religion, & Culture. The goal of this special section is to re-examine the whole of this subject of the bond between attachment and religion and/or spirituality in the cases of those persons suffering from mental health disorders.

Attachment and spiritual coping in patients with chronic schizophrenia

Many studies on spirituality in psychosis have shown that, compared to a nonclinical population, patients make more use of spiritual beliefs/religious practices to deal with their problems. Our research question was to test whether attachment to spiritual figures could be a good explanation for religious coping strategies in patients with psychosis. First, adult attachment was investigated in 28 patients with chronic psychosis and 18 controls, using the Adult Attachment Interview. Diagnostic evaluations were performed with the Structured Clinical Interview for the Diagnostic and Statistical Manual of Mental Disorders (4th edition, Text Revision) Axis I disorders and symptomatic evaluation with the Brief Psychiatric Rating Scale. Results also show a high prevalence of insecure avoidant attachment in patients, and suggest that a significant part of religious coping might be explained by the theory of attachment (64% of the patients, 78% of controls). The implications of these results are interpreted in light of correspondence and compensation hypotheses.

Anxiety, depression, and attachment before and after the first-trimester screening for Down syndrome: comparing couples who undergo ART with those who conceive spontaneously.

OBJECTIVES: This study's aim was to describe the emotional status of parents to be before and after the first-trimester combined prenatal screening test. METHODS: One hundred three couples participated, of which 52 had undergone an in vitro fertilization/intracytoplasmic sperm injection treatment [assisted reproductive technology (ART)] and 51 had conceived spontaneously. Participants completed the state scale of the State-trait Anxiety Inventory, the Edinburgh Depression Scale, and the Maternal and Paternal Antenatal Attachment Questionnaire before the first-trimester combined prenatal screening test at around 12 weeks of gestational age (T1) and just after receiving the results at approximately 14 weeks of gestational age (T2). RESULTS: We observed a significant decrease in anxiety and depression symptoms and a significant increase in attachment from T1 to T2. Results showed no differences between groups at either time point, which suggests that ART parents are more similar to than different from parents conceiving spontaneously. Furthermore, given the importance of anxiety during pregnancy, a subsample of women with clinical anxiety was identified. They had significantly higher rates of clinical depression and lower attachment. CONCLUSIONS: These results indicate that, regardless of whether conception was through ART or spontaneous, clinical anxiety in women over the prenatal testing period is associated with more vulnerability during pregnancy (i.e. clinical depression and less attachment to fetus). © 2015 John Wiley & Sons, Ltd.

Attachment and coping in psychosis in relation to spiritual figures

Background: Studies have found higher levels of insecure attachment in individuals with schizophrenia. Attachment theory provides a framework necessary for conceptualizing the development of interpersonal functioning. Some aspects of the attachment of the believer to his/her spiritual figure are similar to those between the child and his/her parents. The correspondence hypothesis suggests that early child-parent interactions correspond to a person's relation to a spiritual figure. The compensation hypothesis suggests that an insecure attachment history would lead to a strong religiousness/spirituality as a compensation for the lack of felt security. The aim of this study is to explore attachment models in psychosis vs. healthy controls, the relationships between attachment and psychopathology and the attachment processes related to spiritual figures. Methods: Attachment models were measured in 30 patients with psychosis and 18 controls with the AAI (Adult Attachment interview) in relationship with psychopathology. Beliefs and practices related to a spiritual figure were investigated by qualitative and quantitative analyses. Results: Patients with psychosis showed a high prevalence of insecure avoidant attachment. Spiritual entities functioned like attachment figures in two thirds of cases. Interviews revealed the transformation of internal working models within relation to a spiritual figure: a compensation process was found in 7 of the 32 subjects who showed a significant attachment to a spiritual figure. Conclusions: Attachment theory allows us to highlight one of the underlying dimensions of spiritual coping in patients with psychosis.

Hypothyroidism decreases proinsulin gene expression and the attachment of its mRNA and eEF1A protein to the actin cytoskeleton of INS-1E cells

The actions of thyroid hormone (TH) on pancreatic beta cells have not been thoroughly explored, with current knowledge being limited to the modulation of insulin secretion in response to glucose, and beta cell viability by regulation of pro-mitotic and pro-apoptotic factors. Therefore, the effects of TH on proinsulin gene expression are not known. This led us to measure: a) proinsulin mRNA expression, b) proinsulin transcripts and eEF1A protein binding to the actin cytoskeleton, c) actin cytoskeleton arrangement, and d) proinsulin mRNA poly(A) tail length modulation in INS-1E cells cultured in different media containing: i) normal fetal bovine serum - FBS (control); ii) normal FBS plus 1 µM or 10 nM T3, for 12 h, and iii) FBS depleted of TH for 24 h (Tx). A decrease in proinsulin mRNA content and attachment to the cytoskeleton were observed in hypothyroid (Tx) beta cells. The amount of eEF1A protein anchored to the cytoskeleton was also reduced in hypothyroidism, and it is worth mentioning that eEF1A is essential to attach transcripts to the cytoskeleton, which might modulate their stability and rate of translation. Proinsulin poly(A) tail length and cytoskeleton arrangement remained unchanged in hypothyroidism. T3 treatment of control cells for 12 h did not induce any changes in the parameters studied. The data indicate that TH is important for proinsulin mRNA expression and translation, since its total amount and attachment to the cytoskeleton are decreased in hypothyroid beta cells, providing evidence that effects of TH on carbohydrate metabolism also include the control of proinsulin gene expression.