Parent Stress and Observed Parenting in a Parent-Child Interaction Task in a Predominantly Minority and Low-Income Sample.

High stress in parents may affect parenting and subsequent child socioemotional and behavioral development. Previous evidence suggests that highly stressed parents are more likely to engage in negative parenting, which is less structured and more punitive. However, the effects of life stress versus parent specific stress on parent-child interactions in early childhood has not been well studied, especially in minority and low-income samples. Thus, the current study assessed the relationship between perceived life stress, parenting-related stress, and observed parenting responses to young children during a structured, mildly challenging parent-child task. Predominantly minority and low-income parents and their children (2-5 years old; 54 dyads) completed the Perceived Stress Scale, the Parenting Stress Inventory, and participated in a structured 5-minute interaction task, the Toy-Wait Task (TWT), that was video-taped and coded by blind raters. The coding utilized a standardized system with good reliability assessing 1) Affect (parent and child positive and negative affect, shared positive affect), 2) Positive Parenting Behaviors (warmth, structured good involvement, listening/engagement), and 3) Negative Parenting Behaviors (reactivity, judgment, critical parenting). Significant associations were found between perceived life stress and parenting stress, (r (54) = 0.61, p<.01). Parents with higher perceived life stress scores showed more negative affect (r=0.291, p<.05) and lower involvement with the child (r=-0.367, p<.05), while parenting specific stress did not yield significant effects (p's > 0.05). Findings suggest that interventions that reduce stress in minority and low-income parents of young children may also improve parenting of young children with potential impact on decreasing child psychopathology risk.

High-resolution genomic profiling of human papillomavirus-associated vulval neoplasia.

BACKGROUND: The incidence of human papillomavirus-associated vulval neoplasia is increasing worldwide; yet the associated genetic changes remain poorly understood. METHODS: We have used single-nucleotide polymorphism microarray analysis to perform the first high-resolution investigation of genome-wide allelic imbalance in vulval neoplasia. Our sample series comprised 21 high-grade vulval intraepithelial neoplasia and 6 vulval squamous cell carcinomas, with paired non-lesional samples used to adjust for normal copy number variation. RESULTS: Overall the most common recurrent aberrations were gains at 1p and 20, with the most frequent deletions observed at 2q, 3p and 10. Copy-neutral loss of heterozygosity at 6p was a recurrent event in vulval intraepithelial neoplasia. The pattern of genetic alterations differed from the characteristic changes we previously identified in cutaneous squamous cell carcinomas. Vulval neoplasia samples did not exhibit gain at 5p, a frequent recurrent aberration in a series of cervical tumours analysed elsewhere using an identical protocol. CONCLUSION: This series of 27 vulval samples comprises the largest systematic genome-wide analysis of vulval neoplasia performed to date. Despite shared papillomavirus status and regional proximity, our data suggest that the frequency of certain genetic alterations may differ in vulval and cervical tumours.

MicroRNA miR-181a correlates with morphological sub-class of acute myeloid leukaemia and the expression of its target genes in global genome-wide analysis.

MicroRNAs (miRNAs) are short single-stranded RNAs that have a potentially important role in gene regulation. Using a quantitative real-time polymerase chain reaction assay specific to the mature miRNA, the expression level of a selected group of haematopoietic tissue-specific miRNAs was measured across a set of 30 primary adult acute myeloid leukaemia (AML) with a normal karyotype. The expression levels of each miRNA were correlated with the genome-wide mRNA expression profiles in the same leukaemias. This revealed that miR-181a correlated strongly with the AML morphological sub-type and with the expression of genes previously identified through sequence analysis as potential interaction targets. Three other miRNAs, miR-10a, miR-10b and miR-196a-1, showed a clear correlation with HOX gene expression.

Genome-wide detection of recurring sites of uniparental disomy in follicular and transformed follicular lymphoma.

Single-nucleotide polymorphism (SNP) array analysis was performed using the 10K GeneChip array on a series of 26 paired follicular lymphoma (FL) and transformed-FL (t-FL) biopsies and the lymphoma cell lines SCI-1, DoHH2 and RL2261. Regions of acquired homozygosity were detected in 43/52 (83%) primary specimens with a mean of 1.7 and 3.0 aberrations in the FL and t-FL, respectively. A notable feature was the occurrence of recurring sites of acquired uniparental disomy (aUDP) on 6p, 9p, 12q and 17p in cell lines and primary samples. Homozygosity of 9p and 17p arose predominantly in t-FL and in three cases rendered the cell homozygous for a pre-existing mutation of either CDKN2A or TP53. These data suggest that mutation precedes mitotic recombination, which leads to the removal of the remaining wild-type allele. In all, 18 cases exhibited abnormalities in both FL and t-FL samples. In 10 cases blocks of homozygosity were detected in FL that were absent in the subsequent t-FL sample. These differences support the notion that FL and t-FL may arise in a proportion of patients by divergence from a common malignant ancestor cell rather than by clonal evolution from an antecedent FL.

Fingerprinting genomic instability in oral submucous fibrosis.

BACKGROUND: Oral submucous fibrosis (OSF) is a high-risk pre-cancerous condition where 7-13% of these patients develop head and neck squamous cell carcinoma (HNSCC). To date there is no cancer predictive markers for OSF patients. Genomic instability hallmarks early genetic events during malignant transformation causing loss of heterozygosity (LOH) and chromosomal copy number abnormality. However, to date there is no study on genomic instability in OSF. Although this condition is known as a high-risk pre-cancerous condition, there is no data regarding the genomic status of this disease in terms of genetic susceptibility to malignant transformation. METHODS: In this study, we investigated the existence of genetic signatures for carcinogenesis in OSF. We employed the high-resolution genome-wide Affymetrix Mapping single nucleotide polymorphism microarray technique to 'fingerprint' global genomic instability in the form of LOH in 15 patient-matched OSF-blood genomic DNA samples. RESULTS: This rapid high-resolution mapping technique has revealed for the first time that a small number of discrete hot-spot LOH loci appeared in 47-53% of the OSF tissues studied. Many of these LOH loci were previously identified regions of genomic instability associated with carcinogenesis of the HNSCC. CONCLUSION: To our knowledge, this is the first evidence that genomic instability in the form of LOH is present in OSF. We hypothesize that the genomic instability detected in OSF may play an important role in malignant transformation. Further functional association studies on these putative genes may reveal potential predictive oral cancer markers for OSF patients.

Genomic disruption of the histone methyltransferase SETD2 in chronic lymphocytic leukaemia.

Histone methyltransferases (HMTs) are important epigenetic regulators of gene transcription and are disrupted at the genomic level in a spectrum of human tumours including haematological malignancies. Using high-resolution single nucleotide polymorphism (SNP) arrays, we identified recurrent deletions of the SETD2 locus in 3% (8/261) of chronic lymphocytic leukaemia (CLL) patients. Further validation in two independent cohorts showed that SETD2 deletions were associated with loss of TP53, genomic complexity and chromothripsis. With next-generation sequencing we detected mutations of SETD2 in an additional 3.8% of patients (23/602). In most cases, SETD2 deletions or mutations were often observed as a clonal event and always as a mono-allelic lesion, leading to reduced mRNA expression in SETD2-disrupted cases. Patients with SETD2 abnormalities and wild-type TP53 and ATM from five clinical trials employing chemotherapy or chemo-immunotherapy had reduced progression-free and overall survival compared with cases wild type for all three genes. Consistent with its postulated role as a tumour suppressor, our data highlight SETD2 aberration as a recurrent, early loss-of-function event in CLL pathobiology linked to aggressive disease.

Normal c-abl gene protein--a nuclear component.

The subcellular distribution of the c-abl and bcr-abl gene products from KG1A and K562 cells has been studied by two different techniques. Firstly, physical disruption followed by subcellular fractionation was used to demonstrate that normal c-abl (p145) was recovered from the cytosol and the nuclear fractions of KG1A cells. In contrast, bcr-abl products were recovered exclusively from the cytosol fraction of K562 cells. Secondly, indirect immunofluorescence was used to localize c-abl protein to the cytoplasm, nuclear membrane and infrequently to the nucleus of KG1A cells and bcr-abl protein to only the cytoplasm of K562 cells. Thus both the approaches indicate that there is a component of normal c-abl products which appears to be nuclear and this is not reflected in the distribution of the bcr-abl 210 kDa protein, which remains cytosolic.

The cloning, mapping and expression of a novel gene, BRL, related to the AF10 leukaemia gene.

The MLL gene is reciprocally translocated with one of a number of different partner genes in a proportion of human acute leukaemias. The precise mechanism of oncogenic transformation is unclear since most of the partner genes encode unrelated proteins. However, two partner genes, AF10 and AF17 are related through the presence of a cysteine rich region and a leucine zipper. The identification of other proteins with these structures will aid our understanding of their role in normal and leukaemic cells. We report the cloning of a novel human gene (BRL) which encodes a protein containing a cysteine rich region related to that of AF10 and AF17 and is overall most closely related to the previously known protein BR140. BRL maps to chromosome 22q13 and shows high levels of expression in testis and several cell lines. The deduced protein sequence also contains a bromodomain, four potential LXXLL motifs and four predicted nuclear localization signals. A monoclonal antibody raised to a BRL peptide sequence confirmed its widespread expression as a 120 Kd protein and demonstrated localization to the nucleus within spermatocytes.

Expression pattern and cellular distribution of the murine homologue of AF10.

We have cloned Af10, the murine homologue of the MLL partner gene AF10. The predicted open reading frame of Af10 contains 1069 aa which are 90% identical to those of AF10. Af10 contains an N-terminal cysteine-rich region with a LAP/PHD finger, a leucine zipper domain and a glutamine-rich region at the C-terminus, features also found in the human proteins AF10 and AF17. A single 5. 5-kb transcript was detected in murine tissues with the highest level of expression in the testes. A polyclonal antibody raised to the cysteine-rich region of AF10 was able to identify a double band of 140 kDa on Western analysis in mouse testicular extracts. After subcellular separation Af10 was identified in both the nuclear and cytoplasmic extracts, again as a double band of 140 kDa in size. In situ hybridisation studies were performed with sense and antisense digoxigenin-labelled oligonucleotides. High levels of expression were noted in postmeiotic germ cells, especially in spermatids from around stage VI to stage VIII. High levels of expression were also seen in the white matter of the cerebellum, extending into the granular layer. The expression in differentiated rather than in proliferating cells suggests that the role of Af10 may lie in the suppression of proliferation rather than in differentiation. Since the LAP/PHD finger domains are lost in the MLL-AF10 fusion, arguably such a function could be carried out by this domain.

BCR-ABL and BCR proteins: biochemical characterization and localization.

The Philadelphia translocation results in the expression of a family of chimaeric proteins in which a portion of the bcr protein is fused to c-abl protein. Using antibodies which recognize different portions of the bcr gene and abl gene products we have compared the normal bcr products with their chimaeric counterparts. We first conclude that the enhanced kinase activity of the rearranged bcr-abl products (p210 and p190) is recovered almost exclusively from the cytosolic fraction. This methodology was confirmed by the demonstration that in cells transformed by the Abelson murine leukemia virus (A-MuLV) the gag-abl kinase activity was recovered equally from the membrane and cytosolic fractions, in agreement with previous studies. To determine whether the distribution of kinase activity reflected the bulk distribution of the bcr-abl proteins, in vivo labeling followed by subcellular fractionation was performed. Both normal bcr proteins and the p210 bcr-abl protein were recovered from the cytosolic fraction with little detectable amounts present in other fractions. In vivo labeling was also used to demonstrate that both normal bcr products and the p210 bcr-abl had a relatively long half-life. It is concluded that bcr-abl products, like normal bcr products are located in the cytosolic fraction.

Establishment of a lymphoblastoid cell line, SD-1, expressing the p190 bcr-abl chimaeric protein.

A lymphoblastoid cell line (SD-1) has been established by Epstein-Barr virus immortalisation of Philadelphia chromosome positive acute lymphoblastic leukaemia (ALL) cells. Using newly derived anti-bcr monoclonal and anti-abl polyclonal antibodies it was demonstrated that both the original leukaemic cells and the derived cell line expressed the p190 form of the bcr-abl protein found in a proportion of cases of Philadelphia chromosome positive ALL. Interestingly, the leukaemia and the derived cell line each displayed different, clonal patterns of immunoglobulin gene rearrangements providing direct evidence that the t(9;22) translocation which results in the expression of the p190 bcr-abl protein must occur before immunoglobulin heavy chain gene rearrangement. In contrast to the leukaemia, which had multiple chromosome abnormalities in addition to the t(9;22), the cell line had the t(9;22) translocation as its sole abnormality. Although SD-1 cells were demonstrated to express continuously the p190 bcr-abl protein, they were unable to form colonies in soft agar and did not cause tumours in splenectomised nude mice. This cell line therefore represents an appropriate target cell line in which to examine the cooperativity of the p190 bcr-abl protein with other activated oncogene products.

Detection and significance of bcr-abl mRNA transcripts and fusion proteins in Philadelphia-positive adult acute lymphoblastic leukemia.

Blast cells from an unselected consecutive series of 84 adults presenting with acute lymphoblastic leukemia (ALL) to St Bartholomew's Hospital over a seven year period were tested prospectively by cytogenetic and retrospectively by RT-PCR analysis for the presence of the Ph translocation and bcr-abl mRNA. This combination gave an overall figure of 20.3% for bcr-abl-positive and/or Ph-positive ALL. The incidence of bcr-abl-positive/Ph-positive ALL was most common between the ages of 31 and 50 years, becoming less common after the age of 50. Eight out of ten bcr-abl-positive patients expressed the e1a2 mRNA transcript, the other two expressed the b3a2 and b2a3 transcripts respectively. Cells from all patients with bcr-abl mRNA transcripts expressed the appropriate p190 or p210 bcr-abl protein and all were Ph-positive.

Identification and molecular characterisation of a CALM-AF10 fusion in acute megakaryoblastic leukaemia.

The t(10;11)(p13;q14-21) is a non-random translocation described in acute lymphoblastic and myeloid leukaemias. It results in the fusion of the gene CALM, which encodes a clathrin assembly protein, on 11q14 to the gene AF10, a putative transcription factor on 10p13. Here we describe for the first time, the occurrence of a CALM-AF10 fusion in a case of acute megakaryoblastic leukaemia. Fluorescence in situ hybridisation and reverse transcriptase polymerase chain reaction were used to confirm the presence of a CALM-AF10 fusion. A novel splice variant of CALM missing nt 1927-2091 was also detected. Though CALM is a cytoplasmic protein, the chimaeric fusion product is able to localise to both the nucleus and cytoplasm. Analysis of the fusion variants suggests, however, that the critical fusion product is likely to be cytoplasmic and contain the interactive leucine zipper of AF10.

Therapy-related adult acute lymphoblastic leukemia with t(4;11)(q21; q23): MLL rearrangement, p53 mutation and multilineage involvement.

A diagnosis of pro-B acute lymphoblastic leukemia (ALL) with CD15+ was made in a 42-year-old woman, 12 months after the treatment of uterine adenocarcinoma by carboplatinum, anthracyclines, etoposide and radiotherapy. Molecular cytogenetic studies revealed a karyotype with multiple chromosome changes, including the t(4;11)(q21;q23) and a 17p-chromosome, with MLL disruption and 17p13/p53 gene deletion in 86% of the cells. A p53 exon 6 mutation was documented, resulting in p53 protein stabilization, with 20% of the cells reacting with the 1801 anti-p53 monoclonal antibody. Dual-color FISH using MLL and p53 probes was performed on peripheral blood smears, providing direct evidence of the involvement of the blast cells and of the granulocytic lineage. Only a partial, shortlasting response was obtained by induction treatment, confirming that a poor prognosis is associated with therapy-related ALL with the 4;11 translocation.

Representation of central and peripheral vision in the primate cerebral cortex: Insights from studies of the marmoset brain.

How the visual field is represented by neurons in the cerebral cortex is one of the most basic questions in visual neuroscience. However, research to date has focused heavily on the small part of the visual field within, and immediately surrounding the fovea. Studies on the cortical representation of the full visual field in the primate brain are still scarce. We have been investigating this issue with electrophysiological and anatomical methods, taking advantage of the small and lissencephalic marmoset brain, which allows easy access to the representation of the full visual field in many cortical areas. This review summarizes our main findings to date, and relates the results to a broader question: is the peripheral visual field processed in a similar manner to the central visual field, but with lower spatial acuity? Given the organization of the visual cortex, the issue can be addressed by asking: (1) Is visual information processed in the same way within a single cortical area? and (2) Are different cortical areas specialized for different parts of the visual field? The electrophysiological data from the primary visual cortex indicate that many aspects of spatiotemporal computation are remarkably similar across the visual field, although subtle variations are detectable. Our anatomical and electrophysiological studies of the extrastriate cortex, on the other hand, suggest that visual processing in the far peripheral visual field is likely to involve a distinct network of specialized cortical areas, located in the depths of the calcarine sulcus and interhemispheric fissure.

Histone acetylation-mediated regulation of genes in leukaemic cells.

Histone deacetylase (HDAC) and histone acetyltransferase (HAT) functions are associated with various cancers, and the inhibition of HDAC has been found to arrest disease progression. Here, we have investigated the gene expression profiles of leukaemic cells in response to the HDAC inhibitor trichostatin A (TSA) using oligonucleotide microarrays. Nucleosomal histone acetylation was monitored in parallel and the expression profiles of selected genes were confirmed by quantitative polymerase chain reaction (PCR). A large number of genes (9% of the genome) were found to be similarly regulated in CCRF-CEM and HL-60 cells in response to TSA, and genes showing primary and secondary responses could be distinguished by temporal analysis of gene expression. A small fraction of genes were highly sensitive to histone hyper-acetylation, including XRCC1, HOXB6, CDK10, MYC, MYB, NMI and CBFA2T3 and many were trans-acting factors relevant to cancer. The most rapidly repressed gene was MKRN3, an imprinted gene involved in the Prader-Willi syndrome.

Preventing faking in phallometric assessments of sexual preference.

This study evaluated a method of preventing sexual preference faking in phallometric assessments employing audiotaped stimuli. The stimuli were stories describing neutral heterosocial interactions, consenting heterosexual activity, rape, and nonsexual violence. Sixteen normal heterosexual males were each tested with ordinary instructions, with fake instructions (i.e., to appear sexually interested in rape and nonsexual violence but not in consenting sex), and with fake instructions while performing a secondary semantic tracking task. The tracking task was to press one button whenever sexual activity was being described and another button whenever violence occurred. This simple task was designed to focus subjects' attention on only the critical elements of the stories. Group data indicated that subjects could fake inappropriate preferences when instructed to do so without the semantic tracking task but could not when the task was required. The implications of these findings for ethical practice and for the theoretical interpretation of phallometric assessment data were discussed.

Inhibition of P210BCR/ABL Expression in K562 Cells by Electroporation with an Antisense Oligonucleotide.

We designed experiments to study the effects on P210BCR/ABL expression of introducing antisense oligonucleotides into K562 cells. We used two antisense oligonucleotides: one (AS1) is complementary to the first coding codon of the BCR/ABL mRNA and the two 5' and three 3' codons, and the other (AS2) to BCR coding codons 5 to 11 inclusive. To facilitate entry of the oligonucleotides the K562 cells were subjected to electroporation on three occasions at 24 hr intervals (0, 24 and 48 hr). P210BCR/ABL expression was assayed by in vivo phosphorylation followed by immune precipitation with a BCR antibody. Introduction of AS1 inhibited P210BCR/ABL expression at 72 and 96 hrs, whereas AS2 and the control oligonucleotide had no effect. AS1 also killed K562 cells. We conclude that selected antisense oligonucleotides can modify leukaemia-specific protein expression in K562 cells. This approach could prove valuable for purging CML bone marrow cells in vitro.

Salient victim suffering and the sexual responses of child molesters.

In phallometric testing, audiovisual stimuli in which the audio component depicted a child victim's suffering described from the child's point of view were compared with similar scenarios described from the adult male offender's point of view and to portrayals of consenting adult heterosexual activity. Fifteen male participants who had sexually assaulted girls and 15 nonoffenders were also administered questionnaire measures of empathy and opinions regarding sex with children. Consistent with predictions, maximal discrimination between groups was obtained using deviance indices based on stimuli that emphasized victim trauma. Also consistent with predictions, the questionnaire measures were related to sexual deviance. Recommendations are made regarding stimuli for phallometric testing. The results point to the relevance of changing deviant preferences and enhancing empathy for the victim in the treatment of child molesters.

Viewing time as a measure of sexual interest among child molesters and normal heterosexual men.

Although phallometric assessment is the best scientific method for measuring male sexual interest, it is intrusive and highly technical. We examined viewing time as an unobtrusive and technically simple measure of sexual preference and compared the discrimination obtained by viewing time measures with that obtained by phallometric measures. Slides of nude males and females of various ages were shown to child molesters and normal men while their viewing times were recorded. Subjects then rated the sexual attractiveness of the stimulus persons. Phallometric assessments using the same stimulus categories were also given to some of the Ss. Deviance scores calculated from the viewing time data significantly discriminated between the child molesters and the normals, although the discrimination achieved was less than that obtained using phallometric measures. Sexual attractiveness ratings did not differentiate the two groups. Among the normal men, viewing time and sexual attractiveness ratings were highly correlated; but the correlation was much lower for child molesters. Viewing time shows considerable promise as an unobtrusive measure of male sexual interest.