Multiplex PCR analysis of in vivo-arising deletion mutations in the hprt gene of human T-lymphocytes

A multiplex polymerase chain reaction (PCR) procedure was adapted for the rapid and efficient evaluation of deletions of the hypoxanthine guanine phosphoribosyltransferase (hprt) gene in human T-lymphocytes. The hprt clonal assay was used to isolate in vivo-arising hprt-deficient T-cells from six healthy males. Mutant frequencies ranged from 9-27 × 10^?6. Simple crude cellular extracts from 223 mutants were analyzed for hprt gene deletion. Sixteen (7.2%) were found to be due to total gene deletion and 22 (9.9%) were due to partial gene deletion. The relatively high frequency of total gene deletions was caused by replicate isolates of a single mutational event as shown by single-strand conformation polymorphism (SSCP) analysis of rearranged T-cell receptor (TCR)-γ genes. Eighteen of the 22 partial hprt gene deletion mutants were determined to be of independent origin based on a unique hprt mutation or SSCP-TCR-γ pattern. One-half (9/18) of the partial deletion mutants involved all or part of exon 4 alone, suggesting that this region of the hprt gene is prone to deletion. The small deletions effecting exon 1 (1 mutant), exon 2 (2 mutants), and exon 4 (6 mutants) would not have been detected by conventional Southern blot analysis and may represent a new, previously unrecognized class of mutations. The ready isolation of such intragenic deletions will allow the characterization of breakpoint junctions and may provide insights into the important processes of DNA breakage and rejoining. © 1994 Wiley-Liss, Inc.     

Challenging the Presumption of Diminished Capacity to Parent: Does Divorce Really Change Parenting Practices?*

Abstract: The purpose of this paper is to determine whether divorced parents exhibit a diminished capacity to parent in the period following divorce. Using 2 waves of data from a national survey of Canadian children, the current study prospectively follows 5,004 children living in 2–biological parent households at initial interview and compares changes in parenting practices between households that subsequently divorce and those that remain intact. Results show that divorce is unrelated to changes in parenting behavior, suggesting that there are more similarities than differences in parenting among recently divorced and continuously married parents.     

CRITERIA ADOPTED BY DENTISTS TO INDICATE THE EXTRACTION OF PERIODONTALLY INVOLVED TEETH

When dealing with patients with periodontal disease of variable severities, dentists must often choose between treating and restoring the involved tooth or indicating its extraction. Different criteria have been adopted in this decision-making process. The purpose of this study was to evaluate the criteria adopted by dentists to indicate the extraction of teeth with periodontitis. Dentists were interviewed at their private practices in three cities of the state of Rio Grande do Sul, Brazil. The evaluated criteria included severity of attachment loss, tooth mobility, furcation involvement, prosthetic planning, periodontal-endodontic lesion, possible systemic involvement due to the presence of periodontitis, referral to a periodontist for evaluation, radiographic bone loss greater than 50%, presence of extensive caries, socio-economic and cultural status of the patient, among others. The most often adopted criteria to indicate the extraction of periodontally affected teeth were the presence of mobility (37.5%), severity of attachment loss (24.3%) and radiographic bone loss greater than 50% (21.2%). The results of the present study demonstrated the difficulties faced by dentists to indicate the extraction of teeth with severe attachment loss, in addition to the establishment of an adequate prognosis. Aspects associated with the past disease were still the most often reported to indicate the extraction of teeth for periodontal reasons.     

Cloning and expression analysis of the chick DAN gene, an antagonist of the BMP family of growth factors.

Differential screening-selected gene aberrative in neuroblastoma (DAN) is a member of a cystine knot protein family that includes Cerberus and Gremlin. First isolated in a screen to identify genes down-regulated in transformed rat fibroblasts, DAN has subsequently been cloned in Xenopus, mouse, and human. Overexpression of DAN suppresses the transformed phenotype and retards the cell's entry into S phase. Biochemical analyses have demonstrated DAN's ability to bind bone morphogenetic proteins and antagonize their signaling activity. In this study, chick DAN was cloned and sequenced, revealing a conserved cystine knot region as well as an N-glycosylation site. A riboprobe was designed from the 3' chick DAN coding sequence and used for analysis of DAN in the developing chick embryo by in situ hybridization. Chick DAN was expressed beginning at stage 10 in the developing somites and the medial otic epithelium. Expression in the neural layer of the eye became apparent at stage 14. By stage 17, expression had expanded to the base of the hindbrain. Limb bud labeling began at stage 20, whereas expression in the branchial arches appeared at stage 25. Chick DAN expression generally corresponded to that of mouse DAN expression as shown by comparative in situ hybridization. However, chick DAN was found in the otic epithelium and notochord, whereas mouse DAN was restricted to the overlying otic ectomesenchyme and was absent from the notochord. This observation suggests that DAN may play different roles in chick and mouse otic and notochord development.     

The Mental Health of Homeless Children

The literature addressing the mental health of homeless children is critically reviewed. Higher rates of behavioral, emotional, and cognitive problems have been reported for this population, but research results are qualified by significant methodological problems. Minimal attention has been given to the development and evaluation of specific intervention and prevention strategies. Recommendations are made for conducting research that will overcome previous methodological problems and focus more directly on identifying risk and protective mechanisms within the population of children exposed to homelessness.     

In vivo vs in vitro anticariogenic behavior of glass-ionomer and resin composite restorative materials.

OBJECTIVE: To evaluate the in vivo vs the in vitro anticariogenic potential of glass-ionomer and resin composite restoratives, utilizing a standardized interfacial gap model. METHODS: (a) In vitro study. Box shaped cavities were prepared at the buccal surfaces of extracted premolars limited to enamel. The incisal cavity walls received no treatment and were covered with 40 microm-thick metal spacers. The cavities were restored with a glass-ionomer (Ketac-Fil, n=8) and a fluoride-free resin composite (Scotchbond MP Plus/Z100, n=8). After 4 weeks immersion in an acidic gel (pH 4), thin sections were produced and examined under polarized-light microscopy.(b) In vivo study. Four low caries activity volunteers, with first four premolars, each planned to be extracted for orthodontic reasons, participated in the study. Cavities were prepared as before and filled contralaterally per patient with glass-ionomer (n=8) and resin composite (n=8). After 6 months in vivo, the teeth were extracted, sectioned and investigated by polarized-light microscopy, Raman microspectroscopy and SEM-EDS X-ray microanalysis. Unpaired t-test (lesion dimensions) and one-way ANOVA and Newman-Keuls tests (Ca, P wt%, Ca/P ratios) were used to identify statistically significant differences in lesion analysis (alpha=0.05). RESULTS: (a) In vitro study. All restorations developed lesions at incisal and cervical margins. At gap-free regions glass-ionomers showed reduced lesion dimensions compared to those of composites (p<0.05). At regions with gaps, no significant differences were found in lesion depth between the restorative groups tested. Lesion length was increased in composite, and decreased in glass-ionomer, whereas lesion depth in both restorative groups was increased in comparison to gap-free regions (p<0.05).(b) In vivo study. No lesions were observed at gap-free regions. At gap regions, 75.5% of glass-ionomer and 62.5% of composite restorations developed lesions. The lesion dimensions were significantly greater in glass-ionomer (p<0.05). A reduction in PO4(3-), CO3(2-), Ca and P was found in lesions compared to intact tissues. No F was detected and no CaF2 lattice vibrations were found at the enamel margins facing the gap adjacent to glass-ionomers. SIGNIFICANCE: In the presence of a standardized interfacial gap, no preventive effect was exerted in vivo from the glass-ionomer to protect the adjacent enamel wall from secondary caries attack. The lack of any correlation between the in vivo and in vitro models tested implies that artificial caries experiments have a negligible clinical relevance in predicting the in vivo effect.