Genetic linkage of the dentinogenesis imperfecta type III locus to chromosome 4q.

Dentinogenesis imperfecta type III (DGI-III) is an autosomal-dominant disorder of dentin formation which appears in a tri-racial southern Maryland population known as the "Brandywine isolate". This disease has suggestive evidence of linkage to the long arm of human chromosome 4 (LOD score of 2.0) in a family presenting with both juvenile periodontitis and DGI-III. The purpose of this study was to screen a family presenting with only DGI-III to determine if this locus was indeed on chromosome 4q. Furthermore, we wanted to determine if DGI-III co-localized with dentinogenesis imperfecta type II (DGI-II), which has been localized to 4q21-q23. Therefore, a large kindred from the Brandywine isolate was identified, oral examination performed, and blood samples collected from 21 family members. DNA from this family was genotyped with 6 highly polymorphic markers that span the DGI-II critical region of chromosome 4q. Analysis of the data yielded a maximum two-point LOD score of 4.87 with a marker for the dentin matrix protein 1 (DMP1) locus, a gene contained in the critical region for DGI-II. Our results demonstrated that the DGI-III locus is on human chromosome 4q21 within a 6.6 cM region that overlaps the DGI-II critical region. These results are consistent with the hypothesis that DGI-II is either an allelic variant of DGI-III or the result of mutations in two tightly linked genes.

Role of individual N-linked glycosylation sites in the function and intracellular transport of the human alpha folate receptor.

Glycosylation is a structural feature of all three isoforms of the human folate receptor. We have used site-directed mutagenesis to study the role of individual glycosylation sites in the assembly and function of the a isoform of the human folate receptor (alpha(h)FR). Three potential N-linked glycosylation sites in the alpha(h)FR sequence were disrupted by conservative mutation of the S or T residues in the consensus sequence (N-X-S/T) to A or V, respectively. Constructs with the single mutations S(71)-->A (alpha(h)FR(-1)), T(163)-->V (alpha(h)FR(-2)), and S(203)-->A (alpha(h)FR(-3)); the double mutation S(71)--> A/S(203)-->A (alpha(h)FR(-1-3)); and the triple mutation S(71)--> A/S(203)--> A/T(163)--> V (alpha(h)FR(-1-2-3)) were stably transfected into Chinese hamster ovary (CHO) cells. The proteins produced in CHO cells by the mutated cDNAs have apparent molecular weights that are reduced relative to the wild type and are consistent with the loss of carbohydrate residues. The triple mutant, which lacks all three consensus glycosylation sites, yields protein that comigrates with the enzymatically deglycosylated native protein. Determinations of the K(D) for folic acid by Scatchard analyses of the glycosylation mutants indicate that folic acid binding affinity is not significantly affected in the single mutants alpha(h)FR(-1) and alpha(h)FR(-2). However, in the single mutant, alpha(h)FR(-3), and the double mutant, alpha(h)FR(-1-3), folic acid binding affinity is respectively 2.7- and 3.5-fold lower than that in wild type. Deglycosylation by mutation of all three consensus sites (alpha(h)FR(-1-2-3) eliminates both folic acid binding and cell surface expression. In contrast, enzymatic deglycosylation of purified wild-type alpha(h)FR with endoglycosidase F does not significantly affect folate binding affinity. Thus, while carbohydrate residues are not essential for the folate binding activity of the mature folate receptor, at least one of the three core glycosylated residues is necessary for the synthesis of alpha(h)FR in its active conformation.

An open trial of paroxetine in patients with noncombat-related, chronic posttraumatic stress disorder.

The symptom overlap between posttraumatic stress disorder (PTSD) and other pharmacotherapy-responsive disorders suggests that pharmacotherapy might be effective. Nevertheless, of the eight published placebo-controlled trials investigating the pharmacotherapy of PTSD, only four found statistically significant efficacy for the treatment being studied. This literature possesses a number of methodologic limitations, including the fact that most studies have been conducted with war veterans, who may constitute a more treatment-refractory population. Several open trials and one controlled trial with selective serotonin reuptake inhibitors have reported improvement in some or all core PTSD symptoms (reexperiencing, avoidance, numbing, and hyperarousal). The authors hypothesized that paroxetine might be effective in PTSD, based on findings of its particular efficacy for anxiety and agitation in studies of depressed patients. The study presented here summarizes a 12-week, open-label trial of paroxetine among patients with noncombat-related, chronic PTSD. Outcome was assessed by an independent evaluator, the treating physician, and the patient, with the use of established rating scales for depression, anxiety, general symptoms, and PTSD core symptoms. A repeated-measures analysis of variance revealed highly significant improvement in all three symptom clusters, as well as in associated anxiety, depressive, and dissociative symptoms, with 11 of 17 (65%) patients rated as much or very much improved. The mean reduction in PTSD symptom scores was 48%. Exploratory analyses revealed that cumulative childhood trauma was negatively correlated with pharmacotherapy response (r = -0.52, p = 0.03). There was also significant variation in the time course of response across symptom clusters, which is suggestive of multiple mechanisms of response. Because paroxetine seems a highly promising treatment for all three symptom clusters of PTSD, a placebo-controlled clinical trial is warranted.

Reassignment of the 92-kDa type IV collagenase gene (CLG4B) to human chromosome 20.

The collagenase type IV B gene (CLG4B) was previously mapped to human chromosome 16 by hybridization of a cDNA probe to DNAs from a somatic cell hybrid panel. We have relocalized CLG4B to chromosome 20 based on three different lines of evidence: screening a somatic cell hybrid mapping panel, fluorescence in situ hybridization (FISH), and linkage analysis using a newly identified polymorphism.

Epithelioid hemangioendothelioma with multiple site involvement. Literature review and observations.

This report is a case of epithelioid hemangioendothelioma presenting as multiple lytic lesions of the ilium with radiographic findings of diffuse, bilateral lung involvement and biopsy-proven scalp involvement. Histologically, the tumor within bone and skin exhibited cords and nests of plump, epithelioid-appearing cells exhibiting rudimentary vascular differentiation within a myxohyaline stroma. Aggressive histologic features were not present. Immunohistochemical reactivity for Factor VIII-related antigen, Q-bend 10 (CD34), and cytokeratin were demonstrated. Ultrastructural studies revealed abundant intermediate cytoplasmic filaments, pinocytotic vacuoles, and Weibel-Palade bodies. The concurrent bone, skin, and lung involvement, low-grade histologic type, and female sex of the patient aroused speculation about the role of hormones in the development and possible treatment of the tumor, but estrogen and progesterone receptors were not detected. Despite intense combination chemotherapy, the patient died of widely metastatic disease. This report demonstrates the aggressive potential of histologically low-grade epithelioid hemangioendothelioma and the need for a thorough evaluation for metastases.

Decreased expression of the human folate receptor mediates transport-defective methotrexate resistance in KB cells.

The human membrane-associated folate binding protein, or folate receptor (hFR), is a necessary component of folate and methotrexate (MTX) transport in some cell lines. To investigate the role of hFR in acquired MTX resistance in human cells, we characterized nine MTX-resistant clones selected from human nasopharyngeal epidermoid carcinoma (KB) cells (cells which transport folates/anti-folates via the hFR) cultured in media containing low folate concentrations. Compared with wild type KB cells, the level of resistance of the clones ranges from 2- to 80-fold higher and the resistant phenotypes of the clones are characterized as follows. 1) DHFR levels are increased (3-13-fold) in four of nine clones; 2) MTX polyglutamation is not detectably different; 3) the extents of MTX efflux are similar; 4) initial rates of MTX efflux are similar except for two clones which exhibit slightly faster efflux rates (approximately 2-fold); and 5) the Vmax for specific MTX and 5-methyltetrahydrofolate transport are decreased (2-18-fold) in all mutants. The Kt values for MTX transport of each mutant are similar to the Kt of KB cells. These results indicate that all nine MTX-resistant clones exhibit defective MTX transport and that four clones also have increased DHFR levels. Based on folic acid binding assays, the hFR is reduced by 1.8-24-fold in these clones relative to KB cell hFR expression. Western, Northern, and Southern analyses are consistent with decreased hFR expression in these clones rather than mutations, resulting in alterations in the size or ligand binding affinities of the hFR. The decrement in hFR expression correlates closely with the degree of reduction in MTX transport Vmax for each clone. Since folate and MTX influx proceed via hFR in KB cells and in these mutants, the correlation (R2 = 0.90) between hFR expression and the MTX transport Vmax of each clone indicates that hFR expression is an important determinant of acquired MTX resistance in this human tumor cell line. These studies demonstrate that defective transport (manifested by decreased Vmax) resulting from decreased expression of the hFR is frequent in KB cells cultured under these conditions and suggest that modulation of hFR may be relevant to MTX cytotoxicity or resistance in tissues or cells expressing functionally significant levels of hFR.

Mothers' working models of infant feeding: description and influencing factors.

This study examined four aspects of mothers' working models of infant feeding: (1) infant behaviour that cues feeding decisions; (2) infant self-regulative behaviour; (3) importance of infant self-regulative behaviour and maternal effort directed to it; and (4) maternal effort and value given to task-oriented and efficient feeding. The relation of these aspects to maternal experience, age, formal education, family income, and feeding method was also explored. Subjects were 122 mothers of healthy, term infants between 14 and 60 days old. A telephone interview obtained demographic and attribute data and assessed the four aspects with 30 scaled items. Crying before and sleepiness during feeding were relatively compelling cues for maternal action. Most mothers gave only moderate ratings to the importance of infant self-regulative behaviour and to task-oriented and efficient feeding. Parity and feeding method affected response to specific items, with primiparae more concerned about length and regularity of feedings. Mothers who bottle-fed their infants were more concerned about maintaining a feeding once initiated than mothers who breast-fed their infants. Four-factor analysis yielded two item clusters with good internal consistency: Cluster a. Maternal Effort to Accomplish Feeding Goals; and Cluster b. Importance of Infant Self-Regulative Behaviour. Cluster a. and b. were strongly correlated. Feeding method influenced both clusters, and the interaction of parity and feeding method had an effect on Cluster a. Mothers with lower family income had higher scores on Cluster b; multiparae with lower family income on Cluster a. How and when infant self-regulation develops as a goal is a question in need of study.

Future directions for antifolate drug development.

We have discussed potential ways by which new antifolates could be designed and utilized to effect both sensitive and resistant cell/tumor inhibition. Targeting of alternative folate-dependent enzymes and increasing net intracellular accumulation (transport) and polyglutamation of antifolates should be useful approaches. In addition, antibodies to and cloned cDNAs of the transport components (e.g., folate transport proteins) will allow better characterization and understanding of de novo and acquired antifolate resistance states and may provide insights into new drug development.

Decision rules for infant feeding: the influence of maternal expertise, regulating functions, and feeding method.

Decision rules used by mothers to determine actions for tasks and events of infant feeding were described and factors contributing to their use (mothers' functions in regulating the feeding, maternal experience, and feeding method) were explored. A telephone interview surveyed 122 mothers of healthy term infants 14-60 days old. Both Decision Rules and Regulating Function (Maternal-, Infant-, or Mutual-Regulation) varied by feeding task or event examined. Decision Rules and Regulating Functions were associated only for burping during a feeding. Maternal experience (primiparity, multiparity) was not associated with either Decision Rule or Regulating Functions. For several feeding tasks or events, Decision Rule was more likely to be predicted by Regulating Function when feeding method (breast, bottle) was controlled. A mother's agenda for the feeding may be an important component of her internal working model of infant feeding and a factor in making decisions for action.

Training community developmental disabilities associates: a collaborative model.

Direct care staff play critical roles in contributing to the successful community adjustment of individuals with developmental disabilities. The current shortage of qualified personnel for these positions, however, will hinder future community integration efforts, particularly as individuals with more intensive needs attempt to live in the community. Improvements, both in the training of staff and in the pay and other incentives they receive, are needed. One response to this growing need is associate degree training that is being provided by a few community colleges and technical schools throughout the country. This article briefly describes the implementation and major components of such a program now being offered through Wisconsin's Vocational, Technical, and Adult Education System. Developed in response to local needs and representing an ongoing collaborative effort among the academic, advocacy, and service communities, the program also illustrates an important role the University Affiliated Facility can play in promoting exemplary training.

ABO-incompatible bone-marrow transplantation: removal of red blood cells from donor marrow avoiding recipient antibody depletion.

Eight patients with acute leukaemia undergoing allogeneic bone-marrow transplantation from ABO-incompatible donors received red-cell-depleted donor marrow without any procedure to diminish their anti-ABO antibody titres. Successful marrow red-cell removal (mean 98.8%) was achieved by means of a large-volume separation technique on Ficoll-Metrizoate in the IBM 2991 blood-cell processor. Clinically significant ABO-haemolytic reaction was prevented in all patients, and there was neither failure of engraftment nor rejection. This approach used alone is satisfactory for most ABO-incompatible marrow-transplant recipients, although combining this with some method of recipient antibody depletion, such as plasma exchange, is recommended in the occasional patient with high anti-ABO titres.