PubMed Health⌕ Search

Biomedical subjects

K Stephens

Publications and source records attributed to K Stephens.

At least 19 recordsLinked to original sources
PubMed2004

Applications of eTag trade mark assay platform to systems biology approaches in molecular oncology and toxicology studies.

We have developed a universal eTag trade mark multiplex assay platform that can be uniquely applied to survey the molecule profiles of biologic systems in sub-global large-scale analyses. The effectiveness of eTag trade mark assays when applied to focused system biology studies in molecular oncology and predictive toxicology is herein described while reviewing the current methods commonly used. The multi-analyte and multi-parameter assay approach for parallel analysis will form the basis of an emerging paradigm of multiplexed molecular profiling for signaling pathway networks and various aspects of drug development processes.

Gene Expression Profiling↗
PubMed2004

Communication skills training in postgraduate medicine: the development of a new course.

It has long been accepted that communication is of central importance in healthcare, and a core aspect of clinical competence. Many educational institutions and Royal Colleges now reflect this and consider communication skills a priority in postgraduate examination. The new examination "Practical Assessment of Clinical and Examination Skills" has replaced the Royal College of Physicians MRCP part 2 clinical and oral examination. This examination now consists of five clinical stations, two of which focus on communication skills. A short course for postgraduate trainees has been designed to address the communication skills requirements of the part 2 clinical examination. The aims, development, and content of the course are described. Emphasis is placed on candidates practising skills with patients and receiving feedback during the course. Evidence suggests that practice with feedback is an essential ingredient of communication skills courses, and is more effective than other methods such as observing experts or video examples, or simply discussing issues in communication. Results of a preliminary evaluation indicate that the course was perceived as valuable by candidates and that the aims, format, and content were appropriate. Although the preliminary evaluation was largely positive, it could be argued that the acid test of the effectiveness of a course is an objective evaluation of skills, observed before and after the course, a development that is being considered for future evaluation of the course. Recommendations for applying this type of training to postgraduate trainees in any branch of medicine are given.

Clinical Competence↗
PubMed2001

Recombination hotspot in NF1 microdeletion patients.

Neurofibromatosis type 1 (NF1) patients that are heterozygous for an NF1 microdeletion are remarkable for an early age at onset and an excessive burden of dermal neurofibromas. Microdeletions are predominantly maternal in origin and arise by unequal crossover between misaligned NF1REP paralogous sequence blocks which flank the NF1 gene. We mapped and sequenced the breakpoints in several patients and designed primers within each paralog to specifically amplify a 3.4 kb deletion junction fragment. This assay amplified a deletion junction fragment from 25 of the 54 unrelated NF1 microdeletion patients screened. Sequence analysis demonstrated that each of the 25 recombination events occurred in a discrete 2 kb recombination hotspot within each of the flanking NF1REPs. Two recombination events were accompanied by apparent gene conversion. A search for recombination-prone motifs revealed a chi-like sequence; however, it is unknown whether this element stimulates recombination to occur at the hotspot. The deletion-junction assay will facilitate the prospective identification of patients with NF1 microdeletion at this hotspot for genotype-phenotype correlation studies and diagnostic evaluation.

Alleles↗
PubMed2001

Expression of a truncated keratin 5 may contribute to severe palmar--plantar hyperkeratosis in epidermolysis bullosa simplex patients.

Epidermolysis bullosa simplex are dominant disorders of skin fragility characterized by intraepidermal blistering upon mild mechanical trauma. Skin fragility is caused by expression of either an abnormal keratin 5 or an abnormal keratin 14 protein, which compromises the structure and function of the keratin cytoskeleton of basal cells. We report an epidermolysis bullosa simplex patient with a novel single base substitution (A-->T1414) that changes the lysine residue at amino acid 472 to a non-sense codon (K472X). This change predicts the synthesis of a truncated keratin 5, missing 119 amino acids, including the entire tail domain and the highly conserved KLLEGE motif at the carboxy terminus of the 2B domain of the central rod. Expression of an altered keratin 5, of predicted mass and pI for the product of the K472X allele, was documented by one- and two-dimensional western blots of protein extracts from patient skin. Ultrastructural analysis of the patient's nonhyperkeratotic skin was remarkable for basal keratinocytes with dense and irregular keratin filaments proximal to the basement membrane. Keratinocytes, transfected with a cDNA carrying the A-->T1414 non-sense mutation, overexpressed a truncated keratin 5, and showed a disorganized and collapsed keratin filament cytoskeleton. This is the second epidermolysis bullosa simplex patient reported with a premature termination mutation in the KLLEGE motif. The remarkable occurrence of severe palmar--plantar hyperkeratosis in both patients suggests that the keratin 5 tail domain may have unrecognized, but important, normal functions in palmar-plantar tissues.

Amino Acid Sequence↗
PubMed2000

Evidence that juvenile myelomonocytic leukemia can arise from a pluripotential stem cell.

Children with neurofibromatosis type 1 (NF1) carry germline mutations in one allele of the NF1 gene and are predisposed to myeloid malignancies, particularly juvenile myelomonocytic leukemia (JMML). Disruption of the remaining NF1 allele can be found in malignant cells. Flow cytometric cell sorting techniques to isolate the malignant cell populations and molecular genetic methods to assay for somatic loss of the normal NF1 allele were used to study an unusual child with NF1 and JMML who subsequently had T-cell lymphoma. The data show that malignant JMML and lymphoma cells share a common loss of genetic material involving the normal NF1 gene and approximately 50 Mb of flanking sequence, suggesting that the abnormal T-lymphoid and myeloid populations were derived from a common precursor cell. These data support the hypothesis that JMML can arise in a pluripotent hematopoietic cell.

Cell Differentiation↗
PubMed2000

NF1 microdeletion breakpoints are clustered at flanking repetitive sequences.

Neurofibromatosis type 1 patients with a submicroscopic deletion spanning the NF1 tumor suppressor gene are remarkable for an early age at onset of cutaneous neurofibromas, suggesting the deletion of an additional locus that potentiates neurofibromagenesis. Construction of a 3.5 Mb BAC/PAC/YAC contig at chromosome 17q11.2 and analysis of somatic cell hybrids from microdeletion patients showed that 14 of 17 cases had deletions of 1.5 Mb in length. The deletions encompassed the entire 350 kb NF1 gene, three additional genes, one pseudogene and 16 expressed sequence tags (ESTs). In these cases, both proximal and distal breakpoints mapped at chromosomal regions of high identity, termed NF1REPs. These REPs, or clusters of paralogous loci, are 15-100 kb and harbor at least four ESTs and an expressed SH3GL pseudogene. The remaining three patients had at least one breakpoint outside an NF1REP element; one had a smaller deletion thereby narrowing the critical region harboring the putative locus that exacerbates neurofibroma development to 1 Mb. These data show that the likely mechanism of NF1 microdeletion is homologous recombination between NF1REPs on sister chromatids. NF1 microdeletion is the first REP-mediated rearrangement identified that results in loss of a tumor suppressor gene. Therefore, in addition to the germline rearrangements reported here, NF1REP-mediated somatic recombination could be an important mechanism for the loss of heterozygosity at NF1 in tumors of NF1 patients.

Female↗
PubMed2000

Formula for surgical practice resuscitation in an academic medical center.

BACKGROUND: Managing patient referrals for surgical consultation in an academic practice has traditionally emphasized clinical rather than service expertise. However, assuring both efficiency and accuracy in the initial consultation have become critical early measures of quality care. METHODS: In partnership with the academic medical center administration, current practice was analyzed. Performance and communication standards were established around an ideal patient experience. A new ambulatory consultation process was developed; and flowcharting methods for resource allocation, statistical process control, and pre-visit data collection were used to reduce patient administrative time. Automated referral reports engaged referring physicians throughout the consultation. RESULTS: Accurate insurance and referral authorization have been provided for all patients, including the 4% who are underinsured. Patient, provider, and referring physician satisfaction has increased significantly. Staff time investment has progressively declined from 52 +/- 11 (95% confidence) minutes to 34 +/- 10 minutes for most patients. Realignment of tasks has reduced the administrative time spent by the patient by 32% without compromising clinical time. New patient volume increased by 29% per year, maintaining regional market share. CONCLUSIONS: Expertise in the process of consultation delivery is feasible and will be increasingly critical to the survival of academic surgical practice in a competitive market.

Academic Medical Centers↗
PubMed2000

Refined mapping of the human serotonin transporter (SLC6A4) gene within 17q11 adjacent to the CPD and NF1 genes.

The SLC6A4 gene encodes the serotonin transporter, the target of an important class of antidepressant drugs (serotonin selective reuptake inhibitors). Polymorphisms in the SLC6A4 gene have been reported to be associated with susceptibility to depression and other psychiatric disorders. We have constructed a 1 Mb YAC and PAC contig which harbours both the SLC6A4 and the carboxypeptidase D (CPD) genes. The order of loci within the contig was cen-D17S975-D17S1549-24R-D17S1294-SLC6A4-28L+ ++-(CPD, D17S2009, D17S2004)-D17S2120-ter. Both genes were deleted in one of 17 neurofibromatosis type 1 (NF1) patients carrying submicroscopic NF1 contiguous gene deletions.

Carboxypeptidases↗
PubMed1999

Linkage analysis of candidate genes and the small, dense low-density lipoprotein phenotype.

There is accumulating evidence for the importance of small, dense low-density lipoprotein (LDL), the defining feature of the atherogenic lipoprotein phenotype, as a risk factor for coronary heart disease. Although both family studies and twin studies have demonstrated genetic influences on this phenotype, the specific gene(s) involved remain to be identified. The purpose of this study was to determine whether there was evidence for genetic linkage between small, dense LDL (LDL subclass phenotype B), as determined by gradient gel electrophoresis, and selected candidate genes known to be involved in lipid metabolism. The linkage analyses were based on a sample of 19 families, including 142 individual family members, using a lod score linkage analysis approach. Nine candidate genes were examined, including loci for manganese superoxide dismutase (Mn SOD2), apolipoproteins CIII, AII, and apo CII, lipoprotein lipase, hepatic lipase, microsomal triglyceride transport protein, the insulin receptor and the LDL receptor. The analyses did not provide significant evidence for genetic linkage between markers for any of these genes and LDL subclass phenotype B, nor did it confirm previous reports of linkage between the LDL receptor gene and LDL subclass phenotype B. Using three closely linked markers for the Mn SOD2 locus excluded close linkage between this candidate gene region and LDL subclass phenotype B. These findings demonstrate the complexity of genetically mapping risk factor phenotypes, and emphasize the necessity of identifying new genetic loci, other than known candidate genes, involved in susceptibility to atherosclerosis.

Apolipoproteins↗
PubMed1999

Cyclic ichthyosis with epidermolytic hyperkeratosis: A phenotype conferred by mutations in the 2B domain of keratin K1.

Bullous congenital ichthyosiform erythroderma (BCIE) is characterized by blistering and erythroderma in infancy and by erythroderma and ichthyosis thereafter. Epidermolytic hyperkeratosis is a hallmark feature of light and electron microscopy. Here we report on four individuals from two families with a unique clinical disorder with histological findings of epidermolytic hyperkeratosis. Manifesting erythema and superficial erosions at birth, which improved during the first few months of life, affected individuals later developed palmoplantar hyperkeratosis with patchy erythema and scale elsewhere on the body. Three affected individuals exhibit dramatic episodic flares of annular, polycyclic erythematous plaques with scale, which coalesce to involve most of the body surface. The flares last weeks to months. In the interim periods the skin may be normal, except for palmoplantar hyperkeratosis. Abnormal keratin-filament aggregates were observed in suprabasal keratinocytes from both probands, suggesting that the causative mutation might reside in keratin K1 or keratin K10. In one proband, sequencing of K1 revealed a heterozygous mutation, 1436T-->C, predicting a change of isoleucine to threonine in the highly conserved helix-termination motif. In the second family, a heterozygous mutation, 1435A-->T, was found in K1, predicting an isoleucine-to-phenylalanine substitution in the same codon. Both mutations were excluded in both a control population and all unaffected family members tested. These findings reveal that a clinical phenotype distinct from classic BCIE but with similar histology can result from K1 mutations and that mutations at this codon give rise to a clinically unique condition.

DNA Mutational Analysis↗
PubMed1999

Team approach to teaching participatory group process involving natural resources and agriculture.

Contemporary issues are defined by people who share diverse and often strongly defended views about the topic. In Oregon, citizens are increasingly being asked or expected to participate in complex decisions that require a consensus. Rather than teach one professor's synthesis of a contemporary natural resource issue, faculty from six disciplines coach group process, interactive learning skills, and systems thinking as a way to address complex issues from multiple perspectives. Students learn by grappling with a natural resource issue of their choice within groups based on a diversity among majors, degree status, and gender. Students define situation (S), brainstorm new or different targets (T), and analyze two or more pathways (P), using an STP learning and action process. Exploring potential pathways involves defining possible consequences, stakeholder views, feasibility (ecological, social, economic, and political), and planning that includes expected behavior of the improved system over time. Students present their topics and improvements showing systemic relationships, systematic analysis, and integration of scientific facts and secondary data at midterm and during finals. Reflective learning is fostered throughout the course with prompted questions in a journal notebook. Grading criteria promote meaningful inquiry and participation in group process combined with integration of scientific facts and reflective learning.

Agriculture↗
PubMed1998

Electrophysiological monitoring during selective dorsal rhizotomy, and spasticity and GMFM performance.

The relation between abnormal electrophysiological responses to intraoperative stimulation during selective dorsal rhizotomy (SDR) and the degree of spasticity and motor dysfunction was explored in 92 children with spastic cerebral palsy (CP) who underwent SDR at a single center. The proportion of abnormally responding rootlets was compared with the degree of spasticity measured with the modified Ashworth Scale (MAS) and with the spasticity measurement system (SMS) at discrete segmental levels. Motor impairment measured with the Gross Motor Function Measure (GMFM) was also compared with the proportion of abnormally responding dorsal rootlets. A consistent relation between the proportion of abnormally responding rootlets and the degree of spasticity and gross motor abnormality at the corresponding muscles could not be demonstrated. There was also no consistent association between the proportion of rootlets ablated during SDR and the change in spasticity measured with the MAS and SMS, or to the change in motor function as measured with the GMFM. These data suggest that the intraoperative monitoring technique most commonly used for SDR is unlikely to identify accurately those neural elements which contribute to spasticity in children with CP.

Adolescent↗
PubMed1997

Familial neurofibromatosis 1 microdeletions: cosegregation with distinct facial phenotype and early onset of cutaneous neurofibromata.

A notable subset of the recent literature on the disorder neurofibromatosis type 1 (NF1) describes patients with NF1, facial anomalies, and other unusual findings. We describe a molecular re-evaluation of two such families reported previously by Kaplan and Rosenblatt [1985], who suggested that their NF1 manifestations, facial phenotype, and other findings could result from a disorder distinct from NF1. Submicroscopic deletions involving the NF1 gene were identified in both families by fluorescent in situ hybridization and analysis of somatic cell hybrids. Affected subjects of the first family were heterozygous for a microdeletion of approximately 2 Mb, which included the entire NF1 gene and flanking contiguous sequences. The family was remarkable for cosegregation of the NF1 microdeletion with facial abnormalities and a pattern of early onset of cutaneous neurofibromata upon transmission from an affected mother to her three affected children. The propositus of the second family carried a deletion that at the least involved NF1 exon 2 through intron 27, which is > 200 kilobases in length. Because all persons in the family were deceased, the size of the deletion could not be determined precisely. Facial anomalies were observed in the propositus and his NF1-affected mother and sister. The data from these families support our hypothesis, which was initially based solely on sporadic deletion cases, that deletion of the entire NF1 gene, or in conjunction with deletion of unknown contiguous genes, causes the facial anomalies and early onset of neurofibromata observed in this subset of NF1 patients. In addition, other features observed in the persons in these families suggest that some NF1 microdeletion patients may be at increased risk for connective tissue abnormalities and/or neoplasms.

Adolescent↗
PubMed1997

Primers for exon-specific amplification of the KRT5 gene: identification of novel and recurrent mutations in epidermolysis bullosa simplex patients.

The KRT5 and KRT14 genes encode the proteins keratin 5 and 14, respectively, which are the primary structural components of the 10-nm intermediate filaments of the mitotic epidermal basal cells. A single mutation in either gene can disrupt the keratin intermediate filament cytoskeleton, resulting in the skin fragility and blistering that is characteristic of the group of inherited disorders known as epidermolysis bullosa simplex. We have established a mutation detection system that facilitates KRT5 gene analysis from leukocyte genomic DNA, obviating the need for a skin sample or keratinocyte culture for cDNA synthesis. KRT5 intronic regions that flanked each exon were sequenced and sets of facing intronic primers were designed for specific amplification of each of the nine KRT5 exons. Direct sequencing of KRT5-amplified exons identified three novel missense mutations. One mutation recurred in two unrelated patients with sporadic EBS. This glutamate to lysine substitution (E477K), located in the highly conserved KLLEGE motif at the end of the central rod domain, is the third recurrent mutation identified in dominant epidermolysis bullosa simplex disease. The corresponding glutamate in keratin 2e was previously reported to be frequently mutated in ichthyosis bullosa of Siemens, suggesting that this highly conserved residue may be a potential mutational hot spot in other type II keratins or nonkeratin intermediate filament proteins.

Alleles↗
PubMed1996

Genetics, phenotype, and natural history of autosomal dominant cyclic hematopoiesis.

Cyclic hematopoiesis, (CH, or cyclic neutropenia) is a rare disease manifested by transient severe neutropenia that recurs approximately every 21 days. The hematologic profile of families with the autosomal dominant form (ADCH) has not been well characterized, and it is unknown if the phenotype is distinct from the more common sporadic congenital or acquired forms of CH. We studied nine ADCH families whose children displayed typical CH blood patterns. Pedigrees confirmed dominant inheritance without evidence of heterogeneity or decreased penetrance; three pedigrees suggested new mutations. Families were Caucasian with exception of one with a Cherokee Native American founder: A wide spectrum of symptom severity, ranging from asymptomatic to life-threatening illness, was observed within families. The phenotype changed with age. Children displayed typical neutrophil cycles with symptoms of mucosal ulceration, lymphadenopathy, and infections. Adults often had fewer and milder chronic neutropenia without distinct cycles. While CH is commonly described as "benign", four children in three of the nine families died of Clostridium or E. coli colitis, documenting the need for urgent evaluation of abdominal pain. Misdiagnosis with other neutropenias was common but can be avoided by serial blood counts in index cases. Genetic counseling requires specific histories and complete blood counts in relatives at risk to assess status regardless of symptoms, especially to determine individuals with new mutations. We propose diagnostic criteria for ADCH in affected children and adults. Recombinant human granulocyte colony-stimulating factor treatment resulted in dramatic improvement of neutropenia and morbidity. The differential diagnosis from other forms of familial neutropenia is reviewed.

Adolescent↗
PubMed1996

Patterns of hematopoietic lineage involvement in children with neurofibromatosis type 1 and malignant myeloid disorders.

Children with neurofibromatosis type 1 (NF1) are at increased risk of developing malignant myeloid disorders, particularly juvenile chronic myelogenous leukemia/juvenile myelomonocytic leukemia (JCML/JMML). We investigated bone marrows from 11 such patients (8 boys and 3 girls) and detected allelic losses at the NF1 locus in 4 of them and probable losses in 2 others. To determine which hematopoietic cell lineages were derived from the abnormal clones, Epstein-Barr virus (EBV)-transformed cell lines and CD34+ cells were analyzed from 3 children with JCML with allelic losses in unfractionated marrow. CD34 cells from these 3 patients lacked the normal NF1 allele, whereas EBV cell lines retained it. Erythroblasts plucked from the burst-forming unit-erythroid colonies of one of these children lacked the normal NF1 allele. We also studied a 10-month-old boy with NF1 who developed an unusual myeloproliferative syndrome. His bone marrow and EBV cell line both showed loss of the normal NF1 allele. In our series and in the literature, male sex and maternal transmission of NF1 were associated with the highest risk of myeloid leukemia. These data (1) provide strong genetic evidence that NF1 functions as a tumor-suppressor in early myelopoiesis, (2) confirm the clonal nature of JCML/JMML, (3) suggest that the elevation in fetal hemoglobin seen in JCML/JMML is a result of primary involvement of erythroid progenitors in the malignant clone, (4) show consistent loss of NF1 in the CD34 cells of affected children and show that the malignant clone may also give rise to pre-B cells in some cases, and (5) implicate epigenetic factors in the development of leukemia in children with NF1.

Acute Disease↗