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Biomedical subjects

R C McGlennen

Publications and source records attributed to R C McGlennen.

At least 19 recordsLinked to original sources
PubMed2002

Leukemia in donor cells after allogeneic hematopoietic stem cell transplant.

The development of leukemia in donor cells after allogeneic hematopoietic stem cell transplant is an extremely rare event. We report here the case of a patient who developed myelodysplastic syndrome/acute myeloid leukemia, in cells of donor origin 3.5 years after related donor HSCT for refractory chronic lymphocytic leukemia and therapy-induced myelodysplastic syndrome. The origin of the leukemia was determined by analysis of minisatillite polymorphism tested on CD34(+) cells.

Adult↗
PubMed2001

Simultaneous allele-specific amplification: a strategy using modified primer-template mismatches for SNP detection--application to prothrombin 20210A (factor II) and factor V Leiden (1691A) gene mutations.

BACKGROUND: Inherited thrombophilia is caused by mutations in genes central to the clotting cascade. Analysis of the factor V Leiden (FVL) and prothrombin G20210A mutations are the most prevalent in thrombophilia. METHODS AND RESULTS: We have optimized an allele-specific PCR assay for the simultaneous detection of both wild-type and mutant alleles. This method is adapted for clinical use with the FVL and prothrombin G20210A assays and is significant in its intentional use of nucleotide mismatches at the 3' end of allele-specific primers. Two internal allele-specific primers are designed to amplify in opposite directions on opposite strands that reduce differential amplification. Our results show concordance with methods involving PCR with restriction endonuclease digestion, yet are simpler to perform. CONCLUSION: The simultaneous allele-specific amplification method allows simultaneous detection of wild-type and mutant alleles by PCR using four distinct primers. Nucleotide mismatches in the primers reduce competitive amplification.

Alleles↗
PubMed2001

Miniaturization technologies for molecular diagnostics.

BACKGROUND: Molecular diagnostics devices are becoming smaller. With the advancement of miniaturization technologies, microchip-based systems will soon be available for genetic testing. The purpose of this review is to highlight the underlying principles in miniaturization, the strategies being developed for bioanalysis, and the potential impact on the practice of this rapidly growing medical discipline. APPROACH: The author discusses DNA microchips and their practical importation into the clinical laboratory, based on his background in medical device and microchip design and development. His discussion is supported by a body of literature covering both biomedical and electrical engineering and more recent publications in the field of molecular genetics and pathology. CONTENT: This review is descriptive and intended to outline the technologic and methodologic approaches to the creation of an integrated genetic analysis instrument based on miniature components. The review draws on published scientific evaluations of these devices without regard to the companies involved in their development. SUMMARY: The intent of this review is that the reader will better understand the variety of technical approaches toward the miniaturization of molecular genetic testing for the clinical laboratory. With insight into the principles underlying the operation of these chips and the integrated systems, the end user can better evaluate the value to the field in terms of making molecular genetics testing simpler, faster, and less expensive.

Biosensing Techniques↗
PubMed2000

Hereditary spastic paraplegia and hereditary ataxia, Part 2: A family demonstrating various phenotypic manifestations with the SCA3 genotype.

BACKGROUND: Clinical descriptions of the dominantly inherited ataxic motor syndromes in a 7-generation family of German origin were first reported in 1951. OBJECTIVE: To provide follow-up clinical, pathological, and genetic data for 9 patients in this family. DESIGN: Clinical histories and neurologic findings, gross and microscopic pathological features, and DNA analysis. RESULTS: Clinical presentations in this closely followed up portion of the family include fairly uniform ataxic and upper motor neuron symptoms. Nystagmus was a conspicuous and early sign, but generational anticipation was not evident. Although often present, amyotrophy was not a major source of disability. Major pathological degeneration was noted in the pons, spinal cord, and upper brainstem, where ubiquitin-immunoreactive intranuclear inclusion bodies were demonstrated. The diagnosis of Machado-Joseph disease (SCA3 [spinocerebellar ataxia type 3] genotype) was established from autopsy tissue in 1 patient and from blood specimens in 6 others. CONCLUSIONS: Clinical variation within this family and between this family and families with the SCA1 and SCA3 genotypes is so broad as to make the genetic diagnosis from clinical criteria alone practically impossible. The pathological definition of Machado-Joseph disease is more reliable, but some findings do overlap those of other genotypes. To our knowledge, the basis for the phenotypic variations in Machado-Joseph disease, genetic or otherwise, has not been established.

Adult↗
PubMed2000

Non-leukemic autologous reconstitution after allogeneic bone marrow transplantation for Ph-positive chronic myelogenous leukemia: extended remission preceding eventual relapse.

Autologous reconstitution is the recovery of autologous hematopoietic function after failure of an allogeneic graft to establish sustained hematopoiesis either with or without preceding donor engraftment. We reviewed 9 years experience of the University of Minnesota and identified 10 of 291 patients who underwent allogeneic BMT for Ph-positive CML and developed non-leukemic autologous reconstitution. All patients received the same preparative regimen with cyclophosphamide and total body irradiation. Eight patients had a 6/6-antigen matched donor. Eight patients received their graft from an unrelated donor. In five cases the graft was T cell-depleted. Non-malignant autologous reconstitution initially manifested as mixed chimerism in nine of 10 patients and lasted for a median of 11 (3-41) months. Eight patients have relapsed and four are still alive. The two relapse-free patients have died 24 and 48 months post transplant. Of the four surviving patients, two are in interferon-induced cytogenetic remission at 53+ and 101+ months of follow-up. Autologous non-leukemic reconstitution is uncommon, but appears to be a distinct clinical syndrome, perhaps occurring more frequently after unrelated donor BMT. Although usually followed by relapse, relapse-free survival may be prolonged.

Adult↗
PubMed2000

Evaluation of an automated technique for assessment of marrow engraftment after allogeneic bone marrow transplantation using a commercially available kit.

Several methods have been used to evaluate engraftment after allogeneic bone marrow transplantation (BMT). We assessed the usefulness of a multiple short tandem repeat (STR) amplification kit combined with a capillary electrophoresis unit for DNA identity analysis in the evaluation of engraftment after BMT. For 17 of 18 patients, at least 1 locus showed unique alleles for the donor and the recipient. In all cases, at least 1 locus was informative for the presence of small amounts of recipient DNA. The results from STR analysis were the same as Southern blot analysis in 14 of 17 cases. Differences included mixed chimerism detected only with STR analysis, informative loci present only with STR analysis, and informative loci present only with Southern blot analysis (1 case each). By using mock mixed chimeras, minor populations of 5% were detected routinely in all loci using the kit manufacturer's default protocol. By increasing the amount of amplified DNA, minor populations of 1% were detected in all cases but not in all loci. This single reaction technique provides for faster results, reduced workforce needs, and greater sensitivity than traditional Southern blot.

Adolescent↗
PubMed2000

Human papillomavirus oncogenesis.

HPVs have evolved to accomplish the task of controlling host cell proliferation and differentiation to the end of producing more infectious virions. Coincident with the viral life cycle, however, is the risk that the viral genome will be disrupted and its DNA integrated into the host cell chromosomes. Integration of the viral genome is potentiated by host factors and extracellular effectors that alone may increase genetic instability. But it is consequent to viral integration that most HPV-associated malignancies develop. Investigations of the potential for HPV to immortalize primary cells or transform immortalized cells in vitro demonstrate two distinct classes of genital viral types: (1) oncogenic, exemplified by HPV 16 and 18; and (2) the nononcogenic types 6 and 11. Subsequently, localization of the HPV oncogene implicated that E6 and E7 act by uncoupling the checkpoint controls of the cell cycle principally by inhibiting the normal functioning of p53 and pRb, respectively. By in large, the nononcogenic viruses do not effect irreversible growth properties through these same viral genes and the same cellular counterparts.

Animals↗
PubMed1999

Analysis of a very large trinucleotide repeat in a patient with juvenile Huntington's disease.

A patient with juvenile Huntington's disease (HD) of probable maternal inheritance is reported. The expanded IT-15 allele was only detected with the use of modified PCR and Southern transfer techniques, which showed a CAG trinucleotide repeat expansion of approximately 250 repeats-the largest CAG expansion reported within the huntingtin gene. This case emphasizes the need for communication between the diagnostic laboratory and the clinician to define the molecular genetics of unusual cases.

Adolescent↗
PubMed1998

bcl-1 gene rearrangements in mantle cell lymphoma: a comprehensive analysis of 118 cases, including B-5-fixed tissue, by polymerase chain reaction and Southern transfer analysis.

We evaluated 118 cases of mantle cell lymphoma by polymerase chain reaction (PCR) for the major translocation cluster (MTC) region and another breakpoint corresponding to probe p94PS, located 24 kb telomeric to the MTC locus on chromosome 11. The specimens included 64 frozen, 19 formalin-fixed, and 9 B-5-fixed lymph nodes and 26 B-5-fixed bone marrow biopsy specimens. We also analyzed DNA from the 64 frozen lymph nodes by Southern transfer analysis (SB) using three separate bcl-1 breakpoint probes. Gene rearrangements were identified in 17 (PCR) and 18 (SB) of 64 frozen lymph nodes and by PCR in 6 of 19 formalin-fixed lymph nodes, 3 of 9 B-5-fixed lymph nodes, and 12 of 26 B-5-fixed bone marrow cores with MTC locus primers and probe. Only one case showed rearrangement with the p11EH probe that corresponds to breakpoints situated 63 kb telomeric to the MTC locus. No rearrangements were detected by PCR or SB for the breakpoint site corresponding to the p94PS probe, but we identified a polymorphic restriction site with HinD III digest in approximately 25% of the cases. In agreement with other studies, these results confirmed that breakpoints in the MTC region of the bcl-1 locus are tightly clustered and associated with 30 to 40% of mantle cell lymphomas. Other breakpoints in the bcl-1 locus seem to be heterogeneous and cannot be detected by PCR or SB with use of existing probes or primer sequences. The most important finding of our study is optimization of the methodology for the detection of immunoglobulin heavy chain gene rearrangement and MTC region breakpoints by PCR from the DNA isolated from B-5-fixed, paraffin-embedded lymph nodes and bone marrow biopsy specimens. The results obtained from these tissues are comparable to those obtained from frozen or formalin-fixed tissue.

Blotting, Southern↗
PubMed1997

Detection of the factor VLeiden mutation. Development of a testing algorithm combining a coagulation assay and molecular diagnosis.

The Coagulation and Molecular Diagnostic laboratories at the University of Minnesota Medical School (Minneapolis) have collaborated to develop a diagnostic algorithm to identify all factor VLeiden mutation carriers without performing unnecessary and expensive genetic testing. The algorithm uses a coagulation assay for activated protein C resistance (APCR) to determine the need for genetic testing. We report the results of our experience validating this program. We compared the sensitivity, specificity, and positive and negative predictive values of two measures of APCR, the APCR ratio and the normalized ratio. We found that the normalized ratio was the more sensitive but less specific parameter to determine the need for genetic testing. By using the normalized ratio as the standard by which to refer patients to the Molecular Diagnostics Laboratory, all mutation carriers were identified. We found a large overlap in both measures of APCR between symptomatic patients with normal genotype and mutation carriers. Furthermore, we demonstrated that increased factor VIII levels with a normal genotype are associated with apparent APCR. In this article we also review other correlates of apparent APCR.

Adult↗
PubMed1997

Correlation of PCR-detected clonal gene rearrangements with bone marrow morphology in patients with B-lineage lymphomas.

Bone marrow biopsy is the conventional staging and posttherapy evaluation method for assessing marrow involvement by lymphoma. Polymerase chain reactions (PCR) for antigen receptor rearrangements have the potential to increase the detection of minimal degrees of marrow involvement. The present study is a concurrent morphologic and PCR evaluation of 225 staging or posttherapy marrow biopsies from 127 patients with B-lineage non-Hodgkin's lymphoma. The biopsies were morphologically categorized into four groups: group 1 (positive for lymphoma), 60 biopsies (27%); group 2 (suspicious for lymphoma), 20 biopsies (9%); group 3 (lymphocytic lesions of indeterminate biology), 22 biopsies (10%); and group 4 (negative for lymphoma), 123 biopsies (54%). Molecular studies were performed on concurrently obtained aspirates and used consensus immunoglobulin-heavy-chain (IgH) and IgH/bcl-2 gene PCR primers. A molecular clone was detected in 53 of the 225 aspirates (24%): group 1, 34 aspirates (57%); group 2, five aspirates (25%); group 3, one aspirate (5%); and group 4, 13 aspirates (11%). A PCR-positive aspirate was present in 47% of follicular lymphomas, 58% of diffuse large cell lymphomas, and 72% of the other lymphomas in the group I specimens. Morphology or PCR was positive in 79 of the 225 cases (35%). The molecular detection of clonality in the aspirate DNA from cases with positive morphologic findings was lower than anticipated. The discordance between morphology and PCR results may be related to sample variation between the trephine biopsy and aspirate, a failure to aspirate sufficient lymphoma cells, or insufficient primer homology for amplification. DNA extracted from trephine sections may provide results more concordant with morphology, because PCR detected a clone in 10 of 11 DNA specimens extracted from trephine biopsies with positive morphologic findings and PCR negative aspirates.

Base Sequence↗
PubMed1996

The E5 gene product of rhesus papillomavirus is an activator of endogenous Ras and phosphatidylinositol-3'-kinase in NIH 3T3 cells.

We examined the effect of two rhesus papillomavirus 1 (RhPV) oncogenes on cytokine-induced signal transduction pathways leading to the possible activation of Ras protein (p21ras) and phosphatidylinositol kinase. p21ras in both the activated (GTP-bound) and inactivated (GDP-bound) states were quantitated. NIH 3T3 cell lines expressing the RhPV 1 E5 gene or epidermal growth factor receptor cDNA had about a sixfold higher ratio of p21ras-bound GTP to p21ras-bound GDP as compared with parental NIH 3T3 cells or a cell line expressing the RhPV 1 E7 gene under normal culture conditions, yet expressed similar levels of p21ras. Quiescent cells had dramatically reduced levels of activated p21ras, except those containing RhPV 1 E7. Levels were restored by stimulation with epidermal growth factor or platelet-derived growth factor. Both epidermal growth factor and platelet-derived growth factor receptor of RhPV 1 E5- and E7-containing cells responded to cytokine stimulation. Endogenous phosphatidylinositol-3'-kinase was up-regulated in NIH 3T3 cells transformed with the E5 genes of RhPV 1 and bovine papillomavirus 1. These results suggest that E5 genes of papillomaviruses play a major role in the regulation of transduction pathways.

3T3 Cells↗
PubMed1996

Posttransplant T-cell lymphoproliferative disorders--an aggressive, late complication of solid-organ transplantation.

T-cell non-Hodgkin's lymphomas are an uncommon occurrence after solid-organ transplantation. We describe a morphologically and immunophenotypically distinct group of T-cell lymphoproliferative disorders that occurred late in the course of six patients with solid-organ transplants. The patients ranged in age from 31 to 56 years (median, 43). Three were male; all were splenectomized. The interval from transplant to the diagnosis of lymphoma ranged from 4 to 26 years (median, 15). Symptoms at presentation were related to sites of involvement. Pulmonary, marrow, and CNS involvement were present in five, four, and one case, respectively. No patient had lymphadenopathy. Five patients had an elevated lactate dehydrogenase level (range, 226 to 4,880 IU/L; median, 1,220 IU/L). Five of six patients had a leukoerythroblastic reaction. All cases had large-cell histology and frequently contained cytoplasmic granules. Those cases tested expressed CD2, CD3, and CD8 and were negative for B-cell antigens. T-cell receptor beta- and gamma-chain genes were clonally rearranged in three of three and one of three cases, respectively. All T-cell posttransplant lymphoproliferative disorders (T-PTLDs) studied were negative for Epstein-Barr virus (EBV), human T-cell leukemia/lymphoma virus type 1 (HTLV-1), human T-cell leukemia/lymphoma virus type 2 (HTLV-2), and human herpes virus type 8 (HHV-8) genomes. Treatment with acyclovir (three patients) or chemotherapy (three patients) resulted in two responses. All patients had an aggressive course, with a median survival duration of 5 weeks. In conclusion, a clinically aggressive T-PTLD may be a late complication of solid-organ transplantation and does not appear to be related to EBV, HTLV-1, HTLV-2, or HHV-8 infection.

Adult↗
PubMed1996

Sebaceous carcinoma of the vulva: a case report and review of the literature.

Although sebaceous glands are prominent on the vulva, sebaceous carcinomas of the vulva rarely occur. In fact, there have been only two cases of sebaceous carcinomas of the vulva reported in the literature. Eighty percent of vulvar cancers are squamous in origin with human papillomavirus (HPV) DNA detected in approximately 60% of these cancers. We present a third patient with sebaceous carcinoma of the vulva and the first to our knowledge that has been analyzed for HPV DNA. The case report and a review of the literature are presented.

Adenocarcinoma, Sebaceous↗
PubMed1996

Dynamic mutations pose unique challenges for the molecular diagnostics laboratory.

Routine clinical molecular testing of diseases associated with unstable or dynamic trinucleotide repeat syndromes poses unique technical, medical, and ethical challenges to the laboratory. Although the pathophysiology of these disorders is to date still largely undefined, the uniformity of their genetics has led to the development of highly informative diagnostic tests. In general, amplification techniques, such as the polymerase chain reaction (PCR), are used to determine the size of alleles within the genes linked to these disorders. Technically, these assays require empirical optimization so that the PCR reactions are both robust and reproducible, and occasionally other methods must be used to confirm diagnoses. Beyond execution of the test, however, the molecular diagnostics laboratory needs also to be fundamentally involved in the process of interpreting these tests in the correct clinical context and in setting policy as to how these data are presented to patients.

Chemistry, Clinical↗