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Biomedical subjects

M R Brown

Publications and source records attributed to M R Brown.

At least 37 records · Page 2Linked to original sources
PubMed2000

HPV subtype analysis in lower genital tract neoplasms of female renal transplant recipients.

OBJECTIVE: Immunocompromised patients, such as female renal transplant recipients, have an increased incidence of neoplasms involving the lower genital tract (i.e., cervix, vagina, vulva). The relationship between lower genital tract neoplasms and human papillomavirus (HPV) infection has been established and high-risk oncogenic subtypes have been identified (HPV 16, 18, 45, and 56). The purpose of this study is to evaluate HPV subtypes present in lower genital tract neoplasms of post renal transplant women and compare HPV subtypes found in these patients with immunocompetent patients having similar neoplasms and normal immunocompetent controls. METHODS: Twenty specimens from lower genital tract neoplasms of 16 renal transplant patients, 13 specimens from 13 immunocompetent patients with similar histology, and 13 patients with normal lower genital tract histology were analyzed for the presence of HPV using polymerase chain reaction. HPV primers including the L1 (late) region consensus primers and primers specific for the HPV E6 (early) region for subtypes 6, 11, 16, and 18 were amplified with DNA from the above patient samples. RESULTS: Overall, HPV was detected in 21/46 specimens tested. Thirteen of the HPV-positive specimens were from transplant patients, and 8 were from immunocompetent patients (5 immunocompetent with disease and 3 normal patients). This difference in the total number of HPV-positive cases was statistically significant between the transplant and immunocompetent group (P = 0.02). Although no difference in HPV 6 and/or 11 was detected between the two groups, HPV subtypes 16 and/or 18 approached statistical significant difference (P = 0.06). CONCLUSIONS: High-risk oncogenic HPV subtypes 16 and/or 18 were found at a higher rate in transplant patients compared with their immunocompetent counterparts. The combination of immunocompromise and increased HPV 16 and/or 18 positivity may place these patients at increased risk for aggressive lower genital tract neoplastic progression.

DNA, Viral↗
PubMed2000

Screening for MEN1 tumor suppressor gene mutations in sporadic pituitary tumors.

The molecular pathogenesis of the majority of sporadic pituitary tumors is largely unknown. Pituitary adenomas can develop sporadically or as a part of multiple endocrine neoplasia type 1 (MEN1). The MEN1 is thought to be a tumor suppressor gene based on loss of heterozygosity (LOH) for polymorphic markers on 11q13 in tumors of the pancreas, parathyroid, and pituitary. Most patients with familial and sporadic MEN1 carry germ-line mutations in the MEN1 gene. Two previous studies and recently a third one have analyzed mutations by sequencing the MEN1 gene in sporadic pituitary tumors but yielded conflicting results. This study was to investigate and clarify the potential role of MEN1 mutations, in sporadic pituitary adenomas. First, we examined 59 sporadic pituitary adenomas by analyzing LOH on 11q13 in the MEN1 minimal interval with microsatellite analysis. We found 3 tumors with LOH in 1 to 4 polymorphic markers in the MEN1 region. Sequencing analysis did not reveal any mutations in the coding region of the MEN1 gene. However, we found 3 polymorphisms, one of which was a novel CAC to CAT transition encoding His433His, in exon 9. The data show that while LOH occurs in some sporadic pituitary tumors, inactivating mutations of the tumor suppressor gene MEN1 are rare. These results also suggest there may be another additional tumor suppressor gene at this locus which is involved in the pathogenesis of sporadic pituitary neoplasms.

Adenoma↗
PubMed2000

Angiogenesis in ovarian cancer.

Angiogenesis, the formation of new vessels from pre-existing vasculature, is critical to ascites development and metastasis in ovarian cancer. Many growth factors important to ovarian cancer invasion are also prominent in its associated angiogenesis. Deregulation of normal angiogenic processes occurs with the cancer's acquisition of the ability to secrete pro-angiogenic factors. The local imbalance of endogenous angio-stimulators and angio-inhibitors promotes vascularization and vascular leak. Assessment of these pro-angiogenic growth factors and enumeration of tumour-associated microvessels have been shown to be prognosticators of ovarian cancer outcome, and may also be surrogates of ovarian cancer tumour burden and/or ascites formation. The process of angiogenesis has been targeted for therapeutics development. Ovarian cancer is a primary cancer against which these new agents are being tested. Thus, further understanding of the molecular and cell biology of angiogenesis in the context of ovarian cancer offers important directions for estimation of patient outcome and for patient treatment.

Angiogenesis Inhibitors↗
PubMed2000

Multiple esophageal rings: an association with eosinophilic esophagitis: case report and review of the literature.

Esophagitis may present endoscopically with erythema, edema, loss of vascular pattern, friability, and ulceration of the esophageal mucosa. Left untreated, chronic esophagitis may result in stricture formation. The presence of multiple concentric rings involving the entire esophagus has been cited as a chronic form of esophagitis. We present a case of an 8-yr-old boy with multiple concentric esophageal rings and histological evidence of eosinophilic esophagitis, who failed medical antireflux treatment and responded to an elimination diet.

Animals↗
PubMed2000

Inductive learning approaches to rainfall-runoff modelling.

Trying to model the rainfall-runoff process is a complex activity as it is influenced by a number of implicit and explicit factors--for example, precipitation distribution, evaporation, transpiration, abstraction, watershed topography, and soil types. However, this kind of forecasting is particularly important as it is used to predict serious flooding, estimate erosion and identify problems associated with low flow. Inductive learning approaches (e.g. decision trees and artificial neural networks) are particularly well suited to problems of this nature as they can often interpret underlying factors (such as seasonal variations) which cannot be modelled by other techniques. In addition, these approaches can easily be trained on the explicit factors (e.g. rainfall) and the inexplicit factors (e.g. abstraction) that affect river flow. Inductive learning approaches can also be extended to account for new factors that emerge over a period of time. This paper evaluates the application of decision trees and two artificial neural network models (the multilayer perceptron and the radial basis function network) to river flow forecasting in two flood prone UK catchments using real hydrometric data. Comparisons are made between the performance of these approaches and conventional flood forecasting systems.

Algorithms↗
PubMed2000

Enhanced H(2)O(2)-induced cytotoxicity in "epithelioid" smooth muscle cells: implications for neointimal regression.

Vascular smooth muscle cells (SMCs) are phenotypically diverse. Although most medial SMCs can be classified as "fusiform," others are of the "epithelioid" phenotype. Proliferation and apoptosis of epithelioid SMCs may contribute importantly to neointimal formation and regression, respectively. Because reactive oxygen species (ROS) are increased in vascular injury and can induce apoptosis of SMCs, we compared the effects of ROS on epithelioid and fusiform SMCs. Epithelioid and fusiform SMC lines were clonally isolated from rat aortic media and studied under similar conditions and passage numbers. H(2)O(2) produced dose- and time-dependent cytotoxicity that was enhanced in epithelioid compared with fusiform cells. After 24-hour exposure to 50 micromol/L H(2)O(2), epithelioid cell numbers were reduced by 34+/-5% versus a 3+/-5% (P<0.05) reduction in fusiform cell numbers. Similar results were obtained whether H(2)O(2) was administered to growth-arrested or growing cells or when epithelioid and fusiform cells were exposed to extracellular O(2)(.-). To investigate whether apoptosis contributed to enhanced ROS-induced cytotoxicity in epithelioid SMCs, terminal deoxyribonucleotidyl transferase (TDT)-mediated dUTP-digoxigenin nick-end labeling (TUNEL) staining was performed. The incidence of TUNEL positivity was 5-fold increased in epithelioid versus fusiform SMCs after treatment with 50 micromol/L H(2)O(2) (19+/-1% epithelioid versus 5+/-1% fusiform, P<0.05). Enhanced H(2)O(2)-induced apoptosis in epithelioid SMCs was confirmed by DNA laddering. Furthermore, when balloon-injured aortas were exposed to H(2)O(2) ex vivo, enhanced apoptosis was observed in neointimal compared with medial SMCs. These results suggest that epithelioid SMCs exhibit enhanced sensitivity to ROS-induced apoptosis, which may play an important role in neointimal regression.

Animals↗
PubMed2000

Growth hormone deficiency type IB caused by cryptic splicing of the GH-1 gene.

We have found a novel mutation in intron 4 of the GH-1 gene in a Bedouin kindred with isolated growth hormone deficiency type IB (IGHD IB). RFLP analysis suggested linkage between the GH-1 gene and IGHD. Nested PCR amplification followed by single stranded conformation polymorphism (SSCP) analysis indicated sequence variation between introns 2 and 4. Sequencing showed a G-->C transversion at the fifth base in the splice donor region of intron 4. Affected individuals were homozygous for the mutation, which creates a new Mae III restriction site. Reverse transcription and PCR of GH-1 transcripts in EBV transformed lymphocytes indicated predominance of a species lacking 73 bp of exon 4. Amplification with a bridging primer showed that the same mRNA species is present in lymphocytes from normal individuals. The first 102 amino acids of the predicted protein are identical to wild-type GH, but the next 94 amino acids are completely divergent.

Alternative Splicing↗
PubMed1999

Overexpression of human catalase inhibits proliferation and promotes apoptosis in vascular smooth muscle cells.

The role of reactive oxygen species, such as superoxide anions (O(2). (-)) and hydrogen peroxide (H(2)O(2)), in modulating vascular smooth muscle cell proliferation and viability is controversial. To investigate the role of endogenously produced H(2)O(2), rat aortic smooth muscle cells were infected with adenoviral vectors containing cDNA for human catalase (AdCat) or a control gene, beta-galactosidase (AdLacZ). Infection with AdCat resulted in dose-dependent increases in intracellular catalase protein, which was predominantly localized to peroxisomes. After infection with 100 multiplicity of infection (MOI) of AdCat, cellular catalase activity was increased by 50- to 100-fold, and intracellular H(2)O(2) concentration was reduced, as compared with control. Infection with AdCat reduced [(3)H]thymidine uptake, an index of DNA synthesis, in cells maintained in medium supplemented with 2% serum (0.37+/-0.09 disintegrations per minute per cell [AdLacZ] versus 0.22+/-0.08 disintegrations per minute per cell [AdCat], P<0.05). Five days after infection with 100 MOI of AdCat, cell numbers were reduced as compared with noninfected or AdLacZ-infected cells (157 780+/-8413 [AdCat], P<0.05 versus 233 700+/-3032 [noninfected] or 222 410+/-5332 [AdLacZ]). Furthermore, the number of apoptotic cells was increased 5-fold after infection with 100 MOI of AdCat as compared with control. Infection with AdCat resulted in induction of cyclooxygenase (COX)-2, and treatment with a COX-2 inhibitor overcame the AdCat-induced reduction in cell numbers. These findings indicate that overexpression of catalase inhibited smooth muscle proliferation while increasing the rate of apoptosis, possibly through a COX-2-dependent mechanism. Our results suggest that endogenously produced H(2)O(2) importantly modulates survival and proliferation of vascular smooth muscle cells.

Animals↗
PubMed1999

Detection of intracellular phosphorylated STAT-1 by flow cytometry.

We have applied flow cytometry to the investigation of interferon-gamma activation of human monocytes. This approach uses monoclonal antibodies that distinguish between the native and phosphorylated forms of STAT-1. It enables rapid and quantitative assessment of STAT-1 phosphorylation on a discrete cell basis and is both more sensitive and less time consuming than immunoblotting. Furthermore, it allows for discrimination between a mixture of cells that differ in their response to interferon-gamma. This approach should allow for the evaluation of different intracellular signaling pathways using a combination of monoclonal reagents that are specific for native and activation modified proteins. Application of this form of testing should prove valuable in screening for signaling defects in selected patients with recurrent infections. In addition, this technique should permit dissection of a full range of cellular signaling pathways at the protein level.

DNA-Binding Proteins↗
PubMed1999

IL-12-Independent costimulation pathways for interferon-gamma production in familial disseminated Mycobacterium avium complex infection.

We have described previously a family with an apparent genetic susceptibility to disseminated Mycobacterium avium complex infection and an underlying defect in IL-12 regulation leading to abnormally low interferon-gamma production. Their T cells appear to act normally when in the presence of normal accessory cells. Cell-to-cell contact was necessary for normal monocytes to complement the familial patient monocyte defect, suggesting the familial defect in interferon-gamma costimulation involves pathways requiring cell surface molecule interactions. In an effort to better characterize the abnormality in these patients, we examined the role of known costimulatory molecules in residual costimulation by patient PBMC compared to normals. Whereas normals utilized CD40/CD40L interactions and IL-12 production for optimal interferon-gamma costimulation in PHA-stimulated cocultures, familial patient interferon-gamma production was low and unaffected by their blockade. CD86 blockade caused a greater than 50% reduction in both normal and familial patient interferon-gamma production, implying that a majority of residual familial patient costimulation required this pathway. Furthermore, selected myelomonocytic cell lines (K562 and THP1) acted as potent accessory cells for interferon-gamma production by familial patient and normal T cells, largely independent of IL-12 production. However, CD86 blockade of K562 cell/familial cell cocultures resulted in less than a 20% reduction in interferon-gamma production, indicating that familial patient cells respond to IL-12- and CD86-independent costimulatory signals for interferon-gamma as well. Thus, we demonstrate that the familial defect also involves interferon-gamma costimulation pathways requiring both CD40/CD40L interaction and IL-12 production, while residual pathways remain that allow low-level interferon-gamma production. Familial Mycobacterium avium patient monocytes and certain myelomonocytic cell lines can be exploited to investigate IL-12-independent costimulation for interferon-gamma production.

Antigens, CD↗
PubMed1999

Allelic loss on chromosome arm 8p: analysis of sporadic epithelial ovarian tumors.

OBJECTIVE: Our objective was to determine the frequency of allelic loss at 8p21 in sporadic epithelial ovarian cancer. We recently described allelic loss at this locus in 7/9 ovarian cancers from patients with BRCA1 gene mutations. METHODS: We anonymously obtained and examined 40 unselected invasive epithelial ovarian cancers and 5 low-malignant-potential (LMP) ovarian tumors for loss of heterozygosity (LOH) at 8p12-22. Pure epithelial and stromal cell populations were procured selectively by laser capture microdissection and extracted DNA was amplified with polymorphic microsatellite markers spanning the region of interest. RESULTS: LOH was highest (50%) at marker D8S136 located at 8p21 with 15 of 30 informative cases exhibiting an allelic deletion. None of the LMP tumors evaluated showed LOH at 8p12-22. A trend toward more frequent LOH at 8p12-22 was identified with increasing disease aggressiveness from LMP to early stage invasive ovarian cancer to advanced stage invasive ovarian cancer (Lehman's test, P2 < 0.024). CONCLUSIONS: Fifty percent allelic loss at the distal portion of 8p21 has not been reported to date for sporadic epithelial ovarian carcinomas. The higher rate of loss in our cohort, in contrast to previous allelotyping studies, is due likely to analysis from homogenous cell populations. These results, in concert with our previous study of BRCA1 mutation-positive patients, suggest a tumor suppressor gene locus at 8p21 for epithelial ovarian cancer.

Adult↗
PubMed1999

Expression of a glycosylphosphatidylinositol-linked Manduca sexta aminopeptidase N in insect cells.

Aminopeptidase N (APN; EC 3.4.11.2) is an exopeptidase that is attached to cell membranes by a hydrophobic amino-terminal stalk in vertebrates or a glycosylphosphatidylinositol (GPI) anchor in insects. In this study, we report the cloning, expression, and characterization of an aminopeptidase N from Manduca sexta midgut. The full-length aminopeptidase N cDNA (APN1a) encodes a 995-amino-acid protein. The predicted amino acid sequence differs by 8 amino acids from M. sexta APN1. These different amino acids do not modify any putative glycosylation or glycosylphosphatidylinositol anchor sites. The full-length cDNA was cloned into an expression plasmid, pHSP-HR5, and transiently expressed in an insect cell line derived from Spodoptera frugiperda (Sf21 cells). Immunoblot analysis with anti-APN antiserum showed that APN1a expressed in Sf21 cells is the same size (120 kDa) as APN found in midgut brush border membranes. After treatment with phosphatidylinositol-specific phospholipase C (PIPLC), anti-cross-reacting determinant antibody specific for PIPLC cleavage products recognized the expressed 120-kDa APN1a, but not endogenous Sf21 proteins, indicating that APN1a has an intact glycosylphosphatidylinositol anchor. These results are evidence that Sf21 cells synthesize few, if any, endogenous GPI-linked proteins. Immunofluorescence staining showed that the expressed APN1a was located on the surface of Sf21 cells.

Amino Acid Sequence↗
PubMed1999

Identification of a Drosophila brain-gut peptide related to the neuropeptide Y family.

A neuropeptide F (NPF) was isolated from the fruit fly, Drosophila mellanogaster, based on a radioimmunoassay for a gut peptide from the corn earworm, Helicoverpa zea. A partial sequence was obtained from the fly peptide, and a genomic sequence coding for NPF was cloned after inverse polymerase chain reaction and shown to exist as a single genomic copy. The encoded, putative prepropeptide can be processed into an amidated NPF with 36 residues that is related to invertebrate NPF's and the neuropeptide Y family of vertebrates. In situ hybridization and immunocytochemistry showed that Drosophila NPF was expressed in the brain and midgut of fly larvae and adults.

Amino Acid Sequence↗
PubMed1999

Insulin stimulates ecdysteroid production through a conserved signaling cascade in the mosquito Aedes aegypti.

Selective activators and inhibitors of insulin signaling cascades in mammalian cells were tested for their effects on insulin stimulated steroidogenesis by ovaries of Aedes aegypti. Bovine insulin in the concentration range of 1.7 microM to 85 microM stimulated ecdysteroidogenesis in vitro. Pervanadate, an inhibitor of tyrosine kinase phosphatase, stimulated ecdysteroid production at concentrations of 250 microM to 1 microM. Okidaic acid, a serine/threonine phosphatase inhibitor, stimulated steroidogenesis with an ED50 of 77.39 nM. A selective inhibitor of tyrosine kinase activity, HNMPA-(AM3), inhibited ecdysteroid production with an IC50 of 14.2 microM. Two selective inhibitors of phosphatidylinositol 3-kinase, wortmannin and LY294002, inhibited ecdysteroid production at low concentrations (IC50 = 1.6 nM and 30 nM, respectively). These concentrations are similar to those inhibiting insulin action in mammalian cells. A selective inhibitor of mitogen-activated protein kinase, PD098059, had no effect on ecdysteroid production even up to 100 microM. Thus, insulin stimulation of ecdysteroid production by ovaries in vitro appears to be controlled by the tyrosine kinase activity of the mosquito insulin receptor and the signaling cascade involving phosphatidylinositol 3-kinase and protein kinase B.

Aedes↗
PubMed1999

Unexplored reservoirs of pathogenic bacteria: protozoa and biofilms.

In the natural, industrial, hospital and domestic environments, there are numerous phenotypes of pathogenic microorganisms, which vary considerably in chemical, physical and biological properties. A link exists between survival, resistance and virulence. In particular, surface-adherent biofilms and bacteria living within protozoa pose potential health problems that are unrecognized by conventional laboratory culture methods.

Animals↗