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Biomedical subjects

D Lew

Publications and source records attributed to D Lew.

At least 19 recordsLinked to original sources
PubMed1995

An N-terminal deletion mutant of estrogen receptor exhibits increased synergism with upstream activators and enhanced binding to the estrogen response element.

To study the role of the N-terminal region of the estrogen receptor (ER) in transcription activation and in DNA binding, we constructed a mutant of the Xenopus laevis ER which lacks amino acids 1-159 (XER160/586). In transient transfections, XER160/586 exhibited < 10% of the activity of wild-type XER on a synthetic promoter containing two estrogen response elements (EREs). To examine transcriptional synergism by XER and by XER160/586, we determined the activity of promoters containing EREs and binding sites for either the vitellogenin activator, NF1, or AP1 upstream activator protein. For the three promoters transcription by XER was 2.8-fold greater than expected for additive activities, and transcription by XER160/586 was 6.2-fold greater. These data demonstrate that an upstream activator protein bound near the promoter can partially compensate for the loss of the internal N-terminal (AF1) transactivation domain in XER160/586. Using a promoter interference assay to study the intracellular interaction between ER and the estrogen response element, we found that XER160/586 exhibited a significant increase in affinity for the ERE. Its low basal activity and enhanced affinity for the ERE make XER160/586 an effective dominant negative mutant. When co-expressed with wild-type XER at 1:1 and 5:1 ratios, XER160/586 suppressed the activity of wild-type XER by 57% and > 80%, respectively.

Animals↗
PubMed1995

Rhytidectomy: use of the short flap.

PURPOSE: This study compares the use of the short and long flap rhytidectomy. PATIENTS AND METHODS: Thirty short flap rhytidectomies and 20 long flap rhytidectomies were compared for indications, operating time, complications, and patient satisfaction. Flaps 6 cm or less in length were considered short flaps. RESULTS: The short flap technique was used more frequently on younger female patients. The use of this technique resulted in fewer complications and shortened operating and anesthesia times. Patient satisfaction was the same with both procedures. CONCLUSION: The short flap technique is appropriate for the younger patient with minimal nasolabial folds, jowls, rhytids, actinic damage, and less extensive signs of aging. When this technique is used on a suitable patient, the results are cosmetically gratifying.

Adult↗
PubMed1995

Cancer gene therapy using plasmid DNA: pharmacokinetic study of DNA following injection in mice.

The fate of plasmid DNA complexed with cationic lipids delivered intravenously in mice was evaluated at selected timepoints up to 6 months postinjection. Blood half-life and tissue distribution of plasmid DNA and potential expression in tissues were examined. Southern blot analyses of blood indicated that intact plasmid DNA was rapidly degraded, with a half-life of less than 5 min for intact plasmid, and was no longer detectable at 1 hr postinjection. Southern analyses of tissue demonstrated that intact DNA was differentially retained in the lung, spleen, liver, heart, kidney, marrow, and muscle up to 24 hr postinjection. After 7 days, no intact plasmid DNA was detectable by Southern blot analysis; however, the plasmid was detectable by the polymerase chain reaction (PCR) in all tissues examined at 7 and 28 days postinjection. At 6 months postinjection, femtogram levels of plasmid were detected only in muscle. Immunohistochemical analyses did not detect encoded protein in the tissues harboring residual plasmid at 1 or 7 days postinjection.

Animals↗
PubMed1995

Cancer gene therapy using plasmid DNA: safety evaluation in rodents and non-human primates.

To evaluate the safety of a plasmid DNA-lipid complex, a series of good laboratory practice (GLP) safety studies were conducted with VCL-1005, a plasmid DNA expression vector containing both the human class I MHC HLA-B7 heavy-chain and the beta 2-microglobulin (beta 2m) light-chain genes formulated with the cationic lipid, DMRIE/DOPE. In mice, the repeated intravenous injection of VCL-1005 at plasmid DNA doses of 0.1, 1.0, or 10 micrograms for 14 days had only incidental effects on clinical chemistry and hematology, and did not result in any organ pathology. Repeated intrahepatic injections of VCL-1005 in mice did not result in significant liver histopathology or significant alterations in liver enzymes. In cynomolgus monkeys, the repeated intravenous administration of VCL-1005 at a cumulative dose of 720 micrograms of DNA had no effects on clinical chemistry, hematology, or organ pathology. Thus, systemic administration of a plasmid DNA expression vector containing the coding sequence for a foreign MHC class I molecule did not result in significant toxicity or a pathological immune response in animals. These results suggest that the direct transfer of VCL-1005, a plasmid DNA-lipid complex, could be used for the safe in vivo delivery of recombinant DNA for a cancer gene therapy trial.

Animals↗
PubMed1994

Intradermal gene immunization: the possible role of DNA uptake in the induction of cellular immunity to viruses.

The skin and mucous membranes are the anatomical sites were most viruses are first encountered by the immune system. Previous experiments have suggested that striated muscle cells are unique among mammalian cell types in their capacity to take up and express free DNA in the absence of a viral vector or physical carrier. However, we have found that mice injected into the superficial skin with free (naked) plasmid DNA encoding the influenza nucleoprotein gene had discrete foci of epidermal and dermal cells, including cells with dendritic morphology, that contained immunoreactive nucleoprotein antigen. A single intradermal administration of 0.3-15 micrograms of free plasmid DNA induced anti-nucleoprotein-specific antibody and cytotoxic T lymphocytes that persisted for at least 68-70 weeks after vaccination. Intradermal gene administration induced higher antibody titers than did direct gene injection into skeletal muscle and did not cause local inflammation or necrosis. Compared with control animals, the gene-injected mice were resistant to challenge with a heterologous strain of influenza virus. These results indicate that the cells of the skin can take up and express free foreign DNA and induce cellular and humoral immune responses against the encoded protein. We suggest that DNA uptake by the skin-associated lymphoid tissues may play a role in the induction of cytotoxic T cells against viruses and other intracellular pathogens.

Amino Acid Sequence↗
PubMed1994

Aqueous humor penetration of ofloxacin given by various routes.

We studied the aqueous humor penetration of ofloxacin after topical, oral, and intravenous administration in 51 consecutive patients undergoing cataract surgery. Aqueous humor concentration (mean +/- SD) was 0.53 +/- 0.35 mg/l when ofloxacin 0.3% eyedrops were instilled topically six times, one drop every three hours, until 90 minutes preoperatively, and 0.63 +/- 0.29 mg/l (P = .45) when two additional instillations were made, one drop every 30 minutes, until 30 minutes before aqueous humor aspiration. Aqueous humor concentration two hours after a single 200-mg oral dose (0.38 +/- 0.15 mg/l) was significantly lower (P = .048) than that 12 hours after the same oral dose (0.58 +/- 0.24 mg/l). Two hours following an intravenous infusion of 200 mg of ofloxacin, aqueous humor concentration was 0.33 +/- 0.19 mg/l. Our results suggest that therapeutic levels above the minimum inhibitory concentration for many bacteria cultured in endophthalmitis can be achieved in aqueous humor after either topical or oral administration, which indicates that this antibiotic passes easily through the corneal and the blood aqueous barriers.

Administration, Oral↗
PubMed1994

Case report 860: Bacillary angiomatosis of the calcaneum.

Bacillary angiomatosis (BA) is newly reported infectious disease observed mainly in HIV-infected patients, caused by a small gram-negative bacillus of the Rochalimea genus. From a purely dermatological presentation similar to that of Kaposi's sarcoma, it may evolve into a systemic disease. Bone lesions seem fairly frequent. We report a case of an isolated osteolytic lesion due to the BA bacillus.

AIDS-Related Opportunistic Infections↗
PubMed1994

A comparative study of osseointegration of titanium implants in corticocancellous block and corticocancellous chip grafts in canine ilium.

PURPOSE: This study was undertaken to compare the relative rates and extent of osseointegration of dental implants when placed simultaneously with either corticocancellous block or particulate corticocancellous bone grafts. MATERIALS AND METHODS: Using the canine ilium as a model site, the implants were placed so that each served as its own control. The implants were harvested at 1, 2, or 3 months for evaluation by light microscopy, microradiography, and histomorphometry. RESULTS: Both types of grafts were determined to be viable by microscopic evaluation of fluorescent labels. Qualitatively there appeared to be greater bone density in the corticocancellous block graft implant sites. At 3 months, the block graft implant sites had a level of osseointegration (59.6%) that approximated the control implant sites (65.2%), but was significantly greater than the particulate graft sites (39.2%). CONCLUSIONS: These results indicate that implants in corticocancellous block grafts develop osseointegration more rapidly than those in particulate bone grafts. The clinical implications of these findings are discussed.

Animals↗
PubMed1994

Prospective randomized comparison of imipenem monotherapy with imipenem plus netilmicin for treatment of severe infections in nonneutropenic patients.

Nosocomial pneumonia and sepsis, as well as severe diffuse peritonitis, must be treated early in order to prevent complications such as septic shock and organ dysfunctions. With the availability of new broad-spectrum and highly bactericidal antibiotics, the need of combining beta-lactams with aminoglycosides for the treatment of severe infections should be reassessed. A prospective randomized controlled study was performed to compare imipenem monotherapy with a combination of imipenem plus netilmicin in the empiric treatment of nosocomial pneumonia, nosocomial sepsis, and severe diffuse peritonitis. A total of 313 patients were enrolled, and 280 were assessable. The antibiotic treatment was successful in 113 of 142 patients (80%) given the monotherapy and in 119 of 138 patients (86%) given the combination (P = 0.19). The failure rates for the most important type of infection, i.e., pneumonia, were similar in the two groups, as well as the number of superinfections. While creatinine increase was associated with factors not related to antibiotic therapy for all eight patients of the monotherapy group, no factor other than the antibiotics could be found for 6 of the 14 cases of nephrotoxicity observed in the combination group (P = 0.014). Finally, the emergence of Pseudomonas aeruginosa resistant to imipenem occurred in 8 monotherapy patients and in 13 combination therapy patients. In conclusion, imipenem monotherapy appeared as effective as the combination of imipenem plus netilmicin for the treatment of severe infection. The addition of netilmicin increased nephrotoxicity, and it did not prevent the emergence of P. aeruginosa resistant to imipenem.

Adult↗
PubMed1994

A classification of one-dimensional electrophoresis gels using wave packet decomposition.

A classification of one-dimensional electrophoresis gels was achieved using a fast approximation of the Karhunen-Loève transformation. The algorithm was based on wave packet decomposition theory, using in particular quadrature mirror filters derived from orthogonal bases of wavelets. Bacterial proteins from two different species of staphylococci were separated by SDS-PAGE electrophoresis. The gel images were analyzed by computer and the protein bands were clustered, after an orthogonal projection onto a space of lower dimension, by standard statistical methods.

Algorithms↗
PubMed1994

Surgical planning using three-dimensional imaging and computer modeling.

The acquisition, processing, and use of three-dimensional (3D) imaging provide new insights into normal and abnormal craniofacial anatomy. In this article, characteristics of CT and MRI scanning are reviewed along with the methods used to delineate tissues and produce 3D patient displays, including contouring, shaded surface, and volumetric processing. The use of 3D model fabrication for surgical planning is discussed with examples of the production of a custom CT-based auricle model and the use of rapid prototyping technology to create models for custom alloplastic implant surgery. The review concludes with some speculation about the future of 3D medical imaging as it will influence surgical training and practice over the coming decades.

Diagnostic Imaging↗
PubMed1993

The effect of pulsed expansion of subfascially placed expanders on the extent and duration of mitosis in the capsule and rat integument.

This study investigated the mitotic activity in the subdermally induced capsule and the epithelium surrounding tissue expanders maintained by pulsed expansions. Tissue expanders were placed into the fascial plane beneath the panniculus muscle of the rat and expanded sequentially. The volume in the expander was rapidly increased to a constant level and maintained for 48 hours; this was followed by a total decrease of volume for 48 hours. The procedure was repeated with volumes of 20, 30, and 40 mL. At the conclusion of each cycle of expander inflation, a group of rats was killed, and biopsy specimens of the induced capsule and epithelium were obtained. The developing capsule exhibited a significant increase in the mitotic index in the layers adjacent to the expander. The highest mitotic index was obtained with an inflation of 30 mL. These observations suggest that cellular proliferation, growth, and development of the extracellular matrix in the induced subdermal capsule can be sustained by pulsed volume changes in the expander.

Animals↗
PubMed1993

GHF-1-promoter-targeted immortalization of a somatotropic progenitor cell results in dwarfism in transgenic mice.

During pituitary development, the homeo domain protein GHF-1 is required for generation of somatotropes and lactotropes and for growth hormone (GH) and prolactin (PRL) gene expression. GHF-1 mRNA is detectable several days before the emergence of GH- or PRL-expressing cells, suggesting the existence of a somatotropic progenitor cell in which GHF-1 transcription is first activated. We have immortalized this cell type by using the GHF-1 regulatory region to target SV40 T-antigen (Tag) tumorigenesis in transgenic mice. The GHF-Tag transgene caused developmental entrapment of somatotropic progenitor cells that express GHF-1 but not GH or PRL, resulting in dwarfism. Immortalized cell lines derived from a transgenic pituitary tumor maintain the characteristics of the somato/lactotropic progenitor in that they express GHF-1 mRNA and protein yet fail to activate GH or PRL transcription. Using these cells, we identified an enhancer that activates GHF-1 transcription at this early stage of development yet is inactive in cells representing later developmental stages of the somatotropic lineage or in other cell types. These experiments not only demonstrate the potential for immortalization of developmental progenitor cells using the regulatory regions from cell type-specific transcription factor genes but illustrate the power of such model systems in the study of developmental control.

Animals↗
PubMed1992

The role of estrogen response elements in expression of the Xenopus laevis vitellogenin B1 gene.

We have used site-directed mutagenesis and a homologous transient transfection system to investigate the role of the two imperfect estrogen response elements (EREs) located at -302/-334 in the 5'-flanking region of the estrogen-regulated Xenopus laevis vitellogenin B1 gene. Deletion of either ERE effectively abolishes estrogen-dependent transcription of the vitellogenin promoter. Neither replacement of the two imperfect EREs with a single consensus ERE at -334, nor insertion of one or two consensus EREs at -359, restores full estrogen responsiveness to the mutant promoter. In competition gel mobility shift assays using the DNA binding domain of the Xenopus estrogen receptor, the consensus ERE was a severalfold more effective competitor than the two imperfect B1 EREs. These data suggest that flanking DNA sequences may exert a significant effect on the activity of EREs as hormone-dependent transcription activators. When the imperfect EREs at -302/-334 were present, an additional consensus ERE at -359 exhibited synergistic activation of transcription. However, two consensus EREs located close to the TATA box showed additive, not synergistic, activation of transcription. In contrast, synergistic activation of transcription was observed in synthetic promoters containing two EREs and either the vitellogenin activator element or the NF1 or AP1 upstream activator elements.

Animals↗
PubMed1992

N-acetylcysteine enhances receptor-mediated phagocytosis by human neutrophils.

The effect of the sulphur compound N-acetylcysteine (NAC) on certain receptor-mediated cellular functions [chemiluminescence (CL), phagocytosis and degranulation] in human neutrophils was studied, to evaluate how a scavenger of certain toxic oxygen product can protect the phagocyte and the bystander tissue cells from oxidative damage. When using IgG-opsonized yeast particles as stimulating agent, preincubating the neutrophils with NAC (0.25 mg/ml = 1.5 mM) increased both the CL response and phagocytosis. Higher concentrations of NAC (0.50-1.00 mg/ml = 3-6 mM) decreased the CL response, whereas the phagocytic capacity was still enhanced. This effect was more pronounced with adherent neutrophils than with neutrophils in suspension. No increased CL or phagocytic activity was, however, induced by NAC when C3bi-opsonized particles were used as a prey. From the fact that NAC (i) inhibited extracellularly localized myeloperoxidase dependent activities, and (ii) had no effect on neutrophils from patients with chronic granulomatous disease (CGD), we conclude that the scavenger effect of NAC not only reduces the accumulation of oxidative metabolites per se, but also enhances receptor-mediated phagocytosis by protecting Fc(IgG)-receptors from oxidative damage mediated by myeloperoxidase (MPO) and hydrogen peroxide (H2O2). Since NAC can increase phagocytosis and reduce the extracellularly produced oxidative metabolites, we furthermore conclude that NAC possesses some ideal properties as an anti-inflammatory agent.

Acetylcysteine↗
PubMed1991

Purified estrogen receptor DNA binding domain expressed in Escherichia coli activates transcription of an estrogen-responsive promoter in cultured cells.

The region of the Xenopus laevis estrogen receptor responsible for interaction with DNA, the DNA binding domain (DBD), has been cloned and overexpressed in Escherichia coli using a T7 RNA polymerase expression system. Extracts from cells transformed with the DBD expression vector contain a single protein which reacts with polyclonal antibodies to estrogen receptor and exhibits sequence-specific binding to a DNA fragment containing a consensus estrogen response element. The DBD protein has been purified to near homogeneity. Determination of the rotational relaxation time of the dansylated DBD by fluorescence polarization and size fractionation by Superdex column chromatography indicate that the DBD is a monomer in solution. The DBD forms a single protein-estrogen response element complex in gel mobility shift assays at DBD concentrations of 18-3,600 nM, suggesting that the DBD is bound to both halves of the palindromic estrogen response element. To investigate the ability of the DBD expressed in bacteria to activate gene expression, we have developed a simple liposome-based system for delivery of protein into cultured cells. Transfected DBD protein elicited large, concentration-dependent increases in transcription of an estrogen receptor regulated reporter gene. These data demonstrate that the bacterially expressed DNA binding domain, which represents a small portion of the Xenopus laevis estrogen receptor, retains significant ability to activate transcription of an estrogen-responsive promoter in vertebrate cells.

Animals↗