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Biomedical subjects

D L Miller

Publications and source records attributed to D L Miller.

At least 145 records · Page 8Linked to original sources
PubMed1997

A new method for straightening DNA molecules for optical restriction mapping.

We have developed an improved method of straightening DNA molecules for use in optical restriction mapping. The DNA was straightened on 3-aminopropyltriethoxysilane-coated glass slides using surface tension generated by a moving meniscus. In our method the meniscus motion was controlled mechanically, which provides advantages of speed and uniformity of the straightened molecules. Variation in the affinity of the silanized surfaces for DNA was compensated by precoating the slide with single-stranded non-target blocking DNA. A small amount of MgCl2 added to the DNA suspension increased the DNA-surface affinity and was necessary for efficient restriction enzyme digestion of the straightened surface-bound DNA. By adjusting the amounts of blocking DNA and MgCl2, we prepared slides that contained many straight parallel DNA molecules. Straightened lambda phage DNA (48 kb) bound to a slide surface was digested by EcoRI restriction endonuclease, and the resulting restriction fragments were imaged by fluorescence microscopy using a CCD camera. The observed fragment lengths showed excellent agreement with their predicted lengths.

Bacteriophage lambda↗
PubMed1997

Association of human papillomavirus type 16 integration in the E2 gene with poor disease-free survival from cervical cancer.

To determine the clinical relevance of human papillomavirus (HPV) integration and E2 function suggested by in vitro studies, we investigated 50 patients with HPV 16-positive primary cervical carcinoma (stage Ib-IV) diagnosed and treated at one institution. The physical state of HPV was determined by colorimetric in situ hybridization and was not found to vary by stage. Overall, 62% of tumors had integrated HPV, 16% had episomal and 22% had both integrated and episomal. The E1/E2 region was evaluated by 8 separate polymerase chain reactions, which resulted in overlapping products. There was no significant variation in ability to amplify the E1/E2 region with stage. E1/E2 amplification correlated with physical state. Nearly all tumors with episomal or mixed HPV 16 DNA amplified all 8 E1/E2 fragments. Half of the tumors with integrated HPV 16 DNA failed to amplify one or more E1/E2 fragments. Disruptions were most frequent in the E2 region. For all 46 patients receiving curative therapy, the Kaplan-Meier estimate of disease-free survival was determined for those whose primary tumors had amplifiable E2 compared with those lacking one or more E2 DNA fragments. Disruption of E2 was associated with significantly shortened disease-free survival.

Adenocarcinoma↗
PubMed1997

Transfection of a reporter plasmid into cultured cells by sonoporation in vitro.

Cultured Chinese hamster ovary cells were exposed to 2.25-MHz ultrasound in sterile 4.5-mL polyethylene chambers and tested for cell lysis, sonoporation and DNA transfection. Ten percent of Albunex, a gas-body-based ultrasound contrast agent, was added to ensure cavitation nucleation, and the chambers were rotated at 60 rpm to promote cavitation activity during the 1-min exposures. Uptake of large fluorescent dextran molecules by some cells was observed for spatial peak pressure amplitudes as low as 0.1 MPa, which indicates transient permeabilization and resealing, i.e., sonoporation, of these cells during exposure. Significant lysis occurred for 0.2 MPa, and increased rapidly for exposures above the apparent cavitation threshold (using the H2O2 production test) of about 0.4 MPa spatial peak pressure amplitude. In the DNA transfection tests, 20 micrograms/mL luciferase reporter plasmid was added to the suspension during exposure, and cells were assayed for proliferation ability and luciferase gene expression 2 days after exposure. Cell proliferation was greatly reduced above the cavitation threshold. Luciferase production was significant for 0.20-MPa exposure, and reached 0.33 ng per 10(6) cells at 0.8-MPa exposure. The luciferase production was great for cells exposed in medium supplemented with serum than for cells exposed in serum-free medium. Cells harvested for exposure either in the log phase or in the stationary phase of culture gave similar proliferation and transfection results. The effects essentially disappeared when the Albunex was omitted from the suspension and the tube was not rotated. Thus, sonoporation by ultrasonic cavitation in the rotating tube system yields plasmid transfection with subsequent transient gene expression.

Animals↗
PubMed1997

Ultrasonically induced hemolysis at high cell and gas body concentrations in a thin-disc exposure chamber.

Ultrasound image contrast may be enhanced by injecting gas bodies into the blood. This in vitro study was undertaken to assess the potential for induction of hemolysis due to ultrasonic activation of the contrast agent gas bodies. Canine whole blood with Albunex (Mallinckrodt Medical, St. Louis, MO, USA) was exposed to near-field ultrasound beams in 1-mm-thick chambers held stationary (i.e., not rotated) in a 37 degrees C water bath. At 2.25 MHz, statistically significant hemolysis occurred in 0.5 hematocrit, 50% Albunex suspensions for 0.28-MPa, 1-s continuous exposure and for 0.58-MPa, 100-s exposures with 10-microsecond pulses and 1.0-ms pulse repetition period. Continuous exposure durations as short as 10 ms produced about 4.5% hemolysis, which only increased slightly to about 5.5% after 100 s. At a constant 1.6 MPa, hemolysis increased with increasing gas body concentration and with decreasing cell concentration. Hemolysis decreased with increasing frequency in a 50/50 mixture of whole blood and Albunex, with thresholds rising from 0.12 MPa continuous (1 s) and 0.47 MPa pulsed (10 microseconds:1.0 ms for 100 s) at 1.06 MHz to 0.47 MPa continuous and 1.9 MPa pulsed at 5.3 MHz.

Albumins↗
PubMed1997

Sensory properties of a nonabsorbable fat substitute did not affect regulation of energy intake.

Many reduced-fat foods retain the sensory properties of their high-fat counterparts through the use of fat substitutes. This study examined whether regulation of energy intake is affected when the nonabsorbable fat substitute olestra is used to uncouple the sensory properties of fat from fat absorption and metabolism. Cream of broccoli soups were developed in three versions: fat-free, fat-free+olestra (33.3 g olestra), and high-fat (33.3 g fat) (923900 and 2150 kJ per serving, respectively). The olestra soup had the nutrient composition of the fat-free soup but the sensory properties of the high-fat soup. Subjects were grouped by sex, body weight, and dietary restraint (total n = 67). Subjects had either no preload (control) or a soup preload (465 g) followed by a self-selection lunch. Intake was measured at lunch, dinner, snack, and breakfast. At lunch, the response to the soup preloads was not affected by sex, dietary restraint, or body weight. Energy intake (soup+lunch) was significantly greater in the high-fat than in the control condition (P < 0.05), but energy intake in the fat-free and olestra-soup conditions was not significantly different from that in the control condition (3570, 3352, 3464, and 4457 kJ in control, fat-free, olestra, and high-fat soup conditions, respectively). Thus, subjects compensated completely for the energy in the fat-free and olestra soups but not for the energy in the high-fat soup. No differences were found in the response to the two fat-free conditions, one with the fatty taste and one without. In this study the sensory properties of fat alone, ie, apart from the physiologic effects of fat, did not affect energy regulation.

Absorption↗
PubMed1997

Comparison of cardiac and 60 Hz magnetically induced electric fields measured in anesthetized rats.

Extremely low frequency magnetic fields interact with an animal by inducing internal electric fields, which are in addition to the normal endogenous fields present in living animals. Male rats weighing about 560 g each were anesthetized with ketamine and xylazine. Small incisions were made in the ventral body wall at the chest and upper abdomen to position a miniature probe for measuring internal electric fields. The calibration constant for the probe size was 5.7 mm, with a flat response from at least 12 Hz to 20 kHz. A cardiac signal, similar to the normal electrocardiogram with a heart rate of about 250 bpm, was readily obtained at the chest. Upon analysis of its spectrum, the cardiac field detected by the probe had a broad maximum at 32-95 Hz. When the rats were exposed to a 1 mT, 60 Hz magnetic field, a spike appeared in the spectrum at 60 Hz. The peak-to-peak magnitudes of electric fields associated with normal heart function were comparable to fields induced by a 1 mT magnetic field at 60 Hz for those positions measured on the body surface (where induced fields were maximal). Within the body, or in different directions relative to the applied field, the induced fields were reduced (reaching zero at the center of the animal). The cardiac field increased near the heart, becoming much larger than the induced field. Thus, the cardiac electric field, together with the other endogenous fields, combine with induced electric fields and help to provide reference levels for the induced-field dosimetry of ELF magnetic field exposures of living animals.

Anesthesia↗
PubMed1997

Human papillomavirus type in anal epithelial lesions is influenced by human immunodeficiency virus.

BACKGROUND AND OBJECTIVE: Infection with human immunodeficiency virus (HIV) increases the risk for human papillomavirus (HPV)-associated genital neoplasia. Human immunodeficiency virus-infected patients also have higher rates of treatment failure and more rapid neoplastic progression. Impaired immune function does not entirely explain these clinical observations. This pilot project was designed to investigate the hypothesis that HIV infection is associated with changes in HPV type and integration within anogenital lesions that could explain the increased risk of neoplastic progression. METHODS: Anal neoplastic lesions from patients with and without HIV infection were analyzed for the presence, type, and integration status of HPV by colorimetric in situ hybridization. Tissue localization of HIV was evaluated by p24 immunohistochemistry and HIV-1 DNA polymerase chain reaction. Results for matched histology were compared for the two patient groups. RESULTS: For all lesions, the presence of high-risk HPV types and multiple HPV types was strongly associated with HIV infection (P = .003 and .0003, respectively). For lesions with matched histology there was no association of HPV integration with HIV status. Tissue localization of HIV did not significantly influence HPV type or integration. CONCLUSIONS: The presence of high-risk HPV types and multiple types within low-grade lesions may explain the increased risk of neoplastic progression in HIV patients. Colocalization of HIV and HPV does not appear to be required for this effect. There is no evidence that HPV integration is influenced by HIV infection.

Adolescent↗
PubMed1997

Is the low-fat message giving people a license to eat more?

In the past decade, technology for modifying the fat content of foods has progressed and led to the development of highly palatable low-fat foods. Many Americans believe that weight loss can be accomplished by eating an ad libitum low-fat diet, and the development of highly palatable foods that are low in fat provides welcome additions to the food supply. However, in order to maintain high palatability, some of these foods have added sugars and other energy-bearing substances resulting in some low-fat foods which remain high in energy content and are energy-dense. Using such products as substitutes for higher-fat versions of foods is likely to bring about only small reductions in energy intake. A better strategy to reduce energy intake is to consume a low-fat diet made up of low-energy density foods such as fruits, vegetables, and other foods high in fiber and complex carbohydrates. Such a diet may include fat-replaced foods which are also reduced in energy since, for weight loss, a sustained reduction in energy intake is necessary. This can be best accomplished by eating a balanced diet not only low in fat content but also lower in energy.

Diet, Fat-Restricted↗
PubMed1997

Effects of toothbrush design and brushing proficiency on plaque removal.

A single-blind, short-term cross-over clinical trial compared the plaque removal performance of three commercially available manual toothbrushes. A sample of 25 dental hygiene students, 19 to 42 years old, served as participants. On 3 separate occasions, participants were instructed to refrain from toothbrushing or flossing for 24 hours before clinical trials. A prebrushing plaque index using disclosing solution was performed on each participant. One of the 3 test brushes was then randomly dispensed to each participant, and they were allowed to brush for 90 seconds without the aid of a mirror. A postbrushing plaque index was then performed on each participant. This procedure was repeated 2 more times at 2-week intervals so that each participant was tested with all 3 toothbrushes. Previous studies of this kind using random participant samples have suggested that brush design does indeed affect the efficacy of plaque removal. This study used participants who were well versed on efficient toothbrushing technique to determine if improved brushing skills would overshadow advantages in toothbrush design. No significant differences in performance were detected between the test brushes for any of the three scored areas.

Adult↗
PubMed1996

Measurement of Goldmann visual fields in older children who received cryotherapy as infants for threshold retinopathy of prematurity.

BACKGROUND: Cryotherapy administered to eyes with severe acute-phase (threshold) retinopathy of prematurity benefits retinal structure and visual acuity compared with the natural course of the retinopathy. OBJECTIVES: To determine the extent of peripheral field abnormalities in eyes with threshold retinopathy of prematurity that had retinal structure preserved by cryotherapy. METHODS: Kinetic perimetry was performed with a Goldmann perimeter by masked testers on patients in whom bilateral threshold retinopathy of prematurity developed and who had been randomly assigned to undergo cryotherapy in one eye and no cryotherapy in the fellow eye. With the V-4-e and the II-4-e targets, eight meridians were tested: 0 degrees, 45 degrees, 90 degrees, 135 degrees, 180 degrees, 225 degrees, 270 degrees, and 315 degrees. The median value of three presentations in each meridian was accepted as the extent in that meridian. RESULTS: Fourteen eyes (eight treated and six control) of eight patients (mean age, 9.9 years; range, 6 to 11 years) had adequate vision to undergo fields testing. Mean (+/-SE) extent of visual field for treated vs control eyes was 36 degrees +/- 3 degrees vs 46 degrees +/- 6 degrees for the II-4-e target and 49 degrees +/- 4 degrees vs 59 degrees +/- 6 degrees for the V-4-e target. This difference was consistent across all eight meridians for either target, and repeated-measures analysis of variance showed that cryotherapy was associated with smaller visual field extent for both target sizes (P=.08). CONCLUSION: The results of this small pilot study suggest that eyes that have retinal structure and acuity preserved by cryotherapy for severe acute-phase retinopathy of prematurity have slightly smaller visual fields than untreated eyes with severe acute-phase retinopathy of prematurity that had vision preserved.

Acute Disease↗
PubMed1996

Miniature-probe measurements of electric fields induced by 60 Hz magnetic fields in rats.

Extremely-low-frequency (ELF) magnetic fields interact with an animal by inducing internal electric fields, which represent the internal dose from an external exposure. In this study, an electric field probe of approximately 2 mm resolution was used to measure fields induced in rat carcasses by a 60 Hz magnetic field at 1 mT. With the rat lying on its side, the probe was inserted through a small hole in the body wall, and scanned at 5 mm increments from the side with frontal and axial exposure (field horizontal) and from the front with lateral exposure (field vertical). The induced electric field declined from a maximum at the entrance to the abdomen and crossed zero to negative (180 degrees phase shift) values within the body as expected. In general, the magnitudes of the measurements inside the abdomen were less than expected from whole-body calculations that used homogeneous-ellipsoidal models of a rat in the three orientations. The low measurements did not appear to be explained by perpendicular field components, by conductivity differences between the tissue and the probe path, or by air in the lungs. The low measurements probably result from inhomogeneities in actual rats that include conductivity differences between tissues and biological membranes. For example, an alternative model considered the abdominal cavity to be electrically isolated from the body by the diaphragm and the peritoneum and calculations from this model were in better agreement with the measurements inside the abdomen (than were the whole-body calculations). Therefore, inhomogeneitics in conductivity and biomembranes such as the peritoneum should be considered in order to fully understand ELF-induced field dosimetry.

Abdomen↗