Search PubMed⌕ Search

Biomedical subjects

K O'Brien

Publications and source records attributed to K O'Brien.

At least 37 records · Page 2Linked to original sources
PubMed1998

Cloning, expression, purification, and characterization of the acid alpha-mannosidase from Trypanosoma cruzi.

The acid alpha-mannosidase of Trypanosoma cruzi is a broad-specificity hydrolase involved in the catabolism of glycoconjugates, presumably in the digestive vacuole. We have cloned the alpha-mannosidase gene from a T.cruzi epimastigote genomic library. The alpha-mannosidase gene was determined to be single copy by Southern analysis, and similar sequences were not detected in genomic digests of either Trypanosoma brucei or Leishmania donovani. The coding region was subcloned into the Pichia pastoris expression vector pPICZ, and alpha-mannosidase activity was detected in the medium of induced cultures. The recombinant alpha-mannosidase demonstrated a pH optimum, inhibition by swainsonine, Km, and substrate specificity consistent with the characteristics of the alpha-mannosidase previously purified from T.cruzi epimastigotes. The recombinant enzyme was purified 103-fold from the culture medium of Pichia pastoris and had a native molecular mass of 359 kDa by gel filtration. A combination of SDS-PAGE, deglycosylation with endo H, and NH2-terminal sequencing indicates that the enzyme is originally synthesized as a homodimeric polypeptide that is subsequently cleaved to form a heterotetramer composed of 57 and 46 kDa subunits. A polyclonal antibody raised to the recombinant enzyme was shown to immunoprecipitate the alpha-mannosidase from T.cruzi cell extracts and will be used in future immunolocalization studies.

Amino Acid Sequence↗
PubMed1998

MerMade: an oligodeoxyribonucleotide synthesizer for high throughput oligonucleotide production in dual 96-well plates.

We have designed and constructed a machine that synthesizes two standard 96-well plates of oligonucleotides in a single run using standard phosphoramidite chemistry. The machine is capable of making a combination of standard, degenerate, or modified oligos in a single plate. The run time is typically 17 hr for two plates of 20-mers and a reaction scale of 40 nM. The reaction vessel is a standard polypropylene 96-well plate with a hole drilled in the bottom of each well. The two plates are placed in separate vacuum chucks and mounted on an xy table. Each well in turn is positioned under the appropriate reagent injection line and the reagent is injected by switching a dedicated valve. All aspects of machine operation are controlled by a Macintosh computer, which also guides the user through the startup and shutdown procedures, provides a continuous update on the status of the run, and facilitates a number of service procedures that need to be carried out periodically. Over 25,000 oligos have been synthesized for use in dye terminator sequencing reactions, polymerase chain reactions (PCRs), hybridization, and RT-PCR. Oligos up to 100 bases in length have been made with a coupling efficiency in excess of 99%. These machines, working in conjunction with our oligo prediction code are particularly well suited to application in automated high throughput genomic sequencing.

Chromosome Mapping↗
PubMed1998

Orthodontic process and outcome: efficacy studies--strategies for developing process and outcome measures: a new era in orthodontics.

The development of process and outcome measures in orthodontics lends itself to retrospective clinical studies. Once valid and reliable measures have been tested, prospective studies may utilize these measures to assess the quality of orthodontic treatment outcome and the duration and process of treatment. The findings from two retrospective studies comparing the quality of outcome using the peer assessment rating (PAR) occlusal index and duration of treatment are reported. Class I and Class II treatment comparisons indicate the duration of treatment time is increased on average 5 months in Class II, division 1 patients. In the second study, the effect of extraction/non-extraction treatment in a sample that included all types of malocclusion indicates that, on average, treatment time is increased by approximately 5 months when extractions are included as part of the orthodontic treatment plan.

Analysis of Variance↗
PubMed1997

Limb defects in homozygous alpha-thalassemia: report of three cases.

Homozygosity for the South-Asian alpha-thalassemia (--SEA/) deletion is a serious hematological condition that results, in most cases, in intrauterine or postnatal death due to anemia and severe hypoxia of prenatal onset. A relationship between congenital abnormalities and intra-uterine hypoxia has been postulated. However, since homozygosity for the (--SEA/) deletion is most common in underdeveloped countries where detailed autopsies are lacking, the incidence of congenital abnormalities among these babies has not been well delineated. We report on three newborn infants, homozygous for the (--SEA/) deletion, who were born with limb defects. We postulate that this combination is the result of prenatal hypoxia which may affect other fetal body organs. This should be taken into consideration when prenatal treatment of affected fetuses, with intrauterine blood transfusion, is suggested.

Adult↗
PubMed1997

Construction and characterization of a one-plasmid system for the controlled expression of genes in mammalian cells by tetracycline.

The two-plasmid system of Gossen and Bujard [Gossen and Bujard (1992) Proc. Natl. Acad. Sci. USA 89, 5547-5551] to express mammalian genes in a tetracycline-repressed fashion was combined into a single-plasmid system. Two variants of this single-plasmid system that differ in the multiple cloning site (MCS) region are described. These vectors were used to stably transfect raf kinase domain into the normal rat kidney epithelial cell line (NRKE) to obtain a conditionally transformed cell line. These vectors were also used to stably transfect wild-type and mutant human p53 into the human osteosarcoma cell line, SAOS-2. Tetracycline repressed gene expression in both cell lines; about 12-fold in NRKE and about 80-fold in SAOS-2 cell line.

Animals↗
PubMed1997

Polycystic ovarian condition in the dehydroepiandrosterone-treated rat model: hyperandrogenism and the resumption of meiosis are major initial events associated with cystogenesis of antral follicles.

BACKGROUND: The purpose of this study was to elucidate the early effects of dehydroepiandrosterone (DHEA) in the polycystic rat model by charting cytological changes in the early antral follicle of the ovary and constructing a serum hormonal profile. Histological examinations of ovaries from DHEA-treated rats for ten consecutive days revealed that the oocyte of antral follicles, ranging from 1.5 mm to 3.4 mm in diameter, had become activated, i.e., had resumed meiosis. Tabulation and statistical analysis revealed a highly significant difference in the percentage of oocyte activation between the ovaries of DHEA-treated and control rats. Granulosa cells associated with those antral follicles included in our statistical analysis showed no evidence of atresia. A few follicles not included in our analysis contained oocytes that had resumed meiosis and whose associated granulosa cells were atretic. The observed resumption of meiosis occurred in the absence of surges of follicle stimulating hormone (FSH) and luteinizing hormone (LH). During meiosis, a period when many oocytes become activated, levels of serum androgens (DHEA, testosterone, and androstenedione) were high, while FSH, LH, and prolactin (PRL) levels did not differ significantly from those in the controls. Follicles that resume meiosis may be members of a group of follicles that produces a signal(s) when the oocyte becomes uncoupled from the granulosa cell. This signal(s) permit(s) a reprogramming of the accompanying granulosa cells of the follicle to engage in certain developmental processes of cystogenesis. Just what cascade of signals is necessary to achieve this selection remains elusive at this time and is the subject of our continuing investigations.

Androstenedione↗
PubMed1997

Web-site review.

Explore the source record for details and available documents.

Australia↗
PubMed1997

Biochemical evidence for a novel low molecular weight 2-5A-dependent RNase L in chronic fatigue syndrome.

Previous studies from this laboratory have demonstrated a statistically significant dysregulation in several key components of the 2',5'-oligoadenylate (2-5A) synthetase/RNase L and PKR antiviral pathways in chronic fatigue syndrome (CFS) (Suhadolnik et al. Clin Infect Dis 18, S96-104, 1994; Suhadolnik et al. In Vivo 8, 599-604, 1994). Two methodologies have been developed to further examine the upregulated RNase L activity in CFS. First, photoaffinity labeling of extracts of peripheral blood mononuclear cells (PBMC) with the azido 2-5A photoaffinity probe, [32P]pApAp(8-azidoA), followed by immunoprecipitation with a polyclonal antibody against recombinant, human 80-kDa RNase L and analysis under denaturing conditions. A subset of individuals with CFS was identified with only one 2-5A binding protein at 37 kDa, whereas in extracts of PBMC from a second subset of CFS PBMC and from healthy controls, photolabeled/immunoreactive 2-5A binding proteins were detected at 80, 42, and 37 kDa. Second, analytic gel permeation HPLC was completed under native conditions. Extracts of healthy control PBMC revealed 2-5A binding and 2-5A-dependent RNase L enzyme activity at 80 and 42 kDa as determined by hydrolysis of poly(U)-3'-[32P]pCp. A subset of CFS PBMC contained 2-5A binding proteins with 2-5A-dependent RNase L enzyme activity at 80, 42, and 30 kDa. However, a second subset of CFS PBMC contained 2-5A binding and 2-5A-dependent RNase L enzyme activity only at 30 kDa. Evidence is provided indicating that the RNase L enzyme dysfunction in CFS is more complex than previously reported.

2',5'-Oligoadenylate Synthetase↗
PubMed1997

A clinical evaluation of the effectiveness of including fluoride into an orthodontic bonding adhesive.

A clinical trial was undertaken to assess the value of incorporating fluoride released from a commercially available bonding adhesive (Rely-a-Bond) to determine the extent of any protection provided against enamel decalcification. Fifty patients undergoing fixed appliance therapy were included in the trial. Contralateral quadrants were used as controls where no fluoride was present in the adhesive. Enamel decalcification after treatment and bond failure rates during treatment were investigated. A total of 366 experimental and 371 control teeth were included in the study. The results showed that 50 per cent of patients and 13.5 per cent of teeth exhibited post-treatment decalcification. The addition of fluoride to the adhesive did not significantly reduce the incidence of enamel decalcification. Bond failure rates were satisfactory for both experimental and control teeth (all under 5 per cent).

Cariostatic Agents↗
PubMed1997

Balance and skeletal alignment in a group of elderly female fallers and nonfallers.

The purpose of this study was to determine whether sagittal plane posture differed between fallers and nonfallers and to explore the relationship between skeletal alignment and balance in elderly females. Forty-eight women > 65 years of age were recruited from various medical and senior citizen centers. Thirteen of these women were classified as fallers. Spinal posture was measured in standing using an inclinometer, and lower extemity joint angles were assessed in standing using a universal goniometer. The Berg Balance Scale, the Functional Reach Test, end a modified Timed Get Up and Go Test were used to measure balance. Spinal alignment did not differ significantly between fallers and nonfallers; however, knee joint angle was significantly greater in fallers compared to nonfallers. Significant, but low, correlations were found between the inclination of the upper thoracic spine and all three balance measures. Lower thoracic slope and knee joint angle in standing were also weakly related to two of the three balance measures. This study supports the hypothesis that a significant but weak relationship exists between balance and skeletal alignment in elderly females.

Accidental Falls↗
PubMed1997

Measuring beliefs about orthodontic treatment: a questionnaire approach.

OBJECTIVES: Few studies have examined what parents and orthodontists expect from and value about orthodontic treatment. In this study, we designed and tested a questionnaire to outline what drives consumer demand for children's orthodontic care. Further, we present data from the questionnaire to illustrate how expectations and values pertaining to orthodontic treatment relate to sociodemographic variables. METHODS: Subjects were 220 Pennsylvania orthodontists and 220 parents at a university orthodontic clinic who were administered a questionnaire designed to assess what parents and orthodontists value about and expect from orthodontic treatment. Items for the questionnaire were developed via a qualitative, telephone interview process. Data were analyzed using factor analysis and reliability analysis for scale development, and analysis of variance for preliminary validity assessment. RESULTS: Through factor analysis, the questionnaire was reduced from 84 to 52 items, and eight scales were examined: expected treatment benefits, expected treatment risks (short- and long-term), expected treatment inconveniences, value of treatment benefits, value of risks (short- and long-term), and value of treatment inconveniences. For parents, the reliability for all scales was in the acceptable range. For orthodontists, only the "short-term risks" scale failed to attain an acceptable reliability. Preliminary validity was assessed through examining relationships between demographic variables and subscale scores. For parents, income, father's education level, and sex of respondent were related to treatment expectations and values. For orthodontists, age, sex, and patient volume were related to treatment values. CONCLUSIONS: The questionnaire developed in the present study was found to be practical and reliable for use with providers and consumers of orthodontic care and can be used to explore factors affecting the demand for orthodontic care. Implications of possible unrealistic treatment expectations on the part of orthodontists and parents also are discussed.

Adolescent↗
PubMed1997

High bone mass as a marker for breast cancer risk.

Postmenopausal women in the highest quartile for metacarpal bone mass were found to have an increased risk of developing breast cancer, after adjusting for age and other variables known to influence breast cancer risk. Although the mechanisms responsible for this relationship have not been identified, postmenopausal bone mass may serve as an indicator of cumulative estrogen exposure.

Bone Density↗