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Biomedical subjects

T O'Brien

Publications and source records attributed to T O'Brien.

At least 109 records · Page 6Linked to original sources

Expression and function of a recombinant endothelial nitric oxide synthase gene in porcine coronary arteries.

OBJECTIVES: Direct gene transfer of exogenous nitric oxide synthase, with the subsequent increase in nitric oxide production, could represent a potential therapeutic strategy in the treatment of vascular proliferative disorders. The goal of the present study was to determine if porcine coronary arteries could be transduced with an adenoviral vector encoding endothelial nitric oxide synthase (Ad.CMVeNOS) resulting in functional expression. METHODS AND RESULTS: Segments of porcine right coronary artery were exposed for 1 h at 37 degrees C to either replication-deficient adenovirus encoding bovine endothelial nitric oxide synthase (Ad.CMVeNOS, 5 x 10(9) pfu/ml) or control adenovirus encoding Escherichia coli beta-galactosidase (Ad.CMVLacZ, 5 x 10(9) pfu/ml). Immunohistochemistry with a monoclonal antibody specific for nitric oxide synthase (NOS) demonstrated recombinant gene expression in both the endothelial and adventitial layers of Ad.CMVeNOS transduced coronaries with only endogenous NOS confirmed in the endothelium of Ad.CMVLacZ arteries. Coronary arteries transduced with Ad.CMVeNOS yielded 517 +/- 110 (mean +/- S.E.M.) nM/ng nitrite while vessels transduced with Ad.CMVLacZ yielded 126 +/- 84 nM/ng (P < 0.05, n = 6). Isometric tension recording, following prostaglandin F2 alpha constriction, documented a reduced tension in Ad.CMVeNOS transduced coronaries, compared to matched Ad.CMVLacZ coronaries (6.10 +/- 1.08 g vs. 8.45 +/- 1.19 g, respectively, P = 0.05, n = 8). This tension differential was eliminated with prior incubation in NG-monomethyl-L-arginine (L-NMMA, 10(-4) M). The EC50 for calcium ionophore relaxation of Ad.CMVeNOS coronary arteries was reduced compared to Ad.CMVLacZ (-7.90 +/- 0.03 logM vs. -7.26 +/- 0.11 logM, respectively, P < 0.05, n = 8). CONCLUSIONS: These studies demonstrate successful transfer of endothelial nitric oxide synthase into porcine coronary arteries as verified by histochemical localization of recombinant protein with an increase of nitric oxide release as demonstrated by enhanced nitrite production and an alteration in vasomotor function.

Adenoviridae↗

Immunosuppression prolongs adenoviral mediated transgene expression in cardiac allograft transplantation.

BACKGROUND: The immune response to adenoviral vectors used in gene transfer limits the duration of transgene expression and thus poses a potential limitation to their effectiveness for gene therapy. The need for immunosuppression in transplantation may modify this immune response and facilitate prolonged transgene expression. This study hypothesizes that in the setting of heart transplantation, the use of routine immunosuppression will prolong adenoviral-mediated transgene expression. METHODS AND RESULTS: In a model of rat heterotopic abdominal heart transplantation, 350 microliters of viral solution (1 x 10(9) pfu/ml) was infused into the coronary arteries of donor hearts at the time of procurement. The duration of transgene expression was examined following (a) syngeneic transplantation in non-immunosuppressed animals (group A): (b) syngeneic transplantation in immunosuppressed animals (group B); and (c) allogeneic transplantation in immunosuppressed animals (group C). After transplantation donor hearts were studied at; 1, 4, 8 and 12 weeks. Transgene expression was assessed by histochemical staining of tissue cross sections for beta-galactosidase activity. In the non-immunosuppressed syngeneic group (group A), transgene expression had largely disappeared by 4 weeks, whereas in both the immunosuppressed syngeneic (group B) and immunosuppressed allogeneic (group C) animals expression of the reporter gene persisted for the 12 weeks of the study, although the level of expression decreased significantly over time. CONCLUSIONS: This study demonstrates that transgene expression using adenoviral vectors is prolonged by immunosuppression in the heart transplantation setting.

Adenoviridae↗

The effect of the treatment of hypothyroidism and hyperthyroidism on plasma lipids and apolipoproteins AI, AII and E.

OBJECTIVE: Although lipid abnormalities are well described in hypothyroidism, effects on apolipoproteins are less well understood. The aim of this study was to examine the effects of thyroid dysfunction on plasma lipids and apolipoproteins. DESIGN: A prospective study of lipids and apolipoproteins before and after treatment of hypothyroidism and hyperthyroidism. PATIENTS: Eighteen patients with hypothyroidism and 5 patients with hyperthyroidism were included. MEASUREMENTS: Plasma cholesterol, triglycerides, HDL cholesterol, apo AI, apo AII, and apo E were measured before and after treatment of the thyroid abnormality. RESULTS: Total and HDL cholesterol, apo AI and apo E decreased with treatment of hypothyroidism, while triglycerides and apo AII levels were unchanged. The total/HDL cholesterol and LDL/HDL cholesterol ratios also decreased with treatment of hypothyroidism. In contrast, treatment of hyperthyroidism was associated with an increase in total and HDL cholesterol, and apo AI. Triglycerides, apo AII and Apo E were unchanged by treatment of hyperthyroidism. The total/HDL cholesterol and the LDL/HDL cholesterol ratios increased with treatment of hyperthyroidism. CONCLUSIONS: Hypothyroidism and hyperthyroidism have opposite effects on plasma lipids and apolipoproteins. In hypothyroidism, total and HDL cholesterol, total/HDL cholesterol ratio, apo AI and apo E are elevated. The increase in apo AI without a concomitant increase in apo AII suggests selective elevation of HDL2. In contrast, hyperthyroidism is associated with decreased total and HDL cholesterol, total/HDL cholesterol ratio, and apo AI levels. These effects are reversible with treatment of the underlying thyroid disorder.

Adult↗

Lithium dilution cardiac output measurement: a comparison with thermodilution.

OBJECTIVE: To compare the results of cardiac output measurements obtained by lithium dilution and thermodilution. DESIGN: Case series, observational study. SETTING: High-dependency postoperative unit and intensive care unit of a teaching hospital. PATIENTS: Forty patients were studied. Thirty-four patients had undergoing heart surgery requiring cardiopulmonary bypass within the previous 2 days; the diagnoses in the other patients were myocardial infarct (n = 2), septicemia (n = 2), adult respiratory distress syndrome, and pericardectomy. INTERVENTIONS: Cardiac output was measured five times in each patient, using lithium dilution (single measurement) and bolus thermodilution (series of three to six measurements according to standard clinical practice, taking the average of the closest three). In a subgroup of 14 patients, cardiac output was also measured using "continuous thermodilution." MEASUREMENTS AND MAIN RESULTS: Comparing lithium dilution with bolus thermodilution, the mean of the differences (lithium dilution-thermodilution) was -0.25 +/- 0.46 [SD] L/min. Linear regression analysis gave y = 0.31 + 0.89x (r2 = .94) for lithium dilution vs. thermodilution. CONCLUSIONS: The overall agreement between the two methods was good. The variability of the thermodilution measurements was greater than that of the lithium dilution measurements. The lithium dilution method is at least as accurate as bolus thermodilution and, since pulmonary artery catheterization is not needed, it has the advantages of being safe and quick to perform.

Adult↗

Expression and function of recombinant endothelial NO synthase in coronary artery smooth muscle cells.

Smooth muscle cells (SMCs) play a key role in the pathogenesis of vascular diseases. The objectives of this study were to determine whether transfer of recombinant endothelial nitric oxide synthase (eNOS) gene to porcine coronary artery smooth muscle cell (CSMCs) would result in expression of a functional enzyme and to assess the effect of expression of eNOS on cell proliferation. CSMCs were transduced in vitro with adenoviral vectors encoding cDNA for eNOS (AdeNOS) and beta-galactosidase (Ad beta Gal). In contrast to Ad beta Gal- or sham-transduced cells, CSMCs transduced with AdeNOS stained positive with the NADPH-diaphorase stain, acquired calcium-dependent NOS activity (measured by the conversion of [3H]L-arginine to [3H]L-citrulline), had increasing cyclic 3',5' cGMP levels with increasing concentrations of the vector, and produced increased amounts of nitrite. cGMP production by AdeNOS-transduced cells was augmented by increasing intracellular levels of the eNOS cofactor tetrahydrobiopterin. CSMCs transduced with AdeNOS showed diminished serum-stimulated DNA synthesis as measured by thymidine uptake. Cell proliferation was diminished in AdeNOS-transduced CSMCs as assessed by cell counts 3 and 6 days after serum stimulation of quiescent CSMCs. The present study demonstrates that adenovirus-mediated gene transfer of eNOS to CSMCs results in the expression of a functional enzyme whose activity can be augmented by increasing intracellular levels of tetrahydrobiopterin. Expression of recombinant eNOS in CSMCs results in inhibition of serum-stimulated DNA synthesis and cell proliferation. These findings imply that eNOS gene transfer to SMCs may be a unique mode of increasing local NO production in the arterial wall.

Adenoviridae↗

Enhanced endothelium-dependent relaxations after gene transfer of recombinant endothelial nitric oxide synthase to rabbit carotid arteries.

We tested the effects of overexpression of the endothelial nitric oxide synthase (eNOS) gene in the normal arterial wall by adenoviral-mediated gene transfer. Rabbit carotid arteries were surgically isolated and exposed to adenoviral vectors encoding eNOS (AdeNOS) or beta-galactosidase (Ad betaGal) on the contralateral side. Vector solutions at a concentration of 1 x 10(10) plaque forming units/mL were instilled for 20 minutes before restoration of flow. Arteries were harvested 4 days later for immunostaining, measurement of cGMP, and vasomotor studies. Endothelium-specific gene transfer was confirmed by staining for beta-galactosidase in the Ad betaGal arteries. Immunostaining of en face endothelial cell imprints from AdeNOS-transduced arteries with a monoclonal antibody to eNOS showed increased immunoreactivity. Basal cGMP levels were significantly greater in the AdeNOS-transduced arteries (18.4+/-4.6 versus 4.2+/-0.5 pmol/mg protein; P<.05). Contractions to phenylephrine were significantly reduced in the AdeNOS-transduced arteries (area under curve, 106+/-5 versus 119+/-7; P<.05), but in the presence of the eNOS inhibitor, N(G)-monomethyl-L-arginine (L-NMMA, 3 x 10(-4) mol/L), there was no difference between the two (area under curve, 148+/-5 versus 153+/-6; P=NS). Relaxations to acetylcholine obtained during submaximal contractions to phenylephrine were significantly enhanced in the AdeNOS-transduced arteries (EC50, 7.45+/-0.05 versus 7.23+/-0.03; P<.05). We conclude that overexpression of eNOS in the endothelium results in diminished contractile responses, as well as enhanced endothelium-dependent relaxations. These findings imply a possible role for vascular eNOS gene transfer in the treatment of vasospasm and endothelial dysfunction.

Animals↗

Expression and function of recombinant endothelial nitric oxide synthase gene in canine basilar artery.

Endothelial NO synthase (eNOS) is an enzyme responsible for the production of a potent vasodilator and a key regulator of vascular tone, NO. In peripheral arteries, expression of a recombinant eNOS gene increases production of NO in the blood vessel wall. This approach appears to be a promising strategy for gene therapy of cerebrovascular disease. The major objective of the present study was to determine whether a recombinant eNOS gene (AdCMVNOS) can be functionally expressed in cerebral arteries. Replication-defective recombinant adenovirus vectors encoding bovine eNOS and Escherichia coli beta-galactosidase (AdCMVLacZ) genes, driven by the cytomegalovirus promoter, were used for ex vivo gene transfer. Rings of canine basilar artery were incubated with increasing titers of the vectors in MEM. Twenty-four or forty-eight hours after gene transfer, expression and function of AdCMVNOS were evaluated by (1) immunohistochemical staining, (2) isometric tension recording, and (3) cGMP radioimmunoassay. Transfection with AdCMVNOS resulted in the expression of recombinant eNOS protein in the vascular adventitia and endothelium, associated with significantly reduced contractile responses to UTP and enhanced endothelium-dependent relaxation to calcium ionophore A23187. Basal production of cGMP was significantly increased in the transfected vessels. The reduced contractions to UTP with increased cGMP production were reversed by a NOS inhibitor, N(G)-monomethyl-L-arginine. Contractions to UTP or production of cGMP were not affected in arteries transfected with AdCMVLacZ reporter gene. The results of the present study represent the first successful transfer and functional expression of recombinant eNOS gene in cerebral arteries. Our findings suggest that cerebral arterial tone can be modulated by recombinant eNOS expression in the vessel wall.

Animals↗

A randomized crossover study comparing the efficacy and tolerability of a novel once-daily morphine preparation (MXL capsules) with MST Continus tablets in cancer patients with severe pain.

The efficacy, tolerability and 24-h duration of action of MXL capsules, a novel once-daily morphine preparation, were compared with twice-daily morphine tablets (MST Continus tablets) in patients with severe cancer pain. Eighty-five patients were recruited to this randomized, double-blind, double-dummy, crossover study. There was no significant difference between the two treatment groups in the number of occasions that escape medication was required, the pain scores at each of three time points throughout the day, and the number of nights woken due to pain. Both preparations were well tolerated with no significant difference in the number or severity of reported symptoms and side-effects. Sixteen patients withdrew from the study, of whom 13 withdrew for nontreatment-related reasons. There was no difference between the preparations in terms of expressed treatment preference. MXL capsules were shown to provide effective analgesia over the 24-h dosing interval which was comparable to that of MST Continus tablets administered twice daily.

Analgesics, Opioid↗

Lipids and lipoproteins in women.

Coronary artery disease (CAD) is the most common cause of death in women in the United States. Dyslipidemia is a risk factor for CAD in both men and women. Low levels of high-density lipoprotein (HDL) cholesterol and hypertriglyceridemia, especially in association with a dense low-density lipoprotein (LDL) phenotype, may be of greater importance in women than in men. The relationship between CAD and dyslipidemia and the therapeutic approach to disorders of lipid metabolism in women have unique features because of the effects of exogenous and endogenous hormones on lipid pathways. Estrogen decreases LDL cholesterol and Lp(a) lipoprotein and increases triglyceride and HDL cholesterol levels. Progestogens decrease triglycerides, HDL cholesterol, and Lp(a), and they increase LDL cholesterol. Thus, oral contraceptives increase plasma triglycerides, whereas the effect of these agents on LDL cholesterol and HDL cholesterol levels is related to the androgenicity and dose of progestogen. Postmenopausal hormone replacement therapy increases triglycerides and decreases LDL cholesterol. The effect of hormone replacement therapy on HDL cholesterol is influenced by the addition of progestogen. Although no primary prevention studies have analyzed lipid lowering and CAD in women, secondary prevention studies have suggested that the response to drug treatment and the benefit of lipid lowering are similar in women and in men. Hormone replacement therapy should be considered in the treatment of hypercholesterolemia in postmenopausal women; however, individualization of treatment is important to avoid adverse effects.

Estrogen Replacement Therapy↗

Directing antigen specificity towards botulinum neurotoxin with combinatorial phage display libraries.

The production of antibodies towards antigens with low immunogenicity is enhanced by the intrinsic efficiency of screening combinatorial libraries of immunoglobulins. The need to isolate clones with rare binding specificities has dictated a highly efficient method of screening and isolating antibody clones. The production of recombinant immunoglobulin libraries in bacteria allows for a more controlled selection of antibody specificity and can be used in circumstances where hybridoma fusions are unable to isolate rare clones with the desired epitope specificity. Botulinum neurotoxin (NT) with associated non neurotoxin proteins (non-NT) as a complex was used to immunize mice to obtain mRNA for the production of a recombinant antibody library with a repertoire of specificities. Initial screens of the combinatorial library revealed clones which recognized the non-neurotoxin proteins of the toxin complex similar to monoclonal antibodies produced by conventional hybridoma fusions. The combinatorial library was re-screened in order to isolate antibodies that specifically recognized the neurotoxin component of the toxin complex. The ability to alter the biopanning selection process affords the researcher a measure of control in the selection process not available with traditional hybridoma fusions.

Animals↗

The angiogenic factor midkine is expressed in bladder cancer, and overexpression correlates with a poor outcome in patients with invasive cancers.

Midkine (MK) is a member of a family of heparin-binding growth factors, which are reported to be angiogenic. We have investigated by RNase protection analysis the expression of MK in 47 primary bladder tumors and 7 normal bladder samples. MK mRNA transcripts were detectable in 46 (98%) of 47 of the tumors and in 5 (70%) of 7 of the normal bladder samples. However, median MK expression was 4-fold higher in tumors than in the normal bladder (P < 0.004). In eight tumors (17%), MK expression was elevated more than 10-fold compared with the median value of the normal bladder specimens. There was no statistically significant difference in expression between superficial and invasive tumors (P < 0.50). Seven (32 %) of 22 patients with invasive cancers are alive at 1 year with no evidence of recurrence; in 5 (70%) of these patients, MK expression in the tumor was within the normal range at the time of presentation. By contrast, in only 2 (13%) of 15 patients who died or whose tumors recurred or progressed was MK expression in the normal range (P < 0.01). Overall, median MK expression in invasive tumors that caused death, progressed, or recurred within 12 months was 3-fold higher than that found in the tumors of those patients who were clear of disease at 12 months (P < 0.04). Thus, overexpression of MK is associated with the development of bladder cancer and in invasive cancers predicts a poor clinical outcome in the short term.

Adult↗

HDL subparticles and coronary artery disease in NIDDM.

Decreased HDL cholesterol levels are associated with an increased risk of coronary artery disease (CAD) in non insulin dependent diabetes mellitus (NIDDM). The aim of this study was to compare HDL subparticles with apo A-I (LpAI) and those with apo A-I and apo A-II (LpAI/AII) in subjects with and without NIDDM and to study the relationship between HDL subparticles and CAD in NIDDM. Lipids, apo A-I and HDL subparticles were measured in 240 subjects with NIDDM and in 248 age and gender matched controls. Subjects with NIDDM had higher triglyceride levels (2.5 +/- 1.8 vs. 1.4 +/- 0.8 mmol/1, P < 0.001), lower HDL cholesterol (0.9 +/- 0.3 vs. 1.2 +/- 0.3 mmol/l, P < 0.001), apo A-I (124.7 +/- 22.4 vs. 139.8 +/- 24.1 mg/dl, P < 0.001) and LpAI/AII (82.4 +/- 18.2 vs. 94.9 +/- 16.7 mg/dl, P < 0.001) in comparison to controls. LpAI levels were similar in both groups. Diabetic subjects with CAD (n = 109) had higher triglycerides (2.7 +/- 1.9 vs. 2.3 +/- 1.8 mmol/l, P = 0.02) and lower HDL cholesterol (0.8 +/- 0.2 vs. 1.0 +/- 0.3 mmol/l, P <0.001), apo A-I (115.5 +/- 20.1 vs. 132.3 +/- 21.4 mg/dl, P < 0.001), LpAI (40.2 +/- 9.1 vs. 44.4 +/- 12.4 mg/dl, P = 0.06), and LpAI/AII levels (75.4 +/- 18.0 vs. 88.3 +/- 16.2 mg/dl, P < 0.001) in comparison to diabetic subjects without CAD (n = 131). In a multivariate analysis, apo A-I was found to be the best predictor of CAD in subjects with NIDDM. In conclusion, reduced HDL cholesterol levels found in NIDDM are, principally, due to reduced concentrations of apo A-I and apo A-II-containing particles (LpAI/AII). While LpAI and LpAI/AII levels were lower in NIDDM subjects with CAD, plasma apo A-I is the best predictor of CAD in NIDDM.

Aged↗

Results of treatment of pituitary disease in multiple endocrine neoplasia, type I.

OBJECTIVE: The aim of the present study was to examine the clinical and pathological features of pituitary disease in patients with multiple endocrine neoplasia, Type I (MEN I) and to assess the prognosis. METHODS: Fifty-two patients with pituitary disease and MEN I were studied retrospectively. Medical records were reviewed, and all of the patients known to be alive were sent a questionnaire to ascertain current disease status. RESULTS: In 12 patients, pituitary disease was the initial manifestation of MEN I. The most common lesion was prolactinoma, followed, in frequency, by acromegaly and nonsecretory adenoma. Thirty-four of the patients had surgical treatment at the Mayo Clinic, Rochester, MN, as primary treatment, 3 had radiotherapy, and 12 received no specific therapy. Twelve patients had adjunctive radiotherapy postoperatively. Of the 34 patients receiving surgical treatment, 33 had adenoma and 1 had adenoma and pituitary hyperplasia. Immunocytochemical examination demonstrated that many tumors showed reactivity for more than one pituitary hormone. On survival analysis, no excess pituitary-related mortality was found, either in the surgically treated group or in the group as a whole. CONCLUSION: On the basis of this study, we conclude that pituitary disease is frequently the initial manifestation of MEN I; that adenomas, particularly prolactinomas, are the rule and hyperplasia is rare; that a significant proportion of tumors are plurihormonal; and that excess pituitary-related mortality is not a factor in patients with MEN I.

Acromegaly↗

Realtime three-dimensional remote medical skills acquisition.

Three-dimensional techniques are gaining widespread use in a number of disciplines, including medicine, where the visualization of depth information is not only useful but important. Both computer-generated 3-D imaging and 3-D video are set to become increasingly important tools for both diagnosis and teaching. In this paper we will present some of the work that we are currently undertaking using 3-D techniques for remote surgical skills acquisition. This work is based on the use of a small headset with twin miniature video cameras, video codecs, two different 3-D visualization systems and BT's ISDN telecommunications network.

Depth Perception↗

Methylphenidate dosing: twice daily versus three times daily.

OBJECTIVE: To evaluate the short-term efficacy and side effects associated with two methylphenidate hydrochloride (MPH) dosing patterns. METHODS: Twenty-five boys with attention deficit hyperactivity disorder (ADHD) participated in a 5-week, triple-blind, placebo-controlled, crossover evaluation of MPH administered twice (b.i.d.) versus thrice (t.i.d.) per day (mean dose = 8.8 +/- 5 mg, .30 +/- .1 mg/kg/dose). Four dosing conditions (placebo, titration [gradual increase to target dose], b.i.d., and t.i.d.) were used. Dependent measures obtained on a weekly basis included: parent and teacher ratings of child behavior, parent-child conflicts, parent report of stimulant side effects, child self-report of mood symptoms, a sleep log, laboratory measures of attention, and actigraphic recording of sleep activity. RESULTS: All dosing conditions resulted in significant effects on ADHD symptoms when compared with baseline. Relative to placebo, t.i.d. dosing was characterized by improvement on the greatest number of behavioral measures, and both b.i.d. and t.i.d. were generally more effective than titration. Direct comparisons of b.i.d. and t.i.d. dosing revealed that t.i.d. was associated with greater improvement on the Conners Parent Rating Scale Impulsivity/Hyperactivity factor, with a similar marginally significant effect for the ADD-H Teacher Rating Scale Hyperactivity factor. The analysis of clinically significant change favored a three-times-a-day dosing schedule over placebo on both parent and teacher ratings of impulsivity/hyperactivity and attention. Compared with placebo, appetite suppression was rated, on average, as more severe in the t.i.d. and titration conditions, but not in the b.i.d. condition. However, the number of subjects who exhibited any or severe appetite suppression did not differ significantly between the b.i.d. and t.i.d. schedules. Although there was no difference in sleep duration for children on b.i.d. and t.i.d. schedules, total sleep time appeared to decrease slightly on t.i.d. relative to placebo according to both parent ratings and actigraphic assessment. There were no significant differences between b.i.d. and t.i.d. on any other side effects or sleep variables. CONCLUSIONS: For many children with ADHD, t.i.d. dosing may be optimal. There are few differences in acute side effects between b.i.d. and t.i.d. MPH dosing. The dosing schedule should be selected according to the severity and time course of ADHD symptoms rather than in anticipation of dosing schedule-related side effects.

Attention Deficit Disorder with Hyperactivity↗

Coactivator and promoter-selective properties of RNA polymerase I TAFs.

Human ribosomal RNA synthesis by RNA polymerase I requires the activator UBF and the promoter selectivity factor SL1, which consists of the TATA binding protein (TBP) and three associated subunits, TAFI110, TAFI63, and TAFI48. Here it is shown that both TAFI110 and TAFI63 contact the promoter, whereas TAFI48 serves as a target for interaction with UBF and is required to form a transcriptionally active SL1 complex responsive to UBF in vitro. TAFI48 also alters the ability of TBP to interact with TATA box elements, and the resulting complex fails to support transcription by RNA polymerase II. Thus, TAFI48 may function both as a target to mediate UBF activation and as a class-specific promoter selectivity factor.

DNA-Binding Proteins↗