Biomedical subjects
Publications and source records attributed to H A Smith.
The framework for regulating DNA vaccines has been in place since the first clinical trial was initiated in the mid-1990s. American and European regulatory guidance has evolved on the basis of insights provided by ongoing preclinical and clinical studies. These include analyses of the safety of DNA vaccines in normal volunteers, and recent data concerning the tissue distribution, persistence, and integration potential of DNA plasmids.
SUMMARY: Although the neuroprotective effects of hypothermia have been known for a long time, the molecular correlates of this neuroprotection are poorly understood. In this study, the authors investigated how hypothermia affects inflammatory responses in the brain elicited by systemic injection of IL-1 beta. Leukocyte rolling and adhesion were quantified in pial venules (20 to 50 microm) of C57/Bl6 mice 4 hours after intraperitoneal injection of IL-1 beta (5 microg/kg) using an open cranial window and intravital microscopy. Animals were subjected to moderate hypothermia (32 degrees C) or normothermia (37 degrees C) for 1 or 4 hours after IL-1 beta injection. Significant increases in leukocyte rolling and adhesion were observed in IL-1 beta-injected animals as compared with sham controls. Whereas 1-hour hypothermia did not affect IL-1 beta-induced leukocyte rolling and adhesion, 4-hour hypothermia caused a reduction in both rolling and adhesion. Molecular mechanisms of hypothermic effects were investigated in cultured human cerebral endothelial cells exposed to IL-1 beta (50 U/mL) for 4 hours at 37 degrees C or 32 degrees C followed by 18 hours at 37 degrees C. Human cerebral endothelial cells exposed to IL-1 beta at 32 degrees C showed attenuated NF-kappa B activation determined by the Luciferase yellow reporter gene assay and reduced expression of IL-8 and IL-1 beta measured by reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay. Intracellular adhesion molecule-1 was induced to similar levels (threefold over control) at both temperatures. The expression of CD18 on neutrophils in vitro was not affected by either IL-1 beta or hypothermia. These findings suggest that mechanisms by which hypothermia reduces leukocyte rolling and adhesion include suppression of inflammatory gene transcription in brain endothelial cells.
BACKGROUND: dysphagia is common in acute stroke. Accurate detection of the presence or absence of aspiration by bedside swallowing assessment is difficult without objective methods, tending to over-diagnose aspiration. As a result, some patients suffer restricted oral intake unnecessarily. OBJECTIVE: we examined the predictive values of pulse oximetry and speech and language therapy bedside swallowing assessment in the detection of aspiration compared with videofluoroscopy. DESIGN: a double-blind observational study. SETTING: two university teaching hospitals. SUBJECTS: we studied 53 patients whose acute strokes were confirmed by computed tomography scan. METHODS: Each subject had initial standard bedside swallowing assessment, closely followed by simultaneous and mutually blinded pulse oximetry, swallowing assessment and videofluoroscopy. RESULTS: 15 of 53 subjects aspirated. Bedside swallowing assessment and saturation assessment at > or = 2% desaturation gave good sensitivity (80% and 87% respectively), but low positive predictive values (50% and 36% respectively). Both assessments mistook laryngeal penetration for aspiration. Re-analysis with aspiration +/- penetration as a new endpoint improved bedside swallowing assessment positive predictive values to 83% (chi2 =3.59, P=0.032). Sensitivity of saturation assessment was maintained at 86%, positive predictive values of saturation assessment improved to 69% (chi2=6.74, P=0.009). The combination of bedside swallowing assessment and saturation assessment versus aspiration + penetration gave a positive predictive value of 95%. CONCLUSIONS: screening by saturation assessments detects 86% of aspirators/penetrators and should be followed immediately by bedside swallowing assessment, as the combination of the two assessments gives the best positive predictive value. For patients with acute stroke, we advocate a 10 ml water-swallow screening test with simultaneous pulse oximetry by suitably trained medical and nursing staff. Use of this screening test would improve dysphagia detection whilst minimizing unnecessary restriction of oral intake in stroke patients.
Polyethylene mulch with a reflective silver stripe and a yellow summer squash, Cucurbita pepo L., trap crop were tested alone and in combination as tactics to reduce densities of Bemisia argentifolii Bellows & Perring eggs and nymphs, and incidence of bean golden mosaic geminivirus on snap bean, Phaseolus vulgaris L. Egg densities were consistently higher on squash than on bean, but egg densities and virus incidence were not lower on bean grown with squash than on bean grown in monoculture. Silver reflective mulch reduced egg densities compared with bean grown on bare ground during the first week after crop emergence for 2 of the 3 yr that the study was conducted. However, egg suppression by silver mulch was not enhanced by the presence of a squash trap crop when both tactics were combined. The obstacles to suppressing B. argentifolii through the use of trap crops are discussed.
BACKGROUND AND PURPOSE: Atherosclerosis of the major intracranial arteries is an important cause of ischemic stroke. We established measurement criteria to assess percent stenosis of a major intracranial artery (carotid, middle cerebral, vertebral, basilar) and determined the interobserver/intraobserver agreements and interclass/intraclass correlations of these measurements. METHODS: We defined percent stenosis of an intracranial artery as follows: percent stenosis = [(1 - (D(stenosis)/D(normal)))] x 100, where D(stenosis) = the diameter of the artery at the site of the most severe stenosis and D(normal) = the diameter of the proximal normal artery. If the proximal segment was diseased, contingency sites were chosen to measure D(normal): distal artery (second choice), feeding artery (third choice). Using a hand-held digital caliper, three neuroradiologists independently measured D(stenosis) and D(normal) of 24 stenotic intracranial arteries. Each observer repeated the readings 4 weeks later. We determined how frequently two observers' measurements of percent stenosis of each of the 24 diseased arteries differed by 10% or less. RESULTS: Among the three pairs of observers, interobserver agreements were 88% (observer 1 versus observer 2), 79% (observer 1 versus observer 3), 75% (observer 2 versus observer 3) for the first reading and were 75% (observer 1 versus observer 2), 100% (observer 1 versus observer 3), and 71% (observer 2 versus observer 3) for the second reading. Intraobserver agreement for each of the observers was 88%, 83%, and 100%. Interclass correlation was 85% (first reading) and 87% (second reading). Intraclass correlation was 92% (first and second readings combined). CONCLUSION: This method shows good interobserver and intraobserver agreements for the measurement of intracranial stenosis of a major artery. If validated in subsequent studies, this method may serve as a standard for the measurement of percent stenosis of an intracranial artery.
Research efforts to develop DNA vaccine products have progressed to the clinical evaluation of several vaccine candidates. These clinical trials are assessing safety and immunogenicity and whether different routes and schedules of administration can improve immune responses. Vaccine candidates under study include those for hepatitis B, HSV-2, HIV-1, influenza, and malaria. Unique safety concerns include the potential for genomic integration, biodistribution, tolerance and auto-immunity. CBER has issued a Points to Consider (PTC) document outlining regulatory policy, recommendations for the development of consistent manufacturing processes, and the design of preclinical studies to establish the safety of DNA vaccine products. This presentation will provide an overview of DNA vaccine products currently under clinical evaluation and the regulatory policy that has guided their development.
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Far-infrared hydrogen recombination lines H15 alpha (169.4 micrometers), H12 alpha (88.8 micrometers), and H10 alpha (52.5 micrometers) were detected in the peculiar luminous star MWC 349A from the Kuiper Airborne Observatory. Here it is shown that at least H15 alpha is strongly amplified, with the probable amplification factor being greater than or about equal to 10(3) and a brightness temperature that is greater than or about equal to 10(7) kelvin. The other two lines also show signs of amplification, although to a lesser degree. Beyond H10 alpha the amplification apparently vanishes. The newly detected amplified lines fall into the laser wavelength domain. These lasers, as well as the previously detected hydrogen masers, may originate in the photoionized circumstellar disk of MWC 349A and constrain the disk's physics and structure.
The expression of genes in mammalian cells depends on many factors including position in the cell cycle, stage of differentiation, age, and environmental influences. As different groups of genes are expressed, their packaging within chromatin changes and may be detected at the chromosomal level. The organization of DNA within a chromosome is determined to a large extent by the positively charged, highly conserved histones. Histone subtypes and the reversible chemical modifications of histones have been associated with gene activity. Active or potentially active genes have been associated with hyperacetylated histones and inactive genes with nonacetylated histones. Sodium butyrate increases the acetylation levels of histones in cell cultures and acts as both an inducer of gene activity and as a cell-cycle block. We describe a method to label the interphase distribution of DNA associated with various histone acetylation stages on chromosomes. Nucleosomes from untreated and butyrate-treated HeLa cells were fractionated by their acetylation level and the associated DNA labeled, and hybridized to normal human chromosomes. In the sodium butyrate-treated cells the resulting banding patterns of the high- and low-acetylated fractions were strikingly different. DNA from low-acetylated chromatin labeled several pericentric regions, whereas hybridization with DNA from highly acetylated chromatin resulted in a pattern similar to inverse G-bands on many chromosomes. The results from noninduced cells at both high and low acetylation levels were noticeably different from their induced counterparts. The capture and hybridization of DNA from interphase chromatin at different acetylation states provides a "snapshot" of the distribution of gene activity on chromosomes at the time of cell harvest.
Transcranial doppler (TCD) ultrasonography is often used to guide the management of patients with subarachnoid hemorrhage (SAH). However, the correlation between increased blood velocity as measured by TCD ultrasonography and angiographic vasospasm was established before the routine use of hypervolemia/hemodilution and administration of nimodipine and did not address blood flow. The relationship of blood velocity as measured by TCD ultrasonography and local cerebral blood flow (LCBF) in SAH managed with these modalities is unknown. Patients presenting with aneurysmal SAH between January 1992 and September 1993 who underwent TCD ultrasonography and xenon computed tomographic (Xe/CT) LCBF studies within 12 hours were retrospectively studied. Fifty patients underwent a total of 94 paired studies, encompassing 709 vascular territories. All were treated with nimodipine and hypervolemia/hemodilution. Hematocrit, blood pressure, and partial carbon dioxide pressure were similar at the time of TCD ultrasonography and Xe/CT measurement of LCBF. When LCBF in the middle cerebral artery (MCA) was < or = 31 ml/100 g/min, the corresponding peak systolic velocity measured by TCD ultrasonography was 119 cm/s, whereas those > 31 ml/100 g/min had a velocity of 169 cm/s (P = 0.006). High LCBF was associated with high velocity in all vascular territories, reaching significance in all but the internal carotid artery. At the time of each study, 41 neurological examinations were focal and 53 were nonfocal. The Xe/CT measurement of LCBF in the MCA contralateral to a deficit was significantly less than in territories without corresponding clinical deficits (P = 0.01), whereas peak systolic velocities in the MCA were not significantly different (P = 0.71). Territories with increases in blood velocity in the MCA of > 50 cm/s/24 h did not have statistically different LCBF (P = 0.183). Our results suggest that increased blood velocity revealed by TCD ultrasonography correlates with increased LCBF and not with ischemia. No difference in LCBF was found in territories with and without rapid increases in blood velocity in the MCA. Furthermore, although focal neurological deficits corresponded with decreased contralateral LCBF in the MCA, increased velocity did not correlate with neurological findings. Therapeutic decisions based solely on blood velocity revealed by TCD ultrasonography might be inappropriate and potentially harmful. Xe/CT studies of LCBF are useful in guiding the management of SAH.
PURPOSE: The purpose of this study was to determine whether the hemodynamic consequences of extracranial carotid disease correlate with the risk of subsequent cerebral infarction. METHODS: In 95 patients with symptoms who had greater than or equal to 70% stenosis (31 patients) or who had occlusion (64 patients) of the ipsilateral carotid artery, cerebral blood flow was measured by the stable xenon/computed tomography technique both at baseline and after vasodilatory challenge with intravenous acetazolamide. Patients were stratified into group 1, 43 patients with no more than a 5% decrease in flow in any vascular territory, and group 2, 52 patients with greater than a 5% decrease in one or more vascular territories after an acetazolamide challenge. RESULTS: In group 2, 15 (28.9%) of 52 patients had a new stroke, but only one (2.3%) of 43 patients in group 1 did (p = 0.0005). Of patients with total carotid occlusion 10 (26%) of 38 in group 2 and none (0%) of 26 in group 1 had a new stroke (p = 0.003). Of patients with greater than or equal to 70% stenosis, five (36%) of 14 in group 2 and only one (6%) of 17 in group 1 had a stroke (p = 0.067). CONCLUSION: The loss of cerebral reactivity in patients with symptoms who had greater than or equal to 70% carotid stenosis or occlusion is an important predictor of impending cerebral infarction.
We have assessed the measurement of oxygen saturation (SaO2) as a means of detecting aspiration in patients with stroke. For 10 weeks all acute stroke [AS] admissions were seen within 48 hours. Basal SaO2 was measured by pulse oximetry. Patients swallowed 10ml water while sitting up and SaO2 was noted for 2 minutes. Two control groups [young, fit (YF) and inpatient age- and sex-matched, non-neurological disease (IP)] underwent the same assessment. AS subjects underwent independent assessment of swallowing by a speech and language therapist (SLT). Exclusion criteria comprised impaired consciousness, other neurological disease and chest infection. Forty-nine AS subjects [20 men; aged 46-93 (mean 71) years], 55 YF [26 men; aged 18-55 (mean 32) years] and 65 IP [28 men; aged 53-96 (mean 71) years] were studied. Mean (SD) SaO2 fall in AS subjects [2.6 (2.9)%)] was significantly more than in YF [1.1 (0.8)%] or IP [1.1 (0.9)%]. The lower 95% confidence limit for variation in SaO2 did not differ between YF and IP (3.0% 'fall'); 19 (39%) AS subjects desaturated below this 95% lower confidence limit. Mean (SD) SaO2 fall was significantly more in SLT-graded 'aspirators' [4.6 (2.7)%] than 'nonaspirators' [1.4 (1.0)%]. We conclude that (1) a fall in SaO2 on swallowing fluid is common in patients with acute stroke; (2) the presence or absence of desaturation agrees statistically with SLT assessment of aspiration; (3) SaO2 measures may aid bedside assessment of swallowing.
For regulatory purposes nucleic acid vaccines are considered biological products and will be regulated by the Center for Biologics Evaluation and Research (CBER). Vaccines derived through the use of this technology may ultimately find broad application as preventive vaccines for infectious disease or as therapeutic vaccines for treatment of disease. The regulations that govern the use of biological products as well as other guidance documents available from CBER are applicable to the regulation of nucleic acid vaccines. The regulatory concerns associated with the manufacture, preclinical evaluation and clinical studies for these vaccines are similar to those for other biological products. The following discussion will provide an overview of the organization of CBER and how nucleic acid vaccines will be reviewed within this organization. This discussion will also describe the regulations encoded in the US Code of Federal Regulations which govern the use of biological products and additional guidance provided in Points to Consider Documents and in specific Guidelines. In addition, this discussion will note specific concerns regarding the manufacture, lot release and preclinical evaluation to assess the safety of polynucleotide vaccines. Finally, the process for submission of an Investigational New Drug application and the design of protocols for clinical studies will be described.
Haploid leaf tissue of tobacco cultivars K326 and K149 was transformed with several transgenes containing cDNA of the potato virus Y (PVY) coat protein (CP) open reading frame (ORF). The various transgenes containing the PVY CP ORF sequence produced (1) the expected mRNA and CP product, (2) an mRNA rendered untranslatable by introduction of a stop codon immediately after the initiation codon, or (3) an antisense RNA that was untranslatable as a result of the incorrect orientation of the PVY CP ORF behind the transcriptional promoter. Homozygous doubled haploid (DH) (diploid) plants were generated, and selfed progeny from these plants were examined. Resistance was virus specific, functioning only against PVY. An inverse correlation between transgene-derived PVY transcript steady state levels and resistance was generally noted with lines expressing the untranslatable sense version of the PVY CP ORF. A collection of DH lines, derived from a single transformation event of a common haploid plant and isogenic for the PVY transgenes expressing untranslatable sense RNA, displayed different levels of PVY resistance. Lines with actively transcribed, methylated transgene sequences had low steady state levels of transgene transcript and a virus-resistant phenotype. These results are discussed within the context of sense suppression in plants.
Varicella-zoster virus (VZV) open reading frame 4 (ORF4) encodes a putative immediate-early protein which is homologous to herpes simplex virus type 1 (HSV-1) ICP27 on the basis of gene location and similarity in amino acid sequence. In transient expression assays, however, ORF4 and ICP27 exhibit different properties. ICP27 alone has little activity on target plasmids, but it acts as a transactivator or a transrepressor in the presence of other HSV-1 transactivators. In contrast, ORF4 directly transactivates plasmids containing homologous or heterologous promoters and has no apparent transrepressing activity. To further illuminate the functional similarities and differences between ORF4 and ICP27, Vero cell lines which express ORF4 under the inducible metallothionein promoter were constructed. Cell lines expressing functionally active ORF4 protein upregulated the expression of transfected VZV target plasmids but were unable to efficiently complement HSV-1 ICP27 mutants. These results indicate that, despite structural similarities, VZV ORF4 and HSV-1 ICP27 behave differently in transient expression assays and may play different roles in virus replication.
BACKGROUND AND PURPOSE: A chronic compromise of cerebral hemodynamics has been shown to identify a group of patients at an increased risk for stroke. Because a "steal phenomenon" induced by a vasodilatory challenge has characterized the group at greatest risk, it was hypothesized that these individuals would also have a severe compromise of primary collaterals and an increased dependence on leptomeningeal collaterals. METHODS: Twenty-three patients with symptomatic cerebrovascular disease underwent angiography and xenon-enhanced computed tomographic cerebral blood flow studies before and after 1 g IV acetazolamide within 6 months of each other. Cerebral blood flow vasoreactivity was classified by whether cerebral blood flow increased (> 5%) or was unchanged (+/- 5%) (group 1) or fell by > 5% (group 2) in any vascular territory. Angiographic collateralization was classified into four types: normal (type 1), willisian (type 2), ophthalmic (type 3), and leptomeningeal (type 4). RESULTS: Twenty percent (2/10) of group 1 patients and 69% (9/13) of group 2 patients (P = .0009) had leptomeningeal collaterals. CONCLUSIONS: A negative flow reactivity is significantly associated with a dependence on leptomeningeal collaterals and implies a state of maximal hemodynamic compromise.