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Biomedical subjects

D Smith

Publications and source records attributed to D Smith.

At least 199 records · Page 11Linked to original sources
PubMed2000

Evidence for adaptive response and implication in pulse-simulated low-dose-rate radiotherapy.

PURPOSE: Pulsed-dose-rate (PDR) brachytherapy as a substitute for continuous low-dose-rate (LDR) brachytherapy has a number of clinical advantages. However, early results show that some cells can exhibit an adaptive response to radiation and in PDR where many pulses are given such an adaptive response may play an important role in the outcome. METHODS AND MATERIALS: Nine human cell lines (two normal fibroblast and seven tumor) were evaluated for an adaptive response. Cells were given either a single adapting dose before a challenge dose or given PDR sequences for which the average dose rate matched the LDR dose rate. Response was assessed using the colony survival assay. RESULTS: Five of the nine cell lines showed an adapting response to single small doses of radiation. Three of these cell lines were further investigated for adapting response to PDR and two of the three lines (one ovarian carcinoma and one glioma) showed an adaptive response which was dependent on pulse size and interval. CONCLUSION: The data show that an adaptive response can occur in human cells and that it can vary among cell lines. In addition, PDR sequences also produced an adaptive response which could have an affect on PDR therapy if such a response is found in tissues.

Adaptation, Physiological↗
PubMed2000

Pedicle morphology in thoracic adolescent idiopathic scoliosis: is pedicle fixation an anatomically viable technique?

STUDY DESIGN: A radiographic study of thoracic pedicle anatomy in a group of adolescent idiopathic scoliosis (AIS) patients. OBJECTIVE: To investigate the anatomic constraints of the thoracic pedicles and determine whether the local anatomy would routinely allow pedicle screw insertion at every level. SUMMARY OF BACKGROUND DATA: In spite of the clinical successes reported with limited thoracic pedicle screw-rod constructs for thoracic AIS, controversy exists as to the safety of this technique. MATERIAL AND METHODS: Twenty-nine patients with right thoracic AIS underwent preoperative thoracic CT scans and plain radiographs. Anatomic parameters were measured from T1 to T12. RESULTS: Information on 512 pedicles was obtained. The transverse width of the pedicles from T1 through T12 ranged from 4.6-8.25 mm. The medial pedicle to lateral rib wall transverse width from T1 through T2 ranged from 12.6 to 17.9 mm. Measured dimensions from the CT scans showed the actual pedicle width to be 1-2 mm larger than would have been predicted from the plain radiographs. Age, Risser grade, curve magnitude, and the amount of segmental axial rotation did not correlate with the morphology or size of the thoracic pedicles investigated. In no case would pedicle morphology have precluded the passage of a pedicle screw. CONCLUSION: Based on the data identified in this group of adolescent patients, it is reasonable to consider pedicle screw insertion at most levels and pedicle-rib fixation at all levels of the thoracic spine during the treatment of thoracic AIS.

Adolescent↗
PubMed2000

Virological and immunological effects of combination antiretroviral therapy with zidovudine, lamivudine, and indinavir during primary human immunodeficiency virus type 1 infection.

Forty-seven patients presenting with primary human immunodeficiency virus (HIV) infection were treated with zidovudine 200 mg 3 times a day, lamivudine 150 mg 2 times a day, and indinavir 800 mg 3 times a day for 1 year. From a mean pretreatment viral RNA level of 4.93 log(10) copies/mL, the proportions of patients having <500 copies/mL at 24 and 52 weeks were 92.0% and 89.2%, respectively. For the 35 patients with data available at 24 and 52 weeks, the corresponding proportions for the <50 copies/mL analysis were 86.6% and 79.3%, respectively. The change in virus load was -2.19 and -2.41 log(10) copies/mL at weeks 8 and 52, respectively. CD4 cell counts increased, from a mean of 546 cells/mm(3), by 142 cells/mm(3) at week 24 and by 210 cells/mm(3) at week 52. Three patients discontinued the study because of drug-related toxicity. Six (12.8%) patients had adverse experiences associated with nephrolithiasis. Combination therapy with zidovudine, lamivudine, and indinavir during primary HIV infection results in a profound and sustained reduction in virus load with concurrent recovery of the CD4 cell population.

Adult↗
PubMed2000

Identification of domains responsible for signal recognition and transduction within the QUTR transcription repressor protein.

QUTR (qutR-encoded transcription-repressing protein) is a multi-domain repressor protein active in the signal-transduction pathway that regulates transcription of the quinic acid utilization (qut) gene cluster in Aspergillus nidulans. In the presence of quinate, production of mRNA from the eight genes of the qut pathway is stimulated by the activator protein QUTA (qutA-encoded transcription-activating protein). Mutations in the qutR gene alter QUTR function such that the transcription of the qut gene cluster is permanently on (constitutive phenotype) or is insensitive to the presence of quinate (super-repressed phenotype). These mutant phenotypes imply that the QUTR protein plays a key role in signal recognition and transduction, and we have used deletion analysis to determine which regions of the QUTR protein are involved in these functions. We show that the QUTR protein recognizes and binds to the QUTA protein in vitro and that the N-terminal 88 amino acids of QUTR are sufficient to inactivate QUTA function in vivo. Deletion analysis and domain-swap experiments imply that the two C-terminal domains of QUTR are mainly involved in signal recognition.

Base Sequence↗
PubMed2000

The effect of alpha-tocopherol on the oxidative cleavage of beta-carotene.

Two cleavage pathways of beta-carotene have been proposed, one by central cleavage and the other by random (excentric) cleavage. The central cleavage pathway involves the metabolism of beta-carotene at the central double bond (15, 15') to produce retinal by beta-carotene 15, 15'-dioxygenase (E.C.888990988). The random cleavage of beta-carotene produces beta-apo-carotenoids, but the mechanism is not clear. To understand the various mechanisms of beta-carotene cleavage, beta-carotene was incubated with the intestinal postmitochondrial fractions of 10-week-old male rats for 1 h, and cleavage products of beta-carotene were analyzed using reverse-phase, high-performance liquid chromatography (HPLC). We also studied the effects of alpha-tocopherol and NAD(+)/NADH on beta-carotene cleavage. In addition to beta-carotene, we used retinal and beta-apo-14'-carotenoic acid as substrates in these incubations. Beta-apo-14'-carotenoic acid is the two-carbon longer homologue of retinoic acid. In the presence of alpha-tocopherol, beta-carotene was converted exclusively to retinal, whereas in the absence of alpha-tocopherol, both retinal and beta-apo-carotenoids were formed. Retinoic acid was produced from both retinal and beta-apo-14'-carotenoic acid incubations only in the presence of NAD(+). Our data suggest that in the presence of an antioxidant such as alpha-tocopherol, beta-carotene is converted exclusively to retinal by central cleavage. In the absence of an antioxidant, beta-carotene is cleaved randomly by enzyme-related radicals to produce beta-apo-carotenoids, and these beta-apo-carotenoids can be oxidized further to retinoic acid via retinal.

Animals↗
PubMed2000

CEP-1347 increases ChAT activity in culture and promotes cholinergic neurone survival following fimbria-fornix lesion.

Recent evidence suggests that the activation of the Jun N-terminal kinase (JNK) signal transduction pathway may be important in neuronal responses to stresses such as trophic factor deprivation. Preventing the activation of JNK and expression of c-Jun may, therefore, be neuroprotective. Here, we report that the small molecule CEP-1347, which has been shown to inhibit the JNK signalling pathway, promotes cholinergic activity in cultured embryonic septal neurones. In vivo, we have shown that CEP-1347, administered either by sub-cutaneous (s.c.) injection or by continuous infusion, is partially neuroprotective, for cholinergic neurones in the medial septum, following fimbria-fornix transection. These data suggest that small molecules such as CEP-1347 may have beneficial effects in treating neurodegenerative diseases.

Animals↗
PubMed2000

Deficient long-term memory and long-lasting long-term potentiation in mice with a targeted deletion of neurotrophin-4 gene.

We examined the learning and memory of neurotrophin-4 (NT4)-/- mice by using fear conditioning. In both cue and context conditioning, we found significant deficits in the NT4 mutants at 2 and 24 h after training but not at 30 min. Hippocampal slices from the mutant mice showed normal basal synaptic transmission, short-term plasticity, and decremental long-term potentiation (LTP) at the Schaffer collateral-CA1 synapses. These findings, together with the normal short-term memory, suggest that the hippocampal development of NT4-/- mice is largely unaffected. However, consistent with the long-term memory defects, the long-lasting LTP at the same synapses was attenuated significantly in the mutant mice. Our results suggest that NT4 plays a physiological role essential for hippocampus- and amygdala-dependent long-term memory and hippocampal long-lasting LTP and that NT4 may be useful in the therapy of acquired disorders of learning and memory.

Amygdala↗
PubMed2000

Preclinical studies with the photon radiosurgery system (PRS).

PURPOSE: Determine the radiobiological effectiveness (RBE) for low-energy X-rays (average energy of 23 KeV) produced by the Photon Radiosurgery System (PRS). METHODS AND MATERIALS: RBE values were assessed by comparison with survival data obtained for cells irradiated with either low-energy X-rays from a GE Maxitron 100 machine or high-energy photons from a clinically used Varian 6 MV LINAC. The output of the GE and PRS sources was determined using Baldwin-Farmer and Markus thin window ionization chambers calibrated with 50 kVp X-rays and cross-checked against figures supplied by Photoelectron Corporation. The dose-rate for the PRS was 1.2 Gy/min at a distance of 35 mm with a field flatness of +/-2%. RESULTS: The RBE for the PRS low-energy X-ray source (at 1-mm depth) was greater than either the GE or Varian machines and varied with cell survival. For Chinese hamster ovary (CHO) cells, the PRS was 1.25 and 3.3 times more effective than 90 kVp X-rays and 6 MeV photons at 0.5% cell survival, respectively; by comparison, the PRS was 1.2 and 1.9 times more effective at 0.05% cell survival, respectively. Similar RBE values of 1.4 and 1.2 were obtained for human U373 and T98 glioblastoma cells grown in vitro irradiated with the PRS or GE sources, respectively. Other studies showed that the RBE for the PRS low-energy X-ray source increased with depth. The RBEs for the PRS source at 1-mm and 4-mm depth were 1.2 and 2.5 (0.5% survival) and 1. 2 and 1.9 (0.05% survival). CONCLUSIONS: The biological and physical properties of the PRS low-energy X-rays offer, under the right conditions, a significant advantage for patient treatment over conventional external beam, stereotactic, or brachytherapy treatment.

Animals↗
PubMed2000

Long-term immunological response in HIV-1-infected subjects receiving potent antiretroviral therapy.

OBJECTIVE: To determine the long-term T-lymphocyte response to highly active antiretroviral therapy (HAART) and to define predictors of the immunological response. DESIGN: Cohort study, including 135 HIV-1-infected subjects at a city general practice who commenced HAART between 1996 and 1998. METHODS: Collection of plasma HIV-1 RNA, CD4+ and CD8+ T-lymphocyte data at 3-6 monthly time intervals over 2 years. RESULTS: Seventy-three subjects (54%) achieved suppression of plasma HIV-1 RNA to levels below 400 copies/ml during the observation period, 31 individuals (23%) had detectable plasma HIV-1 RNA below 10,000 copies/ml and 31 subjects (23%) had virological failures with viral loads above 10,000 copies/mL. Median CD4+ T lymphocytes increased from 246 to 463 x 10(6) cells/l, showing a median rise of 20 x 10(6) cells/l per month in the first 3 months and 7 x 10(6) cells/l per month thereafter. The proportion of individuals who reached CD4+ cell counts above 500 x 10(6) cells/l increased from 8% at baseline to 54% at 2 years. Treatment-naïve individuals, subjects with a large reduction of HIV-1 RNA or a large early CD8+ increase had better early CD4+ responses. Long-term CD4+ T-cell increases were inversely correlated with mean plasma HIV-1 RNA levels. Baseline CD4+ T-cell count was the most important determinant of reaching CD4+ cell counts above 500 x 10(6) cells/l. Nineteen per cent of subjects had no further CD4+ T-cell increases in the second year of therapy despite undetectable viral load. CONCLUSIONS: Immune reconstitution is a slow process, showing a large individual variability. The virological response to HAART was the most important determinant of the immunological short- and long-term response.

Adult↗
PubMed2000

Satellite measurements of sea surface temperature through clouds

Measurements of sea surface temperature (SST) can be made by satellite microwave radiometry in all weather conditions except rain. Microwaves penetrate clouds with little attenuation, giving an uninterrupted view of the ocean surface. This is a distinct advantage over infrared measurements of SST, which are obstructed by clouds. Comparisons with ocean buoys show a root mean square difference of about 0.6 degrees C, which is partly due to the satellite-buoy spatial-temporal sampling mismatch and the difference between the ocean skin temperature and bulk temperature. Microwave SST retrievals provide insights in a number of areas, including tropical instability waves, marine boundary layer dynamics, and the prediction of hurricane intensity.

Journal Article↗
PubMed2000

Enzymatic amplification staining for flow cytometric analysis of cell surface molecules.

BACKGROUND: Flow cytometric analysis is a powerful technique for the single cell assessment of cell surface expression of selected molecules. The major deficiency of flow cytometry has been its relative insensitivity. Only molecules expressed in abundance have been readily observed. METHODS: We have developed an enzymatic amplification procedure for the analysis of cell surface molecules by flow cytometry. Transformed and nontransformed cells expressing MHC class I, CD5, CD3, CD4, CD6, CD7, CD34, CD45, MHC class II, Fas ligand, and phosphatidylserine were assessed. RESULTS: Our enzymatic amplification technology increased the fluorescence signal between 10 and 100-fold for all surface molecules tested. CONCLUSIONS: Enzymatic amplification staining produces a significant enhancement in the resolving power of flow cytometric analysis of cell surface molecules. Using this technique, we have been able to detect the presence of molecules that could not be observed by the standard procedure.

Annexin A5↗
PubMed2000

Retrotransposition of a bacterial group II intron.

Self-splicing group II introns may be the evolutionary progenitors of eukaryotic spliceosomal introns, but the route by which they invade new chromosomal sites is unknown. To address the mechanism by which group II introns are disseminated, we have studied the bacterial L1.LtrB intron from Lactococcus lactis. The protein product of this intron, LtrA, possesses maturase, reverse transcriptase and endonuclease enzymatic activities. Together with the intron, LtrA forms a ribonucleoprotein (RNP) complex which mediates a process known as retrohoming. In retrohoming, the intron reverse splices into a cognate intronless DNA site. Integration of a DNA copy of the intron is recombinase independent but requires all three activities of LtrA. Here we report the first experimental demonstration of a group II intron invading ectopic chromosomal sites, which occurs by a distinct retrotransposition mechanism. This retrotransposition process is endonuclease-independent and recombinase-dependent, and is likely to involve reverse splicing of the intron RNA into cellular RNA targets. These retrotranspositions suggest a mechanism by which splicesomal introns may have become widely dispersed.

Bacterial Proteins↗
PubMed2000

Clustering of beta(2)-integrins on human neutrophils activates dual signaling pathways to PtdIns 3-kinase.

The beta(2)-integrins on leukocytes can serve as a signaling unit during cell adhesion and locomotion, and to further clarify this important property we investigated the possible mechanisms of beta(2)-integrin-induced activation of PtdIns 3-kinase. It has previously been demonstrated that clustering of beta(2)-integrins activates p21(ras) by a tyrosine kinase-dependent pathway, and here we show that active p21(ras) interacts with its downstream target, PtdIns 3-kinase. Engagement of beta(2)-integrins also activates the tyrosine kinases p58(c-fgr) and p59/61(hck) and causes them to associate with the p85 subunit of PtdIns 3-kinase. These findings suggest a mechanism whereby p58(c-fgr) and p59/61(hck) are directly involved in the activation of PtdIns 3-kinase. No coupling between p58(c-fgr) and p59/61(hck) could be detected; hence these kinases probably trigger independent but parallel signals to PtdIns 3-kinase. The effect of beta(2)-integrin clustering on PtdIns 3-kinase activity was monitored as the activation of protein kinase B (PKB). Stimulation of PKB by beta(2)-integrins was abolished by genistein and wortmannin but not by using methyl transferase inhibitors to abrogate the influence of p21(ras)-related proteins. Thus, even if PtdIns 3-kinase is not activated by p21(ras), it can maintain full enzyme activity due to the mentioned interaction with p58(c-fgr) or p59/61(hck). These tyrosine kinases apparently activate similar pathways that operate in parallel and therefore have the potential to substitute for each other in mediating adhesion and regulating cell locomotion.

Antibodies, Monoclonal↗
PubMed2000

Signals from trunk paraxial mesoderm induce pronephros formation in chick intermediate mesoderm.

We used Pax-2 mRNA expression and Lim 1/2 antibody staining as markers for the conversion of chick intermediate mesoderm (IM) to pronephric tissue and Lmx-1 mRNA expression as a marker for mesonephros. Pronephric markers were strongly expressed caudal to the fifth somite by stage 9. To determine whether the pronephros was induced by adjacent tissues and, if so, to identify the inducing tissues and the timing of induction, we microsurgically dissected one side of chick embryos developing in culture and then incubated them for up to 3 days. The undisturbed contralateral side served as a control. Most embryos cut parallel to the rostrocaudal axis between the trunk paraxial mesoderm and IM before stage 8 developed a pronephros on the control side only. Embryos manipulated after stage 9 developed pronephric structures on both sides, but the caudal pronephric extension was attenuated on the cut side. These results suggest that a medial signal is required for pronephric development and show that the signal is propagated in a rostral to caudal sequence. In manipulated embryos cultured for 3 days in ovo, the mesonephros as well as the pronephros failed to develop on the experimental side. In contrast, embryos cut between the notochord and the trunk paraxial mesoderm formed pronephric structures on both sides, regardless of the stage at which the operation was performed, indicating that the signal arises from the paraxial mesoderm (PM) and not from axial mesoderm. This cut also served as a control for cuts between the PM and the IM and showed that signaling itself was blocked in the former experiments, not the migration of pronephric or mesonephric precursor cells from the primitive streak. Additional control experiments ruled out the need for signals from lateral plate mesoderm, ectoderm, or endoderm. To determine whether the trunk paraxial mesoderm caudal to the fifth somite maintains its inductive capacity in the absence of contact with more rostral tissue, embryos were transected. Those transected below the prospective level of the fifth somite expressed Pax-2 in both the rostral and the caudal isolates, whereas embryos transected rostral to this level expressed Pax-2 in the caudal isolate only. Thus, a rostral signal is not required to establish the normal pattern of Pax-2 expression and pronephros formation. To determine whether paraxial mesoderm is sufficient for pronephros induction, stage 7 or earlier chick lateral plate mesoderm was cocultured with caudal stage 8 or 9 quail somites in collagen gels. Pax-2 was expressed in chick tissues in 21 of 25 embryos. Isochronic transplantation of stage 4 or 5 quail node into caudal chick primitive streak resulted in the generation of ectopic somites. These somites induced ectopic pronephroi in lateral plate mesoderm, and the IM that received signals from both native and ectopic somites formed enlarged pronephroi with increased Pax-2 expression. We conclude that signals from a localized region of the trunk paraxial mesoderm are both required and sufficient for the induction of the pronephros from the chick IM. Studies to identify the molecular nature of the induction are in progress.

Animals↗
PubMed2000

Drug-mediated toxicity caused by genetic deficiency of UDP-glucuronosyltransferases.

Human gene families encoding UDP-Glucuronosyltransferases (UGTs) have been identified and partially characterised. This family of enzymes catalysed the glucuronidation of drugs, xenobiotics and endobiotics. Genetic mutations and polymorphisms have been identified in several UGT genes and examples should be anticipated in all UGT genes. A common genetic defect in the TATA box promoter of the UGT1A1 gene is associated with Gilbert's Syndrome (GS) causing mild hyperbilirubinaemia. Recently, adverse effects of anticancer agents have been observed in Gilbert's patients due to reduced drug or bilirubin glucuronidation.

Antineoplastic Agents↗
PubMed2000

Rules for DNA target-site recognition by a lactococcal group II intron enable retargeting of the intron to specific DNA sequences.

Group II intron homing occurs primarily by a mechanism in which the intron RNA reverse splices into a DNA target site and is then reverse transcribed by the intron-encoded protein. The DNA target site is recognized by an RNP complex containing the intron-encoded protein and the excised intron RNA. Here, we analyzed DNA target-site requirements for the Lactococcus lactis Ll.LtrB group II intron in vitro and in vivo. Our results suggest a model similar to yeast mtDNA introns, in which the intron-encoded protein first recognizes a small number of nucleotide residues in double-stranded DNA and causes DNA unwinding, enabling the intron RNA to base-pair with the DNA for reverse splicing. Antisense-strand cleavage requires additional interactions between the protein and 3' exon. Key nucleotide residues are recognized directly by the intron-encoded protein independent of sequence context, and there is a stringent requirement for fixed spacing between target site elements recognized by the protein and RNA components of the endonuclease. Experiments with DNA substrates containing GC-clamps or "bubbles" indicate a requirement for DNA unwinding in the 3' exon but not the distal 5' exon region. Finally, by applying the target-site recognition rules, we show that the L1.LtrB intron can be modified to insert at new sites in a plasmid-borne thyA gene in Escherichia coli. This strategy should be generally applicable to retargeting group II introns and to delivering foreign sequences to specific sites in heterologous genomes.

Base Pairing↗