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Biomedical subjects

C M Harris

Publications and source records attributed to C M Harris.

At least 73 records · Page 4Linked to original sources

Signal-dependent noise determines motor planning.

When we make saccadic eye movements or goal-directed arm movements, there is an infinite number of possible trajectories that the eye or arm could take to reach the target. However, humans show highly stereotyped trajectories in which velocity profiles of both the eye and hand are smooth and symmetric for brief movements. Here we present a unifying theory of eye and arm movements based on the single physiological assumption that the neural control signals are corrupted by noise whose variance increases with the size of the control signal. We propose that in the presence of such signal-dependent noise, the shape of a trajectory is selected to minimize the variance of the final eye or arm position. This minimum-variance theory accurately predicts the trajectories of both saccades and arm movements and the speed-accuracy trade-off described by Fitt's law. These profiles are robust to changes in the dynamics of the eye or arm, as found empirically. Moreover, the relation between path curvature and hand velocity during drawing movements reproduces the empirical 'two-thirds power law. This theory provides a simple and powerful unifying perspective for both eye and arm movement control.

Animals↗

Highly mutagenic bypass synthesis by T7 RNA polymerase of site-specific benzo[a]pyrene diol epoxide-adducted template DNA.

We have previously developed an in vitro system that allows quantitative evaluation of the fidelity of transcription during synthesis on a natural template in the presence of all four nucleotides. Here, we have employed this system using a TAA ochre codon reversion assay to examine the fidelity of transcription by T7 RNA polymerase past an adenine residue adducted at the N6-position with (-)-anti-trans- or (+)-anti-trans-benzo[a]pyrene diol epoxide (BPDE). T7 RNAP was capable of transcribing past either BPDE isomer to generate full-length run-off transcripts. The extent of bypass was found to be 32% for the (-)-anti-trans-isomer and 18% for the (+)-anti-trans-isomer. Transcription past both adducts was highly mutagenic. The reversion frequency of bypass synthesis of the (-)-anti-trans-isomer was elevated 11,000-fold and that of the (+)-anti-trans-isomer 6000-fold, relative to the reversion frequency of transcription on unadducted template. Adenine was misinserted preferentially, followed by guanine, opposite the adenine adducted with either BPDE isomer. Although base substitution errors were by far the most frequent mutation on the adducted template, three- and six-base deletions were also observed. These results suggest that transcriptional errors, particularly with regard to damage bypass, may contribute to the mutational burden of the cell.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗

Styrene oxide adducts in an oligodeoxynucleotide containing the human N-ras codon 12: minor groove structures of the R(12,1)- and S(12,1)-alpha-(N2-guanyl) stereoisomers determined by 1H nuclear magnetic resonance.

The R- and S-alpha-(N2-guanyl)styrene oxide (SO) adducts at X5 in d(G1G2C3A4X5G6T7G8G9T10G11).d(C12A13C14C15A16C17C18T19G20C21C22 ), encompassing codon 12 of the human N-ras protooncogene (underlined), were examined using 1H NMR spectroscopy. These were the R(12,1) and S(12,1) adducts, indicating the location of the R or S adduct at the first position of codon 12. These differed from the R- and S(12, 2)-alpha-SO adducts [Zegar, I. S., Setayesh, F. R., DeCorte, B. L., Harris, C. M., Harris, T. M., and Stone, M. P. (1996) Biochemistry 35, 4334-4348] in that the base pair 5' to the lesion was changed from G.C to A.T, while the base pair 3' to the lesion was changed from T.A to G.C. Comparison of the R- and S(12,1) adducts with the R- and S(12,2) adducts allowed the effects of flanking bases on the conformations of the alpha-SO adducts to be examined. This change in flanking base affected the R-SO lesion. The R(12,1) adduct structure was disordered at the adduct site, and a refined structure could not be obtained. NOE and chemical shift data suggested that the styrenyl moiety was oriented in the minor groove and in the 3'-direction from the site of adduction. In contrast, this change in flanking base did not affect the S-SO lesion. The S(12,1) adduct yielded a refined structure, with the styrenyl moiety edgewise in the minor groove and oriented in the 5'-direction relative to the site of adduction. A total of 232 interproton distances, including 13 styrene-DNA distances, were obtained. A total of 12 NOE-restrained molecular dynamics calculations converged with pairwise root-mean-square deviation of 1.10 A. The sixth-root residual index between calculated and experimental NOE intensities was 8.0 x 10(-)2 A. The styrene aromatic protons appeared as three resonances, suggesting rapid rotation. The possibility of a hydrogen bond between the styrene hydroxyl and C18 O2 in the S(12,1) adduct could not be confirmed. This work illustrates the dual roles of stereochemistry and sequence in modulating the properties of guanine N2 alpha-SO adducts.

Codon↗

Investigating relationships between a range of potential indicators of general practice prescribing: an observational study.

OBJECTIVES: (i) to describe the variation in a number of potential prescribing indicators across a large number of general practices, (ii) to determine whether particular indicators are associated with higher or lower overall prescribing costs, (iii) to determine how particular indicators are correlated with each other, (iv) to determine whether the indicators are stable over time. DESIGN: Observational study based on the analysis of PACT (Prescribing Analysis and Cost) data for general practices. SETTING: The Trent region of England. SUBJECTS: 809 general practices. MAIN OUTCOME MEASURES: Variations in a range of potential prescribing performance indicators; correlations between these indicators and NIC (net ingredient costs) per ASTRO-PU (Age, sex and temporary resident originated prescribing unit); change in indicators between the financial year 1994-5 and 1995-6. RESULTS: Wide variations were found between practices in a range of potential prescribing performance indicators. There were positive correlation (P < 0.001) between NIC per ASTRO-PU and indicators of relatively high cost prescribing within different therapeutic areas. There were negative correlations (P < 0.001) with indicators of relatively low cost prescribing. There were positive intercorrelations (P < 0.001) between indicators of relatively high cost prescribing. For most indicators there were significant changes (P < 0.001) in values between the two years. CONCLUSION: There are considerable variations between general practices in prescribing performance indicators and the indicators show changes over time. We have demonstrated correlations between different indicators that lend support to the idea that such indicators might act as proxies for more general prescribing patterns. However, most of the correlations we observed were not strong. Further research is needed into the development and use of indicators based on PACT data.

Drug Prescriptions↗

Blood transfusion and total joint replacement surgery: T helper 2 (TH2) cytokine secretion and clinical outcome.

Surgery and blood transfusions have both been reported to cause decreases in various measures of cell-mediated immunity. A study of in vitro T helper lymphocyte type 2 (Th2) cytokine secretion after major joint replacement surgery was performed because these cytokines (IL4 and IL10) generally down-regulate cellular immune function. Th1 cytokines such as IL2 tend to up-regulate cellular immunity. Forty-three patients undergoing elective joint replacement surgery had pre- and multiple post-operative levels of IL2, IL4 and IL10 secretion measured and analysed with regard to demographic and clinical outcome data. Total joint replacement alone without allogeneic transfusions led to substantial increases in peak mean IL4 (2.1 times the pre-operative level) and IL10 secretion in vitro (4.3-fold) compared with much more modest increases in IL2 (1.36-fold) (P < 0.0001 for changes from baseline for each cytokine). In 14 patients who received allogeneic transfusions, these changes were greater than those in recipients of only autologous blood for IL4 (5.0-fold; P = 0.0036 vs. no allogeneic transfusion) and IL10 (15.7-fold; P = 0.079) but not for IL2 (1.38-fold; P = 0.38). The dramatic increase in Th2 cytokine secretion and minimal change in Th1 cytokine secretion after total joint replacement, with or without allogeneic transfusions, was seen regardless of type of anaesthetic, duration of surgery or whether knee or hip replacement occurred. These changes in cytokine patterns may contribute to the decreases in cellular immune function seen after surgery. Allogeneic transfusions but not autologous transfusions appear to exacerbate this immune deviation toward a T helper 2 (Th2) type response, and thus probably contribute to down-regulation of cellular immunity in the setting of joint replacement surgery.

Aged↗

Familial congenital saccade initiation failure and isolated cerebellar vermis hypoplasia.

The underlying lesion in congenital saccade initiation failure (c-SIF) ('congenital ocular motor apraxia', 'Cogan's apraxia') is uncertain. Often no abnormality can be found, yet in others a midline cerebellar abnormality has often been reported. We examined this cerebellar association in a brother and sister. In addition to standard ophthalmological and neurological examinations, both siblings underwent ocular motor testing and neuroradiological investigations including CT and MRI. Both siblings exhibited the typical signs of c-SIF, including headthrusting, synkinetic blinking, missed-nystagmus quick phases, mild developmental delay, and speech difficulties. CT and MRI revealed cerebellar vermis hypoplasia in the brother, but appeared normal in the sister. No other neuroradiological abnormalities were detected. These cases highlight the wide variability in the association of vermis abnormalities with c-SIF, despite the inheritance and similar clinical manifestations. They show that either: (1) the vermis is causal in saccade triggering, but that c-SIF may result from very subtle damage that is beyond MRI resolution in some cases; or (2) that a vermis abnormality per se is not causative but only a marker of another subtle abnormality, either structural or possibly biochemical.

Apraxias↗

Saccadic strategies in children with hemianopia.

Multiple hypometric (undershooting) saccades are generally reported as a compensatory strategy in adults with homonymous hemianopia. However, hypermetric (overshooting) saccades have been reported to develop spontaneously as a beneficial strategy in response to predictable targets. We examined the saccades of 10 children (aged 5 to 16 years) with homonymous hemianopia to determine the type of compensatory eye-movement strategies employed 6 months to 16 years after hemianopia onset. Homonymous hemianopia was identified using perimetry and/or pattern visual evoked potentials and supported with results of neuroimaging. Eye movements were recorded using bitemporal electrooculography. Saccades were elicited to a red light source in a semipredictable paradigm. We found that hypermetria was not a consistent compensatory strategy in our patients. In spite of the predictability of our paradigm and the long follow-up period, multiple hypometric saccades into the blind field appeared to be the preferred strategy.

Adolescent↗

Physician use of an ambulatory medical record system: matching form and function.

An ambulatory medical record system was implemented as a pilot project to determine the clinical and operational impacts of using interactive, computer-based systems in the outpatient setting. Of the 58 physicians who attempted to use the AMR system, 28 continue as active users. Physicians who discontinued use of the system reported a variety of reasons for their dissatisfaction. As a result of the pilot, the AMR implementation team and physicians understand better how to fit the AMR system to a physician's specific requirements, as well as enhance the operation of the physician's practice. Armed with the experience gained during the early phases of the implementation, the Cleveland Clinic Foundation now intends a cautious expansion and continued evaluation of the AMR system, with the objective of meeting organizational needs with appropriate technical functions.

Ambulatory Care Information Systems↗

Lack of correlation between in vitro and in vivo replication of precisely defined benz-a-anthracene adducted DNAs.

Like other polycyclic aromatic hydrocarbons, certain metabolites of benz[a]anthracene have been implicated as potent carcinogens. These effects are thought to be caused by the covalent binding of these species to nucleophilic groups on the bases of DNA. To address the molecular mechanisms by which these molecules induce mutations, this study employed oligonucleotides containing four site-specific N6 adenine-benz[a]anthracene diol epoxide adducts. Using a prokaryotic in vivo replication system, we have shown that both non-bay region anti-trans-benz[a]anthracene adducts are essentially nonmutagenic. In contrast, the bay region anti-trans-benz[a]anthracene lesions do induce point mutations at the adduct site. The mutagenic frequency of these bay region lesions is dependent on the stereochemistry about the adduct-forming bond, as well as the strain of Escherichia coli in which they are replicated. The ability of the bacterial replication machinery to bypass the lesions does not correlate with the differences observed in their mutagenesis. While both non-bay region adducts are readily bypassed in vivo, the bay region adducts are both blocking to approximately the same degree. In vitro studies of the interactions of E. coli DNA polymerase III with these adducts have also been undertaken to further dissect the relationship between adduct structure and biological activity.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗

Effect of doctors' ethnicity and country of qualification on prescribing patterns in single handed general practices: linkage of information collected by questionnaire and from routine data.

OBJECTIVES: To test whether Asian general practitioners who qualified in the Indian subcontinent prescribe items more often, more expensive items, and fewer generic drugs than their British trained Asian and non-Asian counterparts. DESIGN: Linkage study using data collected by questionnaire and from routine sources. SETTING: General practices in England. SUBJECTS: 155 single handed general practitioners: 42 Asian doctors qualified in United Kingdom (group 1), 58 white doctors qualified in United Kingdom (group 2), and 55 Asian doctors qualified in Indian subcontinent (group 3). MAIN OUTCOME MEASURES: Prescribing cost (cost per ASTRO-PU), prescribing frequency (number of items per ASTRO-PU), and generic prescribing (percentage of drugs prescribed that are generic). RESULTS: Doctors in group 1 were significantly younger than those in the other groups and had a higher proportion of patients who were from deprived wards. There was no difference between the groups in the proportion of female doctors and total list size. After adjustment for confounding factors, there were no significant differences between the three groups for prescribing cost (16.58 (95% confidence interval 6.39 to 26.77) for group 1, 17.31 (6.92 to 27.69) for group 2, 17.80 (7.22 to 28.38) for group 3, P = 0.55); prescribing frequency (6.58 (4.60 to 8.40), 6.45 (4.70 to 8.30), 7.89 (6.16 to 9.64), P = 0.34); and generic prescribing (44.44 (38.95 to 49.93), 47.41 (42.12 to 52.70), 44.04 (38.75 to 49.33), P = 0.37). CONCLUSIONS: Asian doctors qualified from the Indian subcontinent did not differ from British trained doctors in their prescribing practice. This study refutes the common belief that Asian doctors are high volume and high cost prescribers.

Accreditation↗

Altered expression and activity of G1/S cyclins and cyclin-dependent kinases characterize squamous cell carcinomas of the head and neck.

Progressive deregulation of the cell-division cycle is thought to contribute to the establishment and progression of neoplasia. Previously, we have documented the in vivo inactivation of p16INK4A, an inhibitor of G1 cyclin-dependent kinases, in squamous cell carcinomas of the head and neck region. In the present study, we extend these findings by examining the expression and functional activity of cyclin-dependent kinases (CDKs) and their regulatory subunits using a model system of cell lines derived from squamous cell carcinomas. Increased activity of CDK4 and 6 was universal in tumor cells compared with normal keratinocytes, reflecting over-expression of either or both kinases. In contrast to other studies, over-expression of cyclin D1, a regulatory subunit of CDK4 and 6, was not observed. Increased activity of CDK2 was less frequent and was related to over-expression of cyclin A and/or E. All tumor cell lines showed increased expression of proliferating cell nuclear antigen compared to normal keratinocytes. Four SCC cell lines, including one tumor-metastasis pair derived from a single patient, failed to express the p15INK4B transcript. Western blot analysis of cell lysates revealed normal or reduced levels of p27KIP1 in tumor cells compared to normal keratinocytes. However, failure to express wild-type p53 was not reflected by lower levels of p21WAF1. Our data suggest that cell-cycle deregulation is likely to occur by multiple mechanisms during the genesis of head and neck squamous cell carcinomas. Furthermore, p16INK4A is likely to be the primary target for inactivation on chromosome 9p21 in these tumors as p15INK4B loss occurs less frequently.

Carcinoma, Squamous Cell↗

The oxidative half-reaction of xanthine dehydrogenase with NAD; reaction kinetics and steady-state mechanism.

The reaction between reduced xanthine dehydrogenase (XDH) from bovine milk and NAD has been studied in detail. An understanding of this reaction is necessary for a complete description of XDH turnover with its presumed natural electron acceptor and to address the preference of XDH for NAD over oxygen as a substrate. The reaction between pre-reduced XDH and NAD was studied by stopped-flow spectrophotometry. The reaction was found to involve two rounds of oxidation with 2 eq of NAD. The first round goes to completion, and the second round reaches a slightly disfavored equilibrium. Rapid binding of NAD with an apparent Kd of 25 +/- 2 microM is followed by NAD reduction at a rate constant of 130 +/- 13 s-1. NADH dissociation at a rate constant of 42 +/- 12 s-1 completes a round of oxidation. These steps have been successfully tested and modeled to repeat themselves in the second round of oxidation. The association rate constant for NAD binding was estimated to be much greater than any rate constant measured in the oxidation by molecular oxygen, thus explaining how NAD competes with oxygen for reducing equivalents. Rate constants for NAD reduction and NADH dissociation are respectively 21- and 7-fold greater than kcat, indicating that the reductive half-reaction of the enzyme by xanthine is mostly rate-limiting in xanthine/NAD turnover. A steady-state mechanism for XDH is discussed.

Animals↗

Epileptic nystagmus in infancy.

Epileptic nystagmus (EN) is a rare form of nystagmus that occurs only during epileptic seizures. We report an infantile case in which EN was first noted at 10 days of age. Electronystagmography showed a right-beating nystagmus with predominantly linear slow phases that traversed the midline. Neuro-imaging revealed dysplasia of the left middle temporal gyrus extending posteriorly into the parieto-occipital cortex. The right hemisphere and subcortical structures appeared normal. Perfusion studies demonstrated interictal hypoperfusion with ictal hyperperfusion in the left temporal lobe. Electrocorticography demonstrated spiking over the left temporal-parieto-occipital region. Following extensive surgical resection of this area and weaning of anti-convulsants, the child has remained seizure-free without nystagmus. This case demonstrates the cortical origin of EN, and shows that infant cortex has functioning efferent connections to brainstem oculomotor centres from 10 days of age.

Electroencephalography↗

Kinetic isotope effects and electron transfer in the reduction of xanthine oxidoreductase with 4-hydroxypyrimidine. A comparison between oxidase and dehydrogenase forms.

Isolated from bovine milk, xanthine oxidase (XO) and xanthine dehydrogenase (XDH) are two interconvertible forms of the same protein, differing in the number of protein cysteines versus cystines. Most differences between XO and XDH are localized to the FAD center, the site at which the oxidizing substrates NAD and molecular oxygen react. A comparative study of the reduction of XO and XDH has been performed to assess differences in reactivity of the molybdopterin site, as well as subsequent electron-transfer events from molybdenum to 2Fe/2S and FAD centers. The compound 4-hydroxypyrimidine (4-OH-P) was chosen as reducing substrate because its higher Km value raised the possibility of binding weak enough to measure kinetically, and its high kcat value could allow detection of intramolecular electron-transfer reactions. As measured by stopped flow spectrophotometry, XO and XDH react with the first equivalent of 4-OH-P via similar mechanisms, differing in the magnitude of rate and dissociation constants. Using [2-2H]4-OH-P as substrate, a D(k/Kd) isotope effect of 1.9 to 2.3 suggests that movement of the hydrogen abstracted from substrate appreciably limits the rate of initial enzyme reduction from Mo(VI) to Mo(IV). Monitoring the visible spectrum of the enzymes, the first observed step is reduction of a single 2Fe/2S center and presumably re-oxidation of Mo(IV) to Mo(V). This suggests a common pathway for electron transfer involving reduction of a 2Fe/2S center prior to reduction of the second 2Fe/2S and FAD centers. Rates of the first electron transfer from molybdenum to the 2Fe/2S center are rapid, 290 s-1 with XO and 180 s-1 with XDH, and are consistent with rates measured by flash photolysis (Walker, M. C., Hazzard, J. T., Tollin, G., and Edmondson, D. E. (1991) Biochemistry 30, 5912-5917) allowing discrete observation of the electron-transfer reactions that occur during turnover. This step also exhibits a modest primary kinetic isotope effect of 1.5 to 1.6 when [2-2H]4-OH-P is used, possibly due to deprotonation of the molybdenum center prior to electron transfer. A second one-electron transfer, presumably oxidizing Mo(V) to Mo(VI), follows in a step coincident with product dissociation, consistent with a role for product release in controlling electron transfer events. The kinetics of this complex system are described and interpreted quantitatively in models that are consistent with all the data.

Animals↗

The reaction of reduced xanthine dehydrogenase with molecular oxygen. Reaction kinetics and measurement of superoxide radical.

Xanthine dehydrogenase (XDH) from bovine milk contains significant activity in xanthine/oxygen turnover assays. The oxidative half-reaction of XDH with molecular oxygen has been studied in detail, at 25 degrees C, pH 7.5, to determine the basis of the preference of XDH for NAD over oxygen as oxidizing substrate. Spectral changes of XDH accompanying oxidation were followed by stopped-flow spectrophotometry. The amount of superoxide radicals formed during oxidation was investigated to assess the ability of XDH to catalyze production of oxygen radicals. Reduced XDH reacts with oxygen in at least 4 bi-molecular steps, with 1.7-1.9 mol of superoxide per mol of XDH formed from the last 2 electrons oxidized. A model is discussed in which the flavin hydroquinone transfers electrons to oxygen to produce hydrogen peroxide at a rate constant of at least 72,000 M-1 s-1, whereas flavin semiquinone reduces oxygen to form superoxide as slow as 16 M-1 s-1. Steady-state kinetics of xanthine/oxygen and NADH/oxygen turnover of XDH were determined to have kcat values of 2.1 +/- 0.1 and 2.5 +/- 0.9 s-1, respectively, at 25 degrees C, pH 7.5. XDH is therefore capable of catalyzing the formation of reduced oxygen species at one-third the rate of xanthine/NAD turnover, 6.3 s-1 (Hunt, J., and Massey, V. (1992) J. Biol. Chem. 267, 21479-21485), in the absence of NAD. As XDH contains a significant and intrinsic xanthine oxidase activity, estimates of relative amounts of XO and XDH based solely upon turnover assays must be made with caution. Initial-rate assays containing varying amounts of xanthine, NAD, and oxygen indicate that at 100% oxygen saturation, NADH formation is only inhibited at concentrations of xanthine and NAD below Km for each substrate.

Animals↗

Differential tolerance to DNA polymerization by HIV-1 reverse transcriptase on N6 adenine C10R and C10S benzo[a]pyrene-7,8-dihydrodiol 9,10-epoxide-adducted templates.

To determine the effect of various stereoisomers of benzo[a]pyrene-7,8-dihydrodiol 9,10-epoxide (BPDE) on translesion bypass by human immunodeficiency virus-1 reverse transcriptase and its alpha-helix H mutants, six 33-mer templates were constructed bearing site- and stereospecific adducts. This in vitro model system was chosen to understand the structure-function relationships between the polymerase and damaged DNA during replication. Comparison of the replication pattern between wild type human immunodeficiency virus-1 reverse transcriptase and its mutants, using primers which were 3' to the lesion, revealed essentially similar patterns. While these primers terminated with all three of the C10R and two of the C10S BPDE-adducted templates 1 base 5' and 1 base 3' to the damaged site respectively, (+)-anti-trans-(C10S) BPDE-adducted DNA alone permitted the formation of full-length products. Utilization of a primer with its 3'-hydroxyl 1 base beyond the lesion resulted in full-length products with all the C10S BPDE-adducted templates and the (-)-syn-trans-(C10R)-BPDE-adducted template, following replication with either the wild type or mutant enzymes. However, the other two C10R BPDE-adducted templates failed to allow any primer extension, even with the wild type enzyme. Although T.P depletion studies further confirmed the differential primer extension abilities using the C10R and C10S adducted templates, their binding affinities were similar, yet distinct from the unadducted template.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗