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Biomedical subjects

J E Fleming

Publications and source records attributed to J E Fleming.

At least 19 recordsLinked to original sources
PubMed1992

Role of oxidative stress in Drosophila aging.

We review the role that oxidative damage plays in regulating the lifespan of the fruit fly, Drosophila melanogaster. Results from our laboratory show that the lifespan of Drosophila is inversely correlated to its metabolic rate. The consumption of oxygen by adult insects is related to the rate of damage induced by oxygen radicals, which are purported to be generated as by-products of respiration. Moreover, products of activated oxygen species such as hydrogen peroxide and lipofuscin are higher in animals kept under conditions of increased metabolic rate. In order to understand the in vivo relationship between oxidative damage and the production of the superoxide radical, we generated transgenic strains of Drosophila melanogaster that synthesize excess levels of enzymatically active superoxide dismutase. This was accomplished by P-element transformation of Drosophila melanogaster with the bovine cDNA for CuZn superoxide dismutase, an enzyme that catalyzes the dismutation of the superoxide radical to hydrogen peroxide and water. Adult flies that express the bovine SOD in addition to native Drosophila SOD are more resistant to oxidative stresses and have a slight but significant increase in their mean lifespan. Thus, resistance to oxidative stress and lifespan of Drosophila can be manipulated by molecular genetic intervention. In addition, we have examined the ability of adult flies to respond to various environmental stresses during senescence. Resistance to oxidative stress, such as that induced by heat shock, is drastically reduced in senescent flies. This loss of resistance is correlated with the increase in protein damage generated in old flies by thermal stress and by the insufficient protection from cellular defense systems which includes the heat shock proteins as well as the oxygen radical scavenging enzymes. Collectively, results from our laboratory demonstrate that oxidative damage plays a role in governing the lifespan of Drosophila during normal metabolism and under conditions of environmental stress.

Aging↗
PubMed1992

Outcome, prognosis, and risk in a longitudinal follow-up study.

This study reports the results of a 4-year follow-up of a community sample of children who were ages 4 to 12 in 1983 at the first wave of data collection. Results on outcomes revealed that conduct disorder showed the greatest stability especially from late childhood to early adolescence. In multivariate analyses, both family dysfunction and problems getting along with others significantly predicted the persistence of one or more psychiatric disorders 4 years later, and low income predicted one or more psychiatric disorders among children free of disorder 4 years earlier. The implications of the results for the child psychiatric field, especially prevention, are discussed.

Adolescent↗
PubMed1992

Predicting substance use in late adolescence: results from the Ontario Child Health Study follow-up.

OBJECTIVE: The purpose of this study was to evaluate the relationship between use of tobacco, alcohol, marijuana, and hard drugs (substance use) and psychiatric disorder in early adolescence and substance use in late adolescence. METHOD: Adolescents included in the study were identified by means of a household sampling frame and participated in the Ontario Child Health Study in 1983 and the follow-up in 1987. There were 726 12-16-year-olds (369 boys and 357 girls) in 1983 who had complete information in 1987. Data on substance use were collected from adolescents by using a structured, self-administered questionnaire. Data on psychiatric disorder were collected in 1983 from both adolescents and their parents by using problem checklists to assess conduct disorder, attention deficit disorder, and emotional disorder. RESULTS: Prior substance use in 1983 was associated strongly with subsequent use in 1987. Among the psychiatric disorders assessed in 1983, only conduct disorder made an independent contribution to predicting use of marijuana (relative odds = 3.46) and other hard drugs (relative odds = 6.82) in 1987, after prior use of these substances and coexisting attention deficit and emotional disorders were controlled. Corresponding estimates of attributable risk (the expected contribution of exposure to conduct disorder to the development of substance use) were 5.7% and 11.1%, respectively. CONCLUSIONS: Although a statistically significant relationship existed between conduct disorder in early adolescence and use of marijuana and hard drugs in late adolescence, the potential is limited for preventing substance use in the general population by treating conduct disorder early on.

Adolescent↗
PubMed1992

Stress resistance of Drosophila transgenic for bovine CuZn superoxide dismutase.

Several oxidative and non-oxidative stresses were applied to two transgenic strains of Drosophila melanogaster (designated P(bSOD)5 and P(bSOD)11) that express superoxide dismutase (SOD) at elevated levels, and control strains that express normal SOD levels. Transgenic strain P(bSOD)5 exposed to paraquat (1,1'-dimethyl-4,4'-bipyridinium dichloride), a redox cycling agent that generates superoxide anion when metabolized in vivo, was significantly more resistant to this xenobiotic than control flies. When test flies were subjected to 100% oxygen for 20 min each day, the mean lifespan was 3.62 days for control strain 25, but 4.35 days for both transgenic strains. The mortality curves of strains fed 1% H2O2 were similar, but the median lifespan of 72 h for controls and 64 h for transgenics suggests that the transgenic flies were slightly more sensitive to H2O2. The activity of catalase was the same for all strains. Using starvation resistance as a non-oxidative stress, flies maintained on water without any food had identical survival curves; for all strains, the median lifespan was 72 h. Throughout the lifespan, no statistically significant difference in physical activity was displayed for transgenic versus control flies. Collectively, these data suggest that the increased lifespan previously observed in SOD transgenics is specifically related to resistance to oxidative stresses.

Animals↗
PubMed1992

Changes in superoxide dismutase and catalase in aging heat-shocked Drosophila.

We have examined the age-dependent expression of CuZn superoxide dismutase (SOD) and catalase (CAT) in Drosophila melanogaster following heat shock. Quantitative northern blot analysis was performed after heat shock on 2-, 23- and 49-day-old flies, using a 0.48 kb Sal1 EcoR1 fragment of the Drosophila SOD cDNA and a 1.4 kb fragment of the human catalase cDNA. Heat shock induction was monitored with a 5.4 kb DNA fragment of the Drosophila hsp70 gene. After exposure to 37 degrees C for 30 min and 60 min, the level of SOD RNA in young flies was elevated above that of nonstressed conditions. Changes in the transcription of SOD gene with age paralleled the expression of hsp70 RNA. The SOD RNA was elevated in heat-shocked middle aged (23-25 days old) flies compared to young Drosophila (2 days old), then it decreased in 49-50-day-old flies. The relative expression of CAT RNA did not change with age or after heat shock. Enzymatic activities of these two antioxidant enzymes were evaluated in nondenaturing polyacrylamide electrophoresis gels. SOD migrates on this gel as three different electromorphs. These were designated as fast, intermediate and slow migrating bands. The most intense activity was associated with the fast band in these flies. In the absence of heat shock, there was an age-dependent decrease in the intermediate, but not in the slow or fast bands. Heat shock does not affect the intensity of the fast band in young or old flies, however; in middle aged flies, there is a shift in this band toward the slow position. No change was detected in the activity of catalase with age or heat shock, although flies of all ages exhibited a shift toward a faster-migrating electromorph with increasing time of heat shock. This effect was also observed in flies fed H2O2 and is more pronounced in insects fed higher concentrations. These results are discussed in relation to the role of these antioxidant enzymes in protecting against age-induced oxidative stresses.

Aging↗
PubMed1992

Drosophila with postponed aging as a model for aging research.

Species of the genus Drosophila, commonly known as "fruitflies," are good model systems for research in aging. Drosophila are extremely well-known genetically, developmentally, and otherwise. They are also genetically analogous to mammalian species in most important respects. Previous work with Drosophila has been hampered by inbreeding depression, but more recent work using selection has created Drosophila with postponed aging that is inherited normally. Genetic transformation has also increased Drosophila life spans in some cases. Several biologic approaches have been applied to the analysis of genetically postponed aging in Drosophila: quantitative genetics, organismal physiology, and protein electrophoresis. Ultimately, these different approaches will be integrated into an overall analysis of aging in Drosophila, one that could be valuable for research with other taxa as well.

Aging↗
PubMed1991

Aging affects expression of 70-kDa heat shock proteins in Drosophila.

We examined the effect of cellular aging on adult mortality and hsp70 gene expression in Drosophila melanogaster under thermal stress. The results showed that flies exposed to 37 degrees C for various time intervals had reduced survival rate with age. The level of hsp70 mRNA increases in flies up to 23-28 days of age, but then declines as they get older. When flies are shifted to 25 degrees C after 30 min of heat stress, the time-dependent decrease in hsp70 mRNA levels occurs more rapidly in young flies than in old ones. The hsp70 mRNA present during this recovery period is translated into protein, and senescent flies continue to synthesize this protein for up to 5 h after heat shock. The prolonged expression of hsp70 RNA during recovery from heat shock was also observed in young flies fed canavanine, an arginine analogue. These data suggest that in old insects, the accumulation of conformationally altered proteins plays a role in the regulation of hsp70 RNA expression. These results are discussed in relation to the finding that old flies are more sensitive to thermal stress than young ones.

Aging↗
PubMed1991

Genetic differentiation of Drosophila melanogaster populations as assessed by two-dimensional electrophoresis.

Seven populations of Drosophila melanogaster, representing a worldwide distribution, were compared using two-dimensional protein gel electrophoresis. A total of 611 protein spots was scored, which probably represent a sample of over 500 loci that were surveyed. Of the protein spots scored, 521 spots were found to be invariant, but another 90 spots were found to be variable among the populations. Of these variable protein spots, 12 were found to be present in only one population. All the populations, except one, had at least one protein spot restricted to itself. However, the Japanese population had by far the most, with five protein spots restricted to this one population, which has been observed in previous studies of private alleles in oriental populations. The mean genetic similarity (F) found among the seven populations was 0.965, with a range of between 0.956 and 0.977. This is similar to previous reports of lower variation found in population genetic surveys using two-dimensional electrophoresis. It was found that the historical relationships among these populations was somewhat congruent with the geographic distribution of the populations, but as in previous studies, it was not exactly coincident.

Animals↗
PubMed1991

Expression of bovine superoxide dismutase in Drosophila melanogaster augments resistance of oxidative stress.

Superoxide dismutases (SOD) play a major role in the intracellular defense against oxygen radical damage to aerobic cells. In eucaryotes, the cytoplasmic form of the enzyme is a 32-kDa dimer containing two copper and two zinc atoms (CuZn SOD) that catalyzes the dismutation of the superoxide anion (O2-) to H2O2 and O2. Superoxide-mediated damage has been implicated in a number of biological processes, including aging and cancer; however, it is not certain whether endogenously elevated levels of SOD will reduce the pathological events resulting from such damage. To understand the in vivo relationship between an efficient dismutation of O2- and oxidative injury to biological structures, we generated transgenic strains of Drosophila melanogaster overproducing CuZn SOD. This was achieved by microinjecting Drosophila embryos with P-elements containing bovine CuZn SOD cDNA under the control of the Drosophila actin 5c gene promoter. Adult flies of the resulting transformed lines which expressed both mammalian and Drosophila CuZn SOD were then used as a novel model for evaluating the role of oxygen radicals in aging. Our data show that expression of enzymatically active bovine SOD in Drosophila flies confers resistance to paraquat, an O2(-)-generating compound. This is consistent with data on adult mortality, because there was a slight but significant increase in the mean lifespan of several of the transgenic lines. The highest level of expression of the active enzyme in adults was 1.60 times the normal value. Higher levels may have led to the formation of toxic levels of H2O2 during development, since flies that died during the process of eclosion showed an unusual accumulation of lipofuscin (age pigment) in some of their cells. In conclusion, our data show that free-radical detoxification has a minor by positive effect on mean longevity for several strains.

Animals↗
PubMed1990

Changes in protein turnover after heat shock are related to accumulation of abnormal proteins in aging Drosophila melanogaster.

Adult Drosophila melanogaster kept at 24 degrees C show a progressive decline in the synthesis and degradation of proteins with age. After exposure of young, 7-10 days old flies to 20 min of heat shock at 37 degrees C, the incorporation of [35S]-methionine into trichloroacetic acid precipitable proteins decreases to more than 60% of that observed in non-stressed flies. This decrease is also accompanied by a lower protein degradation rate. In contrast, the same stress in old, 49 days old insects results in a 3-fold increase in protein synthesis as compared to either non-heat shocked senescent flies or to young heat-shocked flies. The older flies also have faster protein turnover than unshocked controls. An effect similar to that observed in senescent Drosophila also occurs in young flies that have been fed canavanine, an arginine analogue, before and during heat shock. These results suggest that an age dependent accumulation of abnormal proteins may be responsible for the changes in protein turnover observed in the heat-shocked old flies.

Aging↗
PubMed1990

Post-extrasystolic potentiation in a human fetus detected during measurement of systolic time intervals in labour.

Measurement of the pre-ejection period (PEP), a non-invasive index of cardiac function, was made in a human fetus with a disturbance of cardiac rhythm manifested by: (i) intermittent mechanically ineffective extrasystoles (ventricular premature beats) and (ii) pauses. The mean interval between the sinus beats before and following extrasystoles was virtually the same as that before and following pauses. The PEP was shortened at the first sinus beat following both extrasystoles and pauses, but to a greater extent following extrasystoles. The PEP was still shortened at the second sinus beat following extrasystoles, but prolonged at the same cardiac cycle following pauses. These data demonstrate that fetal cardiac post-extrasystolic potentiation was taking place and that this was due in part to inotropic potentiation of the post-extrasystolic beats. Inotropic potentiation of post-extrasystolic beats is due to a fundamental property of cardiac muscle relating frequency of stimulation to strength of contraction (the so-called 'force-interval' relationship). Our data present the first evidence, to our knowledge, that the inotropic state of the human fetal heart in vivo is modulated by this mechanism.

Cardiac Complexes, Premature↗
PubMed1990

Ascorbic acid uptake by young and aged guinea pig lenses.

The dynamics of in vitro uptake of [14C]-ascorbic acid into intact lenses of young (50 days old) and old (730 days old) guinea pigs was evaluated in this study. Two-dimensional protein gel analysis of [35S]-methionine labeled proteins provided evidence that the isolated lenses were viable throughout the culture period. It was found that the in vitro uptake of ascorbate into these lenses follows similar saturation kinetics for lenses from both age groups. Moreover, the linear uptake rate was identical. Ascorbate transport was inhibited by phloretin, p-chloromercuribenzoate, insulin, glucose and ouabain; however, no inhibition was observed with 2,4-dinitrophenol or NaF. These results suggest that the lenticular ascorbate concentration is regulated by a facilitated diffusion process and it not energy-dependent. Data on ascorbate transport into lenses from aged guinea pigs provide strong evidence that the abnormally low ascorbic acid concentrations in lenses of old mammals is not likely the result of a decreased lenticular uptake ability.

Aging↗
PubMed1990

Epidemiology of childhood depressive disorders: a critical review.

The methodology of 14 recent epidemiological studies of childhood and adolescent depressive disorders was critically reviewed and findings summarized for prevalence, comorbidity, correlates, risk factors, and outcome. Shortcomings in sampling and considerable inconsistency in the measurement of depression in the studies made it difficult to draw firm conclusions about the prevalence and correlates of depression in young people. Nonetheless, it is likely that major depressive disorder is relatively uncommon in prepubertal children, increases in frequency in adolescents, and is significantly associated with such variables as family dysfunction and low self-esteem. Comorbidity of depression with other psychiatric disorders was also high in these nonpatient samples and it will be important in future work to assess the implications of this for the etiology, treatment, and prognosis of depressive disorders in children and adolescents.

Adolescent↗
PubMed1989

Preparation of antibodies against xanthine oxidase from human milk.

1. Human xanthine oxidase [XO; EC 1.2.3.2.] was isolated by a non-proteolytic method from fresh human milk. Final purification of the protein was achieved by hydroxyapatite chromatography. Most (less than 95%) of the enzyme was released in the 0.40 M phosphate fraction at pH 6.8. 2. The specific activity of this preparation was found to be 0.047 microM min-1 mg-1 with xanthine as substrate. 3. Sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) separated two subunits, each with a mol. wt approximately 122 kDa. 4. On non-denaturing acrylamide gels both of these subunits exhibited oxidase-like activity with xanthine as substrate in the presence of nitroblue tetrazolium and molecular oxygen. 5. Immunoconjugates of XO were prepared by the keyhole limpet hemocyanin (KLH)- and glutaraldehyde-crosslinking techniques. 6. Polyclonal antibodies to XO were raised by i.m. injection of these conjugates into female New Zealand rabbits. 7. Western blot analysis using the semi-dry technique was employed to confirm the specificity of the antibody.

Animals↗
PubMed1989

Ontario Child Health Study. Summary of selected results.

Selected results from the Ontario Child Health Study (OCHS), a cross-sectional community survey of Ontario children four to 16 years of age, are presented in the areas of prevalence, risk indicators and service utilization. The six month prevalence of one or more of four psychiatric disorders (conduct disorder, hyperactivity, emotional disorder, and somatization), in children four to 16 years of age, in Ontario was 18.1%. The highest rate was in 12 to 16 year old girls, and the lowest rate in four to 11 year old girls. Co-morbidity among these four disorders was high while the proportion of disorders identified by more than one respondent was low. Psychiatric disorders co-occurred significantly with other morbidities in children, including poor school performance, chronic health problems, substance use and suicidal behaviour. Chronic medical illness in the child as well as single parent status, living in a family on social assistance and residing in subsidized housing, were all strong indicators of increased rates of psychiatric disorders in children. Specialized mental health/social services, over a six month period, reached fewer than one of five children with psychiatric disorders, as measured in the study. In contrast, ambulatory medical care (primarily visits to family doctors and pediatricians) served almost 60% of Ontario children four to 16 years old, over the same six month period. The results are compared with those in the literature.

Adolescent↗
PubMed1989

Prevalence of childhood and adolescent depression in the community. Ontario Child Health Study.

Data from a cross-sectional community survey of 2852 children were used to provide estimates of the prevalence of a 'DSM-III-like' major depressive syndrome in children aged 6 to 16. The severity of symptoms required to define a 'case' was varied to generate three levels of diagnostic certainty (DC). The overall estimates of prevalence made with high DC were 0.6% for pre-adolescents and 1.8% for adolescents. Corresponding rates determined with medium DC were 2.7% and 7.8%, whereas the figures for low DC were 17.5% and 43.9%. Utilisation of mental health and social services, comorbidity (combined disorders), poor school performance, problems in getting along with others and need for professional help all increased as diagnostic certainty increased. There was wide disagreement in data supplied by the different groups of respondents, i.e. parents, teachers and adolescents.

Adolescent↗
PubMed1988

Aging results in an unusual expression of Drosophila heat shock proteins.

We used high-resolution two-dimensional polyacrylamide gel electrophoresis to evaluate the effect of aging on the heat shock response in Drosophila melanogaster. Although the aging process is not well understood at the molecular level, recent observations suggest that quantitative changes in gene expression occur as these fruit flies approach senescence. Such genetic alterations are in accord with our present data, which clearly show marked differences in the synthesis of heat shock proteins between young and old fruit flies. In 10-day-old flies, a heat shock of 20 min results in the expression of 14 new proteins as detectable by two-dimensional electrophoresis of [35S]methionine-labeled polypeptides, whereas identical treatment of 45-day-old flies leads to the expression of at least 50 new or highly up-regulated proteins. In addition, there is also a concomitant increase in the rate of synthesis of a number of the normal proteins in the older animals. Microdensitometric determinations of the low molecular weight heat shock polypeptides on autoradiographs of five age groups revealed that their maximum expression occurs at 47 days for a population of flies with a mean life span of 33.7 days. Moreover, a heat shock effect similar to that observed in senescent flies occurs in young flies fed canavanine, an arginine analogue, before heat shock.

Aging↗