Biomedical subjects

E L Burns

Publications and source records attributed to E L Burns.

14 recordsLinked to original sources

PubMed1996

Random mutagenesis of the sheep Na,K-ATPase alpha1 subunit generating the ouabain-resistant mutant L793P.

The polymerase chain reaction was used to randomly mutagenize a cDNA cassette encoding amino acids 691-946 of the sheep Na,K-ATPase alpha subunit. The mutagenized cassettes were used to replace the wild-type region in the full-length cDNA, and pools of mutants were transfected into HeLa cells. After the generation of resistant cells via selection in 0.5 microM ouabain, polymerase chain reaction was used to amplify the mutagenized cassette from the genomic DNA of the stable transfectants. Sequence analysis of the polymerase chain reaction product revealed three amino acid substitutions: I729V, L793P, and K836R. Subsequent site-directed mutagenesis experiments showed that only L793P was important for resistance. To elucidate the role of L793 in ouabain inhibition, additional mutations at this position were prepared. L793A and L793I mutants were constructed and expressed in HeLa cells. Only L793A survived selection using ouabain, which suggested that resistance is not due to the specific substitution of leucine with proline. To explore the mechanism of resistance, apparent affinities of the L793P mutant for sodium and potassium were compared to the wild-type HeLa pump. Although the apparent affinities were comparable for sodium, the mutant had a 2-fold higher apparent affinity for potassium. This suggests that the mechanism of ouabain insensitivity of L793P is due to a perturbation in the region of the enzyme that may include the K+ binding site.

Amino Acid Sequence↗

PubMed1994

Patterns of biosynthesis and accumulation of hydrocarbons and contact sex pheromone in the female German cockroach, Blattella germanica.

De novo synthesis of contact female sex pheromone and hydrocarbons in Blattella germanica was examined using short in vivo incubations. Accumulation of pheromone on the epicuticular surface and the internal pheromone titer were related to age-specific changes in hydrocarbon synthesis and accumulation in normal and allatectomized females. The incorporation of radiolabel from [1-14C]propionate into the cuticular methyl ketone pheromone fraction was positively related to corpora allata activity during two gonotrophic cycles. During peak pheromone production the total internal lipid fraction contained greater titers of pheromone than the cuticular surface, and it too exhibited a cycle internally, preceding the rise in external pheromone. This suggests that synthesis and accumulation of pheromone internally are followed by transport of pheromone to the epicuticular surface where it accumulates. Radiolabel was incorporated efficiently into both cuticular and internal hydrocarbons after the imaginal molt and until the peak of pheromone synthesis, but it declined to lower levels before ovulation and throughout pregnancy. The internal hydrocarbon titer decreased 58% after oviposition, suggesting deposition in the egg case. It remained relatively unchanged during pregnancy and increased again during the second gonotrophic cycle. In allatectomized females, hydrocarbon synthesis was reduced relative to control females until oviposition in the latter. However, subsequent rates of hydrocarbon synthesis in allatectomized females (without oothecae) exceeded the rates in sham-operated females (with oothecae). In the absence of ovarian uptake of hydrocarbons, the internal titer increased without the decline found in control females at oviposition. As internal hydrocarbons increased, so did cuticular hydrocarbons and both internal and cuticular methyl ketone pheromones. These patterns corresponded well with feeding patterns in sham-operated and allatectomized females, suggesting that pheromone production is normally regulated by stage-specific feeding-induced hydrocarbon synthesis (precursor accumulation internally) and juvenile hormone-induced conversion of hydrocarbon to pheromone. They also suggest that both the cuticle and the ovaries might be target sites for hydrocarbon and possibly methyl ketone deposition.

Aging↗

PubMed1993

Random mutagenesis of the sheep Na,K-ATPase alpha-1 subunit generates a novel T797N mutation that results in a ouabain-resistant enzyme.

Site-directed mutagenesis of the Na,K-ATPase has provided a rational approach to identify several amino acids that appear to be involved in ouabain sensitivity. In order to identify additional amino acids that play a role in ouabain binding, we used formic acid to randomly mutagenize a cDNA cassette encoding amino acids 691-946 of the sheep Na,K-ATPase alpha subunit. This was then used to replace the wild type cDNA cassette in the full-length cDNA, and pools of mutants were electroporated into HeLa cells. Ouabain-resistant cells were selected in 0.5 microM ouabain, and the polymerase chain reaction was used to amplify the region corresponding to the mutagenized cassette from the genomic DNA of the resistant cells. Sequence analysis of the polymerase chain reaction product revealed a single amino acid substitution, T797N. Site-directed mutagenesis was used to reproduce this change in the wild type sheep alpha subunit, and this construct was able to confer resistance to HeLa cells, verifying the role of the mutation in ouabain resistance. The mutant sheep enzyme was found to be as resistant to ouabain as is the rat Na,K-ATPase. These data suggest that T797N, predicted to be in the fifth putative transmembrane domain, is involved in the interaction between ouabain and the Na,K-ATPase.

Amino Acid Sequence↗

PubMed1991

Sexual differentiation of nymphal corpora allata and the effects of ovariectomy on adult gland morphometrics in Blattella germanica.

Changes in the number of corpus allatum (CA) cells were investigated in nymphs and in intact and ovariectomized adult female Blattella germanica. The CA of intact adult females exhibit cyclic changes in volume in relation to juvenile hormone (JH) synthesis, while the CA of ovariectomized females become significantly hypertrophied as a result of a gradual and continuous increase in volume that is independent of JH biosynthesis. In both intact and ovariectomized females changes in JH synthesis and CA volume are not related to total cell number which remains relatively constant. However, adult females have twice as many CA cells as do adult males as a result of a female-specific increase in total cell number late in the last nymphal instar.

Aging↗

PubMed1991

Cyclic juvenile hormone biosynthesis in the cockroach, Blattella germanica: effects of ovariectomy and corpus allatum denervation.

The corpora allata (CA) of ovariectomized adult Blattella germanica females exhibited delayed but high rates of juvenile hormone biosynthesis in vitro. Using the onset of sexual receptivity as a probe of the degree of CA activation in females, we demonstrated at least one cycle of CA activity in the experimentally synchronized ovariectomized females. Following their activation, the CA exhibited a partial and transient decline in activity, but in contrast to the CA of intact females, this decline was not accompanied by a regression in CA volume. CA of intact and ovariectomized females that were denervated from the brain were activated, but the subsequent decline in CA activity at the end of the cycle was prevented in ovariectomized females. The presence of an egg-case suppressed the reactivation of the inactive CA in intact females but not in CA-denervated females. We conclude that activation of the CA in B. germanica is not dependent upon either the presence of the ovary or intact nervous connections between the CA and the brain. The brain exerts a partial inhibition on CA activity through intact nerves which is relieved (by disinhibition) in the presence of a young ovary but is enhanced and sustained in the presence of the egg-case. Inhibition of the CA also occurs independently of nervous connections with the brain through factors that originate in the mature ovary and affect both CA activity and morphology.

Animals↗

PubMed1991

Developmental regulation of juvenile hormone synthesis: ovarian synchronization of volumetric changes of corpus allatum cells in cockroaches.

The corpus allatum (CA) cells of adult Blattella germanica females undergo cyclic volumetric changes in relation to juvenile hormone (JH) synthesis. In intact females the size of CA cells changes synchronously during the gonotrophic cycle, resulting in cyclic JH synthesis. In ovariectomized females volumetric changes among CA cells become asynchronous, resulting in highly variable but high rates of JH synthesis. Injection of the steroid hormone 20-hydroxyecdysone into ovariectomized females with active CA resulted in a transient decline followed by an increase in both CA volume and JH biosynthesis. This response was due to a change in the size distribution of CA cells and not in the total number of CA cells. In ovariectomized females, CA cells can be re-synchronized into a uniform population of small inactive cells with injection of 20-hydroxyecdysone and implantation of an artificial egg-case, mimicking the successive events of ovulation, oviposition and pregnancy.

Animals↗

PubMed1991

Absentee patterns among OR staff. Identifying aspects of the problem.

Absenteeism continues to be a great concern to hospital administrators. Nursing personnel should have a clear idea of acceptable attendance parameters based on written policies that are routinely administered. Efforts are now directed toward exploring strategies that reward personnel for good attendance records. Early development of professional attitudes related to absenteeism should continue to be supported and fostered. Implications from this preliminary study indicate that it may be more cost-effective to hire RNs than STs to work in ORs.

Absenteeism↗

PubMed1974