Aspects of audit. 3: Audit in British general practice.

Compared with hospitals, general practice has many handicaps in developing medical audit. This is especially true with regard to defining objectives and outcomes of care. Many methods have been proposed to overcome these problems, and several audit studies that have been published are presented here. Further work should concentrate on evaluating the effectiveness of audit in bringing about change rather than on generating information.     

Blocking of acidosis-mediated apoptosis by a reduction of lactate dehydrogenase activity through antisense mRNA expression.

Lactic acid produced from the cells is a potential cause of extra- and intracellular acidification. Due to scarce technical tools, lactic acid that leads to acidification could not be reduced and direct evidence of the relationship between metabolic lactate and apoptosis has not yet been elucidated. In this study, we designed a cellular pH regulation system in CHO cells by a reduction of lactate dehydrogenase (LDH) activity through LDH antisense mRNA expression. This inhibited lactate production and, therefore, acidification of the cytosol. Under HCO3(-)-buffered growth conditions, both the parent CHO cells and the engineered CHO cells maintained their extracellular pH and intracellular pH fairly well. However, upon acidification of the cytosol, only the parent CHO cells underwent apoptosis under HCO3(-)-free conditions. In fact, we observed a number of apoptosis-related events only in control cells, including mitochondrial dysfunction, cytochrome c release, and an increase in caspase-3 enzymatic activity.     

DNA cleavage and religation by human topoisomerase II alpha at high temperature.

A common DNA religation assay for topoisomerase II takes advantage of the fact that the enzyme can rejoin cleaved nucleic acids but cannot mediate DNA scission at suboptimal temperatures (either high or low). Although temperature-induced DNA religation assays have provided valuable mechanistic information for several type II enzymes, high-temperature shifts have not been examined for human topoisomerase IIalpha. Therefore, the effects of temperature on the DNA cleavage/religation activity of the enzyme were characterized. Human topoisomerase IIalpha undergoes two distinct transitions at high temperatures. The first transition occurs between 45 and 55 degrees C and is accompanied by a 6-fold increase in the level of DNA cleavage at 60 degrees C. It also leads to a loss of DNA strand passage activity, due primarily to an inability of ATP to convert the enzyme to a protein clamp. The enzyme alterations that accompany the first transition appear to be stable and do not revert at lower temperature. The second transition in human topoisomerase IIalpha occurs between 65 and 70 degrees C and correlates with a precipitous drop in the level of DNA scission. At 75 degrees C, cleavage falls well below amounts seen at 37 degrees C. This loss of DNA scission appears to result from a decrease in the forward rate of DNA cleavage rather than an increase in the religation rate. Finally, similar high-temperature alterations were observed for yeast topoisomerase II and human topoisomerase IIbeta, suggesting that parallel heat-induced transitions may be widespread among type II topoisomerases.     

Origin of the cell nucleus,mitosis and sex: roles of intracellular coevolution

Background  

The transition from prokaryotes to eukaryotes was the most radical change in cell organisation since life began, with the largest ever burst of gene duplication and novelty. According to the coevolutionary theory of eukaryote origins, the fundamental innovations were the concerted origins of the endomembrane system and cytoskeleton, subsequently recruited to form the cell nucleus and coevolving mitotic apparatus, with numerous genetic eukaryotic novelties inevitable consequences of this compartmentation and novel DNA segregation mechanism. Physical and mutational mechanisms of origin of the nucleus are seldom considered beyond the long-standing assumption that it involved wrapping pre-existing endomembranes around chromatin. Discussions on the origin of sex typically overlook its association with protozoan entry into dormant walled cysts and the likely simultaneous coevolutionary, not sequential, origin of mitosis and meiosis.     

Cloning of genes that suppress an Escherichia coli K-12 alanine auxotroph when present in multicopy plasmids.

To facilitate molecular analyses of a previously uncharacterized gene involved in alanine synthesis, attempts were made to clone the wild-type allele of this gene, alaA, with a mini-Mu plasmid element used for in vivo cloning. Seventy-six independent Ala+ plasmids were isolated and characterized. Physiological, enzymological, and restriction endonuclease analyses indicated that three different genes, none of them alaA, were cloned. These genes were avtA+, which encodes the alanine-valine transaminase (transaminase C); tyrB+, which encodes the tyrosine-repressible transaminase (transaminase D); and a previously undescribed gene, called alaB, which encodes an alanine-glutamate transaminase.     

Astrocyte–neuron interactions in neurological disorders

Astrocytes have long been considered as just providing trophic support for neurons in the central nervous system, but recently several studies have highlighted their importance in many functions such as neurotransmission, metabolite and electrolyte homeostasis, cell signaling, inflammation, and synapse modulation. Astrocytes are, in fact, part of a bidirectional crosstalk with neurons. Moreover, increasing evidence is stressing the emerging role of astrocyte dysfunction in the pathophysiology of neurological disorders, including neurodegenerative disease, stroke, epilepsy, migraine, and neuroinflammatory diseases.