Exercise-induced alterations of hepatic mitochondrial function.

In order to examine the effect of a single bout of exercise on hepatic mitochondrial function, starved untrained male rats swam at 34-35 degrees C with a tail weight (5% of body wt.) for 100 min. The rates of ADP-stimulated and uncoupled respiration were higher in the mitochondria isolated from the exercised rats regardless of the substrate utilized. Succinate-linked Ca2+ uptake was 48% greater in the exercised group; however, Ca2+ efflux was markedly depressed. The inhibition of Ca2+ uptake by Mg2+ was higher in the control group, so that the difference in Ca2+ uptake between the two groups was greater in the presence of Mg2+ than in its absence. The response of phosphorylating respiration and Ca2+ fluxes to exogenous phosphate and the pH of the assay medium differed in the exercise group. These observations with the exercised group were not related to non-specific stress. The exercise-induced mitochondrial-functional alterations are reminiscent of those obtained from mitochondria isolated from glucagon- or catecholamine-treated sedentary rats. Thus, adrenergic stimulation as well as other factors may be operating during exercise, leading to an alteration of mitochondrial function in vitro.     

Neurochemical alterations during age-related anorexia.

Unexplained weight loss during the latter stages of aging is commonly preceded by a spontaneous diminution in food intake. Multiple etiologies of age-related anorexia in humans, ranging from social isolation to impaired gastrointestinal function, have been proposed. The observation of this phenomenon in older laboratory animals suggests that physiological changes play a significant causal role. A continually expanding body of information on the neurochemical control of food intake supports a contribution of altered neurochemistry to dysregulated feeding behavior. This review provides an update on the relationship between declining food intake during advanced age and physiological (specifically neurochemical) function. The complexity of the control of food intake as well as the variety of investigative methods used in this field of study render the identification of definitive causes difficult. Evidence presented here is evaluated and possible etiologic factors are suggested.     

Receptor alterations and drug tolerance.

A brief review of studies which have employed chronic administration of anticholinesterases, benzodiazepines, amphetamine, antidepressants, neuroleptics and opiates is presented to illustrate the deficiencies in our current knowledge about the degree to which alterations in neurotransmitter receptors can account for tolerance development to these agents. Although numerous investigators have reported alterations in receptor binding following chronic drug treatment and others have reported tolerance development, these two phenomena have seldom been studied in the same experiment. It is argued that a multidisciplinary approach is required to elucidate the role of receptor alterations in drug tolerance.     

IgG subclass alterations in adult asthma.

Immunoglobulin levels were measured in serum samples of 12 black adult non-smoking asthmatic patients, 11 females and 1 male, and compared with 15 age-, sex-matched normal controls. Their total IgG, IgA and IgM levels were within the normal range. However, on quantitation of subclasses, IgG1 levels were significantly above normal, while IgG2 and IgG3 levels were significantly lower than those of controls. No significant differences were found between the two groups when IgG4 levels were compared. These studies as well as those of others suggest that immunoglobulin administration, particularly of individual subclasses, might prove to be a beneficial addition in the management of this condition.     

Mechanisms of oncogene-mediated alterations in metastatic ability.

Transfected ras oncogenes have been shown to induce metastatic properties in some cells. This altered behavior is likely due to changes in ras-mediated signal transduction pathways, resulting in altered expression of genes important to metastasis. Clarification of the mechanisms by which ras is able to induce metastatic ability in model systems will improve our understanding of tumor progression, even in those cells in which ras activation has not been implicated. Many of the consequences of ras expression also have been detected in cells that have become metastatic in the absence of altered ras, suggesting that there is a set of common changes that can lead to metastasis, with multiple signals capable of eliciting these changes. We have identified several changes in metastatic, ras-transformed NIH 3T3 cells that may contribute to their increased malignancy, including expression of proteolytic enzymes and their inhibitors, and adhesive and calcium-binding proteins. Not all cells, however, respond in this way to expression of oncogenic ras. We have found that murine LTA cells, which are tumorigenic but nonmetastatic, are ras resistant and remain nonmetastatic when expressing high levels of transfected ras, in contrast to NIH 3T3 cells, which are ras sensitive and become both tumorigenic and metastatic in response to comparable levels of ras. LTA cells differ in their patterns of gene expression in response to ras when compared with NIH 3T3 cells, suggesting that the two cell lines process the ras signal differently. Here we review our results with ras-transfected NIH 3T3 and LTA cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Neurotransmitter receptor alterations in Parkinson's disease.

Neurotransmitter receptor binding for GABA, serotonin, cholinergic muscarinic and dopamine receptors and choline acetyltransferase (ChAc) activity were measured in the frontal cortex, caudate nucleus, putamen and globus pallidus from postmortem brains of 10 Parkinsonian patients and 10 controls. No changes in any of these systems were observed in the frontal cortex. In the caudaye nucleus, only the apparent dopamine receptor binding was altered with a significant 30% decrease in the Parkinsonian brain. Both cholinergic muscarinic and serotonin receptor binding were significantly altered in the putamen, the former increasing and the latter decreasing with respect to controls. In addition, ChAc activity was decreased in the putamen. In the globus pallidus, only ChAc activity was significantly changed, decreasing about 60%, with no change in neurotransmitter receptor binding. The results suggest that a progressive loss of dopaminergic receptors in the caudate nucleus may contribute to the decreased response of Parkinsonian patients to L-dopa and dopamine agonist therapy.     

Meiotic alterations in CAG repeat tracts.

We have investigated meiotic changes in CAG repeat tracts embedded in a yeast chromosome. Repeat tracts undergo either conversion events between homologs or expansion and contraction events that appear to be confined to a single chromatid. We did not find evidence for conversion of tract interruptions or excess exchange of flanking markers.     

Visualization of freezing damage. II. Structural alterations during warming

There is a growing amount of indirect evidence which suggests that the loss in viability of rapidly cooled cells is due to recrystallization of intracellular ice. This possibility was tested by an evaluation of the formation of morphological artifacts in rapidly cooled cells to determine whether this process can account for the loss in viability. Samples of the common yeast Saccharomyces cerevisiae were frozen at 1.8 or 1500 °C/min, and the structure of the frozen cells was examined by the use of freeze-fracturing techniques. Other cells cooled at the same rate were warmed to temperatures ranging from ?20 ° to ?50 °C and then rapidly cooled to ?196 °C, a procedure that should cause small ice crystals to coalesce by the process of migratory recrystallization. Cells cooled at 1500 °C/min and then warmed to temperatures above ?40 °C formed large intracellular ice crystals within 30 min, and appreciable recrystallization occurred at temperatures as low as ?45 °C. Cells cooled at 1.8 °C/min and warmed to temperatures as high as ?20 °C underwent little structural alteration. These results demonstrate that intracellular ice can cause morphological artifacts. The correlation between the temperature at which rapid recrystallization begins and the temperature at which the cells are inactivated indicates that recrystallization is responsible for the death of rapidly cooled cells.     

Radiation induced alterations in the endotoxin of S. typhimurium.

The lipopolysaccharide (LPS) of S. typhimurium has been shown to be significantly detoxified after in vivo irradiation at 500 krad. Radiation is thus a useful method for converting endotoxin into toxoid. The structural alterations in the detoxified LPS are shown to be mainly in the lipid A molecule, resulting in the loss of beta-hydroxymyristic acid.     

Chicken liver amidophosphoribosyltransferase. Ligand-induced alterations in molecular properties.

A homogeneous amidophosphoribosyltransferase (EC 2.4.2.14) preparation, which was sensitive to purine nucleotide inhibitors, was obtained from chicken liver. From the result of sodium dodecyl sulfate polyacrylamide gel electrophoresis, the subunit weight was estimated to be approximately 58 000. In Tris-HCl buffer, the predominant form of the enzyme had an S20,w of 6.5, Strokes radius of 40 A, and estimated molecular weight of 110 000. Incubation with 5-phosphoribosyl 1-pyrophosphate or Pi resulted in an increase in the S20,w to 9.1--9.5, Strokes radius 50 A, and estimated molecular weight to 200 000. Incubation of the large form with AMP led to a decrease in the molecular wight of the enzyme. It is concluded that chicken liver amidophosphoribosyltransferase is an allosteric protein whose activity is regulated by a series of conformational changes induced by a number of ligands.     

Temperature-induced alterations in phospholipids of Fusarium oxysporum f. sp. lycopersici.

Ten phospholipids were identified in hyphal membrane preparations of Fusarium oxysporum f. sp. lycopersici when the cells were grown to the late log phase at 15, 25, and 37 degrees C, respectively. The major phospholipids present were phosphatidylcholine (PC) and phosphatidylethanolamine (PE), which together made up about 70% of the total membrane phospholipids. The degree of unsaturation in the acyl group of the phospholipids was inversely related to growth temperature. The polar head group composition was also affected by growth temperature. Cells grown at 15 and 25 degrees C contained the same relative proportions of PC and PE, but when the growth temperature was raised to 37 degrees C, the ratio of PC to PE was doubled. A methylating system capable of converting PE to PC was demonstrated in vitro.     

Freeze-fracture study of adenovirus-induced KB cell surface alterations.

A technique of destabilization of KB cell plasma membrane by treatment with hypotonic citrate buffer was employed. The technique did not alter the viability or ability of the cells to synthesize macromolecules or produce virus, but did permit the visualization, by the freeze-fracture technique, of plasma membrane modifications occurring in response to adenovirus adsorption. The modifications consisted of a rearrangement of the membrane particles (MPs) on both protoplasmic and external leaflets of the plasma membrane. The rearrangement delineated bare areas, 139 nm in mean diameter, devoid of MPs and protruding outwards. The membrane changes were transient and were only observed when KB cells were maintained with adenovirus particles at 0 °C. The changes disappeared rapidly upon warming to 37 °C, reforming the ‘random pattern’ of MPs, normally visible on the cell plasma membrane. The same type of study was carried out with purified adenovirus capsid components (hexon, penton, penton base and fiber), with smaller virus particles (poliovirus) and with larger ‘artificial’ adenovirus particles made of latex beads coated with adenovirus pentons. The dimensions of the bare regions devoid of MPs appeared to be related to the size of the particles used, suggesting the existence of a ‘recognition pattern’ specific for virus particles.     

Frequent alterations of visual pigment genes in adrenoleukodystrophy.

Both adrenoleukodystrophy (ALD) and red/green color blindness have been mapped to the distal long arm of the human X chromosome (Xq28). Color-vision defects are frequently associated with ALD, and study of the red and green visual pigment genes in eight ALD kindreds has shown frequent structural changes including deletions and possible intragenic recombinations. Such changes may reflect chromosomal events underlying both ALD and the associated visual defects and should help define both the structural gene responsible for ALD and physical genetic relationships in the Xq28 region.     

Structural alterations of pathologically or physiologically modified DNA.

We have studied the alterations of DNA conformation in in vitro depurinated or methylated topological isomers of the plasmid pAT 153. Depurination by heat/acid treatment or alkylation by methyl methanesulfonate (pathological modifications) result in DNA unwinding detected as a reduction in the degree of supercoiling of DNA topoisomers as measured by the alteration of electrophoretic mobility on agarose gel. On the contrary, in vitro enzymic methylation at the C-5 position of cytosine (physiological modification) does not measurably alter the tertiary structure of the circular substrates. From the average number of modified sites needed to remove one superhelical twist from each single topoisomer of a population of partially relaxed DNA molecules, we have calculated an unwinding angle smaller than -3.4 degree per methylated purine and of approximately -12.0 degree per apurinic site. These results, together with previously reported values of unwinding by pyrimidine dimers, suggest a possible mechanism of recognition of damaged sites by repair mechanisms that are not single-damage specific.     

Mitochondrial DNA alterations as ageing-associated molecular events.

Mitochondrial DNA (mtDNA) is a naked double-stranded circular extrachromosomal genetic element continuously exposed to the matrix that contains great amounts of reactive oxygen species and free radicals. The age-dependent decline in the capability and capacity of mitochondria to dispose these oxy-radicals will render mtDNA more vulnerable to mutations during the ageing process. During the past 3 years, more than 10 different types of deletions have been identified in the mtDNA of various tissues of old humans. Some of them were found only in a certain tissue but some others appeared in more than one organ or tissue. The 4977-bp deletion is the most prevalent and abundant one among these deletions. Skeletal muscle is the target tissue of most ageing-associated mtDNA deletions and has often been found to carry multiple deletions. The onset age of the various deletions in mtDNA varies greatly with individual and type of the deletion. The 4977-bp deletion has been independently demonstrated to occur in the mtDNA of various tissues of the human in the early third decade of life. However, the 7436-bp deletion was only detected in the heart mtDNA of human subjects in their late thirties. The others appeared only in older humans over 40 years old. No apparent sex difference was found in the onset age of these ageing-associated mtDNA deletions. The various ageing-associated deletions could be classified into two groups. Most of the deletions belong to the first group, in which the 5'- and 3'-end breakpoints of the deletion are flanked by 4-bp or longer direct repeats. The deletion in the second group occurs less frequently and shows no distinct repeat sequences flanking the deletion sites. These two groups of mtDNA deletions may occur by different mechanisms. The first group is most probably caused by internal recombination or slippage mispairing during replication of mtDNA by the D-loop mechanism. The deleted mtDNA and the deleted DNA fragment may be further degraded or escape from the mitochondria and get translocated into the nucleus. The latter route has been substantiated by many observations of inserted mtDNA sequences in the nuclear DNA. Thus, the fragments of migrating mtDNA may change the information content and expression level of certain nuclear genes and thereby promote the ageing process or cause cancer. Similar ageing-associated alterations of mtDNA have also been observed in aged animals and plants. I suggest that mtDNA deletions and other mutations to be discovered are molecular events generally associated with the ageing process.