The nature of high frequency sister chromatid exchange cells (HFCs)

We employed the three-way differential staining technique (TWD),which allows SCEs to be distinguished on a per generation basisby scoring third metaphases (M3), in order to study the spontaneouslevels of SCEs in normal and high frequency cells (HFCs) thatoccurred in the first (S1), second (S2) and third (S3) S phases.Fifty one of 900 lymphocytes from 37 healthy donors were definedas HFCs by calculating the 95th percentile of the distributionof SCEs in S1 + S2. ‘Normal’ cells presented almostthe same number of SCEs after the first, second and third cellcycles (SCE averages of 2.43, 2.04 and 3.53 respectively). Incontrast, HFCs showed a higher SCE count in SI, which decreasedrapidly through the cycles and reached baseline level at S3(SCE averages of 7.18, 4.29 and 3.45 respectively). This wouldsuggest that the lesions responsible for the higher SCE frequencyin HFCs were effectively removed after two cell cycles and stronglysupport the hypothesis that HFCs are lymphocytes which accumulatehigher levels of DNA lesions through time. ¹To whom correspondence should be addressed     

Vascular pulsations stimulating nitric oxide release during cyclic exercise may benefit health: a molecular approach (review)

It is widely assumed that all exercise, regardless of the degree of difficulty or strenuousness, is good (no pain-no gain). In this speculative review of the literature and our research findings we highlight the fact that strenuous exercise taken to the extreme initiates an immune and vascular proinflammatory situation. However, mild cyclic exercise appears to produce health benefits for an individual. In part, this is due to vascular cyclic pulsations, occurring in mild exercise, stimulating constitutive nitric oxide synthase derived nitric oxide release. This in turn down-regulates vascular endothelial cells and immunocytes, as well as their interaction and inhibits the disassociation of NF-kappaB, preventing the production of proinflammatory cytokines. The nitric oxide so generated may even scavenge excess free radicals, preventing tissue damage. Prolonged strenuous exercise appears to limit these positive phenomena because of the maintained and prolonged high blood pressure that reduces the cyclic pulsations, limiting nitric oxide production. We further note that pathological conditions, i.e., Parkinson's disorder, may benefit from mild exercise, i.e., cyclic nitric oxide production, since the inactivity associated with this disease may lead to compromised nitric oxide production, initiating a progressive deterioration of tissues, including peripheral adrenergic neurons, due to a lack of adequate basal nitric oxide levels required to maintain the vascular microenvironment in a mild state of inhibition. We conclude that mild exercise represents an alternate and economical therapy to preserve health and/or diminish the rate of decline of the normal physiological processes that may even be associated with aging.     

Association of single nucleotide polymorphisms of the interleukin-10 promoter gene and susceptibility to primary biliary cirrhosis: immunogenetic differences in Italian and Japanese patients

Several lines of data suggest that genetic factors play an important role in the onset and/or progression of primary biliary cirrhosis (PBC). Since PBC is an autoimmune disease, it is reasoned to assume that genes encoding cytokines may confer susceptibility to disease. Amongst these factors, interleukin-10 (IL-10) has received significant attention. The promoter region of IL-10 gene has three single nucleotide polymorphisms (SNPs) at positions -1082, -819 and -592. To elucidate the association of the three SNPs of IL-10 promoter region with susceptibility of PBC in two different genetic populations, 159 unrelated patients with PBC (94 Italian and 65 Japanese) and 143 local controls (72 Italian and 71 Japanese) were enrolled. SNPs were determined using allele-specific PCR/RFLP. In Italian PBC patients, the frequency of homozygosity for G/G at position -1082 was significantly higher than that of local controls (p < 0.041, OR = 2.44, 95% C.I.; 1.02-5.86). The frequencies of haplotype GCC in PBC patients, possibly linked to higher IL-10 production, were also significant higher than local controls (p < 0.033). However, in Japanese population, there were no significant differences in the three SNPs and haplotypes between PBC patients and controls. Excessive production of IL-10 may play an important role in some populations in modulating the onset of PBC. Further, immunogenetic studies of PBC should take into account ethnic and geographic variations; this makes such studies in heterogeneous population, like the USA, more difficult.     

Multifunctional microcapsules for pancreatic islet cell entrapment: design,preparation and in vitro characterization

Great advances in cell transplantation have been made, including the recent, remarkable success in pancreatic islet transplantation for the treatment of type 1 diabetes mellitus. Unfortunately, the transplanted cells are very susceptible to oxidative stress that cause severe damage to either allo- or xenogeneic islets upon graft in diabetic patients. Consequently, the transplanted islet functional life span is significantly shortened. The aim of this study was to examine the possible effects of antioxidants on in vitro cultured adult rat islets, and to evaluate the effects of a prolonged-release formulation, in form of cellulose acetate (CA) microspheres, on Vitamin D(3) activity. Isolated rat islets, both free and entrapped in microspheres were treated with Vitamin D(3). The effects of the vitamin were studied at 3, 6 and 9 days of in vitro cell culture. According to insulin secretory patterns, treatment with Vitamin D(3) of both free and CA entrapped microspheres, increased the insulin output as compared to untreated controls. Such positive effects were confirmed under islet static incubation with glucose at day 6. These results suggest that pancreatic islets can be advantageously treated with anti-oxidising vitamins before implantation, and speculatively, with the help of special delivery systems, throughout the islet cell life span, in the post-transplant time period.     

Nitrosative stress in the bronchial mucosa of severe chronic obstructive pulmonary disease

BACKGROUND: Reactive nitrogen species, formed via the reaction of nitric oxide (NO) with superoxide anion and via (myelo)peroxidase-dependent oxidation of NO(2)(-), have potent proinflammatory and oxidizing actions. Reactive nitrogen species formation and nitrosative stress are potentially involved in chronic obstructive pulmonary disease (COPD) pathogenesis. OBJECTIVES: To investigate the expression of markers of nitrosative stress, including nitrotyrosine (NT), inducible NO synthase (iNOS), endothelial NO synthase (eNOS), myeloperoxidase (MPO), and xanthine oxidase (XO) in bronchial biopsies and bronchoalveolar lavage from patients with mild to severe stable COPD compared with control groups (smokers with normal lung function and nonsmokers). METHODS: The expression of NT, iNOS, eNOS, MPO and XO in the bronchial mucosa and bronchoalveolar lavage of patients was measured by using immunohistochemistry, Western blotting, and ELISA and correlated with the inflammatory cell profile. RESULTS: Patients with severe COPD in stable phase had higher numbers of NT(+) and MPO(+) cells in their bronchial submucosa compared with mild/moderate COPD, smokers with normal lung function, and nonsmokers (P < .01). iNOS(+) and eNOS(+) but not XO(+) cells were significantly increased in smokers with COPD or normal lung function compared with nonsmokers (P < .05 and P < .01, respectively). In patients with COPD, the number of MPO(+) cells was significantly correlated with the number of neutrophils (r = +0.61; P < .0025) in the bronchial submucosa. Furthermore, the number of NT(+) and MPO(+) cells was negatively correlated with postbronchodilator FEV(1). CONCLUSION: These data suggest that nitrosative stress, mainly mediated by MPO and neutrophilic inflammation, may contribute to the pathogenesis of severe COPD.     

Association of BDNF with anorexia, bulimia and age of onset of weight loss in six European populations

Several genes with an essential role in the regulation of eating behavior and body weight are considered candidates involved in the etiology of eating disorders (ED), but no relevant susceptibility genes with a major effect on anorexia nervosa (AN) or bulimia nervosa (BN) have been identified. Brain-derived neurotrophic factor (BDNF) has been implicated in the regulation of food intake and body weight in rodents. We previously reported a strong association of the Met66 allele of the Val66Met BDNF variant with restricting AN (ANR) and low minimum body mass index in Spanish patients. Another single nucleotide polymorphism located in the promoter region of the BDNF gene (-270C>T) showed lack of association with any ED phenotype. In order to replicate these findings in a larger sample, we performed a case-control study in 1142 Caucasian patients with ED consecutively recruited in six different centers from five European countries (France, Germany, Italy, Spain and UK) participating in the 'Factors in Healthy Eating' project. We have found that the Met66 variant is strongly associated to all ED subtypes (AN, ANR, binge-eating/purging AN and BN), and that the -270C BDNF variant has an effect on BN and late age at onset of weight loss. These are the first two variants associated with the pathophysiology of ED in different populations and support a role for BDNF in the susceptibility to aberrant eating behaviors.     

Spatial and temporal properties of neurons of the lateral suprasylvian cortex of the cat

Neurons in the posteromedial lateral suprasylvian cortex (PMLS) of cats were recorded extracellularly to investigate their response to stimulation by bars and by sinusoidal gratings. Two general types of cells were identified: those that modulated in synchrony with the passage of drifting bars and gratings and those that responded with an unmodulated increase in discharge. Both types responded to contrast reversed gratings with a modulation of activity: the cells that modulated to drifting gratings modulated to the first harmonic of contrast reversed gratings (at appropriate spatial phase and frequency), whereas those that did not modulate to drifting gratings always modulated to the second harmonic of contrast reversed gratings. No cell had a clear null point. Nearly all cells were selective for spatial frequency. The preferred frequency ranged from 0.1 to 1 cycles per degree (cpd), and selectivity bandwidths (full width at half height) were around two octaves. Preferred spatial frequency was not correlated with receptive field size, but bandwidth and receptive field size were positively correlated. Preferred spatial frequency decreased with eccentricity, at about 0.05 octaves/deg. The response of all cells increased as a function of grating contrast up to a saturation level. The contrast threshold for response to a grating of optimal parameters was approximately 1% for most cells and the saturation contrast approximately 10%. The contrast gain was approximately 25 spikes/s per log unit of contrast. All cells were tuned for temporal frequency, preferring frequencies from approximately 3 to 10 Hz, with a selectivity bandwidth approximately 2 octaves. For some cells, the spatial selectivity did not depend on the temporal frequency and vice versa. Others were spatiotemporally coupled, with the preferred temporal frequency being lower at high than at low spatial frequencies, and the preferred spatial frequency lower at high than at low temporal frequencies. Previous results showing broad velocity tuning to a bar were replicated and found to be predictable from the combined spatial and temporal tuning of PMLS cells and the Fourier spectrum of a bar. Preferred temporal frequency steadily decreased with eccentricity, at 0.025 octaves/deg. The results for PMLS cells are compared with those of other visual areas. Acuity and spatial preference and selectivity bandwidth is comparable to all areas except area 17, where they are a factor of about two higher. Temporal selectivity in PMLS is as fine as observed in other areas. The possibility that PMLS cells may be involved with motion detection and detection of motion in depth is discussed.     

Ultrastructural alterations induced in quail ciliary neurons by postganglionic nerve crush and by Ricinus toxin administration, separately and in combination

The response to postganglionic nerve crush and Ricinus toxin administration by the ciliary neurons of the quail ciliary ganglion was investigated at the ultrastructural level. The toxin was either applied at the crush site on the postganglionic nerves or injected into the anterior eye chamber without any other operative intervention. Crush of postganglionic nerves without toxin administration and saline injection into the anterior eye chamber served as controls for the two toxin administration procedures. Postganglionic nerve crush caused a distinct chromatolytic reaction, accompanied by massive detachment of the preganglionic axon terminals from the ciliary neurons and loss of most of the synapses, both chemical and electrical. This process does not induce cell death and is reversible. Saline injection in the anterior eye chamber caused a moderate retrograde reaction in some of the ciliary neurons, presumably as a consequence of paracentesis. The changes consisted mainly of an increase of perikaryal neurofilaments with, at most, a minor detachment of the preganglionic boutons from a small portion of the cell body at the nuclear pole. Ricinus toxin administration induced neuronal degeneration following a pattern common to both delivery modes. The degenerative process consisted of disruption and detachment of polyribosomes from the rough endoplasmic reticulum, an increase of smooth cisterns and tubules, a dramatic increase of neurofilament bundles, compartmentalization of the cytoplasmic organelles and, finally, karyorrhexis and cell lysis. The final stages of Ricinus toxin degeneration involve a progressive accumulation of extracellular flocculo-filamentous material and cell lysis. After administration of Ricinus toxin to the crush site, ricin-affected neurons showed withdrawal of the preganglionic boutons from a portion of the ciliary neuron, especially at the nuclear pole. After Ricinus toxin injection into the anterior eye chamber, however, the bouton shell surrounding the affected ciliary neurons remained intact in the early stages of degeneration. Detachment of the preganglionic terminals and disruption of the cell junctions, therefore, is the consequence of nerve crush and not of the toxin itself.

This study demonstrates that quail ciliary neurons are a suitable model for experimental neuropathology and neurotoxicology.