Expression of apoptosis and proliferating cell nuclear antigen (PCNA) in the cardiac conduction system of crib death (SIDS).

Aim of this study is to determine the expression of apoptosis and Proliferating Cell Nuclear Antigen (PCNA) in the cardiac conduction system in crib death and explained death (ED) cases. Postnatal morphogenesis of the conducting tissue is an important part of its normal development. In the atrio-ventricular node (AVN) and His bundle (HB) it consists of degeneration, cell death and replacing in an orderly programmed way. However, its nature and its relation to crib death is not yet fully explained. Apoptosis and PCNA were investigated in 8 heart conduction systems of infants dying of crib death and in 3 conduction systems of infants dying of ED as controls. The cardiac conduction system was removed in two blocks: the first included the sino-atrial node (SAN) and the crista terminalis, the second contained the atrio-ventricular node (AVN), His bundle (HB), bifurcation, and bundle branches. In the conduction systems as well as in the common myocardium the PCNA Labeling Index (PCNA-LI) was found to be negative in all cases. The apoptotic indices (AI) in SIDS and in ED were found to have no statistically significant differences (p>0.05). The SAN, in both groups, showed an AI similar to the one detected in common myocardium. In almost all cases, TUNEL labeling was detected in peripheral region of the AVN, close to the atrial myocardium. The AI was higher in the AVN, HB and the initial tract of bundle branches than in the common myocardium (p<0.05; Student's t test).

The role of different biomarkers (DNA, PCNA, apoptosis and karyotype) in prognostic evaluation of superficial transitional cell bladder carcinoma.

BACKGROUND: In order to clarify the variable behaviour of transitional cell bladder carcinomas (TCBC) with same clinico-pathologic pattern, we investigated the prognostic significance of various biomarkers (PCNA, DNA, apoptosis, karyotype). MATERIALS AND METHODS: We studied 177 superficial TCBC (stage T1) undergoing transurethral resection (TUR). Analysis of biological indicators was performed on serial paraffin sections: DNA by static cytometry, karyotype by fluorescence in situ hybridisation (FISH), PCNA and apoptosis by immunohistochemistry. RESULTS: The most salient results are represented by prevalence of diploidy (56%), a mean PCNA labeling index (PCNA-LI) of 8.2%, nonrandom trisomies and tetrasomies of chromosome 7, and a high presence of apoptosis (in 72% of cases). These data were not related to histological grading. Multivariate analysis showed that only PCNA-LI is a an independent prognostic marker for patient survival (p = 0.01). Besides, we observed a worse prognosis in the presence of both very high PCNA indices and low or absent apoptosis. CONCLUSIONS: The results of this study suggest that tumor prognostic potential in TCBC should be evaluated on the basis of the association between PCNA cell kinetics and cell apoptosis information.

Chromosomal alterations in atherosclerotic plaques.

Alterations of chromosomes 7 and 11 have been involved in the progression of atherosclerosis. Twenty-three carotid endarterectomy specimens were studied for the presence of alterations in chromosomes 7 and 11, and fibroblastic growth factor-3 (FGF-3) gene amplification. Besides classic histological stainings, immunophenotyping of cellular and vascular components and fluorescence in situ hybridization (FISH) were performed. At the caps, unstable plaques (n=18) showed inflammatory infiltration of macrophages, smooth muscle cells, and T-lymphocytes. Specifically in these regions, the FISH showed varying percentages of trisomy (15/18) and tetrasomy (8/15) of chromosome 7. In four cases polisomy 7 was noted in some nuclei. Monosomy of chromosome 11 and gene amplification of FGF-3 gene was observed. The FISH of the five stable plaques and normal arterial walls showed no chromosome alterations; furthermore, chromosome 3, which is not involved in atherosclerotic progression, presented a normal ploidy of smooth muscle cells in stable and unstable plaques and normal arterial walls. In conclusion, chromosome 7 and 11 alterations and FGF-3 gene amplification are components of unstable plaques, and might contribute to the evolution of stable plaques into complicated plaques.

Inflammatory cells, apoptosis and Chlamydia pneumoniae infection in atherosclerosis.

Chlamydia pneumoniae (CP), chromosomal alterations and apoptosis were suggested as contributing factors in the pathogenesis of atherosclerosis. Early (EP) and unstable plaques (UP) were studied in order to assess infiltrate composition, the apoptotic index, chromosome 7 stability and to investigate the concurrent presence of CP in EP and UP. Paraffin embedded sections of three iliac arteries and four aortas from young donors (EP), and four coronaries and nine carotid arteries (UP) were used. Aside from histological techniques, immunophenotypification for macrophages, T and B cells, smooth muscle and endothelial cells; FISH and DNA nick end labeling were performed. The amplifications with PCR for CP infection were negative in all specimens. In the EP, a focal myointimal thickening with foam cells and scarce smooth muscle cells was observed. Macrophages were most frequent in the intima (10.8%) while T and B cells were found in 2.3 and 1.5%. In the UP a thin cap covering a lipid-rich core with widespread vascularization and with severe luminal obstruction was observed. Macrophages were increased (21%), and T (1.5%) and B cells (3.5%) in the caps and inner areas of the lipid cores. At these sites, the FISH showed trisomy and tetrasomy of chromosome 7 and apoptosis was very frequent (10-30%). Macrophages in intimal lesions is one of the most prominent, consistent and permanent features in EP, and an elevated apoptotic index and chromosome 7 instability might contribute to evolution from stable to complicated plaques, while CP seems to play no role. However, further studies are needed with more cases to confirm this last observation.

Beta-endorphin levels in peripheral blood mononuclear cells and long-term naltrexone treatment in autistic children.

We assessed the clinical and biological effects of high-dose, long-term Naltrexone (NTX) treatment in 11 children (3-11 years), who had been diagnosed as autistic. The drug was given following an open design, for 12 weeks. Beta-Endorphin (beta-END) was assayed in peripheral blood mononuclear cells after 1 and 3 months of treatment, and 6 months after the completion of the course. Baseline beta-END levels were higher than in healthy age-matched controls. In seven patients treatment reduced beta-END, whose levels rose in four children. Autistic symptoms were considerably attenuated in all cases, with functional improvements involving several areas. There was a close correlation between the reduction in beta-END levels and the decrease of social withdrawal, and an evident--though weak--correlation between increases in beta-END and decreases in stereotypy and abnormal speech. Both effects persisted after treatment stopped.

p53 over-expression and its correlation with PCNA index in nasal polyps.

Our knowledge about the etiopathogenesis of nasal polyps (Nps) is still limited. In this study, in order to define the biological features of these neoformations, we investigated with immunohistochemistry the p53 over-expression and the proliferating cell nuclear antigen (PCNA) in 32 cases of Nps and in normal mucosa of 11 control cases. The evaluation of PCNA showed a wide range of indices (0.5-18.2%) with a mean value (6.8%) significantly higher than in normal mucosa (2.9%). Over-expression of the p53 oncoprotein, observed in 50% of Nps, was statistically related to a high PCNA-index (> 6.8%). Our results suggest that Nps can behave, in a high percentage of cases, like tumours.

Prognostic significance of different biological markers (DNA index, PCNA index, apoptosis, p53, karyotype) in 126 adenocarcinoma gastric biopsies.

BACKGROUND: The different clinical evolution of gastric adenocarcinomas with the same clinico-pathologic characteristics prompted the authors to investigate the prognostic significance of different biological markers. METHODS: One hundred twenty-six preoperative cancer gastric biopsies, selected according the stage evaluated after following gastrectomy, were examined for DNA content by means the static cytometry, proliferating cell nuclear antigen (PCNA), p53 mutation, apoptosis by immunohistochemistry and karyotype using fluorescence in situ hybridization (FISH) techniques. RESULTS: The gastric cancers, ail belonging at stage III, were adenocarcinomas histologically typed as: 26 well differentiated (G1), 45 moderately differentiated (G2), 43 poorly differentiated (G3) and 12 of undifferentiated type with signet-ring cells. The tumours showed a prevalence of diploidy (68%), a mean PCNA-LI of 4.8%, trisomy of chromosome 7 in 40% of the cases, low presence of apoptosis (30%) and p53 mutation (17%). Only apoptosis was significantly correlated to histological diagnosis (p = 0.009). A multivariate analysis showed that the DNA content and PCNA-LI were the only independent prognostic markers for survival (p = 0.005 and p = 0.0002 respectively). CONCLUSIONS: The evaluation of these two biological variables, especially of the PCNA index, on gastric cancer biopsies may be useful in predicting the aggressiveness of each tumor and in identifying patients in need of additional perioperative therapies.

Detection of trisomy 7 with fluorescence in situ hybridization and its correlation with DNA content and proliferating cell nuclear antigen-positivity in prostate cancer.

The authors applied fluorescence in situ hybridization (FISH), using specific DNA probes for chromosome 7, to routine paraffin-embedded tissue sections obtained from 35 radical-prostatectomy specimens. Proliferative activity was also evaluated using static cytometry to assess DNA content and immunohistochemistry for proliferating cell nuclear antigen (PCNA) positivity. These results were correlated with each other and with the morphologic parameters. The presence of three or more of chromosome 7 was observed in 71% of the cases, as was a high S phase, with a triploid prevalent DNA content and a PCNA index above mean value in 66% of the cases. No correlation was detected between these findings and histologic grade; conversely, there was a significant correlation with stage (chi-square = 5.33; p = 0.021). From these results, the authors maintain that the presence of an extra chromosome 7-correlated in most cases with an increase in cell kinetics and an advanced stage-may be an additional prognostic marker of aggressive behavior.

Cytogenetic aspects of cell proliferation in atherosclerotic plaques.

Fluorescence in situ hybridization (FISH) using chromosome specific alpha-satellite DNA probes for chromosome 7, was performed on paraffin-embedded sections of 21 cases of human "unstable" atherosclerotic plaques, in order to determine whether trisomy 7, found by some Authors with conventional cytogenetics, is a biological characteristic of the atherosclerotic plaque. In 15 cases (71%) the FISH method showed smooth muscle cells with three or more spots. In particular the trisomy 7 was the most significant and constant result. The presence of an extra chromosome 7 is strictly correlated with an overexpression of the gene for chain A of the platelet-derived growth factor (PDGF), which was mapped at pter-7q22 region. Therefore trisomy 7 seems to correspond to an increase in PDGF synthesis and consequently to an increase in proliferative activity of smooth muscle cells.

Beta-endorphin and cholecystokinin 8 concentrations in peripheral blood mononuclear cells of autistic children.

beta-Endorphin (beta-EP) and cholecystokinin 8 (CCK-8) concentrations in peripheral blood mononuclear cells were measured in 12 drug-free autistic (AU) children, in 10 drug-free children with pervasive developmental disorders (PDD) and in 11 healthy controls. The aim of the study was to see whether or not there was an alteration of beta-EP and CCK-8 concentrations in this peripheral compartment, in which it has been suggested that secretion and regulation of the two peptides mimic those of neurons in the central nervous system. Mean beta-EP values were significantly higher in AU than in PDD and control children, while there were no differences in CCK-8 values of the three groups.

Growth hormone response to growth hormone releasing hormone and to clonidine stimulation in peripubertal patients with major depressive disorder.

The responses of growth hormone (GH) to administration of growth hormone-releasing hormone (GHRH-1 micrograms/kg b.w.) and of clonidine (clon-2.5 micrograms/kg b.w.) and basal levels of somatomedin C (SmC) were measured in nine peripubertal patients with Major Depressive Disorder (MDD) and in 9 age- and gender-matched controls. Basal GH and SmC levels, and GH response to GHRH did not differ in patients and controls, whereas responses to clonidine were significantly higher in some and lower in other patients than in controls.

Cortisol responses of depressed children and adolescents to clonidine administration.

The responses of cortisol to acute administration of saline and of clonidine (2.5 micrograms/kg B.W.) were examined in 10 children and adolescents with major depressive disorder and in 10 age- and sex-matched controls. Clonidine administration did not inhibit cortisol secretion in controls or in patients, contrary to what has been observed in depressed adults. In controls, but not in patients, clonidine administration induced a transitory increase of the steroid.

A developmental study of the behaviour of mice fostered to normal and alcoholic virgin females.

Two conditions of temporary litter fostering, to water-drinking subjects and 28%-ethanol-drinking subjects, were compared to stay-with-mother condition and no-adult-present condition. Significant differences were found in the following variables: body weight during growth, self-feeding latency at weaning, timidity at one-month age, and, in adulthood, latency of females to contact an infant stimulus object and aggressive activities between males. It seems that the normal and the alcoholic virgin females had similar, long-lasting effect on emotionality of the fostered mice.

Developmental course of brain-stem auditory evoked potentials in the first days of full term infants.

Modifications of brain-stem acoustic evoked potentials (BAEPs) in the first 5 days of life of normal full-term infants are reported. BAEPs were recorded using rarefaction clicks at 70, 60, 40, 20 dB HL. Multiple linear regression analysis was performed to evaluate chronological and gestational age differences for all positive and negative peaks, interpeak latencies, amplitudes and amplitude ratios. The percentage of newborn infants with auditory threshold of 20 dB HL increased with the number of days. At all intensities the latency of PIII and PV decreased significantly between the 2nd and 3rd day, while the latency of PI decreased significantly between the 3rd and the 4th day; PIII and PV latencies decreased also between the 4th and the 5th day. The longer the period of gestation the longer was the PI latency recorded between the 2nd and 5th day after birth. The anatomical and physiological changes developing shortly after birth are probably responsible for the above findings. These changes most probably appear firstly in the cochlear and trapezoid nuclei and later on in the organ of Corti.

Effects of carbamazepine and valproate on immunological assessment in young epileptic patients.

Serum immunoglobulin levels and peripheral blood lymphocyte subsets were determined in 25 epileptic children treated with anticonvulsant drugs [carbamazepine (CBZ) or sodium valproate (VPA)], 17 untreated patients and 18 healthy subjects. The treated and untreated patients did not differ significantly from the controls with respect to the mean IgA, IgG, IgM values or lymphocyte subsets. The patients on carbamazepine had lower serum concentrations of IgG (though not significantly lower) than the untreated patients and children on valproate. OKT8 subset, in absolute terms and as a percentage of total lymphocytes, was significantly higher in generalized than in partial epilepsy, but this was probably due to the antiepileptic treatment. We found a sex difference in serum immunoglobulins (Ig M, IgG) in the epileptic group. Our data do not seem to argue for major changes in immune status related to clinical type of epilepsy or to VPA or CBZ treatment.

The acute and chronic administrations of piracetam affect the movement-related brain macropotentials.

Neurophysiological and neurochemical studies have demonstrated that piracetam improves learning and memory both in animals and humans. In recent years it has been shown that when a subject is engaged in a motor perceptual task, some psychomotor functions are correlated with a consistent pattern of brain electrical activity. Given the relationship between the movement-related brain macropotentials (MRBMs) and the cognitive processes associated with them, we considered the MRBMs particularly suitable for the neurophysiological assessment of the efficacy of piracetam in man. The aim of this study was to test the acute and chronic effect of piracetam administration on the MRBMs in normal children during the performance of a motor perceptual task. The design was a triple-blind study, during which the subjects took either placebo or piracetam in random sequence, with a washout period of 3 weeks. The dose was 170 mg/kg for the acute treatment and 140 mg/kg/day for chronic treatment. No side-effects were reported by the children during or after acute or chronic treatment with piracetam. There was no statistically significant difference between placebo or piracetam treatment with regard to 'performance', which was already optimal at baseline, and to electromyographic activity. On the contrary, the MRBMs were significantly modified by treatment. In particular the Bereitschaftspotential was present as a positive shift during acute treatment with piracetam and increased after chronic treatment. Skilled performance positivity (SPP) amplitudes were significantly increased and SPP latency reduced by chronic treatment with piracetam. piracetam appears to act on the catecholaminergic and cholinergic systems via an increase of the inhibitory hyperpolarizing processes.

Neuroendocrine investigation in children and adolescents with dysthymic disorders: the DST, TRH and clonidine tests.

A neuroendocrine study was conducted in eight children and adolescents with dysthymic disorders (three females and five males) and in eight age- and sex-matched psychologically normal controls. The dexamethasone suppression test (DST), TSH and GH responses to TRH stimulation and GH response to clonidine stimulation were studied in parallel in each patient. Depressive symptomatology was monitored with the Poznanski Rating Scale. The DST, TRH and clonidine tests revealed normal responses in each patient. TRH induced abnormal GH rises in five of the eight patients. There were no correlations between neuroendocrine parameters and degree of depression, age, sex or weight of the patients, age of onset, duration and family history of the disease.

Sex and ear differences of brain-stem acoustic evoked potentials in a sample of normal full-term newborns. Normative study.

The present paper reports a normative study of the BAEPs recorded from 80 normal full-term newborns by using a rarefaction click at 70, 60, 40, 20 dB HL. Positive, negative peaks and the relative amplitudes were measured. Means, standard deviations and 95% and 99% confidence limits were calculated for each component, for positive and negative IPLs and amplitude ratios. A normality test distribution showed that each component had normal distribution except for wave PIII at 70 dB and at 60 dB HL because of its index of kurtosis and for IPLs PII-PIII and NII-NIII at 70 and 60 dB HL. MLRA was performed and significant statistical differences were found for sex, ears and intensities. Females had the latencies of waves PIII, PIV, PV, NII and NIV shorter than males. The BAEPs obtained from the left ear had shorter latencies for positive and negative peaks. Latencies, amplitude and morphology of the BAEPs vary with variations of the intensity. The IPLs PII-PV, PIII-PV and PIV-PV were not affected by changes of intensity.