Analysis of transactivation capability and conformation of p53 temperature-dependent mutants and their reactivation by amifostine in yeast.

The p53 gene is often mutated during cancer development. Frequency and functional consequences of these mutations vary in different tumor types. We analysed conformation and temperature dependency of 23 partially inactivating temperature-dependent (td) p53 mutants derived from various human tumors in yeast. We found considerable differences in transactivation capabilities and discriminative character of various p53 mutants. No correlations in transactivation rates and conformations of the td p53 proteins were detected. Amifostine-induced p53 reactivation occurred only in 13 of 23 td mutants, and this effect was temperature dependent and responsive element specific. The most of the p53 mutations (10/13) reactivated by amifostine were located in the part of the p53 gene coding for hydrophobic beta-sandwich structure of the DNA-binding domain.

[Heart rate variability during sleep]. / Variabilita srdecní frekvence ve spánku.

BACKGROUND: Changes in the tone of the autonomic nervous system during sleep occur and characterize individual sleep stages and probably also sleep cycles. The spectral analysis of the heart rate variability (SA HRV) is a tool for exact assessment of autonomic nervous activity giving us precise information on the activity of the autonomic nervous system--on its sympathetic and parasympathetic component. METHODS AND RESULTS: All night polysomnographic recording was performed in 11 healthy subjects, during which the SA HRV was carried out. The total spectral power of the heart rate variability and relative values of its individual components were evaluated: the very low frequency component (VLF), the low frequency component (LF), and high frequency component (HF). The absolute value of the RR-interval duration was assessed. The LF spectral band in normalized units was significantly higher during REM sleep than in non-REM sleep. On the other hand, the HF spectral band in normalized units was significantly higher during non-REM sleep compared to REM sleep. The LF/HF ratio, which reflects the sympathovagal balance, had a maximal value during REM sleep and reached its minimum in non-REM sleep. A gradual lengthening of the RR-interval and lowering of the LF/HF ratio during night was observed. CONCLUSIONS: The SA HRV showed to be a sensitive method for detection of activity of the autonomic nervous system during sleep. The sympathovagal balance was shifted to prevailing sympathetic activity in REM sleep. On the contrary, during non-REM sleep this balance was shifted towards prevailing parasympathetic influence. A gradual increase of parasympathetic influence during night was also observed.

[Spectral analysis of the variations in heart rate and cardiac activation on waking up in sleepwalking]. / Análisis espectral de la variabilidad del ritmo cardíaco y activación cardíaca durante el despertar en el sonambulismo.

INTRODUCTION: In sleepwalking, a disorder that is characterised by partial waking, the subject experiences an alteration of the microstructure of sleep that can affect autonomous activity during sleep and the waking state. AIMS: In order to evaluate any possible upset in the regulation of autonomous functioning in sleepwalkers during sleep and the waking state, we conducted a spectral analysis of their heart rate variability (HRV) during both sleep and the waking state. SUBJECTS AND METHODS: Spectral analysis of HRV was conducted in the group of 10 sleepwalkers and 10 normal controls during sleep and during the waking state in both the horizontal and vertical positions. Their pattern of cardiac activation was also analysed during different types of arousal. RESULTS: There were no differences between the group of sleepwalkers and the control group in the parameters used in the spectral analysis of HRV during sleep and in the horizontal position during the waking state. Sleepwalkers showed a greater shift in the sympathovagal balance in favour of sympathetic activity, as a response to standing. During the 5-minute sequences immediately before the start of pathological arousal in sleepwalkers, the total energy in the spectral analysis of HRV was seen to increase. No differences were found between the patterns of cardiac activation displayed by the groups of patients and normal subjects during several different types of arousal. CONCLUSIONS: Autonomous reactivity was seen to be altered as a response to the orthostatic load in sleepwalkers, which could be the consequence of the instability of these patients' sleep. The increase in the total energy in the spectral analysis of HRV immediately before pathological arousal during NREM 4 sleep in sleepwalkers suggests that autonomous activation precedes cortical arousal.

Análisis espectral de la variabilidad del ritmo cardíaco y activación cardíaca durante el despertar en el sonambulismo / Spectral analysis of the variations in heart rate and cardiac activation on waking up in sleepwalking

Introducción. En el sonambulismo, un trastorno caracterizado por un despertar parcial, se refiere una alteración de la microestructura del sueño que puede influir en la actividad autónoma durante el sueño y la vigilia. Objetivo. Para evaluar una posible desregulación autónoma en los pacientes con sonambulismo durante el sueño y la vigilia, realizamos un análisis espectral de la variabilidad del ritmo cardíaco (VRC) durante el sueño y la vigilia. Sujetos y métodos. En el grupo de 10 sonámbulos y de 10 sujetos normales se realizó el análisis espectral de la VRC durante el sueño y durante la vigilia en la posición horizontal y vertical, y un análisis del patrón de la activación cardíaca durante varios tipos de despertar. Resultados. El grupo de sonámbulos y el grupo de control no difirieron en los parámetros del análisis espectral de la VRC durante el sueño y en la posición horizontal durante la vigilia. En los sonámbulos se demostró un mayor cambio del equilibrio simpaticovagal en beneficio de la actividad simpática, en respuesta a la verticalización. Durante las secuencias de 5 minutos inmediatamente anteriores al inicio del despertar patológico en los sonámbulos, se observó el incremento de la energía total del análisis espectral de la VRC. El patrón de la activación cardíaca durante varios tipos de despertar no difirió entre los pacientes y los sujetos normales. Conclusiones. Se evidenció una alteración de la reactividad autónoma en respuesta a la carga ortostática en sonámbulos, lo que puede ser la consecuencia de la inestabilidad del sueño en estos pacientes. El incremento de la energía total del análisis espectral de la VRC, inmediatamente antes del despertar patológico durante el sueño NREM 4 en sonámbulos, sugiere que la activación autónoma precede el despertar cortical (AU)

Introduction. In sleepwalking, a disorder that is characterised by partial waking, the subject experiences an alteration of the microstructure of sleep that can affect autonomous activity during sleep and the waking state. Aims. In order to evaluate any possible upset in the regulation of autonomous functioning in sleepwalkers during sleep and the waking state, we conducted a spectral analysis of their heart rate variability (HRV) during both sleep and the waking state. Subjects and methods. Spectral analysis of HRV was conducted in the group of 10 sleepwalkers and 10 normal controls during sleep and during the waking state in both the horizontal and vertical positions. Their pattern of cardiac activation was also analysed during different types of arousal. Results. There were no differences between the group of sleepwalkers and the control group in the parameters used in the spectral analysis of HRV during sleep and in the horizontal position during the waking state. Sleepwalkers showed a greater shift in the sympathovagal balance in favour of sympathetic activity, as a response to standing. During the 5-minute sequences immediately before the start of pathological arousal in sleepwalkers, the total energy in the spectral analysis of HRV was seen to increase. No differences were found between the patterns of cardiac activation displayed by the groups of patients and normal subjects during several different types of arousal. Conclusions. Autonomous reactivity was seen to be altered as a response to the orthostatic load in sleepwalkers, which could be the consequence of the instability of these patients’ sleep. The increase in the total energy in the spectral analysis of HRV immediately before pathological arousal during NREM 4 sleep in sleepwalkers suggests that autonomous activation precedes cortical arousal (AU)

Hypocretin deficiency in Prader-Willi syndrome.

Four patients with clinically and genetically confirmed Prader-Willi syndrome (PWS) underwent nocturnal polysomnograpy (PSG), multiple sleep latency test (MSLT), human leukocyte antigens (HLA) typing and estimation of cerebrospinal fluid (CSF) hypocretin-1 (Hcrt-1) level to investigate if a role of hypothalamic dysfunction and sleep disturbance might be functionally connected through the hypocretin (orexin) system. In all four patients physical examination confirmed extreme obesity (increasing with age) with dysmorphogenetic features. Excessive daytime sleepiness (EDS) was manifested in only two subjects without any imperative feature. None of the patients under study suffered from cataplexy. Nocturnal PSG revealed fragmented sleep with low efficiency, the hypopnea and apnea indexes increasing from borderline up to very high values in direct proportion to the patients' age. MSLT latency was shortened in two patients with clinically expressed EDS, only one sleep onset rapid eye movements (REM) period (SOREM) was found. HLA typing showed DQB1*0602 positivity in two patients; the further two were negative. Mean value of CSF Hcrt-1 in the patients group was down to 164 +/- 46.8 pg/ml (in comparison with 265.8 +/- 48.8 pg/ml in 10 young healthy subjects, P=0.02). The deficiency of CSF Hcrt-1 level correlated in PWS patients with their EDS severity.

Spectral analysis of the heart rate variability in sleep.

Spectral analysis of heart rate variability (HRV) during overnight polygraphic recording was performed in 11 healthy subjects. The total spectrum power, power of the VLF, LF and HF spectral bands and the mean R-R were evaluated. Compared to Stage 2 and Stage 4 non-REM sleep, the total spectrum power was significantly higher in REM sleep and its value gradually increased in the course of each REM cycle. The value of the VLF component (reflects slow regulatory mechanisms, e.g. the renin-angiotensin system, thermoregulation) was significantly higher in REM sleep than in Stage 2 and Stage 4 of non-REM sleep. The LF spectral component (linked to the sympathetic modulation) was significantly higher in REM sleep than in Stage 2 and Stage 4 non-REM sleep. On the contrary, a power of the HF spectral band (related to parasympathetic activity) was significantly higher in Stage 2 and Stage 4 non-REM than in REM sleep. The LF/HF ratio, which reflects the sympathovagal balance, had its maximal value during REM sleep and a minimal value in synchronous sleep. The LF/HF ratio significantly increased during 5-min segments of Stage 2 non-REM sleep immediately preceding REM sleep compared to 5-min segments of Stage 2 non-REM sleep preceding the slow-wave sleep. This expresses the sympathovagal shift to sympathetic predominance occurring before the onset of REM sleep. A significant lengthening of the R-R interval during subsequent cycles of Stage 2 non-REM sleep was documented, which is probably related to the shift of sympathovagal balance to a prevailing parasympathetic influence in the course of sleep. This finding corresponds to a trend of a gradual decrease of the LF/HF ratio in subsequent cycles of Stage 2 non-REM sleep.

Low cerebrospinal fluid hypocretin (Orexin) and altered energy homeostasis in human narcolepsy.

Hypocretins (orexins) are hypothalamic neuropeptides involved in sleep and energy homeostasis. Hypocretin mutations produce narcolepsy in animal models. In humans, narcolepsy is rarely due to hypocretin mutations, but this system is deficient in the cerebrospinal fluid (CSF) and brain of a small number of patients. A recent study also indicates increased body mass index (BMI) in narcolepsy. The sensitivity of low CSF hypocretin was examined in 38 successive narcolepsy-cataplexy cases [36 human leukocyte antigen (HLA)-DQB1*0602-positive] and 34 matched controls (15 controls and 19 neurological patients). BMI and CSF leptin levels were also measured. Hypocretin-1 was measurable (169 to 376 pg/ml) in all controls. Levels were unaffected by freezing/thawing or prolonged storage and did not display any concentration gradient. Hypocretin-1 was dramatically decreased (<100 pg/ml) in 32 of 38 patients (all HLA-positive). Four patients had normal levels (2 HLA-negative). Two HLA-positive patients had high levels (609 and 637 pg/ml). CSF leptin and adjusted BMI were significantly higher in patients versus controls. We conclude that the hypocretin ligand is deficient in most cases of human narcolepsy, providing possible diagnostic applications. Increased BMI and leptin indicate altered energy homeostasis. Sleep and energy metabolism are likely to be functionally connected through the hypocretin system.

Increased REM density in narcolepsy-cataplexy and the polysymptomatic form of idiopathic hypersomnia.

The present work is focused on REM sleep density in patients with primary hypersomnia in comparison with non-hypersomnia subjects. 28 unmedicated patients with narcolepsy-cataplexy (NC) and 10 unmedicated patients suffering from the polysymptomatic form of idiopathic hypersomnia (IH) and their age- and sex-matched controls were included in the study. The clinical diagnosis was confirmed by MSLT and nocturnal PSG, HLA typing was performed in a respective group of narcoleptic patients. Polygraphical recordings were visually scored with particular regard to the two most characteristic phasic features of REM sleep: the number of rapid eye movements (REMs) and chin muscle twitches (Tws) per minute. These events were evaluated according to recognized criteria; a closer look was taken at both their frequency and their distribution across all the nocturnal REM periods (REMPs). The following main differences between hypersomniac patients (of both groups examined) and healthy controls were found in terms of phasic activity: (I) REM density (expressed in REMs/min and Tws/min in each REM period) was significantly increased in the hypersomniac patients in comparison with the controls. (p>0.05).(II) The intra-night phasic activity distribution was found rising more conspicuously in the hypersomniacs than in the controls.

[Hypocretin (orexin) deficiency in narcolepsy-cataplexy]. / Deficit hypocretinu (orexinu) u narkolepsie-kataplexie.

A mutation in the HCRT locus was proved in 18-yrs old male suffering from narcolepsy-cataplexy. He has demonstrated cataplectic attacks (brief spells of head dropping provoked by laughter) as well as imperative sleep in spells of several minutes up to one hour since the age of six months. He has suffered from severe bulimia since five years; later hypnagogic hallucinations, sleep paralysis and unquiet nocturnal sleep accompanied by periodic limb movements appeared. Symptoms are partially controlled with methylphenidate and either imipramine, clomipramine or fluoxetine. Periodic leg movements poorly responded to L-DOPA and clonazepam treatment. He is HLA-DQB1*0602 negative. Repeated MSLT (over 16 years followed-up period) showed extremely short latency with predominant SOREMPs and also nocturnal PSG recordings revealed fragmented sleep with SOREMPs. This case report demonstrates that hypocretin (orexin) mutations in human can produce the full narcolepsy phenotype and validates data recently reported in dog and mouse models suggesting a role for hypocretin (orexin) in the pathophysiology of narcolepsy and the regulation of REM sleep.

Application of Bathurin and borax in the biological control of Monomorium pharaonis in housing estates.

Of two apartment houses infested with ants of the species Monomorium pharaonis, one was treated with a biological, the other with a chemical substance. In the first case we used Bathurin in combination with borax at a concentration of 1.3% for both substances. In the first week, borax was added to the bait (minced beef), within the next three weeks, the bait was mixed with Bathurin. The procedure was repeated 5 times. After this period, we achieved complete control of the ants on the infested premises.--Our results obtained with the chemical substance (the insecticide Anthrix) were less successful. This indicates that the first method, although time-consuming and requiring a perfect collaboration with all inhabitants, is safer, because it leads to the complete eradication of ants in the infested premises.

Determination of exotoxin in Bacillus thuringiensis cells.

The presence of exotoxin in Bacillus thuringiensis was demonstrated and its quantity in the cells determined. The concentration of exotoxin in the producing microorganism is approximately half the concentration of ATP. Exotoxin is produced at such a rate that the cell excretes 1/5 to 1/4 of its exotoxin content into the medium per minute.