Effectiveness of an intensive weight-loss program for severe OSA in patients undergoing CPAP treatment: a randomized controlled trial.

STUDY OBJECTIVES: To determine whether an intensive weight-loss program (IWLP) is effective for reducing weight, the severity of obstructive sleep apnea (OSA), and metabolic variables in patients with obesity and severe OSA undergoing continuous positive airway pressure treatment. METHODS: Forty-two patients were randomized to the control (CG, n = 20) or the intervention group (IG, n = 22), who followed a 12-month IWLP. The primary outcome was a reduction in the apnea-hypopnea index (AHI) as measured at 3 and 12 months by full polysomnography. Metabolic variables, blood pressure, body fat composition by bioimpedance, carotid intima media thickness, and visceral fat by computed tomography were also assessed. RESULTS: Mean age was 49 (6.7) years, body mass index 35 (2.7) kg/m², and AHI 69 (20) events/h. Weight reduction was higher for the IG than the CG at 3 and 12 months, -10.5 versus -2.3 kg (P < .001), and -8.2 versus -0.1 kg (P < .001), respectively, as was loss of visceral fat at 12 months. AHI decreased more in the IG at 3 months (-23.72 versus -9 events/h) but the difference was not significant at 12 months, though 28% of patients from the IG had an AHI < 30 events/h compared to none in the CG (P = .046). At 12 months, the IG showed a reduction in C-reactive protein (P = .013), glycated hemoglobin (P = .031) and an increase in high density lipoprotein cholesterol (P = .027). CONCLUSIONS: An IWLP in patients with obesity and severe OSA is effective for reducing weight and OSA severity. It also results in an improvement in lipid profiles, glycemic control, and inflammatory markers. CLINICAL TRIAL REGISTRATION: Registry: ClinicalTrials.gov; Title: Effectiveness of an Intensive Weight Loss Program for Obstructive Sleep Apnea Syndrome (OSAS) Treatment; Identifier: NCT02832414; URL: https://clinicaltrials.gov/ct2/show/record/NCT02832414.

Partial cloning of CB1 cDNA and CB1 mRNA changes in stress responses in the Solea solea.

Endogenous cannabinoids, through the CB1 receptor, are involved in the control of several functions including stress responses. The aim of this study was to investigate the presence of cannabinoid receptor CB1 in the sole ovary by partial cloning of brain CB1 cDNA; in a stress paradigm of disturbance by handling, which consisted in catching, netting and hand-sorting, changes of CB1 mRNA were related with those of proopiomelanocortin (POMC) mRNA; the trend and timing of stress responses and adaptation were monitored by measuring plasma cortisol levels. We characterized two forms of CB1-like receptor, termed CB1A and CB1B. The two sole CB1 (both 799bp) share 76% identity in their cDNAs, and the deduced amino acid sequences are 80% identical. The handling stress induced a sustained increase in plasma cortisol levels 1h after the handling began and decreased to low levels 12h after initiation of handling, showing the same trend of ovarian POMC mRNA expression. In addition, while CB1A mRNA did not show any significant changes during handling stress, significantly lower levels of CB1B mRNA were found in stressed fish 1h after the beginning of handling, with CB1 expression increased 24h after stress induction, both in the ovary and brain. It can be concluded that endocannabinoid system is involved in the modulation of adaptive responses to environmental conditions.

[Criteria of medical surveillance of the thyroid in occupational exposure to ionizing radiation: contribution of a caseload]. / Criteri di sorveglianza medica della tiroide nella esposizione lavorativa a radiazioni ionizzanti: contributo casistico.

In this paper preliminary results obtained by the application of a scheduled procedure for the diagnosis and management of thyroid nodular diseases in the medical surveillance of radiation workers are discussed. The study demonstrates the utility of the schedule in early diagnosis of thyroid cancer in medical surveillance of radiation protection.

[Method remarks and observations on the use of the respiratory function tests in occupational medicine]. / Appunti di metodo e riflessioni per l'esecuzione delle prove di funzionalità respiratoria in Medicina del Lavoro.

FVC test represents a preliminary test used to approach the 2nd diagnostic level of non invasive screening in occupational pneumopathy (DLCO, N2 wash out, closure volume). Quality, validity and ability to be replicated of the test in spirometry have been evaluated. Results have been examined by Fisher exact test and by linear regression analysis. It was possible to demonstrate the validity and reproducyibility of this simple, diagnostic and non invasive screening in Occupational Health.

Increase of the ornithine decarboxylase/polyamine system and transglutaminase upregulation in the spinal cord of aged rats.

We have investigated changes in ornithine decarboxylase (ODC) activity and in polyamine levels in the central nervous system of aged rats. We measured a significant increase of ODC catalytic activity in the spinal cord from 30 month-old rats (+105%) as compared to 4 month-old rats. No changes were noticed in the cerebellum, cortex and hippocampus from the same animals. A related putrescine increase was measured in the spinal cord of 30 month-old rats (+168%), together with a smaller increase of spermidine (+33%). A parallel increase (+78%) of the Ca2+-dependent transglutaminase activity was detected in the spinal cord of 30 month-old rats, while no changes were apparent in the cortex and cerebellum. Our observations indicate a possible role of the ODC/polyamine system during the normal process of ageing in rats and point to the spinal cord as the most sensitive area for this kind of modification. A possible role of protein polyamination by transglutaminase is discussed.

Regional distribution of nitric oxide synthase and NADPH-diaphorase activities in the central nervous system of teleosts.

Neuronal nitric oxide synthase (nNOS) and NADPH-diaphorase activities were investigated in discrete areas of the central nervous system of goldfish and brown trout. Both species showed a similar distribution pattern of nNOS activity with regional differences in all examined areas. Telencephalon and hypothalamus showed the highest nNOS values, while in the goldfish cerebellum and its valvula nNOS was not detectable. In both species, NADPH-diaphorase activity showed a lower regional variability, compared to nNOS. The highest activity was measured in the olfactory bulbs where, conversely, low levels of nNOS activity were present. The non close correspondence between NOS and NADPH-diaphorase activities confirms the discrepancies indicated by morphological data. Western blot analysis revealed the presence of a nNOS isoform of about 150 kDa mol. wt. corresponding to that of mammals. The pattern of nNOS expression in the considered brain regions of the goldfish and trout was comparable to the levels of the nNOS activity.

Topography of neurochemical alterations in the CNS of aged rats.

We have performed a general survey study on alterations of neurotransmitter-related and glia-related neurochemical markers in various regions of the CNS of aged (30-month-old) as compared to adult (4-month-old) rats. We have found significant decreases in the level of neurochemical parameters related to the cholinergic and GABAergic systems in several regions of the CNS of aged rats. Only few of the alterations present at the age of 30 months, were present in a group of rat of intermediate age (20 months) included in the present study. Less widespread alterations were found concerning the glutamatergic neurotransmission system. Neurochemical markers related to glial cells (astrocytes and oligodendrocytes) showed a remarkable stability in aged rats as compared to neurotransmitter-related markers. Considering the various CNS areas examined in the present study, the spinal cord of the aged rats was the region showing the most profound alterations of neurochemical parameters, as compared to the various brain areas of the same rats. The present results suggest that moderate and region-specific alterations of neurotransmitter-related parameters occur during normal aging and that glia-related markers are fundamentally stable in the absence of specific pathologies.

Partial neuroprotection of in vivo excitotoxic brain damage by chronic administration of the red wine antioxidant agent, trans-resveratrol in rats.

The antioxidant compound trans-resveratrol, is found in substantial amount in several types of red wine and is considered one of the substances responsible for the lower incidence of coronary heart diseases among regular consumers of such wines, an effect also known as the French paradox. It has also been proposed that resveratrol may have beneficial effects against neurodegenerative diseases. We report here that chronic administration of resveratrol to young-adult rats, significantly protects from the damage caused by systemic injection of the excitotoxin kainic acid, in the olfactory cortex and the hippocampus. The same treatment, however, is not able to give any significant protection in an ex vivo model of simulated ischemia on hippocampal slices in vitro. This first evidence of a partial neuroprotective action of chronic administration of resveratrol in vivo, suggests that other models of neurodegenerative injury, and in particular of excitotoxic brain damage, should be investigated in order to assess the potentiality for resveratrol to be used as a pharmacological tool for neuroprotection.

Developmental effects of in vivo and in vitro inhibition of nitric oxide synthase in neurons.

The diffusible chemical messenger nitric oxide (NO) is involved in neuronal plasticity and it is, therefore, supposed to play a role in brain development. A shortage of NO during the critical period of brain maturation may theoretically have long-lasting consequences on the organization of the adult brain. We have performed in neonatal rats a chronic inhibition of the enzyme responsible for NO production, nitric oxide synthase (NOS), from postnatal day 3 to postnatal day 23, through administration of the competitive antagonist N-nitro-L-arginine methylester (L-NAME). The calcium-dependent catalytic activity resulted almost completely inhibited throughout the period of treatment and it took more than 4 days after its suspension to get a full recovery. The expression of the neuronal isoform of the enzyme (nNOS), revealed by immunoblotting, was unchanged during the treatment and after it. The histochemical reaction for NADPH diaphorase was reduced at the end of the treatment and recovered in concomitance with the recovery of the catalytic NOS activity. No gross structural alterations were detected in brain morphology. The levels of three neurotransmitter-related and one astrocytic marker were unchanged in the cerebellum, hippocampus and cortex of 60-day-old rats which had been neonatally treated. A similar lack of significant effects on neurochemical brain maturation was also noticed in a parallel series of experiments, in which a short pulse of NOS inhibition was performed at a critical prenatal time of brain development, from gestational day 14 to gestational day 19. In vitro, chronic exposure of cerebellar granule cells to L-NAME (500 microM) resulted in slight decrease of surviving neurons after 8 days in culture and in better resistance to the challenge of stressful culture conditions. The present results suggest that the basic plan of brain organization can be achieved despite an almost complete NOS inhibition during the maturation period. In vitro, NOS inhibition may bring to more pronounced consequences on neuronal viability and function.

Ornithine decarboxylase activity during development of cerebellar granule neurons.

Ornithine decarboxylase (ODC), the key enzyme for polyamine biosynthesis, dramatically decreases in activity during normal cerebellar development, in parallel with the progressive differentiation of granule neurons. We have studied whether a similar pattern is displayed by cerebellar granule neurons during survival and differentiation in culture. We report that when granule cells were kept in vitro under trophic conditions (high K+ concentration), ODC activity progressively decreased in parallel with neuronal differentiation. Under nontrophic conditions (cultures kept in low K+ concentration), the enzymatic activity dropped quickly in parallel with an increased apoptotic elimination of cells. Cultures kept in high K+ but chronically exposed to 10 mM lithium showed both an increased rate of apoptotic cell death at 2 and 4 days in vitro and a quicker drop of ODC activity and immunocytochemical staining. A short chronic treatment of rat pups with lithium also resulted in transient decrease of cerebellar ODC activity and increased programmed cell death, as revealed by in situ detection of apoptotic granule neurons. The present data indicate that a sustained ODC activity is associated with the phase of survival and differentiation of granule neurons and that, conversely, conditions that favor their apoptotic elimination are accompanied by a down-regulation of the enzymatic activity.

Alteration of neuronal nitric oxide synthase activity and expression in the cerebellum and the forebrain of microencephalic rats.

Microencephalic rats were obtained through gestational (for the forebrain) or neonatal (for the cerebellum) administration of the DNA-alkylating agent methylazoxymethanol acetate (MAM), which selectively kills dividing cells during neurogenesis. In the microencephalic cerebellum the specific activity of calcium-dependent nitric oxide synthase (NOS) was decreased by 35-40% at 12, 28 and 70 days of age. Other neurochemical markers not related to granule cells (the neuronal population selectively compromised by neonatal MAM treatment), choline acetyltransferase (ChAT) and glutamate decarboxylase (GAD) were not decreased, but actually increased when determined as specific activity. In agreement with the decreased catalytic activity measured in the tube, the expression of neuronal NOS protein was attenuated as judged from immunohistochemistry and Western blotting. In the microencephalic forebrain, the specific calcium-dependent NOS activity measured in homogenates of the whole hemisphere was significantly increased as compared to normal animals. Accordingly, immunohistochemistry for neuronal NOS, as well as NADPH-diaphorase histochemistry revealed an apparent increase in the density of strongly reactive neurons in the underdeveloped cortex and striatum of microencephalic rats. The results reported here demonstrate that permanent alterations of neuronal NOS activity and expression occur when the development of the brain and its neuronal circuits are severely compromised. Furthermore, the permanent downregulation of neuronal NOS in the cerebellum of microencephalic rats may be exploited for the study of the role of NO in mechanisms of synaptic plasticity such as long term depression (LTD).

Simulated ischaemia-reperfusion conditions increase xanthine dehydrogenase and oxidase activities in rat brain slices.

Xanthine dehydrogenase and oxidase activities increased by 87% in rat brain slices after 30 min in vitro ischaemia. A further 41% increase was induced by 30 min simulated reperfusion of ischaemic slices. No conversion from the dehydrogenase to the oxidase activity was observed. The increment of enzyme activity was not due to neosynthesis of the enzyme, since it was not affected by the addition of cycloheximide during the ischaemic incubation. The increased oxygen-dependent form of the enzyme could aggravate the ischaemic brain injury by free radicals production, in particular after reperfusion.

Neuronal nitric oxide synthase is permanently decreased in the cerebellum of rats subjected to chronic neonatal blockade of N-methyl-D-aspartate receptors.

Pharmacological blockade of the (NMDA) receptor at critical stages of brain development may have long-lasting effects on brain chemistry and on animal behavior. We report here experiments in which the competitive NMDA receptor antagonist CGP 39551 was administered to rat pups from postnatal day 7 (P7) to P18. The stage of treatment was selected to primarily target the cerebellum, whose granule cells undergo post-mitotic migration and establishment of synaptic connections during this period. We focused our study on the long-term consequences of CGP 39551 treatment on the neuronal isoform of nitric oxide synthase (nNOS) since nNOS is highly expressed in the cerebellum and it is functionally linked to the NMDA receptor. Treated rats exhibited a long-lasting (up to P70) decrease in the intensity of nNOS immunocytochemical staining in the cerebellar cortex accompanied by a decrement of calcium-dependent NOS catalytic activity. A comparable decrease of enzyme activity was measured in the cerebral cortex, but not in the hippocampus, of adult rats. Other neurochemical markers (glutamatergic, gabaergic, purinergic) and glutamine synthetase were unchanged, while a cholinergic marker was slightly increased in the cerebellum of CGP 39551 treated animals. Taken together these data show that blockade of NMDA receptor during the critical period of formation and stabilization of neuronal circuits preferentially affects long-term nNOS expression and catalytic activity in the cerebellum.

Ischemic and excitotoxic damage to brain slices from normal and microencephalic rats.

Brain slices (olfactory cortex, fronto-parietal cortex and hippocampus) taken from normal or microencephalic rats, obtained by gestational administration of the DNA-alkylating agent methylazoxymethanol acetate (MAM), were subjected to in vitro simulated ischemia or exposed to glutamate (5 mM) or kainate (1 mM). All these neurotoxic insults resulted in decreased viability of the slices, as quantitatively assessed by decrease in the rate of protein synthesis. Hippocampal slices subjected to ischemia and olfactory cortex slices exposed to glutamate or kainate were significantly less sensitive to the neurotoxic insult in microencephalic rats than in controls. The increased efflux of neurotransmitter amino acids (glutamate, aspartate and GABA) in the medium from slices subjected to ischemia or exposed to kainate, showed no significant differences among microencephalic and control rats. The present results suggest that the decreased excitotoxic sensitivity of microencephalic rats is, at least in part, related to intrinsic structural and/or functional alterations of some brain regions which undergo decrease in size as a consequence of the gestational treatment.

Effects of chronic lithium treatment on ornithine decarboxylase induction and excitotoxic neuropathology in the rat.

Young adult rats were chronically treated with lithium (2.5 mmol/kg/day) for 16 days. The day after the last lithium administration, rats were injected s.c. with the excitotoxic convulsant kainic acid (10 mg/kg). As compared to saline controls, lithium-treated rats had no apparent attenuation of convulsions. Furthermore, the induction of brain ornithine decarboxylase and the consequent increase of putrescine levels, an index related to the convulsant effects of kainic acid, were similar in saline- and lithium-treated rats. Other rats were unilaterally injected with ibotenic acid into the nucleus basalis magnocellularis: no differences were measured in cortical choline acetyltransferase (ChAT) decrease among saline- and lithium-treated rats. In both the above experiments, apoptotic cell death was monitored in relevant brain regions of saline- or lithium-treated rats through a specific in situ labeling method for fragmented DNA. Whilst morphological evidence for a reduced damage in the olfactory cortex and hippocampus of kainic acid-injected rats was not obtained, lithium-treated rats showed a lower decrease of specific neurochemical markers: [3H]D-aspartate uptake and glutamate decarboxylase. This result suggests that mechanisms of recovery, absent in saline-treated animals, are elicited by the excitotoxic insult in lithium-treated rats.

Neurochemical changes in cerebellum of goldfish exposed to various temperatures.

Acclimation of goldfish at 35 degrees C increased the cerebellar content of aspartate, glutamate, and taurine and [3H]glutamate uptake. Acclimation at 4 degrees C increased the levels of glutamine, serine, and alanine and glutamine synthetase (GS) activity. Adenosine content increased in cerebellum of fish acclimated to warm temperature. K+-evoked release of endogenous and exogenous glutamate from cerebellar slices increased in fish acclimated at 35 degrees C compared to 4 degrees C. The basal level of cyclic adenosine 3':5'-monophosphate (cAMP) in perfused cerebellar slices in fish acclimated at 35 degrees C was much higher than in fish acclimated at 5 degrees and 22 degrees C. It is concluded that variations of environmental temperature produces large neurochemical changes in goldfish cerebellum.

Absence of excitotoxic neuropathology in microencephalic rats after systemic kainic acid administration.

We have s.c. injected with kainic acid (12 mg/kg) normal adult rats as well as rats rendered microencephalic by selectively timed administration of the DNA alkylating agent methylazoxymethanol acetate (MAM) to the mother during pregnancy. Histological examination of the brains revealed that normal animals underwent neurodegeneration in brain regions sensitive to kainic acid excitotoxicity, such as the olfactory cortex and the hippocampus, while no damage was apparent in the same regions of microencephalic rats. Evaluation of the neurotoxic outcome consequent to the excitotoxic stimulation, was quantitatively performed by measuring the levels of appropriate neurochemical markers 15 days after kainic acid injection. In normal animals, this resulted in significant decrease (up to 60% in the olfactory cortex and 30% in the hippocampus) of markers related to glutamatergic and GABAergic neurons, whereas in MAM-treated rats the same markers were not significantly affected, thus demonstrating a substantial protection against the excitotoxic insult in the microencephalic condition.

Effects of gestational or neonatal treatment with alpha-difluoromethylornithine on ornithine decarboxylase and polyamines in developing rat brain and on adult rat neurochemistry.

Pregnant rats were treated for five consecutive days during gestation with s.c. injections of the ornithine decarboxylase (ODC) inhibitor alpha-difluoromethylornithine (DFMO). Treatment beginning at gestational days 13 or 14 was effective in inhibiting ODC and altering polyamine levels, and resulted in relatively small decreases in body and forebrain weight, but not in significant differences in adult neurochemistry. Neonatal rats were treated with DFMO from postnatal day 0 (PD 0) to PD 24. In addition to some somatic effects (decreased body weight, delayed eyelid opening and delayed fur growth) the postnatal treatment resulted in a permanent decrease in brain weight, which was mainly due to a dramatic decrease in cerebellar size. During treatment, and 3 days after the end of it, the levels of putrescine and spermidine, but not those of spermine, were consistently lower in the cerebellum and forebrain of DFMO-treated rats than in controls. On the other hand, ODC appeared strongly inhibited only during the first phase of the treatment and showed recovery, and also rebound of the activity, during the second part of the treatment. A screening of neurochemical markers related to cholinergic, GABAergic and glutamatergic neurons, as well to astrocytes and oligodendrocytes was performed in several brain regions (cerebellum, olfactory bulbs, cortex, striatum, hippocampus) of some of these rats once they became adults. Significant alterations for all the parameters tested, with the exception of the marker for the glutamatergic transmission, were measured in the undersized cerebellum of the neonatally DFMO-treated rats. A shorter neonatal treatment with DFMO (from PD 1 to 6) resulted, in the adult, in decreased cerebellar size and in neurochemical alterations, both very similar to those occurring after the prolonged treatment. In the other brain regions a few minor differences were noticed. The present results show that: (1) the brain polyamine system is differently regulated in foetuses with respect to newborns; (2) the effects of chronic ODC blockade are different on prenatally or postnatally proliferating neurons, due either to a lower sensitivity of gestationally proliferating neurons or to a subsequent recovery; and (3) chronic postnatal ODC inhibition has a strong effect on proliferating neurons, but little effect on further maturation of postmitotic neurons.

Neurotoxic effects of DSP-4 on the noradrenergic system of the goldfish brain.

The substance N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) is a neurotoxin with selective and long-lasting effects on the noradrenergic (NA) neurons of mammalian brains. The present study examines the effects of this toxin on the noradrenergic system of the goldfish brain. Single doses (50 mg/kg body weight) of DSP-4 reduce the immunoreactivity of the NA synthesizing enzyme dopamine-beta-hydroxylase (DBH), as revealed by immunohistochemistry 7 and 12 days after toxin administration. The depletion involves the DBH-positive fibres and spares the DBH-positive cell bodies. Dopamine-beta-hydroxylase immunoreactivity, 40 days after toxin administration, showed a complete recovery. Ultrastructural investigations confirmed that DSP-4 toxicity affects only nervous fibres and terminals, sparing cell bodies. Administration of DSP-4 also produced a marked decrease of noradrenaline (NA) levels in the goldfish brain, seven days later, while dopamine (DA) and serotonin (5-HT) levels were unaffected by toxin injection. The reduction of NA levels induced by DSP-4 was prevented by the concomitant administration of the NA uptake inhibitor desipramine. Noradrenaline levels measured 40 days after toxin administration show that DSP-4 toxicity was completely reversed. The results suggest a pronounced plasticity of the noradrenergic system in the goldfish brain.

Excitotoxic increase of xanthine dehydrogenase and xanthine oxidase in the rat olfactory cortex.

Excitotoxic lesions induced by systemic injection of kainic acid, resulted in 2-3-fold increase of xanthine dehydrogenase and xanthine oxidase activities in the rat olfactory cortex 48-72 h after drug administration. A significant increase of the xanthine oxidase/dehydrogenase ratio was also observed at 4 and 48 h post-injection. No similar changes were noticed in the hippocampus. The enhancement of enzyme activity seems to be primarily a consequence of the altered cell composition in damaged area. Free radicals produced by the increased oxygen-dependent form of the enzyme could in turn aggravate the excitotoxic brain injury.