Cell-autonomous regulation of hematopoietic stem cell cycling activity by ATP.

Extracellular nucleotides regulate many cellular functions through activation of purinergic receptors in the plasma membrane. Here, we show that in hematopoietic stem cell (HSC), ATP is stored in vesicles and released in a calcium-sensitive manner. HSC expresses ATP responsive P2X receptors and in vitro pharmacological P2X antagonism restrained hematopoietic progenitors proliferation, but not myeloid differentiation. In mice suffering from chronic inflammation, HSCs were significantly expanded and their cycling activity was sensitive to treatment with the P2X antagonist periodate-oxidized 2,3-dialdehyde ATP. Our results indicate that ATP acts as an autocrine stimulus in regulating HSCs pool size.

BSCC, Bethesda or other? Terminology in cervical cytology European panel discussion.

The European panel agreed that reproducibility and translatability of terminology in cervical cytology were essential, arguing well for harmonization of reporting systems. The majority at this meeting use a modification of the Bethesda system (BS). Local modifications involved reporting subcategories within high grade and low grade lesions, which would not alter the overall translatability of their systems both with each other and BS. The majority agree that low grade lesions with and without koilocytosis should be managed similarly as should high grade lesions (moderate dysplasia/CIN2 or worse). Those systems linking moderate dysplasia with mild rather than severe dysplasia would need to define moderate dysplasia as such, if their results were to be translatable, which would be preferable to their using a different definition of low grade and high grade lesions. Translation between systems might anyway be facilitated by reporting moderate dysplasia as a subcategory within high grade, which was favoured by most of those present. Therefore, there is no need for exact agreement of terminology if broad principles are agreed. This useful discussion adds weight to the British Society for Clinical Cytology recommendation that the new classification should be adopted by the UK National Health Service Cervical Screening Programme. If the new classification is adopted, the UK would join the European consensus opinion on terminology.

Chronic blockade of glutamate receptors enhances presynaptic release and downregulates the interaction between synaptophysin-synaptobrevin-vesicle-associated membrane protein 2.

During development of neuronal circuits, presynaptic and postsynaptic functions are adjusted in concert, to optimize interneuronal signaling. We have investigated whether activation of glutamate receptors affects presynaptic function during synapse formation, when constitutive synaptic vesicle recycling is downregulated. Using primary cultures of hippocampal neurons as a model system, we have found that chronic exposure to both NMDA and non-NMDA glutamate receptor blockers during synaptogenesis produces an increase in miniature EPSC (mEPSC) frequency, with no significant changes in mEPSC amplitude or in the number of synapses. Enhanced synaptic vesicle recycling, selectively in glutamatergic nerve terminals, was confirmed by the increased uptake of antibodies directed against the lumenal domain of synaptotagmin. No increased uptake was detected in neuronal cultures grown in the chronic presence of TTX, speaking against an indirect effect caused by decreased electrical activity. Enhanced mEPSC frequency correlated with a reduction of synaptophysin-synaptobrevin-vesicle-associated membrane protein 2 (VAMP2) complexes detectable by immunoprecipitation. Intracellular perfusion with a peptide that inhibits the binding of synaptophysin to synaptobrevin-VAMP2 induced a remarkable increase of mEPSC frequency in control but not in glutamate receptor blocker-treated neurons. These findings suggest that activation of glutamate receptors plays a role in the downregulation of the basal rate of synaptic vesicle recycling that accompanies synapse formation. They also suggest that one of the mechanisms through which this downregulation is achieved is an increased interaction of synaptophysin with synaptobrevin-VAMP2.

Evidence of a role for cyclic ADP-ribose in calcium signalling and neurotransmitter release in cultured astrocytes.

Astrocytes possess different, efficient ways to generate complex changes in intracellular calcium concentrations, which allow them to communicate with each other and to interact with adjacent neuronal cells. Here we show that cultured hippocampal astrocytes coexpress the ectoenzyme CD38, directly involved in the metabolism of the calcium mobilizer cyclic ADP-ribose, and the NAD+ transporter connexin 43. We also demonstrate that hippocampal astrocytes can release NAD+ and respond to extracellular NAD+ or cyclic ADP-ribose with intracellular calcium increases, suggesting the existence of an autocrine cyclic ADP-ribose-mediated signalling. Cyclic ADP-ribose-induced calcium changes are in turn responsible for an increased glutamate and GABA release, this effect being completely inhibited by the cyclic ADP-ribose specific antagonist 8-NH2-cADPR. Furthermore, addition of NAD+ to astrocyte-neuron co-cultures results in a delayed intracellular calcium transient in neuronal cells, which is strongly but not completely inhibited by glutamate receptor blockers. These data indicate that an astrocyte-to-neuron calcium signalling can be triggered by the CD38/cADPR system, which, through the activation of intracellular calcium responses in astrocytes, is in turn responsible for the increased release of neuromodulators from glial cells.

Covalently conjugated VEGF--fibrin matrices for endothelialization.

Vascular endothelial growth factor (VEGF) is a key factor in endothelial cell biology and blood vessel formation and a candidate therapeutic for the stimulation of angiogenesis-dependent tissue regeneration. The objective of this study was to confer the angiogenic activity of VEGF(121) upon the biomaterial fibrin, a natural substrate for endothelial cell growth and clinically accepted as 'fibrin glue'. To achieve this, we engineered fibrin-based hydrogels that were covalently modified with VEGF(121). Our laboratory has recently developed novel methodology that allows the covalent incorporation of exogenous bioactive peptides by the transglutaminase activity of factor XIIIa into fibrin during coagulation. Here, this ability of factor XIIIa to crosslink additional proteins within fibrin was employed to covalently incorporate VEGF(121). By recombinant DNA methodology, a mutant VEGF(121) variant, alpha(2)-PI(1--8)-VEGF(121), which contains an additional factor XIIIa substrate sequence NQEQVSPL at the aminoterminus, was expressed in E. coli. In soluble form, the mutant protein fully retained its mitogenic activity for endothelial cells. Using (125)I-labeled alpha(2)-PI(1--8)-VEGF(121), its covalent incorporation and the efficiency of incorporation into fibrin was demonstrated and characterized. The immobilized, fibrin-conjugated VEGF(121) protein remained an active and very efficient mitogen for human endothelial cells grown on two-dimensional VEGF(121)-modified fibrin surfaces, and the incorporation of increasing amounts of alpha(2)-PI(1--8)-VEGF(121) resulted in dose-dependent enhancement of endothelial cell growth. The VEGF-modified fibrin matrices can be formed as injectable gels in a single-step reaction under physiological conditions in vivo. When used as a ingrowth matrix, such VEGF incorporating materials could be useful in a variety of clinical situations that require an angiogenic response into an ischemic region or inplant.

A common exocytotic mechanism mediates axonal and dendritic outgrowth.

Outgrowth of the dendrites and the axon is the basis of the establishment of the neuronal shape, and it requires addition of new membrane to both growing processes. It is not yet clear whether one or two exocytotic pathways are responsible for the respective outgrowth of axons and dendrites. We have previously shown that tetanus neurotoxin-insensitive vesicle-associated membrane protein (TI-VAMP) defines a novel network of tubulovesicular structures present both at the leading edge of elongating dendrites and axons of immature hippocampal neurons developing in primary culture and that TI-VAMP is an essential protein for neurite outgrowth in PC12 cells. Here we show that the expression of the N-terminal domain of TI-VAMP inhibits the outgrowth of both dendrites and axons in neurons in primary culture. This effect is more prominent at the earliest stages of the development of neurons in vitro. Expression of the N-terminal domain deleted form of TI-VAMP has the opposite effect. This constitutively active form of TI-VAMP localizes as the endogenous protein, particularly concentrating at the leading edge of growing axons. Our results suggest that a common exocytotic mechanism that relies on TI-VAMP mediates both axonal and dendritic outgrowth in developing neurons.

Mitochondrial nitric-oxide synthase stimulation causes cytochrome c release from isolated mitochondria. Evidence for intramitochondrial peroxynitrite formation.

Nitric oxide (NO) is synthesized by members of the NO synthase (NOS) family. Recently the existence of a mitochondrial NOS (mtNOS), its Ca(2+) dependence, and its relevance for mitochondrial bioenergetics was reported (Ghafourifar, P., and Richter, C. (1997) FEBS Lett. 418, 291-296; Giulivi, C., Poderoso, J. J., and Boveris, A. (1998) J. Biol. Chem. 273, 11038-11043). Here we report on the possible involvement of mtNOS in apoptosis. We show that uptake of Ca(2+) by mitochondria triggers mtNOS activity and causes the release of cytochrome c from isolated mitochondria in a Bcl-2-sensitive manner. mtNOS-induced cytochrome c release was paralleled by increased lipid peroxidation. The release of cytochrome c as well as increase in lipid peroxidation were prevented by NOS inhibitors, a superoxide dismutase mimic, and a peroxynitrite scavenger. We show that mtNOS-induced cytochrome c release is not mediated via the mitochondrial permeability transition pore because the release was aggravated by cyclosporin A and abolished by blockade of mitochondrial calcium uptake by ruthenium red. We conclude that, upon Ca(2+)-induced mtNOS activation, peroxynitrite is formed within mitochondria, which causes the release of cytochrome c from isolated mitochondria, and we propose a mechanism by which elevated Ca(2+) levels induce apoptosis.

Rapid detection of mutated E-cadherin in peritoneal lavage specimens from patients with diffuse-type gastric carcinoma.

Tumor cells in abdominal lavage specimens from patients with gastric carcinoma strongly predict subsequent peritoneal metastasis and poor prognosis. Reverse transcription (RT)-polymerase chain reaction (PCR) detection of wild-type E-cadherin has been claimed to be superior to conventional cytology for the detection of patients who subsequently develop peritoneal metastases. The present study tested this hypothesis and determined whether or not the detection of mutated, tumor-specific E-cadherin messenger RNA in abdominal lavage specimens serve as a useful diagnostic tool. Preoperative lavage specimens from 52 patients with diffuse-type gastric carcinoma and from 5 patients with benign disease were analyzed by conventional cytology and by RT-PCR for amplification of E-cadherin. Tumor cells were detected by cytology in 8 (15.3%) of the 52 patients with gastric cancer. The E-cadherin was detected in all 57 samples by RT-PCR. Two of these had abnormal E-cadherin amplification products confirmed to be mutations by direct sequencing, which were identical in the primary tumors. These findings suggest that the detection of wild-type E-cadherin is not sufficiently tumor specific. Also, for diffuse gastric carcinomas with confirmed E-cadherin mutations, detection of mutant E-cadherin by RT-PCR is a potentially valuable method for tumor cell detection in lavage specimens.

Ceramide induces cytochrome c release from isolated mitochondria. Importance of mitochondrial redox state.

In the present study we show that N-acetylsphingosine (C2-ceramide), N-hexanoylsphingosine (C6-ceramide), and, to a much lesser extent, C2-dihydroceramide induce cytochrome c (cyto c) release from isolated rat liver mitochondria. Ceramide-induced cyto c release is prevented by preincubation of mitochondria with a low concentration (40 nM) of Bcl-2. The release takes place when cyto c is oxidized but not when it is reduced. Upon cyto c loss, mitochondrial oxygen consumption, mitochondrial transmembrane potential (Delta Psi), and Ca2+ retention are diminished. Incubation with Bcl-2 prevents, and addition of cyto c reverses the alteration of these mitochondrial functions. In ATP-energized mitochondria, ceramides do not alter Delta Psi, neither when cyto c is oxidized nor when it is reduced, ruling out a nonspecific disturbance by ceramides of mitochondrial membrane integrity. Furthermore, ceramides decrease the reducibility of cyto c. We conclude that the apoptogenic properties of ceramides are in part mediated via their interaction with mitochondrial cyto c followed by its release and that the redox state of cyto c influences its detachment by ceramide from the inner mitochondrial membrane.

Skin microcirculation in patients with type I diabetes with and without neuropathy after neurovascular stimulation.

1. Neurovascular inflammation is impaired in patients suffering from diabetic neuropathy. The aim of our study was to evaluate the distribution of nutritive and total skin blood flow in diabetic patients with and without neuropathy after neurovascular stimulation with acetylcholine. 2. Twenty patients with Type I diabetes, 10 with and 10 without neuropathy, and 10 age-matched non-diabetic control subjects, underwent microvascular investigations before and after neurovascular stimulation by intracutaneous application of acetylcholine. The capillary blood cell velocity in the nailfold of the hallux was measured by videophotometric capillaroscopy, and the total skin microcirculation in the same area by laser Doppler flowmetry. 3. The increase in total skin blood flow was significantly impaired in the group of neuropathic diabetic patients compared with the non-neuropathic diabetic patients (17.5 +/- 8.3 versus 51.0 +/- 16.2; P < 0.05) and the non-diabetic subjects (17.5 +/- 8.3 versus 67.8 +/- 19.7; P < 0.01). The increase in capillary blood flow was not significantly impaired in Type I diabetes patients with neuropathy. 4. The ratio between capillary blood flow and total skin perfusion decreased significantly in the control group (from 0.82 +/- 0.15 to 0.47 +/- 0.11; P < 0.05) and in the Type I diabetes patients without neuropathy (from 0.79 +/- 0.12 to 0.43 +/- 0.12; P < 0.05), whereas the decrease in the neuropathic group was statistically insignificant (from 1.05 +/- 0.19 to 0.72 +/- 0.16). 5. Diminished total skin perfusion in the foot after intracutaneous stimulation with acetylcholine in Type I diabetes patients is associated with diabetic neuropathy, indicating a disturbance in the neurovascular reflex arc. This impaired neurovascular response is caused by a diminished total and subpapillary blood flow and not by a diminished nutritive capillary flow. There is no evidence of a diminished nutritive capillary blood flow during neurogenic inflammation in Type I diabetes patients suffering from diabetic neuropathy.

In vitro effects of glucose polymer-containing peritoneal dialysis fluids on phagocytic activity.

Commercially available peritoneal dialysis fluids (PDFs) are known to impair peritoneal cellular defense mechanisms. We have investigated the influence of glucose polymer-containing PDFs on phagocytic function in vitro. Polymorphonuclear neutrophils (PMNLs) and monocytes (MNs) from 10 continuous ambulatory peritoneal dialysis patients and 10 healthy donors were incubated in PDFs containing either 7.5% icodextrin (glucose polymer) or 1.5% glucose at original pH and pH 7.4. Chemiluminescence response and H202 production were measured following stimulation with preopsonized Staphylococcus epidermidis or phorbol myristate acetate. Phagocytosis of radiolabeled bacteria and killing capacity of the cells were determined. A comparison of the impact of glucose polymer versus glucose-containing solutions at their original pH on the oxidative metabolism of the cells showed a highly significant difference (P < 0.0001) in favor of glucose polymers for H202 production of PMNLs (7.78 +/- 4.5 nmol cytochrome C reduction/10(6) cells/min v 1.11 +/- 0.67 nmol cytochrome C reduction/10(6) cells/min) and MNs (7.66 +/- 3.6 nmol cytochrome C reduction/10(6) cells/min v 1.29 +/- 0.86 nmol cytochrome C reduction/10(6)cells/min). Correspondingly, PMNLs and MNs incubated in glucose polymers showed a significantly higher chemiluminescence response irrespective of the stimulant used (P < 0.0001). Applying the killing assay on PMNLs also revealed a significantly higher percentage of inactivated bacteria (45.5% +/- 11.0% v 29.2% +/- 15.5%; P < 0.05). After adjustment of pH to 7.4, a significant difference could only be found for H202 production of PMNLs in favor of glucose polymers (16.73 +/- 6.98 nmol cytochrome C reduction/10(6) cells/min v 11.65 +/- 5.37 nmol cytochrome C reduction/10(6) cells/min; P < 0.05). In addition, we compared the glucose-polymer solution to an otherwise equally composed equiosmolar solution that contained 0.274% glucose instead of glucose polymers. No significant differences were detected with any of the tests applied. Our data suggest that glucose polymer solutions are comparatively less suppressive to phagocytic function than currently used glucose-containing PDFs. This effect may be attributed to the low osmolarity of these solutions.

The response of perennial ryegrass/white clover swards to elevated atmospheric CO2 concentrations 1. Effects on competition and species composition and interaction with N supply.

The effects of long-term carbon dioxide enrichment on competition for nutrients and light in a ryegrass/clover association were determined for simulated swards of perennial ryegrass (Lolium perenne L. cv. Parcour) and white clover (Trifolium repens L. cv. Karina), which were grown as monocultures and in three mixtures (25/75, 50/50, 75/25), according to the replacement design, at two levels of nitrogen (N) supply (no additional N and 200 kg N ha-1 ) and at season-long ambient (380 ppm) and elevated (670 ppm) CO2 concentrations, in open-top chambers. Stands were cut four times, at about monthly intervals, to a height of 5 cm. Plant material was separated into different species, fresh and dry weights were determined and the content of macroelements (N, P, K, S, Mg) in both species was measured. In addition, plant height of both species at harvest dates and during several regrowth periods was monitored. Results indicate that both species made demand on different resources and profited from growth in a mixed sward. Co2 related yield increase amounted to 16-4-2 % for white clover whereas the effect of high CO2 on ryegrass yield ranged between -33% and +9% depending on N supply, mixture and year. As a result the contribution of white clover to total yield in mixed swards was significantly enhanced by CO2 enrichment at many harvests in both N supply treatments. Without additional N supply, shoot competition for light was intensified by CO2 enrichment to the disadvantage of ryegrass, since clover petioles grew longer and ryegrass was shorter at elevated CO2 With N fertilization, no marked effect of CO2 enrichment on interspecific competition could be observed. Since clover and total yield were increased by CO2 enrichment, nutrient requirements were also increased and potassium deficiency and increased intraspecific competition of clover for K was observed in the mixtures under elevated CO2 which had the highest nutrient withdrawal. Although white clover profited much more from CO2 enrichment in both N fertilization treatments, the suppression of ryegrass in mixed swards could only be observed under low N conditions. Generally, the effect of N fertilization on competitive interference between both species was much greater than the effect of CO2 enrichment and it is suggested that the effect of elevated CO2 on the balance of species and the outcome of competition in a grass/clover sward is mainly dependent on the N status.

Low-dose intradermal versus intramuscular hepatitis B vaccination in patients with end-stage renal failure. A preliminary study.

Patients with end-stage renal disease (ESRD) are at high risk of hepatitis B infection. Only 50-60% of the patients respond adequately to the routinely performed intramuscular (i.m.) hepatitis B vaccination. We examined whether low dose intradermal (i.d.) application of the vaccine is equivalent to regular i.m. administration. Thirty-two patients with ESRD of different etiologies were investigated at the onset of dialysis treatment [11 patients on continuous ambulatory peritoneal dialysis (CAPD) and 21 patients on hemodialysis (HD)]. Patients were vaccinated at month 0, 1, 3 and 6 with either 40 micrograms HBs Ag (2 ml Engerix B, 14 patients) i.m. or with 10 micrograms HBsAg (0.5 ml Engerix B, 18 patients) i.d. The i.m. vaccination was applied in the deltoid muscle, while for i.d. vaccination the vaccine was injected into the skin of the deltoid region. Six weeks after the last vaccination anti-HBs titers were measured. 61% (11 patients) of the patients vaccinated i.d. and 64% (9 patients) of the patients vaccinated i.m. developed protective titers. Neither the height of the titers nor the proportion of patients responding to the vaccination differed significantly between the two vaccination schedules. No difference regarding the height of titers achieved or the rate of seroconversion could be found when CAPD and HD patients were analyzed separately. Only minor side effects have been observed. According to these preliminary data i.d. hepatitis B vaccination in patients with ESRD may be equivalent to i.m. administration of the vaccine. Given equivalency i.d. vaccination may be a cost-saving alternative to i.m. vaccination (only a quarter of the dose of i.m. administered vaccine is needed) with a good practicability (vaccination can be performed during HD) and a low rate of side effects.

In vitro effects of bicarbonate- versus lactate-buffered continuous ambulatory peritoneal dialysis fluids on peritoneal macrophage function.

At present, lactate is the most commonly used buffer in peritoneal dialysis fluids (PDFs). The high lactate concentration in combination with low original pH was demonstrated to suppress phagocytic function. We evaluated the in vitro effects of a newly formulated bicarbonate-buffered PDF containing glycylglycine (BiGG15 and BiGG40; Pierre Fabre Medicament, Castres, France) on peritoneal macrophage (PMO) function, and compared them with those of equiosmolar lactate-buffered PDF (1.5% and 4.25% glucose; pH 5.4 and pH 7.4) and control buffer. Peritoneal macrophages were isolated from the effluents of 10 continuous ambulatory peritoneal dialysis patients and tested for luminol- and lucigenin-enhanced chemiluminescence, superoxide (O2-) generation measured by cytochrome c reduction, killing capacity, and phagocytosis after incubation in the PDF used. Exposure of PMO to lactate-buffered PDF with an original pH of 5.4 resulted in a significant suppression of all PMO functions measured, compared with bicarbonate- and lactate-buffered PDFs with a pH of 7.4. At physiological pH (7.4), chemiluminescence generation of PMO exposed to BiGG15/40 was significantly higher compared with the corresponding equiosmolar lactate-buffered PDF (1,992 +/- 858 x 10(3) cpm/10(4) cells v 856 +/- 398 x 10(3) cpm/10(4) cells; P < 0.004). O2- generation, killing capacity, and phagocytosis were not significantly different after PMO exposure to bicarbonate compared with exposure to lactate-buffered PDF with a neutral pH. Irrespective of the buffer used, high-osmolality PDFs suppressed PMO function significantly more than low-osmolar PDFs. In conclusion, bicarbonate-buffered PDFs are less detrimental to PMO function than lactate-containing PDFs; these preliminary in vitro results need to be confirmed in vivo.

In vitro effects of low-calcium peritoneal dialysis solutions on peritoneal macrophage functions.

The use of low-calcium peritoneal dialysis solutions (PDS) for continuous ambulatory peritoneal dialysis is becoming widely accepted to reduce the risk of serum hypercalcemia in patients taking calcium salts as phosphate binders. We compared the in vitro effects of low-calcium PDS (1,000 mumol calcium/L), calcium-free buffer, and buffers with increasing calcium concentrations (500 to 5,000 mumol calcium/L) on peritoneal macrophage (PMO) functions. Peritoneal macrophages isolated from 10 continuous ambulatory peritoneal dialysis patients were incubated in the different solutions and tested for phagocytic and killing capacity, superoxide generation (cytochrome-C reduction and lucigenin-enhanced chemiluminescence), and the rate of myeloperoxidase-dependent oxidative metabolism (luminol-enhanced chemiluminescence). All functions of the PMO incubated in calcium-free buffer were significantly suppressed compared with the PMO incubated in calcium buffers. No dose-dependent increase of a single PMO function could be found after incubating the PMO in calcium buffer with increasing concentrations. Incubation of PMO in otherwise identical PDS containing 1,000, 1,450, or 1,750 mumol calcium/L did not result in significantly different PMO functions. Acidic PDS (pH 5.3 to 5.5) suppressed all measured PMO functions as compared with their neutralized counterparts (pH 7.4), irrespective of the calcium concentration. Results of our in vitro study show that low-calcium PDS does not suppress PMO functions any more than standard-calcium PDS (1,750 mumol calcium/L) does.

In vitro effects of amino-acid-based versus glucose-based continuous ambulatory peritoneal dialysis fluids on peritoneal macrophage function.

We studied the in vitro effects of peritoneal dialysis fluids (PDF) containing 1 and 1.5% amino acids (AA) as compared to approximately equiosmolar glucose (GLU)-based PDF (1.5 and 4.25%) and control buffer, respectively, on peritoneal macrophage (PMO) function. The media were tested at original pH (5.3-5.5) and after pH adjustment to 7.4. PMO were isolated from the effluents of 10 on continuous ambulatory peritoneal dialysis (CAPD) patients and tested for luminol- and lucigenin-enhanced chemiluminescence (CL), superoxide generation measured by cytochrome c reduction, killing capacity and phagocytosis after incubation (30 min, 37 degrees C) in the PDF used. All AA-based PDF exhibited a statistically significant depressive effect on integral CL response, O2- production and bacterial killing of PMO at pH 7.4 in comparison with pH-adjusted GLU-based PDF of similar osmolality and buffer. Exposure of PMO to acidic AA-based media did not result in a significantly different suppression of the oxidative metabolism and the killing capacity as compared to fresh GLU-based fluids. Phagocytosis of PMO did not show significant differences after incubation in the solutions studied. Thus, the AA-based PDF employed compromise the oxidative metabolism and the killing capacity of PMO at pH 7.4 in vitro significantly more than GLU-based fluids. Since pH-identical and almost equiosmolar PDF were compared, the specific composition of the AA-based fluids, especially the high content of lactate and several essential AA, could be responsible for this detrimental impact.

Incidence and significance of pneumoperitoneum in continuous ambulatory peritoneal dialysis.

In a retrospective study we reviewed upright chest x-ray films of 101 continuous ambulatory peritoneal dialysis (CAPD) patients to determine the incidence and significance of free subdiaphragmal air. A pneumoperitoneum (PP) was diagnosed if a minimal shadow of free air was detected under the diaphragm. The amount of free air was determined by measuring the height and width of the subdiaphragmal air shadow. Of all CAPD patients, 33.6% (34 of 101) had at least one occurrence of PP. Thirteen of these 34 patients (38.2%) were diagnosed within 30 days after catheter implantation, 10 patients (29.5%) acquired a PP during an episode of peritonitis, and in 11 patients (32.4%) no additional risk factor could be determined. Patients radiographed within 30 days after catheter implantation showed a statistically significant higher incidence of PP compared with the same patients radiographed later (22% v 10%; P < 0.05). The incidence of PP in CAPD patients suffering from peritonitis (33%) was significantly higher than in patients without peritonitis (10%; P < 0.001). The amount of free air did not differ statistically significantly between the investigated groups. Only two patients with PP and peritonitis had surgically confirmed visceral perforation. Therefore, the main reason for PP seemed to be handling faults during CAPD bag exchange. There was no correlation between the organisms causing peritonitis and PP or the CAPD connector system and PP. In conclusion, a PP occurs in approximately one third of all CAPD patients and a visceral perforation cannot be diagnosed by the occurrence and amount of free subdiaphragmal air.

Influences of partial REM sleep deprivation and awakenings on nocturnal cortisol release.

Decreasing endogenous plasma cortisol levels during rapid eye movement (REM) sleep have been recently reported, suggesting a diminished or absent secretory activity of the adrenals during this period. On the other hand, episodes of light sleep (Stage 1) and intermittent wakefulness have been found to be associated with increasing plasma cortisol levels. The present experiments in 10 adult men were designed to examine whether or not REM sleep inhibits adrenocortical activity and if short periods of wakefulness increase nocturnal cortisol release. Somnopolygraphic recordings were obtained from each subject under three experimental sleep conditions: a baseline night, an REM deprivation night in which the subject's sleep was disturbed contingent upon the occurrence of REM, and a control night in which sleep was disturbed both during REM deprivation and non-REM (NREM) epochs, i.e., mostly during Stage 2 sleep. This last condition was introduced to distinguish the effects of REM deprivation from those of arousals that may per se act as stressful stimuli for cortisol release. Contrary to expectation, we found that both REM sleep deprivation and arousals in NREM epochs reduced rather than enhanced mean plasma cortisol levels as compared with baseline conditions. These findings do not support the hypothesis of an inhibitory effect of REM sleep on cortisol secretion, though present data do not refute this hypothesis. The awakenings, or the light sleep subsequent to sleep disturbance, appear to have no stimulatory effect on adrenocortical secretion. Awakenings during sleep at night may even reflect the activity of mechanisms inhibiting sleep-related increases in plasma cortisol concentration.