High-flow nasal oxygen therapy in intensive care and anaesthesia.

Oxygen therapy is first-line treatment for hypoxaemic acute respiratory failure (ARF). High-flow nasal oxygen therapy (HFNO) represents an alternative to conventional oxygen therapy. HFNO provides humidified, titrated oxygen therapy matching or even exceeding the patients' inspiratory demand. The application of HFNO is becoming widespread in Intensive Care Units (ICUs), favoured by increasing evidence based on numerous studies supporting its efficacy. The mechanisms of action and physiological effects of HFNO are not yet fully understood. Pharyngeal dead space washout, decrease in airway resistance, generation of a positive end-expiratory pressure, and enhanced delivery of oxygen are all alleged to be potential mechanisms. The emerging evidence suggests that HFNO is effective in improving oxygenation in most patients with hypoxaemic ARF of different aetiologies. Notwithstanding the potential benefit of HFNO in the management of hypoxaemia, further large cohort studies are necessary to clarify the indications, contraindications and factors associated with HFNO failure. HFNO may also be valuable in reducing the need for tracheal intubation in the management of post-extubation ARF. In addition, HFNO has been proposed to limit oxygen desaturation by prolonging apnoeic oxygenation during intubation both in ICUs and operating theatres.

Telemonitoring in chronic ventilatory failure: a new model of survellaince, a pilot study.

BACKGROUND AND AIM: The efficiency of tele-monitoring or tele-assistance in patients with severe chronic ventilatory failure in home mechanical ventilation (HMV) is still being investigated. Our aim was to test the feasibility of a model which consisted in: 1) once a week nocturnal telemonitoring, supervised by a doctor in charge in a Respiratory Intensive Care Unit, who also provided a telephone-counselling (24/7) on demand; 2) a scheduled visit every two months. METHODS: A 2-year observational study was carried out on 16 patients ventilated for at least 1 year and for > or = 8 hours/day. Once a week patients underwent a nocturnal monitoring during HMV. The compliance was evaluated by regular transmission of data and regular follow-up, the level of satisfaction by a telephone-questionnaire. RESULTS: The adherence to the protocol study was good in 9/16 (56%) and poor in 7/16 (44%) patients. For each patient, the mean number of connections was 46.12 +/- 36.39 (70.7% of that expected), in those with good compliance it increased to 63.8 +/- 32.7 (114% of that expected). The median hours of connection was 343 (138-1019) and 89 (0-521) for patients with good and poor compliance respectively, p = 0.038. The mean scheduled visits for patient with good compliance was 6.9 +/- 4.14 (100% of that expected). Emergency visits were avoided in 62.5% of cases. The satisfaction score was higher in compliant versus non compliant patients (p = 0.019). CONCLUSION: This pilot study showed that the telemonitoring system employed was feasible and effective in more compliant patients who claimed a high rate of satisfaction.

Omalizumab decreases exacerbation frequency, oral intake of corticosteroids and peripheral blood eosinophils in atopic patients with uncontrolled asthma.

Omalizumab is a humanized monoclonal anti-IgE antibody approved in 2005 by the European Medicine Agency (EMA) for the treatment of severe persistent allergic asthma, which remains inadequately controlled despite optimal therapy with high doses of inhaled corticosteroids and long-acting ß2-adrenergic agonists. Within this context, the present observational study refers to 16 patients currently treated with omalizumab at the Respiratory Unit of "Magna Græcia" University Hospital located in Catanzaro, Italy, whose anti- IgE therapy was started in the period included between March 2007 and February 2010, thus lasting at least 10 months. After 40 weeks of add-on treatment with omalizumab, very relevant decreases were detected, in comparison with pre-treatment mean (± standard deviation) values, in monthly exacerbation numbers (from 1.1 ± 0.6 to 0.2 ± 0.4; p < 0.01) and oral corticosteroid consumption (from 22.6 ± 5.0 to 1.2 ± 2.9 mg/day of prednisone; p < 0.01). These changes were associated with stable improvements in lung function, expressed as increases of both FEV1 (from 53.6 ± 14.6% to 77.0 ± 14.9% of predicted values; p < 0.01) and FEV1/FVC ratio (from 56.3 ± 9.5% to 65.8 ± 9.2%; p < 0.01). Moreover, in 5 patients who persistently had increased numbers of eosinophils (mean ± SD: 15.9 ± 8.0% of total WBC count; absolute number: 1,588.0 ± 956.9/µl) despite a long-lasting therapy with inhaled and systemic corticosteroids, the peripheral counts of these cells decreased down to near normal levels (mean ± SD: 6.3 ± 2.3% of total WBC count; absolute number: 462.0 ± 262.3/µl) after 16 weeks of treatment with omalizumab. Therefore, this descriptive evaluation confirms the efficacy of add-on omalizumab therapy in selected patients with exacerbation-prone, chronic allergic uncontrolled asthma, requiring a continuous intake of oral corticosteroids.

Effects of budesonide on P38 MAPK activation, apoptosis and IL-8 secretion, induced by TNF-alpha and Haemophilus influenzae in human bronchial epithelial cells.

Non-typeable Haemophilus influenzae (NTHi) is one of the most frequently involved pathogens in bacterial exacerbations of chronic obstructive pulmonary disease (COPD). In the airways, the main tissue target of NTHi is bronchial epithelium, where this pathogen can further amplify the inflammatory and structural changes induced by proinflammatory cytokines such as tumour necrosis factor-alpha (TNF-alpha). Therefore, the aim of this study is to investigate, in primary cultures of human bronchial epithelial cells, the effects of NTHi on signal transduction pathways, apoptotic events and chemokine production activated by TNF-alpha. Moreover, we also evaluated the effects exerted on such cellular and molecular phenomena by a corticosteroid drug. p38 mitogen-activated protein kinase (MAPK) phosphorylation was analyzed by Western blotting, using an anti-phospho-p38 MAPK monoclonal antibody. Apoptosis was assayed by active caspase-3 expression. Interleukin-8 (IL-8/CXCL8) was detected in cell-free culture supernatants by ELISA. TNF-alpha induced a significant increase in p38 MAPK phosphorylation. NTHi was able to potentiate the stimulatory actions of TNF-alpha on caspase-3 expression and, to a lesser extent, on IL-8 secretion. These effects were significantly (P less than 0.01) inhibited by a pharmacological pre-treatment with budesonide. These results suggest that TNF-alpha is able to stimulate, via activation of p38 MAPK signalling pathway, IL-8 release and airway epithelial cell apoptosis; the latter effect can be markedly potentiated by NTHi. Furthermore, budesonide can be very effective in preventing, through inhibition of p38 MAPK phosphorylation, both structural and proinflammatory changes elicited in bronchial epithelium by TNF-alpha and NTHi.

Long-term oxygen therapy in COPD: evidences and open questions of current indications.

Long term oxygen therapy (LTOT) has been shown to improve the survival rate in Chronic Obstructive Pulmonary Disease (COPD) patients with severe resting hypoxemia by NOTT and MRC studies, published more than 25 years ago. The improved survival was found in patients who received oxygen for more than 15 hours/day. The effectiveness of LTOT has been documented only in stable COPD patients with severe chronic hypoxemia at rest (PaO2 < 55 mmHg (7.3 kPa) or PaO2 ranging from 56 to 59 mmHg (7.4-7.8 kPa) in presence of signs of Cor Pulmonale, hematocrit > 55%. In fact no evidence supports the use of LTOT in COPD patients with moderate hypoxemia (55 < PaO2 < 65 mmHg), and in those with decreased oxygen saturation (SO 2 <90%) during exercise or sleep. Furthermore, it is generally accepted without evidence that LTOT in clinical practice is warranted in other forms of chronic respiratory failure not due to COPD when a-terial blood gas criteria match those established for COPD patients. The prescription of oxygen in these circumstances, as for unstable patients, increases the number of patients receiving supplemental oxygen and the related costs. Comorbidities are likely to affect both prognosis and health outcomes in COPD patients, but at the moment we do not know if LTOT in these patients with complex chronic diseases and mild-moderate hypoxemia could be of any use. For these reasons a critical revision of the actual guide lines indications for LTOT in order to optimise effectiveness and costs, and future research in the areas that have not previously been addressed by NOTT and MRC studies, are mandatory.

Immunolocalization of alpha-synuclein in the rat spinal cord by two novel monoclonal antibodies.

This study provides the first immunohistochemical evidence of the presence and distribution patterns in the rat spinal cord of alpha-synuclein (alpha-Syn), a soluble acidic protein, widely expressed in the CNS and closely associated to the pathogenesis of neurodegenerative conditions such as Parkinson's and Alzheimer's diseases. We used two novel homemade monoclonal antibodies (2E3 and 3D5) recognizing two different epitopes of alpha-Syn. Both antibodies localized alpha-Syn within the nerve terminals, whereas 3D5 alone also localized it within the neuronal nuclei. alpha-Syn-immunoreactive nervous elements were widely recognized throughout rat spinal cord and in almost all the gray matter laminae. However, they appeared particularly concentrated within laminae I, II, VII and X and more scattered in the others. Double immunofluorescent labeling showed that alpha-Syn colocalized with synaptophysin in the presynaptic nerve terminals, with neuropeptide Y (NPY) in lamina I, II, IX and X, and had close relationships with tyrosine hydroxylase (TH) immunoreactive neurons in laminae VII and X. Interestingly, the alpha-Syn-immunoreactive nerve elements, in lamina X, contained little of calbindin-28KD and calretinin-31KD. Our findings could help in understanding the genesis of some early clinical symptoms of Parkinson's disease (PD), such as pain and dysautonomic disorders, and indicate the spinal cord as their probable starting point, according to the ascending theory of PD, proposed by Braak.

Endocrine cells in atresic chick embryo intestine: histochemical and immunohistochemical study.

Intestinal motility disorders are an important problem in the postoperative management of patients with intestinal atresia. Intestinal motility could be initiated by luminal factors that activate intrinsic and extrinsic primary afferent nerves involved in the peristaltic reflex. Endocrine cells act as a key point, because they transfer information regarding the intestinal contents and intraluminal pressure to nerve fibers lying in close proximity to the basolateral surface of the epithelium. In chick embryo, experimental intestinal atresia is associated with disorders in the development of the enteric nervous system, related to the severity of intestinal dilation. Our aim was to investigate the distribution pattern of endocrine cells in the developing endocrine system of chick embryo small intestine with experimentally-induced atresia on day 12 and on day 16. Changes in enteroendocrine population were examined in gut specimens (excised proximal and distal to the atresia) from experimental embryos 19 days old and in control sham-operated chick embryos at the same age. Sections from proximal and distal bowel and control bowel were stained with Grimelius silver stain, a valuable histochemical method for detecting the argyrophil and argentophilic cells, and with an immunohistochemical procedure for detecting serotonin and neurotensin immunoreactive cells. In chick embryo proximal bowel, intestinal dilation differed in the various embryos. We found significantly higher enteroendocrine cell counts in proximal bowel than in distal and control bowel. The differences depended on the precociousness of surgery and the severity of dilation. Considering the major contribution of enteroendocrine cells to the peristaltic reflex, our data may help to explain the pathogenesis of motility disorders related to intestinal atresia.

Increased activation of p38 MAPK in COPD.

Inflammation, oxidative stress and apoptosis, which are involved in chronic obstructive pulmonary disease (COPD) pathogenesis, may activate the p38 subgroup of mitogen-activated protein kinases (MAPKs). Therefore, the aim of the present study was to evaluate the expression of the phosphorylated, active form of p38 MAPK (phospho-p38) in the lungs of COPD patients. Surgical specimens were obtained from 18 smokers with COPD at different stages of disease severity, plus nine smoking and eight nonsmoking subjects with normal lung function. Phospho-p38+ cells were quantified by immunohistochemistry in both alveolar spaces and alveolar walls. Moreover, a Western blot analysis of phospho-p38 and total p38alpha isoform expressed by alveolar macrophages was performed. Phospho-p38+ alveolar macrophages and phospho-p38+ cells in alveolar walls were increased in patients with severe and mild/moderate COPD, compared with smoking and nonsmoking controls. Moreover, they were inversely correlated to values of forced expiratory volume in one second (FEV(1)) and FEV(1)/forced vital capacity. Western blot analysis showed that phosphorylated p38, but not the total p38alpha isoform, was specifically increased in alveolar macrophages from COPD patients. Activation of the p38 mitogen-activated protein kinase pathway appears to be involved in the pathogenesis of chronic obstructive pulmonary disease. The present findings suggest that this protein may be a suitable pharmacological target for therapeutic intervention.

Early development of chick embryo respiratory nervous system: an immunohistochemical study.

The extrinsic and intrinsic respiratory nervous systems receive specific contributions from the vagal and sympathetic components. Using specific markers for vagal and sympathetic structures, we studied the distribution patterns of immunoreactivity to galanin (GAL), pituitary adenylate cyclase-activating polypeptide-27 (PACAP) and the tachykinin substance P in extrinsic and intrinsic nerve of chick embryo respiratory system, during development from the very early age to hatching. All peptides studied appeared in the intrinsic and extrinsic nervous systems early. We found substance P in both the vagal and sympathetic systems, PACAP in vagal components alone and GAL mainly in the sympathetic system. The intrinsic nervous system showed high immunoreactivity for all peptides studied. These data accord with the well known early trophic functions that peptides have on the development of nervous networks and modulatory activity on the intrinsic nervous system. The GAL again proves to be the main peptide in chick embryo sympathetic respiratory system.

Postnatal development of [D-Ala2]deltorphin-I-like immunoreactive structures in the rat brain.

[D-Ala2]deltorphin-I, a highly selective ligand for delta opioid receptors, is a heptapeptide originally purified from frog skin. Previous immunohistochemical studies indicate that [D-Ala2]deltorphin-I-like molecule(s) may be present in adult rat brain, including specific neuronal cells and fibers partially overlapping with the mesocortical and nigrostriatal dopaminergic systems. Here, we examined the developmental aspect of such immunoreactive brain structures in early postnatal rats. In newborn to 21-day-old rats, positive staining in the brain occurred mainly in subpopulations of neurons and occasionally in tanycytes. On postnatal day 0, neuronal cell bodies containing [D-Ala2]deltorphin-I-like immunoreactivity were found in various brain regions, including the olfactory tubercle, ventral pallidum, hippocampus, ventral tegmental area, pars compacta of the substantia nigra, supramammillary nucleus, and dorsal raphe nucleus. Immunoreactive nerve fibers were observed in the main and accessory olfactory bulbs, olfactory tubercle, prelimbic area, anterior cingulate cortex, neostriatum, accumbens, lateral septal nucleus, lateral habenular nucleus, and superior colliculus. As pups grew, positive staining of cell bodies decreased gradually in both density and intensity, and those in the olfactory tubercle and ventral pallidum were no longer visible on postnatal day 14. On postnatal day 21, positive cells were found only in the ventral midbrain, including the pars compacta of the substantia nigra, ventral tegmental area, A8 region, and supramammillary nucleus. Positive fibers also decreased in density with age except in the accessory olfactory bulb, olfactory tubercle, prelimbic area, and anterior cingulate cortex.

Transient expression of [D-Ala2] deltorphin I-like immunoreactivity in prenatal rat small intestine.

We studied the distribution of immunoreactive elements for [D-Ala2] deltorphin I (DADTI), a delta-opioid receptor ligand, in fetal and postnatal rat small intestine. DADTI-like immunoreactive cells were detected transiently on embryonic Days 20 and 21. Electron microscopic examination revealed that positive staining occurred in mucous epithelial cells, either mature goblet cells or undifferentiated cells containing only a few mucous granules. Positive immunoreaction products in mature goblet cells were confined in their apical cytoplasm to the luminal parts of mucous granule aggregates. The result suggests that a DADTI-like molecule(s) is synthesized in rat intestinal goblet cells and is secreted in a diacrine fashion into the intestinal lumen at a late fetal period. The molecule(s) thus secreted may be important for the intestine of rats just before birth, because DADTI-like immunopositive goblet cells are no longer seen at any postnatal period.

Ontogeny of (D-Ala(2))-deltorphin I-like immunoreactive neurones in foetal rat brain.

Foetal rat brain from embryonic day (ED) 12-22 was immunohistochemically studied to describe the time of first appearance and further distribution patterns of (D-Ala(2))-deltorphin-I-immunoreactive (DADTI-IR) nerve elements. The primary antiserum used in this study was a polyclonal antibody against DADTI previously used in adult and postnatal rat brain mapping. DADTI-IR nerve elements first appeared in the neuroepithelium of ventral mesencephalon on ED 13. From there, positive cell bodies migrated towards the mantle layer until they invaded the whole ventral mesencephalic tegmentum. They then reached their definitive position, corresponding to a subpopulation of the A8, A9 and A10 dopaminergic neurones that had been constantly observed also in the adult age. From ED 15-17, DADTI-positive nerve fibres appeared in the medial forebrain bundle, the neostriatum anlage, the accumbens nucleus, the olfactory tubercle, the fasciculus retroflexus, and the prefrontal cortex. All these locations have also been found in adult rats. From ED 14 onwards, transient DADTI-IR somata and nerve fibres were observed in retinal neuroepithelium, optic pathways as far as the superior colliculus, CA3 hippocampal field, reticular formation in the medulla oblongata. All these locations gradually disappeared either before birth (medulla oblongata) or within the first 3 weeks after birth. These results suggest that the DADT-like molecule recognised by our antibody has during the embryonic development a regulatory function in neuronal growth and differentiation.

Autoradiographic study on [3H]-[D-Ala2]-deltorphin-I binding sites in the rat brain.

Previous biochemical and pharmacological studies have shown that [D-Ala2]-deltorphin-I (DADTI) has a high affinity and selectivity for delta-opioid receptors. In this study, designed to provide morphological details, the distribution of DADTI binding sites was examined by autoradiography on coronal, sagittal and horizontal frozen sections of adult rat brain. The sections were incubated with tritiated DADTI solution and exposed for 12 weeks to a 3H-sensitive film. DADTI labelling clearly demonstrated selective and high affinity binding sites of delta-opioid type in several brain regions, including olfactory system, neostriatum, nucleus accumbens, and cortical layers I-II and V-VI.

Production of antiserum to [D-Ala2]deltorphin I and its immunohistochemical application to the mouse brain.

Antiserum to haptenic [D-Ala2]deltorphin I (DADTI: Tyr-D-Ala-Phe-Asp-Val-Val-Gly.NH2), a highly selective ligand for delta opioid receptors, was produced in rabbits. By immunospot assay, the antiserum recognized 62.5 pmol DADTI but failed to react even with 4 nmol carrier protein of the immunogen. Although the antiserum reacted equally with an isomer [L-Ala2]deltorphin I, virtually no cross-reaction occurred with other analogues such as [D-Ala2]deltorphin II (Tyr-D-Ala-Phe-Glu-Val-Val-Gly.NH2) and similar peptides lacking the C-terminal glycine amide. Therefore, the major epitope for immunorecognition appeared to be in the C-terminal region which is known to be the specific domain for delta opioid receptor selectivity. Immunohistochemical study using this antiserum revealed positive neuronal structures in some specific systems of the mouse brain.

Ontogeny of PACAP immunoreactivity in extrinsic and intrinsic innervation of chicken gut.

An immunohistochemical study was conducted on the ontogeny of pituitary adenylate cyclase-activating polypeptide-27 (PACAP) immunoreactive elements within the extrinsic and intrinsic nerve supply of the chicken embryo gut. The first PACAP-immunoreactivity was detected in the extrinsic nerve supply at E 4 within the pharyngeal region and the primary sympathetic chain. At E 5.5 it appeared in the vagus nerve, the spinal cord, the secondary sympathetic chain, some perivascular plexuses and the Remak ganglion. In the intrinsic nerve supply, the first PACAP-immunoreactive elements were shown at E 4.5-E 5 in the mesenchymal bud of the proventriculus/gizzard. Then they gradually appeared also cranially and caudally both in myenteric and submucous plexuses.

Production and immunohistochemical application of antiserum against Tyr-D-Ala-Phe, a N-terminal tripeptide common to dermorphin/deltorphin family.

Tyr-D-Ala-Phe is a N-terminal sequence commonly found in a peptide family including dermorphin and deltorphin. The tripeptide was synthesized and conjugated with poly L-lysine. Nuclear magnetic resonance (NMR) indicated that approximately 38 molecules of the tripeptide were bound to each molecule of poly L-lysine. The conjugate was used to immunize rabbits, and high titer antisera were obtained. An IgG fraction was purified by protein G affinity chromatography. A specific antibody to the tripeptide was then obtained by affinity chromatography using formylcellulofine conjugated with Tyr-D-Ala-Phe. On immunospot assay, the best IgG antibody was capable of detecting 125 ng of Tyr-D-Ala-Phe but failed to react even with 2.0 microg of Tyr-L-Ala-Phe or poly L-lysine. Our immunohistochemical examination selectively localized the secretory glands of frog skin.

Immunohistochemical localization of amylin in rat brainstem.

Immunohistochemical studies were conducted on rat brainstem using a specific polyclonal antiserum against the COOH-terminal (25-37) of human amylin. Amylin-immunoreactive cell bodies were observed in the vestibular, cochlear, trapezoid, and inner cerebellar nuclei and in the mesencephalic nucleus of trigeminal nerve. Positive cell bodies were also found in lateral, gigantocellular and magnocellular reticular nuclei. Numerous amylin-immunoreactive nerve fibers were shown in the trigeminal spinal tract, in the solitary area and in the area postrema. Amylin-immunoreactive cell bodies were often surrounded by a network of tyrosine hydroxylase-immunoreactive nerve fibers. These results provide morphologic evidence that amylin may play a role in some discrete sensory functions.

Immunohistochemical demonstration of [D-Ala2]deltorphin-I in amacrine cells of rat retina.

In an immunohistochemical study, a specific antiserum raised against [D-Ala2]deltorphin-I (DADTI), a highly selective ligand for delta opioid receptors, was used to demonstrate immunoreactive structures in the rat retina. [D-Ala2]Deltorphin-I immunoreactivity occurred in a subpopulation of the retinal amacrine cells situated in the inner nuclear layer. Their stained processes were mainly distributed in the sublaminae 1 and 3 of the inner plexiform layer. A few positive cells, probably displaced amacrine cells, were also seen in the ganglion cell layer. Double immunostaining revealed that 12.8% of DADTI-immunoreactive cells costored GABA and 27.7% costored neuropeptide Y, whereas only few DADTI-positive cells colocalized tyrosine hydroxylase (0.3%) and almost no other peptides. These findings suggest that some retinal amacrine cells possess DADTI-like molecule(s), possibly acting as neurotransmitter(s) or neuromodulator(s).

Demonstration of [D-Ala2]deltorphin I-like immunoreactivity in mucosal epithelial cells of the rat gastrointestinal tract.

Using a specific antiserum to [D-Ala2]deltorphin I (DADTI), a delta-opioid receptor ligand, the localization of positive structures was studied in rat gastrointestinal tract by immunocytochemistry. Immunoreactive staining was not detected in the stomach, colon, or neuronal elements of any gastrointestinal tissue. However, positive cells were distributed in the mucosal epithelium of the small intestine, including the duodenum, jejunum, and ileum. The density of positive cells was highest at a proximal part of the jejunum and was gradually decreased toward the duodenum or the distal end of the intestine. These positive cells had spindle-like somata that tended to locate more closely to the lumen compared with nonimmunoreactive cells. Some of the positive cells extended cytoplasmic basal processes toward the lamina propria. Immunoelectron microscopy revealed that positive reaction products occurred within the secretory granules as well as in the cytoplasm. Because these positive granules were frequently observed in the apical cytoplasm beneath the microvilli, it is suggested that the DADTI-like molecule(s) may be secreted to the lumen.

Sauvagine/urotensin I-like immunoreactivity in the caudal neurosecretory system of a seawater fish Diplodus sargus L. in normal and hyposmotic milieu.

We report the presence of sauvagine/urotensin I-like immunoreactive (SV/UI-LI) elements in the caudal neurosecretory system of a teleost (Diplodus sargus L.) collected from aquaria tanks of the Aquaculture Center (Talassographic Institut of CNR) of Messina or maintained in an hyposmotic milieu for different periods. In normal specimens, SV/UI-LI material was recognizable in discrete or little amounts both in Dahlgren cell cytoplasm and in their axons that reach the urophysis. On the contrary, the specimens transferred in an hyposmotic milieu showed a fast and dramatic increase of immunoreactivity mainly in neurohemal endings of the urophysis. This suggests a physiological role of caudal neurosecretory products on osmoregulatory mechanisms.