Non-invasive ventilation for acute respiratory failure.

Non-invasive ventilation has become an increasingly utilised tool for the treatment of acute respiratory failure. Potential benefits include a decreased incidence of intubation, duration of hospital stay and mortality. Non-invasive ventilation is also being used more and more outside the intensive care environment. Successful use of non invasive ventilation involves knowledge of its indications, contraindications and limitations, and appropriate patient selection. This article reviews these issues as well as the practical application of non invasive ventilation in the acute setting.

Exploring the sialome of the blood-sucking bug Rhodnius prolixus.

Rhodnius prolixus is a Hemiptera that feeds exclusively on vertebrate blood in all life stages. Its salivary glands produce potent pharmacological substances that counteract host hemostasis, including anti-clotting, anti-platelet, and vasodilatory substances. To obtain a further insight into the salivary biochemical and pharmacological complexity of this insect, a cDNA library was randomly sequenced, and salivary gland homogenates were fractionated by HPLC to obtain aminoterminal sequences of abundantly expressed proteins. Results indicate a remarkable expansion of the lipocalin family in Rhodnius salivary glands, among other protein sequences described. A summary of 31 new full length proteins deducted from their mRNA sequence is described, including several new members of the nitrophorin, triabin, and pallidipin families. The electronic version of the complete tables is available at http://www.ncbi.nlm.nih.gov/projects/vectors/rhodnius_prolixus.

Toward a description of the sialome of the adult female mosquito Aedes aegypti.

To describe the set of mRNA and protein expressed in the salivary glands (sialome) of Aedes aegypti mosquitoes, we randomly sequenced a full-length cDNA library of this insect and performed Edman degradation of PVDF-transferred protein bands from salivary homogenates. We found 238 cDNA clusters which contained those coding for 10 of the 11 proteins found by aminoterminal degradation. All six previously described salivary proteins were found in this library. Full-length sequences of 32 novel cDNA sequences are reported, one of which is the product of a transposable element. Among the 31 novel protein sequences are 4 additional members of the D7 protein family; 4 novel members of the antigen 5 family (a protein family not reported in Aedes); a novel serpin; a novel member of the 30-kDa allergen of Ae. Aegypti; a secreted calreticulin; 2 proteins similar to mammalian angiopoietins; adenosine deaminase; purine hydrolase; lysozyme; a C-type lectin; 3 serine proteases, including one with high similarity to Bombyx prophenoloxidase activating enzyme; 2 proteins related to invertebrate immunity; and several sequences that have no significant matches to known proteins. The possible role of these proteins in blood and sugar feeding by the mosquito is discussed.

Seasonal variation in admission rates to intensive care units.

Intensive care physicians perceive that there is seasonal variation in the number of admissions to critical care services. There is, however, little published evidence to support this belief. Data were therefore collected from five adjacent critical care units in the eastern region over a period of 8 years, in order to quantify any seasonal variation that may exist. Data on 16 355 critically ill patients were obtained between 1992 and 2000. Analysis showed clear winter peaks; December had a 30% higher admission rate than the quietest month, February. There was a small, but increasing, summer peak. The admission rate also exhibits an increasing linear trend, equivalent to a 6.6% annual increase in admissions per critical care bed. We conclude that there is significant seasonal variation in critical care activity, and that this is important to consider when planning services.

Toward a defined anti-Leishmania vaccine targeting vector antigens: characterization of a protective salivary protein.

Leishmania parasites are transmitted to their vertebrate hosts by infected phlebotomine sand fly bites. Sand fly saliva is known to enhance Leishmania infection, while immunity to the saliva protects against infection as determined by coinoculation of parasites with vector salivary gland homogenates (SGHs) or by infected sand fly bites (Kamhawi, S., Y. Belkaid, G. Modi, E. Rowton, and D. Sacks. 2000. Science. 290:1351-1354). We have now characterized nine salivary proteins of Phlebotomus papatasi, the vector of Leishmania major. One of these salivary proteins, extracted from SDS gels and having an apparent mol wt of 15 kD, was able to protect vaccinated mice challenged with parasites plus SGH. A DNA vaccine containing the cDNA for the predominant 15-kD protein (named SP15) provided this same protection. Protection lasted at least 3 mo after immunization. The vaccine produced both intense humoral and delayed-type hypersensitivity (DTH) reactions. B cell-deficient mice immunized with the SP15 plasmid vaccine successfully controlled Leishmania infection when injected with Leishmania plus SGH. These results indicate that DTH response against saliva provides most or all of the protective effects of this vaccine and that salivary gland proteins or their cDNAs are viable vaccine targets against leishmaniasis.

Non-invasive positive pressure ventilation for severe thoracic trauma.

A 35-year-old man was admitted to the intensive care unit (ICU) following a road traffic accident. He had sustained severe trauma to the left side of his chest, as well as other musculoskeletal injuries. After a short initial period of ventilation of the lungs via a tracheal tube, he was managed using a combination of continuous positive airway pressure and non-invasive positive pressure ventilation. He avoided ventilator-associated pneumonia, and spent a large part of his time on the ICU without any invasive monitoring lines, another potential focus of infection. He was discharged from the ICU after 25 days without having suffered any septic complications. The role of non-invasive positive pressure ventilation in severe thoracic trauma is discussed.

An assessment of the staffing level required for a high-dependency unit.

High-dependency units are increasing in number and becoming an ever more important part of a hospital's facilities. The optimum staffing ratio is unknown, but the Department of Health and the Intensive Care Society recommend a level of one nurse to two patients. We recorded Therapeutic Intervention Scoring System-28 scores and Nurse Dependency Scores for all admissions to our adult, general high-dependency unit over 7 months. We found a weak correlation between the nurse dependency score and the Therapeutic Intervention Scoring System-28 score. The median Therapeutic Intervention Scoring System-28 score was 23 points (interquartile range 19-26), and the median Nurse Dependency Score was 1.0. These results are approximately two-thirds of those for European intensive care units. We conclude that a nurse-to-patient ratio of 1:2 may be insufficient for an adult general high-dependency unit, and would recommend a nurse-to-patient ratio of 2:3.

Endogenous peptides with distinct amino acid anchor residue motifs bind to HLA-A1 and HLA-B8.

Distinct amino acid (aa) residue motifs for peptides binding to HLA-A1 and HLA-B8 were identified by sequence analyses of reversed-phase HPLC fractions containing endogenous peptides derived from these HLA molecules. Fifteen different primary sequences were determined for HLA-A1-associated peptides, 12 of which were nine aa in length. Common features among these peptide sequences were Tyr at the COOH-terminus, a negatively charged aa (usually Glu) at position 3 (P3), and Pro at P4. Twenty-seven different primary sequence assignments were made for HLA-B8-associated peptides, most of which were eight aa in length. Lys, and in a few cases Arg, predominated at P3 and P5; Leu and Pro predominated at P2, and Leu was the preferred COOH-terminal residue. Unlike all other human class I molecules whose peptide-binding properties have been studied, both HLA-A1 and HLA-B8 endogenous peptide sequences have a dominant anchor residue at P3, and these aa are opposite in charge to the aa at position 156 of the peptide-binding site. Synthetic peptides corresponding to endogenous peptide sequences bound to their respective HLA molecules in vitro, indicating that they derive from peptides bound to HLA and not from copurifying contaminants. Eight of the HLA-A1 and HLA-B8 endogenous peptide sequences matched intracellularly expressed proteins found in protein sequence data bases. The HLA-A1 peptide-binding motif was then used to identify potential antigenic peptides from influenza A viral proteins that bound to HLA-A1 in vitro.

The murine Niemann-Pick type C lesion affects testosterone production.

We have determined the effects of the Niemann-Pick type C (NPC) lesion, which impairs transport of cholesterol from lysosomes, on the androgenic status of male NPC mice. The mice have low serum testosterone levels resulting from decreased testosterone secretion. Testosterone secretion is reduced in NPC mouse testes incubated with 8-bromo-cAMP, 20 alpha-hydroxycholesterol, and pregnenolone compared to testosterone release by normal mouse testes under identical conditions. Ultrastructural examination of testes revealed a paucity of lipid droplets, extensive accumulation of inclusion bodies, and distorted endoplasmic reticulum in Leydig cells of adult NPC mice. The hypoandrogenemia caused systemic deficiencies in NPC mice. Seminal vesicles, a testosterone-responsive tissue, were underdeveloped in NPC male mice. The testosterone-responsive kidney beta-glucuronidase activity was also underexpressed. Seminal vesicle mass and beta-glucuronidase activity were increased by testosterone treatment of NPC mice. Many hepatic proteins, identified by microsequencing, were also deficient in NPC male mice. Levels of alpha 2-mu-globulin, glutathione S-transferase-pi, carbonic anhydrase-III, and selenium-binding protein increased in normal male mice during puberty, but did not increase in the NPC male mice. Based on the increases in protein expression during puberty, differential expression in males and females, and the reported involvement of androgens in regulating expression of some of these proteins, deficient expression of most of these proteins in male NPC mice appears to result from low testosterone levels. We conclude that a defect in testicular testosterone production in NPC male mice causes a pleiotropic deficiency in androgen-sensitive expression of proteins in various organs.

Rhodococcus species fatal infection in an immunocompetent host.

A 24-year-old woman had fatal pneumonia-associated adult respiratory distress syndrome caused by Rhodococcus species. Histological examination of lung biopsy tissue showed intracellular coccobacillary microorganisms. Antimicrobial susceptibility tests on the patient's blood isolate showed that it was resistant to clindamycin and norfloxacin but susceptible to several other antimicrobial agents. Also, the isolate's biochemical reactions and pattern of RNA gene-containing restriction fragments were significantly different from those of the 20 recognized Rhodococcus species, suggesting that this patient's infection was caused by an as yet uncharacterized Rhodococcus species. Of the 17 human cases of nonequi Rhodococcus species infection reported to date (including the current case), nine patients were immunocompetent, five had disseminated infection, and four died. Further studies will be required to unequivocally establish the species status of this patient's Rhodococcus isolate biochemically and genetically.

Carotid endarterectomy: operative risks, recurrent stenosis, and long-term stroke rates in a modern series.

To determine whether carotid endarterectomy (CEA) safely and effectively maintained a durable reduction in stroke complications over an extended period, we reviewed our data on 478 consecutive patients who underwent 544 CEA's since 1976. Follow-up was complete in 83% of patients (mean 44 months). There were 7 early deaths (1.3%), only 1 stroke related (0.2%). Perioperative stroke rates (overall 2.9%) varied according to operative indications: asymptomatic, 1.4%; transient ischemic attacks (TIA)/amaurosis fugax (AF), 1.3%; nonhemispheric symptoms (NH), 4.9%; and prior stroke (CVA), 7.1%. Five and 10-year stroke-free rates were 96% and 92% in the asymptomatic group, 93% and 87% in the TIA/AF group, 92% and 92% in the NH group, and 80% and 73% in the CVA group. Late ipsilateral strokes occurred infrequently (8 patients, 1.7%). Late deaths were primarily cardiac related (51.3%). Stroke-free rates were significantly (p less than 0.0001) greater than stroke-free survival rates, confirming a non-stroke related cause for late death. Restenoses greater than 50% according to duplex scanning developed in 13%, most (67%) within 2 years after CEA. Most of these (77%) were asymptomatic, and only 0.3% (1 patient) presented with a permanent neurologic deficit. The results of carotid endarterectomy are superior to those of optimal medical management in symptomatic and asymptomatic patients in terms of long-term stroke prevention. When low perioperative stroke mortality/morbidity rates are achieved, carotid endarterectomy is justified for treatment of patients with carotid bifurcation disease.

Endothelins belong to the assortment of mast cell-derived and mast cell-bound cytokines.

Local accumulation of endothelins (ETs) as cytokine-like factors via autocrine/paracrine mechanisms seems to represent an important aspect of their pathophysiological action. This assumption prompted us to investigate mast cells as a possible source of these peptides. With the use of a combination of high-performance liquid chromatography and a radioimmunoassay specific for endothelin-1 (ET-1), 3-week-old cultures of primary murine bone marrow mast cells (BMMC) as well as various mast cell lines were shown to contain and secrete immunoreactive ET-1. The amounts of this peptide were constitutively high in cellular extracts of BMMC, while there was considerable variation in the basal cellular content among mast cell lines, ranging from high (C57) to undetectable (RBL) levels. Treatment of the cells with the combination of phorbol myristate acetate (PMA) and A23187 for 5 h led to induction of ET-1 production in all cases tested. In contrast to the rapid stimulation by PMA/A23187 of histamine release from BMMC or C57 cells, however, no ET-1 secretory response was noted as early as 30 min after this combined treatment. Moreover, stimulation of mast cells with crosslinked IgE for 30 min or 5 h did not affect ET-1 secretion, suggesting that mast cell ET-1 release is not directly related to mast cell degranulation. After exposure of the cells to crosslinked IgE for 20 h, however, there was a distinct increase in immunoreactive ET-1 in the medium, to approximate 10 times the basal level. Polymerase chain reaction (PCR) analysis of mRNA expression in mast cells revealed that the amount of ET-1 PCR product, which is low or undetectable under nonstimulated conditions, is enhanceable by both PMA/A23187 and crosslinked IgE. The IgE-mediated induction kinetics for ET-1 mRNA parallel the kinetics obtained with PMA/A23187, albeit at somewhat lower levels. With the use of fluorescent ligand binding/flow cytometry as a screening method and a radioreceptor assay as the confirming method, mast cells were found to express a single class of high affinity ET receptors with distinct selectivity for ET-1 and a pharmacological profile resembling that of the ETA type ET receptor. Stimulation of mast cell ET-1 receptors did not provoke histamine release, nor did it result in a mitogenic response of BMMC. In conclusion, mast cells synthesize and secrete ET-1 and have ET receptors, suggesting that ET-1 may participate in mediating mast cell-related long-term changes in the microenvironment, e.g., in smooth muscle tone or the proliferation rate of fibroblasts.

Poly(A) polymerase and a dissociable polyadenylation stimulatory factor encoded by vaccinia virus.

mRNA made in eukaryotic cells typically has a 3' poly(A) tail that is added posttranscriptionally. To investigate mechanisms by which 3' poly(A) is formed, we identified the genes for the two vaccina virus-encoded polypeptides, VP55 and VP39. Primer-dependent polyadenylation activity was associated exclusively with purified VP55-VP39 heterodimer, which, although stable to column chromatography and glycerol gradient sedimentation, was readily dissociated by antibody to an N-terminal peptide of VP55. Poly(A) polymerase activity was associated with immunopurified VP55, but not with immunopurified or chromatographically purified VP39. VP39 was, however, required for the formation of long poly(A) molecules, in conjunction with either purified VP55 or low concentrations of the heterodimer, and was shown to bind free poly(A). Thus, a catalytic polypeptide and a dissociable poly(A)-binding stimulatory factor each contribute to poly(A) tail formation. No prokaryotic or eukaryotic homologs of either polypeptide were detected in sequence data bases, consistent with the absence of previously reported poly(A) polymerase genes from any source.

Serologic discrimination of human T cell lymphotropic virus infection by using a synthetic peptide-based enzyme immunoassay.

Synthetic peptides corresponding with unique regions of the envelope glycoproteins (gp46) of human T cell lymphotropic viruses (HTLVs) were used in an enzyme immunoassay to determine if HTLV-I and -II infections could be discriminated. Two synthetic HTLV-I sequence-derived peptides, Env-1 (amino acids 191-215) and Env-5 (amino acids 242-257), reacted with 92% and 100% of the serum specimens (n = 52) from HTLV-I-infected persons, respectively. Although a small percentage (8.6%) of serum specimens from persons infected with HTLV-II cross-reacted with Env-1, none of these specimens reacted with Env-5. Peptide Env-2 encoded by the envelope region of HTLV-II (amino acids 187-210) reacted with serum specimens from both HTLV-I (94%)- and HTLV-II (74%)-infected patients, whereas Env-6, another HTLV-II peptide (amino acids 238-254), reacted with less than 6% of the specimens. Therefore, the Env-5 peptide with amino acid sequence SerProAsnValSerValProSerSerSerSerThrProLeuLeuTyr represents an immunodominant domain of HTLV-I that is recognized by serum antibodies from all HTLV-I-infected persons. Moreover, the Env-5-based ELISA allows a categorical distinction between the closely related HTLV-I and -II infections.

Therapeutic response to intravenous infusions of glucocerebrosidase in a patient with Gaucher disease.

Enzyme replacement has been under consideration as a therapeutic strategy for patients with Gaucher disease for more than two decades. Previous studies indicated that single injections of purified glucocerebrosidase reduced the amount of storage material in the liver. It was important to determine whether administration of exogenous enzyme on a regular basis would be of clinical benefit. We report here that weekly i.v. infusions of a macrophage-targeted preparation of human placental glucocerebrosidase in a child with type 1 Gaucher disease increased hemoglobin from 6.9 +/- 0.8 g/dl (+/- 1 SD) to 10.2 +/- 0.4 g/dl (+/- 1 SD) over a 20-week period. The platelet count also increased from a pretreatment value of 30,000 +/- 7000/mm3 (+/- 1 SD) to 54,000 +/- 11,000/mm3 (+/- 1 SD). Phagocytic activity in the spleen decreased during the period of enzyme administration, and there was radiographic evidence of skeletal improvement. These observations document objective clinical responses to enzyme supplementation in a patient with a sphingolipid storage disorder.

Isolation and characterization of TGF-beta 2 and TGF-beta 5 from medium conditioned by Xenopus XTC cells.

TGF-beta 2 and -beta 5 have been purified from medium conditioned by Xenopus cultured cells (XTC) and identified based on their N-terminal amino acid sequence analysis and biological activity. When applied in high concentrations, Xenopus TGF-beta 2, like porcine TGF-beta 2, induces expression of mesodermal markers from cultured Xenopus ectodermal explants, whereas TGF-beta 5 is inactive in this assay. However, the TGF-beta 's could be separated from the major mesoderm-inducing activity present in XTC medium. Xenopus TGF-beta 2 and -beta 5 are approximately equivalent to TGF-beta 1 in their abilities to inhibit the growth of mink lung CCL-64 cells, induce anchorage-independent growth of rat NRK cells, inhibit the proliferation and antibody secretion of human B-lymphocytes, and stimulate chemotaxis of human monocytes. These data establish the functional activity of TGF-beta 5 and suggest that more complex multicellular systems, in contrast to most isolated cells, discriminate between the different TGF-beta s.