Positive airway pressure therapy for sleep-disordered breathing confers short-term benefits to patients with spinal cord injury despite widely ranging patterns of use.

STUDY DESIGN: Prospective, cohort study. OBJECTIVES: To evaluate the effectiveness of bi-level positive airway pressure (PAP) therapy and the patterns of use for sleep-disordered breathing (SDB) in individuals with spinal cord injury (SCI). SETTING: Academic tertiary care center, USA. METHODS: Overall, 91 adults with C1-T6 SCI for ≥3 months were recruited and 74 remained in the study to be evaluated for SDB and follow-up. Individuals with SDB but no nocturnal hypercapnia (NH) were prescribed auto-titrating PAP. Those with NH were prescribed PAP with volume-assured pressure support. Device downloads and overnight transcutaneous capnography were performed at 3, 6, and 12 months to quantify PAP use and effectiveness. Participants kept daily event logs, and quality of life (QOL) questionnaires were performed after 3, 6, and 12 months. RESULTS: Overall, 45% of 91 participants completed the study. There was great diversity among SCI patients in PAP utilization; after 3 months, 37.8% of participants used PAP for ≥70% nights and ≥240 min per night, whereas 42.2% seldom used PAP and 20% used PAP sporadically or for short periods. PAP therapy was effective in improving OSA in 89% and nocturnal hypercapnia in 77%. Higher PAP pressures predicted higher levels of device use. There were marked reductions in symptoms of autonomic dysreflexia (AD) and orthostatic hypotension as well as some improved indices of QOL. CONCLUSIONS: Despite widely diverse patterns of use, PAP therapy may have short-term benefits with regard to QOL and reducing episodes of dizziness and autonomic dysreflexia.

Simplified Approach to Diagnosing Sleep-Disordered Breathing and Nocturnal Hypercapnia in Individuals With Spinal Cord Injury.

OBJECTIVE: To evaluate a strategy of home-based testing to diagnose sleep-disordered breathing and nocturnal hypercapnia in individuals with spinal cord injury (SCI). DESIGN: Case series. SETTING: Referral center. PARTICIPANTS: Adults with C1-T6 SCI (N=81). Individuals were eligible if ≥ 18 years old, with SCI of ≥ 3 months' duration, living within 100 miles of the study site, and not meeting exclusion criteria. Of the 161 individuals recruited from the SCI Model System database who were not enrolled, reasons were not interested in participating, change of location, prior positive pressure ventilation use, or medical contraindication. Ten individuals did not complete the study. INTERVENTIONS: Performance of an unsupervised home sleep apnea test combined with transcutaneous partial pressure of carbon dioxide/oxygen saturation by pulse oximetry monitoring. MAIN OUTCOME MEASURES: Prevalence of sleep-disordered breathing and nocturnal hypercapnia. Clinical and physiological variables were examined to determine which, if any, correlate with the severity of sleep-disordered breathing. RESULTS: Obstructive sleep apnea (OSA) was found in 81.3% of individuals, central sleep apnea (CSA) was found in 23.8%, and nonspecific hypopnea events, where respiratory effort was too uncertain to classify, were present in 35%. Nonspecific hypopnea events correlated strongly with CSA but weakly with OSA, suggesting that conventional sleep apnea test scoring may underestimate central/neuromuscular hypopneas. Nocturnal hypercapnia was present in 28% and oxygen desaturation in 18.3%. Neck circumference was the primary predictor for OSA, whereas baclofen use and obstructive apnea/hypopnea index weakly predicted CSA. Awake transcutaneous partial pressure of carbon dioxide and CSA were only marginally associated with nocturnal hypercapnia. CONCLUSIONS: Unsupervised home sleep apnea testing with transcutaneous capnography effectively identifies sleep-disordered breathing and nocturnal hypercapnia in individuals with SCI.

Domain 2 of uPAR regulates single-chain urokinase-mediated angiogenesis through ß1-integrin and VEGFR2.

How single-chain urokinase (ScuPA) mediates angiogenesis is incompletely understood. ScuPA (≥4 nM) induces phosphorylated (p)ERK1/2 (MAPK44 and MAPK42) and pAkt (Ser(473)) in umbilical vein and dermal microvascular endothelial cells. Activation of pERK1/2 by ScuPA is blocked by PD-98059 or U-0126, and pAkt (Ser(473)) activation is inhibited by wortmannin or LY-294002. ScuPA (32 nM) or protease-inhibited two-chain urokinase stimulates pERK1/2 to the same extent, indicating that signaling is not dependent on enzymatic activity. ScuPA induces pERK1/2, but not pAkt (Ser(473)), in SIN1(-/-) cells, indicating that the two pathways are not identical. Peptides from domain 2 of the urokinase plasminogen activator receptor (uPAR) or domain 5 of high-molecular-weight kininogen compete with ScuPA for the induction of pERK1/2 and pAkt (Ser(473)). A peptide of the integrin-binding site on uPAR, a ß1-integrin peptide that binds uPAR, antibody 6S6 to ß1-integrin, tyrosine kinase inhibitors AG-1478 or PP3, and small interfering RNA knockdown of VEFG receptor 2, but not HER1-HER4, blocked ScuPA-induced pERK1/2 and pAkt (Ser(473)). ScuPA-induced endothelial cell proliferation was blocked by inhibitors of pERK1/2 and pAkt (Ser(473)), antibody 6S6, and uPAR or kininogen peptides. ScuPA initiated aortic sprouts and Matrigel plug angiogenesis in normal, but not uPAR-deficient, mouse aortae or mice, respectively, but these were blocked by PD-98059, LY-294002, AG-1478, or cleaved high-molecular-weight kininogen. In summary, this investigation indicates a novel, a nonproteolytic signaling pathway initiated by zymogen ScuPA and mediated by domain 2 of uPAR, ß1-integrins, and VEGF receptor 2 leading to angiogenesis. Kininogens or peptides from it downregulate this pathway.

Home-based overnight transcutaneous capnography/pulse oximetry for diagnosing nocturnal hypoventilation associated with neuromuscular disorders.

OBJECTIVE: To determine the utility of home-based, unsupervised transcutaneous partial pressure of carbon dioxide (tc-Pco(2)) monitoring/oxygen saturation by pulse oximetry (Spo(2)) for detecting nocturnal hypoventilation (NH) in individuals with neuromuscular disorders. DESIGN: Retrospective case series analyzed consecutively. SETTING: Multidisciplinary neuromuscular respiratory failure (NMRF) clinic at an academic institution. PARTICIPANTS: Subjects (N=35, 68.6% men; mean age, 46.9y) with spinal cord injury (45.7%) or other neuromuscular disorders underwent overnight tests with tc-Pco(2)/Spo(2) monitoring. Fifteen (42.9%) were using nocturnal ventilatory support, either bilevel positive airway pressure (BiPAP) or tracheostomy ventilation (TV). INTERVENTIONS: A respiratory therapist brought a calibrated tc-Pco(2)/Spo(2) monitor to the patient's home and provided instructions for data collection during the subject's normal sleep period. Forced vital capacity (FVC), body mass index (BMI), and exhaled end-tidal Pco(2) (ET-Pco(2)) were recorded at a clinic visit before monitoring. MAIN OUTCOME MEASURES: Detection of NH (tc-Pco(2) ≥50mmHg for ≥5% of monitoring time). Data were also analyzed to determine whether nocturnal oxygen desaturation (Spo(2) ≤88% for ≥5% of monitoring time), FVC, BMI, or daytime ET-Pco(2) could predict the presence of NH. RESULTS: NH was detected in 18 subjects (51.4%), including 53.3% of those using BiPAP or TV. NH was detected in 43.8% of ventilator-independent subjects with normal daytime ET-Pco(2) (present for 49.4%±31.5% [mean ± SD] of the study period), and in 75% of subjects with an elevated daytime ET-Pco(2) (present for 92.3%±8.7% of the study period). Oxygen desaturation, BMI, and FVC were poor predictors of NH. Only 3 attempted monitoring studies failed to produce acceptable results. CONCLUSIONS: Home-based, unsupervised monitoring with tc-Pco(2)/Spo(2) is a useful method for diagnosing NH in NMRF.

Factor XII stimulates ERK1/2 and Akt through uPAR, integrins, and the EGFR to initiate angiogenesis.

Factor XII (FXII) and high molecular weight kininogen (HK) mutually block each other's binding to the urokinase plasminogen activator receptor (uPAR). We investigated if FXII stimulates cells by interacting with uPAR. FXII (3-62nM) with 0.05mM Zn(2+) induces extracellular signal-related kinase 1/2 (ERK1/2; mitogen-activated protein kinase 44 [MAPK44] and MAPK42) and Akt (Ser473) phosphorylation in endothelial cells. FXII-induced phosphorylation of ERK1/2 or Akt is a zymogen activity, not an enzymatic event. ERK1/2 or Akt phosphorylation is blocked upstream by PD98059 or Wortmannin or LY294002, respectively. An uPAR signaling region for FXII is on domain 2 adjacent to uPAR's integrin binding site. Cleaved HK or peptides from HK's domain 5 blocks FXII-induced ERK1/2 and Akt phosphorylation. A beta(1) integrin peptide that binds uPAR, antibody 6S6 to beta(1) integrin, or the epidermal growth factor receptor (EGFR) inhibitor AG1478 blocks FXII-induced phosphorylation of ERK1/2 and Akt. FXII induces endothelial cell proliferation and 5-bromo-2'deoxy-uridine incorporation. FXII stimulates aortic sprouting in normal but not uPAR-deficient mouse aorta. FXII produces angiogenesis in matrigel plugs in normal but not uPAR-deficient mice. FXII knockout mice have reduced constitutive and wound-induced blood vessel number. In sum, FXII initiates signaling mediated by uPAR, beta(1) integrin, and the EGFR to induce human umbilical vein endothelial cell proliferation, growth, and angiogenesis.

An obligate role for membrane-associated neutral sphingomyelinase activity in orienting chemotactic migration of human neutrophils.

For polymorphonuclear neutrophils (PMNs) to orient migration to chemotactic gradients, weak external asymmetries must be amplified into larger internal signaling gradients. Lipid mediators, associated with the plasma membrane and within the cell, participate in generating these gradients. This study examined the role in PMN chemotaxis of neutral sphingomyelinase (N-SMase), a plasma membrane-associated enzyme that converts sphingomyelin to ceramide. A noncompetitive N-SMase inhibitor, GW4869 (5 mM, 5 minutes), did not inhibit PMN motility (as percentage of motile cells, or mean cell velocity), but it abrogated any orientation of movement toward the source of the chemotaxin, formylmethionylleucylphenylanaline (FMLP) (net displacement along the gradient axis in micrometers, or as percentage of total migration distance). This defect could be completely reversed by treatment with lignoceric ceramide (5 µg/ml, 15 minutes). Immunolocalization studies demonstrated that N-SMase (1) distributes preferentially toward the leading edge of some elongated cells, (2) is associated with the plasma membrane, (3) is more than 99.5% localized to the cytofacial aspect of the plasma membrane, (4) is excluded from pseudopodial extensions, and (5) increases rapidly in response to FMLP. Morphologically, the inhibition of N-SMase limited cellular spreading and the extension of sheet-like pseudopods. Elongated PMNs demonstrated a polarized distribution of GTPases, with Rac 1/2 accumulated at, and RhoA excluded from, the front of the cell. This polarity was negated by N-SMase inhibition and restored by lignoceric ceramide. We conclude that N-SMase at the cytofacial plasma membrane is an essential element for the proper orientation of PMNs in FMLP gradients, at least in part by polarizing the distribution of Rac 1/2 and RhoA GTPases.

FcgammaRI ligation leads to a complex with BLT1 in lipid rafts that enhances rat lung macrophage antimicrobial functions.

Leukotriene (LT) B(4) is generated in response to engagement of the Fc gamma receptor (Fc gamma R) and potently contributes to Fc gamma R-mediated antimicrobial functions in pulmonary alveolar macrophages. In this study, we report that the LTB(4) receptor leukotriene B(4) receptor 1 (BLT1) redistributes from nonlipid raft (LR) to LR membrane microdomains upon immunoglobulin G-red blood cell, but not LTB(4), challenge. Cholesterol depletion to disrupt LRs abolished LTB(4)-induced enhancement of phagocytosis, microbicidal activity, and signaling. The dependence on LR integrity for BLT1 signaling correlated with formation of a complex consisting of BLT1, its primary coupled G protein G alpha i3, Src kinase, and Fc gamma RI within LRs. This association was dependent on Src-mediated phosphorylation of BLT1. These data identify a novel form of regulation in which engagement of a macrophage immunoreceptor recruits a stimulatory G protein-coupled receptor into a LR microdomain with resultant enhanced antimicrobial signaling.

Migrating human neutrophils exhibit dynamic spatiotemporal variation in membrane lipid organization.

Highly ordered sphingolipid-enriched lipid raft microdomains (LRMs) within plasma membranes purportedly function as specialized signaling platforms. Leukocyte migration is believed to entail LRM redistribution, but progress in studying LRMs in situ during cell movement has been limited. By using an improved method for imaging the spectral shift of the environmentally sensitive probe, laurdan (expressed as a generalized polarization function), the plasma membrane order (i.e., tight packing of membrane bilayer lipids) of human polymorphonuclear neutrophils (PMNs) was mapped in real time during migration. Morphologically polarized PMNs exhibited prominent LRM clusters at the uropod, where in every instance membrane order was found to oscillate with mean periodicities of 37.0 ± 1.46 and 149.9 ± 9.0 seconds (P < 0.01). LRM aggregates were also demonstrated in punctate and clustered distributions of nonpolarized cells and transiently at the lamellipodia of polarized PMNs. Cellular polarization was not accompanied by an overall increase in membrane order. LRM disorganization with methyl-ß-cyclodextrin had small negative effects on cell velocity, but it abrogated directionally biased migration toward chemotactic gradients of FMLP or leukotriene B(4). LRMs disruption also caused redistribution of Rac 1/2 GTPase and GM3 ganglioside away from the lamellipodium, as well as extension of multiple pseudopods simultaneously or in rapid succession, rather than formation of a defined leading edge. Thus, we demonstrate that the plasma membrane order of migrating PMNs changes dynamically, with prominent oscillations consistently seen at the uropod. These findings solidify the existence of rapidly reorganizing LRMs in situ and support a role for LRMs in chemotaxin responsiveness.

Sleep disordered breathing in spinal cord injury: A systematic review.

CONTEXT: Spinal cord injury commonly results in neuromuscular weakness that impacts respiratory function. This would be expected to be associated with an increased likelihood of sleep-disordered breathing. OBJECTIVE: (1) Understand the incidence and prevalence of sleep disordered breathing in spinal cord injury. (2) Understand the relationship between injury and patient characteristics and the incidence of sleep disordered breathing in spinal cord injury. (3) Distinguish between obstructive sleep apnea and central sleep apnea incidence in spinal cord injury. (4) Clarify the relationship between sleep disordered breathing and stroke, myocardial infarction, metabolic dysfunction, injuries, autonomic dysreflexia and spasticity incidence in persons with spinal cord injury. (5) Understand treatment tolerance and outcome in persons with spinal cord injury and sleep disordered breathing. METHODS: Extensive database search including PubMed, Cochrane Library, CINAHL and Web of Science. RESULTS: Given the current literature limitations, sleep disordered breathing as currently defined is high in patients with spinal cord injury, approaching 60% in motor complete persons with tetraplegia. Central apnea is more common in patients with tetraplegia than in patients with paraplegia. CONCLUSION: Early formal sleep study in patients with acute complete tetraplegia is recommended. In patients with incomplete tetraplegia and with paraplegia, the incidence of sleep-disordered breathing is significantly higher than the general population. With the lack of correlation between symptoms and SDB, formal study would be reasonable. There is insufficient evidence in the literature on the impact of treatment on morbidity, mortality and quality of life outcomes.

Selective localization of recognition complexes for leukotriene B4 and formyl-Met-Leu-Phe within lipid raft microdomains of human polymorphonuclear neutrophils.

Neutrophilic polymorphonuclear leukocytes contain glycosphingolipid- and cholesterol-enriched lipid raft microdomains within the plasma membrane. Although there is evidence that lipid rafts function as signaling platforms for CXCR chemokine receptors, their role in recognition systems for other chemotaxins such as leukotriene B4 (LTB4) and fMLP is unknown. To address this question, human neutrophils were extracted with 1% Brij-58 and fractionated on sucrose gradients. B leukotriene receptor-1 (BLT-1), the primary LTB4 receptor, partitioned to low density fractions, co-isolating with the lipid raft marker, flotillin-1. By contrast, formyl peptide receptor (FPR), the primary fMLP receptor, partitioned to high density fractions, co-isolating with a non-raft marker, Cdc42. This pattern was preserved after the cells were stimulated with LTB4 or fMLP. Fluorescence resonance energy transfer (FRET) was performed to confirm the proximity of BLT-1 and FPR with these markers. FRET was detected between BLT1 and flotillin-1 but not Cdc42, whereas FRET was detected between FPR and Cdc42, but not flotillin-1. Pretreating neutrophils with methyl-beta-cyclodextrin, a lipid raft-disrupting agent, suppressed intracellular Ca(2+) mobilization and ERK1/2 phosphorylation in response to LTB4 but had no effect on either of these responses to fMLP. We conclude that BLT-1 is physically located within lipid raft microdomains of human neutrophils and that disrupting lipid raft integrity suppresses LTB4-induced activation. By contrast, FPR is not associated with lipid rafts, and fMLP-induced signaling does not require lipid raft integrity. These findings highlight the complexity of chemotaxin signaling pathways and offer one mechanism by which neutrophils may spatially organize chemotaxin signaling within the plasma membrane.

Cutting edge: optical microspectrophotometry supports the existence of gel phase lipid rafts at the lamellipodium of neutrophils: apparent role in calcium signaling.

Although much progress has been made in elucidating the biochemical properties of lipid rafts, there has been less success in identifying these structures within living cell membranes, which has led to some concern regarding their existence. One difficulty in analyzing lipid rafts using optical microscopy is their small size. We now test the existence of lipid rafts in polarized neutrophils, which redistribute lipid raft markers into comparatively large lamellipodia. Optical microspectrophotometry of Laurdan-labeled neutrophils revealed a large blue shift at lamellipodia relative to cell bodies. This blue shift disappeared after exposure to methyl-beta-cyclodextrin (m beta CD), which disrupts lipid rafts. The Ca(2+) channel transient receptor potential-like channel-1, a lipid raft marker, traffics to lamellipodia, but redistributes uniformly about cells after exposure to m beta CD. This is accompanied by disruption of Ca(2+) waves normally initiated at lamellipodia. Thus, m beta CD-sensitive lipid-ordered domains are present at and participate in signaling from the lamellipodia of living neutrophils.

Lipid raft compartmentalization of urokinase receptor signaling in human neutrophils.

Urokinase plasminogen activator (uPA) receptors (uPAR) can be engaged for activation signaling either by aggregation or by binding exogenous uPA. These signaling mechanisms require uPAR to associate with two distinct adhesion proteins, L-selectin and complement receptor 3 (CR3), respectively. uPAR contains a glycosylphosphatidylinositol anchor, suggesting that it is concentrated within glycosphingolipid-enriched microdomains, or "lipid rafts". This study was undertaken to determine the extent to which uPAR-mediated signaling is compartmentalized to lipid rafts. Human neutrophil uPAR was cross-linked or stimulated with uPA after pretreatment with the lipid raft-disrupting agents, methyl-beta-cyclodextrin or filipin III. Both agents suppressed increases in intracellular Ca(2+) concentrations ([Ca(2+)](i)) triggered by cross-linking, but did not affect [Ca(2+) ](i) in response to uPA. Neutrophil membranes were separated into lipid raft and non-raft fractions, revealing the presence of uPAR and L-selectin, but the virtual absence of CR3 alpha chain in lipid rafts, either constitutively or in response to uPAR aggregation. Fluorescence resonance energy transfer experiments confirmed close proximity of a lipid raft marker to both uPAR and L-selectin, but not CR3. We conclude that uPAR can engage distinct signaling pathways involving different partner proteins that are functionally and physically segregated from one another in both lipid raft and non-raft domains of the plasma membrane.