The Ca2+ signaling and contractility of airway smooth muscle cells (SMCs) were investigated with confocal microscopy in murine lung slices (∼75-μm thick) that maintained the in situ organization of the airways and the contractility of the SMCs for at least 5 d. 10–500 nM acetylcholine (ACH) induced a contraction of the airway lumen and a transient increase in [Ca2+]i in individual SMCs that subsequently declined to initiate multiple intracellular Ca2+ oscillations. These Ca2+ oscillations spread as Ca2+ waves through the SMCs at ∼48 μm/s. The magnitude of the airway contraction, the initial Ca2+ transient, and the frequency of the subsequent Ca2+ oscillations were all concentration-dependent. In a Ca2+-free solution, ACH induced a similar Ca2+ response, except that the Ca2+ oscillations ceased after 1–1.5 min. Incubation with thapsigargin, xestospongin, or ryanodine inhibited the ACH-induced Ca2+ signaling. A comparison of airway contraction with the ACH-induced Ca2+ response of the SMCs revealed that the onset of airway contraction correlated with the initial Ca2+ transient, and that sustained airway contraction correlated with the occurrence of the Ca2+ oscillations. Buffering intracellular Ca2+ with BAPTA prohibited Ca2+ signaling and airway contraction, indicating a Ca2+-dependent pathway. Cessation of the Ca2+ oscillations, induced by ACH-esterase, halothane, or the absence of extracellular Ca2+ resulted in a relaxation of the airway. The concentration dependence of the airway contraction matched the concentration dependence of the increased frequency of the Ca2+ oscillations. These results indicate that Ca2+ oscillations, induced by ACH in murine bronchial SMCs, are generated by Ca2+ release from the SR involving IP3- and ryanodine receptors, and are required to maintain airway contraction.

Our data demonstrate that dialysate-related factors such as low bacterial contamination can induce the activation of monocytes, resulting in elevated serum levels of IL-6. Dialysate-related cytokine induction might diminish erythropoiesis. The use of pyrogen free ultrapure dialysate resulted in a better response to rHuEpo. Not only would it save money, but it would also help to maintain an optimal haemoglobin level without further increase in rHuEpo dosage.

2ϩ oscillations that were accompanied by airway contraction. After ϳ1 min, the Ca 2ϩ oscillations subsided and the airway relaxed. By contrast, Ն0.5 M adenosine 5Ј-O-(3-thiotriphosphate) (nonhydrolyzable) induced Ca 2ϩ oscillations in the SMCs and an associated airway contraction that persisted for Ͼ2 min. Adenosine 5Ј-O-(3-thiotriphosphate)-induced Ca 2ϩ oscillations occurred in the absence of external Ca 2ϩ but were abolished by the phospholipase C inhibitor U-73122 and the inositol 1,4,5-trisphosphate receptor inhibitor xestospongin. Adenosine, AMP, and ␣,␤-methylene ATP had no effect on airway caliber, and the magnitude of the contractile response induced by a variety of nucleotides could be ranked in the following order: ATP ϭ UTP Ͼ ADP. These results suggest that the SMC response to ATP is impaired by ATP hydrolysis and mediated via P2Y 2 or P2Y 4 receptors, activating phospholipase C to release Ca 2ϩ via the inositol 1,4,5-trisphosphate receptor. We conclude that ATP can serve as a spasmogen of airway SMCs and that Ca 2ϩ oscillations in SMCs are required to sustain airway contraction. calcium signaling; confocal microscopy; ATP hydrolysis ASTHMA IS AN AIRWAY DISEASE characterized by a hyperreactive response of airway smooth muscle cells (SMCs) to spasmogens such as histamine, leukotrienes, and perhaps serotonin (2, 6, 25). A major source of histamine and other agonists is an exocytotic release from inflammatory cells (i.e., mast cells and basophils) within the submucosal tissue, in close proximity to the SMCs. However, mast cell degranulation can simultaneously release another potential spasmogen, ATP, which is packaged in the same exocytotic vesicles (23). In addition, airway epithelial cells have also been identified as a source of extracellular ATP. Although the active release mechanisms of ATP from epithelial cells are not understood (5,7,11,36,37), epithelial trauma associated with asthma (12, 18) can result in damaged cells that passively release ATP.In many other tissues, ATP has been found to stimulate P2X and P2Y receptors, both of which have been identified in lung tissue (19,29,34,35 ] i ) of SMCs usually result in the phosphorylation of myosin light chain and the stimulation of contraction (15, 16), ATP is likely to be a potent spasmogen of airway SMCs.Surprisingly, the effects of ATP on the [Ca 2ϩ ] i of airway SMCs or on airway contraction have not been extensively studied, and the few reports that exist are inconsistent. For example, ATP was found to increase in vivo airway resistance in rats (9) or induce contraction in isolated guinea pig tracheae (8). On the other hand, ATP was reported to relax rabbit tracheal smooth muscle strips (1) and isolated mouse trachea (10, 17) that had been previously contracted with acetylcholine (ACh). These tissue preparations were derived from the large airways and consisted of tracheal rings or strips. As a result, these studies were not able to focus on the responses of individual SMCs. In addition, it is not clear whether results obtained from large air...

Objectives Residual renal function (RRF) is of paramount importance to dialysis adequacy, morbidity, and mortality, particularly for long-term continuous ambulatory peritoneal dialysis (CAPD) patients. Residual renal function seems to be better preserved in patients on CAPD than in hemodialysis (HD) patients. We analyzed RRF in 45 patients with end-stage renal disease (ESRD), commencing either CAPD or HD, to prospectively define the time course of the decline in RRF, and to evaluate dialysis-technique–related factors such as cardiovascular stability and bioincompatibility. Study Design Single-center prospective investigation in parallel design with matched pairs. Materials Fifteen patients starting CAPD and 15 matched pairs of patients commencing HD were matched according to cause of renal failure and RRF. Hemodialysis patients were assigned to two dialyzer membranes differing markedly in their potential to activate complement and cells (bioincompatibility). Fifteen patients were treated exclusively with the cuprophane membrane (bioincompatible) and the other 15 patients received HD with the high-flux polysulfone membrane (biocompatible). Measurements Residual renal function was determined at initiation of dialytic therapy and after 6, 12, and 24 months. Dry weight (by chest x ray and diameter of the vena cava) was closely recorded throughout the study, and the number of hypotensive episodes counted. Results Residual renal function declined in both CAPD and HD patients, although this decline was faster in HD patients (2.8 mL/minute after 6 months and 3.7 mL/min after 12 months) than in CAPD patients (0.6 mL/min and 1.4 mL/ min after 6 and 12 months respectively). It declined faster in patients with bioincompatible than with biocompatible HD membranes (3.6 mL/min vs 1.9 mL/min after 6 months). Eleven percent of the HD sessions were complicated by clinically relevant blood pressure reductions, but there were no differences between the two dialyzer membrane groups. None of the CAPD patients had documented hypotensive episodes. None of the study patients suffered severe illness or received nephrotoxic antibiotics or radiocontrast media. Conclusions The better preservation of RRF in stable CAPD patients corresponded with greater cardiovascular stability compared to HD patients, independently of the membrane used. Furthermore, there was a significantly higher preservation of RRF in HD patients on polysulfone versus cuprophane membranes, indicating an additional effect of biocompatibility, such as less generation of nephrotoxic substances by the membrane. Thus, starting ESRD patients on HD prior to elective CAPD should be avoided for better preservation of RRF.

Proton transfer reaction mass spectrometry (PTR-MS) has been used to analyze the volatile organic compounds (VOCs) emitted by in-vitro cultured human cells. For this purpose, two pairs of cancerous and non-cancerous human cell lines were selected:1. lung epithelium cells A-549 and retinal pigment epithelium cells hTERT-RPE1, cultured in different growth media; and 2. squamous lung carcinoma cells EPLC and immortalized human bronchial epithelial cells BEAS2B, cultured in identical growth medium. The VOCs in the headspace of the cell cultures were sampled: 1. online by drawing off the gas directly from the culture flask; and 2. by accumulation of the VOCs in PTFE bags connected to the flask for at least 12 h. The pure media were analyzed in the same way as the corresponding cells in order to provide a reference. Direct comparison of headspace VOCs from flasks with cells plus medium and from flasks with pure medium enabled the characterization of cell-line-specific production or consumption of VOCs. Among all identified VOCs in this respect, the most outstanding compound was m/z = 45 (acetaldehyde) revealing significant consumption by the cancerous cell lines but not by the non-cancerous cells. By applying multivariate statistical analysis using 42 selected marker VOCs, it was possible to clearly separate the cancerous and non-cancerous cell lines from each other.