Investigating the correlation of delirium after cardiac surgery with memories and posttraumatic stress disorder consequences of intensive care unit: A prospective cohort study.

OBJECTIVES: To explore the differences in post-intensive care unit memory and posttraumatic stress disorder symptoms between patients with and without delirium, and assess the correlations between the two. DESIGN: Prospective cohort observation study. SETTING: A cardiac intensive care unit of a tertiary hospital in China. We enrolled 318 consecutive patients after cardiac surgery between December 2017 and March 2019. MAIN OUTCOME MEASURES: Delirium was assessed using the Confusion Assessment Method for the ICU from intensive care unit admission to discharge. Intensive care unit memory was assessed using the ICU-Memory Tool through face-to-face interviews one week after discharge. Posttraumatic stress disorder was measured telephonically using the Impact of Events Scale-revised questionnaire at three months post-discharge. RESULTS: Eighty patients each in the delirium and non-delirium groups were enrolled for follow-up interviews. Patients with delirium had vaguer memories of pre-intensive care unit admission and of their stay, and recollected more memories of feelings (vs. without delirium). Posttraumatic stress disorder was diagnosed in 14 patients with and in seven without delirium, with non-significant differences between groups. Delirium did not influence post-intensive care unit factual, feeling, and delusional memories, nor posttraumatic stress disorder and hyperarousal, intrusion, and avoidance. The memories of feelings were positively correlated with the last three (r = 0.285, r = 0.390 and r = 0.373, respectively). CONCLUSION: Patients with delirium had vague intensive care unit memories. Memories of feelings were positively correlated with symptoms of hyperarousal, intrusion, and avoidance. Delirium did not influence factual, feeling, or delusional memories nor posttraumatic stress disorder incidence and symptoms. IMPLICATIONS FOR CLINICAL PRACTICE: Interventions are needed to reduce the impact of vague memory in patients with post-intensive care unit delirium. Memories of feelings should be focused on because of their correlation with hyperarousal, intrusion, and avoidance. Delirium prevention and early recognition measures are suggested.

Plasma biomarkers and delirium in critically ill patients after cardiac surgery: A prospective observational cohort study.

BACKGROUND: Delirium is common in postoperative critically ill patients and may affect by intraoperative events. Biomarkers are vital indicators in the development and prediction of delirium. OBJECTIVES: This study aimed to investigate the associations between various plasma biomarkers and delirium. METHODS: We performed a prospective cohort study on cardiac surgery patients. Delirium assessment was performed twice daily using the confusion assessment method for the intensive care unit (ICU), and the Richmond Agitation Sedation Scale was used to assess the depth of sedation and agitation. Blood samples were collected on the day after ICU admission, and the concentrations of cortisol, interleukin (IL)-1ß, IL-6, tumor necrosis factor α, soluble tumor necrosis factor receptor-1 (sTNFR-1), and sTNFR-2 were measured. RESULTS: Delirium in the ICU was noted in 93 (29.2%, 95% CI 24.2-34.3) out of 318 patients (mean age 52 years, SD 12.0). The longer duration of cardiopulmonary bypass, aortic clamping and surgery, and higher plasma, erythrocytes, and platelet transfusion requirements were among the significant differences in intraoperative events between patients with and without delirium. Median levels of IL-6 (p = 0.017), TNF-α (p = 0.048), sTNFR-1 (p < 0.001), and sTNFR-2 (p = 0.001) were significantly higher in patients with delirium than in those without it. After adjusting for demographic variables and intraoperative events, only sTNFR-1 (odds ratio 6.83, 95% CI: 1.14-40.90) was associated with delirium. CONCLUSIONS: Plasma IL-6, TNF-α, sTNFR-1, and sTNFR-2 levels were higher in ICU-acquired delirium patients after cardiac surgery. sTNFR-1 was a potential indicator of the disorder.

Improving emulsifying properties of carboxylated microcrystalline cellulose by calcium bridging to hydrophobic peptides.

This study aimed to improve the emulsifying properties of microcrystalline cellulose by carboxylating (CC) and bridging to hydrophobic oat globule peptides (HP) via Ca2+ (CC-Ca-HP). FTIR and XRD spectra analysis proved the successful attachment of HP to CC through a salt bridge. The Ca2+-bridging significantly changed the particle characteristics of CC-Ca-HP, including particle size, ζ-potential, and wettability. The Ca2+-bridged composite CC-Ca-HP demonstrated remarkable emulsifying stability compared with the nonbridged blend (CC-HP). Further analysis of the steady flow characteristics and dynamic viscoelastic properties revealed a network structure formed in the CC-Ca-HP emulsion. Moreover, the CC-Ca-HP emulsion showed a marked release of free fatty acids, increased bioaccessibility of zeaxanthin in the simulated gastrointestinal digestion, and less oil oxidation under the accelerated oxidation condition, indicating that the stable structure of CC-Ca-HP imparted by Ca2+-bridging prevented the aggregation of oil droplets as collision occurred under the harsh gastric conditions.

Risk factors for inadvertent intraoperative hypothermia in patients undergoing laparoscopic surgery: A prospective cohort study.

BACKGROUND: Inadvertent intraoperative hypothermia is frequent during open surgeries; however, few studies on hypothermia during laparoscopic abdominal surgery have been reported. We aimed to investigate the incidence and risk factors for hypothermia in patients undergoing laparoscopic abdominal surgery. METHODS: This single-center prospective cohort observational study involved patients undergoing laparoscopic surgery between October 2018 and June 2019. Data on core body temperature and potential variables were collected. A multivariate logistic regression analysis was performed to identify the risk factors associated with hypothermia. A Cox regression analysis was used to verify the sensitivity of the results. RESULTS: In total, 690 patients were included in the analysis, of whom 200 (29.0%, 95% CI: 26%-32%) had a core temperature < 36°C. The core temperature decreased over time, and the incident hypothermia increased gradually. In the multivariate logistic regression analysis, age (OR = 1.017, 95% CI: 1.000-1.034, P = 0.050), BMI (OR = 0.938, 95% CI: 0.880-1.000; P = 0.049), baseline body temperature (OR = 0.025, 95% CI: 0.010-0.060; P < 0.001), volume of irrigation fluids (OR = 1.001, 95% CI: 1.000-1.001, P = 0.001), volume of urine (OR = 1.001, 95% CI: 1.000-1.003, P = 0.070), and duration of surgery (OR = 1.010, 95% CI: 1.006-1.015, P < 0.001) were significantly associated with hypothermia. In the Cox analysis, variables in the final model were age, BMI, baseline body temperature, volume of irrigation fluids, blood loss, and duration of surgery. CONCLUSIONS: Inadvertent intraoperative hypothermia is evident in patients undergoing laparoscopic surgeries. Age, BMI, baseline body temperature, volume of irrigation fluids, and duration of surgery are significantly associated with intraoperative hypothermia.

Sulfatides are endogenous ligands for the TLR4-MD-2 complex.

Many endogenous molecules, mostly proteins, purportedly activate the Toll-like receptor 4 (TLR4)-myeloid differentiation factor-2 (MD-2) complex, the innate immune receptor for lipopolysaccharide (LPS) derived from gram-negative bacteria. However, there is no structural evidence supporting direct TLR4-MD-2 activation by endogenous ligands. Sulfatides (3-O-sulfogalactosylceramides) are natural, abundant sulfated glycolipids that have variously been shown to initiate or suppress inflammatory responses. We show here that short fatty acid (FA) chain sulfatides directly activate mouse TLR4-MD-2 independent of CD14, trigger MyD88- and TRIF-dependent signaling, and stimulate tumor necrosis factor α (TNFα) and type I interferon (IFN) production in mouse macrophages. In contrast to the agonist activity toward the mouse receptor, the tested sulfatides antagonize TLR4-MD-2 activation by LPS in human macrophage-like cells. The agonistic and antagonistic activities of sulfatides require the presence of the sulfate group and are inversely related to the FA chain length. The crystal structure of mouse TLR4-MD-2 in complex with C16-sulfatide revealed that three C16-sulfatide molecules bound to the MD-2 hydrophobic pocket and induced an active dimer conformation of the receptor complex similar to that induced by LPS or lipid A. The three C16-sulfatide molecules partially mimicked the detailed interactions of lipid A to achieve receptor activation. Our results suggest that sulfatides may mediate sterile inflammation or suppress LPS-stimulated inflammation, and that additional endogenous negatively charged lipids with up to six lipid chains of limited length might also bind to TLR4-MD-2 and activate or inhibit this complex.

Adenosine monophosphate deaminase 3 null mutation causes reduction of naive T cells in mouse peripheral blood.

Adenosine monophosphate deaminase 3 (Ampd3) encodes the erythrocyte isoform of the adenosine monophosphate (AMP) deaminase gene family. Mutations in this gene have been reported in humans, leading to autosomal-recessive erythrocyte AMP deaminase deficiency. However, the mutation is considered clinically asymptomatic. Using N-ethyl-N-nitrosourea mutagenesis to find mutations that affect peripheral lymphocyte populations, we identified 5 Ampd3 mutations (Ampd3guangdong, Ampd3carson, Ampd3penasco, Ampd3taos, and Ampd3commanche) that strongly correlated with a reduction in naive CD4+ T and naive CD8+ T-cell populations. Causation was confirmed by targeted ablation of Ampd3. Knockout mice had reduced frequencies of CD62LhiCD44lo CD4+ naive and CD8+ naive T cells. Interestingly, these phenotypes were restricted to T cells circulating in peripheral blood and were not seen in T cells from secondary lymphoid organs (lymph nodes and spleen). We found that reduction of naive T cells in the peripheral blood of Ampd3-/- mice was caused by T-cell-extrinsic factor(s), which we hypothesize to be elevated levels of adenosine triphosphate released by Ampd3-deficient erythrocytes. These findings provide an example in which disruption of an erythrocyte-specific protein can affect the physiological status of lymphocytes in peripheral blood.

Genetic and structural studies of RABL3 reveal an essential role in lymphoid development and function.

The small GTPase RABL3 is an oncogene of unknown physiological function. Homozygous knockout alleles of mouse Rabl3 were embryonic lethal, but a viable hypomorphic allele (xiamen [xm]) causing in-frame deletion of four amino acids from the interswitch region resulted in profound defects in lymphopoiesis. Impaired lymphoid progenitor development led to deficiencies of B cells, T cells, and natural killer (NK) cells in Rabl3xm/xm mice. T cells and NK cells exhibited impaired cytolytic activity, and mice infected with mouse cytomegalovirus (MCMV) displayed elevated titers in the spleen. Myeloid cells were normal in number and function. Biophysical and crystallographic studies demonstrated that RABL3 formed a homodimer in solution via interactions between the effector binding surfaces on each subunit; monomers adopted a typical small G protein fold. RABL3xm displayed a large compensatory alteration in switch I, which adopted a ß-strand configuration normally provided by the deleted interswitch residues, thereby permitting homodimer formation. Dysregulated effector binding due to conformational changes in the switch I-interswitch-switch II module likely underlies the xm phenotype. One such effector may be GPR89, putatively an ion channel or G protein-coupled receptor (GPCR). RABL3, but not RABL3xm, strongly associated with and stabilized GPR89, and an N-ethyl-N-nitrosourea (ENU)-induced mutation (explorer) in Gpr89 phenocopied Rabl3xm.

Mutual inhibition between Prkd2 and Bcl6 controls T follicular helper cell differentiation.

T follicular helper cells (TFH) participate in germinal center (GC) development and are necessary for B cell production of high-affinity, isotype-switched antibodies. In a forward genetic screen, we identified a missense mutation in Prkd2, encoding the serine/threonine kinase protein kinase D2, which caused elevated titers of immunoglobulin E (IgE) in the serum. Subsequent analysis of serum antibodies in mice with a targeted null mutation of Prkd2 demonstrated polyclonal hypergammaglobulinemia of IgE, IgG1, and IgA isotypes, which was exacerbated by the T cell-dependent humoral response to immunization. GC formation and GC B cells were increased in Prkd2-/- spleens. These effects were the result of excessive cell-autonomous TFH development caused by unrestricted Bcl6 nuclear translocation in Prkd2-/- CD4+ T cells. Prkd2 directly binds to Bcl6, and Prkd2-dependent phosphorylation of Bcl6 is necessary to constrain Bcl6 to the cytoplasm, thereby limiting TFH development. In response to immunization, Bcl6 repressed Prkd2 expression in CD4+ T cells, thereby committing them to TFH development. Thus, Prkd2 and Bcl6 form a mutually inhibitory positive feedback loop that controls the stable transition from naïve CD4+ T cells to TFH during the adaptive immune response.

Essential cell-extrinsic requirement for PDIA6 in lymphoid and myeloid development.

In a forward genetic screen of N-ethyl-N-nitrosourea (ENU)-induced mutant mice for aberrant immune function, we identified mice with a syndromic disorder marked by growth retardation, diabetes, premature death, and severe lymphoid and myeloid hypoplasia together with diminished T cell-independent (TI) antibody responses. The causative mutation was in Pdia6, an essential gene encoding protein disulfide isomerase A6 (PDIA6), an oxidoreductase that functions in nascent protein folding in the endoplasmic reticulum. The immune deficiency caused by the Pdia6 mutation was, with the exception of a residual T cell developmental defect, completely rescued in irradiated wild-type recipients of PDIA6-deficient bone marrow cells, both in the absence or presence of competition. The viable hypomorphic allele uncovered in these studies reveals an essential role for PDIA6 in hematopoiesis, but one extrinsic to cells of the hematopoietic lineage. We show evidence that this role is in the proper folding of Wnt3a, BAFF, IL-7, and perhaps other factors produced by the extra-hematopoietic compartment that contribute to the development and lineage commitment of hematopoietic cells.

Short peptides secreted by Bacillus subtilis inhibit the growth of mold on fresh-cut pumpkin (Cucurbita pepo).

BACKGROUND: This study investigates the efficacy of short peptides secreted by Bacillus subtilis for fungal inhibition in fresh-cut pumpkin and for maintaining its shelf life. RESULTS: Low-molecular-weight filtrate (LC < 1000 Da) of B. subtilis culture (BC) significantly lowered the total number of molds on fresh-cut pumpkin compared with the untreated control and a BC group after storage. Low-molecular-weight filtrate prevented the deterioration of sensory quality in a pumpkin incision, and reduced pectinase activity. It also inhibited the growth of Phytophthora capsici and Penicillium chrysogenum, and the activity of ß-1,3-glucan synthase (GS) secreted by both molds. Fifty-seven GS-inhibiting peptides were screened from 95 LC peptides with two to five amino acid residues. The two most potent peptides, AWYW and HWWY, had strongly suppressive effects on the growth of P. capsici and P. chrysogenum. CONCLUSION: Our study demonstrated that short peptides present in B. subtilis culture can play an important role in the maintenance of fresh-cut pumpkin by suppressing fungal growth. © 2019 Society of Chemical Industry.

[Risk factors for intensive care unit delirium after cardiac operation].

OBJECTIVE: To analyze the risk factors of delirium in patients in cardiac surgery intensive care unit (CSICU). METHODS: A prospective observational study was performed. Patients admitted to CSICU of Fujian Medical University Union Hospital from March to August in 2017 were enrolled. The combination of the Richmond agitation sedation scale (RASS) and the ICU-confusion assessment method (CAM-ICU) were used to evaluate delirium. The patient was assessed on the second day after CSICU admission, twice a day, the evaluation was stopped, and the follow-up observation was terminated after the patient was discharged from CSICU. The patients were divided into two groups according to whether delirium occurred in CSICU. The general and clinical treatment data (including condition, operation, anesthesia and CSICU treatment) of the two groups were compared. The related factors of delirium were identified by univariate analysis and multifactor Logistic regression analysis. RESULTS: A total of 318 cases were included in this study. Among them, 93 cases had delirium and the incidence of delirium was 29.2%. It was shown by univariate analysis that age, history of hypertension, type of surgery, surgical procedure, American Society of Anesthesiologists (ASA) anesthesia classification, usage of propofol, plasma transfusion, red blood cells, platelet transfusion, blood loss, operative time, cardiopulmonary bypass (CPB) time, myocardial block time, acute physiology and chronic health evaluation II (APACHE II), duration of mechanical ventilation, the length of intensive care unit (ICU) stay, postoperative usage of diazepam, midazolam, fentanyl, morphine, chlorpromazine, etc. which were related to delirium, and occupation (on-the-job or self-employed), medical insurance (city or provincial medical insurance), education (primary to junior high school, high school or above) could reduce the risk of delirium. Colinearity diagnosis was performed on variables with statistically significant differences, and variables with variance expansion factor (VIF) < 3 were included in multivariate Logistic regression analysis. The results showed that age, education level, type of surgery, ASA classification, CPB time, APACHE II, ICU mechanical ventilation time, and post operation usage of midazolam were independently related to delirium [age: odds ratio (OR) = 1.625, 95% confidence interval (95%CI) = 1.303-2.026; education level: OR = 0.293, 95%CI = 0.171-0.504; type of surgery: OR = 2.194, 95%CI = 1.052-4.576; ASA classification: OR = 1.916, 95%CI = 1.032-3.559; CPB time: OR = 2.125, 95%CI = 1.105-4.088; APACHE II: OR = 2.091, 95%CI = 1.005-4.349; ICU mechanical ventilation time: OR = 1.943, 95%CI = 1.269-2.975; midazolam: OR = 2.653, 95%CI = 1.328-5.299; all P < 0.05], among which, high education level has a good protective effect on delirium. CONCLUSIONS: Age, type of surgery, ASA classification, CPB time, APACHE II, ICU mechanical ventilation time, post operation usage of midazolam were independent risk factors for delirium, and high education level had a good protective effect. Among them, the educational level, CPB time, duration of mechanical ventilation, and midazolam are intervention factors. In clinical treatment, not only the risk factors should be identified, but also intervention should be taken to prevent the occurrence of delirium.

Structural Basis of TLR2/TLR1 Activation by the Synthetic Agonist Diprovocim.

Diprovocim is a recently discovered exceptionally potent, synthetic small molecule agonist of TLR2/TLR1 and has shown significant adjuvant activity in anticancer vaccination against murine melanoma. Since Diprovocim bears no structural similarity to the canonical lipopeptide ligands of TLR2/TLR1, we investigated how Diprovocim interacts with TLR2/TLR1 through in vitro biophysical, structural, and computational approaches. We found that Diprovocim induced the formation of TLR2/TLR1 heterodimers as well as TLR2 homodimers in vitro. We determined the crystal structure of Diprovocim in a complex with a TLR2 ectodomain, which revealed, unexpectedly, two Diprovocim molecules bound to the ligand binding pocket formed between two TLR2 ectodomains. Extensive hydrophobic interactions and a hydrogen-bonding network between the protein and Diprovocim molecules are observed within the defined ligand binding pocket and likely underlie the high potency of Diprovocim. Our work shed first light into the activation mechanism of TLR2/TLR1 by a noncanonical agonist. The structural information obtained here may be exploited to manipulate TLR2/TLR1-dependent signaling.

Diprovocims: A New and Exceptionally Potent Class of Toll-like Receptor Agonists.

A screen conducted with nearly 100000 compounds and a surrogate functional assay for stimulation of an immune response that measured the release of TNF-α from treated human THP-1 myeloid cells differentiated along the macrophage line led to the discovery of the diprovocims. Unique to these efforts and of special interest, the screening leads for this new class of activators of an immune response came from a compound library designed to promote cell-surface receptor dimerization. Subsequent comprehensive structure-activity relationship studies improved the potency 800-fold over that of the screening leads, providing diprovocim-1 and diprovocim-2. The diprovocims act by inducing cell-surface toll-like receptor (TLR)-2 dimerization and activation with TLR1 (TLR1/TLR2 agonist), bear no structural similarity to any known natural or synthetic TLR agonist, and are easy to prepare and synthetically modify, and selected members are active in both human and murine systems. The most potent diprovocim (3, diprovocim-1) elicits full agonist activity at extraordinarily low concentrations (EC50 = 110 pM) in human THP-1 cells, being more potent than the naturally derived TLR1/TLR2 agonist Pam3CSK4 or any other known small molecule TLR agonist.

Adjuvant effect of the novel TLR1/TLR2 agonist Diprovocim synergizes with anti-PD-L1 to eliminate melanoma in mice.

Successful cancer immunotherapy entails activation of innate immune receptors to promote dendritic cell (DC) maturation, antigen presentation, up-regulation of costimulatory molecules, and cytokine secretion, leading to activation of tumor antigen-specific cytotoxic T lymphocytes (CTLs). Here we screened a synthetic library of 100,000 compounds for innate immune activators using TNF production by THP-1 cells as a readout. We identified and optimized a potent human and mouse Toll-like receptor (TLR)1/TLR2 agonist, Diprovocim, which exhibited an EC50 of 110 pM in human THP-1 cells and 1.3 nM in primary mouse peritoneal macrophages. In mice, Diprovocim-adjuvanted ovalbumin immunization promoted antigen-specific humoral and CTL responses and synergized with anti-PD-L1 treatment to inhibit tumor growth, generating long-term antitumor memory, curing or prolonging survival of mice engrafted with the murine melanoma B16-OVA. Diprovocim induced greater frequencies of tumor-infiltrating leukocytes than alum, of which CD8 T cells were necessary for the antitumor effect of immunization plus anti-PD-L1 treatment.

NLRP3 activation and mitosis are mutually exclusive events coordinated by NEK7, a new inflammasome component.

The NLRP3 inflammasome responds to microbes and danger signals by processing and activating proinflammatory cytokines, including interleukin 1ß (IL-1ß) and IL-18. We found here that activation of the NLRP3 inflammasome was restricted to interphase of the cell cycle by NEK7, a serine-threonine kinase previously linked to mitosis. Activation of the NLRP3 inflammasome required NEK7, which bound to the leucine-rich repeat domain of NLRP3 in a kinase-independent manner downstream of the induction of mitochondrial reactive oxygen species (ROS). This interaction was necessary for the formation of a complex containing NLRP3 and the adaptor ASC, oligomerization of ASC and activation of caspase-1. NEK7 promoted the NLRP3-dependent cellular inflammatory response to intraperitoneal challenge with monosodium urate and the development of experimental autoimmune encephalitis in mice. Our findings suggest that NEK7 serves as a cellular switch that enforces mutual exclusivity of the inflammasome response and cell division.

Mixed lineage kinase domain-like protein MLKL causes necrotic membrane disruption upon phosphorylation by RIP3.

Programmed necrotic cell death induced by the tumor necrosis factor alpha (TNF-α) family of cytokines is dependent on a kinase cascade consisting of receptor-interacting kinases RIP1 and RIP3. How these kinase activities cause cells to die by necrosis is not known. The mixed lineage kinase domain-like protein MLKL is a functional RIP3 substrate that binds to RIP3 through its kinase-like domain but lacks kinase activity of its own. RIP3 phosphorylates MLKL at the T357 and S358 sites. Reported here is the development of a monoclonal antibody that specifically recognizes phosphorylated MLKL in cells dying of this pathway and in human liver biopsy samples from patients suffering from drug-induced liver injury. The phosphorylated MLKL forms an oligomer that binds to phosphatidylinositol lipids and cardiolipin. This property allows MLKL to move from the cytosol to the plasma and intracellular membranes, where it directly disrupts membrane integrity, resulting in necrotic death.

Regulation of lung surfactant secretion by microRNA-150.

P2X7 receptor (P2X7R) is a purinergic ion-channel receptor. We have previously shown that the activation of P2X7R in alveolar type I cells stimulates surfactant secretion in alveolar type II cells. In this study, we determined whether miR-150 regulates P2X7R-mediated surfactant secretion. The miR-150 expression level in alveolar type II cells was much higher than alveolar type I cells, which was inversely correlated with the P2X7R protein level. An adenovirus expressing miR-150 significantly reduced the P2X7R protein expression in E10 cells, an alveolar type I cell line. Furthermore, pre-treatment of E10 cells with the adenovirus reduced the surfactant secretion induced by E10 cell conditioned medium. Our study demonstrates that miR-150 regulates surfactant secretion through P2X7R.

Purinergic P2X7 receptor regulates lung surfactant secretion in a paracrine manner.

Alveolar epithelium is composed of alveolar epithelial cells of type I (AEC I) and type II (AEC II). AEC II secrete lung surfactant by means of exocytosis. P2X(7) receptor (P2X(7)R), a P2 purinergic receptor, has been implicated in the regulation of synaptic transmission and inflammation. Here, we report that P2X(7)R, which is expressed in AEC I but not AEC II, is a novel mediator for the paracrine regulation of surfactant secretion in AEC II. In primary co-cultures of AEC I and AEC II benzoyl ATP (BzATP; an agonist of P2X(7)R) increased surfactant secretion, which was blocked by the P2X(7)R antagonist Brilliant Blue G. This effect was observed in AEC II co-cultured with human embryonic kidney HEK-293 cells stably expressing rat P2X(7)R, but not when co-cultured with AEC I in which P2X(7)R was knocked down or in co-cultures of AEC I and AEC II isolated from P2X(7)R(-/-) mice. BzATP-mediated secretion involved P2Y(2) receptor signaling because it was reduced by the addition of the ATP scavengers apyrase and adenosine deaminase and the P2Y(2) receptor antagonist suramin. However, the stimulation with BzATP might also release other substances that potentially increase surfactant secretion as a greater stimulation of secretion was observed in AEC II incubated with BzATP when co-cultured with E10 or HEK-293-P2X(7)R cells than with ATP alone. P2X(7)R(-/-) mice failed to increase surfactant secretion in response to hyperventilation, pointing to the physiological relevance of P2X(7)R in maintaining surfactant homeostasis in the lung. These results suggest that the activation of P2X(7)R increases surfactant secretion by releasing ATP from AEC I and subsequently stimulating P2Y(2) receptors in AEC II.

UTP regulation of ion transport in alveolar epithelial cells involves distinct mechanisms.

UTP is known to regulate alveolar fluid clearance. However, the relative contribution of alveolar type I cells and type II cells to this process is unknown. In this study, we investigated the effects of UTP on ion transport in type I-like cell (AEC I) and type II-like cell (AEC II) monolayers. Luminal treatment of cell monolayers with UTP increased short-circuit current (I(sc)) of AEC II but decreased I(sc) of AEC I. The Cl(-) channel blockers NPPB and DIDS inhibited the UTP-induced changes in I(sc) (DeltaIsc) in both types of cells. Amiloride, an inhibitor of epithelial Na(+) channels (ENaC), abolished the UTP-induced DeltaI(sc) in AEC I, but not in AEC II. The general blocker of K(+) channels, BaCl(2), eliminated the UTP-induced DeltaI(sc) in AEC II, but not in AEC I. The intermediate conductance (IK(Ca)) blocker, clofilium, also blocked the UTP effect in AEC II. The signal transduction pathways mediated by UTP were the same in AEC I and AEC II. Furthermore, UTP increased Cl(-) secretion in AEC II and Cl(-) absorption in AEC I. Our results suggest that UTP induces opposite changes in I(sc) in AEC I and AEC II, likely due to the reversed Cl(-) flux and different contributions of ENaC and IK(Ca). Our results further imply a new concept that type II cells contribute to UTP-induced fluid secretion and type I cells contribute to UTP-induced fluid absorption in alveoli.