Charles W. Emala

Differential protective effects of Volatile anesthetics against renal ischemia-reperfusion injury in vivo

Background:Volatile anesthetics protect against cardiac ischemia–reperfusion injury via adenosine triphosphate–dependent potassium channel activation. The authors questioned whether volatile anesthetics can also protect against renal ischemia–reperfusion injury and, if so, whether cellular adenosine triphosphate–dependent potassium channels, antiinflammatory effects of volatile anesthetics, or both are involved. Methods:Rats were anesthetized with equipotent doses of volatile anesthetics (desflurane, halothane, isoflurane, or sevoflurane) or injectable anesthetics (pentobarbital or ketamine) and subjected to 45 min of renal ischemia and 3 h of reperfusion during anesthesia. Results:Rats treated with volatile anesthetics had lower plasma creatinine and reduced renal necrosis 24–72 h after injury compared with rats anesthetized with pentobarbital or ketamine. Twenty-four hours after injury, sevoflurane-, isoflurane-, or halothane-treated rats had creatinine (± SD) of 2.3 ± 0.7 mg/dl (n = 12), 1.8 ± 0.5 mg/dl (n = 6), and 2.4 ± 1.2 mg/dl (n = 6), respectively, compared with rats treated with pentobarbital (5.8 ± 1.2 mg/dl, n = 9) or ketamine (4.6 ± 1.2 mg/dl, n = 8). Among the volatile anesthetics, desflurane demonstrated the least reduction in plasma creatinine after 24 h (4.1 ± 0.8 mg/dl, n = 12). Renal cortices from volatile anesthetic–treated rats demonstrated reduced expression of intercellular adhesion molecule 1 protein and messenger RNA as well as messenger RNAs encoding proinflammatory cytokines and chemokines. Volatile anesthetic treatment reduced renal cortex myeloperoxidase activity and reduced nuclear translocation of proinflammatory nuclear factor &kgr;B. Adenosine triphosphate–dependent potassium channels are not involved in sevoflurane-mediated renal protection because glibenclamide did not block renal protection (creatinine: 2.4 ± 0.4 mg/dl, n = 3). Conclusion:Some volatile anesthetics confer profound protection against renal ischemia–reperfusion injury compared with pentobarbital or ketamine anesthesia by attenuating inflammation. These findings may have significant clinical implications for anesthesiologists regarding the choice of volatile anesthetic agents in patients subjected to perioperative renal ischemia.

Anaphylaxis and anesthesia: controversies and new insights.

PERIOPERATIVE anaphylaxis may be a life-threatening clinical condition and is typically a result of drugs or substances used for anesthesia or surgery. After anaphylaxis, allergologic assessment is essential to identify the offending agent and prevent recurrences, because no preemptive therapeutic strategies exist. This review seeks to (1) identify the clinical diagnostic pathway necessary to distinguish anaphylaxis from confounding clinical diagnoses, (2) discuss the more common allergens that cause anaphylaxis during anesthesia, (3) discuss a rational approach to the identification of the offending allergen through blood and skin testing that allows for the safe future clinical management of patients experiencing perioperative anaphylaxis, and (4) discuss new therapeutic perspectives for the management of patients whose hemodynamic collapse is unresponsive to catecholamines, the initial recommended pharmacologic intervention.

A1 Adenosine Receptor Activation Inhibits Inflammation, Necrosis, and Apoptosis after Renal Ischemia-Reperfusion Injury in Mice

It was previously demonstrated that preischemic A(1) adenosine receptor (AR) activation protects renal function after ischemia-reperfusion (IR) injury in rats. The role of the A(1) AR in modulating inflammation, necrosis, and apoptosis in the kidney after IR renal injury was further characterized. C57BL/6 mice were subjected to 30 min of renal ischemia, with or without pretreatment with 1,3-dipropyl-8-cyclopentylxanthine or 2- chlorocyclopentyladenosine (selective A(1) AR antagonist and agonist, respectively). Plasma creatinine levels and renal inflammation, necrosis, and apoptosis were compared 24 h after renal injury. C57BL/6 mice that had been pretreated with the A(1) AR agonist demonstrated significantly improved renal function and reduced expression of inflammatory markers, necrosis, and apoptosis 24 h after IR injury. In contrast, C57BL/6 mice that had been pretreated with the A(1) AR antagonist demonstrated significantly worsened renal function and increased expression of inflammatory markers, necrosis, and apoptosis. In conclusion, it was demonstrated that endogenous and exogenous preischemic activation of the A(1) AR protects against IR injury in vivo, through mechanisms that reduce inflammation, necrosis, and apoptosis.

Reverse remodeling and enhanced adrenergic reserve from passive external support in experimental dilated heart failure

OBJECTIVES We sought to test the efficacy of a passive elastic containment device to reverse chronic chamber remodeling and adrenergic down-regulation in the failing heart, yet still maintaining preload reserve. BACKGROUND Progressive cardiac remodeling due to heart failure is thought to exacerbate underlying myocardial dysfunction. In a pressure-volume analysis, we tested the impact of limiting progressive cardiac dilation by an externally applied passive containment device on both basal and adrenergic-stimulated function in failing canine hearts. METHODS Ischemic dilated cardiomyopathy was induced by repeated intracoronary microembolizations in six dogs. The animals were studied before and three to six months after surgical implantation of a thin polyester mesh (cardiac support device [CSD]) that surrounded both cardiac ventricles. Pressure-volume relations were measured by a conductance micromanometer catheter. RESULTS Long-term use of the CSD lowered end-diastolic and end-systolic volumes by -19 +/- 4% and -22 +/- 8%, respectively (both p < 0.0001) and shifted the end-systolic pressure-volume relation to the left (p < 0.01), compatible with reverse remodeling. End-diastolic pressure and chamber diastolic stiffness did not significantly change. The systolic response to dobutamine markedly improved after CSD implantation (55 +/- 8% rise in ejection fraction after CSD vs. -10 +/- 8% before CSD, p < 0.05), in conjunction with a heightened adenylyl cyclase response to isoproterenol. There was no change in the density or affinity of beta-adrenergic receptors. Diastolic compliance was not adversely affected, and preload-recruitable function was preserved with the CSD, consistent with a lack of constriction. CONCLUSIONS Reverse remodeling with reduced systolic wall stress and improved adrenergic signaling can be achieved by passive external support that does not generate diastolic constriction. This approach may prove useful in the treatment of chronic heart failure.

Actin reorganization in airway smooth muscle cells involves Gq and Gi-2 activation of Rho

Extracellular stimuli induce cytoskeleton reorganization (stress-fiber formation) in cells and Ca2+ sensitization in intact smooth muscle preparations by activating signaling pathways that involve Rho proteins, a subfamily of the Ras superfamily of monomeric G proteins. In airway smooth muscle, the agonists responsible for cytoskeletal reorganization via actin polymerization are poorly understood. Carbachol-, lysophosphatidic acid (LPA)-, and endothelin-1-induced increases in filamentous actin staining are indicative of actin reorganization (filamentous-to-globular actin ratios of 2.4 +/- 0.3 in control cells, 6.7 +/- 0.8 with carbachol, 7.2 +/- 0.8 with LPA, and 7.4 +/- 0.9 with endothelin-1; P < 0.001; n = 14 experiments). Although the effect of all agonists was blocked by C3 exoenzyme (inactivator of Rho), only carbachol was blocked by pertussis toxin. Although carbachol-induced actin reorganization was blocked in cells pretreated with antisense oligonucleotides directed against Galphai-2 alone, LPA- and endothelin-1-induced actin reorganization were only blocked when both Galphai-2 and G(q)alpha were depleted. These data indicate that in human airway smooth muscle cells, carbachol induces actin reorganization via a Galphai-2 pathway, whereas LPA or endothelin-1 induce actin reorganization via either a Galphai-2 or a Gqalpha pathway.

Abnormal pattern of Tie-2 and vascular endothelial growth factor receptor expression in human cerebral arteriovenous malformations

OBJECTIVEHuman cerebral arteriovenous malformations (AVMs) are speculated to result from abnormal angiogenesis. Vascular endothelial growth factor receptors (VEGF-Rs) and Tie-2 play critical roles in vasculogenesis and angiogenesis. We hypothesized that the abnormal vascular phenotype of AVMs may be associated with abnormal expression of VEGF-Rs and Tie-2. METHODSWe measured the expression of Tie-2, VEGF-R1, and VEGF-R2 in AVMs and normal brain tissue, using immunoblotting. To assess active vascular remodeling, we also measured endothelial nitric oxide synthase expression. CD31 expression was used to control for endothelial cell mass for Tie-2, VEGF-Rs, and endothelial nitric oxide synthase. Immunoblotting data were presented as relative expression, using normal brain tissue values as 100%. RESULTSCD31 was expressed to similar degrees in AVMs and normal brain tissue (99 ± 29% versus 100 ± 20%, mean ± standard error, P = 0.98). Tie-2 expression was markedly decreased in all AVMs, compared with normal brain tissue (16 ± 9% versus 100 ± 37%, P = 0.04). VEGF-R1 expression was decreased in four of five AVMs, but the difference between the mean values was not significant (35 ± 8% versus 100 ± 42%, P = 0.14). VEGF-R2 expression was decreased in all AVMs, compared with normal brain tissue (28 ± 6% versus 100 ± 29%, P = 0.03). There was no difference in endothelial nitric oxide synthase expression between AVMs and normal brain tissue (106 ± 42% versus 100 ± 25%, P = 0.91). CONCLUSIONAVM vessels exhibited abnormal expression of Tie-2 and VEGF-Rs, both of which may contribute to the pathogenesis of AVMs.

Propofol Directly Increases Tau Phosphorylation

In Alzheimers disease (AD) and other tauopathies, the microtubule-associated protein tau can undergo aberrant hyperphosphorylation potentially leading to the development of neurofibrillary pathology. Anesthetics have been previously shown to induce tau hyperphosphorylation through a mechanism involving hypothermia-induced inhibition of protein phosphatase 2A (PP2A) activity. However, the effects of propofol, a common clinically used intravenous anesthetic, on tau phosphorylation under normothermic conditions are unknown. We investigated the effects of a general anesthetic dose of propofol on levels of phosphorylated tau in the mouse hippocampus and cortex under normothermic conditions. Thirty min following the administration of propofol 250 mg/kg i.p., significant increases in tau phosphorylation were observed at the AT8, CP13, and PHF-1 phosphoepitopes in the hippocampus, as well as at AT8, PHF-1, MC6, pS262, and pS422 epitopes in the cortex. However, we did not detect somatodendritic relocalization of tau. In both brain regions, tau hyperphosphorylation persisted at the AT8 epitope 2 h following propofol, although the sedative effects of the drug were no longer evident at this time point. By 6 h following propofol, levels of phosphorylated tau at AT8 returned to control levels. An initial decrease in the activity and expression of PP2A were observed, suggesting that PP2A inhibition is at least partly responsible for the hyperphosphorylation of tau at multiple sites following 30 min of propofol exposure. We also examined tau phosphorylation in SH-SY5Y cells transfected to overexpress human tau. A 1 h exposure to a clinically relevant concentration of propofol in vitro was also associated with tau hyperphosphorylation. These findings suggest that propofol increases tau phosphorylation both in vivo and in vitro under normothermic conditions, and further studies are warranted to determine the impact of this anesthetic on the acceleration of neurofibrillary pathology.

Carbachol-induced actin reorganization involves Gi activation of Rho in human airway smooth muscle cells

To determine whether M2 muscarinic receptors are linked to the monomeric G protein Rho, we studied the effect of carbachol on actin reorganization (stress fiber formation) in cultured human airway smooth muscle cells that expressed mainly M2 muscarinic receptors by dual- fluorescence labeling of filamentous (F) and monomeric (G) actin. F-actin was labeled with FITC-labeled phalloidin, and G-actin was labeled with Texas Red-labeled DNase I. Carbachol stimulation induced stress fiber formation (increased F-actin staining) in the cells and increased the F- to G-actin ratio 3.6 ± 0.4-fold (mean ± SE; n = 5 experiments). Preincubation with pertussis toxin, Clostridium C3 exoenzyme, or tyrosine kinase inhibitors reduced the carbachol-induced increase in stress fiber formation and significantly decreased the F- to G-actin ratio, whereas a mitogen-activated protein kinase inhibitor, a phosphatidylinositol 3-kinase inhibitor, and a protein kinase C inhibitor were without effect. This study demonstrates that in cultured human airway smooth muscle cells, muscarinic-receptor activation induces stress fiber formation via a pathway involving a pertussis-sensitive G protein, Rho proteins, and tyrosine phosphorylation.To determine whether M2 muscarinic receptors are linked to the monomeric G protein Rho, we studied the effect of carbachol on actin reorganization (stress fiber formation) in cultured human airway smooth muscle cells that expressed mainly M2 muscarinic receptors by dual-fluorescence labeling of filamentous (F) and monomeric (G) actin. F-actin was labeled with FITC-labeled phalloidin, and G-actin was labeled with Texas Red-labeled DNase I. Carbachol stimulation induced stress fiber formation (increased F-actin staining) in the cells and increased the F- to G-actin ratio 3.6 +/- 0.4-fold (mean +/- SE; n = 5 experiments). Preincubation with pertussis toxin, Clostridium C3 exoenzyme, or tyrosine kinase inhibitors reduced the carbachol-induced increase in stress fiber formation and significantly decreased the F- to G-actin ratio, whereas a mitogen-activated protein kinase inhibitor, a phosphatidylinositol 3-kinase inhibitor, and a protein kinase C inhibitor were without effect. This study demonstrates that in cultured human airway smooth muscle cells, muscarinic-receptor activation induces stress fiber formation via a pathway involving a pertussis-sensitive G protein, Rho proteins, and tyrosine phosphorylation.

Adenosine regulation of alveolar fluid clearance

Adenosine is a purine nucleoside that regulates cell function through G protein-coupled receptors that activate or inhibit adenylyl cyclase. Based on the understanding that cAMP regulates alveolar epithelial active Na+ transport, we hypothesized that adenosine and its receptors have the potential to regulate alveolar ion transport and airspace fluid content. Herein, we report that type 1 (A1R), 2a (A2aR), 2b (A2bR), and 3 (A3R) adenosine receptors are present in rat and mouse lungs and alveolar type 1 and 2 epithelial cells (AT1 and AT2). Rat AT2 cells generated and produced cAMP in response to adenosine, and micromolar concentrations of adenosine were measured in bronchoalveolar lavage fluid from mice. Ussing chamber studies of rat AT2 cells indicated that adenosine affects ion transport through engagement of A1R, A2aR, and/or A3R through a mechanism that increases CFTR and amiloride-sensitive channel function. Intratracheal instillation of low concentrations of adenosine (≤10−8M) or either A2aR- or A3R-specific agonists increased alveolar fluid clearance (AFC), whereas physiologic concentrations of adenosine (≥10−6M) reduced AFC in mice and rats via an A1R-dependent pathway. Instillation of a CFTR inhibitor (CFTRinh-172) attenuated adenosine-mediated down-regulation of AFC, suggesting that adenosine causes Cl− efflux by means of CFTR. These studies report a role for adenosine in regulation of alveolar ion transport and fluid clearance. These findings suggest that physiologic concentrations of adenosine allow the alveolar epithelium to counterbalance active Na+ absorption with Cl− efflux through engagement of the A1R and raise the possibility that adenosine receptor ligands can be used to treat pulmonary edema.

Local Anesthetics Reduce Mortality and Protect against Renal and Hepatic Dysfunction in Murine Septic Peritonitis

Background:Mortality from sepsis frequently results from multiple organ injury and dysfunction. Cecal ligation and puncture is an established murine model of septic peritonitis that produces septic shock characterized by an initial hyperinflammatory response. In addition to their anesthetic properties, local anesthetics have been shown to attenuate inflammatory responses both in vivo and in vitro. In the current study, the ability of local anesthetic infusions to protect against sepsis-induced mortality, as well as renal and hepatic dysfunction after cecal ligation and puncture, was investigated. Methods:C57BL/6 mice received mini-osmotic pumps containing saline (vehicle), 10% lidocaine, or 1% bupivacaine and were subjected to cecal ligation and puncture. Twenty-four hours after cecal ligation and puncture, renal and hepatic functions were assessed as well as markers of inflammation (proinflammatory cytokine protein and mRNA concentrations and myeloperoxidase activity). Renal apoptosis and 7-day survival was also assessed. Results:Mice treated with lidocaine or bupivacaine infusion showed improved survival and had significantly lower plasma creatinine, aspartate aminotransferase, and alanine aminotransferase concentrations compared with mice receiving vehicle alone. Significant reduction in plasma tumor necrosis factor-&agr; and keratinocyte-derived chemokine, as well as reductions in myeloperoxidase activity, intracellular adhesion molecule-1 protein expression, mRNA concentrations of proinflammatory markers, and apoptosis were observed in renal cortices from both local anesthetic groups. Conclusions:The current data demonstrate that local anesthetic infusions confer a protective effect in mice from septic peritonitis by attenuating the hyperacute inflammatory response. This suppression resulted in improved mortality and less progression to acute kidney and liver injury and dysfunction.