Diagnosis and treatment of allergic bronchopulmonary aspergillosis.

Allergic bronchopulmonary aspergillosis (ABPA) is an underdiagnosed pulmonary disorder in asthmatic patients and patients with cystic fibrosis. Its clinical and diagnostic manifestations arise from an allergic response to multiple antigens expressed by fungi, most commonly Aspergillus fumigatus, colonizing the bronchial mucus. The clinical course is one of recurrent exacerbations characterized by chest infiltrates evident on chest x-ray films and associated with cough, wheeze, and sputum production that usually respond to oral corticosteroid treatment. Specific immunologic and radiologic markers of disease include elevation of the total serum IgE levels, presence of aspergillus IgE antibodies, and the occurrence of central bronchiectasis. Long-term treatment with corticosteroids is often required for effective management. The adverse effects of chronic corticosteroid use have led to attempts at treatment with antifungal agents such as itraconazole. Itraconazole has been reported anecdotally to be effective, and evidence for its effectiveness in randomized trials is still accruing. Consideration should be given to its use as a corticosteroid-sparing agent or for treatment of patients in whom corticosteroid response is poor. The natural history and prognosis of ABPA are not well characterized but may be complicated by progression to bronchiectasis and pulmonary fibrosis. If ABPA is diagnosed and treated before the development of bronchiectasis and fibrosis, these complications may be prevented.

Invited review: plasma membrane stress failure in alveolar epithelial cells.

In this review, we examine the hypothesis that plasma membrane stress failure is a central event in the pathophysiology of injury from alveolar overdistension. This hypothesis leads us to consider alveolar micromechanics and specifically the mechanical interactions between lung matrix and alveolar epithelial cell cytoskeleton and plasma membrane. We then explore events that are central to the regulation of plasma membrane tension and detail the lipid-trafficking responses of in vitro deformed and/or injured cells. We conclude with a reference to upregulation of stress-responsive genes after membrane injury and resealing.

Peak calf blood flow estimates are higher with Dohn than with Whitney plethysmograph.

Estimates of calf blood flow with venous occlusion plethysmography vary widely between studies, perhaps due to the use of different plethysmographs. Consequently, we compared calf blood flow estimates at rest and during reactive hyperemia in eight healthy subjects (four men and four women) with two commonly used plethysmographs: the mercury-in-silastic (Whitney) strain gauge and Dohn air-filled cuff. To minimize technical variability, flow estimates were compared with a Whitney gauge and a Dohn cuff on opposite calves before and after 10 min of bilateral femoral arterial occlusion. To account for any differences between limbs, a second trial was conducted in which the plethysmographs were switched. Resting flows did not differ between the plethysmographs (P = 0.096), but a trend toward lower values with the Whitney was apparent. Peak flows averaged 37% lower with the Whitney (27.8 +/- 2.8 ml.dl-1.min-1) than with the Dohn plethysmograph (44.4 +/- 2.8 ml.dl-1.min-1; P < 0.05). Peak flow expressed as a multiple above baseline was also lower with the Whitney (10-fold) than with the Dohn plethysmograph (14.5-fold; P = 0.02). Across all flows at rest and during reactive hyperemia, estimates were highly correlated between the plethysmographs in all subjects (r2 = 0.96-0.99). However, the mean slope for the Whitney-Dohn relationship was only 60 +/- 2%, indicating that over a wide range of flows the Whitney gauge estimate was 40% lower than that for the Dohn cuff. These results demonstrate that the same qualitative results can be obtained with either plethysmograph but that absolute flow values will generally be lower with Whitney gauges.

Validation of a new live cell strain system: characterization of plasma membrane stress failure.

Motivated by our interest in lung deformation injury, we report on the validation of a new live cell strain system. We showed that the system maintains a cell culture environment equivalent to that provided by conventional incubators and that its strain ouput was uniform and reproducible. With this system, we defined cell deformation dose (i.e., membrane strain amplitude)-cell injury response relationships in alveolar epithelial cultures and studied the effects of temperature on them. Deformation injury occurred in the form of reversible, nonlethal plasma membrane stress failure events and was quantified as the fraction of cells with uptake and retention of fluorescein-labeled dextran (FITC-Dx). The undeformed control population showed virtually no FITC-Dx uptake at any temperature, which was also true for cells strained by 3%. However, when the membrane strain was increased to 18%, ~5% of cells experienced deformation injury at a temperature of 37 degrees C. Moreover, at that strain, a reduction in temperature to 4 degrees C resulted in a threefold increase in the number of cells with plasma membrane breaks (from 4.8 to 15.9%; P < 0.05). Cooling of cells to 4 degrees C also lowered the strain threshold at which deformation injury was first seen. That is, at a 9% substratum strain, cooling to 4 degrees C resulted in a 10-fold increase in the number of cells with FITC-Dx staining (0.7 vs. 7.5%, P < 0.05). At that temperature, A549 cells offered a 50% higher resistance to shape change (magnetic twisting cytometry measurements) than at 37 degrees C. We conclude that the strain-injury threshold of A549 cells is reduced at low temperatures, and we consider temperature effects on plasma-membrane fluidity, cytoskeletal stiffness, and lipid trafficking as responsible mechanisms.

Integrin α9ß1 promotes malignant tumor growth and metastasis by potentiating epithelial-mesenchymal transition.

The integrin α9ß1 binds a number of extracellular matrix components to mediate cell adhesion, migration and tissue invasion. Although expressed in a variety of normal human cells including endothelium, it is also expressed in cancer cells. We have previously shown that α9ß1 binds VEGF-A to facilitate angiogenesis, an important component of the tumor microenvironment. As α9ß1 induces accelerated cancer cell migration, we wished to determine what role it played in cancer growth and metastasis. In this study, we show that α9ß1 expression induces molecular changes consistent with epithelial-mesenchymal transition. In addition, we found that α9ß1 forms a tri-partite protein complex with ß-catenin and E-cadherin, which dissociates following integrin activation and subsequent src and ß-catenin phosphorylation. These findings were consistent in cells in which: α9ß1 was exogenously over-expressed, or when its expression was suppressed in cancer cells endogenously expressing α9ß1. These in vitro results are biologically significant as α9ß1-expressing cancer cells induce greater tumor growth and metastases in mice as compared to the cells without α9ß1 expression or when integrin expression is suppressed. Furthermore, integrin α9ß1 is expressed in primary human small cell lung cancer and patients having a high expression of α9ß1 demonstrated significantly worse long-term survival compared with patients with low α9ß1 expression. These findings highlight a novel mechanism of integrin α9ß1 function in human cancer.

Deformation-induced lipid trafficking in alveolar epithelial cells.

Mechanical ventilation with a high tidal volume results in lung injury that is characterized by blebbing and breaks both between and through alveolar epithelial cells. We developed an in vitro model to simulate ventilator-induced deformation of the alveolar basement membrane and to investigate, in a direct manner, epithelial cell responses to deforming forces. Taking advantage of the novel fluorescent properties of BODIPY lipids and the fluorescent dye FM1-43, we have shown that mechanical deformation of alveolar epithelial cells results in lipid transport to the plasma membrane. Deformation-induced lipid trafficking (DILT) was a vesicular process, rapid in onset, and was associated with a large increase in cell surface area. DILT could be demonstrated in all cells; however, only a small percentage of cells developed plasma membrane breaks that were reversible and nonlethal. Therefore, DILT was not only involved in site-directed wound repair but might also have served as a cytoprotective mechanism against plasma membrane stress failure. This study suggests that DILT is a regulatory mechanism for membrane trafficking in alveolar epithelia and provides a novel biological framework within which to consider alveolar deformation injury and repair.

Stretch induces cytokine release by alveolar epithelial cells in vitro.

Mechanical ventilation can injure the lung, causing edema and alveolar inflammation. Interleukin-8 (IL-8) plays an important role in this inflammatory response. We postulated that cyclic cell stretch upregulates the production and release of IL-8 by human alveolar epithelium in the absence of structural cell damage or paracrine stimulation. To test this hypothesis, alveolar epithelial cells (A549 cells) were cultured on a deformable silicoelastic membrane. When stretched by 30% for up to 48 h, the cells released 49 +/- 34% more IL-8 (P < 0.001) than static controls. Smaller deformations (20% stretch) produced no consistent increase in IL-8. Stretch of 4 h duration increased IL-8 gene transcription fourfold above baseline. Stretch had no effect on cell proliferation, cell viability as assessed by (51)Cr release assay, or the release of granulocyte-macrophage colony-stimulating factor and tumor necrosis factor-alpha. We conclude that deformation per se can trigger inflammatory signaling and that alveolar epithelial cells may be active participants in the alveolitis associated with ventilator-induced lung injury.