Gerald S. Davis

Efficacy of infliximab in extrapulmonary sarcoidosis: results from a randomised trial

The aim of the present study was to investigate the efficacy of infliximab for the treatment of extrapulmonary sarcoidosis. A prospective, randomised, double-blind, placebo-controlled trial was conducted, with infliximab at 3 and 5 mg·kg−1 body weight administered over 24 weeks. Extrapulmonary organ severity was determined by a novel severity tool (extrapulmonary physician organ severity tool; ePOST) with an adjustment for the number of organs involved (ePOSTadj). In total, 138 patients enrolled in the trial of infliximab versus placebo for the treatment of chronic corticosteroid-dependent pulmonary sarcoidosis. The baseline severity of extrapulmonary organ involvement, as measured by ePOST, was similar across treatment groups. After 24 weeks of drug-therapy study, the change from baseline to week 24 in ePOST was greater for the combined infliximab group compared with the placebo group. After adjustment for the number of extrapulmonary organs involved, the improvement in ePOSTadj observed in the combined infliximab group was also greater than that observed in placebo-treated patients, after 24 weeks of therapy. The improvements in ePOST and ePOSTadj were not maintained during a subsequent 24-week washout period. Infliximab may be beneficial compared with placebo in the treatment of extrapulmonary sarcoidosis in patients already receiving corticosteroids, as assessed by the severity tool described in the present study.

Pathogenesis of silicosis: current concepts and hypotheses.

Silicosis is caused by the inhalation of crystalline silica (silicon dioxide, SiO2) in various forms. This review proposes that the cascade of inflammatory and fibrotic events involved in cell-mediated, and possibly humoral, immune responses also produces silicosis. The hypothesis rests on the central concept that interactions between silica and pulmonary macrophages are the pivotal events in the pathogenesis of silicosis. Resident and recruited pulmonary macrophages demonstrate intimate contact with silica from the moment of deposition, and throughout the time the particles remain in the lung. The silica probably exerts its effects on the macrophages that ingest it by altering their function while they are alive, rather than merely by disrupting them. The macrophage appears to be stimulated to secrete mediator substances, such as interleukin-1 (IL-1), which alter the function and behavior of other cells. Lymphocytes and macrophages appear in close proximity to one another in developing silicotic nodules, and increased proportions of lymphocytes are found in bronchoalveolar lavage specimens from animals and humans with silica dust exposure. Hypothetically, macrophages influence and activate lymphocytes, which then feed back to amplify the response by stimulating the same or other recruited macrophages. An expanded and activated population of lymphocytes under the influence of IL-1 in turn can secrete a variety of lymphokines which profoundly alter monocyte/macrophage function. The macrophage has been implicated as a major cause for the fibrosis that accompanies silicosis. The products of activated T-lymphocytes, particularly interferon-gamma (IF-g), are potent stimulators of secretion from macrophages of a fibroblast growth competence factor, macrophage-derived growth factor (MDGF). IL-1 may have an additional stimulatory effect on fibroblasts. Neutrophils and macrophages also may be important in silicosis because of their potent ability to cause lung tissue injury. Silicosis provides a model of chronic diffuse interstitial immunologic and fibrotic lung disease in which the cause is known, can be applied in defined doses, and tracked in the lung throughout the course of the disease. Further studies should provide better understanding of the mechanisms that goven pulmonary injury, inflammation, repair, and fibrosis.

Rapid early onset lymphocyte cell death in mice resistant, but not susceptible to Leishmania major infection

Leishmania major (Lm) infection in mice is a prototypical model for the role of immune deviation in disease resistance. Resistant strains of mice develop a Th1 response to Lm infection, distinguished by secretion of IL-12 and interferon γ. In contrast, susceptible strains display sustained IL-4 expression characteristic of a Th2 response. However, when mechanisms of cell death are blocked, mice display a susceptible phenotype even in the presence of a strong Th1 response, suggesting that cell death, and not cytokine bias, may be an importnt factor in disease resistance. Here, we investigated this hypothesis by comparing lymphocyte cellularity, cell death and Fas expression in resistant CBA and susceptible BALB/c mice during the course of Lm infection. We found that delayed onset of cell death and late Fas induction correlated with massive lymphocyte accumulation and susceptibility to leishmaniasis, while early cell death and rapid Fas induction occurred in resistant mice.

Inhibition of LPS-induced tumor necrosis factor-alpha production by colchicine and other microtubule disrupting drugs.

Colchicine has been shown to act as an antiinflammatory agent. In this study, we examined whether colchicine and other microtubule-depolymerizing drugs affected the production of TNF-alpha. When rat peritoneal macrophages were stimulated by LPS, addition of colchicine, vincristine, vinblastine or nocodazole was found to inhibit TNF-alpha release in a concentration-dependent manner. Suppression of TNF-alpha release was not due to interference with secretion as the cytokine did not accumulate intracellularly following colchicine treatment. Colchicine markedly enhanced PGE2 release from LPS-stimulated macrophages. However, addition of the cyclooxygenase inhibitor indomethacin only partially reversed the suppressive effect of colchicine on TNF-alpha production. Colchicine caused a strong reduction of LPS-induced TNF-alpha mRNA accumulation, suggesting that a pretranslational effect may represent the primary mechanism by which colchicine reduced TNF-alpha production. These observations could have clinical relevance in ameliorating undesirable effects due to excessive TNF-alpha production, for example following LPS stimulation of monocytes/macrophages in gram-negative sepsis. Furthermore, these drugs may provide useful tools to study the apparent involvement of the microtubular system in cytokine gene expression and cytokine production.

QUANTITATIVE IMAGE ANALYSIS OF LUNG CONNECTIVE TISSUE IN MURINE SILICOSIS

Pulmonary brosis is a disabling consequence of many lung diseases but is difficult to quantify. Lucifer yellow CH uorescent dye (LY) appears to stain connective tissue matrix macromolecules selectively. Laser scanning confocal microscopy can quantify the intensity of uorescence and determine the area of uorescent material. We hypothesized that the abundance of lucifer yellow stained matrix macromolecules in lung tissue sections could be measured by laser scanning confocal microscopy, would reflect differences between varying degrees of pulmonary brosis, and could be compared directly to biochemical measurements of lung collagen. We exposed C57B1/6 and 129 strains of mice by aerosol to cristobalite silica (70 mg/m3, 12 days, 5 hours/day) or sham-air and examined them 2 and 16 weeks after exposure. The area of LY-stained matrix in tissue sections was quantitated by laser scanning confocal microscopy, and total lung collagen was measured biochemically as hydroxyproline (OH-proline). The LY-stained connective tissue matrix appeared as bright linear bands in the alveolar septae, and was increased significantly by image analysis in C57B1/6 and 129 mice with silicosis 16 weeks after exposure. Total lung OH-proline was significantly increased in silica-exposed mice from both stains at both time points. Comparing all 8 groups, there was a signicant linear correlation between the average area of connective tissue measured by LY stain and the total OH-proline per lung measured by chemical analysis (r =. 72, P =. 042). LY staining and confocal microscopy with image analysis offers a rapid technique for quantitative measurements of the extent of pulmonary brosis.

Lipids in bronchoalveolar lavage fluid from patients with sarcoidosis.

The recovery of protein and two specific surfactant lipids, phosphatidylcholine and phosphatidylglycerol, from bronchoalveolar lavage fluid is altered in chronic and acute non-granulomatous interstitial lung disease. This study set out to determine whether the same is true for patients with sarcoidosis. The median value for recovery of protein from lavage fluid was significantly higher in 21 patients with sarcoidosis than in 19 normal subjects (18 v 11 mg), while the median value for phospholipid recovery was significantly lower (4 v 1.7 mg). There were no changes in the proportions of phosphatidylcholine and phosphatidylglycerol. In addition, significantly less of the neutral lipid, cholesterol, was recovered (3.2 v 1.5 mg). The combined values of three biochemical measurements, non-phospholipid polar lipid, non-polar lipid, and protein, correctly classified all 40 subjects in our series; in a further group of nine normal subjects and 11 patients with sarcoidosis it allowed all but one normal subject to be classified correctly. These results are discussed in terms of alterations in epithelial cell function in interstitial disease.

Interleukin-12 is not essential for silicosis in mice

BackgroundSilicosis features foci of inflammation where macrophages and lymphocytes precede and accompany fibroblast proliferation, alveolar epithelial hyperplasia, and increased deposition of connective tissue matrix material. In the mouse following silica inhalation there is recruitment of natural killer-, B-, and CD4+ and CD8+ lymphocytes to the alveolar spaces, enlargement of bronchial-associated lymphoid tissues (BALT), and aggregation of lymphocytes surrounding small airways and blood vessels. A substantial fraction of the recruited lung lymphocytes produce interferon-γ (IFN-γ), and IFN-γ gene-deleted mice develop less silicosis than wild-type mice. Interleukin-12 (IL-12) is an important pathway for driving the adaptive immune response towards a TH1-like phenotype. We hypothesized that IL-12 might stimulate lymphocyte activation and the up-regulation of IFN-γ, and consequently be an essential mediator for silicosis.ResultsC57Bl/6 wild-type (WT) and IL-12 deficient (IL-12 KO) mice were exposed to sham-air or crystobalite silica (61 mg/m3) by inhalation for 5 hours/day for 12 days and then studied from 1 to 112 days after exposure. Mice exposed to sham-air had normal lung histology at all time points. WT mice exposed to titanium dioxide (72 mg/m3) showed pulmonary macrophage recruitment but no increase in lung collagen. Both WT and IL-12 KO mice exposed to silica showed similar progressive lung pathology, increased wet lung weight and increased total lung collagen (hydroxyproline). IL-12 p35 mRNA was not increased in either strain after silica exposure; IL-12 p40 mRNA was up-regulated after silica in WT mice and constitutively absent in the IL-12 KO mice. IL-18 mRNA was not increased after silica exposure. The expression of IL-15 (an important driver for innate immunity, Natural Killer cell activation, and IFN-γ production) was abundant in air-exposed mice and was increased slightly in the lungs of mice with silicosis.ConclusionThe axis of IL-12 driving IFN-γ production is not essential for the full manifestations of silicosis in mice exposed to a crystobalite silica aerosol.

Suppression of LPS-Induced Tumor Necrosis Factor-α Gene Expression by Microtubule Disrupting Agents

Microtubule disrupting agents such as colchicine have been shown to reduce TNF-alpha production in macrophages. To examine molecular mechanisms underlying the action of colchicine, TNF-alpha gene expression was studied in the murine macrophage cell line PU5-1.8. An LPS stimulation caused an intense up-regulation of TNF-alpha gene expression which was followed by a high TNF-alpha protein production. Simultaneous addition of colchicine (10 microM) suppressed LPS-induced TNF-alpha mRNA accumulation by one-third and TNF-alpha protein release by two-thirds. This effect was shared by vinblastine and vincristine, chemically different agents that also disrupt microtubule polymerization. For full suppressive activity on TNF-alpha gene expression, colchicine had to be present for 3 h in LPS-stimulated macrophage cultures. With nuclear run-on transcription experiments we could demonstrate that colchicine primarily inhibited de novo gene transcription and did not accelerate degradation of TNF-alpha mRNA in actinomycin D-treated macrophages. Thus, the well-known antiinflammatory action of microtubule depolymerizing agents may be largely due to a reduced TNF-alpha gene expression.

Changes in the Surface Morphology of Human Alveolar Macrophages Induced by Tobacco and Marijuana Smoking

The alveolar macrophage (AM) is pleomorphic in shape, with complex surface folds and ruffles. Two morphologic types of human AM from nonsmokers were identified in scanning electron microscopic studies of fresh plastic-adherent mono layers: flat cells (55%) and round cells (45%). The assumption of a flat, or spread, cell shape was complete after 30 min in tissue culture, and was dependent on active cell metabolism. The flat cell appeared to be the more active phagocyte, and the round cell relatively quiescent. Distinct morphologic changes in individual cells in the population were found in specimens from tobacco and marijuana smokers. These studies demonstrate qualitative and quantitative heterogeneity of the human bronchoalveolar lavage AM population, and indicate that this population can change in response to chronic inhaled pollutant exposure.

The role of endobronchial ultrasound‐guided transbronchial needle aspiration (EBUS TBNA) in the diagnosis of sarcoidosis

Gerald S. Davis, MD, FACP, FACCP, is a Professor of Medicine with tenure at the University of Vermont College of Medicine and former Division Director of the Pulmonary Disease and Critical Care Medicine Unit. He is the Director of the Bronchoscopy Service and the Physician Leader of the Lung Cancer Multi-Disciplinary Program at Fletcher Allen Health Care, the affiliated university hospital. He has carried out National Institutes of Health-sponsored and industry-sponsored clinical and translational laboratory research related to sarcoidosis, idiopathic pulmonary fibrosis, and silicosis for more than 30 years. He serves on advisory panels and data and safety monitoring boards for both the National Heart Lung and Blood Institute and for industry sponsors.