Knight. Acid and particulate-induced aspiration lung injury in mice: importance of MCP-1.

SM, Knight PR 3rd. Acid aspiration-induced lung inflammation and injury are exacerbated in NADPH oxidase-deficient mice.

In this study, we examined the mitogen-activated protein kinase (MAPK) cascade in micrometastatic cell lines generated from rib bone marrow (RBM) of patients undergoing resection of esophagogastric malignancies. The molecular mechanism(s) involved in esophagogastric MAPK activation have not previously been investigated. Constitutive activation of both ERK1 and -2 isoforms was evident in each of the five RBM cell lines. Elk-1, a transcription factor activated by the ERK1/2 pathway was also found to be constitutively activated. Cell lines generated from metastases of involved lymph nodes (OC2) and ascites (OC1) of patients with esophageal cancer do not display, however, hyperphosphorylation of ERK1/2. Constitutive RBM ERK1/2 activation is protein kinase C and phosphatidylinositol 3-kinase dependent. Surprisingly, constitutive ERK1/2 activation is MEK-independent. Pharmacological inhibition of MEK with two specific inhibitors, PD 98059 and U0126, were both ineffective in blocking ERK activation. Similarly, the use of a dominant negative MEK mutant was without effect. Interestingly, experiments overexpressing two different dominant negative Pak1 mutants significantly reduced RBM ERK1/2 activation, albeit not to the same extent for all cell lines. We also examined the role of three different phosphatases, PAC1, MKP-1, and -2. While RBM ERK1/2 activation was found to be PAC1-and MKP-2-independent, surprisingly, MKP-1 was down-regulated in all five RBM cell lines. In conclusion, we provide evidence for the first time for a MEK-independent constitutive ERK1/2 activation pathway in esophagogastric RBM cell lines. These findings have important implications for drug treatment strategies which currently target MEK in other forms of cancer.Esophageal cancer is an aggressive tumor which responds poorly to treatment and has a poor prognosis (1, 2). Approximately half of patients diagnosed with localized esophageal cancer die of metastatic disease within the first 2 years following tumor resection. We have succeeded in developing a number of esophagogastric cell lines from the rib bone marrow of patients (3). We have previously shown that these cells are viable, proliferate, and grow independently in tissue culture and form malignant tumors in athymic nude mice (4). These metastatic cells are representative of the disseminated progenitors of secondary tumors and are the appropriate targets for treatment.The precise molecular events leading to the acquisition of the metastatic phenotype remain largely unknown. Members of the Ras superfamily of small GTP-binding proteins have been implicated in tumor progression and are found to be activated in 20 to 30% of tumors (5). In its active GTP-bound state, Ras activates the serine threonine kinase Raf (6). Raf upon activation in turn phosphorylates the dual specific kinase MEK 1 (also known as MAP kinase kinase or MAPKK) which in turn phosphorylates the MAP kinases ERK1 and -2 (for extracellular regulated kinases 1 and 2). Although MEK is the only known kinase directly downstream of Raf...

Recruitment of neutrophils and release of reactive oxygen species are considered to be major pathogenic components driving acute lung injury (ALI). However, NADPH oxidase, the major source of reactive oxygen species in activated phagocytes, can paradoxically limit inflammation and injury. We hypothesized that NADPH oxidase protects against ALI by limiting neutrophilic inflammation and by activating Nrf2, a transcriptional factor that induces anti-oxidative and cytoprotective pathways. Our objective was to delineate the roles of NADPH oxidase and Nrf2 in modulating acute lung inflammation and injury in clinically relevant models of acute gastric aspiration injury, a major cause of ALI. Acid aspiration caused increased ALI (as assessed by bronchoalveolar lavage fluid albumin concentration) in both NADPH oxidase-deficient (p47phox−/−) mice and in Nrf2−/− mice compared to wild-type mice. NADPH oxidase reduced airway neutrophil accumulation, but Nrf2 decreased ALI without affecting neutrophil recovery. Acid injury resulted in a 120-fold increase in mitochondrial DNA, a pro-inflammatory and injurious product of cellular necrosis, in cell-free bronchoalveolar lavage fluid. Pharmacologic activation of Nrf2 by the triterpenoid, CDDO-Im, limited aspiration-induced ALI in wild-type mice and reduced endothelial cell injury caused by mitochondrial extract-primed human neutrophils, leading to the conclusion that NADPH oxidase and Nrf2 have coordinated, but distinct, functions in modulating inflammation and injury. These results also point to Nrf2 as a therapeutic target to limit ALI by attenuating neutrophil-induced cellular injury.