Julie Negri

Microarray analysis of distinct gene transcription profiles in non-eosinophilic chronic sinusitis with nasal polyps.

Background Recent literature has indicated the feasibility of microarray analysis in the characterization of chronic sinusitis. We hypothesized that previously unexplored inflammatory mechanisms would be involved in the pathophysiology of noneosinophilic chronic rhinosinusitis with nasal polyps (NE-CRSwNP) and that this technology could be used to identify the gene expression of these novel and previously known mediators. Methods Patients with CRSwNP failing medical therapy were prospectively enrolled and NP tissue was removed at time of surgery. NE-CRSwNP was diagnosed based on clinical parameters including absence of allergic disease and confirmed with histopathology showing lack of eosinophilic infiltration. Messenger RNA (mRNA) transcripts extracted from study and control patients were then subjected to microarray analysis using Affymatrix based chips. Validation of findings was then confirmed via quantitative reverse transcription polymerase chain reaction (qRT-PCR). Results Microarray analysis revealed activation of pathways involved in antigen presentation, cellular movement, hematopoiesis, carcinogenesis, apoptosis, and cell signaling. Previously unexplored genes of interest were identified and their differential regulation was validated via qRT-PCR. Our data showed up-regulation of innate inflammation genes (IL-6, IL-8, and monocyte chemoattractant protein 1), hypoxia-induced inflammation 1α, and fibrosis (tenascin) and lack of up-regulation of genes associated with allergic, eosinophilic inflammation (IL-4 and IL-13). Additionally, the genes for CXCL1 and autocrine motility factor receptor were novelly identified to be up-regulated Conclusion This study explores the utility of gene microarray technology in identifying unexplored targets of immune dysregulation in NE-CRSwNP. Furthermore, the data characterize the immunologic profile of NE-CRSwNP as it differs from other forms of CRSwNP, in particular, those known to be associated with eosinophilic inflammation.

Lack of a Role for Cross-Reacting Anti-Thyroid Antibodies in Chronic Idiopathic Urticaria

The etiology of chronic idiopathic urticaria (CIU) is attributed to autoantibodies directed against the alpha-chain of the high-affinity IgE receptor (FcepsilonRIalpha) or IgE on mast cells in 30-60% of patients. Approximately 30% of CIU patients have Hashimotos thyroiditis (HT). We investigated the pathophysiologic relationship of anti-thyroid and anti-FcepsilonRIalpha antibodies. Nine individuals with both CIU and HT underwent autologous serum skin testing (ASST) and sera were assayed for thyroid autoantibodies, thyroid-stimulating hormone, and anti-FcepsilonRIalpha antibodies. Serum samples were studied for their ability to activate a human mast cell line (LUVA) as determined by cysteinyl leukotriene (CysLT) production. Experiments were performed to determine whether epitope cross-reactivity could explain the high incidence of HT found in CIU patients. A significant proportion of CIU patients had a positive ASST (nine of six) and anti-FcepsilonRIalpha antibodies (six of nine). Incubation of patient sera with FcepsilonRIalpha, but not thyroglobulin or thyroid peroxidase, resulted in the decreased ability to detect anti-FcepsilonRIalpha antibodies. Incubation with thyroid antigens did not inhibit CysLT production by mast cells. Epitopic cross-reactivity does not explain the increased prevalence of HT found in CIU patients. The frequent concurrence of HT and CIU likely reflects a genetic tendency toward autoimmune diseases.

Biological Effects of Leukotriene E4 on Eosinophils

Studies demonstrate the existence of novel receptors for cysteinyl leukotrienes (CysLTs) that are responsive to leukotriene (LT) E4 and might be pathogenic in asthma. Given the eosinophilic infiltration in this disorder, we investigated eosinophil expression of P2Y12 and gpr99 and their capacity to respond to LTE4. Receptor transcript expression was investigated via quantitative PCR and surface protein expression via flow cytometry. We investigated LTE4 influences on eosinophils including Ca(+2) flux, cAMP induction, modulation of adhesion molecule expression, apoptosis and degranulation. Eosinophils displayed both transcript and surface protein expression of P2Y12 and gpr99. We could not find evidence of LTE4 activation of eosinophils, however, LTE4 induced cAMP expression, and preincubation of eosinophils with LTE4 inhibited degranulation. Even though eosinophils are an important source of CysLTs in AERD, eosinophils are not themselves the pro-inflammatory biological target and, in contrast, LTE4 via cAMP primarily elicits anti-inflammatory responses.