Frida Schain

15-Lipoxygenase-1 induces expression and release of chemokines in cultured human lung epithelial cells

15-Lipoxygenase-1 (15-LOX-1) has been proposed to be involved in various physiological and pathophysiological activities such as inflammation, atherosclerosis, cell maturation, and tumorigenesis. Asthma and chronic obstructive pulmonary disease are associated with increased expression of 15-LOX-1 in bronchial epithelial cells, but the potential functions of 15-LOX-1 in airway epithelial cells have not been well clarified. To study the function of 15-LOX-1 in bronchial epithelial cells, we ectopically expressed 15-LOX-1 in the human lung epithelial cell line A549. We found that overexpression of 15-LOX-1 in A549 cells leads to increased release of the chemokines MIP-1alpha, RANTES, and IP-10, and thereby to increased recruitment of immature dendritic cells, mast cells, and activated T cells. These results suggest that an increased expression and activity of 15-LOX-1 in lung epithelial cells is a proinflammatory event in the pathogenesis of asthma and other inflammatory lung disorders.

Hodgkin Reed-Sternberg cells express 15-lipoxygenase-1 and are putative producers of eoxins in vivo

Classical Hodgkin lymphoma has unique clinical and pathological features and tumour tissue is characterized by a minority of malignant Hodgkin Reed–Sternberg cells surrounded by inflammatory cells. In the present study, we report that the Hodgkin lymphoma‐derived cell line L1236 has high expression of 15‐lipoxygenase‐1 and that these cells readily convert arachidonic acid to eoxin C4, eoxin D4 and eoxin E4. These mediators were only recently discovered in human eosinophils and mast cells and found to be potent proinflammatory mediators. Western blot and immunocytochemistry analyses of L1236 cells demonstrated that 15‐lipoxygenase‐1 was present mainly in the cytosol and that the enzyme translocated to the membrane upon calcium challenge. By immunohistochemistry of Hodgkin lymphoma tumour tissue, 15‐lipoxygenase‐1 was found to be expressed in primary Hodgkin Reed–Sternberg cells in 17 of 20 (85%) investigated biopsies. The enzyme 15‐lipoxygenase‐1, however, was not expressed in any of 10 biopsies representing nine different subtypes of non‐Hodgkin lymphoma. In essence, the expression of 15‐lipoxygenase‐1 and the putative formation of eoxins by Hodgkin Reed–Sternberg cells in vivo are likely to contribute to the inflammatory features of Hodgkin lymphoma. These findings may have important diagnostic and therapeutic implications in Hodgkin lymphoma. Furthermore, the discovery of the high 15‐lipoxygenase‐1 activity in L1236 cells demonstrates that this cell line comprises a useful model system to study the chemical and biological roles of 15‐lipoxygenase‐1.

Transcriptional Regulation of 15-Lipoxygenase Expression by Histone H3 Lysine 4 Methylation/Demethylation

15-Lipoxygenase-1 (15-LOX-1) oxidizes polyunsaturated fatty acids to a rich spectrum of biologically active metabolites and is implicated in physiological membrane remodelling, inflammation and apoptosis. Its deregulation is involved in the pathogenesis of diverse cancer and immune diseases. Recent experimental evidence reveals that dynamic histone methylation/demethylation mediated by histone methyltransferases and demethylases plays a critical role in regulation of chromatin remodelling and gene expression. In the present study, we compared the histone 3 lysine 4 (H3-K4) methylation status of the 15-LOX-1 promoter region of the two Hodgkin lymphoma (HL) cell lines L1236 and L428 with abundant and undetectable 15-LOX-1 expression, respectively. We identified a potential role of H3-K4 methylation in positive regulation of 15-LOX-1 transcription. Furthermore, we found that histone methyltransferase SMYD3 inhibition reduced 15-LOX-1 expression by decreasing promoter activity in L1236 cells. SMYD3 knock down in these cells abolished di−/trimethylation of H3-K4, attenuated the occupancy by the transactivator STAT6, and led to diminished histone H3 acetylation at the 15-LOX-1 promoter. In contrast, inhibition of SMCX, a JmjC-domain-containing H3-K4 tri-demethylase, upregulated 15-LOX-1 expression through induction of H3-K4 trimethylation, histone acetylation and STAT6 recruitment at the 15-LOX-1 promoter in L428 cells. In addition, we observed strong SMYD3 expression in the prostate cancer cell line LNCaP and its inhibition led to decreased 15-LOX-1 expression. Taken together, our data suggest that regulation of histone methylation/demethylation at the 15-LOX-1 promoter is important in 15-LOX-1 expression.

Evidence for a pathophysiological role of cysteinyl leukotrienes in classical Hodgkin lymphoma

Classical Hodgkin lymphoma (cHL) is characterized histologically by a minority of malignant Hodgkin Reed‐Sternberg cells surrounded by abundant inflammatory cells, generally believed to be of major importance in the pathophysiology of the disease. Here, we present data that link inflammatory cell‐derived arachidonic acid metabolites, the cysteinyl leukotrienes (CysLT), to the pathogenesis of cHL. Two HL cell lines, L1236 and KMH2, were shown to express functional CysLT1 receptors, responding with a robust calcium signal upon leukotriene (LT) D4 challenge. LTD4 stimulated protein release of tumor necrosis factor‐α, interleukin‐6 and ‐8 by L1236 cells and interleukin‐8 by KMH2 cells. Importantly, all these LTD4‐induced effects were blocked by the CysLT1 receptor‐specific antagonist zafirlukast. Immunohistochemical studies of cHL biopsies and microarray analysis of microdissected cells revealed that the CysLT1 receptor is expressed also by primary Hodgkin Reed‐Sternberg cells. As these cells are surrounded by CysLT‐producing eosinophils, macrophages and mast cells, our results suggest the CysLTs as mediators in the pathogenesis of cHL, contributing to the aberrant cytokine network of this lymphoma.

Interleukin-13 stimulation of the mediastinal B-cell lymphoma cell line Karpas-1106P induces a phenotype resembling the Hodgkin lymphoma cell line L1236.

OBJECTIVE Primary mediastinal B-cell lymphoma (PMBCL) and Hodgkin lymphoma (HL) share many biological and clinical characteristics supporting a common pathogenetic pathway. Interleukin (IL)-13 has an important pathophysiological role in HL. In this study, we asked the question of whether IL-13 is a major contributor to the observed difference in features of inflammation between HL and PMBCL. MATERIALS AND METHODS Expression of IL-13 and IL-4 receptors was studied by flow cytometry, expression of a functional cysteinyl leukotriene receptor type 1 (CysLT1R) was investigated by calcium flux measurement, expression and activity of 15-lipoxygenase type 1 (15-LO-1) was determined by Western blot and reversed-phase high-performance liquid chromatography, respectively, and cytokines were quantified by Bioplex detection. RESULTS Stimulation of the PMBCL cell line Karpas-1106P with IL-13 or IL-4 induced a proinflammatory phenotype similar to that of the HL cell line L1236. Upon interleukin stimulation of the PMBCL cell line, the cellular size increased and cells became multinucleated. Cells also expressed CysLT1R and 15-LO-1, and produced the proinflammatory eoxins. The IL-13 or IL-4 treated PMBCL cell line and the HL cell line secreted a similar set of cytokines such as IL-6, tumor necrosis factor-alpha, interferon-inducible protein-10, interferon-gamma, and RANTES. CONCLUSIONS IL-13 or IL-4 stimulation of the PMBCL cell line Karpas-1106P induced an inflammatory phenotype that resembles that of the HL cell line. Our results suggest that the autocrine release of IL-13 in HL is one critical factor that can at least partly explain the difference in phenotype between these two lymphoma entities.

Differential Expression of Cysteinyl Leukotriene Receptor 1 and 15-Lipoxygenase-1 in Non-Hodgkin Lymphomas

BACKGROUND Arachidonic acid metabolites have been suggested to play an important role in carcinogenesis. We have recently reported that the cysteinyl leukotriene receptor 1 (CysLT1) and 15-lipoxygenase-1 (15-LO-1) are expressed by the malignant Hodgkin Reed-Sternberg cells of Hodgkin lymphoma and certain Hodgkin lymphoma cell lines, and that these cells convert arachidonic acid to the novel proinflammatory eoxins. MATERIALS AND METHODS The expression of the CysLT1 receptor and 15-LO-1 was investigated in a broad range of non-Hodgkin lymphomas (NHLs) by immunohistochemistry. The functionality of the CysLT1 receptor in primary mediastinal B-cell lymphoma (PMBCL) cell lines was studied by calcium mobilization assays. RESULTS Primary mediastinal B-cell lymphoma was the only NHL entity showing tumor cells positive for the CysLT1 receptor (9 of 10 tumors), and the PMBCL cell line Med-B1 expressed functional CysLT1 receptors, responding with a robust calcium signal upon cysteinyl leukotriene challenge. Furthermore, the tumor cells in 1 of 4 T-cell-derived anaplastic large-cell lymphomas, in contrast to all other studied NHLs, strongly expressed 15-LO-1. CONCLUSION Among the NHL entities included in this study, the CysLT1 receptor was exclusively expressed by the tumor cells of PMBCL. Thus, this further corroborates the pathologic overlap between PMBCL and classical Hodgkin lymphoma.

Human 15-lipoxygenase-1 is a regulator of dendritic-cell spreading and podosome formation

Dendritic cells (DCs) involved in proinflammatory immune responses derive mainly from peripheral monocytes, and the cells subsequently mature and migrate into the inflammatory micromilieu. Here we report that suppressing of 15‐lipoxygenase‐1 led to a substantial reduction in DC spreading and podosome formation in vitro. The surface expression of CD83 was significantly lower in both sh‐15‐lipoxygenase‐1 (15‐LOX‐1)‐transduced cells and DCs cultivated in the presence of a novel specific 15‐LOX‐1 inhibitor. The T‐cell response against tetanus‐pulsed DCs was only affected to a minor extent on inhibition of 15‐LOX‐1. In contrast, endocytosis and migration ability of DCs were significantly suppressed on 15‐LOX‐1 inhibition. The expression of 15‐LOX‐1 in DCs was also demonstrated in affected human skin in atopic and contact dermatitis, showing that the enzyme is indeed expressed in inflammatory diseases in vivo. This study demonstrated that inhibiting 15‐LOX‐1 ledtoanimpairedpodosome formation in DCs, and consequently suppressed antigen uptake and migration capacity. These results indicated that 15‐LOX‐1 is a potential target for inhibiting the trafficking of DCs to lymphoid organs and inflamed tissues and decreasing the inflammatory response attenuating symptoms of certain immunologic and inflammatory disorders such as dermatitis.—Han, H., Liang, X., Ekberg, M., Kritikou, J. S., Brunnström, Å., Pelcman, B., Matl, M., Miao, X., Andersson, M., Yuan, X., Schain, F., Parvin, S., Melin, E., Sjåberg, J., Xu, D., Westerberg, L. S., Bjårkholm, M., Claesson, H.‐E. Human 15‐lipoxygenase‐1 is a regulator of dendritic‐cell spreading and podosome formation. FASEB J. 31, 491–504 (2017). www.fasebj.org