Glioma-Associated Oncogene Homologue 3, a Hedgehog Transcription Factor, Contributes to the Classical Hodgkin Lymphoma Microenvironment through the Modulation of a Subset of Inflammatory Chemokines and Cytokines
Khaled Alayed, Kranthi Kunkalla, Changju Qu, Zaher Chakhachiro, L Jeffrey Medeiros, Rajesh Singh, Francisco Vega. MD Anderson Cancer Center, Houston
Background: Glioma associated oncogene homologue 3 (GLI-3), a hedgehog (HH) transcription factor, is consistently and strongly expressed in Hodgkin and Reed-Sternberg cells of classical Hodgkin lymphoma (CHL) but is rarely expressed in other B- or T-cell lymphomas (Hum Pathol, 2011). The function of GLI-3 in CHL is unknown. However, Theodorides et al described GLI-3 expression in stromal thymic cells and proposed that GLI-3 regulates proliferation and differentiation of thymocytes (Blood 2005, 106:1296-1304). Here, we investigated the potential contribution of GLI-3 to the inflammatory microenvironment of CHL.
Design: We used two EBV negative CHL-derived cell lines, HDLM-2 and L428. These cell lines were transiently transfected in duplicate using a sMARTpool mix of GLI3-specific siRNA and non-specific scrambled siRNA (control). Efficiency of the silencing was confirmed by measurement of GLI-3 mRNA levels by quantitative RT-PCR. Quantitative mRNA expression analysis of 84 inflammatory cytokines and chemokines was performed in duplicate using the human inflammatory cytokines and receptors RT2 profiler™ PCR array (SuperArray Bioscience Corporation, Frederick, MD, USA). Genes with a 2-fold or higher change were considered to be down- or up-regulated.
Results: In the HDLM-2 cell line, 14 genes were reproducibly under-expressed after silencing GLI-3, of which 7 genes (CCL1, CCL13, CCL2, CCL20, CCL24, CCL25 and CCL26) are chemokines and 3 (CCR1, CCR3 and CX3CR1) are chemokine receptors. These chemokines are mainly inflammatory chemotactic factors for T-cells, eosinophils, IgA producing B-cells/plasma cells and monocytes. In the L428 cell line, 8 genes were reproducibly under-expressed after silencing GLI-3, of which 6 genes (CCL18, CCL21, CCL25, CXCL13, CXCL2 and XCR1) are chemokines with a chemotactic role for T-cells, B-cells, IgA producing plasma cells and granulocytes. The chemokines CCL25 and IL1B were consistently down-regulated in both cell lines after silencing GLI-3. Silencing of GLI-3 resulted in no up-regulated genes.
Conclusions: Silencing of GLI-3 in CHL resulted in down-regulation of a subset of inflammatory chemokines and cytokines. These results support a role for GLI-3 in generating or maintaining the inflammatory microenvironment in CHL. The down-regulation of CCL25 and IL1B in both CHL cell lines suggests that these genes may be directly controlled by GLI-3.
Wednesday, March 21, 2012 9:30 AM
Poster Session V # 203, Wednesday Morning