Real-time polymerase chain reaction determination of cytokine mRNA expression profiles in Hodgkin's lymphoma

Abstract

BACKGROUND AND OBJECTIVES:Classical Hodgkin's lymphoma (HL) is a malignant disorder characterized by a small number of tumor cells and inflammatory cells. Both the tumor cells and the inflammatory cells produce cytokines which are thought to contribute to the clinical parameters of HL. Quantification of these cytokines at a protein level is still somewhat imprecise. We, therefore, used a method to quantify cytokine mRNA expression in HL cell lines and lymph node biopsies. DESIGN AND METHODS: We used real-time quantitative polymerase chain reaction (RQ-PCR) to investigate mRNA expression for interleukin (IL)-1alpha, IL-1beta, IL-2, IL-4, IL-5, IL-8, IL-12p35, IL-12p40, IL-13, IL-15, interferon (IFN)-gamma and tumor necrosis factor (TNF-alpha) in lymph node tissue from 15 patients with classic Hodgkin's lymphoma (c-HL) and one with lymphocyte predominance (LP) HL. HL-derived cell lines L1236, L540, and L428 were also investigated. Reactive lymphatic tissue (n=6) and peripheral blood mononuclear cells (PBMC) from healthy donors (n=4) before and after stimulation were used as controls. In 5 c-HL samples the cytokine expression in T lymphocytes was also studied by flow cytometry. RESULTS: All c-HL samples (but not LP) expressed IL-13 mRNA. This cytokine was not found in non-stimulated PBMC or in reactive lymphatic tissue. Expression of IL-10, IL-1beta, IL-15 and IL-12p35 mRNA was higher in HL samples than in PBMC and reactive lymphatic tissue. Expression of IL-10, IL-1beta, TNF-alpha and IFN-gamma mRNA was significantly higher in the EBV+ HL samples (n=6) than in the EBV- cases. All HL cell lines showed high expression of IL-13, IL-12p35, TNF-alpha and IL-15 mRNA. IFNg mRNA levels were high in L428 and L540 cells, IL-10 in L1236 cells and L540 cells, IL-5 in L428 cells and IL-4 in L1236 cells. INTERPRETATION AND CONCLUSIONS: Cytokine mRNA levels can be measured by RQ-PCR using a limited amount of tissue. This method gives valuable information on biological variation between different HL samples and may contribute to unraveling the complex cytokine network contributing to the clinical and biological heterogeneity of this disease.