Abstract
To get a comprehensive picture of cytokine expression in health and disease is difficult, cytokines are transiently and locally expressed, and protein analyses are burdened by biological modifications, technical issues, and sensitivity to handling of samples. Thus, alternative methods, based on molecular techniques for cytokine mRNA analyses, are often used. We compared cytokine mRNA and protein expression to evaluate whether cytokine mRNA profiles can be used instead of protein analyses. In kinetic experiments, cytokine mRNA and protein expression of IL-1β, IL-6, IL-8, TNF-α, and TNF-β/LTA were studied using real-time RT-qPCR and Luminex® microarrays in the ovarian cancer cell lines OVCAR-3, SKOV-3 and the T-cell line Jurkat, after activation of transcription by thermal stress. In addition, we analyzed IL-6 and IL-8 mRNA and protein in a small number of ovarian cancer patients. Ovarian cancer cells can express cytokines on both mRNA and protein level, with 1-4hours' time delay between the mRNA and protein peak and a negative Spearman correlation. The mRNA and protein expression in patient samples was poorly correlated, reflecting previous studies. Cytokine mRNA and protein expression levels show diverging results, depending on the material analyzed and the method used. Considering the high sensitivity and reproducibility of real-time RT-qPCR, we suggest that cytokine mRNA profiles could be used as a proxy for protein expression for some specific purposes, such as comparisons between different patient groups, and in defining mechanistic pathways involved in the pathogenesis of cancer and other pathological conditions.
Highlights
The involvement and importance of the immune system in cancer and its treatment have righteously and been acknowledged as an essential factor, playing a decisive role on the curability, prognosis and outcome of this disease
Considering the high sensitivity and reproducibility of real-time RT-qPCR, we suggest that cytokine mRNA profiles could be used as a proxy for protein expression for some specific purposes, such as comparisons between different patient groups, and in defining mechanistic pathways involved in the pathogenesis of cancer and other pathological conditions
Despite the fact that the biological effects of cytokines are driven by their protein/peptide form, the assessment of cytokine mRNA expression signatures for defining functional cytokine pathways of pathogenic importance has gained a wide use.[6,7,8,9,10,11,12]
Summary
The involvement and importance of the immune system in cancer and its treatment have righteously and been acknowledged as an essential factor, playing a decisive role on the curability, prognosis and outcome of this disease. Real-time quantitative RT-PCR is one of the methods often used as a proxy to cytokine protein analyses.[6,7,8,9,10,11,12] It has been an important technique in identifying different diagnostic tests and predicting outcomes for different malignant and non-malignant diseases.[13] Recently, we used this method to gain knowledge on the cytokine mRNA expression profile in paired ovarian cancer tissue samples and peripheral blood mononuclear cell (PBMC) samples of women suffering from HGSC.[6] Our results indicated elevated cytokine mRNA levels in the ovarian cancer microenvironment compared with non-malignant conditions and normal ovarian tissue, where inflammation, immune suppression, and promotion of T reg cells prevailed. The correlation between cytokine mRNA and protein expression in kinetic experiments has to our knowledge not been previously investigated in ovarian cancer cells
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