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Published on: August 1, 2016
Leukemia Inhibitory Factor and Interleukin-6 downregulate sarcoplasmic reticulum Ca2+ ATPase (SERCA2) in cardiac
S Villegas1, F J Villarreal, W H Dillmann
1University of California, San Diego, La Jolla 92103-0618, USA.
Insights
Cytokines Leukemia Inhibitory Factor (LIF) and Interleukin-6 (IL-6) significantly downregulate sarcoplasmic reticulum calcium ATPase (SERCA2) gene and protein expression in cardiac myocytes. This suggests a transcriptional mechanism contributing to cardiac hypertrophy and heart failure.
Area of Science:
- Cardiovascular Biology
- Molecular Cardiology
- Cellular Signaling
Background:
- Cardiac hypertrophy and heart failure are associated with altered gene expression, notably reduced sarcoplasmic reticulum calcium ATPase (SERCA2).
- Elevated cytokines, including Leukemia Inhibitory Factor (LIF) and Interleukin-6 (IL-6), can induce hypertrophic responses in cardiac cells.
Purpose of the Study:
- To investigate the impact of LIF and IL-6 on the regulation of SERCA2 levels in cardiac myocytes.
- To elucidate the molecular mechanisms underlying cytokine-mediated effects on SERCA2 expression.
Main Methods:
- Neonatal rat ventricular myocytes were transfected with a SERCA2 promoter-reporter construct.
- Cells were treated with varying concentrations and durations of LIF and IL-6.
- SERCA2 promoter activity, mRNA, and protein levels were assessed using reporter assays, Northern, and Western blot analyses, respectively.
Main Results:
- LIF and IL-6 significantly inhibited SERCA2 promoter activity by 23-36%.
- Both cytokines reduced SERCA2 mRNA levels in a time- and dose-dependent manner, with maximal effects at 48 hours.
- Western blot analysis revealed a significant 60% decrease in total SERCA2 protein levels following cytokine treatment.
Conclusions:
- The cytokines LIF and IL-6 downregulate SERCA2 gene expression and protein levels in cardiac myocytes.
- The observed downregulation of SERCA2 by these cytokines is, at least partly, mediated at the transcriptional level.
- These findings provide insight into the molecular pathways linking inflammation and cardiac dysfunction.
Abstract:
Alterations in gene expression are a hallmark of cardiac hypertrophy and heart failure. Among these, the decreased expression of the sarcoplasmic reticulum calcium ATPase (SERCA2) has been described. Elevated levels of cytokines in particular, Leukemia Inhibitory Factor (LIF) and Interleukin-6 (IL-6) have been shown to have the capacity to elicit hypertrophic responses in cultured cardiac myocytes. In this study, we investigated the effects of these cytokines (LIF & IL-6) on the regulation of SERCA2 levels in cardiac myocytes. Cultured neonatal rat ventricular myocytes were transfected with a 3.2 kb promoter plasmid construct containing the SERCA2 promoter linked to a chloramphenicol acetyltransferase (CAT) reporter gene, and subsequently treated with 10 ng/ml LIF or 10 ng/ml IL-6. LIF and IL-6 independently caused a significant (p < or = 0.05) 23-36% inhibition in SERCA2 promoter activity. LIF and IL-6 induced inhibition was also evident in SERCA2 mRNA levels as assessed by Northern analysis. Time course of inhibition of SERCA2 mRNA levels showed the most prominent decrease occurring after 48 hours of treatment, with both cytokines having a dose dependent effect on the inhibitory response. Western analysis using a polyclonal antibody to SERCA2 protein indicate a significant, 60% decrease in the amount of total SERCA2 protein in cultured myocytes treated with 10 ng/ml LIF or IL-6. In conclusion, the cytokines LIF and IL-6 downregulate SERCA2 gene expression and protein levels. The molecular mechanism responsible for cytokine induced downregulation of SERCA2 is at least partly transcriptional.
