Background
The role of inflammation in the pathogenesis and progression of chronic heart failure (CHF) is well established [
1,
2]. Typical hallmarks for the involvement of immune mechanisms in CHF pathogenesis are the activation/migration of leukocytes from the circulation to the areas of myocardial inflammation and an increased expression of proinflammatory cytokines, such as tumor necrosis factor α, interleukin-1, and interleukin-6 from a damaged myocardium [
3‐
6]. In heart failure such cytokine levels increase in association with disease severity and may contribute to impair cardiac function through cardiomyocyte apoptosis, inflammatory response, cardiac hypertrophy and matrix metalloproteinase activation [
1,
7‐
9].
Proinflammatory cytokines may cause myocyte apoptosis and necrosis; interleukin-6 induces a hypertrophic response in myocytes [
10], whereas TNF-α causes left ventricular dilatation, apparently through the activation of matrix metalloproteinases [
11]. Elevated levels of CXC and C-C chemokines such as GROα, IL-8, MCP-1, RANTES and MIP-1α were also found in the most severe cases of heart failure, which would suggest a possible role for such molecules in CHF progression [
12,
13]. Peripheral blood mononuclear cells are important players in the CHF inflammatory process [
14,
15]. However, the mechanisms that, in heart disease, ultimately cause the transition from cardiac disease to heart failure still need further investigation.
The identification of the cytokines associated with chronic heart failure is not only important to get deeper insights into the inflammatory pathways involved, but it could also lead to the identification of disease's biomarkers. The biomarkers found in heart failure include BNP, NT pro-BNP, C-reactive protein, tumor necrosis factor α, interleukins 1 and 6, TNF-α, matrix metalloproteinases, neuroepinephrine, renin, galectin-3 [
16‐
18], some of which are also important tools in the diagnosis and pathogenesis of heart failure, in the identification of subjects at risk of heart failure, risk stratification, therapeutic agents response monitoring [
17].
The aim of this study was to investigate the inflammatory pathways associated with CHF, and to assess whether disease etiology affects cytokine cascades. To this end, we analyzed plasma cytokine levels in patients with stable ischaemic or non ischaemic dilated cardiomyopathy CHF vs healthy donors, by a multiplex cytokine/growth factor immunoassay, and then assessed their relationship to echocardiographic parameters and disease duration.
Discussion
The aim of this study was to further our understanding of the inflammatory pathways associated with CHF and to assess whether disease etiology affects cytokine cascades. To this end we analyzed several cytokines and growth factors' plasma levels with the XMAP technology. This tecnique has previously been used in cardiovascular research, specifically, in coronary artery disease, to evaluate selected cytokine ability in predicting long-term prognosis [
23], and in Primary Graft Dysfunction [
24].
In our study, we described two different modulation patterns, i.e. increased or decreased cytokine plasma levels, in chronic heart failure (ICM and NIDCM) patients as compared to controls. The first group was composed of MIP-1 β, MCP-1, IL-8, VEGF and IL-9, that were increased in both the ICM and NIDCM patients vs controls.
The second group was composed of IL-5, IL-7, IFN-γ, which showed lower plasma levels in both the ICM and NIDCM patients as compared to controls. Furthermore, IL-6 increased only in ICM while IL-1 β decreased only in NIDCM, both changes with respect to controls. Previous study results on IL-1 β systemic level changes in heart failure have been inconsistent and reported as either unchanged [
25], increased [
26], or decreased (present study) in such patients.
Several previous reports have described the role of cytokines and growth factors in the development and/or progression of chronic heart failure [
1,
5,
9,
11]; further, a number of such cytokines are regarded as disease biomarkers [
17,
18]. It is worth noting that our study, for the first time, has associated IL-5, IL-7, IL-9, and IFN-γ plasma levels with chronic heart failure.
It has been reported that IL-5 cardiac protein levels increase four days after Myocardial Infarction (MI) in a mouse model [
27]; on the contrary, as yet no data is available on IL-5 serum or plasma levels in chronic heart failure patients. IL-7 is produced by stromal cells in lymphoid tissues and is required for T cell development and persistence in the periphery. Interestingly, local production of IL-7 has been associated with the maintenance and predominance of CD8+ T cells, which cause tissue damage in human chronic Chagas' disease cardiomyopathy, an inflammatory-dilated cardiomyopathy [
28]. IL-9 is a mostly T-cell produced cytokine that has a functional role in allergic disease and resistance to intestinal nematodes [
29], but no data is yet available on its role in heart disease. IFN-γ is an important pro-inflammatory cytokine produced by Th1 cells, that increases the expression of MHC class I and class II molecules. An increase in the percentage of IFN-γ-positive CD4 (+) T cells, has previously been described in patients with CHF [
30]. Furthermore, an increase in IFN γ serum levels has also been described [
31], in a mouse model, one week after MI, i.e. not in chronic heart failure experimental conditions.
Our study recorded decreased IL-5 plasma levels in both ICM and NIDCM groups vs controls. Interestingly, IL-5 plasma levels inversely correlated with disease duration in NYHA III/IV ICM patients. In fact, patients with longer disease duration had smaller concentrations of plasmatic IL-5 in comparison with patients more recently diagnosed. Furthermore, a statistically significant correlation with IL-5 plasma levels was found only among NYHA III/IV ICM patients, but not among patients with less severe chronic heart failure. This would suggest that decreased levels of this cytokine are associated with disease progression.
We here showed an increase in IL-9 levels in both ICM and NIDCM groups vs controls and that IL-9 levels inversely correlated with ejection fraction in ICM patients, but not in NIDCM patients. This would suggest that increased levels of this cytokine are associated with left ventricular dysfunction and, thus, with disease progression toward organ failure in ICM patients. Further, we reported a decrease in plasma IFN-γ levels and of IL-7 in both ICM and NIDCM CHF patients vs controls, but no correlation with echocardiographic parameters or duration of disease was found (not shown).
Nevertheless, our conclusions cannot broadly apply to the entire chronic heart failure population as only a small number of women were enrolled and analysed in both the ICM and NIDCM groups.
Interestingly, the cytokines that were identified in the present study are part of a gene network of expression/function regulation that link them to Norepinephrine, Endothelin-1, IL-1A, Galectin-3, TNFα, IL-6 and IL-18 [
13], which are molecules associated with chronic heart failure [
1,
4,
7,
10,
11,
17,
18]; thus we hypothesize that IL-5, IL-7, IL-9 and IFN-γ are part of signaling cascades that contribute to the development and/or progression of chronic heart failure.
Notwithstanding the ICM and NIDCM difference in pathogenetic causes there was a similar cytokine expression pattern when compared to controls. In fact, among all cytokines previously reported as being associated with CHF, our study showed only IL-6 and IL-1 β modification to be associated uniquely with ICM or NIDCM, respectively. All the other cytokines instead similarly increased or decreased, vs controls, regardless of the etiology. Further, we found that IL-1Ra was able to discriminate between the two patient groups, since it was statistically higher in the ICM group, although only a small increase was observed (data not shown).
In conclusion, the novelty in our study results is the increase of IL-9 and decrease of IL-5, IL-7, and IFN-γ plasma levels, in both ICM and NIDCM CHF patients vs controls and the inverse correlation of IL-5 and IL-9 plasma levels with duration of disease and LVEF, respectively, in ICM patients. Further studies are needed to assess whether the modification of IL-5, IL-7, IL-9 and IFN-γ plasma levels has a null, a protective, or a pathogenetic role in chronic heart failure and may constitute a potential target for therapeutic intervention.
Competing interests
The authors declare that they have no competing interests.
This article conforms to ethics in the authorship and publishing of scientific articles [
32].
Authors' contributions
MN conceived the study design, coordinated the study and drafted the manuscript. CC isolated plasmas, performed statistical analysis and contributed to drafting the manuscript. LDV collected the clinical data of patients and performed statistical analysis. FC, GL, MCC and RM critically revised the manuscript's draft. GM performed statistical analysis, LS enrolled patients. EC and AF performed and interpreted the multiplex assay. All authors read and approved the final manuscript.