Joint analysis of left ventricular expression and circulating plasma levels of Omentin after myocardial ischemia

Louis A Saddic, Sarah M Nicoloro, Olga T Gupta, Michael P Czech, Joshua Gorham, Stanton K Shernan, Christine E Seidman, Jon G Seidman, Sary F Aranki, Simon C Body, Timothy P Fitzgibbons, Jochen D Muehlschlegel, Louis A Saddic, Sarah M Nicoloro, Olga T Gupta, Michael P Czech, Joshua Gorham, Stanton K Shernan, Christine E Seidman, Jon G Seidman, Sary F Aranki, Simon C Body, Timothy P Fitzgibbons, Jochen D Muehlschlegel

Abstract

Background: Omentin-1, also known as Intelectin-1 (ITLN1), is an adipokine with plasma levels associated with diabetes, obesity, and coronary artery disease. Recent studies suggest that ITLN1 can mitigate myocardial ischemic injury but the expression of ITLN1 in the heart itself has not been well characterized. The purpose of this study is to discern the relationship between the expression pattern of ITLN1 RNA in the human heart and the level of circulating ITLN1 protein in plasma from the same patients following myocardial ischemia.

Methods: A large cohort of patients (n = 140) undergoing elective cardiac surgery for aortic valve replacement were enrolled in this study. Plasma and left ventricular biopsy samples were taken at the beginning of cardiopulmonary bypass and after an average of 82 min of ischemic cross clamp time. The localization of ITLN1 in epicardial adipose tissue (EAT) was also further characterized with immunoassays and cell fate transition studies.

Results: mRNA expression of ITLN1 decreases in left ventricular tissue after acute ischemia in human patients (mean difference 280.48, p = 0.001) whereas plasma protein levels of ITLN1 increase (mean difference 5.24, p < 0.001). Immunohistochemistry localized ITLN1 to the mesothelium or visceral pericardium of EAT. Epithelial to mesenchymal transition in mesothelial cells leads to a downregulation of ITLN1 expression.

Conclusions: Myocardial injury leads to a decrease in ITLN1 expression in the heart and a corresponding increase in plasma levels. These changes may in part be due to an epithelial to mesenchymal transition of the cells that express ITLN1 following ischemia. Trial Registration Clinicaltrials.gov ID: NCT00985049.

Keywords: Adipokine; Cardiovascular; Ischemia; Omentin; RNA-seq.

Figures

Fig. 1
Fig. 1
ITLN1 expression is decreased in the left ventricle and circulating plasma levels of ITLN1 are increased following acute ischemia in the human heart. Stripcharts display the change in ITLN1 expression in left ventricular tissue samples (a) and in circulating ITLN1 protein levels following acute ischemia in the heart (b). FPKM: fragments per kilobase per million mapped reads
Fig. 2
Fig. 2
Venn diagram of differentially regulated genes between subcutaneous adipose tissue (SAT) vs. visceral adipose tissue (VAT) and epicardial adipose tissue (EAT) vs. SAT. In the SAT vs. VAT comparison, 845 genes were differentially regulated (FC > 2.0, p < 0.05). In the EAT vs. SAT comparison 188 genes were differentially regulated. 46% of these were commonly differentially regulated in VAT in comparison to SAT (middle overlapping segment). The numbers in each circle indicate the number of genes whose expression was altered; numbers in bold italic are upregulated, those in red font are contraregulated, and those in underlined plain font are downregulated in the respective comparisons. The specific genes are listed in Table 3
Fig. 3
Fig. 3
ITLN1 is specifically expressed in the stromal vascular fraction of visceral fat in humans. a Microarrays were queried for expression of ITLN1 in visceral adipose tissue (VAT) vs. subcutaneous adipose tissue (SAT) in paired samples (n = 8) (*** p < 0.001). bITLN1 expression from microarrays of the adipocyte and stromal vascular fraction (SVF) of VAT in paired samples (n = 6) (** p < 0.01). c Expression of ITLN1 from microarrays of murine SAT and VAT
Fig. 4
Fig. 4
Immunohistochemistry of ITLN1 in epicardial adipose tissue (EAT) shows enrichment within the mesothelial cell layer. Subcutaneous adipose tissue (SAT) stained for ITLN1 (a). EAT stained for ITLN1 (b) with magnification of the mesothelial cell layer of the visceral pericardium (c). Secondary antibody alone was used as a control (d). ITLN1 red, DAPI blue
Fig. 5
Fig. 5
ITLN1 expression in primary mesothelial cells is dramatically reduced following an epithelial to mesenchymal transition. a qRT-PCR for ITLN1 in mesothelial cells ± TGFβ1 and in a human adipocyte cell line (SGBS) before (D0) and after differentiation (D14) (n = 6 per group). b qRT-PCR for the adipocyte specific gene PLIN1 in mesothelial cells ± TGFβ1 and in a human adipocyte cell line (SGBS) before (D0) and after differentiation (D14)(n = 6 per group). c Expression of the epithelial cell marker CDH-1 and the mesenchymal cell markers VMAC, SNAI1, and SNAI2 before and after treatment of mesothelial cells with TGFβ1 (n = 6 per group)

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