Up-regulation of a hydrogen peroxide-responsive pre-mRNA binding protein in atherosclerosis and intimal hyperplasia

Abstract

Background: Multiple lines of investigation have implicated hydrogen peroxide (H(2)O(2)) as an important endogenous mediator of cell proliferation in the vessel wall. Heterogeneous nuclear ribonucleoprotein C (hnRNP-C), a nuclear pre-mRNA binding protein that plays roles in vertebrate cell proliferation and differentiation, has been identified as a component of a vascular cell signaling pathway activated by low physiologic levels of H(2)O(2). The expression of hnRNP-C in human arteries has not previously been assessed.

Methods: Segments of human proximal internal carotid arteries were evaluated for the expression of hnRNP-C by immunohistochemistry.

Results: In normal proximal internal carotid arteries, hnRNP-C is expressed predominantly by the endothelium, with significantly lower expression by medial smooth muscle. In preatherosclerotic intimal hyperplasia, hnRNP-C is up-regulated in the artery wall, due to the robust expression by the intimal smooth muscle cells, without up-regulation in the medial smooth muscle cells. In arteries with atherosclerotic lesions, there is strong expression of hnRNP-C not only by intimal cells but also by medial smooth muscle cells.

Conclusions: The H(2)O(2) responsive pre-mRNA binding protein hnRNP-C is up-regulated in atherosclerosis and in preatherosclerotic intimal hyperplasia in humans, supporting the hypothesis that H(2)O(2) is a regulator of vascular cell proliferation in these conditions. These data also suggest that hnRNP-C may be useful as a marker of vascular cell activation.

Fig. 1

Normal artery. Shown are histologic sections (400× magnification) stained with (A) hematoxylin and eosin, or with immunohistochemical stains for (B) hnRNP-C, (C) smooth muscle actin, and (D) the endothelial marker CD31. The arrows indicate the intima / media boundaries. Staining for hnRNP-C is markedly enhanced in endothelial nuclei compared with medial smooth muscle cells.

Fig. 2

Artery with intimal hyperplasia. Shown are histologic sections (200× magnification) stained with (A) hematoxylin and eosin, or with immunohistochemical stains for (B) hnRNP-C, (C) smooth muscle actin, and (D) the macrophage marker CD68. The arrows indicate the intima / media boundaries. The arrowhead indicates a CD68⁺ macrophage. Staining for hnRNP-C is markedly enhanced in endothelium and intimal smooth muscle cells compared with medial smooth muscle cells.

Fig. 3

The intima in atherosclerosis. Shown are histologic sections (200× magnification) of the intima of an artery with a Type IV atheroma stained with (A) hematoxylin and eosin, or with immunohistochemical stains for (B) hnRNP-C, (C) smooth muscle actin, and (D) the macrophage marker CD68. The necrotic / lipid core is present in the lower left corner (*). There is strong staining of endothelial cells, intimal smooth muscle cells (arrows) and macrophages (arrowheads) for hnRNP-C.

Fig. 4

The media in atherosclerosis. Shown are histologic sections (200× magnification) of the media of an artery with a Type IV atheroma stained with (A) hematoxylin and eosin, or with immunohistochemical stains for (B) hnRNP-C, (C) smooth muscle actin, and (D) the macrophage marker CD68. The arrows indicate the intima / media boundaries. The necrotic / lipid core is present in the upper left corner (*). There is strong staining of both intimal and medial smooth muscle cells for hnRNP-C.

Fig. 5

Quantitation of hnRNP-C upregulation. Shown is bar graph indicating the percentage of nuclei staining positively for hnRNP-C in each lesion type: (N) normal, (IH) intimal hyperplasia, and (A) atherosclerosis. Data have been plotted both combining the intima and media (intima + media) and for the intima and media separately. Data were assessed by ANOVA: * P < 0.05 versus normal artery intima+media, ** P < 0.001 versus both normal artery intima+media and intmal hyperplasia intima+media, *** P < 0.001 versus both the media of normal arteries and the media of arteries with intimal hyperplasia.