C-type natriuretic peptide (CNP) is released abluminally by human vascular endothelial cells to effect relaxation of smooth muscle cells (SMC) and is important in the physiological maintenance of vascular tone. However, it is not known whether other human vascular cell types have the capacity to produce CNP. The aim of this study was to establish whether or not human vascular smooth muscle cells express CNP.

Vascular specimens were obtained from coronary artery bypass graft patients, from whom consent was obtained, and for which local Ethics Committee approval was granted. CNP expression was examined in cultured human vascular SMCs by RT-PCR, compared against -smooth muscle actin expression, and CNP protein was probed using immunohistochemical (IHC) and immunocytochemical (ICC) staining, in transverse sections (0.8 µm thickness) of human saphenous vein (SV), internal mammary artery (IMA) and radial artery (RA), and in SMCs cultured from these vessels, respectively. A negative control (CNP knockout mouse vs wild type) for CNP expression by PCR was established using RNA extracted from mouse myocardium.

Results: Expression of CNP by RT-PCR was demonstrated in SMCs derived from SV(n=3), IMA (n=2) and RA (n=3), which produced a PCR product 334bp in length, the same size as the product produced from the positive control (endothelial cells; HUVEC), although the intensity of the CNP product was markedly less in SMCs than in HUVECs. Myocardium from CNP knockout mouse did not express CNP, while expression was positive in tissue from the wild type mouse. Expression of -smooth muscle actin was similar in both positive and negative controls. Confirmation of the PCR results was provided by detection of CNP protein ICC in 100% of SMCs derived from SV (n=6), IMA (n=2) and RA (n=4). By IHC staining, CNP was detected strongly in endothelium, and to a lesser degree throughout the medial layer of whole vessel sections of the three vessel types (n=2 for each). No CNP staining was detected in adventitia.

As far as we are aware this is the first report of expression of CNP in at gene and protein level by human vascular SMCs. These findings may have implications for synthesis of CNP in endothelial dysfunction.