CRP was shown to be the most studied biomarker of cardiovascular pathologies, particularly atherosclerosis and ischemic heart diseases [8]. However, there is still ongoing discussion about the possible role of CRP as a possible bystander or inducer of cardiovascular and metabolic alterations [13]. Recently, human CRP transgenic SHR-CRP rats were introduced to assess CRP’s role in the pathogenesis of metabolic syndrome [13]. Indeed, the presence of human CRP in these rats resulted in higher blood pressure, glucose levels, oxidative stress, and inflammation compared to non-transgenic animals [13]. However, until now, no study has been conducted to investigate the potential effects of CRP in the aorta.

Thus, we hypothesized that CRP affects the protein expression of biomarkers of endothelial dysfunction, inflammation, and oxidative stress in the aorta.

In this study, the presence of human CRP was associated with significantly decreased insulin-stimulated glycogenesis in skeletal muscle, increased insulin plasma levels, increased muscle and hepatic accumulation of TAG and decreased plasmatic cGMP concentrations, reduced adiponectin levels and increased monocyte chemoattractant protein-1 (MCP-1) levels in the blood, suggesting pro-inflammatory effects and presence of multiple features of metabolic syndrome in SHR-CRP animals as was partially shown previously [13]. Indeed, adiponectin is an important insulin-sensitizing adipokine expressed almost exclusively in adipose tissue and has also been reported to exert anti-diabetic, anti-atherosclerotic, anti-inflammatory, and cardioprotective actions. It has been shown that decreased levels are found in people with cardiovascular disease. Several studies showed that the weight and metabolic activity of visceral adipose tissue are important and may be related to reduced adiponectin concentrations [18]. However, we did not detect significant changes in these parameters, including the relative weight of epididymal adipose tissue, the relative weight of perirenal adipose tissue, and NEFA between SHR and SHR-CRP rats. Interestingly, the changes in adiponectin levels were associated with inflammation and oxidative stress, which was increased in the aorta in this study as well.

Indeed, adiponectin mediates several functions in vascular endothelium, e.g., production of NO via phosphorylation of eNOS at Ser1177 or attenuation of production of ROS [18, 19], suggesting that reduced levels of adiponectin might also be related to NO levels in aorta. Thus, we studied the expression of proteins related to the proper function of vascular endothelium. eNOS and its phosphorylated form p-eNOS (phosphorylated on Ser1177) are necessary for the proper production of NO and the regular function of the endothelium. It was demonstrated that CRP reduced the expression of eNOS (both gene and protein) in vitro in human aortic endothelial cells (HAECs), suggesting the direct participation of CRP in the development of endothelial dysfunction [6]. Furthermore, the reverse association of CRP and eNOS activity has been shown in rats after intraperitoneal administration of human CRP [5]. Western blot analysis demonstrated a significant decrease in the p-eNOS/eNOS protein expression, which is a typical hallmark of alteration of endothelial function. Interestingly, it was shown that ENG supports eNOS activity and expression, thus, it might be involved in endothelial protection [20], and its reduced expression was related to the aggravation of endothelial function [21]. However, western blot analysis conducted in the present study did not reveal significant changes in ENG expression between SHR-CRP and SHR, which suggests that ENG is not related to eNOS expression in these rats. It is of interest to mention that ENG expression in rat aorta has not been studied so far when compared to mice. Therefore, we cannot exclude the possibility of the different role of ENG in rat endothelial function.

The inflammation represents the hallmark of endothelial dysfunction, and it is characterized by the increased activity and expression of NFkB, which regulates the expression of cell adhesion molecules involved in the transmigration of leukocytes in early endothelial dysfunction [22]. Indeed, CRP was also shown to increase the expression of cell adhesion molecules in HAECs [6]. However, the presence of human CRP in SHR in this study did not affect NFkB, P-selectin, and COX2 expression in the aorta, suggesting that cell adhesion molecules and inflammation are not induced by the presence of CRP in the aorta of SHR-CRP rats.

The presence of CRP is also related to oxidative stress, as published previously [13]. Thus, we focused on oxidative stress-related biomarkers in rat aorta. HO-1 is a highly inducible vascular protective enzyme activated by various stimuli, including oxidative stress (e.g., LDL oxidation) and inflammation (e.g., TNF-α) [23]. Its expression represents the hallmark of oxidative stress, however, it may also be responsible for the protection [11]. We found significantly increased expression of HO-1 in rat aorta, suggesting the development of oxidative stress in SHR-CRP rats. In addition, the expression of extracellular SOD3 was reduced in SHR-CRP. It was proposed that SOD3, in cooperation with catalase, improves endothelial-dependent vasodilatation in various conditions by protecting the NO-mediated signaling, and restoration of SOD3 is necessary for the correction of vascular structure and function [24,25,26]. Thus, reduced expression of important superoxide scavenger SOD3 and increased HO-1 expression suggests oxidative stress induction in SHR-CRP rats.

Our study demonstrated, for the first time, that CRP is only partially able to affect biomarkers of endothelial dysfunction in SHR rats, which are genetically predisposed to the development of hypertension and metabolic syndrome. Thus, we suggest that CRP alone cannot fully induce the expression of endothelial dysfunction biomarkers, suggesting other risk factors of cardiovascular disorders are necessary to be involved in inducing endothelial dysfunction with CRP.