**1. Introduction**

About 50% of adults in the United States of America (USA), aged over 30 years, are a ffected by periodontitis, and almost 10% of the world population have a severe form of periodontal disease [1,2]. Periodontitis can be defined as a chronic inflammatory multifactorial disease caused by periodontal bacteria that determine the destruction of the tooth-supporting tissues, including alveolar bone, and which can lead to tooth loss [3]. Some observational studies during the last few decades have shown a direct and positive association between periodontitis and coronary heart disease, known also as ischemic heart disease (CAD), including myocardial infarction, stroke, and cardiovascular disease (CVD) [4,5]. More specifically, recent large cohort studies and a systematic review highlighted a positive graded association between periodontitis and increased risk of stroke and CAD [6–8].

The etiology of periodontitis comprises inflammatory and immunological processes that cause dysregulation in the host response due to the superinfection of periodontal bacteria [9]. Moreover, periodontitis has been positively associated with higher serum levels of di fferent inflammatory biomarkers, such as interleukin 6 (IL-6), IL-17, C-reactive protein, and prostaglandins [10].

vitamin C has been shown, together with some other antioxidant agents, to be an endogenous modulator of the metabolism of nitric oxide (NO) and subsequent endothelium-dependent vasodilation [11]. NO is one of the important mediators that regulate function and vasodilatation of the endothelium, because it controls the level of inflammation in the vessels, vascular tone, and cell proliferation, and it modulates the release of di fferent growth factors [12]. vitamin C has been extensively used to evaluate and predict early signs of endothelial dysfunction and CAD events [13]. A prospective study on 200 patients with heart failure showed that patients with high serum vitamin C deficiency moderated the relationship between inflammation and CAD events [14]. Furthermore, a multi-trial study on 134 subjects with CAD showed a therapeutic use of vitamins C and E against the reperfusion damage produced during angioplasty [15].

Few reports have associated periodontitis with CAD, endothelial dysfunction, and augmented the risk of CVD [16–18]. It has previously been hypothesized that several inflammatory mediators are systemically released during periodontitis, including CRP, metalloproteases, and prostaglandins, into the bloodstream and decrease the production of NO [19]. The reduced production of NO negatively impacts the vascular endothelial cells, whose impairment determines, finally, endothelial dysfunction, vasodilatation, and CAD [20,21]. Hence, this has aroused interest in assessing possible oral factors that influence and regulate endothelial changes as subclinical signs of CAD.

Previous studies have demonstrated an indirect association between high serum vitamin C and a direct association between high CRP levels and consequent endothelial damage in patients with periodontitis [22,23].

The local production of NO has an essential role in the development and progression of periodontitis. Both increases and decreases in the production of salivary NO metabolites during periodontitis in gingival tissue against periodontal bacteria and periodontal tissues have been reported to be associated with impaired endothelium-dependent vasodilatation [24]. More specifically, it has been shown that vitamin C and several antioxidants act as a competitive stimulator of the NO synthase, and that lower serum vitamin C levels have been reported in several metabolic disorders, including periodontitis [25,26].

All of these studies were performed only on serum vitamin C. To date, few studies have evaluated salivary vitamin C levels during periodontitis. Moreover, there are insu fficient data on the association of periodontitis on both serum and salivary vitamin C levels during periodontitis and CAD.

The aims of this study were to evaluate a possible association of gingival health, periodontitis, CAD, or a combination of both diseases on saliva and serum vitamin C and antioxidant levels. Moreover, the association between both saliva and serum vitamin C levels were assessed, and whether salivary or serum vitamin C levels were mediated by serum CRP in patients with periodontitis and with CAD.

#### **2. Materials and Methods**
