Beyond the conventional view of Hp as a marker of hemolysis, several findings point toward an immunomodulatory effect of Hp in B-cell mediated progression of atherosclerosis [13]. It has been demonstrated that the HP*2/*2 phenotype is associated with markedly higher peripheral B-cell counts than HP*1/*1 [14]. In contrast, B-cell percentages in bone marrow are lowest in HP*2/*2 subjects [14]. A negative correlation between serum Hp 1–1 concentration and peripheral B-cell counts was also observed [14]. The number of free CD22 binding sites on circulating B cells was estimated to be higher in HP*2/*2 individuals, who display lower serum Hp levels than those with HP*1/*1. Furthermore, the concentrations of large, multimeric proteins such as IgM and Hp 2–2 are considerably lower in extravascular fluids than in plasma, because diffusion of these macromolecules into the interstitial compartment is limited. Thus, it is conceivable that some interactions of B cells that are inhibited in the blood stream are enabled within atherosclerotic plaque of HP*2/*2 subjects.

However, it is not known whether and how the different Hp phenotypes can influence the B-T cell dialogue and T cell activation by interfering with CD22 function. Notably, higher peripheral T helper lymphocyte counts are observed in HP*2/*2 subjects [14]. However, Hp plays a direct effect on T cells through a dose-dependent suppression of induced T-cell proliferation [6]. Hp exhibits a strong inhibitory effect on T_h2 cytokine release, thus promoting a dominant T_h1 activation over T_h2 activation and playing a modulating role on the T_h1/T_h2 balance [6]. The majority of the T cells in atherosclerotic lesions produce T_h1cytokines, such as IL-2, IL-12, interferon-γ, TNF-α, and TNF-β, supporting cell-mediated responses [15]. In contrast, human atheromas contain only modest quantities of the T_h2 cytokines IL-4, IL-5, and IL-10, which can promote humoral responses [15]. Whether and how the different polymorphic Hp forms can attenuate or enhance atherogenesis by direct modulation of Th1/Th2 balance requires further investigation. Some pro-atherogenic properties related to Hb binding of Hp 2–2 can have indirect effects on B- and T-cell responses as well as antibody production against oxidized LDL. First, Hp 2–2 polymers are less efficient in the clearance of free Hb after intraplaque hemorrhage, causing higher susceptibility of LDL to Hb-driven oxidation [16]. Oxidatively modified LDL epitopes are highly immunogenic and stimulate B- and T-cell responses that can promote humoral as well as cellular immunity [17]. Second, Hb/Hp 2–2 complexes display higher affinity for CD163 on monocyte-macrophages, leading to intracellular accumulation of Hb-derived iron [18]. Such iron-loaded macrophages may, in the arterial intima, expose LDL to reactive oxygen species and thus generate an inflammatory stimulus [19]. Finally, via CD163 binding, the Hb–Hp 1–1 complex stimulates the macrophage to secrete the antiinflammatory cytokine IL-10 and heme oxygenase-1 to a markedly greater degree than the Hb–Hp 2–2 complex [19]. Release of IL-10 has been suggested to reduce inflammatory cell infiltration and macrophage accumulation in atherosclerotic plaques of Hp 1–1 mice [19]. IL-10 inhibits the T_h1 response in atheroma. In contrast, macrophages activated by Hb–Hp 2–2 binding to CD163 shift the T helper response towards T_h1 cytokines [20].

Inflammation with infiltrates of macrophages and lymphocytes is an important feature of abdominal aortic aneurysms (AAA) [21, 22]. Elevated levels of various plasma markers of inflammation have been reported in patients with AAA as compared with healthy controls or patients with cardiovascular disease [23–25]. Positive correlations between inflammatory markers and the degree of aortic dilatation have been reported in cross-sectional studies [26–29].

In this context, Hp may play a crucial role in modulation the inflammatory response characterizing AAA, both as a susceptibility factor (by its genetic polymorphism) and as a prognostic marker.

The first study on Hp’s role in AAA was performed in early 1980, when Norrgärd and co-workers found that there was an increased frequency of the HP*2/*1 phenotype in Northern Swedish individuals with AAA [30]. Following this first report, Powell et al. [31] investigated whether the different Hp phenotypes influence the degradation of aortic connective tissue. The HP*1 allele frequency was significantly increased in patients with aneurysms compared to control subjects (0.51 vs. 0.35, P  <0.05). However, HP*2/*2 patients had the highest mean age at aneurysm resection. Moreover, they found that Hps containing an α1-chain accelerated the degradation of aortic elastin by elastases two- to four-fold in vitro.

This preliminary evidence led Adamson et al. [32] to test for the usefulness of the HP polymorphism as a family-based informative genetic marker of AAA, without significant results. After this study, the role of Hp in AAA was almost ignored until 2001, when Wiernicki et al. [33] analyzed the influence of Hp phenotypes on serum elastase activity, neutrophil count, and elastin concentration in the aorta of Polish AAA (N  =  52) and aortoiliac atherosclerotic occlusive disease (AOD; N  =  37) patients. HP phenotype distribution did not differ between the two groups and the third control group of 37 subjects without atherosclerosis. Nevertheless, significant increases in serum elastase activity and neutrophil count were measured in the HP*2/*1 phenotype of AAA patients, thus supporting the association of AAA susceptibility with the HP*2/*1 phenotype as postulated by Norrgӓrd et al. [30]. Nearly 10 years later, the same research group provided further evidence for the association of the HP*2/*1 genotype with AAA [34]. They found that HP*2/*1 patients had a significantly higher growth rate [3.69 (2.40)  mm/year] of AAA compared with patients with HP*2/*2 [1.24 (0.79), P  <0.00001] and HP*1/*1 [1.45 (0.68), P  =  0.00004]. Elevated elastase serum activity was also evident in AAA patients with HP*2/*1 [0.119 (0.084) arbitrary units] in contrast to HP*2/*2 [0.064 (0.041), P  <0.00001] and HP*1/*1 [0.071 (0.040), P = 0.0006] patients. CRP serum levels (mg/l) were significantly higher in patients with HP*2/*1 (7.2 [7.1]) than with HP*2/*2 [3.4 (3.1), P  =  0.0058] and HP*1/*1 [2.8 (4.1), P = 0.044]. In 2011, Pan et al. [35] conducted another study aiming to assess the association of the HP polymorphism with AAA in the Taiwanese population. Forty-five patients with AAA and 49 non-AAA subjects were included. They found that plasma Hp concentrations were significantly higher in AAA patients compared with non-AAA subjects (254  ±  158 vs. 186  ±  108 ng/ml; P  =  0.017), in particular for HP*2/*2 carriers compared with corresponding non-AAA subjects (238  ±  144 vs. 163 ± 86 ng/ml; P  =  0.024).

Beside studies based on the HP genetic polymorphism, three reports have been conducted only at the protein level. The first was performed on a large Swedish cohort, the “Malmö Preventive Study,” including 6,075 men with information on 5 inflammation-sensitive plasma proteins (ISPs; fibrinogen, orosomucoid, α1-antitrypsin, haptoglobin, and ceruloplasmin) [36]. A total of 63 men had AAA (0.49 per 1,000 person/years). Fifty were non-fatal cases whose aneurysm was repaired in an open vascular or endovascular surgical procedure, and 13 of those 50 (26 %) were ruptured. The remaining 13 cases were fatal. The mean time from the baseline examination to aneurysm repair or death from AAA was 18.8  ±  4.9 years (range 1.3–26.6). Age at the time of the AAA was 67.1  ±  5.3 years (range: 55–80 years). The Hp level (g/l) was significantly higher in men who subsequently had AAA as compared with the controls (1.69  ±  0.79 vs. 1.38  ±  0.68, p  <0.001). Moreover, a higher Hp level conferred an increased risk of fatal AAA compared with repaired AAA (O.R  =  2.500, p  <0.010). Another study examined the Hp level after postoperative surgery in elective repair of infrarenal aortic aneurysm patients [37]; AAA patients had a significant postoperative rise in IL-10 levels and a significant decrease in plasma Hp levels. Lastly, very recently, Gamberi et al. [38] performed a comprehensive plasma proteomic profiling on eight patients scheduled for AAA repair through elective aortic reconstructive surgery, identifying a significant Hp increase in plasma from AAA patients.

Overall, the emerging picture from these studies is a striking involvement of Hp in the pathophysiology of AAA. First, the HP*2/*1 genotype seems to be an important predisposing factor of AAA liability, also influencing the prognosis. On the other side, the higher Hp level found in AAA patients is a clear demonstration of the proinflammatory nature of the AAA phenomenon. Indeed, given the antiinflammatory properties of Hp, it is conceivable that the rising of Hp level is a counteracting response to the increased inflammation due to the presence of AAA. Concerning the role of the HP*2/*1 genotype in AAA liability, the molecular and genetic complexity of the HP polymorphism needs to be taken into account. Most of the authors failed to explain why the HP*2/*1 heterozygous phenotype was different from both homozygous phenotypes and speculated that the HP*2/*1 phenotype might cumulate harmful features associated with allele HP*1 (stimulation of the elastin hydrolysis) and allele HP*2 (increased risk of atherosclerosis) to yield a combination that is particularly efficient in promoting AAA growth. The reason underlying the association of the HP*2/*1 phenotype with AAA resides in the phenomenon of molecular heterosis at the HP 1/2 polymorphism [39, 40] reported by several genetic association studies [40, 41] and further confirmed by the molecular structure of the haptoglobin protein (Fig. 8.1) [1]. Molecular heterosis occurs when subjects heterozygous for a specific genetic polymorphism show a significantly greater effect (positive heterosis) or lesser effect (negative heterosis) for a quantitative or dichotomous trait than subjects homozygous for either allele. Comings and MacMurray estimated that molecular heterosis may occur in up to 50 % of gene associations [40]. In this context, the HP 1/2 polymorphism is characterized by the production of three distinct biochemical phenotypes, each one possessing different molecular configurations (Fig. 8.1) and functions [1]. The disease-modifying effect of Hp 2–1 proteins can be related to their scarce distribution in the extravascular environment in that, being more highly polymeric proteins than Hp 1–1 dimers and displaying asymmetric structure compared with cyclic polymers produced by Hp 2–2, they are limited by their molecular mass and stereo configuration. Therefore, they have lower efficacy in both preventing oxidative tissue damage and downregulating the inflammatory processes in general. Moreover, we suggest that Hp 2–1 molecules would have lower binding to CD163 receptors and thus lower scavenging activity and angiogenic activity than Hp 2–2 molecules. They would have lower binding to hemoglobin than Hp 1–1 molecules and thus would confer a higher risk of AAA than both Hp 2–2 and Hp 1–1 molecules. Unfortunately, most of the functional studies compared Hp 1–1 with Hp 2–2, and only a few studies were applied to purified Hp 2–1 [6]. Thus, little is known about real behavior of Hp 2–1. New functional studies are clearly needed to further clarify the role of Hp 2–1 in AAA and in the inflammatory process in general.