*3.3. Role of Sclerostin*

Sclerostin, a protein produced by osteocytes, and coded by the SOST gene on chromosome 17q12-q21, is an inhibitor of wingless-type mouse mammary tumor virus integration site (Wnt) pathway in osteoblasts, which is responsible for osteoblastogenesis [123]. In this way, sclerostin inhibits bone formation in a healthy state. Although previously described to be secreted as 27 kDa monomer only by osteocytes [124,125], later research pointed to the secretion also by other cells (osteoblasts, osteoclasts, chondrocytes, cementocytes) [126,127]. Interestingly, the SOST gene is found to be also expressed in other tissues and organs and, besides bone, primarily in heart, lung, aorta, and kidney [128,129]. Based on these data, sclerostin was no longer considered to be a bone-specific protein and marker of bone turnover, but the topic of further research aiming to understand its role in extraosseal tissues and organs. Unfortunately, the exact nature of sclerostin in those are not fully understood, neither in health, nor in a disease. Some of the limiting factors are the weak association between protein expression in the tissue and mRNA levels and different nature of sclerostin in different parts of the same tissue [130].

Clinical data on the association of sclerostin with CV risk and mortality are not very clear. It is known that patients with CKD have increased serum sclerostin levels already from the initial stages [131]. As the SOST gene is present in the heart and vascular tissue, the potential association of serum sclerostin with increased CV risk in CKD patients has also been investigated and is still an important topic in experimental and clinical research. However, compared to the studies investigating the association of FGF23 and klotho with LVH, most studies linked sclerostin with the presence of atherosclerosis and VC in CKD. Studies investigating the heart in CKD referred to the relationship between sclerostin and valvular calcification. In addition, sclerostin may exert an indirect e ffect on heart disease in CKD, by taking part in VC development and, hence, through increased peripheral vascular resistance and heart failure.

Elevated serum sclerostin levels were seen in patients with aortic valve calcification with increased upregulation of sclerostin mRNA [132]. Sclerostin is shown to be an independent risk factor for heart valve calcification in patients with CKD stages 3–5 and is increased in serum before the increase in serum phosphate and PTH is seen [133]. In addition, in patients with CKD stages 2–5, serum sclerostin was reported to be associated with inflammation markers, phosphate, FGF23, indoxylsulphate and p-cresyl sulphate, β2-microglobulin, and arterial sti ffness [134], emphasizing its role in CKD-MBD development.

High sclerostin serum levels (>200 pg/mL) were reported to be associated with increased carotid-femoral pulse wave velocity (>9.5 m/s) in HD patients [135]. Although during 2-year follow-up HD patients who died had higher sclerostin levels, sclerostin did not predict survival [136]. Similarly, to this study, it was reported that higher CV risk in HD patients was associated with sclerostin values above the median (>84pmol/L) during the five-year follow up period [137].

Recent experimental data sugges<sup>t</sup> a positive correlation between the presence of VC in CKD rats and vascular Wnt3a and β-catenin expression together with blood pressure variability, but no association with sclerostin was seen [138]. In CKD patients, sclerostin was positively associated with VC (coronary arteries and thoracic aorta, but not with those at the aortic or mitral valves and it did not predict cardiovascular events) [139]. Meta-analysis performed by Kanbay et al. showed that serum sclerostin was not associated with all-cause and CV mortality [140]. Previously, it has been shown that serum sclerostin values were associated with fatal and nonfatal CV events in non-dialysis CKD patients [141]. On the other hand, the NECOSAD study indicated that incident dialysis patients with higher sclerostin level had better CV survival [142].

Up to now, there are some data suggesting the association of serum sclerostin with vascular and valvular calcification in CKD patients and the number of studies is very scarce with conflicting results. On the other hand, data on the potential relationship of sclerostin with uremic cardiomyopathy are lacking. Taken together, clinical studies on the role of sclerostin in CKD report inconclusive data and the exact role of sclerostin in CKD-MBD and CV risk is ye<sup>t</sup> not clear with a need for further investigation.

At present, it cannot be clearly stated whether serum sclerostin turns into a toxin in CKD and increases CV risk and mortality, or if it is only a marker of disturbed bone and (cardio)vascular and valvular metabolism. The critical point here is the ability to confirm the origin of high serum sclerostin levels and then to explain the reason for such increased values. Similarly, CKD-MBD treatment for reducing sclerostin levels is a double-edged sword. Although it has been shown that the application of anti-sclerostin antibodies improves bone and mineral density and reduces fracture risk in osteoporosis [143], there is also important data indicating that such treatment can increase CV risk in patients with primary osteoporosis [144].

Nevertheless, new studies on this topic should reveal the real physiological and pathophysiological roles of sclerostin in heart and vascular disease in patients with CKD and will direct future therapeutic strategies.

### *3.4. Role of OPG-RANK-RANKL System in CKD-MBD*

Bone disease is an important component of CKD-MBD, that is linked to vascular disease and described as a calcification paradox [145] (depicted in Figure 3).

**Figure 3.** Mechanism of increased mortality in patients with chronic kidney disease. ↑ increases/increased; ↓ decreases/decreased.

The disturbed OPG-RANK-RANKL pathway might be one of the contributors to bone disease and VC development in CKD. In physiological conditions, osteoprotegerin (OPG) is a protein which inhibits activation and differentiation of osteoclasts by blocking the binding of receptor activator of nuclear factor kappa–B ligand (RANKL) to RANK expressed on osteoclast precursors [145]. It has been shown that osteoprotegerin is produced by the arterial wall and other tissues [146].

Experimental data indicate that OPG knockout in mice is responsible for osteoporosis and VC development [147]. Moreover, OPG knockout mice displays higher RANKL and RANK levels, as well as OPG downregulation detected in calcified human arteries [148,149]. An important mediator of the opposite OPG-RANK-RANKL system regulation in bone and vasculature might be TGF-β, as it increases the OPG/RANKL ratio in bone and decreases in vasculature, disabling the VC inhibition by OPG [145]. Clinical data showed that coronary artery calcification score correlated positively with serum osteoprotegerin and negatively with RANKL, and serum osteoprotegerin correlated positively with the progression of coronary artery calcification score in hemodialysis patients [150].

Nevertheless, the calcification paradox seems to be very complex and most likely disturbs several pathways deserving more detailed experimental and clinical explanation.
