INTERRELATIONS BETWEEN GROWTH DIFFERENTIATION FACTOR 15, P-SELECTIN AND GALECTIN-3 AND CLINICAL COURSE IN PATIENTS WITH ARTERIAL HYPERTENSION AND TYPE 2 DIABETES MELLITUS

The aim of our study was to determine the base levels of Growth Differentiation Factor 15, P-selectin and Galectin-3 in blood plasma in patients with AH and T2DM and to assess their association with the diseases clinical course. Materials and methods. A total of 121 patients were included in our study (60 female and 61 male, mean age 64.7± ±10.6 years, with AH and/or T2DM). Patients were divided into three groups: 1st group with AH only (51 patient), 2nd group with AH and T2DM (57 patients) and 3rd group with T2DM only (13 patients, control group). GDF-15, Galectin-3 and P-selectin tests were performed using standard enzyme-linked immunosorbent assay kits (ELISA). Results. Compared with AH without T2DM and T2DM only groups, AH with T2DM group had a statistically significant higher level of GDF-15. Grade 3 hypertension group had a significantly lower level of GDF-15 compared with Grade 1&2 hypertension groups. P-selectin mean level was significantly higher in Grade 3 hypertension group GDF-15 compared with Grade 1&2 hypertension groups. We observed weak correlation between Galectin-3 and GDF-15 in blood plasma, which was confirmed by linear regression analysis. Conclusions. A combination of hypertension and type 2 diabetes mellitus revealed a significant increase of GDF-15 levels in compare with patients with only hypertension or type 2 diabetes mellitus, which may be due to a greater response to oxidative stress and low-intensity systemic inflammation. P-selectin mean level was higher in patients with grade 3 hypertension, which reflects a greater platelet activation as a part of the systemic inflammatory response. Galectin-3 mean level was higher in patients with stage 3 hypertension compared with patients with stages 1 and 2 due to possibly more pronounced fibrosis progression.


Introduction
For decades, one of the most popular research fields in medicine is the impact of low-grade systemic inflammation in patients with arterial hypertension and type 2 diabetes mellitus, which leads to atherosclerosis and atherothrombosis. One of the major topics to be investigated in this field is finding the novel inflammatory markers, which will have greater prognostic significance in cardiovascular events and evaluation of treatment effectiveness.
In order to confirm significant difference between groups we performed ANOVA test ( Table 2). GDF-15 had a significant difference between different groups. There were no statistically significant differences of other marker levels between groups. We analysed inflammatory marker levels according to hypertension grade classification [8] ( Table 3). Grade 3 hypertension group had a significantly higher level of GDF-15 compared with Grade 1&2 hypertension groups. P-selectin mean level was significantly higher in Grade 3 hypertension group GDF-15 compared with Grade 1&2 hypertension groups. There were no statistically significant differences in Galectin-3 levels between groups.
In order to confirm significant difference between different stages we performed ANOVA test ( Table 5). According to ANOVA test, Galectin-3 had a significant difference between different hypertension stages (F=3.398; p=0.037). There were no statistically significant differences of other marker levels between groups.

Discussion
Correlation and regression analysis between inflammatory markers showed a weak relationship between galectin-3 and RDF-15 levels in the plasma and no significant correlations with P-selectin and standard systemic inflammation marker hs-CRP. We consider that our results create the prerequisites for searching an additional diagnostic and prognostic information.
In general, GDF-15 levels were significantly higher in patients with group of patients with hypertension and type 2 diabetes. Other studies have shown that GDF-15 activates atherosclerotic injury through interleukin-6 (IL-6) pro-inflammatory mechanisms [11,12].
For increasing GDF-15 levels, it is necessary to activate a promoter with 2 p53 transcription factor binding sites, which is the fundamental response of cells to inflammation, oxidative stress, and oncogenic activation [13,14]. It is interesting to note that we detected maximum expression of GDF-15 in patients with hypertension and type 2 diabetes due to, probably, a greater damage to cells, which is associated with oxidative stress and low-intensity systemic inflammation. We also revealed a significant increase of P-selectin levels in patients with grade 3 hypertension. It is known that P-selectin, which is released on activated platelets and endothelial cells, can act as a counter ligand for GPIbα and, therefore, may mediate platelet adhesion and thrombus formation [15,16]. We suggest that the increase of P-selectin level reflects the activation of the platelet link of systemic inflammation in patients with grade 3 hypertension.
An increase of Galectin-3 level is primarily associated with the fibrosis progression as well as the development of heart failure [17,18] and diabetes mellitus [19,20]. The revealed increase in Galectin-3 in patients with stage 3 hypertension may be associated with the development of fibrosis and the initial stages of heart failure.
Study limitations. First, while the total sample size in our study was relatively large, the sample size of patients only with T2DM was small due to low frequency of patients without AH in this age.
Second, some plasma samples were stored more than 6 months. On this basis, we conclude that the new inflammatory markers GDF-15, P-selectin and Galectin-3 reflect different pathogenetic mechanisms activation which are responsible for low-intensity systemic inflammation in patients with hypertension and type 2 diabetes and may provide various new diagnostic and prognostic information.
Further studies with larger sample sizes are necessary to confirm our findings. 2. P-selectin mean level was higher in patients with grade 3 hypertension (grade 3 -133.95±28.13 Normal BP -100.50±42.73 ng/mL; р<0.05), which reflects a greater platelet activation as a part of the systemic inflammatory response.
3. Galectin-3 mean level was higher in patients with stage 3 hypertension compared with patients with stages 1 and 2 (13.31±5.52; 11.25±3.40 and 9.85±3.86 ng/mL; р<0.05) due to possibly more pronounced fibrosis progression. 4. A weak correlation between GDF-15 and Galectin-3 was revealed (r=0.260, p=0.022). There was no significant correlation between P-selectin and Galectin-3 as with the standard marker of inflammation hs-CRP which reflect the activation of various pathogenetic mechanisms of low-intensity systemic inflammation in patients with hypertension and type 2 diabetes. Therefore, this information creates the prerequisites for individualizing therapeutic goals.

Conflict of interests
The authors declare that they have no conflicts of interest.