Korelacja pomiędzy adipokinami i nieswoistymi mediatorami stanu zapalnego u pacjentów z cukrzycą typu 2 w zależności od stopnia kontroli glikemii

Alina Urbanovych, Galyna Suslyk

Department of Endocrinology, Lviv Danylo Halytskyi National Medical University, Lviv, Ukraine

 

ABSTRACT

Introduction: The relationship between glycaemic control in type 2 diabetes and the risk of its complications has been proven in many studies. However, the role of adipose tissue hormones and non-specific inflammatory mediators in type 2 diabetes compensation has not been studied completely.

The aim: To evaluate the correlation between the content of selected adipose tissue hormones and mediators of nonspecific inflammation, depending on the glycemic control in type 2 diabetes.

Materials and methods: the study has been focused on the analysis of contents and correlations between leptin, resistin, IL-2, IL-6, TNF-α in 305 patients with type 2 diabetes, who were divided into the following groups (according to their glycemic control): group 1 – with optimal glycaemic control (HbA1c ≤ 7%), group 2 – with suboptimal glycaemic control (HbA1c 7.1-8%), and group 3 – with poor glycaemic control (HbA1c ≥ 8.1%).

Results: the group of type 2 diabetic patients with poor glycaemic control showed a higher resistin level compared with the patients with optimal (+29.43%; p <0.05) and suboptimal glycaemic control (+33.45%, p < 0.05). Statistically, when comparing groups of type 2 diabetic patients with the different glycaemic control we have noticed no significant changes in the leptin concentration (all p> 0.05). The level of circulating insulin was significantly lower in the group of type 2 diabetic patients with poor glycemic control of diabetes, compared to those with suboptimal glycaemic control (-20.87%; p <0.05).

Conclusions: In patients with type 2 diabetes an impaired glycaemic control does not influence the leptin level. Though impaired glycaemic control significantly raises resistin level in the blood serum. Studying the concentration of cytokines (TNF-α, IL-2 and IL-6) in the blood serum of type 2 diabetic patients showed that glycaemic control does not provoke any significant differences in their content. The study has proved that resistin is more closely interconnected to cytokines in case of worsening the diabetes compensation – there has been found a significant positive correlation between the content of TNF-α and resistin in the group with optimal glycaemic control, then between IL-6 and resistin in the group with suboptimal glycaemic control; and in the group with unsatisfactory glycaemic control there has been detected a correlation between TNF-α, IL-2, IL-6 and resistin.

 

Wiad Lek 2018, 71, 6, -1174

 

Introduction

Recent scientific research showed that adipose tissue is an independent endocrine organ and therefore produces cytokines, adipokines, chemokines and other biologically active compounds that play an important role in the regulation of carbohydrate and lipid metabolism, non-specific inflammation and immune system disorders. It has been proven that a number of biologically active compounds of adipose tissue, including resistin, leptin, tumor necrotic factor alpha (TNF-a), interleukin-6 (IL-6), interleukin-2 (IL-2) have proinflammatory potential and are markers of acute phase response, but little is known how they associate with the level of glycemic control of type 2 diabetes [1, 2, 3].

Resistin is a low-molecular-weight protein, consisting of 108 amino acids; belongs to adipocytokines. It was first described in 2001 almost simultaneously by several scientific groups. Animal studies have shown that resistin acts as an antagonist of insulin because it inhibits insulin-mediated glucose uptake by target tissues, and is associated with obesity, insulin resistance, and type 2 diabetes.

These effects of resistin in the human body are still not finally confirmed and not refuted. Despite nowadays numerous studies of resistin, very little is known about its intracellular signalling pathways. In animals, this hormone is produced mainly by adipocytes, in humans, by monocytes, macrophages and, partly, adipocytes [4-7].

Macrophage cytokines are cells of the subpopulation Th1, which produce proinflammatory cytokines – IL-2, IL-6, TNF-α. These cytokines are mediators of inflammation and tissue destruction, they enhance cellular and inhibit humoral immunity. There is an assumption that type 2 diabetes is the result of acute phase inflammatory reactions, during which there is a release of cytokines [8].

Leptin is one of the most studied hormones of adipose tissue, there is evidence that it plays a significant role in regulating the immunological response. Despite a large number of studies, the role of leptin in systemic inflammation, the pathogenesis of type 2 diabetes and the development of complications of this disease remains unclear and still unexplored.

Leptin modulates the concentration of proinflammatory and anti-inflammatory cytokines, since it activates such inflammatory cells as macrophages, neutrophil granulocytes, and T-lymphocytes. According to the study results, leptin is associated with the synthesis and activation of acute systemic inflammation reagents – TNF-α, IL-6, IL-12, IL-10, C-reactive protein (CRP).

There is a direct correlation between the level of TNF-α, CRP, IL-6 and leptin in healthy individuals. Leptin, TNF-α, IL-6, IL-2 and other cytokines are interconnected and regulated by the same mechanisms – through PPAR (peroxisome proliferator-activated receptor) γ-activated receptors. It is likely that leptin, directly or indirectly through the immune system, may alter the activation and synthesis of cytokines that contribute significantly to the onset and progression of type 2 diabetes [9-13].

The aim

Studying the correlation between the content of selected adipose tissue hormones and nonspecific inflammatory mediators, depending on the glycemic control of type 2 diabetes, will determine their role in the pathogenesis of type 2 diabetes and its complications, which was the purpose of this work.

Materials and methods

During the study, 305 patients with type 2 diabetes have been examined. The patients were diagnosed with type 2 diabetes according to WHO criteria. Based on the glycaemic control, the patients have been divided into 3 following groups:

Group 1– optimal glycaemic control (HbA1c ≤ 7 %);

Group 2– suboptimal glycaemic control (HbA1c 7,1-8 %);

Group 3– poor glycaemic control (HbA1c ≥ 8,1 %).

All patients have overgone anthropometric and general clinical examinations. The content and interactions of leptin, resistin, IL-2, IL-6, and TNF-α in these groups of patients have been analyzed.

The level of HbA1c in the blood was determined by high-pressure cation-exchange liquid chromatography using a semi-automatic BIO-RADD-10 analyzer and BIO-RAD (USA) reagent kits.

The immunoassays (solid-phase immuno-enzyme assay method) included the determination in the blood the levels of insulin, leptin, (“ELISA”) using DRG (Germany) reagent kits; resistin (“ELISA”) using a set of reagents from “BioVendor” (Czech Republic); IL-2, IL-6, TNF-α using reagent kits from the company “Diaclone” (France) on the HumaReader Single Plus (Germany) enzyme-immunoassay analyzer.

When performing the statistical analysis of the data obtained, the following methods were used: the calculation of the mean arithmetic and its mean error (M ± m); assessment of the probability of the difference between the results obtained in the comparable groups using the Student’s and Kolmogorov-Smirnov’s criteria; correlation analysis – calculation of pair correlation coefficients. If the primary data were given in quantitative values, then the Pearson method was used; if qualitative – the method of Spearman with the following definition of the degree of its statistical significance. While comparing the groups with each other there has been an ANOVA test performed.

The clinical study was performed keeping the safety measures provided in such cases for the patient’s health, protection of his/her rights, human dignity and moral and ethical standards in accordance with the principles of the Helsinki Declaration of Human Rights, the Council of Europe Convention on Human Rights and Biomedicine, the relevant laws of Ukraine; authorization of the Bioethics Commission at Lviv Danylo Halytskyi National Medical University.

Results and discussion

Table I represents general characteristics of examined patients. The groups were compared in age, BMI, biochemical indicators. The results of this study show that the content of resistin increases with the impaired glycaemic control of diabetes: in the group of patients with poor glycemic control of diabetes, there was seen a higher level of this hormone in adipose tissue compared with patients with an optimal (+29.43%, p <0.05) and with suboptimal glycemic control (+33.45%, p <0.05).

However, there are no statistically significant changes detected in the concentration of leptin when comparing the groups (all p> 0.05). So, the content of leptin did not change statistically depending on the glycemic control of type 2 diabetes.

The level of circulating insulin was significantly lower in the group patients with poor glycemic control, compared to those with suboptimal patients (-20.87%; p <0.05) and tends to be accurate compared with optimal control of diabetes (-14.58%; p = 0.07), Table II.

Analysis of the content of cytokines in type 2 diabetic patients, depending on the state of glycemic control is shown in Table III.

It should be noted that TNF-α and IL-6 levels in the blood serum increase with impaired glycemic control, but these changes were statistically unreliable.

Correlation analysis revealed a positive correlation between the content of TNF-α and resistin in the group with optimal glycemic control (r = 0.37; p <0.05); between IL-6 and resistin in the group with suboptimal glycemic control of diabetes (r = 0.53; p <0.01) (Fig. 1).

In the group with the poor glycaemic control there has been detected a positive correlation between TNF-α and resistin (r = 0,41; p < 0,0001) (Fig. 2), between IL-2 and resistin (r = 0,29; p < 0,001), and between IL-6 and resistin (r = 0,29; p < 0,0001). No correlation between leptin levels and cytokines in any group of examined patients has been noticed.

They say that the level of resistin affects glucose level and FFA and reduces the sensitivity of tissues to insulin. The combination of these factors causes resistance to insulin [14, 15]. Numerous studies conducted in mice indicate that resistin influences glucose metabolism [16]. Researchers explain this by the fact that resistin has the function of a protein that is involved in the regulation of glucose homeostasis, and, accordingly, its chronically high concentration in the blood leads to the development of fasting hyperglycemia and glucose intolerance.

Our previous studies have found that resistin levels are higher in patients with type 2 diabetes compared to those with no carbohydrate metabolism and that the concentration of this fatty acid hormone increases with the duration of type 2 diabetes [17].

With increased resistin level, insulin-stimulated glucose uptake is disturbed, while antibodies to resistin inhibit this effect. Furthermore, the studied adipokine neutralizes the inhibitory effect of insulin on glucose production by the liver and reduces its absorption by skeletal muscles.

Accordingly, we have established a significant reduction in insulin content in the blood serum and increase in the level of resistin in type 2 diabetic patients with impaired glycemic control of diabetes. It was established that when the blood serum resistin content is lowered, homeostasis of glucose improves as a result of inhibition of gluconeogenesis in the liver. This effect is partly due to increased activity of AMP-activated protein kinase and decreased expression of enzymes in hepatic gluconeogenesis.

In the course of the study, we have noticed an increase in the levels of IL-2, IL-6, TNF-α in the blood serum of type 2 diabetic patients with impaired glycaemic control, but these data were statistically not reliable. It is known from the scientific data that the effect of cytokines results in impaired function and apoptosis induction in pancreatic β-cells [18].

The synthesis is influenced by TNF-α, secretion and activity of other cytokines; affects insulin sensitivity and stimulates lipolysis. The glucose metabolism is affected by IL-6, which is due to its action on skeletal muscle cells, adipocytes, hepatocytes, pancreatic β-cells. It is believed that elevation of IL-6 leads to suppression of GLUT4 [430], which is believed to lead to the development of hyperglycemia and IR (insulin resistance). Probably, this also concerns decompensation of type 2 diabetes.

Resistin induces translocation of the transcription factor NF-kB, which stimulates the release of proinflammatory cytokines from macrophages and monocytes [19]. In our study, we have found a positive correlation between resistin content and

non-specific inflammatory markers specifically in the group with the highest resistin level.

Conclusions

In patients with type 2 diabetes, impaired glycaemic control does not influence the leptin level. Though the resistin level is significantly raising with impaired glycaemic control.

Studying the concentration of cytokines (TNF-α, IL-2 and IL-6) in the blood serum of patients with type 2 diabetes, we have not detected any significant differences in their content depending on the glycaemic control.

The study has proved that resistin is more closely interconnected to cytokines in diabetes management – there is a significant positive correlation between the content of TNF-α and resistin in the group with optimal glycaemic control, then between IL-6 and resistin in the group with suboptimal glycaemic control; and in the group with unsatisfactory glycaemic control there is a correlation between TNF-α, IL-2, IL-6 and resistin.

References

1. Donath M.Y., Shoelson S.E. Type 2 diabetes as an inflammatory dis’ease. Nat. Rev. Immunol. 2011; 11 (2): 98—107.

2. Su S.C., Pei D., Hsieh C.H. et al. Circulating proinflammatory cytokines and adiponectin in young men with type 2 diabetes. Acta Diabetol. 2011; 48 (2): 113—119.

3. Bo S., Gambino R, Pagani A. et al. Relationships between human serum resistin, inflammatory markers and insulin resistance. Int. J Obes. 2005;29(11): 1315-1320

4. Bogkareva M., Nagaev I., Dahlber L., Resistine, an adipokine with potent inflammatory properties. J Immunol 2005; 174 (9): 5789-5795.

5. Lehrke M., Reilly MP, Millington SC et al. An inflammatory cascade leading to hiperresistinemia in humans. PloS med.2004; 1(2):45.

6. Shetty GK, Economides PA, Horton ES. Circulating adiponectin and resistin levels in relation to metabolic factors, inflammatory markers and vascular reactivity in diabetic patients and subject as risk for diabetes. Diabetes Care. 2004; 27(10):2450-2457.

7. Lauders M, Oberhauser F, Schulte DM et al. Visfatin/PBEF/Nampt and resistin expressions in circulating blood monocytes are differentially related to obesity and type 2 diabetes in humans. Horm Metab Res. 2010; 42(4): 268-273.

8. Alexandraki K, Piperi C, Kalofoutis C et al. Inflammatory process in type 2 diabetes. Annals of the New York Academy of Sciences. 2006; 1: 89-117.

9. Shoelson SE, Lee J, Goldfine AB. Inflammation and insulin resistance. J Clin Invest. 2006; 116: 1793-1801.

10. Mankovsky B., Urbanovych A. The Content of Blood Leptin and Activity of Systemic Inflammatory Response in Patients with Type 2 Diabetes Mellitus depending on Weight and Length of the Process. International Journal of Physiology and Pathophysiology. 2015; 6 (3): 213-219.

11. Urbanovych A.M. Leptin Role in the Pathogenesis of diseases, accompanied by insulin resistance. Experimental and Clinical physiology and biochemistry. 2010; 1: 57-63.

12. Tilg H, Moschen AR. Inflammatory mechanisms in the regulation of insulin resistance. Mol Med. 2008; 14(3-4): 222-231.

13. Fantuzzi G, Mazzone T. Adipose tissue and atherosclerosis: Exploring the connection. Arterioscler Thromb Vasc Biol. 2007; 27: 996-1003.

14. Cao H., Hegele R. Single nucleotide polymorphisms of the resistin (RSTN) gene. J. Hum Genet 2001; 46: 553-555.

15. Meier U., Gressner AM. Endocrine regulation of energy metabolism: rewiew of pathobiochemical and clinical chemical aspects of leptin, ghrelin, adiponectin and resistin. Clin Chem 2004; 50: 1511-1525.

16. Rangwala S., Rich A., Rhoades B. et al. Abnormal glucose homeostasis due to chronic hyperresistinemia. Diabetes 2004; 53: 1937-1941.

17. Urbanovych A. The blood resistin level in patients with DM2, depending on the duration of the disease. Current Issues in Pharmacy and Medical Sciences. 2015; 28 (2):111-114.

18. Barthson J., Germano C.M., Moore F. et al. Cytokines tumor necrosis factor-α and interferon-γ induce pancreatic β-cell apoptosis through STAT1-mediated Bim protein activation J. Biol. Chem. 2011; 286 (45): 39632-39643.

19. Lehrke M, Reilly M. P., Millington S. C. et al An inflammatory cascade leading to hiperresistinemia in humans. PloS med. 2004; 1: 2-45.

Authors’ contributions:

According to the order of the Authorship.

Conflict of interest:

The Authors declare no conflict of interest.

CORRESPONDING AUTHOR

Alina Urbanovych

Department of Endocrinology,

Lviv Danylo Halytskyi National Medical University

str. Svyentsits’koho, 3, 79011, Lviv, Ukraine

e-mail: alinaur@dr.com

Received: 10.05.2018

Accepted: 09.08.2018

Table I. Classification of type 2 diabetic patients according to their glycaemic control, (M ± m)

Indicator

Group, number of patients

1

(HbA1c ≤ 7 %),

n=52

2

(HbA1c 7,1-8 %),

n=80

3

(HbA1c ≥ 8,1 %),

n=173

F:M

29:23

40:40

85:88

Age, years

55,03±0,87

р1-2>0,05

p1-3>0,05

52,76±1,30

p2-3>0,05

53,56±0,47

BMI (body mass index), kg/m2

32,15±4,89

р1-2>0,05

p1-3>0,05

31,28±0,98

p2-3>0,05

31,55±0,48

waste/hip ratio

0,97±0,01

р1-2>0,05

p1-3>0,05

0,98±0,02

p2-3>0,05

0,99±0,01

HbA1c, %

6,43±0,07

р1-2<0,01

р1-3<0,0001

7,55±0,04

р2-3<0,0001

10,82±0,15

AST, U/L

22,82±1,09

р1-2>0,05

p1-3>0,05

25,56±3,42

p2-3>0,05

27,33±1,77

ALT, U/L

29,64±5,69

р1-2>0,05

p1-3>0,05

29,62±3,71

p2-3>0,05

33,69±2,07

Creatinine, μmol/L

90,19±3,13

р1-2>0,05

p1-3>0,05

92,38±5,36

p2-3>0,05

90,84±2,09

Urea, mmol/L

5,34±0,27

р1-2>0,05

p1-3>0,05

5,43 ±0,35

p2-3>0,05

5,44±0,19

Note: р – difference between groups

Table II. The content of insulin, leptin and resistin in the blood serum of type 2 diabetic patients depending on glycaemic control, (M ± m).

Data

Group 1

(HbA1c ≤ ٧ ٪)

Group 2

(HbA1c 7,1-8 %)

Group 3

(HbA1c ≥ 8,1 %)

Insulin, mcU/mL

21,61 ± 1,91

р1-2>0,05

р1-3>0,05

23,33±2,32

р2-3<0,05

18,46 ± 0,71

Leptin, ng/mL

23,26 ± 2,82

р1-2>0,05

р1-3>0,05

35,18 ± 6,37

р2-3>0,05

28,18 ± 2,68

Resistin, ng/mL

2,99±0,27

р1-2>0,05

р1-3<0,05

2,90±0,23

р2-3<0,05

3,87±0,18

Note. р – difference between groups.

Table III. The content of non-specific inflammation mediators in type 2 diabetic patients depending on glycaemic control, (M±m)

Data

Group 1

(HbA1c ≤ ٧ ٪)

Group 2

(HbA1c 7,1-8 %)

Group 3

(HbA1c ≥ 8,1 %)

TNF-α, pg/mL

7,83±0,76

р1-2>0,05

р1-3>0,05

7,99±1,22

р2-3>0,05

9,38±0,88

IL-2, pg/mL

6,15±0,39

Р1-2>0,05

р1-3>0,05

7,73±0,96

р2-3>0,05

7,07±0,63

IL-6, pg/mL

1,76±0,16

Р1-2>0,05

р1-3>0,05

2,34±0,52

р2-3>0,05

3,32±0,58

Note. р – difference between groups.

Fig. 1. Correlations between resistin concentration and IL-6 content in patients with suboptimal glycaemic control of type 2 diabetes.

Fig. 2. Correlations between resistin concentration and IL-6 content in patients with poor glycaemic control of type 2 diabetes.