- Email: [email protected]
Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy
G Model DSX 851 No. of Pages 4
Diabetes & Metabolic Syndrome: Clinical Research & Reviews xxx (2017) xxx–xxx
Contents lists available at ScienceDirect
Diabetes & Metabolic Syndrome: Clinical Research & Reviews journal homepage: www.elsevier.com/locate/dsx
Original Article
Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy Gandhipuram Periyasamy Senthilkumara,* , Melepallappil Sabeenakumari Anithalekshmia , Md. Yasira , Sreejith Parameswaranb , Rajaa muthu Packirisamya , Zachariah Bobbya a b
Department of Biochemistry, Jawaharlal Institute of Postgraduate Medical Education and Research, Puducherry-605006, India Department of Nephrology, Jawaharlal Institute of Postgraduate Medical Education and Research, Puducherry-605006, India
A R T I C L E I N F O
A B S T R A C T
Article history: Available online xxx
Aims: Diabetic nephropathy (DN) is one of the major chronic vascular complication of T2DM and leading cause of end-stage renal disease. Inflammation is one of the proposed pathway which explains microvascular complications in T2DM but exact mechanism is still unclear. Omentin-1 is an antiinflammatory adipokine which promotes insulin signaling. IL-6 is a multifunctional cytokine having role in immune and inflammatory responses. The present study was conducted to elucidate the role of omentin-1 and IL-6 in the pathogenesis of DN and its association with insulin resistance. We aimed to assess and compare the serum levels of omentin-1 and IL-6 in T2DM patients with and without DN. Materials & Methods: Our study comprised of 2 groups of 41 each. Group A (controls) included T2DM without nephropathy patients and group B (cases) included T2DM nephropathy patients. Parameters studied were serum omentin-1, insulin, IL-6, fasting blood glucose, urea, creatinine, lipid profile, HOMAIR, eGFR and BMI. Results & Conclusion: Omentin-1 (p=0.03) was significantly decreased; concomitantly, significant increase in levels of insulin (p=0.004), IL-6 (p=0.023) and HOMA-IR (p=0.0004) were found in cases compared to controls. Bivariate analysis showed eGFR correlating positively with omentin-1 and negatively with insulin in the study population. Our study results, based on serum omentin-1 and IL-6 data suggest important role played by inflammatory mechanism and insulin resistance in the pathogenesis of diabetic nephropathy in type 2 diabetes mellitus patients. © 2017 Diabetes India. Published by Elsevier Ltd. All rights reserved.
Keywords: Diabetic nephropathy Omentin-1 IL-6
1. Introduction Worldwide, 415 million people are affected by diabetes mellitus, out of which, India alone account for 69 million cases [1]. A long period of diabetes causes vascular changes and dysfunction [2]. Diabetic nephropathy (DN) is the one of the major chronic vascular complication of diabetes mellitus and leading cause of end-stage renal disease [3]. A study from India has shown that 31.3% of renal failure in India is caused by DN [4]. DN is characterized by excessive extracellular matrix deposition in tubules, interstitial tissue and glomerulus and this deposition may develop into interstitial fibrosis and glomerular sclerosis [5].
* Corresponding author at: Department of Biochemistry, Jipmer Academic Centre, JIPMER, Dhanvantri Nagar, Gorimedu, Puducherry-605006, India. E-mail address: [email protected] (G.P. Senthilkumar).
Hyperglycaemia, hypertension, family history and smoking are major risk factors for the development of DN in type 2 diabetes mellitus (T2DM) patients. Higher-normal albumin excretion and time duration are important indicators of T2DM patients progressing to nephropathy. Omentin-1 is a protein composed of 313-amino acid. It is an anti-inflammatory adipokine, predominantly expressed in stromal vascular cells of visceral adipose tissue and it promotes insulin signaling [6,7]. Circulating omentin-1 level is related with pathological mechanisms of obesity, vascular and metabolic disorders, atherosclerosis, hypertension and cardiovascular disease [8]. It is also associated with macro- and microvascular complications of diabetes mellitus. Interleukin-6, formerly called B-cell stimulatory factor- 2, is a multifunctional cytokine. IL-6 has a variety of functions, including growth and differentiation of hematopoietic cells and the immune and inflammatory responses [9]. Recently, studies have shown that
http://dx.doi.org/10.1016/j.dsx.2017.08.005 1871-4021/© 2017 Diabetes India. Published by Elsevier Ltd. All rights reserved.
Please cite this article in press as: G.P. Senthilkumar, et al., Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy, Diab Met Syndr: Clin Res Rev (2017), http://dx.doi.org/10.1016/j.dsx.2017.08.005
G Model DSX 851 No. of Pages 4
2
G.P. Senthilkumar et al. / Diabetes & Metabolic Syndrome: Clinical Research & Reviews xxx (2017) xxx–xxx
IL-6 is implicated in glomerulonephritis, including mesangial proliferative glomerulonephritis. Its level is highly elevated in DN and associated with albumin excretion and renal hypertrophy [10]. The present study was conducted to elucidate the role of omentin-1 and IL-6 in the pathogenesis of DN and its association with insulin resistance. We aimed to assess and compare the serum levels of omentin-1 and IL-6 in T2DM patients with and without DN.
using commercially available ELISA kits and readings taken using Biotech ELISA reader. eGFR (estimated glomerular filtration rate) was calculated using CKD- EPI creatinine equation [12]: eGFR = 141 * min {Scr/k, 1}a* max {Scr/k, 1} {If female} * 1.159 {If black}
1.209
* 0.993 Age * 1.018
Scr = Serum creatinine in mg/dl
2. Materials and methods This case-control study was conducted by the Department of Biochemistry, Jawaharlal Institute of Postgraduate Medical Education and Research (JIPMER), Puducherry, in collaboration with the Department of Nephrology, JIPMER, during 2015 2016. Ethical approval was obtained from the Institute Ethics Committee (no. JIP/IEC/SC/2015/23/843) and written informed consent was obtained from all the study subjects. Sample size required for the study was estimated as 41 in each groups. It was calculated within expected differences in omentin-1 levels between the groups at 5% level of significance and 90% power based on the study done by Tekce et al., 2014 [11]. 2.1. Study protocol and laboratory analysis The study subjects were divided into 2 groups of 41 each. Group A (controls) included diabetic without nephropathy patients. Group B (cases) included already diagnosed diabetic nephropathy patients (DN). All patients were between the age of 18 to 60 years. Patients with pregnancy, malignancy, cardiovascular diseases, active infectious disease, rheumatoid arthritis, SLE, other inflammatory diseases were not included in the present study. Clinical data and anthropometric parameters were collected after obtaining informed written consent from each study patients. About 5 ml of venous blood was drawn after 12 h fasting from all patients. The sample was centrifuged at 4000 rpm for 5 mintues and serum was separated. Routine biochemical parameters like fasting blood glucose, urea, creatinine, albumin, lipid profile levels were estimated using the auto-analyser (Beckman-coulter AU680) and haemoglobin measured by another auto-analyser (Sysmex XS1000i). Serum levels of Omentin-1 (RayBiotech, USA), IL-6 (Diaclone SAS, France) and Insulin (DIAsource, Belgium) estimated
k = 0.7 for females or 0.9 for males
a = 0.329 for females and
0.411 for males
HOMA-IR was calculated using the formula: HOMA-IR = [Fasting insulin (mIU/ml) Fasting glucose (mg/dl)]/ 405 2.2. Statistical analysis Statistical analysis was performed using SPSS version 20 software for windows. The distribution of continuous data were calculated and expressed as mean SD. The comparisons of these variables between the groups were done by using independent student’s t- test. The correlation analysis between variables were done by Pearson’s correlation. All statistical tests were two-tailed and P < 0.05 were considered statistically significant. 3. Results 3.1. Baseline anthropometric and laboratory parameters of patients Table 1 shows the baseline anthropometric and routine laboratory data of cases and controls. We found statistically significant age difference between study groups. Cases have significantly decreased weight, waist circumference and BMI compared to controls. Systolic and diastolic blood pressures in cases are significantly higher compared to controls. Cases have significantly higher serum levels of fasting serum glucose, urea,
Table 1 Baseline anthropometric and laboratory parameters of patients. Parameters
Group A (controls) (n = 41)
Group B (cases) (n = 41)
P value
Age (years) Height (cm) Weight (kg) BMI (kg/m2) SBP (mm/Hg) DBP (mm/Hg) Waist circumference (cm) Fasting serum glucose (mg/dl) Urea (mg/dl) Creatinine (mg/dl) Albumin (mg/dl) eGFR (ml/min) Total cholesterol (mg/dl) Triglyceride (mg/dl) HDL cholesterol (mg/dl) LDL cholesterol (mg/dl) VLDL cholesterol (mg/dl) Haemoglobin (g/dl)
46.34 7.1 154.48 11.2 66.62 12.2 27.2 3.7 124.04 10.70 78.75 9.01 93.90 10.69 154.42 52.52 23.10 4.43 0.80 0.09 4.14 0.45 99.80 29.58 179.04 37.68 141.60 47.28 43.75 9.54 106.97 39.7 28.32 9.45 12.58 1.09
50.65 7.13 160 7.3 59.75 8.3 23.30 2.7 135.75 5.35 88.31 8.34 87.07 8.64 196.36 89.26 62.53 22.34 2.88 0.64 2.98 0.33 26.14 7.74 208.48 38.91 176.04 88.62 41.53 13.74 130.27 33.24 35.20 17.72 11.13 1.11
0.008** 0.100 0.004** 0.001** 0.001** 0.001** 0.0028** 0.012* 0.001** 0.001** 0.001** 0.001** 0.001** 0.032* 0.339 0.005** 0.032* 0.001**
*Data are mean SD. *P < 0.05; **P < 0.01. BMI: Body mass index; SBP: Systolic blood pressure; DBP: Diastolic blood pressure; eGFR: estimated glomerular filtration rate; HDL: High density lipoprotein; LDL: Low density lipoprotein; VLDL: Very low density lipoprotein.
Please cite this article in press as: G.P. Senthilkumar, et al., Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy, Diab Met Syndr: Clin Res Rev (2017), http://dx.doi.org/10.1016/j.dsx.2017.08.005
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Table 2 Results of special serum parameters assayed by ELISA. Parameters
Group A (controls) (n = 41)
Group B (cases) (n = 41)
P value
Insulin (mU/ml) IL-6 (pg/ml) Omentin-1 (ng/ml) HOMA-IR
7.36 3.71 8.71 3.07 21.07 9.63 2.87 1.8
11.24 7.30 10.5 3.8 16.30 9.89 5.94 4.9
0.004** 0.023* 0.030* 0.0004***
Data are mean SD. *P < 0.05; **P < 0.01; ***P < 0.001 IL-6: Interleukin 6; HOMA-IR: Homeostatic model assessment of insulin resistance.
creatinine, total cholesterol, triglyceride, LDL, VLDL and haemoglobin in comparison to controls. Serum albumin level was significantly decreased in cases compared to controls. Estimated GFR was found to be significantly decreased in cases compared to controls. There was no difference between groups in term of serum HDL level.
3.3. Diabetic nephropathy stages and serum levels of omentin-1, insulin and IL-6 Table 3 shows the comparison of our specific study parameters between stages of nephropathy. All our cases were either stage 3 or stage 4. We found no significant difference between stage 3 and stage 4 in terms of insulin, IL-6 and omentin-1.
3.2. Results of special serum parameters assayed by ELISA 3.4. Relationship of estimated GFR with omentin-1, insulin and IL-6 Table 2 shows the comparison of our special study parameters between groups. We found significantly higher levels of serum insulin and IL-6 in cases compared to controls. Serum omentin-1 level was significantly decreased in cases compared to controls. We calculated HOMA-IR using fasting glucose and insulin values and found it to be significantly higher in cases compared to controls. Table 3 Diabetic nephropathy stages and serum levels of insulin, IL-6 and omentin-1. Parameters
Stage 3 (n = 14)
Stage 4 (n = 27)
P value
Insulin (mU/ml) IL-6 (pg/ml) Omentin-1 (ng/ml)
12.8 7.9 9.5 3.4 18.1 10.1
10.3 6.9 11.03 4.03 15.3 9.8
0.335 0.230 0.406
Data are mean SD.
Table 4 Relationship of estimated GFR with omentin-1, insulin and IL-6. Parameters
R value
P value
Omentin-1 (ng/ml) Insulin (mU/ml) IL-6 (pg/ml)
0.238 0.285 0.206
0.032* 0.010* 0.064
*P < 0.05.
Table 4 shows the results of correlation study done between eGFR and omentin-1, insulin and IL-6 in all the study population. Significant positive correlation were found between eGFR and omentin-1 (Fig. 1); concomitantly eGFR were found to have significant negative correlation with insulin. eGFR also correlated negatively with IL-6 but it was not statistically significant. 4. Discussion In this study we proposed to find the role of omentin-1 and IL-6 in the pathogenesis of diabetic nephropathy and its association with insulin resistance. We found 4 main findings from this study: (1) Serum Omentin-1 level was significantly decreased; concomitantly IL-6 and insulin levels were significantly increased in DN patients compared to diabetic controls. (2) HOMA-IR, a measure of insulin resistance, is found to be significantly higher in DN patients compared to diabetic controls. (3) No significant difference in the levels of omentin-1 were found between stage 3 and stage 4 nephropathy. (4) Estimated GFR, which is a prevalent marker of kidney function, is found to have significant positive correlation with omentin-1 and significant negative correlation with insulin. Out of 2 types of obesity, visceral obesity is mainly linked with metabolic disorders, diabetes mellitus and insulin resistance.
Fig. 1. Scatter plot showing correlation of omentin-1 with estimated GFR in whole study population. R = 0.238, P = 0.032*.
Please cite this article in press as: G.P. Senthilkumar, et al., Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy, Diab Met Syndr: Clin Res Rev (2017), http://dx.doi.org/10.1016/j.dsx.2017.08.005
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Omentin-1 is a novel adipocytokine having a role in insulin resistance, micro vascular complications of DM, obesity, atherosclerosis and inflammation. It has been found that omentin-1 is an insulin sensitizing molecule [13]. Studies have shown a negative correlation between omentin-1 level and metabolic conditions like diabetes mellitus [14]. In this study, we found that serum omentin1 level in case group (T2DM nephropathy) was significantly decreased as compared with control group (T2DM without nephropathy). We also found that serum levels of insulin and fasting glucose and HOMA-IR were significantly higher in cases than the control group. These results show that decrease in serum omentin-1 level leads to increased insulin resistance and subsequently poor control of diabetes in type 2 diabetes mellitus patients leading to its complications like nephropathy. Role of different marker molecules involved in inflammation have been explored and inflammation has been proposed as a potential mechanism of diabetic nephropathy [15,16]. Through the work done on vascular smooth muscle cell lines, researchers have shown that omentin-1 plays an anti-inflammatory role[17]. Our study findings of decreased omentin-1 and increased IL-6 in cases compared to controls substantiate the role of inflammation in the pathogenesis of diabetic nephropathy. All patients in our case group were either stage-3 or stage-4 nephropathy. We did not find any significant difference in the level of omentin-1. This can be accounted for due to worsened excretory function of kidneys in stage 4 nephropathy. Similar results were obtained in a study done with patients of end-stage renal disease who were on haemodialysis [18]. Despite this exception, we still found omentin-1 to have positive correlation with eGFR in whole study population. We also found the significant increase in total and LDL cholesterol in diabetic nephropathy group compared to controls. Along with insulin resistance, diabetic nephropathy has been found to be a potential risk factor for the development of atherosclerosis [19,20]. This is probably the first study done in Indian population to find out the potential role of omentin-1 and IL-6 in the mechanism of diabetic nephropathy in T2DM patients. Still, our study has some limitations. It is a cross-sectional study, so causal association cannot be elucidated. Also, all the diabetic nephropathy cases are of stage 3 or 4 which accounts for macroalbuminuria. We can expect more significant results with early stages of nephropathy cases showing microalbuminuria. 5. Conclusion In conclusion, decreased serum level of omentin-1 and increased serum levels of IL-6 and insulin and raised HOMA-IR in cases suggest that insulin resistance and inflammation are important mechanisms in the pathogenesis of diabetic nephropathy in type 2 diabetes mellitus patients. Disclosure
Acknowledgements We would like to thank JIPMER, Puducherry, India for funding this study. References [1] International Diabetes Federation. Diabetes Atlas. 7th edition 2015. . (Accessed June 2 2017) http://www.diabetesatlas.org. [2] Mauer SM, Steffes MW, Ellis EN, Sutherland DE, Brown DM, Goetz FC. Structural-functional relationships in diabetic nephropathy. J Clin Invest 1984;74(4):1143–55. [3] Rajapurkar MM, John GT, Kirpalani AL, Abraham G, Agarwal SK, Almeida AF, et al. What do we know about chronic kidney disease in India: first report of the Indian CKD registry. BMC Nephrol 2012;13:10. [4] Ritz E, Zeng X. Diabetic nephropathy epidemiology in Asia and the current state of treatment. Indian J Nephrol 2011;21(2):75–84. [5] Hovind P, Rossing P, Tarnow L, Smidt UM, Parving HH. Progression of diabetic nephropathy. Kidney Int 2001;59(2):702–9. [6] Lesná J, Tichá A, Hyšpler R, Musil F, Bláha V, Sobotka L, et al. Omentin-1 plasma levels and cholesterol metabolism in obese patients with diabetes mellitus type 1: impact of weight reduction. Nutr Diabetes 2015;5(11):e183. [7] Yang X-H, Cao R-F, Yu Y, Sui M, Zhang T, Xu J-Y, et al. A study on the correlation between MTHFR promoter methylation and diabetic nephropathy. Am J Transl Res 2016;8(11):4960–7. [8] Li Y, Gong G, Geng H, Qian Y. phox C242T gene polymorphism and overt diabetic nephropathy: a meta-analysis of 1,452 participants. Korean J Intern Med 2017 Published online on 2016 December 8[Internet] 2016 [cited 2017 Jun 3]; Available from: http://www.kjim.org/journal/view.php?number=169759. [9] Scheller J, Chalaris A, Schmidt-Arras D, Rose-John S. The pro- and antiinflammatory properties of the cytokine interleukin-6. Biochim Biophys Acta 2011;1813(5):878–88. [10] Yan P, Liu D, Long M, Ren Y, Pang J, Li R. Changes of serum omentin levels and relationship between omentin and adiponectin concentrations in type 2 diabetes mellitus. Exp Clin Endocrinol Diabetes 2011;119(4):257–63. [11] Tekce H, Tekce BK, Aktas G, Alcelik A, Sengul E. Serum omentin-1 levels in diabetic and nondiabetic patients with chronic kidney disease. Exp Clin Endocrinol Diabetes 2014;122(8):451–6. [12] Levey AS, Stevens LA, Schmid CH, Zhang YL, Castro AF, Feldman HI, et al. A new equation to estimate glomerular filtration rate. Ann Intern Med 2009;150 (9):604–12. [13] Moreno-Navarrete JM, Catalán V, Ortega F, Gómez-Ambrosi J, Ricart W, Frühbeck G, et al. Circulating omentin concentration increases after weight loss. Nutr Metab 2010;7:27. [14] Gürsoy G, Kırnap NG, Eşbah O, Acar Y, Demirbaş B, Akçayöz S, et al. The relationship between plasma omentin-1 levels and insulin resistance in newly diagnosed type 2 diabetıc women. Clin Rev Opin 2010;2(4):49–54. [15] Forbes JM, Cooper ME. Mechanisms of diabetic complications. Physiol Rev 2013;93(1):137–88. [16] Lim AKH, Tesch GH. Inflammation in diabetic nephropathy [Internet]. Mediators Inflamm 2012;2012:12 Article ID 146154[cited 2017 Jul 12]. Available from: https://www.hindawi.com/journals/mi/2012/146154/. [17] Kazama K, Usui T, Okada M, Hara Y, Yamawaki H. Omentin plays an antiinflammatory role through inhibition of TNF-a-induced superoxide production in vascular smooth muscle cells. Eur J Pharmacol 2012;686(13):116–23. [18] Alcelik A, Tosun M, Ozlu MF, Eroglu M, Aktas G, Kemahli E, et al. Serum levels of omentin in end-stage renal disease patients. Kidney Blood Press Res 2012;35 (6):511–6. [19] Shinohara K, Shoji T, Emoto M, Tahara H, Koyama H, Ishimura E, et al. Insulin resistance as an independent predictor of cardiovascular mortality in patients with end-stage renal disease. J Am Soc Nephrol JASN 2002;13(7):1894–900. [20] Wu T, McGrath KC, Death AK. Cardiovascular disease in diabetic nephropathy patients: cell adhesion molecules as potential markers? Vasc Health Risk Manage 2005;1(4):309–16.
The authors declared no conflicts of interest. This study was funded by the JIPMER Intramural Research Grant (JIP/Res/IntraMSc/02/2015-16 dated 31/12/2015).
Please cite this article in press as: G.P. Senthilkumar, et al., Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy, Diab Met Syndr: Clin Res Rev (2017), http://dx.doi.org/10.1016/j.dsx.2017.08.005
Diabetes & Metabolic Syndrome: Clinical Research & Reviews xxx (2017) xxx–xxx
Contents lists available at ScienceDirect
Diabetes & Metabolic Syndrome: Clinical Research & Reviews journal homepage: www.elsevier.com/locate/dsx
Original Article
Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy Gandhipuram Periyasamy Senthilkumara,* , Melepallappil Sabeenakumari Anithalekshmia , Md. Yasira , Sreejith Parameswaranb , Rajaa muthu Packirisamya , Zachariah Bobbya a b
Department of Biochemistry, Jawaharlal Institute of Postgraduate Medical Education and Research, Puducherry-605006, India Department of Nephrology, Jawaharlal Institute of Postgraduate Medical Education and Research, Puducherry-605006, India
A R T I C L E I N F O
A B S T R A C T
Article history: Available online xxx
Aims: Diabetic nephropathy (DN) is one of the major chronic vascular complication of T2DM and leading cause of end-stage renal disease. Inflammation is one of the proposed pathway which explains microvascular complications in T2DM but exact mechanism is still unclear. Omentin-1 is an antiinflammatory adipokine which promotes insulin signaling. IL-6 is a multifunctional cytokine having role in immune and inflammatory responses. The present study was conducted to elucidate the role of omentin-1 and IL-6 in the pathogenesis of DN and its association with insulin resistance. We aimed to assess and compare the serum levels of omentin-1 and IL-6 in T2DM patients with and without DN. Materials & Methods: Our study comprised of 2 groups of 41 each. Group A (controls) included T2DM without nephropathy patients and group B (cases) included T2DM nephropathy patients. Parameters studied were serum omentin-1, insulin, IL-6, fasting blood glucose, urea, creatinine, lipid profile, HOMAIR, eGFR and BMI. Results & Conclusion: Omentin-1 (p=0.03) was significantly decreased; concomitantly, significant increase in levels of insulin (p=0.004), IL-6 (p=0.023) and HOMA-IR (p=0.0004) were found in cases compared to controls. Bivariate analysis showed eGFR correlating positively with omentin-1 and negatively with insulin in the study population. Our study results, based on serum omentin-1 and IL-6 data suggest important role played by inflammatory mechanism and insulin resistance in the pathogenesis of diabetic nephropathy in type 2 diabetes mellitus patients. © 2017 Diabetes India. Published by Elsevier Ltd. All rights reserved.
Keywords: Diabetic nephropathy Omentin-1 IL-6
1. Introduction Worldwide, 415 million people are affected by diabetes mellitus, out of which, India alone account for 69 million cases [1]. A long period of diabetes causes vascular changes and dysfunction [2]. Diabetic nephropathy (DN) is the one of the major chronic vascular complication of diabetes mellitus and leading cause of end-stage renal disease [3]. A study from India has shown that 31.3% of renal failure in India is caused by DN [4]. DN is characterized by excessive extracellular matrix deposition in tubules, interstitial tissue and glomerulus and this deposition may develop into interstitial fibrosis and glomerular sclerosis [5].
* Corresponding author at: Department of Biochemistry, Jipmer Academic Centre, JIPMER, Dhanvantri Nagar, Gorimedu, Puducherry-605006, India. E-mail address: [email protected] (G.P. Senthilkumar).
Hyperglycaemia, hypertension, family history and smoking are major risk factors for the development of DN in type 2 diabetes mellitus (T2DM) patients. Higher-normal albumin excretion and time duration are important indicators of T2DM patients progressing to nephropathy. Omentin-1 is a protein composed of 313-amino acid. It is an anti-inflammatory adipokine, predominantly expressed in stromal vascular cells of visceral adipose tissue and it promotes insulin signaling [6,7]. Circulating omentin-1 level is related with pathological mechanisms of obesity, vascular and metabolic disorders, atherosclerosis, hypertension and cardiovascular disease [8]. It is also associated with macro- and microvascular complications of diabetes mellitus. Interleukin-6, formerly called B-cell stimulatory factor- 2, is a multifunctional cytokine. IL-6 has a variety of functions, including growth and differentiation of hematopoietic cells and the immune and inflammatory responses [9]. Recently, studies have shown that
http://dx.doi.org/10.1016/j.dsx.2017.08.005 1871-4021/© 2017 Diabetes India. Published by Elsevier Ltd. All rights reserved.
Please cite this article in press as: G.P. Senthilkumar, et al., Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy, Diab Met Syndr: Clin Res Rev (2017), http://dx.doi.org/10.1016/j.dsx.2017.08.005
G Model DSX 851 No. of Pages 4
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G.P. Senthilkumar et al. / Diabetes & Metabolic Syndrome: Clinical Research & Reviews xxx (2017) xxx–xxx
IL-6 is implicated in glomerulonephritis, including mesangial proliferative glomerulonephritis. Its level is highly elevated in DN and associated with albumin excretion and renal hypertrophy [10]. The present study was conducted to elucidate the role of omentin-1 and IL-6 in the pathogenesis of DN and its association with insulin resistance. We aimed to assess and compare the serum levels of omentin-1 and IL-6 in T2DM patients with and without DN.
using commercially available ELISA kits and readings taken using Biotech ELISA reader. eGFR (estimated glomerular filtration rate) was calculated using CKD- EPI creatinine equation [12]: eGFR = 141 * min {Scr/k, 1}a* max {Scr/k, 1} {If female} * 1.159 {If black}
1.209
* 0.993 Age * 1.018
Scr = Serum creatinine in mg/dl
2. Materials and methods This case-control study was conducted by the Department of Biochemistry, Jawaharlal Institute of Postgraduate Medical Education and Research (JIPMER), Puducherry, in collaboration with the Department of Nephrology, JIPMER, during 2015 2016. Ethical approval was obtained from the Institute Ethics Committee (no. JIP/IEC/SC/2015/23/843) and written informed consent was obtained from all the study subjects. Sample size required for the study was estimated as 41 in each groups. It was calculated within expected differences in omentin-1 levels between the groups at 5% level of significance and 90% power based on the study done by Tekce et al., 2014 [11]. 2.1. Study protocol and laboratory analysis The study subjects were divided into 2 groups of 41 each. Group A (controls) included diabetic without nephropathy patients. Group B (cases) included already diagnosed diabetic nephropathy patients (DN). All patients were between the age of 18 to 60 years. Patients with pregnancy, malignancy, cardiovascular diseases, active infectious disease, rheumatoid arthritis, SLE, other inflammatory diseases were not included in the present study. Clinical data and anthropometric parameters were collected after obtaining informed written consent from each study patients. About 5 ml of venous blood was drawn after 12 h fasting from all patients. The sample was centrifuged at 4000 rpm for 5 mintues and serum was separated. Routine biochemical parameters like fasting blood glucose, urea, creatinine, albumin, lipid profile levels were estimated using the auto-analyser (Beckman-coulter AU680) and haemoglobin measured by another auto-analyser (Sysmex XS1000i). Serum levels of Omentin-1 (RayBiotech, USA), IL-6 (Diaclone SAS, France) and Insulin (DIAsource, Belgium) estimated
k = 0.7 for females or 0.9 for males
a = 0.329 for females and
0.411 for males
HOMA-IR was calculated using the formula: HOMA-IR = [Fasting insulin (mIU/ml) Fasting glucose (mg/dl)]/ 405 2.2. Statistical analysis Statistical analysis was performed using SPSS version 20 software for windows. The distribution of continuous data were calculated and expressed as mean SD. The comparisons of these variables between the groups were done by using independent student’s t- test. The correlation analysis between variables were done by Pearson’s correlation. All statistical tests were two-tailed and P < 0.05 were considered statistically significant. 3. Results 3.1. Baseline anthropometric and laboratory parameters of patients Table 1 shows the baseline anthropometric and routine laboratory data of cases and controls. We found statistically significant age difference between study groups. Cases have significantly decreased weight, waist circumference and BMI compared to controls. Systolic and diastolic blood pressures in cases are significantly higher compared to controls. Cases have significantly higher serum levels of fasting serum glucose, urea,
Table 1 Baseline anthropometric and laboratory parameters of patients. Parameters
Group A (controls) (n = 41)
Group B (cases) (n = 41)
P value
Age (years) Height (cm) Weight (kg) BMI (kg/m2) SBP (mm/Hg) DBP (mm/Hg) Waist circumference (cm) Fasting serum glucose (mg/dl) Urea (mg/dl) Creatinine (mg/dl) Albumin (mg/dl) eGFR (ml/min) Total cholesterol (mg/dl) Triglyceride (mg/dl) HDL cholesterol (mg/dl) LDL cholesterol (mg/dl) VLDL cholesterol (mg/dl) Haemoglobin (g/dl)
46.34 7.1 154.48 11.2 66.62 12.2 27.2 3.7 124.04 10.70 78.75 9.01 93.90 10.69 154.42 52.52 23.10 4.43 0.80 0.09 4.14 0.45 99.80 29.58 179.04 37.68 141.60 47.28 43.75 9.54 106.97 39.7 28.32 9.45 12.58 1.09
50.65 7.13 160 7.3 59.75 8.3 23.30 2.7 135.75 5.35 88.31 8.34 87.07 8.64 196.36 89.26 62.53 22.34 2.88 0.64 2.98 0.33 26.14 7.74 208.48 38.91 176.04 88.62 41.53 13.74 130.27 33.24 35.20 17.72 11.13 1.11
0.008** 0.100 0.004** 0.001** 0.001** 0.001** 0.0028** 0.012* 0.001** 0.001** 0.001** 0.001** 0.001** 0.032* 0.339 0.005** 0.032* 0.001**
*Data are mean SD. *P < 0.05; **P < 0.01. BMI: Body mass index; SBP: Systolic blood pressure; DBP: Diastolic blood pressure; eGFR: estimated glomerular filtration rate; HDL: High density lipoprotein; LDL: Low density lipoprotein; VLDL: Very low density lipoprotein.
Please cite this article in press as: G.P. Senthilkumar, et al., Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy, Diab Met Syndr: Clin Res Rev (2017), http://dx.doi.org/10.1016/j.dsx.2017.08.005
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Table 2 Results of special serum parameters assayed by ELISA. Parameters
Group A (controls) (n = 41)
Group B (cases) (n = 41)
P value
Insulin (mU/ml) IL-6 (pg/ml) Omentin-1 (ng/ml) HOMA-IR
7.36 3.71 8.71 3.07 21.07 9.63 2.87 1.8
11.24 7.30 10.5 3.8 16.30 9.89 5.94 4.9
0.004** 0.023* 0.030* 0.0004***
Data are mean SD. *P < 0.05; **P < 0.01; ***P < 0.001 IL-6: Interleukin 6; HOMA-IR: Homeostatic model assessment of insulin resistance.
creatinine, total cholesterol, triglyceride, LDL, VLDL and haemoglobin in comparison to controls. Serum albumin level was significantly decreased in cases compared to controls. Estimated GFR was found to be significantly decreased in cases compared to controls. There was no difference between groups in term of serum HDL level.
3.3. Diabetic nephropathy stages and serum levels of omentin-1, insulin and IL-6 Table 3 shows the comparison of our specific study parameters between stages of nephropathy. All our cases were either stage 3 or stage 4. We found no significant difference between stage 3 and stage 4 in terms of insulin, IL-6 and omentin-1.
3.2. Results of special serum parameters assayed by ELISA 3.4. Relationship of estimated GFR with omentin-1, insulin and IL-6 Table 2 shows the comparison of our special study parameters between groups. We found significantly higher levels of serum insulin and IL-6 in cases compared to controls. Serum omentin-1 level was significantly decreased in cases compared to controls. We calculated HOMA-IR using fasting glucose and insulin values and found it to be significantly higher in cases compared to controls. Table 3 Diabetic nephropathy stages and serum levels of insulin, IL-6 and omentin-1. Parameters
Stage 3 (n = 14)
Stage 4 (n = 27)
P value
Insulin (mU/ml) IL-6 (pg/ml) Omentin-1 (ng/ml)
12.8 7.9 9.5 3.4 18.1 10.1
10.3 6.9 11.03 4.03 15.3 9.8
0.335 0.230 0.406
Data are mean SD.
Table 4 Relationship of estimated GFR with omentin-1, insulin and IL-6. Parameters
R value
P value
Omentin-1 (ng/ml) Insulin (mU/ml) IL-6 (pg/ml)
0.238 0.285 0.206
0.032* 0.010* 0.064
*P < 0.05.
Table 4 shows the results of correlation study done between eGFR and omentin-1, insulin and IL-6 in all the study population. Significant positive correlation were found between eGFR and omentin-1 (Fig. 1); concomitantly eGFR were found to have significant negative correlation with insulin. eGFR also correlated negatively with IL-6 but it was not statistically significant. 4. Discussion In this study we proposed to find the role of omentin-1 and IL-6 in the pathogenesis of diabetic nephropathy and its association with insulin resistance. We found 4 main findings from this study: (1) Serum Omentin-1 level was significantly decreased; concomitantly IL-6 and insulin levels were significantly increased in DN patients compared to diabetic controls. (2) HOMA-IR, a measure of insulin resistance, is found to be significantly higher in DN patients compared to diabetic controls. (3) No significant difference in the levels of omentin-1 were found between stage 3 and stage 4 nephropathy. (4) Estimated GFR, which is a prevalent marker of kidney function, is found to have significant positive correlation with omentin-1 and significant negative correlation with insulin. Out of 2 types of obesity, visceral obesity is mainly linked with metabolic disorders, diabetes mellitus and insulin resistance.
Fig. 1. Scatter plot showing correlation of omentin-1 with estimated GFR in whole study population. R = 0.238, P = 0.032*.
Please cite this article in press as: G.P. Senthilkumar, et al., Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy, Diab Met Syndr: Clin Res Rev (2017), http://dx.doi.org/10.1016/j.dsx.2017.08.005
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Omentin-1 is a novel adipocytokine having a role in insulin resistance, micro vascular complications of DM, obesity, atherosclerosis and inflammation. It has been found that omentin-1 is an insulin sensitizing molecule [13]. Studies have shown a negative correlation between omentin-1 level and metabolic conditions like diabetes mellitus [14]. In this study, we found that serum omentin1 level in case group (T2DM nephropathy) was significantly decreased as compared with control group (T2DM without nephropathy). We also found that serum levels of insulin and fasting glucose and HOMA-IR were significantly higher in cases than the control group. These results show that decrease in serum omentin-1 level leads to increased insulin resistance and subsequently poor control of diabetes in type 2 diabetes mellitus patients leading to its complications like nephropathy. Role of different marker molecules involved in inflammation have been explored and inflammation has been proposed as a potential mechanism of diabetic nephropathy [15,16]. Through the work done on vascular smooth muscle cell lines, researchers have shown that omentin-1 plays an anti-inflammatory role[17]. Our study findings of decreased omentin-1 and increased IL-6 in cases compared to controls substantiate the role of inflammation in the pathogenesis of diabetic nephropathy. All patients in our case group were either stage-3 or stage-4 nephropathy. We did not find any significant difference in the level of omentin-1. This can be accounted for due to worsened excretory function of kidneys in stage 4 nephropathy. Similar results were obtained in a study done with patients of end-stage renal disease who were on haemodialysis [18]. Despite this exception, we still found omentin-1 to have positive correlation with eGFR in whole study population. We also found the significant increase in total and LDL cholesterol in diabetic nephropathy group compared to controls. Along with insulin resistance, diabetic nephropathy has been found to be a potential risk factor for the development of atherosclerosis [19,20]. This is probably the first study done in Indian population to find out the potential role of omentin-1 and IL-6 in the mechanism of diabetic nephropathy in T2DM patients. Still, our study has some limitations. It is a cross-sectional study, so causal association cannot be elucidated. Also, all the diabetic nephropathy cases are of stage 3 or 4 which accounts for macroalbuminuria. We can expect more significant results with early stages of nephropathy cases showing microalbuminuria. 5. Conclusion In conclusion, decreased serum level of omentin-1 and increased serum levels of IL-6 and insulin and raised HOMA-IR in cases suggest that insulin resistance and inflammation are important mechanisms in the pathogenesis of diabetic nephropathy in type 2 diabetes mellitus patients. Disclosure
Acknowledgements We would like to thank JIPMER, Puducherry, India for funding this study. References [1] International Diabetes Federation. Diabetes Atlas. 7th edition 2015. . (Accessed June 2 2017) http://www.diabetesatlas.org. [2] Mauer SM, Steffes MW, Ellis EN, Sutherland DE, Brown DM, Goetz FC. Structural-functional relationships in diabetic nephropathy. J Clin Invest 1984;74(4):1143–55. [3] Rajapurkar MM, John GT, Kirpalani AL, Abraham G, Agarwal SK, Almeida AF, et al. What do we know about chronic kidney disease in India: first report of the Indian CKD registry. BMC Nephrol 2012;13:10. [4] Ritz E, Zeng X. Diabetic nephropathy epidemiology in Asia and the current state of treatment. Indian J Nephrol 2011;21(2):75–84. [5] Hovind P, Rossing P, Tarnow L, Smidt UM, Parving HH. Progression of diabetic nephropathy. Kidney Int 2001;59(2):702–9. [6] Lesná J, Tichá A, Hyšpler R, Musil F, Bláha V, Sobotka L, et al. Omentin-1 plasma levels and cholesterol metabolism in obese patients with diabetes mellitus type 1: impact of weight reduction. Nutr Diabetes 2015;5(11):e183. [7] Yang X-H, Cao R-F, Yu Y, Sui M, Zhang T, Xu J-Y, et al. A study on the correlation between MTHFR promoter methylation and diabetic nephropathy. Am J Transl Res 2016;8(11):4960–7. [8] Li Y, Gong G, Geng H, Qian Y. phox C242T gene polymorphism and overt diabetic nephropathy: a meta-analysis of 1,452 participants. Korean J Intern Med 2017 Published online on 2016 December 8[Internet] 2016 [cited 2017 Jun 3]; Available from: http://www.kjim.org/journal/view.php?number=169759. [9] Scheller J, Chalaris A, Schmidt-Arras D, Rose-John S. The pro- and antiinflammatory properties of the cytokine interleukin-6. Biochim Biophys Acta 2011;1813(5):878–88. [10] Yan P, Liu D, Long M, Ren Y, Pang J, Li R. Changes of serum omentin levels and relationship between omentin and adiponectin concentrations in type 2 diabetes mellitus. Exp Clin Endocrinol Diabetes 2011;119(4):257–63. [11] Tekce H, Tekce BK, Aktas G, Alcelik A, Sengul E. Serum omentin-1 levels in diabetic and nondiabetic patients with chronic kidney disease. Exp Clin Endocrinol Diabetes 2014;122(8):451–6. [12] Levey AS, Stevens LA, Schmid CH, Zhang YL, Castro AF, Feldman HI, et al. A new equation to estimate glomerular filtration rate. Ann Intern Med 2009;150 (9):604–12. [13] Moreno-Navarrete JM, Catalán V, Ortega F, Gómez-Ambrosi J, Ricart W, Frühbeck G, et al. Circulating omentin concentration increases after weight loss. Nutr Metab 2010;7:27. [14] Gürsoy G, Kırnap NG, Eşbah O, Acar Y, Demirbaş B, Akçayöz S, et al. The relationship between plasma omentin-1 levels and insulin resistance in newly diagnosed type 2 diabetıc women. Clin Rev Opin 2010;2(4):49–54. [15] Forbes JM, Cooper ME. Mechanisms of diabetic complications. Physiol Rev 2013;93(1):137–88. [16] Lim AKH, Tesch GH. Inflammation in diabetic nephropathy [Internet]. Mediators Inflamm 2012;2012:12 Article ID 146154[cited 2017 Jul 12]. Available from: https://www.hindawi.com/journals/mi/2012/146154/. [17] Kazama K, Usui T, Okada M, Hara Y, Yamawaki H. Omentin plays an antiinflammatory role through inhibition of TNF-a-induced superoxide production in vascular smooth muscle cells. Eur J Pharmacol 2012;686(13):116–23. [18] Alcelik A, Tosun M, Ozlu MF, Eroglu M, Aktas G, Kemahli E, et al. Serum levels of omentin in end-stage renal disease patients. Kidney Blood Press Res 2012;35 (6):511–6. [19] Shinohara K, Shoji T, Emoto M, Tahara H, Koyama H, Ishimura E, et al. Insulin resistance as an independent predictor of cardiovascular mortality in patients with end-stage renal disease. J Am Soc Nephrol JASN 2002;13(7):1894–900. [20] Wu T, McGrath KC, Death AK. Cardiovascular disease in diabetic nephropathy patients: cell adhesion molecules as potential markers? Vasc Health Risk Manage 2005;1(4):309–16.
The authors declared no conflicts of interest. This study was funded by the JIPMER Intramural Research Grant (JIP/Res/IntraMSc/02/2015-16 dated 31/12/2015).
Please cite this article in press as: G.P. Senthilkumar, et al., Role of omentin 1 and IL-6 in type 2 diabetes mellitus patients with diabetic nephropathy, Diab Met Syndr: Clin Res Rev (2017), http://dx.doi.org/10.1016/j.dsx.2017.08.005