Menoufia Medical Journal

ORIGINAL ARTICLE
Year
: 2016  |  Volume : 29  |  Issue : 1  |  Page : 22--29

The role of serum apelin in diabetic patients with retinopathy


Sanaa Sayed Gazareen1, Alaa Eldin Abdl El-salam Dawood1, Amin Faisal Ellakwa1, Dalia Hosny Abou Elela1, Amany Anwar Mohamed Serag2,  
1 Internal Medicine Department, Faculty of Medicine, Menoufiya University, Menoufiya, Egypt
2 Internal Medicine Department, Mansoura Insurance Hospital, Mansoura, Egypt

Correspondence Address:
Amany Anwar Mohamed Serag
MBBCh, Biala, 33571 Kafer El-Shiekh
Egypt

Abstract

Objective The aim of this work is to study serum apelin in patients with diabetic retinopathy and its possible pathophysiological role in such patients. Background Diabetic retinopathy is the most common microvascular complication of diabetes, and it remains a leading cause of blindness and visual impairment in the working-age population in the developed world. Apelin is a peptide isolated from bovine stomach extracts that acts as an endogenous ligand for the previously orphaned G protein-coupled receptors (angiotensin 1-related putative receptors). Apelin has been recognized as a factor promoting angiogenesis, but its role in the development of proliferative diabetic retinopathy (PDR) has not been fully examined. Materials and methods Sixty patients were included in the study and classified into three groups. Group 1, the control group, included individuals without diabetes mellitus (10 patients), group 2 included patients with diabetes mellitus but without retinopathy (20 patients), and group 3 included patients classified into two subgroups: group 3a, which included patients with nonproliferative diabetic retinopathy (NPDR) (15 patients), and group 3b, which included patients with PDR (15 patients). Results Apelin was significantly higher among patients with diabetes without retinopathy, NPDR, and PDR than the control group (P < 0.001); the mean value of apelin was significantly higher in patients with PDR than in those with diabetes without retinopathy (P < 0.05). Also, the mean value of apelin was significantly higher in patients with PDR than NPDR (P < 0.001). Conclusion Serum apelin was significantly higher in patients with PDR compared with those without nonproliferative retinopathy and in those with diabetes without retinopathy, and apelin plays a role in the development of PDR. Ischemic retinopathy could be trated with Inhibition of the apelinergic system. Serum apelin correlated significantly with serum creatinine in patients with retinopathy, which may indicate the role of apelin in diabetic nephropathy.



How to cite this article:
Gazareen SS, Dawood AE, Ellakwa AF, Elela DH, Serag AA. The role of serum apelin in diabetic patients with retinopathy.Menoufia Med J 2016;29:22-29


How to cite this URL:
Gazareen SS, Dawood AE, Ellakwa AF, Elela DH, Serag AA. The role of serum apelin in diabetic patients with retinopathy. Menoufia Med J [serial online] 2016 [cited 2024 Mar 28 ];29:22-29
Available from: http://www.mmj.eg.net/text.asp?2016/29/1/22/178940


Full Text

 Introduction



Diabetic retinopathy (DR) is the most common complication of diabetes and the leading cause of irreversible visual loss in individuals of working age in the USA [1]. DR involves damage to the microvasculature of the retina as a result of the prolonged exposure to the metabolic changes induced by diabetes [2].

The two main types of DR are the less severe form, nonproliferative diabetic retinopathy (NPDR), and the severe form, proliferative diabetic retinopathy (PDR), which involves the growth of new blood vessels, 'angiogenesis', in the retina [2].

PDR is commonly defined by retinal neovascular events and impairment of vision. The mechanisms of DR-related vision loss include vitreous hemorrhage, tractional retinal detachment, development of a fibrovascular membrane in the vitreous, and macular edema [3]. Retinal neovascularization, of PDR, is considered a major risk factor for severe vision loss in patients with diabetes mellitus [4].

Apelin mRNA and immunoreactive apelin were detected in various tissues and organs including the brain, heart, lung, stomach, uterus, and ovary [5]. The apelin system is distributed in diverse tissues, where it exerts a broad range of physiological and pathological actions, including angiogenesis, vasodilatation and vasoconstriction, myocardial ischemia, hypertension, appetite, the hypothalamus-hypophysis axis and fluid homeostasis, retinal vascular development and neovascularization, neuroprotection, and cell proliferation/survival [5],[6].

Apelin has been recognized as a factor promoting angiogenesis, but its role in the development of PDR has not been fully examined [7].

The apelin receptors (APLNR) are expressed in endothelial cells at the leading edge of vessels during early embryogenesis [8] and apelin in combination with vascular endothelial growth factor (VEGF) induces the proliferation of endothelial cells and the formation of new blood vessels (angiogenesis) [9],[10].

Progression of ischemic retinal diseases, such as DR, is associated closely with pathological retinal angiogenesis mainly induced by VEGF and erythropoietin [10],[11].

Antiangiogenic therapies targeting these factors are effective in the treatment of PDR [12],[13], but research has shown that anti-VEGF alone cannot completely prevent the development of new vessels, which indicates that factors other than VEGF may also participate in the process of neovascularization [14].

This work was carried out to study serum apelin in patients with DR and its possible pathophysiological role in such patients.

 Materials and methods



This study was carried out on 60 patients recruited from the inpatient department and outpatient clinics of the Internal Medicine Department and Ophthalmology Department in Menoufiya University Hospital; there were 25 men and 35 women between 26 and 68 years of age. The selected participants provided consent for participation in the study before they were subjected to examinations and investigations, and the study was approved by the Ethics Committee of Menoufiya University Hospital. The study was carried out from February 2013 to August 2013.

The selected participants were classified into three groups:

Group 1: control group of individuals without diabetes mellitus (10 patients, five men and five women). Group 2: patients with diabetes mellitus without retinopathy (20 patients, 15 women and five men). Group 3: patients with diabetes mellitus and retinopathy (30 patients) divided into two subgroups: group a: patients with NPDR (15 patients, eight women and seven men). Group b: patients with PDR (15 patients, seven women and eight men).

Inclusion criteria

Patients with type II diabetes mellitus (receiving oral hypoglycemic treatment) with different stages of DR were included in this study.

Exclusion criteria

Patients with advanced liver disease, heart failure, pregnancy, renal failure, steroid and nitrate usage, and non-DR were excluded from this study.

All patients were subjected to the following:

Assessment of history: Assessment of history was performed for all participants in the study group, with a special focus on the age and sex of the patients, duration of diabetes mellitus, and the presence or absence of specific diabetic complications and treatment.Complete clinical examination: All participants were subjected to a complete clinical examination including measurement of weight and height. BMI was calculated as the weight in kilograms divided by the square of the height in meters (kg/m 2 ), systolic and diastolic blood pressures were measured using a mercury sphygmomanometer with the patient in the sitting position after the patient had rested for at least 15 min; arterial hypertension was defined as a systolic blood pressure of 140 mmHg or higher, a diastolic blood pressure of 90 mmHg or higher, or treatment with antihypertensive medication.Indirect biomicroscopic fundus examination and fundus fluorescein angiography were performed for all patients to classify them according to the severity of their retinopathy.

Laboratory investigations included the following:

Glycosylated hemoglobin (HbA1c)

3 ml of venous blood was collected under completely aseptic conditions. HbA1c was measured using a spectrophotometer (Clinical Chemistry Analyzer 7; Germany).

Lipid profile and serum creatinine

Blood was collected in a plain tube without an anticoagulant and serum was separated; lipid profile and serum creatinine were measured using an autoanalyzer (Beckman Coulter Synchron CX 9; Beckman Coulter Inc., Mallaustrasse 69-73 68219, Mannheim, Germany).

Albumin/creatinine ratio

Morning 10 ml midstream urine samples were collected and were centrifuged at 3000 rpm for 10 min before analysis.

Microalbuminuria levels were measured using an autoanalyzer (Beckman Coulter Synchron CX 9; Beckman Coulter Inc.). Urine creatinine levels were measured in the first morning urine using a spectrophotometer (Clinical Chemistry Analyzer 7) and the ratio was calculated as the urinary albumin/creatinine ratio.

Serum apelin level

The RayBio Apelin-C Terminus Enzyme Immunoassay Kit (Brea, California, United States) is designed to target the C-terminus of the 77-aa apelin peptide; thus, the active forms of apelin including apelin-36 and apelin 13 can be detected.

Blood samples obtained from the patients after an overnight fasting were collected in plain test tubes without an anticoagulant, and these samples were immediately chilled in ice boxes. Then, serum samples were separated by centrifugation and stored in a deep-freeze at -20°C until they were analyzed collectively.

Detection range

0.1-1000 ng/ml.

Statistical analysis

Data were collected, tabulated, and analyzed statistically by a computer using SPSS version 10.

Two types of statistics were calculated:

(1) Descriptive: n (%), mean, SD, and range.

(2) Analytical:

Student t-test:

Quantitative data were analyzed using the Student t-test for the comparison of two groups of normally distributed variables.

The correlation coefficient test (Pearson correlation) (r-test) was used to detect the association between quantitative variables. The results of the correlation coefficient test (r-test) may indicate a positive (+) correlation (reverse) or a negative (-) correlation (inverse).

Analysis of variance test (F-test): Analysis of variance was used for the comparison of more than two groups of normally distributed variables.

P values:

Tests of nonsignificance at P value greater than 0.05.Tests of significance at P value less than 0.05.Tests of high significance at P value less than 0.001.

 Results



There was no significant difference between the four groups studied in age, sex, and BMI (P > 0.05).

The duration of diabetes was significantly higher in patients with PDR compared with those with diabetes without retinopathy (P < 0.001), and its mean was significantly higher in patients with NPDR compared with those with diabetes without retinopathy (P < 0.001) [Table 1] and [Figure 1].

Glycosylated hemoglobin (HbA1c) was significantly higher in patients with diabetes without retinopathy, NPDR, and PDR compared with the control group (P < 0.001) [Table 2] and [Figure 2].{Figure 1}{Figure 2}{Table 1}{Table 2}

Serum creatinine was significantly higher in patients with NPDR and PDR compared with the control group (P < 0.05), but there was no significant difference between patients with diabetes without retinopathy and the control group (P > 0.05) [Figure 3].{Figure 3}

The urinary albumin creatinine ratio was significantly higher in patients with diabetes without retinopathy, NPDR, and PDR compared with the control group (P < 0.05) [Table 3].{Table 3}

Serum cholesterol was significantly higher in patients with diabetes without retinopathy, NPDR, and PDR than the control group (P < 0.05). Low-density lipoprotein was significantly higher in patients with diabetes without retinopathy, NPDR, and PDR than the control group (P < 0.05). High-density lipoprotein was significantly lower in patients with diabetes without retinopathy, NPDR, and PDR than the control group (P < 0.05), and the mean value of patients with diabetes without retinopathy was significantly higher than that of patients with PDR (P < 0.05). Serum triglycerides were significantly higher in patients with diabetes without retinopathy, NPDR, and PDR than control group (P < 0.05) [Figure 4].{Figure 4}

Apelin was significantly higher among patients with diabetes without retinopathy, NPDR, and PDR than the control group (P < 0.001); the mean value of apelin was significantly higher in patients with PDR than diabetes without retinopathy (P < 0.05). Also, the mean value of apelin was significantly higher in patients with PDR than in patients with NPDR (P < 0.001) [Table 4] and [Figure 5].{Figure 5}{Table 4}

The present study showed that in the control group, there was no correlation between apelin and any of the other measurements.

Among patients with diabetes without retinopathy, there was no correlation between apelin and any of the other measurements.

Among patients with NPDR, there was no correlation between apelin and any of the other measurements.

Among patients with PDR, there was a positive correlation between apelin and serum creatinine [Figure 6].{Figure 6}

In group 3 (NPDR and PDR), there was a positive correlation between apelin and serum creatinine [Figure 7]{Figure 7}

 Discussion



DR is the most common complication of diabetes, and the leading cause of irreversible visual loss in individuals of working age in the USA [1].

The progression of ischemic retinal diseases, such as DR, is associated closely with pathological retinal angiogenesis [10]. Angiogenesis is the formation of new blood vessels from pre-existing vasculature and it is a pathologic hallmark of PDR, which in turn has made the control of retinal neovascularization a promising therapeutic target for patients with PDR [15].

Apelin is a peptide isolated from bovine stomach extracts that acts as an endogenous ligand for the previously orphaned G protein-coupled receptors [angiotensin 1-related putative receptor (APJ)] [8].

The apelin receptors (APLNR) are expressed in endothelial cells at the leading edge of vessels during early embryogenesis [8] and apelin in combination with VEGF induces the proliferation of endothelial cells and the formation of new blood vessels (angiogenesis) [9],[10].

The aim of this work was to study serum apelin in patients with DR and its possible pathophysiological role in such patients.

In the current study, the multivariate analysis of PDR and other parameters showed that the mean values of serum triglycerides, serum apelin, low-density lipoprotein, and the duration of diabetes were the most significant in terms of the other parameters.

These results are not in agreement with those of Yonem et al. [16], who found that the values of triglyceride, serum cholesterol, high density-lipoprotein, and low-density lipoprotein cholesterol were similar in all the groups (PDR, NPDR, and diabetes without retinopathy) and were statistically insignificant.

The duration of diabetes is the strongest predictor for the development and progression of retinopathy; the prevalence of PDR was 0% at 3 years and increased to 25% at 15 years from the onset of diabetes mellitus [17].

Yonem et al. [16] reported that patients with a duration of diabetes more than 10 years were 32 times more likely to develop nonproliferative retinopathy and 2 × 10 8 times more likely to develop proliferative retinopathy than patients with a duration of diabetes of less than 5 years; this may probably be related to the magnitude or prolonged exposure or both to hyperglycemia, coupled with other risk factors.

In the present study, the mean value of serum apelin was significantly higher in patients with PDR compared with patients with diabetes without retinopathy and NPDR.

These results are in agreement with those of Kasai et al. [10], who studied the role of endogenous apelin in pathological retinal angiogenesis. In the OIR model, vaso-obliteration of the central retinal vessel during the hyperoxic phase was accompanied by subsequent upregulation of VEGF. Apelin was markedly increased during the hypoxic phase, and this increase in the expression of apelin was higher than that of VEGF and similar to that of erythropoietin. APJ in the retinas of the OIR model mice was highly expressed in capillary endothelial cells, most of which were proliferative. Although the retinas of wild-type mice showed a significant increase in capillary density accompanied by the growth of abnormal vessels in the hypoxic phase, there was no increase in capillary density and abnormal vessels in the retinas of apelin-knockout mice despite upregulation of VEGF and Epo mRNA. Furthermore, apelin siRNA suppressed the proliferation of endothelial cells independent of the VEGF/VEGFR2 signaling pathway. These results strongly suggest that the apelin/APJ system is a prerequisite in pathological retinal angiogenesis [10].

Apelin/APJ signals probably function as an angiogenic factor in retinal vasculature. Retinal neovascularization occurring as a complication of diabetes mellitus can cause vision loss and blindness [18].

The identification of novel molecules involved in retinal angiogenesis may be a new target to suppress retinal neovascularization in diabetes and other ocular diseases and suggests that apelin/APJ could be candidate molecules physiologically or pathologically regulating retinal angiogenesis [10].

Toa et al. [7] and Nada et al. [18] showed that the concentration of apelin in the vitreous of patients with PDR was significantly higher than the level of apelin in the vitreous of patients without diabetes. In the patients with PDR, the vitreous concentrations of apelin were not significantly associated with the vitreous concentrations of VEGF or with the apelin concentrations in the plasma. Correspondingly, the plasma concentrations of apelin did not vary between the study group of patients with PDR and the control group of nondiabetic patients.

The finding of increased intraocular apelin concentrations, in contrast to normal apelin concentrations in the plasma, was paralleled by immunohistochemical findings of positive immunofluorescence staining of apelin and APJ in the endothelial cells of fibrovascular membranes of the patients with PDR, and it was paralleled by a significantly higher expression of apelin mRNA and APJ mRNA in the fibrovascular membranes of patients with PDR than in the membranes of patients with idiopathic epiretinal membranes. This is consistent with previous studies of Kasai et al. [19], Kasai et al. [10], and Deguan et al. [20] that suggested a relationship between apelin and angiogenesis and in the development of PDR.

Immunohistochemistry in the Tao et al. [7] study also showed an immunofluorescence staining of apelin and APJ in the endothelial cells of the fibrovascular membranes of the patients with PDR; this may lead one to infer that the increased apelin concentrations in the vitreous of the eyes with PDR were because of the local production of apelin, and it would suggest that apelin potentially plays a role in the retinal neovascularization and in the development of PDR.

A study of Qian et al. [15] reported no significant differences in the concentrations of vitreous and plasma apelin between two groups: the control group that was subjected to vitrectomy without preoperative intravitreal bevacizumab and the other group that received preoperative intravitreal bevacizumab. Immunohistochemistry showed positive immunofluorescence staining of apelin on endothelial cells in the fibrovascular membranes of both groups. The results suggest that apelin might contribute toward the formation of fibrovascular membranes during the development of PDR and apelin may not be directly regulated by VEGF; consequently, apelin signaling could represent a new promising therapeutic target during pathologic neovascularization associated with DR and associated with the development of epiretinal membranes in PDR and may not directly correlate with VEGF [15].

In the study by Erdem et al. [21] plasma concentrations of apelin were significantly lower in patients with newly diagnosed type II diabetes mellitus than in healthy control patients. In another investigation, however, the basal plasma apelin concentrations were significantly higher in patients with impaired glucose tolerance and in diabetic patients than in nondiabetic control patients. The reasons for the discrepancies between the studies may be differences in the patient inclusion criteria and in the study composition, such as the inclusion of patients with advanced diabetes mellitus with proliferative retinopathy as in our study, in contrast to patients with newly diagnosed type II diabetes mellitus in the study by Erdem et al. [21].

However, Yonem et al. [16] found that apelin levels were similar in all the groups (PDR, NPDR, and diabetic without retinopathy).

In our study, in the control group, patients with diabetes without retinopathy, and patients with NPDR, there was no correlation between apelin and any of the other measurements, but among patients with PDR, there was a positive correlation between apelin and serum creatinine.

Zhang et al. [22] reported that serum apelin levels are correlated positively with microalbuminuria (early nephropathy) in patients with type II diabetes. These results indicated that apelin/APLNR might be a promoting factor for diabetic nephropathy.

 Conclusion



Serum apelin is significantly higher in patients with PDR compared with patients with nonproliferative retinopathy and patients with diabetes without retinopathy, and apelin may play a role in the development of PDR; inhibition of the apelinergic system could offer new therapeutic opportunities against ischemic retinopathy.

Serum apelin significantly correlated with serum creatinine in patients with retinopathy, which may indicate the role of apelin in diabetic nephropathy.

 Acknowledgements



Conflicts of interest

No conflict of Interest.

References

1Yang X, Zhu W, Zhang P, et al. Apelin-13 stimulates angiogenesis by promoting cross-talk between AMP-activated protein kinase and Akt signaling in myocardial microvascular endothelial cells. Mol Med Rep 2014; 5 :1590-1596.
2 Sivaprasad S, Gupta B, Crosby-Nwaobi R, Evans J. Prevalence of diabetic retinopathy in various ethnic groups a worldwide perspective. Surv Ophthalmol 2012; 4 :347-370.
3 Tang J, Kern T. Inflammation in diabetic retinopathy. Prog Retin Eye Res 2011; 30:343-358.
4 Abdulla W, Fawzi A. Anti-VEGF therapy in proliferative diabetic retinopathy. Int Ophthalmol Clin 2009; 49 :95-107.
5 Wang H, Liang B, Peng X, et al. Risk factors for diabetic retinopathy in a rural Chinese population with type 2 diabetes. The Handan Eye Study. Acta Ophthalmologica 2011; 4 :336-343.
6 Laurell I, Dray C, Knauf C, Duparc T, Knauf C, Valet P. Apelin, diabetes, and obesity. Endocrine 2011; 40 :1-9.
7 Tao Y, Lu Q, Jiang Y et al. Apelin in plasma and vitreous and in fibrovascular retinal membranes from patients with proliferative diabetic retinopathy. Invest Ophthalmol Vis Sci 2010; 8 :4237-4241.
8 Sato K, Takahashi T, Kobayashi Y, Hagino A, Roh S, Katoh K. Apelin is involved in postprandial responses and stimulates secretions of arginine-vasopressin, adrenocorticotropic hormone and growth hormone in the ruminant. Domest Anim Endocrinol 2012; 42 :165-172.
9 Cox CM, D′Agostino SL, Miller MK, Heimark RL, Krieg PA. Apelin, the ligand for the endothelial G-protein-coupled receptor, APJ, is a potent angiogenic factor required for normal vascular development of the frog embryo. Dev Biol 2006; 296 :177-189.
10Kasai A, IshimaruY , Kinjo T, SatookaT, MatsumotoN, Yoshioka Y, et al. Apelin is a crucial factor for hypoxia-induced retinal angiogenesis. Arterioscler Thromb Vasc Biol 2010; 30 :2182-2187.
11Watanabe D, Suzuma K, Matsui S, Kurimoto M, Kiryu J, Kita M, et al. Erythropoietin as a retinal angiogenic factor in proliferative diabetic retinopathy. N Engl J Med 2005; 353 :782-792.
12Chen J, Connor M, Aderman M, Willett L, Aspegren P, Smith E. Suppression of retinal neovascularization by erythropoietin siRNA in amouse model of proliferative retinopathy. Invest Ophthalmol Vis Sci 2009; 50 :1329 -1335.
13Simó R, Hernández C. Intravitreous anti-VEGF for diabetic retinopathy: hopes and fears for a new therapeutic strategy. Diabetologia 2008; 51 :1574-1580.
14Zhao T, Lu Q, Tao Y, Liang XY, Wang K, Jiang YR. Effects of apelin and vascular endothelial growth factor on central retinal vein occlusion in monkey eyes intravitreally injected with bevacizumab: a preliminary study. Mol Vis 2011; 17 :1044-1055.
15Qian J, Lu Q, Tao Y, Jiang YR. Vitreous and plasma concentrations of apelin and vascular endothelial growth factor after intravitreal bevacizumab in eyes with proliferative diabetic retinopathy. Retina 2011; 31 :161-168.
16Yonem A, Duran C, Unal M, Ipcioglu OM, Ozcan O. Plasma apelin and asymmetric dimethylarginine levels in type 2 diabetic patients with diabetic retinopathy. Diabetes Res Clin Pract 2009; 84 :219-223.
17American Diabetes Association. Standards of medical care in diabetes; 2006.
18Nada W, Abdel Moety D, Abo Almeaty M, Hadhoud K. Evaluation of the role of apelin in diabetic retinopathy. Egypt Ophthalmol Soc 2014; 4 :245-248.
19Kasai A, Shintani N, Oda M, et al. Apelin is a novel angiogenic factor in retinal endothelial cells. Biochem Biophys Res Commun 2004; 325 :395-400.
20Deguan L, Hening S, Chen L. Apelin and APJ a novel critical factor and therapeutic target for a therosclerosis. Acta Biochemica Biophysica Sinica 2013; 7 :527-533.
21Erdem G, Dogru T, Tasci I, Sonmez A, Tapan S. Low plasma apelin levels in newly diagnosed type 2 diabetes mellitus. Exp Clin Endocrinol Diabetes 2008; 116 :289-292.
22Zhang B, Wang W, Wang H, Yin J. Promoting effects of the adipokine apelin on diabetic nephropathy. Plos One 2013; 4 :6-45.