Objectives: Apelin, a cytokine mainly secreted by adipocytes and several tissues, includes the gastrointestinal tract, adipose, brain, kidney, liver, lung, and various sites within the cardiovascular system. Apelin is closely related to glucose metabolism, and has been proposed to be a promising therapeutic agent in treating insulin resistance. Apelin and orphaned G-protein–coupled apelin exhibit roles in regulating fluid homeostasis. Circulating serum apelin suppresses insulin secretion by binding to the G-protein–coupled apelin receptor on B cells of islets of Langerhans. Several studies also have documented the altered level of serum apelin in type 2 diabetic patients, but the results remain controversial. This study sought to analyze apelin levels in new-onset diabetes after transplant.
Materials and Methods: Forty-seven diabetic renal transplant recipients were compared with 40 nondiabetic renal transplant recipients. Data were collected for positive family history of diabetes, body weight, body mass index, blood pressure, and blood chemistry including apelin level. Logistic multiple analysis were made for statistically significant data on univariate analysis.
Results: Apelin levels were significantly higher among obese, hypercholesterolemia new-onset diabetes after transplant patients, 428.7 ± 193.29, 256.8 ± 128 (P > .001). There was appositive cor-relation between serum apelin and proteinuria.
Conclusions: Serum apelin has a high level in new-onset diabetes after transplant, than nondiabetic patients, and they positively correlate with proteuria in new-onset diabetes after transplant patients.
Key words : Renal transplant, Posttransplant diabetes mellitus, Apelin, Adipocytes
Adipocytokines related to adhesion molecules might support the importance of inflammation/endothelial cell injury in the pathogenesis of atherosclerosis and its consequences in chronic kidney disease.1 Apelin, a cytokine mainly secreted by adipocytes, is closely related to glucose metabolism, and has been proposed as a promising therapeutic agent for treating insulin resistance. Apelin is expressed in several tissues, including the gastrointestinal tract, adipose, brain, kidney, liver, lung, and various sites within the cardiovascular system.2 Consistent with this pattern of distribution, apelin, and orphaned G-protein – coupled apelin (APJ) – exhibit roles in regulating fluid homeostasis.3
Studies suggest that the most potent isoform of the apelin is pyroglutamated form of apelin-13 (Pyro apelin-13), which is considered to be the principle active biological ligand.4 Circulating serum apelin suppresses insulin secretion by binding to the APJ receptor on B cells of islets of Langerhans.5 Apelin production by adipocytes is decreased during fasting and increases again in response to eating or insulin injection.2 In addition to its effects on pancreatic cells, apelin decreases the blood pressure by inducing nitrous oxide-dependant vasodilation and suppressing thirst.3 Several studies also have documented the altered level of serum apelin in type 2 diabetic patients, but the results remain controversial.6 New-onset diabetes mellitus after transplant is a serious and common complication after solid-organ transplant; kidney transplant recipients who develop new-onset diabetes after transplant have variably been reported to be at increased risk of fatal and nonfatal cardiovascular events and other adverse outcomes including infection, reduced patient survival, and graft rejection.7
Materials and Methods
Forty-seven diabetic renal transplant recipients were compared with 40 nondiabetic renal transplant recipients at Mansoura Urology and Nephrology Center regarding positive family history of diabetes, body weight, body mass index, pretransplant dialysis duration, intraoperative ischemia time, blood pressure, blood chemistry, complete blood count, and apelin level.
Apelin level was done by morning venous blood sample. We obtained apelin (3 mL on EDETA) and centrifuged it at 3000 rpm for 10 minutes to separate plasma in an Eppendorf tube and placed at -20°C to measure apelin-12 level using a solid-phase enzyme linked immunosorbent assay (Glory Science Co., Ltd; Del Rio, TX, 78840 USA). The kit uses a double antibody sandwich enzyme-linked immunosorbent assay to assay the level of human apelin-12 in plasma.8
Monitoring for new-onset diabetes after transplant
All patients had blood glucose measurements performed routinely during initial hospitalization at the time of transplant. Blood glucose measurements were made every 12 hours for the first 3 days after the transplant. Thereafter, blood glucose levels were measured daily while the recipient was an inpatient, twice weekly for the first posttransplant month as an outpatient, weekly for the second posttransplant month, and whenever patients had routine chemistry studies thereafter. New-onset diabetes after transplant was diagnosed according to the American Diabetes Association Guidelines, 2011. Briefly, diagnostic criteria include (1) symptoms of diabetes plus random plasma glucose ≥ 200 mg/dL (11.1 mmol/L), A1C ≥ 6.5 percent (48 mmol/mol) (symptoms include polyuria, polydipsia, and unexplained weight loss); (2) fasting plasma glucose ≥ 126 mg/dL (7.0 mmol/L) (fasting is defined as no caloric intake for at least 8 hours) and (3) two-hour plasma test in which the glucose is ≥ 200 mg/dL (11.1 mmol/L) during an oral glucose tolerance test.9
Qualitative data were displayed in cross tabulation and quantitative data were described in terms of arithmetic mean ± SD. Bivariate techniques were used for initial evaluation of contrasts. Thus, the chi-square and Fisher exact tests were used to compare the qualitative variables, and the unpaired t test was used to compare the means 2 quantitative variables. A P value less than .05 was considered statistically significant. All analyses were carried out using the computer package SPPS for windows (version 10, SPSS Inc. Chicago, IL, USA). The data were expressed as mean ± SD. Means were compared by the t test or the nonparametric test if the variable was not normally distributed. Logistic multiple analysis were made for statistically significant data on univariate analysis.
Apelin levels in the new-onset diabetes after transplant group were significantly higher than those of the non-diabetic group (428.7 ± 193.29 vs 256.8 ± 128 pg/mL, P < .001). Regarding the demographics data of different groups, family history of diabetes mellitus and body mass index show a high statistical significance (P = .03, P < .001); new-onset diabetes after transplant patient had a higher body mass index (28.9 ± 2.8). Serum apelin correlated significantly with fasting blood sugar and proteinuria (P < .001, P = .001). Also there was a statistical significance regarding electrocardiogram changes (P = .01) and hypertensive treatment (P = .001), although there was no statistical significance regarding pretransplant hemodialysis duration and intraoperative ischemia time (Table 1).
Oral hypoglycemic drugs had the highest percentage regarding treatment of diabetes mellitus in the new-onset diabetes after transplant patients (70.3%) (Table 2). The nondiabetic group had a better graft function at last follow-up with no significance (Table 3).
In our study, we report that diabetic kidney allograft recipients show a higher apelin level than do their nondiabetic counterparts; this finding is similar to results reported.10,11 Apelin showed to be closely related to insulin resistance and glucose uptake metabolism, the underlying mechanism of how apelin affects insulin resistance is poorly understood. Elucidating a molecular role for apelin in insulin resistance is extremely important. Insulin-stimulated glucose uptake is extremely important, which was defined as insulin sensitivity, and is considered to be the criterion standard for evaluating insulin resistance at the cellular level. Cells with a significant reduction in insulin-stimulated glucose uptake have commonly been accepted to be in an insulin-resistant state. Furthermore, apelin preincubation with p13k specific inhibitor also increased adipocytes and insulin-stimulated glucose uptake.6
We found that serum level of apelin is lower in the nondiabetic group who had undergone a shorter length of hemodialysis pretransplant, our results agree with those reported by El-Shehaby and associates, who, in 2010, studied the plasma level of apelin in patients with hemodialysis to assess the effect of renal transplant and dialysis session on plasma apelin and found that plasma apelin levels were significantly lower in hemodialysis patients compared with controls while levels increased significantly in the early posttransplant period.12
Leal and associates found no difference between apelin-36 levels in hemodialysis patients and healthy subjects. In contrast apelin-12 levels were significantly higher in patients compared with healthy subjects. It is unknown why apelin levels may be decreased in allograft recipients or patients undergoing dialysis. A role of the kidney in metabolizing apelin has not been investigated.13
Yilmaz and associates found that endothelial dysfunction in type-2 diabetics with early diabetic nephropathy is associated with low circulating adiponectin. Also, apelin receptor mRNA and protein expression significantly decreased in both ischemic and nonischemic kidneys experimentally.14 Apelin may have opposite effects on the activity of NOS and production of nitrous oxide.15 Our results are not helpful to study the relation between renal ischemia and apelin level as both groups have almost equal intraoperative ischemia time with no statistical significance.
The common early signs of diabetic nephropathy are microalbuminuria and overt proteinuria. Our results indicate that serum apelin levels positively correlate with proteinuria in patients with new-onset diabetes after transplant; this is in accordance with Zhang and associates.16 It is generally accepted that endothelial dysfunction is important for diabetic microvascular disease.17 However, there has been little direct evidence of a causative link among endothelial dysfunction, microvascular disease, and diabetic end-organ damage. Microvascular disease can lead to organ damage through impaired vascular function, increased inflammation, or increased apoptosis.18-20 The growth of new blood vessels and increased permeability of microvessels in nephrons are believed to be the main pathogenesis of diabetic nephropathy and glomerular endothelial cells incubated with apelin were more permeable to fluorescein isothiocyanate labeled bovine serum albumin. The observed increase of glomerular permeability with apelin may, therefore, be relevant during the early stages of diabetic nephropathy; thereby supporting the concept that endothelial dysfunction is causally linked to diabetic nephro-pathy.21
In our study, for the ischemic electrocardiogram changes are more common in the diabetic group with higher apelin level; several investigators suggested an association between apelin and heart diseases. Apelin is expressed in cardiac tissue and although not fully clarified, the endothelium of the cardiac vasculature seems to be the dominant cellular source.22 Whether the endothelium in other organs also expresses the apelin gene should be examined, as apelin secretion from the peripheral endothelium cannot be excluded. However, studying the racemic status23 was the determinant for decreased plasma apelin content among heart involvement. In contrast, the recent studies reported by Sheikh and associates24 demonstrated that apelin and APJ were up-regulated in the heart and skeletal muscle after myocardial injury, suggesting that apelin expression remains in the endothelium.
In vitro experiments with cultured endothelial cells showed apelin mRNA and protein levels are increased in hypoxia through a hypoxia-inducible-factor–mediated pathway. The correlation between apelin and some endothelial cell injury markers may be explained by the hypoxia, owing to anemia and other factors among patients on renal replacement therapy. Studies suggest that apelin-expressing endothelial cells respond in conditions associated with heart failure, possibly including local tissue hypoxia, thereby modulating apelin-APJ expression to regulate cardiovascular homeostasis, the apelin-APJ pathway may thus provide a mechanism for systemic endothelial monitoring of tissue perfusion and adaptive regulation of cardiovascular function.24
Because apelin improves glucose uptake and mediates the inflammatory response, apelin may be a promising therapeutic agent for type 2 diabetes. However, further studies that investigate the protein and mRNA levels of the anti-insulin resistant effect of apelin, and in vivo studies of apelin in insulin- resistant animals are necessary to elucidate its pharmaceutical potential.
Our study has some limitations: pretransplant apelin levels are not performed. Also, follow-up levels may be valuable. Serum apelin has a high level in new-onset diabetes after transplant. Patients other than nondiabetic patients and positively correlate with proteinuria in new-onset diabetes after transplant patients. We recommend a randomized, controlled, evaluation of apelin levels pretransplant and periodically postoperative in accordance with other laboratory investigations as plasma glucose, urinalysis (proteinuria), lipid profile, and HbA1c.
Volume : 13
Issue : 4
Pages : 319 - 323
DOI : 10.6002/ect.2014.0276
From the 1Department of Dialysis and Transplantation, The Urology,
Nephrology Center, Mansoura University; the 2Department of Clinical
Pathology, Faculty of Medicine, Mansoura University; and the Department of
3Clinical Pathology, The Urology, Nephrology Center, Mansoura University,
Acknowledgements: The authors declare that they have no sources of funding for this study, and they have no conflicts of interest to declare.
Corresponding author: Ayman Maher Nagib, MD, Urology & Nephrology Center, Mansoura University, El Gomhoria Street, PO Box: 35516 Mansoura, Egypt
Phone: +20 5 022 62222
Fax: +20 5 022 3717
Table 1. Comparison of Different Parameters Between Groups
Table 2. Treatment of Diabetes Mellitus
Table 3. Graft Function at Last Follow-Up