Renalase gene polymorphism is associated with increased blood pressure in preeclampsia

Pregnancy Hypertension: An International Journal of Women’s Cardiovascular Health 6 (2016) 115–120

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Renalase gene polymorphism is associated with increased blood pressure in preeclampsia Binnur Bagci a,b,⇑, Savas Karakus c, Gokhan Bagci d, Enver Sancakdar d a

Department of Nutrition and Dietetics, Faculty of Health Sciences, Cumhuriyet University, 58140 Sivas, Turkey Advanced Technology Research Center (CÜTAM), Cumhuriyet University, 58140 Sivas, Turkey c Department of Obstetrics and Gynecology, Faculty of Medicine, Cumhuriyet University, 58140 Sivas, Turkey d Department of Biochemistry, Faculty of Medicine, Cumhuriyet University, 58140 Sivas, Turkey b

a r t i c l e

i n f o

Article history: Received 15 March 2016 Accepted 18 April 2016 Available online 19 April 2016 Keywords: Renalase Polymorphism Preeclampsia Blood pressure

a b s t r a c t Background: Renalase is a novel enzyme that degrades circulating catecholamines. We aimed to investigate the role of rs2576178 and rs10887800 polymorphisms of the renalase gene in preeclampsia (PE) patients Methods: This case-control study consisted of 110 women with PE and 102 normotensive controls. PCRRFLP method was used for determination of renalase gene polymorphisms. Results: Allele frequency and genotype distribution of rs10887800 polymorphism were found statistically significantly higher in women with PE (p < 0.05). Also G allele and GG genotype of rs10887800 polymorphism were found higher in women with severe PE than that of mild PE (p < 0.05). There was no significant difference for rs2576178 polymorphism in terms of allele frequency and genotype distribution (p > 0.05). In PE patients, systolic blood pressure (SBP) means according to rs10887800 genotypes were found statistically significantly higher (GG vs AA; p = 0.001) and (GG vs GA; p = 0.001). Similarly, diastolic blood pressure (DBP) means were found statistically significantly higher in PE patients (GG vs GA: p = 0.001); (GG vs AA: p = 0.004). For rs2576178 polymorphism, SBP means were found as (GG vs AA; p = 0.012, GG vs GA; p > 0.05) in PE patients. DBP means were not significant according to rs2576178 genotypes in PE patients (p > 0.05). Conclusions: The findings of the present study suggest that blood pressure may be increased by GG genotype and G allele of rs10887800 polymorphism and the polymorphism may increase the susceptibility to PE. Ó 2016 International Society for the Study of Hypertension in Pregnancy. Published by Elsevier B.V. All rights reserved.

1. Introduction Hypertensive disorders account for approximately 10% of all pregnancies and involve in a wide range of situations including chronic hypertension, gestational hypertension, eclampsia, hemolysis, elevated liver enzymes and low platelets (HELLP) syndrome and preeclampsia (PE) [1–3]. PE is defined as the onset of hypertension and proteinuria after the second halfs of the pregnancy [2,4,5]. It is one of the most common medical complications of pregnancy and affects about 2–8% of all pregnancies [6,7]. Despite the developments in antenatal and neonatal care, PE continues to be one of the major causes of maternal and neonatal morbidity and mortality worldwide [8–10].

⇑ Corresponding author at: Department of Nutrition and Dietetics, Faculty of Health Sciences, Cumhuriyet University, 58140 Sivas, Turkey. E-mail address: [email protected] (B. Bagci).

Experimental studies have provided clear evidence that eclampsia and PE are states of sympathetic hyperactivity [11,12]. Both sympathetic nervous and sympathetic adrenal activities are increased in patients with PE [12–14]. The catecholamine concentration has been found to be increased in PE and eclampsia [15,16]. Arterial epinephrine was shown to be increased in PE [12]. It was shown that increased plasma epinephrine level correlated with increased blood pressure (BP) in the development of PE [17]. In the regulation of BP, catecholamines like dopamine, norepinephrine, and epinephrine play a key role. Monoamine oxidase-A (MAO-A), monoamine oxidase-B (MAO-B) and catechol-O-methyl transferase (COMT) are the responsible intracellular enzymes for degradation of catecholamines [18]. In the last decade, researchers identified a novel soluble flavin adenine dinucleotide (FAD)-dependent amine oxidase called as renalase [19]. Renalase is mainly expressed in the kidney and also found in the heart, small intestine and skeletal muscle [19]. Renalase metabolizes catecholamines and catecholamine-like substrates [20].

http://dx.doi.org/10.1016/j.preghy.2016.04.002 2210-7789/Ó 2016 International Society for the Study of Hypertension in Pregnancy. Published by Elsevier B.V. All rights reserved.

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It shows significantly high activity against catecholamines such as epinephrine, dopamine and norepinephrine, but shows little or no activity towards physiologically occurring amines such as serotonin, benzylamine, tyramine, methylamine [21]. The gene encoding human renalase is called as (RNLS) and located on chromosome 10q23.33 and formed 10 exons and 309,469 base pairs (bp). The human renalase protein is 342 amino acids long, and consists of an amine oxidase domain, a FAD-binding region, and a signal peptide [19]. The kidney is primarily responsible organ for maintaining steady levels of the renalase. Renalase circulates in the blood in an inactive form called as prorenalase. Prorenalase is secreted into the blood by the kidney. In the basal state, prorenalase has no amine oxidase activity, and it is rapidly converted to renalase by increased catecholamines and heightened BP. Renalase decreases the BP by degrading the catecholamines including dopamine, epinephrine and norepinephrine [22]. Several animal experiments were designed to clarify the role of renalase in catecholamine degradation and BP regulation [20,23–25]. Recent data have indicated that deficiency of renalase associated with heightened BP and increased circulating catecholamine levels [26,27]. Two polymorphisms selected for the present study are located in the putative functional regions. rs2576178 polymorphism is located at the 50 flanking region and rs10887800 polymorphism is located at the intron 6, near the border of exon/intron, and consequently these polymorphisms might affect regulation and expression of the RNLS gene [28]. To the best of our knowledge, polymorphisms in the renalase gene (rs2576178, rs10887800) were found to be associated with hypertension [28,29], stroke, type 2 diabetes [28], end-stage renal disease (ESRD) [30], coronary heart disease [31], and pregnancy induced hypertension (PIH) [32]. To date, there is no study investigating the role of renalase gene polymorphisms in PE patients. Although the signs and symptoms of the disease are well known, the etiology is still unknown. Therefore it could not possible to prevent the disease [6]. Studies carried out to date have focused on the pathophysiology, prevention and treatment of the disease [6,7]. In the light of this information, we aimed to investigate the possible role of rs2576178 and rs10887800 polymorphisms of the renalase gene on the development of preeclampsia, and to show their possible associations with demographic parameters and laboratory findings, primarily blood pressure, and to find whether there is a relationship with disease severity.

2. Materials and methods This case-control study consisted of 110 women with preeclampsia and 102 normotensive controls. All subjects had been admitted to Obstetrics & Gynecology clinic, Cumhuriyet University Hospital between April 2015 and October 2015 included in the study. All subjects of the present study were Caucasians of Turkish origin. The diagnosis of preeclampsia was determined according to the International Society for the Study of Hypertension in Pregnancy’s (ISSHP) criteria [33]. The following criteria were used for the diagnosis of PE: systolic blood pressure (SBP) and diastolic blood pressure (DBP) shold be above 140/90 mmHg, at least two measurements, minimum 4 h apart, and proteinuria should be above 300 mg per 24 h. Pregnant women who did not meet these criteria were excluded from the study. The control group were recruited from the same center and consisted of normotensive volunteer pregnant women with at least one pregnancy and no history of PE. Ethics Committee of Cumhuriyet University, Faculty of Medicine approved the present study. The present study was conducted in accordance with the Declaration of Helsinki ethical principles. A written, informed consent was obtained from all subjects.

Patients with PE included in the present study were categorized into 2 groups based on the disease severity. Severe PE was defined as having one or more of the following criteria: SBP/ DBP P 160/110 mmHg on two occasions 4 or more hours apart in a pregnant woman on bed rest and having proteinuria with excretion P3 g in 24 h urine sample, visual disturbances, headache, upper abdominal pain, elevated levels of serum creatinine and transaminases, thrombocytopenia, fetal-growth restriction. Mild PE has determined as SBP/DBP between 140–160/90– 110 mmHg and having proteinuria <3 g in 24 h urine sample [3]. The exclusion criteria for preeclamptic patients and controls were as follows: gestational hypertension, diabetes mellitus, gestational diabetes mellitus, multiple pregnancy, isolated proteinuria, thrombocytopenia, elevated levels of transaminases with no hypertension, maternal chronic and inflammatory disorders, stillbirth, intrahepatic cholestasis of pregnancy, and maternal hepatitis. Demographic parameters including maternal age, gestational age, gravidity, parity, SBP and DBP, body mass index (BMI), prior PE history, family history of PE, chronic hypertension and diabetes mellitus (DM) were collected from patients and controls. Fasting blood glucose (FBG), hematocrit (HCT), hemoglobin (Hgb), creatinine, blood urea nitrogen (BUN), white blood cells count (WBC), platelet count (PLT), alanine aminotransferase (ALT), aspartate aminotransferase (AST), urinary dipstick test were collected from routine laboratory tests of the subjects. 2.1. Genotyping PCR-RFLP method was used for determination of renalase gene polymorphisms (rs2576178, rs10887800). Peripheral venous blood samples (3 ml) were taken into tubes with K3EDTA from patient and control subjects. These tubes are stored 20 °C until genetic analysis time. Total genomic DNA was extracted from peripheral venous blood with according to salting out procedure [34]. Template DNAs were amplified for detection of renalase gene polymorphisms (rs2576178, rs10887800). Same PCR protocol was used for the detection of both polymorphisms. PCR were carried out in a total reaction mixture of 25 ll containing 150–300 ng of genomic DNA, 12.5 ll of PCR master mix (Fermentas, Lithuania), 9,5 ll of ddH2O and 10 pmol of each primer (1 ll). Following primers were used for amplification reaction: for rs2576178 polymorphism, forward: 50 -AGCAGAGAAGCAGCTTAACCT-30 , reverse: 50 -TATCTGCAAGTCAGCGTAAC-30 ; and for rs10887800 polymorphism, forward: 50 -CAGGAAAGAAAGAGTTGACAT-30 , reverse: 50 -AAGTTG TTCCAGCTACTGT-30 . Amplification of two polymorphism on the renalase gene was performed an initial denaturation step at 94 °C for 5 min followed by 40 cycles of denaturation step at 94 °C for 45 s, annealing step at 58 °C for 1 min, elongation step at 72 °C for 2 min and followed by a final elongation step at 72 °C for 5 min. Restriction digestion of PCR products for rs2576178 polymorphism was performed in 10 ll volume using 5 units of MspI restriction endonuclease (New England Biolabs, UK) at 37 °C for 16 h. Digestion products were analyzed by electrophoresis on a 2% agarose gel in TAE buffer, and visualized using ethidium bromide staining. Samples with a single 525 bp band were identified as wild type AA genotype while samples with 2 bands (423, 102 bp) were identified as homozygous GG genotype and those with 3 bands (525, 423 and 102 bp) were identified as heterozygous AG genotype. Restriction digestion for rs10887800 polymorphism was performed in 10 ml volume using 10U of PstI restriction endonuclease (New England Biolabs, UK) at 37 °C for 16 h. Digestion products were electrophoresed on a 2% agarose gel in TAE buffer and visualized using ethidium bromide staining. Samples with a single 554 bp band were identified as wild type AA genotype while samples with 2 bands (415, 139 bp) were iden-

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tified as homozygous GG genotype and those with 3 bands (554, 415 and 115 bp) were identified as heterozygous AG genotype.

Table 2 Biochemical and hematological parameters of patients with preeclampsia and controls.

2.2. Statistical analysis

Variables

Preeclampsia (n = 110)

Controls (n = 102)

P

Statistical Package for the Social Sciences (IBM SPSS version 22, Armonk, NY, USA) was used the statistical analysis of the present study. Descriptive values were demonstrated as the mean ± SD. Independent samples t-test was performed to compare the means of demographic, biochemical and hematological parameters of women with PE and controls. Chi-square test or Fisher’s exact test were carried out to compare genotype distributions in all groups. Allele frequencies were calculated by allele counting method. One-way ANOVA with Tukey test were used to compare SBP and DBP means of women with PE according to rs10887800 and rs2576178 genotypes. A p value <0.05 was considered as statistically significant. We calculated the odds ratio (OR) with 95% confidence intervals (CIs).

White blood cells (1000/mL) Hematocrit (%) Hemoglobin (g/dL) Platelet count (1000/lL) BUN (mg/dL) Creatinine (mg/dL) ALT (U/L) AST (U/L) FBG (mg/dL)

11.72 ± 3.62 37.72 ± 3.49 12.6 ± 1.36 225.35 ± 74.64 10.44 ± 6.11 0.77 ± 0.97 32.33 ± 78.89 49.95 ± 110.51 98.64 ± 26.58

11.21 ± 3.04 36.24 ± 3.84 11.98 ± 1.45 231.98 ± 64.84 6.51 ± 2.02 0.50 ± 0.08 12.55 ± 5.26 19.34 ± 4.85 89.73 ± 25.27

0.273 0.004 0.002 0.49 <0.001 0.006 0.012 0.006 0.013

3. Results The total number of individuals included in this study is 212. The patient group and control group were consisted of 110 preeclamptic and 102 normotensive women, respectively. The preeclamptic group comprise of 23 patients including 2 patients with eclampsia, 15 patients with HELLP syndrome, 4 patients with intrauterine growth restriction (IUGR), and 2 patients with preterm labor (PL). Demographic parameters of patients with PE and controls were shown in Table 1. There was no difference in terms of age, BMI, gestational age, gravidity, and parity between patients and controls. As expected, both SBP mean (PE 161.67 ± 15.04 vs control group 109.83 ± 10.21; p < 0.001) and DBP mean (PE 104.23 ± 12.43 vs control group 71.03 ± 10.3; p < 0.001) were statistically significantly different when patients compared to controls. Biochemical and hematological data of the patients and control group were shown in Table 2. There was no statistically significant difference between preeclamptic patients and controls according to WBC and PLT count. However, HCT (p = 0.004), Hgb (p = 0.002), BUN (p < 0.001), creatinine (p = 0.006), ALT (p = 0.012), AST (p = 0.006) and FBG (p = 0.013) were found statistically significantly higher between preeclamptic patients and controls. Genotype distributions and allele frequencies of rs10887800 and rs2576178 polymorphisms in patients and controls were shown in Table 3. Genotype distribution of rs10887800 in the patient group was found as 32 (29.1%), 55 (50%), 23 (20.9%) for

Table 1 Demographic features of women with preeclampsia and controls. Variables

PE (n = 110)

Controls (n = 102)

p

Age (years) BMI (kg/m2) Gestational age (week) Gravidity (number) Parity (number) SBP (mmHg) DBP (mmHg) Prior PE Family history of PE HELLP Eclampsia IUGR PL

29.75 ± 7.20 29.65 ± 6.12 35.18 ± 4.45 3.02 ± 1.82 (1–9) 1.52 ± 1.53 (0–8) 161.67 ± 15.04 104.23 ± 12.43 21 (19.1) 12 (10.9) 15 (13.63) 2 (1.81) 4 (3.63) 2 (1.81)

28.15 ± 5.67 29.13 ± 5.88 38.0 ± 3.71 2.62 ± 1.64 (1–7) 1.38 ± 1.29 (0–6) 109.83 ± 10.21 71.03 ± 10.3 0 0 0 0 0 0

0.076 0.529 0.795 0.123 0.502 <0.001 <0.001 – – – – – –

Data were presented as mean ± SD, (min–max) and percentage as appropriate. PE: preeclampsia; BMI: body mass index; SBP: Systolic blood pressure; DBP: Diastolic blood pressure; HELLP: Hemolysis, elevated liver enzyme levels and low platelets; IUGR: Intrauterine growth restriction; PL: Preterm Labor.

Data were presented as mean ± SD. P < 0.05 vs. controls and preeclampsia. BUN, blood urea nitrogen; ALT, alanine aminotransferase; AST, aspartate aminotransferase; FBG, fasting blood glucose.

AA, AG and GG, respectively. Genotype distribution of rs10887800 was found as 44 (43.1%), 47 (46.1%) and 11 (10.8%) for AA, AG and GG, in the control group respectively. There was statistically significant difference in terms of rs10887800 genotype distribution between preeclamptic patients and controls (X2 = 6.46; p = 0.039). Also, there was a significant difference between patients and controls with regard to allele frequencies (X2 = 6.44; p = 0.011). rs10887800 GG genotype was found 2.18 times higher in women with PE compared with controls (GG vs AG + AA; OR = 2.18; 95% CI = 1.00–4.75; p = 0.048) and G allele increased the disease risk 1.66 times (G vs A; OR = 1.66; 95% CI = 1.12–2.46; p = 0.011). Genotype distribution of rs2576178 polymorphism in the patients was found as 60 (54.5%) for AA, 38 (34.5%) for AG and 12 (11%) for GG, and 57 (55.9%) for AA, 37 (36.3%) for AG and 8 (7.8%) for GG in the control group. When the patient and control groups were compared in terms of rs2576178 genotype distribution, no significant difference was observed (X2 = 0.59; p = 0.75). Furthermore, when the patient and control groups were compared in terms of allele frequencies, there was no significant difference for rs2576178 polymorphism (X2 = 0.37; p = 0.54) Table 3. We compared the means of following parameters according to genotypes of renalase polymorphisms both in women with PE, and in severe/mild PE: gestational age, age, gravidity, parity, BMI, WBC, HCT, Hgb, PLT, creatinine, BUN, ALT, AST, FBG. In women with PE, there was statistically significant difference only for HCT, Hgb and WBC according to rs10887800 and rs2576178 genotypes (all p values >0.05) (data were not shown). In women with severe PE compared to mild PE group, statistically significant difference was found in terms of HCT, Hgb and WBC with regard to rs10887800 genotypes (p < 0.05; data were not shown). Table 4 shows SBP and DBP means in women with PE, according to rs10887800 and rs2576178 polymorphisms. For rs10887800 AA, AG and GG genotypes, SBP means were found as 154.38 ± 14.35, 161.0 ± 14.7 and 173.43 ± 8.81, respectively (GG vs AA; p = 0.001), (GG vs GA; p = 0.001), (AG vs AA; p = 0.077). Likewise, DBP means were found as 102.50 ± 14.14, 101.55 ± 8.86 and 113.04 ± 13.71, respectively. (GG vs GA: p = 0.001); (GG vs AA: p = 0.004; (AG vs AA: p = 0.928). Similarly, for rs2576178 polymorphism, SBP means were 158.25 ± 14.01 for having AA genotype, 163.92 ± 16.62 for having AG genotype and 171.67 ± 8.34 for having GG genotype. (GG vs AA; p = 0.012, GG vs GA; p = 0.246, AG vs AA; p = 0.149). Likewise, DBP means were as 102.17 ± 10.90 for having AA genotype, 105.79 ± 14.82 for having AG genotype 109.58 ± 9.64 for having GG genotype. (GG vs AA; p = 0.141, GG vs GA; p = 0.621, AG vs AA; p = 0.333). Our patients consist of 69 mild and 41 severe preeclamptic women according to disease severity. Genotype distributions and

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Table 3 Genotype distribution and allele frequencies of rs10887800 and rs2576178 polymorphisms in patients with preeclampsia and controls. Polymorphism

Controls (n = 102)

Preeclampsia (n = 110)

p

OR

rs10887800 Genotypes AA AG GG

44 (43.1) 47 (46.1) 11 (10.8)

32 (29.1) 55 (50) 23 (20.9)

X2 = 6.46; p = 0.039

(AA vs AG + GG; OR = 0.63; 0.36–1.10; p = 0.11) (AG vs AA + GG; OR = 1.17; 0.68–2.00; p = 0.56) (GG vs AG + AA; OR = 2.18; 1.00–4.75; p = 0.048)

rs10887800 Alleles A G

0.66 0.34

0.54 0.46

X2 = 6.44; p = 0.013

(G vs A; OR = 1.66; 1.12–2.46; p = 0.011)

rs2576178 Genotypes AA AG GG

57 (55.9) 37 (36.3) 8 (7.8)

60 (54.5) 38 (34.5) 12 (11)

X2 = 0.59; p = 0.75

(AA vs AG + GG; OR = 0.94; 0.55–1.62; p = 0.89) (AG vs AA + GG; OR = 0.92; 0.52–1.62; p = 0.79) (GG vs AA + AG; OR = 1.43; 0.56–3.67; p = 0.44);

rs2576178 Alleles A G

0.69 0.31

0.72 0.28

X2 = 0.37; p = 0.54

(G vs A; OR = 1.11; 0.72–1.71; p = 0.61)

Data were presented as subject number and percentage as appropriate. OR = Odds Ratio; vs: versus. The odds ratio (OR) was given with 95% confidence intervals (CIs).

Table 4 SBP and DBP means regarding to genotypes in women with preeclampsia. Genotypes

SBP (mmHg)

DBP (mmHg)

rs10887800 AA (n = 32) AG (n = 55) GG (n = 23) p

154.38 ± 14.35 161.0 ± 14.7 173.43 ± 8.81 (GG vs GA: p = 0.001); (GG vs AA: P < 0.001; (AG vs AA: p = 0.077)

102.50 ± 14.14 101.55 ± 8.86 113.04 ± 13.71 (GG vs GA: p = 0.001); (GG vs AA: p = 0.004; (AG vs AA: p = 0.928)

rs2576178 AA (n = 60) AG (n = 38) GG (n = 12) p

158.25 ± 14.01 163.92 ± 16.62 171.67 ± 8.34 (GG vs GA: p = 0.246); (GG vs AA: p < 0.012; (AG vs AA: P = 0.149)

102.17 ± 10.90 105.79 ± 14.82 109.58 ± 9.64 (GG vs GA: p < 0.621); (GG vs AA: p < 0.141; (AG vs AA: P = 0.333)

Data were presented as mean ± SD. SBP: Systolic blood pressure; DBP: Diastolic blood pressure.

allele frequencies of rs10887800 and rs2576178 of women with mild and severe PE were shown in Table 5. In the mild group, the distribution of rs10887800 AA, AG and GG genotypes were found as 21 (30.4%), 41 (59.4%) and 7 (10.1%), respectively; and the distribution of AA, AG and GG genotypes were found in the severe group as 11 (26.8%), 14 (34.1%), 16 (39%), respectively. When the renalase rs10887800 genotypes were compared in terms of disease severity, statistically significantly difference was found between severe and mild PE (X2 = 13.65; p = 0.001). Compared to AA + AG genotype, GG genotype had 3.96 times higher risk for severe PE compared with mild PE (GG vs AA + AG; OR = 3.96; 1.58–9.92; p = 0.003). Frequencies of A and G alleles for mild PE were found as 83 (60.1%) and 55 (39.9%), and frequencies of A and G alleles for severe PE were found as 36 (43.9%) and 46 (56.1%). The frequency of G allele was significantly different between severe and mild PE (p = 0.02; OR = 1.92; 95% CI = 1.1–3.35). It was found that G allele of rs10887800 polymorphism creates 1.92 times higher risk in women with severe PE compared with women with mild PE. Allele frequency and genotype distribution of rs2576178 polymorphism were not found statistically significant according to disease severity.

4. Discussion The present study describes the role of renalase polymorphisms in preeclampsia and shows the impact of renalase polymorphisms on the blood pressure. In the current study, we investigated rs10887800 and rs2576178 polymorphism in women with PE

and normotensive controls. We found that allele frequency and genotype distribution of rs10887800 polymorphism were statistically significantly high in women with PE. Also we found that frequencies of G allele and GG genotype of rs10887800 polymorphism were higher in women with severe PE than women with mild PE. We think that rs10887800 polymorphism in renalase gene contribute to the development of preeclampsia. PE is a common complication of pregnancy in both developed and developing countries. Although mild PE usually has a good prognosis for mother and baby, severe PE and eclampsia could have grave consequences for the mother and fetus or newborn [35]. Although several risk factors such as immunologic, genetic, nutrition and infectious origins have been proposed, PE has still remained as a ‘‘disease of theories” [36–38]. Sympathetic nervous system regulates BP by the action of catecholamines such as dopamine, epinephrine and norepinephrine [18]. It has been shown that elevated catecholamine level is related with progress or severity of PE [15]. Renalase, a novel FADdependent amine oxidase, circulates in the blood and regulates the sympathetic tone, BP and cardiac function. Renalase degrades circulating catecholamines and leads to a significant decrease in the BP [18]. There is a great deal of evidence that renalase plays an important role in the pathogenesis of many diseases such as hypertension, diabetes, cardiovascular diseases, and these evidences are on the increase [39]. Renalase action on catecholamines differs significantly from that of COMT, MAO-A and MAO-B. Despite the activity of renalase is FAD dependent, the FAD-dependent monoamine oxidase

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Table 5 Genotype distributions and allele frequencies of women with mild and severe preeclampsia. Polymorphism

Mild preeclampsia (n = 69)

Severe preeclampsia (n = 41)

p

OR

rs10887800 Genotypes AA 21 (30.4) AG 41 (59.4) GG 7 (10.1)

11 (26.8) 14 (34.1) 16 (39)

X2 = 13.65; p = 0.001

AA vs AG + GG; OR = 0.83; 0.35–1.98; p = 0.68 AG vs AA + GG; OR = 0.35; 0.15–0.79; p = 0.011 GG vs AA + AG; OR = 3.96; 1.58–9.92; p = 0.003

Alleles A G

83 (60.1) 55 (39.9)

36 (43.9) 46 (56.1)

p = 0.025

G vs A; OR = 1.92; 1.1–3.35; p = 0.02

rs2576178 Genotypes AA 39 (56.5) AG 25 (36.2) GG 5 (7.2)

21 (51.2) 13 (31.7) 7 (17.1)

X2 = 2.56; p = 0.278

AA vs AG + GG; OR = 0.80; 0.37–1.75; p = 0.58 AG vs AA + GG; OR = 0.81; 0.35–1.85; p = 0.62 GG vs AA + AG; OR = 2.63; 0.77-8.93; p = 0.11

Alleles A G

55 (67.1) 27 (32.9)

X2 = 1.45; p = 0.27

G vs A; OR = 1.44; 0.79–2.63; p = 0.22

103 (74.6) 35 (35.4)

Data were presented as subject number and percentage as appropriate. OR = Odds Ratio; vs: versus. The odds ratio (OR) was given with 95% confidence intervals (CIs).

inhibitors (pargyline and clorgyline) cannot inhibit its activity. In this respect, amine oxidase activity of renalase differs from the previously reported FAD-dependent amine oxidases MAO-A and MAOB [21]. Elsetohy et al. [32] found rs10887800 G allele frequency as 0.66 in Egyptian patients with PIH and found that AG genotype of rs10887800 polymorphism is associated with PIH. In the Northern Han Chinese population, Zhao et al. [29] found G allele frequency as 0.50 in essential hypertension patients. They found no meaningful difference between patients and controls. In patients with type 2 diabetes mellitus with hypertension, type 2 diabetes mellitus without hypertension, and stroke; G allele frequency was found as 0.49, 0.44 and 0.62, respectively. The G allele of the renalase polymorphism rs10887800 appears to be associated with an increased incidence of stroke [28]. In a study conducted with 212 ischemic stroke patients in Northern Chinese Han population, G allele frequency was found as 0.48. Both genotype distribution and allele frequencies were found to be associated with intracranial cerebral atherosclerotic vascular stenosis [40]. In the Polish population, Stec et al. [30] found that G allele frequency in ESRD with hypertension and without hypertension as 0.46 and 0.37, respectively. They found the risk of ESRD with hypertension as higher 1.76 times for G allele carriers and 1.44 times for GG genotype carriers compared to ESRD without hypertension. In Egyptian population, G allele frequency was found as 0.58 in patients with chronic kidney disease [41]. The minor G allele frequency was 0.47 on dbSNP database. In the present study, carriers of rs10887800 GG genotype in women with PE have 2.18 times higher risk compared to controls. G allele frequency of rs10887800 polymorphism was found as approximately 0.46 in the patient group. G allele had 1.66 times higher risk compared with controls. In the current study, rs2576178 minor G allele frequency was found as 0.28 and 0.31 in the patients and control group, respectively. There is no statistically significant difference between patients and control group. The minor G allele frequency was 0.34 on dbSNP database. Our results were found slightly different from dbSNP database. Elsetohy et al. [32] found frequency of the minor G allele of rs2576178 polymorphism as 0.35 in Egyptian patients with PIH. In their study, GG genotype of rs2576178 was found statistically significantly higher in patients with PIH, and there was significant difference in SBP and DBP means with regard to genotypes. Buraczynska et al. [28] found that G allele frequency in patients with type 2 diabetes mellitus with hypertension and type 2 diabetes mellitus without hypertension were 0.37 and

0.40, respectively. Li et al. [31] found that genotype and allele frequencies were statistically significantly different between patients with hypertension and coronary heart disease, and patients with hypertension. They found G allele frequency as 0.37 in the first group and as 0.49 in the second group. They also found that AA genotype of rs2576178 increased the susceptibility to development of coronary heart disease. Zhao et al. [29] investigated rs2576178 polymorphism in essential hypertension in Northern Han Chinese population. They found statistically significant difference between patients and control according to genotype distribution and allele frequencies, and found minor G allele frequency as 0.55 in essential hypertension patients. They also found that the GG genotype increased the essential hypertension risk for 1.58 times. In a study by Li et al. [40], G allele was not found statistically significant in intracranial cerebral atherosclerotic vascular stenosis in ischemic stroke, and frequency of G allele was found as 0.56. In polish population, Stec et al. [30] found that carriers of G allele were associated with a 1.55 times higher risk in hypertension, and found G allele frequency in ESRD patients with hypertension as 0.28 in ESRD patients without hypertension as 0.22. In the Malmö Diet and cancer study, SBP and DBP were not found different according to genotypes [27]. In an Egyptian study, G allele frequency was 0.71 in ESRD patients. In their study, it was found that G allele increased the risk of ESRD [41]. The discrepancy between our findings and other studies might partly explained because of the genetic heterogeneity across populations. In the present study, we analyzed SBP and DBP means in patients with PE with regard to genotypes of the renalase polymorphisms. There was statistically significant difference between SBP and DBP means according to rs10887800 genotypes. SBP means were found as having GG genotype > having AG genotype > having AA genotype. Women having GG genotype were higher DBP means than having AA and AG genotypes. According to rs2576178 genotypes comparison, only the comparison of GG versus AA was statistically significant with regard to SBP means (p = 0.012). DBP means were not statistically significant. In the present study, according to disease severity, rs10887800 minor G allele frequency was found as 0.56/0.40 in severe and mild PE, respectively. When GG genotype compared to AA + AG genotype, GG genotype had 3.96 times higher risk in severe PE than mild PE. These findings suggest that G allele and GG genotype of rs10887800 polymorphism have an additive effect on BP. Because of renalase metabolizes catecholamines, it might be useful in the treatment of conditions associated with increased sympathetic tone. To prevent from hypertension caused maternal

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death, high BP in pregnant women must be treated [35,42]. Hypertension and chronic kidney disease develop in rats subjected to subtotal nephrectomy (5/6 Nx). A single dose of recombinant renalase (subcutaneously administered 1.3 mg/kg) had an effect equivalent to enalapril (5 mg/kg) and decreased both systolic and diastolic BP. In the strokeprone spontaneously hypertensive rat (SHR-sp), administration of a single dose of renalase or enalapril (1 mg/kg) showed similar decreases in BP. Subcutaneously given recombinant renalase decreases BP without any significant change in heart rate, unlike b-blockers and calcium channel blockers [18]. The most important limitation of the present study is that the levels of the renalase and catecholamines such as epinephrine, norepinephrine and dopamine were not measured. Therefore, the possible relationship between renalase polymorphisms and levels of catecholamine, renalase and BP was not established. In addition, despite relatively small sample size, the relationship between renalase polymorphisms and their possible effects on BP may be considered as acceptable. In conclusion, findings of the present study suggest that G allele and GG genotype of rs10887800 polymorphism have an additive impact on the blood pressure. Furthermore, genetic polymorphisms in the renalase gene may contribute to the development and maintenance of preeclampsia, by increasing blood pressure. In addition, findings of the present study suggest that blood pressure may be increased by GG genotype and G allele of rs10887800 polymorphism and the polymorphism may increase the susceptibility to PE. Further researches with larger population should be done to show the effect of renalase polymorphisms in PE pathophysiology. Funding There is no funding for this work. Conflict of interest The authors declare that they have no conflict of interest. References [1] J.T. Drost, A.H. Maas, J. van Eyck, Y.T. van der Schouw, Preeclampsia as a female-specific risk factor for chronic hypertension, Maturitas 67 (2010) 321– 326. [2] C. Dolea, C. AbouZahr, Global Burden of Hypertensive Disorders in Pregnancy in the Year 2000. GBD 2000 Working Paper Geneva, World Health Organization, 2003. [3] J.A. Turner, Diagnosis and management of pre-eclampsia: an update, Int. J. Women Health 2 (2010) 327–337. [4] G.A. Dekker, B.M. Sibai, Etiology and pathogenesis of preeclampsia: current concepts, Am. J. Obstet. Gynaecol. 179 (1998) 1359–1375. [5] J.M. Roberts, D.W. Cooper, Pathogenesis and genetics of preeclampsia, Lancet 357 (2001) 53–56. [6] L. Duley, The global impact of pre-eclampsia and eclampsia, Semin. Perinatol. 33 (2009) 130–137. [7] B. Sibai, G. Dekker, M. Kupferminc, Pre-eclampsia, Lancet 365 (2005) 785–799. [8] ACOG technical bulletin. Hypertension in pregnancy, Int. J. Gynaecol. Obstet. 53 (2) (1996) 175–183. [9] National High Blood Pressure Education Working GroupHigh blood pressure during pregnancy, Am. J. Obstet. Gynaecol. 163 (1990) 1689–1712. [10] O. Demirkiran, Y. Dikmen, T. Utku, S. Urkmez, Critically ill obstetric patients in the intensive care unit, Int. J. Obstet. Anesth. 12 (4) (2003) 266–270. [11] D.A. Davey, M.F. Macnab, Plasma adrenaline, noradrenaline and dopamine in pregnancy hypertension, Br. J. Obstet. Gynaecol. 88 (1981) 611–618. [12] P. Øian, S.E. Kjeldsen, I. Eide, J.M. Maltau, Increased arterial catecholamines in pre-eclampsia, Acta Obstet. Gynaecol. Scand. 65 (6) (1986) 613–617. [13] K.E. Airaksinen, P. Kirkinen, J.T. Takkunen, Autonomic nervous dysfunction in severe pre-eclampsia, Eur. J. Obstet. Gynaecol. Reprod. Biol. 19 (5) (1985) 269– 276. [14] S. Khatun, N. Kanayama, E. Sato, H.M. Belayet, T. Kobayashi, T. Terao, Eclamptic plasma stimulates norepinephrine release in cultured sympathetic nerve, Hypertension 31 (6) (1998) 1343–1349.

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