Serum Levels of C Type Natriuretic Peptide in Reproductive Age, Perimenopausal and Postmenopausal Women
Phase: NA
Conditions: Serum CNP Levels, Healthy Reproductive Age Women, Healthy Perimenopausal Women, Healthy Postmenopausal Women
Sponsor: Near East University, Turkey

Serum Levels of C Type Natriuretic Peptide in Different Reproductive Periods 
NCT04000815 
Date:21.01.2020 
 
 
Serum Levels of C Type Natriuretic Peptide in Different Reproductive Periods 
NCT04000815 
Date:21.01.2020 
Study Protocol and Statistical Analyses Plan 
Recent studies have shown that C type natriuretic peptide (CNP) that is produced from 
granulosa cells of the ovary, has an increasing effect on cyclic guanosine monophosphate 
(cGMP) production by affecting cumulus cells through natriuretic peptide receptors (NPR). It 
has been suggested that the transport of cGMP to oocyte via gap junctions causes an 
inhibition on oocyte phosphodiesterase 3 which leads an increase in cyclic adenosine 
monophosphate (cAMP) levels within the oocyte by hydrolyses. An important role of 
increased cAMP levels in oocyte is shown to suppress meiotic progression. Hormonal 
regulation of CNP secretion has been demonstrated in animal and human studies. 
Deoxyribonucleic acid (DNA) studies in animals have shown that expression of the natriuretic 
peptide precursor (NPPC) increases during the periovulatory period and shows that this 
increase decreases rapidly after Luteinizing hormone(LH) / human chorionic(hCG) 
stimulation. Human studies have shown that after ovulation induction, the CNP level in 
follicular fluid decreases following ovulatory dose of hCG. It has been demonstrated by many 
studies that LH has a suppressing effect on NPPC / NPR2 production. In other studies, 
esradiol has been shown to have a promoting effect on NPPC. Follicle stimulating hormone 
(FSH) alone, has been shown to have no effect on CNP regulation. In fact when there is 
estrogenic effect together with FSH, there is increasing CNP levels. 
C type natriuretic peptide, which has been demonstrated that is affected by E2, LH and FSH 
in the hormonal regulation and is synthesized by the granulosa cells, may be a new ovarian 
reserve test. And also, it may be associated with increased age, which is also an important 
factor in ovarian aging. In this study, it was aimed to determine and to compare serum CNP 
values in three different stages of reproductive life. 
In this prospective study, 71 patients are recruited. After the exclusion of patients, group 1 
consists of 15 healthy reproductive aged women between 18-40 years old, with regular 
menstruation. Group 2 includes 15 patients in perimenopausal time period between 40-49 
ages and in group 3 there is 15 postmenopausal women. 
Age, gravida, parity and body mass index (BMI) data of all patients are recorded. BMI is 
calculated by dividing the body weight in kilograms by the square of the height in meters. All 
patients went under ultrasound examination by the same clinician (ACO). The number of 
antral follicles in group 1 and group 2 were recorded. In addition, on the 2nd or 3rd day of 
menstruation, serum FSH, LH and E2 data of the patients in group 1 and 2 were recorded. 
Patients with drug use that may affect menstruation like oral contraceptives in the last 6 
months, patients with a known cardiac or renal disease and therefore using medicine, patients 

Serum Levels of C Type Natriuretic Peptide in Different Reproductive Periods 
NCT04000815 
Date:21.01.2020 
 
 
suffering from infertility as a result of any reason other than unexplained etiology, patients 
with a history of ovarian surgery, presence of polycystic ovary syndrome or that have an 
irregular menstrual cycle were excluded from this study. In addition, patients with central 
nervous system and endocrine diseases were excluded. 
Morning fasting venous blood samples were taken from the patients on the 2nd or 3rd day of 
the menstruation for group 1 and 2. All blood samples were centrifuged on the day of 
collection and separated serum samples and were kept at -80 degrees celcius until the day of 
CNP test. Serum CNP levels of the patients were analyzed by an enzyme‐linked 
immunosorbent (ELISA) assay for human CNP in accordance with the manufacturer's 
instructions (SEA721Hu, ELISA Kit for Human CNP, Wuhan USCN Business Co., Ltd., 
Cloud-Clone Corp., CCC, USA). 
Data were analyzed using Statistical Package for Social Sciences software (SPSS v15, SPSS 
Inc, Chicago, IL, USA). The variables were investigated using visual and analytical methods 
(histograms, homogeneity of variances test, Kolmogorov-Simirnov/Shapiro-Wilk's test) to 
determine whether or not they were normally distributed. Patient demographics and CNP 
values were presented as median and interquartile range. Gravida and parity were 
demonstrated by frequency distribution. The correlation coefficients and their significance 
were calculated using the Spearman test. P-values less than 0.05 will be regarded as 
statistically significant. 
 
Results 
A total of 45 patiens were evaluated in the statistics. The flow chart of the study is shown in 
figure 1. Table 1 shows the demographic characteristics and table 2 shows frequency 
distribution of gravida and parity of the patients. In table 3 serum CNP levels are shown. The 
median and interquartile range of serum CNP levels in groups 1, 2 and 3 were, 43.68 (35.54-
47.87) pg/mL, 55.98(49.84-61.13) pg/mL and 116.36(59.61-250.00) pg/mL, respectively. 
When the three groups were compared with Kruskal Wallis test, there was a significant 
difference between them in terms of serum CNP values (p<0.001). Post hoc analysis showed 
significant differences between group 1 vs. group 2, group 1 vs. group 3 and group 2 vs. 
group 3. (p values; p <0.001, p <0.001, and p = 0.002, respectively). When all patients were 
evaluated, a positive correlation was found between age and serum CNP levels. (Rs= 0.712, 
p<0.001) (Figure 2) In group 1 and group 2, there was a positive correlation between serum 
CNP and serum FSH, (Rs=0.609, p<0.001) (Figure 3) and a negative correlation between 
serum CNP and AFC. (Rs= -0.603, p<0.001) (Figure 4) There was no correlation between 
serum CNP and serum LH and E2 values in group 1 and 2. 
Figure 1: Flow chart of the study 

Serum Levels of C Type Natriuretic Peptide in Different Reproductive Periods 
NCT04000815 
Date:21.01.2020 
 
 
 
 
Table 1. Demographic characteristics of participants 
 
Group 1(n=15) 
Group 2 (n=15) 
Group 3 (n=15) 
All participants 
(n=45) 
Age (years) 
24.9±2.9 
44.1±2.2 
57.7±5.4 
42.2±14.1 
BMI (kg/m2) 
21.3±1.9 
24.3±2.4 
27.1±4.1 
24.2±3.8 
AFC 
14.2±2.18 
8.33±1.72 
- 
11±7 
FSH (mIU/mL) 
5.28±1.33 
13.12±7.3 
- 
7.65±7.96 
LH(mIU/mL) 
5.71±2.51 
5.16±2.45 
- 
5.29±2.76 
E2(pg/mL) 
41±26 
22±14 
- 
32±27 
BMI: Body mass index, AFC: Antral follicle count, FSH: Follicle stimulating hormone, LH: 
Luteinizing hormone, E2: Estradiol 
 
Table 2: Frequency distribution of gravida and parity among groups 
Frequency  
Group 1 (n=15) 
Group 2 (n=15) 
Group 3 (n=15) 
Gravida 
Parity 
Gravida 
Parity 
Gravida 
Parity 

Serum Levels of C Type Natriuretic Peptide in Different Reproductive Periods 
NCT04000815 
Date:21.01.2020 
 
 
0 
14 
14 
1 
1 
 
 
1 
1 
1 
1 
4 
 
1 
2 
 
 
8 
8 
5 
10 
3 
 
 
4 
2 
4 
2 
4 
 
 
1 
 
3 
1 
5 
 
 
 
 
1 
 
7 
 
 
 
 
 
1 
10 
 
 
 
 
2 
 
 
 
 
 
 
Table 3. Serum CNP levels of participants 
 
Group 1(n=15) 
Group (n=15) 
Group 3 (n=15) 
p 
CNP (pg/mL) 
43.68 (35.54-47.87) 
55.98(49.84-61.13) 
116.36(59.61-250.00) 
<0.001 
CNP:C type natriuretic peptide 
Group 1vs group 2: p<0.001; group 1 vs group 3: p<0.001; group 2 vs group 3: p=0.002.  
Figure 2.Correlation between age and serum CNP  

Serum Levels of C Type Natriuretic Peptide in Different Reproductive Periods 
NCT04000815 
Date:21.01.2020 
 
 
 
 
 
 
 
 
Figure 3.Correlation between serum CNP and serum FSH 

Serum Levels of C Type Natriuretic Peptide in Different Reproductive Periods 
NCT04000815 
Date:21.01.2020 
 
 
 
 
Figure 4.Correlation between serum CNP and AFC