Analyses of Interleukin-6, Presepsin and Pentraxin-3 in the Diagnosis and Severity of Late-onset Preeclampsia
Conditions: Preeclampsia
Sponsor: Cengiz Gokcek Women's and Children's Hospital

Full study protocol and statistical 
analysis plan 
Official Title of the study: 
Analyses of Interleukin-6, Presepsin and Pentraxin-3 in the diagnosis and severity of 
late-onset preeclampsia 
 
Date of the document: 
October 2019. 
Contents: 
1. Participant Flow 
2. Baseline Characteristics 
3. Outcome Measures 
4.Endpoints of the study 
5.Limitations and Strengths 
6. Certain Agreements 
7. Results Point of Contact  
 
1. Participant Flow 
Recruitment Details 
The study was conducted at Cengiz Gokcek Women’s and Children’s Hospital, Gaziantep, Turkey at 
department of obstetric and gynecology between date of June 2018 and January 2019. The study 
protocol was designed according to the Declaration of Helsinki, and the institutional ethical review 
board of Gaziantep University approved the study (Reference number: 2018/393). The investigators 
included subjects consisted of women with a singleton pregnancy who were diagnosed as having 
late-onset preeclampsia between 34+0 and 41+0 weeks of gestation. Pregnant women with 
uncomplicated pregnancies were randomly selected to serve as controls. The study was conducted 
with 44 late-onset preeclampsia patients as study group and  44 patients with normal pregnancies as 
control group. All participants included in the study gave oral and written informed consent. 
Pre‐assignment Details 
The authors firstly were assessed the recruited people to ensure meeting the inclusion and 
exclusion criteria. 
The inclusion criteria were maken according to the official documents. 

Arm/Group Information * 
 
There are two groups in the study. 
 
Arm/Group Title * 
 
Preeclampsia group 
Control group 
2. Baseline Characteristics 
The normal distribution of data is analysed with Kolmogorov Smirnov ve Shapiro Wilk tests. 
Student t test was used for variables that normally distributed for gorup comparison(L-
PrE/kontrol) and Mann Whitney U test was used for variables that is not normally distributed. 
Also, Sperasman correlation was used for relationship of non-normally distributed variables 
with each other. The ROC analysis was applied for determination of cut-off points for  IL6, 
presepsin ve pentraxin-3 variables. SPSS for Windows 22.0 ve Medcalc programs were used 
for statistical analysis. P<0.05 was accepted as statistical significance. 
3. Outcome Measures 
 
Every pariticipant in the study population underwent obstetric ultrasound examination and 
fetal-maternal assesment was carried out by one of the investigators. The obstetric anamnesis 
were obtained from all pariticipants. The demographic data like age, gravidity, parity, body 
mass index (BMI) and gestational age were recorded. The gestational age was determined by 
calculation from the last menstrual period and supported by the ultrasonography 
measurements at first trimester of gestation. Maternal venous blood samples were taken for 
measurement of IL-6, presepsin and pentraxin-3 levels after the diagnosis of L-PrE in 
outpatient clinic. These samples quickly centrifuged at 1,500 g for 10 min, plasma samples 
were seperated and stored at -80°C until the day of measurement. All patients with L-PrE 
were also hospitalized and their pregnancies were terminated. The samples of the control 
groups were obtained during the routine obstetrical care examination in the third trimester of 
pregnancy. Then these pregnant women were followed-up until the delivery. Both groups 
were compared in terms of maternal age, BMI, week of gestation, gravida, parity, live born, 
systolic/diastolic blood pressure, total protein in spot urine sample, hemoglobin, hematocrit, 
platelet count, white blood cell count, neutrophil, lymphocyte, neutrophil-to-lymphocyte ratio 
(NLR), urea,  uric acid, albumin,  blood urea nitrogen, creatinine, liver function tests (AST, 
ALT), lactic acid dehydrogenase, IL-6, presepsin, pentraxin-3, and infant weight at delivery. 
Neutrophil-to-lymphocyte ratio (NLR), which is the ratio of the neutrophils and lymphocytes  in blood 
sample at admission, which can accept a marker of systemic inflammation. Small for gestational age 
(SGA) neonate was defined as birth weight <10th percentile for gestational age with Turkey's 
national nomogram as the reference for fetal growth. The maternal plasma presepsin levels 
were determined using a commercially available ELISA kit, the human presepsin ELISA Kit 
(Rel Assay Diagnostics, Gaziantep, Turkey), according to the manufacturer’s instructions. 
Human presepsin ELISA kit is based on the principle of biotin double antibody sandwich 
technology. IL-6 levels in blood samples were measured by using the ELISA method 
(DIAsource ImmunoAssays SA, Nivelles, Belgium) in accordance with the method described 
by the manufacturer. Pentraxin-3 levels were measured by using the ELISA kits in accordance 
with the manufacturer’s instructions (Sunredbio, Shanghai, PRC).   
 

 
4. Endpoints of the study:  
The primary endpoint in these analyses was IL-6, presepsin and pentraxin-3 levels in L-PrE 
group and control group.  
The secondary endpoint was the IL-6, presepsin and pentraxin-3 levels in mild L-PrE group 
and severity L-PrE group. 
 Tertiary endpoint was the IL-6, presepsin and PTX-3 levels in SGA group and non-SGA 
group. 
 
5.Limitations and strengths: 
 
This study have some limitations. It was a small, observational and single-centered study. 
These three inflammatory markers were measured only during admission, we could not assess the 
changes in IL-6, presepsin, and pentraxin-3 in response to treatment due to lack of serial 
measurements. 
 We demonstrated, significant relations of all three elevated new inflammatory markers with 
both the presence and the severity of PrE at the same time in this study. Another strength of our 
study was that the women with PE had no pharmacological treatment. Because the antihypertensive 
drugs lead to the decrease in presepsin level due to the antiinflammatory effect.  
 
6. Certain Agreements 
 
The authors declare that they have no conflict of interest. 
 
7. Results Point of Contact 
Corresponding Author: Ali OVAYOLU, Cengiz Gokcek Public Hospital, Department of Obstetrics 
and Gynecology, Gaziantep/Turkey, drovayolu@yahoo.com 
Adress: Osmangazi Mahallesi, Cengiz Gökçek Kadın Hastalıkları ve Doğum Hastanesi 27010 
GAZİANTEP 
GSM: +90 (532) 640 40 60