Population and Study Design
The study was conducted in 36 Kebeles of Raya Kobo district, North Wollo Zone, Northern Ethiopia. The district covers an area of 2,001.57 square kilometers and has a total population of 228,798 of which 147,837 are females . A historical cohort study design was employed from March 15 to April 30, 2019, to investigate the effect of prenatal famine exposure on estimated glomerular filtration rate and risk of chronic kidney disease. Self-reported age and or birth date were used to categorize famine exposed and non exposed groups. Participants were categorized as prenatal exposed groups, if their age was 34 to 36 years old during the data collection period and or if their reported birth date was within 8th September 1983 to 30th August 1985. Non-exposed groups were participants aged 30 to 32 years and or their birth date was within 8th September 1987 to 8th October 1988 (suppl.1). To get the optimal washout of famine effect between the groups, participants who were born immediately after the end of the famine peak (between 8 September 1986 to 30 August 1987) were excluded. Further, exclusions were made for subjects with history of household displacement during the famine period, physical disability, including deformity (Kyphosis, Scoliosis, limb deformity) and pregnancy.
Sample size and sampling procedures
The sample size was calculated by applying two population proportion assuming a prevalence of type two diabetes mellitus as proxy risk factor for CKD in fetal exposed group (22.6%) and non-exposed group (9.8%) . Accordingly, the total sample size was 456 (228 exposed and 228 non- exposed). Multistage stratified random sampling technique was used to select the study participants. Details of participants’ selection procedure were depicted in supplementary figure (suppl.1).
The exposure variable was the great Ethiopian famine of the 1983–1985 G.C. Other sociodemographic, biochemical including fasting blood glucose, lipids and lipoproteins and anthropometric variables were also assessed. The outcome variable eGFR was computed from standardized serum creatinine using the CKD Epidemiology Collaboration (CKD-EPI) equation where eGFR (estimated glomerular filtration rate) expressed as ml/min/1.73 m2
eGFR= 141 x min(SCr/κ, 1)α x max(SCr /κ, 1)-1.209 x 0.993Age x 1.018 [if female] x 1.159 [if Black].
Where SCr (standardized serum creatinine) in mg/dL, κ = 0.7 (females) or 0.9 (males), α = -0.329 (females) or -0.411 (males), min = indicates the minimum of SCr/κ or 1, max = indicates the maximum of SCr/κ or 1, age = years .
Data collections and measurements
The data collection procedure and measurements were guided by standard operating procedures outlined by the WHO STEPwise approach to surveillance (STEPS) manual . The data collection was carried out through an interview, biochemical test, blood pressure and anthropometric measurements.
Interview: Pretested structured questionnaire was used to collect sociodemographic data of the participants using face-to-face interview. The questionnaire was first prepared in English and then translated into Amharic (the local language) and back into English to ensure consistency. Eight trained clinical nurses collected the data.
Biochemical measurements: Fasting total cholesterol, plasma glucose, triglycerides and high-density lipoproteins were measured. Five milliliters of venous blood was collected in plane test tubes after overnight fasting participants (8-12 hours). The analysis was made using A-25 bio-system® clinical chemistry analyzer. Low density lipoprotein (LDL) level was determined using Freidwald formula . Standard operating procedures (SOP) were following to collect blood samples, perform laboratory analysis . All laboratory analysis were conducted in Dessie branch of Amhara Public Health Institute (APHI) laboratory.
Anthropometric measurements: The height of participants was measured to the nearest 0.1 cm using a stadiometer (Seca®, Germany) with the subjects positioned at the Frankfurt Plane and the four points (heel, calf, buttocks and shoulder) touching the vertical stand and their shoes taken off. Weight was measured using portable battery operated Seca® digital scale. All anthropometric measurements were done in triplicate and the average value was used for further analyses . The weigh scales were checked read zero and standardized using an object of known weight before measurement.
Blood pressure (BP); Blood pressure was measured using a digital blood pressure apparatus in triplicate, with five minutes of rest in between measurements. The subsequent measurements were done 5 minutes apart. During data analysis, the mean of the second and third readings were calculated .
Data Processing and Analysis
Data were double-entered using EpiData 3.1 and exported to SPSS for windows version 25 ([SPSS Inc. version 25, Chicago, Illinois] for cleaning and analysis. The data were cleaned by checking outliers and missing values. Categorical variables were described as frequencies and percentages and compared using the Pearson chi-square test. Continuous variables with a normal distribution were described using the relevant indicators of central tendency and spread (mean ± SD or median and IQR). Student's t - test was used to evaluate the mean score difference between prenatal famine exposed and non-exposed cohorts.
Linear and logistic regressions were employed to examine the relationship between famine exposure in prenatal life and eGFR or CKD respectively. In order to account for the effect of outstanding biologic covariates four different regression models were evaluated. The first models in both regressions present unadjusted coefficients while model two, three and four were adjusted in a stepwise approach for fasting blood glucose, systolic hypertension and body mass index, respectively. Hosmer-Lemeshow test greater than 0.05 and maximum standard errors (SE) greater than 2 were used to check model fitness and multicollinearity, respectively. Potential effect modification was assessed by interaction terms. All analyses were two sided and p value of 0.05 was used to declare a significant difference. The results were presented as crude as well as adjusted odds ratio or beta coefficients and their 95%confidence intervals
Ethics approval and consent to participate
Ethical clearance was obtained from Jimma University Institutional Review Board (IRB) as per protocol number IHRPGD/443/2018. Written informed consent was taken from each participant. The study participants were assured that they are free to withdraw their consent and discontinue participation without any form of prejudice. Privacy and confidentiality of collected data was ensured throughout the study. Copy of laboratory results were given to the respective participants and referral to the nearby public health facility were made for individuals with laboratory findings beyond the reference range.