Prematurity as Predictor of Children's Cardiovascular-renal Health
Condition: Endothelial Dysfunction; Sublingual Capillary Glycocalyx and Density; Visual Disturbances and Blindness; Ventricular Dysfunction, Left; Renal Impairment
MeSH terms: Blindness; Premature Birth; Renal Insufficiency; Ventricular Dysfunction; Ventricular Dysfunction, Left
Study type: Observational

Summary: 
      Extreme preterm birth interferes with the development of the cardiovascular system. Both
      macro- as well as microvasculature undergoes extensive, organ specific maturation. Under
      normal fetal conditions, microvascular growth drives renal development and continues until
      34-36 weeks of gestational age, while retinal vascular growth continues until term age.
      Studies show that there is association between low birth weight and cardiovascular
      dysfunction. According to the Barker hypothesis, this is due to nutritional shortage. In
      extreme preterm birth cases, this growth restriction is observed in neonatal life.

      In adult life, this suboptimal growth is associated with impaired renal and (micro)vascular
      function, hypertension, glucose intolerance and cardiovascular disease. According to the
      Brenner hypothesis, disrupted renal development results in hyperfiltration and hypertension,
      a process that subsequently promotes itself and leads to renal impairment. We will
      investigate macro- and microvasculature in different organs, including eye, kidney, heart and
      sublingual mucosa in former preterm infants, now aged 8-13 years old and age-matched
      controls.

      The expectation is that the results of this project will identify risk factors for
      cardiovascular-renal disease in the adult life of former preterm infants compared to the
      controls, while further analysis on mediators in neonatal life of this cardiovascular-renal
      outcome may provide new information on perinatal risk factors.
    

Detailed description: 
      STATE OF THE ART The cardiovascular system (both macro- and microcirculation) undergoes
      extensive maturation throughout fetal, perinatal and pediatric life. Extreme preterm birth
      interferes with the normal development of the cardiovascular and microcirculatory systems.
      Disruption in vascular ontogenesis leads to abnormalities in the microvascular structure and
      circulation in various organs, such as retina (retinopathy of prematurity [O'Connor et al.]),
      kidney (abnormal glomerulogenesis [Sutherland et al.]) and glycocalyx in sublingual
      capillaries [Nieuwdorp et al.], among microvascular-driven disruptions observed in other
      organs (e.g. periventricular leukomalacia [Takashima et al.], bronchopulmonary dysplasia
      [Gien et al.]). Besides the well known retinopathy of prematurity, microvascular growth
      drives glomerulogenesis in the kidney and terminates after 34-36 weeks of gestational age
      under normal fetal conditions.

      Perinatal (fetal or neonatal) growth restriction or preterm birth therefore impairs
      glomerulogenesis [Sutherland et al., Abitbol et al., Barker et al., Faa et al., Gubhaju et
      al., Zaffanello et al.]. Other cardiovascular abnormalities following from premature birth in
      later life include decreased heart rate variability [Rakow et al.], endothelial dysfunction
      [Norman et al.] and hypertension [Abitbol et al., Keijzer-Veen et al.].

      Epidemiological observations further confirm that former preterm born infants are indeed at
      increased risk to develop cardiovascular disease and chronic kidney disease during adulthood
      [Brenner et al., Carmody et al., Vieux et al., Zandi-Nejad et al.]. The concept that fetal
      and perinatal conditions affect normal cardiovascular and renal ontogeny is in itself not
      new. Epidemiological studies showed that there is an association between low birth weight and
      vascular dysfunction in later life, suggesting that vascular impairment in early life is a
      harbinger of a poorer long-term prognosis [Sutherland et al., Bacchetta et al., Keijzer-Veen
      et al., Puddu et al.]. Intra-uterine growth retardation and children small for gestational
      age can be regarded as a failure of a fetus to reach the genetic potential of growth due to
      nutritional deprivation, the so-called Barker hypothesis [Barker et al.]. In adult life, this
      early growth retardation is associated with impaired renal and (micro)vascular function,
      hypertension, glucose intolerance and cardiovascular disease [Sutherland et al., Abitbol et
      al., Faa et al., Zaffanello et al., Carmody et al., Keijzer-Veen et al.].

      The same sequence of vascular impairment serving as an indicator for long-term prognosis
      applies to the (early) postnatal life of preterm infants. According to the Brenner
      hypothesis, the decreased number of nephrons causes hyperfiltration, sodium loss with
      activation of the renin-angiotensinaldosterone system and hypertension [Brenner et al.], a
      process that entails further nephron loss, predisposition to develop proteinuria and possibly
      chronic kidney disease [Vieux et al., Puddu et al.].

      MOVING BEYOND THE CURRENT STATE OF THE ART This project aims to move beyond the
      state-of-the-art by studying association between macro- and microvascular structure and
      function in children (8-13 years) born prematurely (extremely low birth weights, ELBW, i.e.
      birth weight below 1000 grams), and sex- and age-matched controls. The specific strengths
      hereby are that this ELBW cohort has been well characterized on its perinatal aspects [George
      et al.]. The characterization in the postnatal period includes biometry, perinatal
      characteristics (e.g. Apgar score, drugs, respiratory support), creatinine trends in the
      first 6 weeks of postnatal life and psychomotor development (Bayley Scales of Infant
      Development) at the age of nine months and two years.

      The phenotypes of interest for the current project include the micro- and macrocirculation,
      cardiac structure and function, endothelial function and renal anatomy and function during
      childhood. We hereby aim to apply state of the art methods (cf methodology section) to assess
      micro- and macrocirculatory function, combined with state of the art statistics and advanced
      tools (metabolomics, epigenetics) to explore epidemiological findings and its pathogenesis.

      The expectation is that the results of this project will identify and quantify risk factors
      in former preterm infants for cardiovascular-renal disease when compared to control children,
      while we can also map early risk factors for cardiovascular-renal disease in adult life and
      pave the way for a better informed prevention of these complications. Finally, data collected
      in this specific cohort can be compared to data collected in other cohorts.

      HYPOTHESIS We hypothesize that former ELBW infants, compared to controls following term
      birth, will be associated with changes in the macro- and microcirculation of the
      cardiovascular-renal system already in young children over and beyond what is currently
      known. These changes - even if subtle - are probably forerunners of cardiovascular-renal
      complications in adulthood. In addition, confounders (e.g. neonatal nutrition, neonatal
      treatment) documented in the early life may identify new approaches for early prevention.

      OBJECTIVES Using a case-control design in a 1/2 proportion, this study aims to detect
      functional and structural changes in children after preterm birth (ELBW) compared with
      children born after normal gestation with normal weight. The phenotypic aspects considered
      cover micro- and macrovascular structures and function.

        1. Endothelial function

        2. Sublingual capillary glycocalyx and density

        3. Retinal imaging and visual acuity

        4. Left ventricular function

        5. Renal anatomy and function

        6. Structure and function of the carotid artery (intima-media thickness, distensibility,
           Young's elastic modulus), aortic pulse wave velocity and the systolic augmentation
           index.

      MODULATORS Next, this project will search for host characteristics, life style and
      environmental factors (see below) that may further modulate the differences in youngsters
      born either prematurely (case, ELBW) and at term, as well as for circulating and urinary
      biomarkers that are associated with the observed differences, and may provide insight into
      the pathogenesis involved.

      IDENTIFICATION OF PREDICTORS Based on the existing database of the early perinatal follow-up
      of the prematurely born infants, this project will attempt to construct models predicting
      increased risk of potentially clinically significant changes associated with preterm birth
      and vice versa (mediators in neonatal life may predict cardiovascular-renal outcome in adult
      life).

      INTENTION TO LINK THESE DATASETS WITH OTHER COHORTS WITH SIMILAR OBSERVATIONS Finally,
      pooling of these cardiovascular phenotypic data in children with other cohorts, either or not
      former ELBW cases may provide opportunities for additional prediction model development,
      validation and subsequent secondary preventive strategies.

      METHODOLOGY Methodology-related issues include phenotyping, database construction/quality
      control, modulators, predictors and statistics.

      PHENOTYPING

        1. Endothelial function will be assessed by 24h urinary microalbumin excretion and digital
           pulse wave amplitude hyperemic response (photoplethysmography,PPG). To determine
           amplitude changes of the digital pulse, the response of the PPG pulse wave amplitude to
           hyperemia will be calculated from the hyperemic fingertip as ratio of post-deflation PPG
           pulse to baseline amplitude (PAht/PAh0, PA=pulse amplitude, h=hyperemic finger, t=time
           interval, 0=baseline). To obtain this ratio, we will divide the PAht/PAh0 ratio by the
           corresponding ratio at the control hand (PAct/PAc0, c=control finger)[Kuznetsova et
           al.].

        2. To measure sublingual capillary glycocalyx and density, videos (10 sec) of sublingual
           capillaries in 2 areas lateral of the frenulum and 3-4 cm anterior to the tongue base
           will be recorded, using orthogonal polarization spectral (OPS) and side stream dark
           field (SDF) imaging [Hubble et al.]. In our hands, the intra- and inter-observer
           reproducibility of capillary density is 10.2 and 13.4% respectively. The
           erythrocyte-endothelium gap is the gold standard for glycocalyx measurement in vivo
           [Nieuwdorp et al.], as endothelial glycocalyx allows limited access to erythrocytes. The
           perfused boundary region (PBR) hereby reflects glycocalyx thickness and integrity,
           increased PBR glycocalyx loss.

        3. Retinal imaging will be performed using a Canon Cr-DGi (Canon Co Ltd, Kyoto, Japan)
           nonmydriatic retinal visualization system. After accommodation to darkness, 1 image/eye
           will be obtained [Liu et al.]. Trained observers will identify individual arterioles and
           venules, using a validated computer-assisted program IVAN (Vasculo-matic Nicola,
           Ophthalmology and Visual Science, University of Wisconsin-Madison[Sherry et al.]). In
           addition, we will investigate visual acuity (clearness of vision, spatial resolution of
           the visual processing system) by the non-invasive adapted Snellen charts without visual
           aids.

        4. All children will undergo detailed assessment of left ventricular (LV) function through
           echocardiography. PREMATCH will hereby focus on early changes in diastolic and systolic
           LV function. In combination with tissue Doppler imaging (TDI), transmitral and pulmonary
           vein blood flows will be used to detect LV filling changes.

        5. Renal anatomy and function will be assessed by 2-dimensional measurement, renal arterial
           Doppler blood flow measurement and 3-dimensional calculations. Creatinine clearance, 24h
           microalbuminuria and Cystatin C on peripheral blood (serum) will be quantified.

        6. The structure and function of the carotid artery, aortic pulse wave velocity (PWV) and
           systolic augmentation index will be assessed as measures of macrovascular function and
           structure. We will use ultrasound to measure the local properties of the common carotid
           artery. diameter, distension, and intima-media thickness will be measured and averaged
           over 3 cardiac cycles in recordings consisting of >4 consecutive beats. Distensibility
           (103/kPa), compliance (mm²/kPa) and Young's elastic modulus will be calculated. Local
           application tonometry will be performed (SpyghmoCor system). Measurements will be
           performed at carotid, radial and femoral arteries. The local arterial pulse wave will be
           recorded as well as carotid-to-femoral and carotid-to-radial PWV (cf- and cr-PWV). The
           carotid and radial augmentation indexes will be measured directly at the carotid and
           radial arteries and the aortic augmentation index will be calculated from the radial
           signal by the validated generalized transfer function [Richart et al., Seidlerova et
           al., Mischak et al.].Furthermore, we will implement ambulatory assessment of central
           hemodynamics, using the Mobil-O-Graph 24h PWA Monitor (IEM GmbH, Stolberg, Germany), a
           validated monitor for 24h blood pressure monitoring, including the ARCSolver
           application, which allows pulse wave analysis of the central blood pressure and
           measuring of aortic PWV [Luzardo et al.].

      DATABASE CONSTRUCTION/QUALITY CONTROL Trained nurses will code questionnaires, technicians
      will enter the data. For quality assurance, 10% of questionnaires will be randomly selected
      and recoded by another nurse. All data will be inputted twice by different technicians.
      Duplicate datasets will be compared with the PROC COMPARE application (SAS software) to trace
      input errors. Data coders and SAS programs will check for internal consistency of
      questionnaire replies. Non-Gaussian distributions will be normalized by proper
      transformation. As part of the quality control, descriptive statistics will be generated at 6
      month intervals.

      MODULATORS

        -  Anthropometric characteristics, by sex, age, height, weight, body mass index, span
           width, waist-tohip ratio, skinfolds (Harpenden Skinfold Caliper, Bedfordshire, UK).

        -  Matrix reasoning and spatial orientation tests (Wechsler), indicators of mental
           capability.

        -  Muscle strength, by grip strength.

        -  Sexual maturation, by Tanner scale.

        -  Office and self-measured home blood pressure, by Mobil-O-Graph 24h PWA monitor (IEM,
           Stolberg, Germany)

        -  Body composition, by Bodystat4000 (Bodystat Ltd, Douglas, UK)

        -  Questionnaire (education, medication use, habits, menarche (girls) and familial and
           personal history).

        -  Measurements on blood and 24h urine samples. Plasma, serum and urine samples will be
           divided into aliquots and stored (-20/-80°C) at the bio-bank of the Studies Coordinating
           Centre (SCC). Routine measurements include hemoglobin, hematocrit, red and white blood
           cell counts, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular
           hemoglobin concentration, differential white blood cells count, serum creatinine, uric
           acid, serum lipids (total, HDL-cholesterol), glycaemia and insulin. Measurements of
           metabolic, inflammatory and oxidative stress include: 8-hydroxy -2'-deoxyguanosine
           (8-OHdG), interleukin-6 and high sensitivity C-reactive protein[Pearson et al.]. Other
           measurements will be considered if enough samples are available (e.g. homeostasis model
           assessment index (HOMA)[Marcelis et al.], leptin, adiponectin [Marcelis et al., Yeon et
           al.], E-selectin, P-selectin, vascular adhesion molecule-1, plasma fibrinogen, tumor
           necrosis factor, superoxide dismutase or serum malondialdehyde[Rao et al., Sharma et
           al.]).

      Measurements of 24h urine samples include volume, electrolytes, creatinine, micro-albumin,
      and aldosterone. Urinary proteomics will be done in collaboration with Prof Harald Mischak,
      SME Mosaiques (mosaiques-diagnostics.de) by capillary electrophoresis coupled to mass
      spectrometry according to standard operating procedures in an environment with proper quality
      control [Mischak et al.].

      - GPS coordinates of residence. Meteorological data and data on airborne pollutants and fine
      particulate collected from the appropriate sources.

      PREDICTORS In our published cohort [George et al.] on creatinemia in ELBW infants in the
      first 6 weeks of life, raised creatinemia reflected immaturity (e.g. gestational age, weight)
      and morbidity (Apgar, ventilation, retinopathy of prematurity, intraventricular hemmorrhage),
      but also treatment modalities (e.g. ibuprofen, steroids, parenteral nutrition). We will link
      the perinatal covariates and creatinine trends to the dataset of this study to explore to
      what extent perinatal data predict cardiovascular and renal outcome. Since also treatment
      modalities are included, this study will provide first data on long term cardiovascular and
      renal outcome following drug exposure.
    

Observational model: Case-Control
Time perspective: Cross-Sectional
Sampling method: Non-Probability Sample
Study population: 
        ELBW (birth weight below 1000 g) neonates and have been well characterized and documented
        in the postnatal period, survivors (n = 140) will be matched with two controls. One control
        will be matched to sex, birth year and residential area and will be suggested by the index
        patient, the second control will be age and sex matched from the area of the field center.
        All children considered are currently between 8 and 15 years of age.

        Based on GCP guidance and national law, parents or custodians will provide informed written
        consent, while the child has to provide informed assent.
      
Biospecimen description: 
      Urine collection Blood collection
    
Keywords: Prematurity; Extremely low birth weight; Phenotyping