Patients

For our analyses we subdivided a group of 101 consecutive children with height below the third percentile into the following diagnostic groups. All of them had been referred between the years 2005 to 2015 to the pediatric outpatient clinic of our hospital:

1. Congenital syndromes (N = 17) like Down's -, Turner-, Noonan-, Rett- and Silver Russell syndrome. These syndromes are known as genetic disorders with impact on height (genomic imprinting).
2. Small for gestational age (SGA; N = 30): SGA is due to malnutrition in utero (gestational imprinting). SGA is related to an enhanced risk of cardiovascular disease and ADHD.
3. Growth hormone deficiency (GHD; N = 11): GHD is related to an autonomic disorder in children and adults with a known enhanced cardiovascular risk.
4. Congenital heart disease (CHD; N = 10): CHD patients have an enhanced risk of growth failure and ADHD in later life related to heart failure in infancy (infant imprinting) [9, 10].
5. Constitutional growth delay (CGD; N = 25): CGD is characterized by delayed bone age maturation. These children are our additional "healthy control" to analyze the sole effect of short stature on the autonomic nervous system.
6. Miscellaneous (N = 8). These children cannot be assigned to the other groups. Most of these patients suffered from gastrointestinal disease like celiac disease.

Furthermore, we investigated the impact of early life stress on HRV in 20 infants with heart failure and 10 infants with failure to thrive in and ex utero in order to demonstrate the autonomic effects of these diagnoses during early life.

Control group

Healthy children (N = 55) were collected from our “normal values heart rate variability project”, therefore data of patients, who attended our outpatient clinic for exclusion of cardiac arrhythmia were analysed retrospectively. The retrospective analysis was approved by the ethical board of our state’s medical chamber (Landesärztekammer Baden Württemberg) and recently published [11]. Children with a body mass index above 96% and height below 3% were excluded from this group of healthy children.

24 hour ECG and analysis of heart rate variability

Measurement and interpretation of HRV parameters in the current sample were standardized according to the Task Force Guidelines [12]. Cardiac autonomic functioning was measured by a 24 hour Holter 12 bit digital ECG (Reynolds Pathfinder II, Spacelabs, Germany; 1024 scans/sec). Daytime and night time periods were defined according to patient’s protocols. All Holter recordings were reviewed by the same experienced cardiologists (RB and CW) and were edited to validate the systems QRS labelling. Measures of HRV were calculated employing only normal-to-normal intervals. QRS-complexes classified as noise were ejected. A minimum of 23 hours of analyzable data and minimally 95% of analyzable NN intervals were required for the data to be included. For time domain measures, mean RR interval, resulting heart rate and the following HRV parameters were calculated as average hourly values and as 24-hour average values: square root of the mean of the sum of squares of differences between adjacent NN-intervals (rMSSD) and the standard deviation of NN intervals (SDNN). RMSSD predominantly reflects changes in vagal tone. SDNN—this parameter reflects global heart rate variability—is dually influenced by cholinergic and adrenergic activity, as well as other physiological inputs. In the following we use the term vagal tone for the part of the autonomic nervous system measured by rMSSD. For frequency domain measures, beat-to-beat fluctuations were transformed to the frequency domain using Fast Fourier Transformation. Spectral power was determined over three frequency regions of interest: Very low frequency (VLF, < 0.04 Hz), low frequency (LF, 0.04–0.15 Hz) and high frequency (HF, 0.15–0.4 Hz) with derived HF/LF ratio. LF reflects mostly sympathetic activity, HF vagal tone. The methods of 24 hour ECG and analysis of heart rate variability have been described earlier [5].

Statistical Analysis

Groups were compared with controls by means, standard deviations and t-testing.

P-values below 0.005 were considered as statistically significant.

From anthropometrical data percentile values and BMI have been calculated using the free myBMI web page (nowadays changed to myBMI4Kids) according to percentile data by Kromeyer-Hauschild et al.

Ethics

The FDA issued a Safety Alert in August 2011, reporting that a French study concluded that people with certain types of short stature treated with recombinant human growth hormone during childhood and who were then followed over a long period of time, were at a small increased risk of death when compared to individuals in the general population of France. This increased mortality rate is related to cardiovascular events [13] and/or stroke [14]. We routinely communicate this enhanced cardiovascular risk to all parents prior to growth hormone treatment and propose a monitoring of cardiovascular risk by 24 hour Holter ECG and NT-BNP measurements as part of routine blood tests for short stature. We received an informed consent from nearly all parents of children with growth failure, but we certainly disregard performing these examinations if the parents refuse. All parents and patients (if appropriate according to patient`s age) enrolled in this analysis have given oral informed consent.

Autonomic imbalance in children with symptomatic short stature

Autonomic imbalance measured by HRV is a predictor of metabolic risk, cardiovascular disease, diabetes and early mortality [5]. We now clearly demonstrate a significant autonomic imbalance in children with short stature due to a history of small for gestational age, congenital syndromes and congenital heart disease. These children have a well-known enhanced cardiovascular risk.

Remarkably different patient groups show different patterns of autonomic imbalance. Increased heart rate is found in the SGA and the congenital syndromes group. Reduced SDNN is found in the syndromes and in the growth hormone deficiency group. RMSSD reduction can be diagnosed in the SGA, the cardiac growth failure and the syndromes group. HF component is reduced in the cardiac growth failure and the syndromes group. These different patterns of autonomic imbalance suggest the affection of different regulatory properties or different types of lesions.

Because HRV measurement can be performed and repeated easily this method provides the possibility of monitoring individual cardiovascular risk at an early stage as well as the impact of pharmacological treatment such as growth hormone. It may be used as an additional diagnostic tool combined with measurement of blood pressure, body weight or serum lipids in order to asses risk stratification. Moreover our data clearly indicate that short stature per se is not a factor for an autonomic imbalance or stress disease as shown in 25 children with constitutional growth delay. We agree with DE Sandberg et al. [15] that short stature as an isolated physical characteristic is no cause of psychological diseases. The impression of an association between short stature and psychological problems has been based largely upon methodologically weak studies that have typically confounded short stature with other medical and neurocognitive problems due to the underlying diseases like small for gestational age, congenital syndromes, congenital heart disease and probably prematurity.

Focusing on the group of SGA patients we provide evidence for autonomic imbalance in our group of patients, which is described by ongoing growth failure—more precisely by the lack of catch up growth. An adult group of SGA in the study of Schäffer et al., which showed normal growth, was autonomically healthy [16]. So the individual ability of an SGA patient to generate catch up growth seems to be related to the integrity of autonomic regulation. By this the lack of catch up growth seems to be a clinical marker for cardiovascular risk within this group of patients.

Early life stress in children with SGA or heart failure

With respect to our groups of patients with heart failure and SGA we are investigating patients who experienced severe stress during different episodes in early life. SGA patients are most likely to have experienced caloric restriction in utero resulting in intrauterine growth restriction. The group of SGA premature infants shows SDNN and rMSSD in the lowest range (Figs 2 and 3). From animal models there are data from literature describing the impact of intrauterine nutrition on an individual’s regulatory properties mainly on the level of hypothalamic regulation [17, 18]. In accordance with Barker’s hypothesis our data indicate that birth weight is an important predictor for autonomic imbalance in later life.

Referring to the heart failure patients there has to be recognized an episode of severe stress resembled by well-known activation of neuro humeral axis, release of renin, cortisol and norepinephrine. Heart failure in this group of infants was severe indicated by extraordinary NT-Pro-BNP value of 16437 ± 7066 pg/l. As shown in Figs 2 and 3 infants with early life stress due to heart failure or SGA have reduced rMSSD in infancy and most of these children with height below the third percentile in later life further have low rMSSD values. Recently we published longitudinal data that show that reduced HRV in later life due to congenital heart disease may be related to reduced HRV in early infancy [19].

As shown in the literature there seems to be a common pathway resulting from early life stress leading to elevation of NT-Pro-BNP. This constellation is shown in children with failure to thrive [20] as well as in infants with heart failure [21] but also in infants with small gestational age at birth [22]. Circulating norepinephrine levels in girls with SGA are elevated and the renin angiotensin aldosterone system in boys with SGA is activated. These hormonal changes correlate with blood pressure at the age of 10 years [23].

A possible example for autonomic imprinting?

We speculate to encounter here an underlying mechanism of early life stress which is imprinting the development of impaired autonomic regulation. This idea may explain concordant effects of nutrition and stress on metabolic regulation, growth and neurodevelopment:

1. We have shown recently that nutrition has prominent and reproducible effects on the autonomic nervous system [24].
2. Heart failure in infancy like other severe diseases causes sympathetic activation, suppresses parasympathetic tone and is related to growth failure, as well as attention deficit disorder [19].

A. Waterland and C. Garza defined the term metabolic imprinting as a “basic biological phenomenon that putatively underlies relations between nutritional experiences during early life and later diseases” [25]. For their definition the following conditions are characterized:

1. “a susceptibility limited to a critical ontogenetic window early in development” (the critical period)
2. “a persistent effect lasting through adulthood” (autonomic disorder)
3. “a specific and measurable outcome” (height, cognition, cardiovascular disease), and
4. “a dose-response or threshold relation between a specific exposure and outcome”.

There is a clear historical precedent for this definition: Konrad Lorenz chose “imprinting” to refer to the setting of certain animalistic behavior that resulted from early experience. Central to his definition was the fact that imprinting may only occur during “a narrowly defined period in the individual’s life” (the critical period), moreover that the imprinted behavior cannot be ‘forgotten’.

We assume different critical periods in early life for autonomic imprinting:

1. Genomic imprinting seems to be an epigenetic phenomenon most of all in some congenital syndromes. One well known mechanism is the parent-of-origin-specific genomic imprinting. If the allele inherited from the father is imprinted only the allele from the mother is expressed and vice versa.
2. Gestational imprinting mainly by fetal malnutrition results in a baby small for gestational age. Barker was the first who used the term fetal programming in his hypothesis [26].
3. Neonatal imprinting during the first 4 weeks of life occurs mostly due to severe disease like premature birth or feeding problems.
4. Infant imprinting during the first year of life is greatly due to severe disease like heart failure. Preoperative heart failure is one of the most stressful life events in infancy and has been neglected as a well-defined early life stress model with longtime consequence like growth failure and ADHD. We use this model owing to the fact that in most infants stress is limited to a well-defined time period and is terminated by an operative procedure at a distinct time point (predominantly with an age of approximately 6 month in heart defects with left-to-right shunts) [24].
5. Childhood imprinting during early childhood due to severe disease, neglect, trauma and abuse.
6. We are fortunate to have only a small number of children with short stature owing to starvation in Germany. However in 2010, 171 million children under 5 years of age suffer from stunting globally, with 98% being from low- and middle-income countries. Weight gain in the first two years of life is an important predictor of schooling outcomes in pooled analyses from five birth cohorts in these countries [1]. The understanding of child growth patterns is critical to the development and evaluation of appropriate interventions. Providing small-quantity lipid based nutrient supplements significantly increased growth and reduced stunting, wasting and anemia prevalence in young burkinabe children [27].

By understanding autonomic imprinting we are aware of the detrimental longtime effects of early life stress on growth and cardiovascular risk. Remarkable in our data is the high incidence of ADHD suggesting a relationship between symptomatic short stature and cognitive and behavioral properties. Improving care and nutrition for mothers and children within the first 1000 days after conception is one of the most promising public health policies [8].

Moreover if early life stress is not preventable due to severe diseases like heart failure or SGA, we may be able to improve long term outcome by targeted treatment of stress with beta blockers or supplementation with omega-3-fatty acids. Measurement of HRV as outlined here may help to monitor and evaluate these treatment options during further trials.

However, our data from a small pediatric department are observational data from a small group of patients. Further prospective clinical studies will be needed to identify critical periods of infant development and to evaluated treatment strategies.