PROGRAMA DE INVESTIGACION
Descripción del programa de actividades de la investigación (Description of the research activity programme)
In the last four years, our network (Red SAMID), and previously most of its research groups, have been studying different aspects on two key areas on paediatric growth and development:
1) the factors causing brain injury in Childhood and thus affecting neurodevelopment, and
2) the early nutritional factors related to the latter development of the metabolic syndrome in preadolescent children.
Although these two topics might not seem to be related at first though, indeed there is a close links between them since both affect the growth and development processes from the foetus to the adolescence, areas of expertise of the research groups that are included in our research network.
1) Factors causing brain injury and thus affecting neurodevelopment (ND). The developing brain is especially vulnerable to many different insults that can induce impairment of its growth and development, and cause later in life neurodevelopment (ND) delays (cerebral palsy, mental retardation, language or learning and behavioural disorders...). Those insults to the brain occurred during the foetal, neonatal or early post-neonatal periods, and although the specific mechanisms are complex and vary, all are related to cerebral hypoxia and ischemia, and reperfusion (Hilario E. Current Pediatr Review 2006;2:131).
The aetiology of the ND disability is multiple, and the most frequent causes are different in the foetus -intrauterine growth restriction (IUGR), exposure to drugs and environmental toxic substances, malformations...)-, in the neonate -hypoxic-ischemic encephalopathy (HIE), infections, and cerebral lesions related to prematurity like severe Intraventricular haemorrhage (IVH), periventricular leukomalacia (PVL)-, or in the early infancy -infections, injuries, toxics, nutrition…- (Mwaniki MK et al. Lancet 2012;379:445).
Although advances in obstetrics and intensive care management have improved the survival of those target populations, impairment of ND has not decrease in the same magnitude. Thus, research on scientific aspects related to its causes and its health consequences, specially related to its prevention are much needed.
The different ND disorders produce great stress to parents, families and communities; consume large resources in the health and educational systems for follow-up, diagnosis, management and rehabilitation. Moreover, affected children might not be able to achieve and ability to self support and care for themselves and thus a full integration in society.
Although the incidence of some of these disorders has been specifically measured, the overall burden and long-term sequelae have not been capture in estimates of the total burden of disease measured by disability-free years, either in Spain or at an international level (Olusanya BO. PloS 2007;4:e84).
2) Early nutritional factors related to the latter development of the metabolic syndrome in preadolescent children. Childhood obesity is one of the main problems affecting children in developed countries and some developing ones, having a real epidemic proportion. In Spain, its prevalence has been increasing steadily, and is been estimated that affects to a 27% of children, one of the highest rates of all European countries (Lobstein T et al. Obes Rev 2003;4:195). The causes of obesity are multiple, been influenced by genetic and environmental factors, although over-feeding is the single most important one.
Obesity plays a key role on insulin resistance and the metabolic syndrome (MS) with severe cardiovascular -hypertension- and metabolic risk factors (abdominal obesity, hyperglycemia due to type-2 diabetes, dyslipidemia) (Simmons R. Pediatr Clin N Amer 2009;56:449). However, only some obese susceptible individuals develop the MS, generally attributed to the interaction of genetic and environmental factors.
Many human and experimental studies have shown the relationship between low-birth-weight and obesity, insulin-resistance and the MS. Recently studies showed that fetal IUGR is an independent risk factor for insulin-resistance that it is amplified by obesity in childhood. In fact, those observations following the initial Barker-hypothesis (Baker DJ et al Lancet 1986;1; 1077) introduced the concept of early programming induced by altered intrauterine environment related to the maternal or fetal protein restrictions, and induces epigenetic gene regulation, in the context of the developmental origins (foetal) of health and cardiovascular disease (Gluckman PD et al 2004;15;183). The concept of “developmental plasticity” is now used to describe the phenomena of generation of alternative genotypes forming a single genotype by the action of environmental factors during development (Gluckman PD et al. J Intern Med 2007;261;46).
On the other hand, produced much efforts to develop effective preventive strategies, but unfortunately little has been achieved so far.
Our research network, by its multidisciplinary approach, is formed by researchers and clinicians in obstetrics, neonatal and childhood health specialists as well as some basic scientists, so it seems especially well equipped to undertake this program and fulfil the objectives of the proposed scientific program. For this reasons, as well as for the experience on working together toward a common goal in these fields for the last four years, we believe on the importance and dimension of our research program both, at a national and international levels, since we have human and equipment resources and expertise to study the topics proposed from the bench -animal models- to the bed side -epidemiological and clinical trials-.
The state of the art in both Spain and in the international context is similar, as far as the clinical management of both cerebral damage and neurodevelopmental delays, and obesity and the MS. However, on the research aspects, in Spain there are no previous multidisciplinary research efforts in this topic, not only in Spain but there are some excellent studies on single aspects of the problems. The horizontal research approach -from the foetus to the adolescence- and the broad multidisciplinary groups involved, makes this research program almost unique even at an international level.
Hypothesis. There are perinatal and postnatal factors that lead to neurologic, nutritional and metabolic disorders from prenatal period to adolescence, that subsequently condition adult life health. The study of some of this factor might contribute to its prevention.
The Scientific Programme has its main strategic objective on the prevention and treatment of the perinatal and post-neonatal risk factors involved in the development of neurologic, nutritional and metabolic problems, from the foetal age to the adolescence.
More precisely, the operative scientific objectives are related to, 1) the factors causing brain injury and thus affecting neurodevelopment and 2) the early nutritional factors related to the latter development of the metabolic syndrome in preadolescent children.
To ensure the accomplishment of these two operative specific objectives, the research programme will be approached in several Work Packages (WP) appropriately linked and coordinated. Each WP will approach a specific objective that will be analysed by a few specific tasks. For each task, an outcome will be produced, that could be measurable to assure the correct progression of the program. Moreover, a timetable and a list of key milestones will be established for similar proposes.
Each of the two main areas of the programme will be studied in a horizontal timeline, from the foetal stage to the adolescence as appropriate. So, prenatal, neonatal and post-neonatal WP will be set for each of the two main study lines, as indicated in the next paragraphs.
WP-1. PRENATAL RISK FACTORS FOR ABNORMAL NEURODEVELOPMENT AND NUTRITIONAL AND METABOLIC OUTCOME.
General objective. To study on IUGR biomarkers able to predict the postnatal development of disorders of ND and MS.
1.1 To establish a protocol for the diagnosis and obstetrical management of IUGR agreed by all network centres.
1.2 To study the interrelation between epidemiological, toxic environmental, socio-economic variables and maternal risk factors with the alteration/change in ND and MS
1.3 To study the relationship between Doppler hemodynamic compromise of umbilical blood flow and alteration of the ND and MS.
1.4 To study the interrelation between immediate perinatal complications with of ND and MS
1.5 To develop a foetal model of IUGR, to study factor affective fetal growth and test possible preventive and therapeutic strategies.
WP-2 NEONATAL RISK FACTORS FOR IMPAIRED NEURODEVELOPMENT OUTCOME.
General objective. To study early biochemical, biophysical and brain image biomarkers able to predict the postnatal development of disorders of ND.
1. To analyse the changes in the Redox status in foetal to neonatal transition, and the oxidative damage to membrane lipids, neuronal cell membranes, nitrosative damage caused to circulating proteins and inflammation caused to DNA.
2. Metabolic changes due to fetal to neonatal transition, oxygen supplementation, hypoxia, hyperoxia, etc. or factors changing Redox status (inflammation, infection, ventilation, etc.),
3. To study neonatal risk factor for brain injury and adverse ND in relation to: cerebral oxygenation, transient cardiovascular instability and hypoxic-ischemic encephalopathy.
4. To developed and integrated data acquisition e-system to validate the development of algorithms to be use of early biomarkers of brain and cardiovascular dysfunction.
5. Stalish animal models of neonatal hypoxic-isquemic brain injury.
WP-3 NEONATAL RISK FACTORS FOR ADVERSE NUTRITIONAL AND METABOLIC OUTCOME.
General objective. To study neonatal risk factors for adverse nutritional and metabolic outcomes and possible preventive nutritional strategies.
3.1. Improve the ND of premature infants by changing the diet with the intake of DHA of the breast milk
3.2 To study the changes experienced by the fresh and pasteurized human milk throughout the administration process
3.3 To study the effect of nutritional status at discharge on body composition and glucose intolerance, and on later ND
3.4 To Know which probiotic allows better colonization of the gastrointestinal tract and better intestinal development, growth and neurodevelopment.
3.5.To learn the effect of a high protein diet on protein synthesis and overall synthesis of certain proteins such as albumin in critical pediatric patient.
WP-4. POST-NEONATAL RISK FACTORS FOR IMPAIR NEURODEVELOPMENT OUTCOME.
General objective. To identify risk factors of postnatal neurological development of children and evaluate the effectiveness of diagnostic methods on prevention and treatment.
4.1 To identify populations and postnatal risk factors for major ND impairment and neurological disorders in childhood: cardiac arrest , heart surgery, traumatic brain injury, stroke and poisoning.
4.2. To study the usefulness of methods for early detection of risk factors and neurological disorders: Neuroimaging methods: ultrasound, CT, MRI, PET, Monitoring of cerebral blood flow, Saturation brain using near-infrared spectroscopy (NIRS) ECG, Biochemical biomarkers.
4,3. To study the efficacy of prevention and treatment methods to reduce neurological disorders,/.Oxygenation: to analyze the influence of the fraction of inspired oxygen during cardiopulmonary resuscitation and treatment of hypoxic-ischemic alterations in the development and prevention of acute neurological injury.
4.4 To develop animal models of diagnosis, prevention and treatment of neurological diseases that occur in pediatric animal models of hypoxic neurological injury, ischemic neurologic injury and cardiac arrest.
WP-5. POST-NEONATAL: RISK FACTORS FOR NUTRITIONAL AND METABOLIC ADVERSE OUTCOME.
General objective. To study risk factors for adverse nutritional and metabolic outcomes in early childhood and possible preventive interventional strategies.
5.1 To investigate subclinical cardiovascular disease in children 8-10 years old, with perinatal developmental factors (IUGR or extreme low birth weight) that potentially influences the future risk of this disease.
5.2. To define non-invasive approaches to identify children with early changes in cardiovascular physiology that potentially affect future cardiovascular outcome, emphasizing their potential applications in childhood.
5.3 To evaluate the biological effects of early and realistic interventions in the selected population, analyzing potential changes in the defined biomarkers.
5.4 To investigate the influence of genetic variants on the development of obesity and yo evaluate the association between those genetic variants and food habits, physical activity and biomarkers of inflammation, cardiovascular diseases risk and oxidative stress.
WP-6: EPIGENETIC, TOXIC AND ENVIRONMENTAL RISK FACTORS FOR ABNORMAL NEURODEVELOPMENTAL, NUTRITIONAL AND METABOLIC OUTCOME.
General objective. Research on prenatal and postnatal environmental factors related to neurological, nutritional and metabolic disorders from prenatal period to adolescence.
6.1. Development of analytical methodology to describe and validate biomarkers of prenatal and postnatal exposure to substances and drugs of abuse, drugs of prescription, persistent organic toxics and heavy metals in different alternative matrix.
6.2 To study the prevalence of prenatal and postnatal exposure these several substances on prospective and retrospective cohorts of newborn infants born in different regions of Spain, and to perform follow-up of the cohorts of prenatally exposed newborns.
6.3 To develop animal models of prenatal exposure to xenobiotics, mainly alcohol to define biomarkers of different substances (parent substances and metabolites) to which foetus, newborn, child and adolescent can be exposed
To assure the accomplished of all specific objectives included in the six wp, a complete and detailed Work plan has been developed. It includes: 1) details about the human resources available (personnel provided by the institutions of the network partners and those specifically hired with the project's funds) as well as equipment (mainly provided by the partners from other sources), 2) details about the tasks planned to achieve the specific objectives just outlined, 3) a time schedule for all tasks and deliverables, 4) a detailed budget, and 5) a set of indicators to measure the research have been developed. Details on all of these different components of the Work plan are described on the appropriate sections of this proposal.
The innovative aspects of this proposed research project are not only related to the horizontal and multidisciplinary approach to be implemented, but also in regard to the real possibilities for several technological advances to be transfer to industry and eventually to clinical use. Some groups have or are developing several devices or systems. In this regard, among others, a device to be used to prevent preterm birth has been developed and proven effective (Goya M et al. Lancet 2012, April 3, 2012 DOI:10.1016/S0140-6736(12)60030-0), prototypes of devices to deliver nebulised exogenous surfactant without need for intubation in preterm and term neonates with severe respiratory failure, and a multichannel integrated system to capture and analyse multiple electronic signals from brain, cardiovascular and pulmonary function, will be eventually used for monitoring and treatment decisions..
Furthermore, several of the research groups have register patents to support liquid ventilation (Alvarez et al. No pat 2006/03096, Alvarez et al. No 1999/01420), that proves their ability to transfer rapidly research findings into technological innovations, assuring that it will also happen in this project.
Both human and equipment resources have been summarised in the previous section “Scientific and technological background of the participating groups”.
Below are the tasks to be developed in relation to the specific objetives of each WP are brieftly descrived below. Moreover outpots for each tasks are also summarised.
WP1. PRENATAL RISK FACTORS FOR ABNORMAL NEURODEVELOPMENT AND NUTRITIONAL AND METABOLIC OUTCOME
In order to achieve the aims of this WP, the network will perfom the following tasks: 1) Establish a protocol for diagnosis and obstetric management of IUGR. 2) Design of an electronic notebook for data collection. 3) Study the correlation between epidemiological, toxic environment, socio-economic variables and maternal risk factors with altered and postnatal metabolic syndrome (MS). 4) Study on the correlation between Doppler hemodynamic compromise with impaired ND and postnatal MS. 5) Study on the correlation between immediate perinatal complications with impaired ND and postnatal MS. 6) Design of a bank from blood samples of: mothers, umbilical cord, neonates and children diagnosed with IUGR. 7) Development of animal IUGR model.
After reaching these tasks, the following outputs will be obtained: 1) Protocol for diagnosis and obstetric management of IUGR. 2) Provide 80% of the epidemiological, toxic environmental, socio-economic variables and maternal risk factors of patients enrolled in the study. 3) Have Doppler hemodynamic variables within a given period of less than 10 days after delivery in 100% of cases and controls. 4) Record of perinatal data in 80% of study population. 5) Disposal of blood samples from mothers in 50% of cases and controls, umbilical cord in 30% of infants and children.6) Using the animal model for testing different treatments with clinical potential.
In WP1, we will implement a clinical guideline for managing risk pregnancies with IUGR and fetal hypoxia.
WP2. NEONATAL RISK FACTORS FOR IMPAIRED NEURODEVELOPMENT OUTCOME
In order to achieve the aims of this WP, the network will develop the following tasks: 1) Study of cerebral hemodynamic imaging, macro- microcirculation and oxygen delivery (Cerebral Monitoring Techniques for the prevention of brain injury of prematurity). 2) Analysis of biomarkers involved in brain injury. 3) Development of strategies to prevent brain injury in neonatal hypoxic-ischemic encephalopathy. 4) Animal model of hypoxic-ischemic brain damage. 5) Design of electronic systems used as early markers of brain and cardiovascular damage.
After reaching these tasks, the following outputs will be obtained: 1) Clinical guidelines for the management of cerebral oxygenation in the extremely preterm during the transitional stage of movement. 2) Score of biomarkers in the premature circulatory failure, used to direct therapeutic interventions. 3) Score adverse prognostic risk based on analysis of parameters of different integrated monitoring systems 4) Pharmacokinetics, tolerability and toxicity of different experimental treatments in asphyxic neonates undergoing moderate hypothermia. 5) Testing of different treatments with clinical potential in animal models, and 6) Patent of electronic systems designed.
In summary, this WP2 will implement a clinical guideline for management of cardiovascular instability and the hypoxic-ischemic encephalopathy.
WP3 NEONATAL RISK FACTORS FOR ADVERSE NUTRITIONAL AND METABOLIC OUTCOME
In order to achieve the related aims of this WP, the network will develop the following tasks:
1) Design of a randomised controlled study to evaluate the effect of dietary advice on the concentration of DHA in breast milk. 2) Description of the relationship between nutritional status at discharge and sensitivity to insulin. 3) Description of the influence of probiotics administered in intestinal colonization in very preterm infants 4) Study of the effect of diet on overall protein metabolism and on specific individual protein synthesis. 5) Establishment of nutritional losses, immunological, antioxidants factors and vitamins that suffer mother's milk after HTS pasteurization and compare it to Holder pasteurization. 6) Evaluation of the effect of speed and infusion time for power management systems, and different homogenization treatments (routine, manual, ultrasonic), change in lipid content, caloric and immunoglobulins and cytokines in thawed milk.
After reaching these tasks, the following outputs will be obtained: 1) Dietary survey. Relation between intake of DHA estimated by survey and longitudinal concentration of DHA. A clinical trial will be flamed. 2) Relationship between insulin sensitivity and nutritional status at discharge. Study sample size estimation to evaluate the effect of nutritional status on neurodevelopment. 3) Effect of administered probiotics on intestinal colonization during the initial admission. Changes in intestinal colonization of very preterm children during the following months of discharge. 4) Effect of diet on overall protein synthesis on the splanchnic sequestration of amino acids and the synthesis of specific proteins. 5) Comparative table of nutritional losses, immune factors, antioxidants and vitamins of Holder pasteurization and HTST. Clinical guideline on the HTST pasteurization process for use of donated human milk banks. 6) Clinical guideline on the management of mother's milk. 7)Design of an infusion system that incorporates a method of homogenization.
In summary, develop a dossier of recommendations on the implementation of human milk banks following Good Laboratory Practice.
WP4. POST-NEONATAL RISK FACTORS FOR IMPAIR NEURODEVELOPMENT OUTCOME
In order to achieve the related aims of this WP, the network will develop the following tasks: 1) Design and development of a multicenter prospective observational studies of risk factors for neurologic impairment secondary to cardiac arrest, congenital heart disease, cardiac surgery, or secondary injury in children with long-term observation and to encourage the participation in the international multicenter study on pediatric stroke and to perform the control of long-term Spanish cohort. 2) Correlation and predictive capacity of alterations in computerized tomography (CT), magnetic resonance imaging (MRI), positron emission tomography (PET), cerebral blood flow, cerebral saturation measured by near-infrared spectroscopy (NIRS), electroencephalographic (EEG) abnormalities and biomarkers of neurodevelopment damage with acute neurological and long-term neurological deficits. 3) Correlation between FiO2, respiratory rate and/or PaCO2 used in cardiopulmonary resuscitation and mechanical ventilation with acute neurological and long-term neurological deficits in children. Multicenter prospective study to analyze the effect of hypothermia in the prevention of acute neurological and long term after cardiac arrest in children beyond the neonatal period. 4) Animal model of pediatric neurological injury induced by: ischemia, hypoxia and cardiac arrest. 5) Comparison of different respiratory rates and FiO2 on cerebral blood flow, cerebral saturation measured by NIRS and biomarkers of neuronal hypoxic-hyperoxic damage on randomized experimental studies of cardiac arrest, hypoxia and ischemia.
After reaching these tasks, the following outputs will be obtained: 1) Multicenter study protocol of neurological disorders in children with: cardiac arrest, congenital heart disease and cardiac surgery. Report of participation in the multicenter study of stroke in children. 2) Clinical guidelines recommending neuroimaging studies in children at risk of acute neurological disorders. Clinical Guide recommendation for the use and interpretation of measurements of cerebral blood flow, cerebral saturation measured by NIRS, EEG and brain damage biomarkers in children with risk of neurological disorders. 3) Clinical guidelines on the FiO2 during cardiopulmonary resuscitation in children. Clinical guidelines on respiratory rate during cardiopulmonary resuscitation in children. Multicenter study protocol of hypothermia after cardiac arrest in children. Report of potentially neuroprotective drugs during hypoxia-ischemia and biomarkers of neurodevelopment damage in childhood to raise future clinical studies. 4) Pediatric animal model for ischemic neurological damage, for hypoxic neurological damage, for cardiac arrest. Report of the alterations of cerebral blood flow and cerebral saturation by NIRS in experimental models of acute neurological injury. Report of hypoxic-hiperoxic biomarkers alterations in experimental models of acute neurological injury. Experimental study protocol of the influence of FiO2 in pediatric cardiac arrest. Experimental study protocol of the influence of respiratory arrest in pediatric cardiac arrest.
In summary, a clinical guideline will be proposed for the early management of the various disorders that can cause brain damage.
WP5. POST-NEONATAL RISK FACTORS FOR NUTRITIONAL AND METABOLIC ADVERSE OUTCOME
In order to achieve the related aims of this WP, the network will develop the following tasks: 1) Subjects recruitment for different risk situations. 2) Assessment of personal and family history of the metabolic syndrome components. 3) Clinical examination, including clinical signs of insulin resistance and maturation status, blood pressure, and anthropometry. 4) Assessment of nutrient intakes and food habits using a standardized interview computer assisted 24h recall and Food Frequency Questionnaire. 5) Measurements of traditional biomarkers associated with insulin resistance and metabolic syndrome. 6) Measurements of biomarkers associated with insulin resistance, inflammation and cardiovascular diseases risk. 7) Measurements of parameters associated with oxidative stress. 8) Analysis of genetic variants. 9) Detection of cardiovascular subclinical alterations (echocardiography), potentially related with future cardiovascular diseases risk.10) Effect of classic intervention (nutrition and physical activity) on plasma and vascular parameters. 11) Statistical analysis.
After reaching these tasks, the following outputs will be obtained: 1) Report on the influence of genetic variants on the development of obesity. Results from the case-control study. 2) Report on the relationship between genetic variants and food habits, physical activity and biomarkers of inflammation, cardiovascular diseases risk and oxidative stress. 3) Report on the noninvasive (echocardiography) cardiovascular measurements and its correlation within the cardiovascular diseases risk factors. 5) Report on the efficacy of the intervention in terms of cardiovascular diseases risk factors reduction.
Protocol to define and to prevent metabolic syndrome in children by the use of early biomarkers. This protocol will be defined by consensus and based in our results.
WP6. EPIGENETIC, TOXIC AND ENVIRONMENTAL RISK FACTORS FOR ABNORMAL NERURODEVELOPMENTAL, NUTRITIONAL AND METABOLIC OUTCOME
In order to achieve the related aims of this WP, the network will develop the following tasks: 1) To define paediatric alternative matrices useful for the determination of biomarkers of damage, or exposure. Furthermore, to create a biobank of clinical samples. 2) To define biomarkers of different substances (parent substances and metabolites) to which foetus, newborn, child and adolescent can be exposed (alcohol, tobacco, drugs, toxics, heavy metals). These biomarkers can be used in studies of prevalence of prenatal or postnatal exposure or in studies of tissue damage due to exposure. 3) To describe and validate the analytical methodology for each biomarker in every alternative matrix. 4) To determine the prevalence of prenatal and postnatal exposure to several substances. 5) Clinical follow up of the cohorts of prenatally exposed newborns and children to several substances. 6) Research on pharmacokinetics of drugs of prescription in children (clinical trials with drugs of prescription). 7) Development of animal models of prenatal exposure to several substances.
After reaching these tasks, the following outputs will be obtained: 1) List, detection time window, analytical methodology, significance and importance, and applicability of the alternative matrices. 2) List and table of biomarkers (parent substance, metabolites) of exposure of different substances. 3) Description of the analytical methodologies and its applicability. 4) Prevalence figures of prenatal and postnatal exposure to several substances. 5) Clinical and analytical results derived from the follow up of several substances prenatal and postnatal exposure. 6) Pharmacokinetic results and prescription recommendations and indications of medicines in children. 7) Description of animal models of prenatal exposure to several substances.
Presentation of clinical guidelines, for example, human milk banks, and environmental toxic (legal and non legal drugs).
In summary, our network propose to study two hot topics in Paediatrics: 1) the factors causing brain injury, affecting neurodevelopment; and 2) the early nutritional factors related to the latter development of the metabolic syndrome in children. Our multidisciplinary research network has been formed by basic researchers and clinicians in obstetrics, neonatal and childhood health specialists to undertake the program and to fulfil the aims, with adequate human and equipment resources. The horizontal research approach (from foetus to adolescence) and the multidisciplinary groups (from bench to bed side), makes this research program almost unique even at international level. These topics will be approached by our research network in six work packages, which are our expertise area. Finally, we propose that the main outcomes obtained from the Work Plan will be freely presented to all Spanish health stakeholders, international scientist and the Spanish society as a whole for its dissemination and spreading.