Probiotics and Breastmilk Are Associated With a Decreased Risk of Atopic Dermatitis in Very Low Birth Weight Premature Infants
Conditions: Atopic Dermatitis
Sponsor: José Uberos Fernández

 
 
 
 
 
 
 
PROBIOTICS AND BREASTMILK ARE 
ASSOCIATED WITH A DECREASED RISK OF 
ATOPIC DERMATITIS IN VERY LOW BIRTH 
WEIGHT PREMATURE INFANTS  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Study protocol and statistical analysis. 
August 9, 2023. 

STUDY PROTOCOL 
 
BACKGROUND  
 
Prematurity is associated with significant morbidity and mortality, including the 
prevalence of sepsis and gastrointestinal problems such as necrotising enterocolitis 
(NEC), among others. The aetiology is mainly due to immaturity, although numerous 
external factors play a role in aggravating its complexity. Some of these causes are 
increased gastrointestinal permeability, immaturity of the immune system, delayed 
colonisation and/or abnormal colonisation (dysbiosis), use of antimicrobials (sometimes 
prolonged), physiological limitation of the initiation of enteral nutrition... Some prenatal 
factors such as premature rupture of membranes or chorioamnionitis may also play a role, 
exposing them very early to pathogenic micro-organisms. These factors justify the higher 
risk of gut microbiota disruptions present in preterm infants compared to term infants. 
 
The human microbiota plays a fundamental role in the development and maturation of 
the immune system. The symbiosis between microbiota and humans is essential in both 
innate and acquired immune maturation, intervening in the maturation of T cells and 
natural killer cells, proper regulation of the Th1-Th2 response, as well as in the tolerance 
of intestinal endotoxins. 
 
Intestinal dysbiosis is conditioned by nutrition and healthy habits. In neonates, the main 
determinant of gut flora composition is diet and antibiotic exposure. Breast milk is the 
paradigm in neonatal nutrition, its microbiological content and macromolecules such as 
oligosaccharides in breast milk promote the growth of Bifidobacterium spp.  
 
The use of probiotics in preterm infants, a population exposed to unfavourable 
environmental factors, may favour a more physiological development of the microbiota. 
 
Numerous studies show that the use of probiotics in this population during the neonatal 
period reduces the development of NEC and late-onset sepsis in the short term. However, 
they also report other long-term beneficial effects on inflammatory, metabolic, 
neurological and gastrointestinal diseases, such as the reduction of allergic diseases 
throughout life, which is the focus of this study. 
 
In recent decades, a significant increase in atopic-based diseases has been observed, in 
the pathophysiology of which metabolic activity triggered by gut dysbiosis seems to play 
a major role, as it has been observed that it may precede the onset of atopic dermatitis. 
 
Atopic dermatitis is a chronic recurrent inflammation of the skin that leads to increased 
permeability, and therefore a loss of the skin barrier and increased risk of allergic 
sensitisation. 
 
Several studies show the benefits of daily oral probiotics in the clinical improvement of 
atopic dermatitis and other atopic-based diseases with chronic inflammation, due to their 
effect on the development of immune tolerance. In the case of atopic dermatitis, a 
significant reduction in symptomatology and severity has been observed, as well as a 
reduction in the use of topical corticosteroids.  
 

Factors involved in the early maturation of the immune system will condition the long-
term development of atopic dermatitis. The use of oral probiotics composed of 
Lactobacillus spp. and Bifidobacterium spp. in the neonatal period may be a promising 
strategy against the development of atopic dermatitis due to their collaboration in the 
maturation and regulation of the immune system. 
 
However, there is a lack of uniformity in the literature regarding the type of strains of 
choice, applicability of probiotics as prophylaxis or treatment, and in some of them even 
notable differences in results. 
 
DESCRIPTION 
 
In recent years there has been an increase in allergic diseases, which can be explained 
pathophysiologically by a dysregulation of the immune system. There are several lines of 
research into the cause of this increase, the best known of which is the "hygiene theory". 
Most agree that it could be due to a lack of immunological tolerance from a very early 
age. 
 
The gut microbiota is directly involved in mediating immune tolerance to all antigens, 
including pathogenic and commensal mycobacteria, exposed at the level of the intestinal 
lumen. Currently, the use of probiotics is very popular in multiple diseases of an 
immunological and inflammatory nature, due to their interaction and modification of the 
microbiota, and therefore, with the immune system.  
 
In neonatology, the use of probiotics is becoming increasingly widespread as a preventive 
measure against necrotising enterocolitis and late-onset sepsis. Based on theories about 
the interaction of probiotics with the immune system, it may be that the use of probiotics 
in this population may offer other medium- to long-term benefits in inflammatory 
diseases. 
 
OBJETIVES  
 
The main objectives are as follows:  
1. To evaluate the association between nutritional factors in the neonatal period and 
atopic dermatitis, in relation to prevention. 
2. To assess whether the use of breastfeeding and probiotic supplementation in very 
low birth weight infants as prophylaxis for necrotising enterocolitis is associated 
with a decrease in atopic dermatitis at preschool and school age.  
3. To determine probiotic strains that offer benefits in reducing the occurrence of 
atopic disease.  
 
The secondary objectives are set out below:  
1. To assess whether the use of breast milk (both fresh and donated) offers a 
synergistic effect, when combined with probiotics, in the reduction of atopic 
dermatitis.  
2. Impact of caloric and protein intake in very low birth weight infants on the 
development of atopic dermatitis later in life. 
 
DESIGN AND METHODS  
 

Observational restrospective cohort study in very low birth weight infants, <1500 grams 
and/or < 32 weeks, hospitalised in the Neonatal Intensive Care Unit at the Hospital 
Universitario Clínico San Cecilio between January 2009 and December 2021. 
 
Enteral and parenteral nutritional data, type of feeding (fresh and/or thawed breast milk, 
donated breast milk or formula), probiotic administration and strain administered are 
recorded. Parental permission is available for all patients included in the study. 
 
Inclusion and exclusion criteria: 
All patients hospitalised in the Neonatal Intensive Care Unit with < 1500 grams birth 
weight and/or < 32 weeks at birth during the indicated time period. Deceased patients, 
incomplete or unavailable medical history and patients with severe congenital diseases 
are excluded. 
 
Nutritional management: 
The unit's nutrition and fluid management protocol, both enteral and parenteral, follows 
the recommendations of the Nutrition and Metabolism Group of the Spanish Society of 
Neonatology. Daily fluid (ml/kg/day), carbohydrate (mg/kg/min), protein (g/kg/day), 
lipid (g/kg/day) and calorie (kcal/day) intakes are calculated.  
Given the characteristics of this group of patients, as described above, nutritional needs 
are often met with parenteral nutrition combined or not with enteral feeding. The 
objective is to cover the minimum needs to guarantee adequate growth and development 
of the newborn. 
In relation to enteral nutrition, the first choice is fresh breast milk, sometimes it is 
necessary to freeze it for preservation, and it is thawed prior to use. In cases of 
unavailability, with parental consent, donated milk that has been pasteurised according to 
Holder's method is administered, or failing that, formula for premature infants.  
Fortification with maternal or donated milk is started from one week of life, once water 
intake of at least 80 ml/kg/day has been achieved. 
In our unit, the use of probiotics was initiated in December 2013 as a preventive measure 
against the development of necrotising enterocolitis. Two strains were available: 
Lactobacillus Rhamnosus GG and the combination of Lactobacillus acidophilus + 
Lactobacillus bifidum. The choice of the type administered depended on the preference 
of the responsible paediatrician. The probiotic was introduced after initiation of 
satisfactory enteral nutrition and continued until 35 weeks postnatal or until discharge 
from the unit. 
 
Atopy registry: 
Atopy is a state of abnormal hypersensitivity to exposure or contact with non-pathogenic 
antigens. We record a history of atopic dermatitis and first-degree family history. 
 
STATISTICAL CONSIDERATIONS 
 
In our study we have two types of descriptive variables: continuous variables and 
categorical variables.  
- 
For continuous variables we will use the median and interquartile range, using the 
Mann-Whitney test for comparison.  
- 
For categorical variables, distribution frequencies will be used, analysing them by 
means of Chi-square.  

In order to study more than one variable on the desired effect, multivariate logistic 
regression analysis will be used.