Analyses of Anemia Risk Factors and Prognosis in VLBW Infants.
Conditions: Very Preterm Maturity of Infant, The Risk Factors for Improving the Prognosis of VLBW Infants
Sponsor: Shenzhen Bao'an Maternal and Child Health Hospital

Clinical trial：NO. NCT05949788
Document date: 2024-1-29
Project BA2023
study protocol and statistical analysis
plan
Title: Analyses of Anemia Risk Factors and Prognosis
in VLBW Infants.
Study center
Shenzhen Bao'an Women’s and Children’s Hospital (ShenzhenBMCHH)
Shenzhen Longhua Women’s and Children’s Hospital
Time frame
Study Start: March 1, 2023
Study Completion: March 24, 2024

Clinical trial：NO. NCT05949788
Document date: 2024-1-29
1. Background
Preterm infants (accounts for 9.9% of newborn infants in 2020) are born with immature organs and immune
system[1]. Especially, 4% of them are extremely preterm infants with a gestational age of less than 28
weeks[2].In recent years, the treatment and management of extremely preterm infants has become the main
task of NICU. Moreover, the morbidity rate for extremely premature infants around the world has been
rising annually.These vulnerable patients are prone to various complications. Anemia is one of the most
common complications that occur during hospitalization in these infants[3]. Therefore, According to a
survey by the Canadian Neonatal Network (CNN), over 75% of extremely preterm infants receive at least
one blood transfusion treatment during hospitalization[4], and WhyteRK et al.[5] even pointed out that over
90% of extremely preterm infants require blood transfusion treatment during hospitalization. Most of the
recipients of blood transfusions are caused by anemia, indicating a very high incidence and severity of
anemia in these immature infants. Anemia of prematurity (AOP) can cause a series of adverse
consequences such as apnoea, prolonged mechanical ventilation, prolonged hospital stay, necrotizing
enterocolitis (NEC) [6], retinopathy of prematurity (ROP) [7], infection, and extrauterine growth restriction
(EUGR). It can be seen that anemia in premature infants, especially in extremely preterm infants, has a
significant impact on their prognosis.
There is a lack of evidence regarding the relationship between blood hemoglobin level at birth and the
neonatal morbidity and mortality of preterm infants despite the mode of delivery and prenatal infection. It
remains unknown the impact of early anemia on the neonatal morbidity and mortality of extremely preterm
infants[8].With this study, We hope to demonstrate the relationship between anemia on admission to the
neonatal intensive care unit (NICU) and the neonatal morbidity and mortality in extremely preterm infants.
2. Objective and hypotheses
Objective: To study the risk factors of anemia in different stages of extremely preterm infants and their
impact on prognosis, which provide a certain evidence for improving the prognosis of these infants.
Hypothesis: Small gestational age and low birth weight are risk factors for anemia and receiving blood
transfusion treatment in extremely preterm infants. The younger the gestational age and the lower the birth
weight, the more severe the anemia, and the greater the likelihood of receiving blood transfusion treatment.
3. Study design
3.1. study subject
From January 1, 2020 to December 31, 2022, extremely preterm infants with a gestational age of less than
28 weeks were admitted to Bao'an Women’ s and Children ’s Hospital affiliated with Jinan University and
Longhua District Women’s and Children’s Hospital.
3.2. The method of study
This was a retrospective study on the maternal and infant clinical and electronic records. We analyzed the
maternal and obstetrical data, demographics, as well as the neonatal morbidity and mortality diagnosed
during hospitalization. The study was approved by the Ethics Committee of our institution.

Clinical trial：NO. NCT05949788
Document date: 2024-1-29
3.3. Inclusion criteria
Extremely preterm infants with admission age ≤24 hours, hospital stay ≥14 days, and gestational age less
than 28 weeks were included.
Anemia of Premature (AOP):

Within 2 weeks of birth, venous blood hemoglobin level (Hb)≤130g/L, capillary blood
≤145g/L.

Age 2 weeks to 1 month, Hb<110g/L.

Age after one month, Hb<90g/L.
3.4. Exclusion criteria

(1) Transfer or automatic discharge within 2 weeks.

(2) Patients with combined neonatal hemorrhagic diseases.

(3) Newborns with severe malformations (such as congenital diaphragmatic hernia, umbilical
protrusion, multiple malformations, etc.) or chromosomal abnormalities.

(4) Incomplete and inaccurate hospitalization data.
3.5. General information
Several variables on mother, pregnancy, delivery,resuscitation, hematologic parameters including the Hct
and Hb at admission, PRBC transfusions, use of vasopressor agents within the first 72 h of life, data on
invasive and noninvasive respiratory support, neonatal morbidity and mortality, and length of NICU stay
were collected and analyzed.
Care practices included antenatal steroid use, the use of antenatal magnesium sulfate, cesarean section, and
delivery room resuscitation.

Antenatal steroids: A full cycle of antenatal steroids was considered when at least 12 h had
elapsed after the last dose of dexamethasone (four doses of 6 mg of dexamethasone given
intramuscularly 12 h apart) administered to the mother [9].

Antenatal magnesium sulfate was defined as the intravenous administration of magnesium
sulfate to the mother during pregnancy at any time before delivery.

Gestational age (GA) was determined according to the first day of the last menstrual period,
an early prenatal ultrasound examination, and a physical examination of the infant after
birth[10].

Maternal chorioamnionitis was diagnosed clinically during pregnancy, labor, or delivery.

Small for gestational age (SGA) was defined as a birth weight below the 10th percentile,
based on the Fenton Growth Chart [11].

Neonatal sepsis was diagnosed in the presence of clinical signs (fever; hypothermia; lethargy;
tachypnea; apnea/bradycardia; cyanosis; and hypoglycemia not explained by other diagnosis),
associated to at least two laboratory markers (thrombocytopenia;leukocytosis or leucopenia;
C-reactive protein elevation; PCT; I/T) according to our National Program of Infection
Control Guidelines [12]. Early-onset sepsis and late-onset sepsis were considered if diagnosed
before and after the first 72 h of life, respectively.

Intraventricular hemorrhage (IVH) was classified according to Volpe [13].

Clinical trial：NO. NCT05949788
Document date: 2024-1-29

Bronchopulmonary dysplasia (BPD) was defined according to the National Institute of Health
[14]. For the analysis of this study,we considered BPD as the dependency for oxygen until 36
weeks of corrected age.

Parenteral nutrition was used as soon as there was clinical stability, preferably since day one
of life, starting with a volume of 70 ml/kg/day, with daily increments until 150 mg/kg/day by
day seven of life. Incubator humidity is usually 70% during the first week.

The modified criteria of Bell were used for the diagnosis and staging of necrotizing
enterocolitis (NEC) [15].

Staging of retinopathy of prematurity (ROP) was done according to the International
Classification [16,17].
3.6. Study outcomes
The study outcomes and endpoints are listed in Table 1.
Table 1. List of study outcomes and endpoints.
Outcomes
Endpoints
General information

Sex (male or female)

Pregnancy mode (In vitro fertilization or not)

Chorioamnionitis (Yes or not)

GBS colonization (Yes or not)

Early onset sepsis (Yes or not)

Antenatal steroids (Yes or not)

Delivery mode (natural delivery or cesarean
section)

Multiple births (Yes or not)

Gestational age (weeks)

Birth weight (grams)

Mother's hemoglobin level before delivery

Mother's HCT level before delivery
Neonatal morbidity and mortality

BPD

ROP

NEC

Early onset sepsis

Late onset sepsis

EUGR

IVH

Death

Clinical trial：NO. NCT05949788
Document date: 2024-1-29
4. Statistical analysis plan
Categorical variables were expressed as proportions n/N (%) and compared using the Chi-square (χ2) test.
Continuous variables were expressed as (mean±SD) or median and interquartile range (P25, P75), and
compared using Student’s t-test or a nonparametric test, as appropriate. Two-tailed p-values < 0.05 were
considered statistically significant. A multivariate analysis by logistic regression was performed to evaluate
predictors of neonatal morbidities. Statistical analyses were performed using IBM SPSS Statistics, version
20.

Clinical trial：NO. NCT05949788
Document date: 2024-1-29
Appendix:

Clinical trial：NO. NCT05949788
Document date: 2024-1-29
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