Even with advances in neonatal intensive care, preterm infants [<37 weeks of gestational age (GA)] still face tremendous challenges to surviving and thriving. Notably, males born preterm are consistently observed to have a higher risk of perinatal complications, with worse mortality and morbidity (1–4), and neurodevelopmental impairments (5–7) than females. The etiology of the “sex bias” in preterm growth and health remains unclear and likely multifactorial (4). Sex variation in nutritional need is increasingly recognized as playing a significant role in the growth and health outcomes of preterm infants (8). Emerging studies show sex-specific responses to nutritional intake (8–13), suggesting potential sex-based nutrition requirements in early life for optimal development.

Mother's own milk (MOM) is ideal for preterm infants' growth, immunity, and neurodevelopment (14, 15). The amount and composition of MOM has been found dynamically adjusted to support the infants' nutritional requirements along with their developmental phase (16). Emerging research shows the interplay between the MOM characteristics and infant sex for tailoring sex-specific needs of the offspring (16). While mothers of term male infants are more likely to produce milk with higher levels of energy, free amino acids, total lipids, phospholipids, and gangliosides, which are significantly associated with infants' neurobehavioral development (16–20). There is limited evidence of sex-specific response to feeding in preterm infants. Moreover, it is challenging to conclude the impact of sex-specific MOM on the growth and development of preterm infants. This challenge arises from the intricate difficulties faced by mothers in terms of breastfeeding and providing MOM for their infants (15, 21).

Our study aims to examine the associations between sex-specific responses to feeding, especially MOM intake, and the growth and neurodevelopment of preterm infants from NICU up to 2 years of corrected age (CA). We hypothesize that male and female preterm infants have distinct feeding needs at the early stage of their lives, which contribute to their growth and neurodevelopmental outcomes. Therefore, we used linear mixed models to evaluate the significance of factors including feeding and clinical data, in predicting the growth and neurodevelopmental outcomes in both male and female infants. Further, we applied SHapley Additive exPlanations (SHAP) to determine the importance of risk factors on predicting neurodevelopmental outcomes.

Sex differences become apparent early in development and persist into later stages in preterm infants (35), however, the underlying mechanisms remain poorly understood. Feeding regimen has emerged as a potential factor that may influence the sex-differentiated growth in infants (36). Therefore, our study investigated sex-specific response, such as early life growth (1 to 16 postnatal weeks), to different feeding regimens. This timeframe aligns with the “window of opportunity” identified in other studies, playing a critical role in shaping infants' health outcomes (37). We also consider week 1 to 8 as “early time” and week 9 to 16 as “later time” to look at even closer to the sex-varied growth during this window of opportunity. We observed the association of both total feeding amount and proportion of MOM with the growth of males and females is sensitive to their postnatal age. MOM proportion during postnatal weeks 1–8 is associated with the growth of both males and females. This association persists in females beyond 8 postnatal weeks, but not in males. In addition, early total feeding is also associated with females' growth but not for males. This could be explained by the different responses of females and males to the nutritional composition of feeding (38). Personalizing essential nutrient intake may help reduce gender differences. Furthermore, sex-differentiated breastmilk components, such as oligosaccharides (HMOs) (39), mineral (40), cortisol (41), and protein profiles (42) amplify this distinction. This disparity was referred to as “male disadvantage” (43). We suspected that females may benefit more from early MOM, with this benefit continuing as they age. This promotes essential nutrient intake and accelerated growth. This consideration arises while recognizing the growth of males might be “delayed” due to reduced MOM intake in their later life, or perhaps they require different nutrients not provided by MOM. Additionally, other components of breast milk, such as 8- hydroxy-2′-deoxyquanosine (8-OHdG), histone deacetylase sirtuins (SIRT3) and serotonin (5-HT) have been associated with inflammation and oxidative stress induced by premature and/or cesarian delivery (44). However, the sex specific effects of these breast milk components on infant growth remain unknown (44). Further research is needed to explore the association between MOM components and infant growth and neurodevelopment.

The earliest indication of sex-differentiated post-birth growth is reflected in males having significantly higher birth weights than females (45). Our study aligns with this pattern; however, we also observed a decrease in body weight over time in male infants, which was consistent with the findings of other study (46). In our study, body weights in both males and females exhibited a significant post-birth increase, although the growth was relatively slower, as suggested by the trajectories of their weight z-scores, possibly due to their non-term equivalent age (47). It is noteworthy that post 9 weeks after birth, females' growth (z-score) was faster than males. Interestingly, females had a higher intake of mother's breastmilk before week 9, and a higher intake of formula post week 9 in comparison to males, potentially contributing to their accelerated catch-up in body weight post week 9 compared to males. Our finding aligns with other studies indicating that formula intake is associated with fast increased weight gain (48). Although six-month exclusive breastfeeding is the recommended gold standard for infants' nutrition (49), our study observed that preterm infants often encountered medical issues and/or lactation difficulties, leading to disrupted breast milk expression. Therefore, supplementing breastfeeding with alternatives like HDM and formula is recommended to ensure their well-being when MOM is limited (50).

Although preterm infants typically undergo catch-up growth after birth, this rapid weight gain could bridge the growth gaps between preterm and term infants (51); however, it does not necessarily eliminate the risk of neurodevelopment disorders (52–54). Our study observed that at 36 to 38 weeks of PMA, male infants demonstrated a higher level of fussing and crying during the assessment. In contrast, females showed a more mature neurobehavioral development if they received a higher amount of MOM before 8 postnatal weeks. This may be explained by the fact that earlier and higher MOM exposure boosts infants' brain structure (55) and connectivity (56). Further, whole brain volume increases in direct proportion to macronutrient and protein intake (57), however, a gender-specific scaling law exists, such that the cerebral cortex in males grows at a relatively slower rate compared to overall brain volume (58). Additionally, compared to females, resting neurophysiological network communication is more significantly altered in very preterm males, which further contribute to their disrupted cognitive outcomes later in childhood (at 8 years old) (59). Our study identified that the positive association between earlier and higher proportion of MOM exposure and better language and cognitive persist until the age of 2 in females. These findings were supported by other studies indicating the positive impact of earlier and longer breastfeeding on language ability performance and cognitive development (18, 60–62, 63).

To the best of our knowledge, this is the first time using SHAP, one of the Machine Learning methods, to determine the importance of total feeding amount, the quantity of MOM, and the proportion of MOM within 8 weeks postnatal in predicting neurodevelopmental outcomes in both males and females from the NICU until their 2 years of age. Other studies found birth complications such as intubation, durations of ventilatory support, length of hospital stay, and whether the infant was receiving breastmilk can also be used as predictors to predict cognitive abilities at 2 years of age (64). Consistent with other studies that have highlighted factors such as birth weight, GA, SNAPPEII, etc. (65), our analysis also identified these as predictors of cognitive and language development. Males in our study had relatively older birth GA and higher birth weight, but experienced delayed growth compared with females, these findings underscore the importance of recognizing that males with older birth GA, yet experiencing delayed weight gain, may be at risk for delayed neurobehavioral outcomes. Notably, these sex-differentiated neurodevelopmental outcomes should be interpreted with caution, as comparable sex differences have also been reported in term infants (66). Our future study will include both preterm and term infants to investigate the impact of preterm birth risk on sex-related neurodevelopmental outcomes.

Various practical barriers and challenges impact breast milk utilization and infant care, including adverse maternal social determinants of health such as lack of family support, availability of caregiver, and knowledge of breastfeeding (67), single mother (68), younger maternal age and lower education level (69), and the inability of extremely preterm infants to breastfeed (69). To improve infant health outcomes, multiple strategies should be considered. First, breast milk intake should be recognized as a key parameter in NICU protocols for monitoring and assessing preterm infant outcomes. Additionally, promoting breast milk intake alongside early intervention programs, such as developmental therapies for preterm infants, may further enhance neurological outcomes (70, 71).

Large sample size with equally distributed sex and race subsamples, and subjects follow up from NICU until 1 and 2 years of CA providing a comprehensive understanding of infants' health outcomes. In addition, the SHAP analysis validated the results obtained from linear mixed models. This approach provided further insights into how each predictor influences the developmental outcomes in infants. However, the study has limitations. The composition such as neurotrophic factors, lipids, and proteins, or calories of MOM, HDM, formula, and fortification were not evaluated, which could offer insights into the biomechanisms underlying the clinical manifestations and sex-specific micronutrients and macronutrients. The duration of breast milk exposure is another factor we will consider in future studies to better understand its impact on infants' growth and neurodevelopment. Additionally, the study did not incorporate maternal social determinants of health, which could potentially broaden our understanding of the maternal factors influencing sex-specific feeding regimens in preterm infants.

Sex-specific MOM intake, but not HDM and formula, was associated with infants' growth and neurodevelopmental outcomes. In addition, sex-specific “window of opportunity” for feeding, growth, and development in early life and sex-specific risk predictors for neurodevelopment from the NICU to 2 years of CA can inform the development of tailored, sex-specific feeding plans. Our study further emphasizes that early initiation of breastfeeding and exclusive breastfeeding beyond 8 weeks after birth are critical for infants' growth and neurodevelopment. To meet their nutritional requirements for growth, infants—especially males —should receive appropriate supplements. Individualized assessment and feeding regimens should be provided to preterm infants during their NICU stay.