This is one of the first published RCT studies investigating the potential effect of different post-discharge nutritional regimens on BMD in 6-year-old children born very preterm. We did not find a difference in bone variables comparing the two human milk-fed groups, but we found a significant difference when comparing human milk feeding with formula feeding. In our previous studies, we found early rapid growth (weight growth > 1 SD from 34 weeks to 2 months CA) to have a potential influence on metabolic outcomes at six years of age [9, 10]. Early rapid growth did not seem to influence bone variables. A similar study, with a cohort of very preterm infants born from 2003 to 2006, did a follow up at 8 years of age. In this study, infants were randomised to a preterm or a term formula post-discharge, and the third group was a human milk-fed group. At six years of age, researchers did not find any difference in metabolic outcomes or bone variables [18].

Several studies have investigated the impact of feeding during the hospitalization period on BMC in preterm infants. Recently Einloft et al. performed a study in which 83 preterm infants with a birth weight of less than 1500 g were randomized to receive HM with or without supplementation with FM85®. DXA was performed at study entry and when the infant reached 2000 g, shortly before discharge. Whole body BMC was established in 38 infants, and was found to be significantly higher at the end of the study in infants fed fortified HM. Also, the level of alkaline phosphatase (ALP) was significantly higher among infants fed HM without supplementation [19]. Higher BMC and lover ALP in the fortified group indicates a need for extra protein. ALP’s applicability in distinguishing the presence of metabolic bone disease in preterm infants is questionable [20]. It is reasonable to assume that the lower BMC and higher ALP in the HM group is caused by a higher bone turnover aiming to maintain the serum values of minerals within the normal range due to insufficient enteral supply. Unfortunately, measurements of ALP were not performed in our study, thereby limiting the evaluation of possible differences in osteoblastic activity according to different post-discharge diets.

In a randomized double blinded study by Koo et al., 89 preterm infants with a GA of 24–34 weeks were randomized to receive nutrient-enriched formula with a higher energy density, higher protein, calcium, and phosphate content compared to a standard formula for term infants. The formulas were given for one year after hospital discharge, and 67 infants completed the study. At 2-, 4-, 6-, and 12-months post-discharge, DXA scans were obtained. Infants fed term formula surprisingly turned out to have higher weight and length Z-scores compared to infants fed the enriched formula. This increase in body size was also reflected in a higher total body BMC in the children fed term formula. The authors did not measure area BMD or volumetric BMD, and the recommended setting TBLH was not used, most likely confounding the results [21]. Figueras-Aloy et al. reported a higher BMD in preterm infants when fed fortified human milk compared with a preterm formula. The study was a prospective study including 336 preterm infants with a GA ≤ 31 weeks and birth weight ≤ 1500 g. DXA was performed before discharge with analysis of the BMD value of the lumbar spine L1-L4. Meanwhile, precaution must be applied in the interpretation of the BMD of the lumbar spine in only a few months old preterm infants, as the vertebrae are largely cartilaginous at birth, which may indeed challenge the edge detection of the vertebrae [22].

A takeaway from these studies is that we don’t need to enrich the formula but continue to include HM as a main component in nutrition for very preterm infants. Further, it is important to choose the most frequently reported data (BMD/BMC) and thereby be able to compare studies. Often BMD is used, as this data gives the bone mineral content per square root of the bone, making it a more precise measure of the bone, regardless of the height of the person having a DXA scan.

Bishop et al. published data on a subgroup of 54 preterm children randomized to banked HM without fortification versus PF as a supplement to maternal HM. By single photon absorptiometry (SPA), and when adjusting for body size, the authors found a strong positive correlation between BMC at the age of five years and a high percentage of HM (maternal milk and banked HM) compared to HM and supplementation with formula [23]. This supports our finding of a higher BMD at six years of age in the group fed HM versus formula feeding, as our results also suggest that HM is important for bone growth.

Fewtrell et al. performed a follow-up study of the same preterm-born children with DXA scans at ages 8–12 years, and this revealed no differences in bone density related to different diets during hospitalization in infancy [24]. In a later study in 20-year-old adults from a preterm cohort by Fewtrell et al., DXA scans showed a higher whole-body BMC associated with HM feeding compared to formula feeding in preterm-born infants during the first month of life [25]. However, this effect disappeared when correcting for whole body bone area, emphasizing the importance of appropriate size adjustments when interpreting DXA results, with children as well adults. As the uncorrected BMC was higher in the group fed HM, the authors conclude that feeding HM during the first months of preterm infants’ lives may contribute to larger bones with a proportional increase in mineral mass at age 20 years [25]. Fewtrell et al. concluded that the increased whole-body bone mass may reflect non-nutritive factors in the mother’s own “low nutrient milk” compared to a very high nutrient content when formula fed [26]. In our study, there was no difference in BMD at six years of age when comparing children fed low nutrient diet versus high nutrient diet (group UHM vs. FHM + PF). We therefore agree that other substances than those added by supplementation to breast milk might be responsible for the achievement of a higher bone density at age six years.

Interestingly, a study of formula fed versus breastfed term born children showed no difference in bone mass by DXA, when examined at the age of 10 years [27]. We have not found other studies showing an effect on bone mineralization among term-born infants fed different diets, but our data indicates a favourable effect of breast versus formula feeding with respect to BMD in children born very preterm. Preterm-born infants are at risk of poor bone mineralization in early life due to mineral deficit at birth and insufficient nutrition while treated at the NICU. However, nutritional intake has been improved in many NICU’s according to recent recommendations from e.g. ESPGHAN on both parenteral and enteral intakes including calcium and vitamin D [6]. In our RCT, all infants received HM with fortification or PF and vitamin D and phosphate during hospitalization as recommended. The results of the present follow-up study primarily illustrate the influence of different post-discharge nutrition until four months, and not solely the nutrition during hospitalization.

Lucas et al. randomized 229 premature infants (GA < 37 weeks with birth weight < 1750 g) to an enriched post-discharge formula or a standard term formula from discharge until nine months corrected age. A reference group of 65 preterm infants were breastfed for at least six weeks post-term. The authors found breastfed infants to be shorter and lighter, but raised the question in 2001 of whether infants breastfed post-discharge need nutritional supplementation [28]. We did not find any difference in anthropometric data at four months of CA [8] or at six years of age associated with the fortification of human milk post-discharge [11]. The evidence in favour of advising fortification of breast milk has been addressed in a recent Cochrane review, concluding that the data available is limited and does not provide strong evidence that supplementing preterm infants with fortification is superior to HM, but provides low-quality evidence of increased growth rates (weight, length, and head circumference) before hospital discharge. Very limited data was available for outcomes beyond infancy [29], and the results seem to be in accordance with our results, showing no effect on growth or bones at six years when infants were fed fortified HM versus HM post-discharge.

Strengths and limitations: Our study was conducted in a prospective design, randomized to supplementation with or without fortification when breastfed post-discharge. It is known that severe premature birth-related co-morbidities and medications could possibly influence bone health later in life in very preterm infants [30]. Medication including details on PN and diseases known to affect bone mineralization led to exclusion from the study. The dose of added fortifier to mother’s milk was relatively low and was not increased during the trial period. The measured values of blood urea nitrogen and growth were not significantly different between the fortified and unfortified groups [8], which also justifies combining the human milk-fed groups for comparison with preterm formula. The differences in level of physical activity, calcium intake and/or vitamin D status during infancy and early childhood may have influenced the BMD at age six years, but this potential effect is expected to be of equal impact in all three groups.

Future studies on the effects of early nutrition on BMD in preterm-born children need to be reported in follow-up studies, since data is still sparse.