MON-623 Insulin Influences Human Lactation and Promotes Mammary Mitochondrial Biogenesis

Abstract Disclosure: F. Hannan: None. X. Meng: None. B. Davies: None. T. Elajnaf: None.Insulin regulates mammary development during puberty and pregnancy. However, its role in lactation is unclear. We hypothesised that insulin enhances mammary cell metabolism to support milk synthesis and that insulin resistant mothers have delayed lactation onset after childbirth. To investigate this, we performed clinical and mammary cell transcriptomic analyses following informed consent in n=63 breastfeeding mothers, of whom n=35 had insulin resistant conditions such as obesity, gestational diabetes or polycystic ovary syndrome. Lactation onset, as assessed by participant reporting of milk coming in after childbirth, was significantly delayed in affected mothers compared to n=28 healthy breastfeeding mothers (2.9±0.1 vs. 2.6±0.1 days postpartum, p<0.05). RNA sequencing of mammary cell RNA isolated from milk samples obtained on postpartum days 3 or 4 from n=15 insulin resistant mothers and n=15 healthy breastfeeding mothers revealed mitochondrial oxidative phosphorylation (OX-PHOS) as the major affected process with downregulation (>1.4-fold decrease, p<0.05) of genes encoding mitochondrial electron transport chain (ETC) complex I (ND1, ND3, ND4, ND5, ND6), complex IV (COX1, COX2) and ATP synthase (ATP6, ATP8). We further evaluated the role of insulin on mammary mitochondrial biology using the milk protein-expressing HC11 mammary cell line. Live imaging of HC11 cells incubated with a mitochondrial fluorophore (PKmito) and stimulated with 100nM insulin showed an increase in total mitochondrial volume (2.7±0.2µm3) vs. control cells (2.0±0.1µm3, p<0.01), suggestive of enhanced mitochondrial biogenesis. Consistent with this, RNA sequencing of insulin-treated HC11 cells showed increased expression (>1.4-fold, p<0.05) of 235 mitochondrial genes encoding ETC complexes and components involved in mitochondrial protein import and cristae synthesis. We next assessed whether insulin-mediated mitochondrial biogenesis led to increased OX-PHOS using an intracellular O2-sensitive fluorophore (MitoXpress Intra). This showed that 100nM insulin caused a 2.5-fold increase in both the basal and maximal O2 consumption rates in HC11 cells (p<0.01), consistent with enhanced OX-PHOS. In summary, this study demonstrates that insulin promotes mammary mitochondrial biogenesis and OX-PHOS, which may be pivotal for initiating milk synthesis after childbirth. Our findings indicate that such effects may be impaired in insulin resistant breastfeeding mothers and delay lactation onset, which is a major cause of low milk supply. Presentation: Monday, July 14, 2025