Cold exposure during early pregnancy alters bile acid and microbial metabolism in breast milk, conferring durable protection in male rat offspring against impaired glycemic control and hepatic fat accumulation.
Maternal cold exposure reprograms offspring metabolism through the microbiota-bile acid-Th17 axis. Study: Maternal cold exposure improves offspring metabolic health via the milk lithocholic acid–microbiota–Th17 axis
In a recent study published in the journal npj biofilm and microbiomeresearchers evaluated whether maternal cold exposure during early pregnancy affects long-term metabolic outcomes in offspring.
This study combined immunophenotyping, transcriptomic analysis, metabolomic analysis, and a cross-breeding approach and revealed that male offspring of cold-exposed dams had significantly improved glucose tolerance, insulin sensitivity, and hepatic lipid profile when fed a Western diet. In a separate cohort fed a standard diet, these metabolic benefits were still detectable at 18 months of age.
Importantly, the study found that these maternal benefits persist into late adulthood in male rats fed a standard diet. Mechanism evaluation identified lithocholic acid (LCA)-rich breast milk as a potential mediator and showed that its benefits are transmitted, at least in part, through postnatal factors associated with breastfeeding. These in vivo model findings were complemented by observational analyzes in humans showing that winter conception, used as a surrogate for early pregnancy cold exposure, is associated with a reduced risk of metabolic dysfunction-associated fatty liver disease (MASLD).
background
The modern shift to Western-style diets has been highlighted as a major cause of the increasing global burden of metabolic diseases such as obesity. Remarkably, symptoms of type 2 diabetes (T2D) and MASLD, previously associated with adulthood, are increasingly being reported in childhood and early life, suggesting intergenerational changes in metabolic risk.
The Developmental Origins of Health and Disease (DOHaD) framework aims to explain these observations by positing that environmental exposures during the prenatal and perinatal periods can have lasting effects on an individual’s physiology. Negative factors such as pollutants and inadequate diet have been shown to predispose offspring to adult diseases, whereas beneficial maternal interventions such as exercise and dietary changes can improve metabolic resilience in offspring.
Separately, studies in adults have shown that cold exposure activates thermogenic and lipid metabolic pathways. However, whether maternal cold exposure can program sustained metabolic health in offspring remained unclear.
About research
The present study aimed to address this knowledge gap by experimentally exposing pregnant rats to room temperature (25 ± 1 °C) or cold conditions (4 ± 1 °C) during the first 10 days of pregnancy. Their offspring were kept at room temperature until the end of weaning (21 days), after which dietary treatment was initiated.
The intervention consisted of feeding pups in the experimental cohort a high-fat Western diet containing 60% of energy from fat and 10% of weight/volume (w/v) fructose in their drinking water for 5 weeks. Another cohort continued to receive a standard diet until 18 months of age to assess outcomes in late adulthood.
A cross-species rearing experiment was conducted to separate prenatal and postnatal effects of maternal exposure. Within 24 hours of birth, a subset of newborn pups were swapped between mothers exposed to cold and mothers at room temperature.
Long-term metabolic changes and their potential physiological mechanisms were evaluated using next-generation molecular assays, specifically liquid chromatography tandem mass spectrometry (LC-MS/MS) for metabolomic profiling of breast milk and offspring plasma.
In addition, progeny immune cells were assessed by flow cytometry (immunophenotyping), and ribonucleic acid (RNA) sequencing and transcriptome profiling by quantitative real-time polymerase chain reaction (qRT-PCR) were used to elucidate concurrent liver tissue changes.
The researchers then examined whether the animal findings were reflected in human observational data from 33,915 UK Biobank participants and 3,209 China Health and Retirement Longitudinal Study (CHARLS) participants.
Research results
The study’s statistical analysis revealed that male offspring born to mothers exposed to cold environments showed a significant metabolic advantage compared to male offspring born to mothers reared at ambient temperatures. Oral glucose tolerance and insulin tolerance tests showed that the former cohort exhibited significantly better glucose tolerance and insulin sensitivity when fed a Western diet (p < 0.05).
At the same time, organ weight measurements, liver triglyceride assays, and histological analyzes revealed statistically significant reductions in liver weight, liver fat accumulation, and liver triglyceride levels in pups born and raised by mothers exposed to cold environments. No comparable liver differences were observed in female offspring, so subsequent mechanistic experiments focused on males.
Remarkably, these metabolic benefits persisted for 18 months into late adulthood. Members of a separate standard-fed cohort continued to show better glucose tolerance and insulin sensitivity, reduced hepatic lipid content, and lower expression of lipid-related genes CD36, FABP1, and FAS (p < 0.05).
Cross-fostering experiments showed that these metabolic benefits were mediated, at least in part, by postnatal factors associated with the adoptive mother, supporting a role for lactation and breast milk components. Through metabolomics, we confirmed that LCA, a secondary bile acid, is highly abundant in the milk of cold-exposed dams.
LCA itself did not directly reverse palmitic acid-induced transcriptional changes in the HepG2 hepatoma cell line or the Jurkat T-cell leukemia cell line. However, antibiotic treatment abolished the metabolic effects of LCA supplementation in rats, supporting the need for microbial conversion of LCA to active metabolites.
Targeted bile acid profiling identified higher plasma levels of the LCA derivatives 3-oxo-LCA and isoallo-LCA in male offspring of cold-exposed dams. A bioinformatics screen of 1,520 representative human gut bacterial genomes identified Clostridium sindens, Ruminococcus gnabus, and Eggertella lenta as prominent carriers of genes encoding related 3α-hydroxysteroid dehydrogenase enzymes.
Supplementation with 3-oxo-LCA reduced hepatic interleukin-17 (IL-17) pathway gene expression and improved glucose tolerance, insulin sensitivity, and hepatic lipid outcomes. The effect was not abolished by antibiotics. This indicates that 3-oxo-LCA acts after the microbiota-dependent LCA conversion step.
The researchers then tested Clostridium scindens experimentally in rats. Supplementation with this bacterium increased fecal 3-oxo-LCA and recapitulated some of these protective effects.
In an observational human analysis, winter pregnancy was associated with a 24.9% lower risk of developing MASLD in the UK Biobank cohort, with a hazard ratio (HR) of 0.751 and a 95% confidence interval (CI) of 0.568 to 0.991. In the CHARLS trial, winter conception was associated with lower odds of MASLD in cold northern China (OR = 0.580; 95% CI = 0.338-0.995), but no association was observed in southern China.
conclusion
These results support a maternal cold milk bile acid-microbiota-T helper 17 (Th17) axis that may influence the long-term metabolic health of male rat offspring. The human observations, which are broadly consistent with the animal findings, indicate that the microbiota and bile acid-related pathways are potential targets for further investigation rather than establishing transgenerational therapeutic strategies.
Human analyzes were associative and did not directly measure maternal cold exposure, LCA, 3-oxo-LCA, gut microbial transformation, or Th17 activity. Winter pregnancy was estimated from month of birth, whereas MASLD in CHARLS was determined using metabolic proxies rather than liver imaging. Also, in heterogeneous feeding, milk-derived LCA cannot be completely separated from other maternal, milk-related, or postnatal factors. Absorption kinetics of supplemented LCA and 3-oxo-LCA were also not determined.
Therefore, further research is needed to establish whether the proposed mechanisms work in humans and whether related interventions are safe, effective, and durable.
