Paternal folate deficiency induces hepatic insulin resistance in offspring mice

Background Folate (vitamin B9) is a water-soluble vitamin necessary for one-carbon metabolism, supporting the synthesis, repair, and methylation of DNA. While maternal folate status is well-studied for its role in fetal development and metabolic programming, the impact of inadequate folate intake in males on offspring development and metabolic diseases remains poorly understood. This study investigates the effects of folate deficiency in male parents on developing hepatic insulin resistance in offspring, focusing on molecular and metabolic disruptions within the liver. Methods Three-week-old C57BL/6 male mice were categorized into two groups: Group I received a folate-normal (FN) diet, and Group II was fed a folate-deficient (FD) diet for four weeks before mating. F1 offspring from Group I (FN diet) were mated to produce F2 offspring (PNMN: paternal normal, maternal normal). F1 males from Group II (lifetime FD diet) were mated with F1 females on an FN diet to produce F2 offspring (PDMN: paternal deficient, maternal normal). F2 offspring from both groups were maintained on an FN diet and monitored for body weight. The study assessed systemic markers of insulin resistance, lipid and glucose metabolism, and gene expression profiles and proteins associated with insulin signaling in the liver. Mechanistic pathways involving lipid-induced and ER stress-triggered hepatic insulin resistance were explored. Results Male offspring born to folate-deficient fathers (PDMN) exhibited significantly elevated fasting glucose and insulin levels, impaired glucose tolerance, and increased insulin resistance indices (HOMA-IR, QUICKI) at 10 weeks. Hepatic insulin signaling was disrupted, as evidenced by downregulated p-AKT levels in 7-week PDMN males. Lipogenic pathways were upregulated, with increased expression of transcription factors Srebf1c and Chrebp (both at gene and protein levels), contributing to hepatic steatosis. Gluconeogenic genes, including Foxo1 and Fbp1 , were also upregulated, indicating elevated hepatic glucose output and exacerbation of hyperglycemia. Chronic endoplasmic reticulum (ER) stress, marked by upregulation of Perk and Atf6 (both at gene and protein levels), further impaired hepatic insulin signaling possibly by activating stress pathways and disrupting protein folding. Conclusion This study provides the first evidence that paternal folate deficiency predisposes offspring to hepatic insulin resistance by disrupting insulin signaling, promoting lipid dysregulation, and activating ER stress pathways. These effects are more severe in males, underscoring sex-specific susceptibility. The findings emphasize the importance of balanced paternal folate intake during reproduction to prevent intergenerational metabolic disorders and suggest potential therapeutic targets to mitigate hepatic insulin resistance caused by paternal nutritional deficiencies.