Upregulated hepatic lipogenesis from dietary sugars in response to low palmitate feeding supplies brain palmitate

Palmitic acid (PAM), a saturated fatty acid, is crucial for brain development and function, contributing to myelination and cell signaling. PAM can be obtained from the diet or synthesized endogenously through de novo lipogenesis (DNL), primarily from glucose. Preclinical studies on PAM's origin in the developing brain are scarce and yield conflicting results compared to adult studies. Some studies suggest that the developing brain synthesizes PAM endogenously, while others indicate that dietary PAM can be incorporated into brain tissue. This study aimed to clarify the origin of brain PAM during development using a novel approach. The researchers leveraged naturally occurring carbon isotope ratios (¹³C/¹²C; δ¹³C) in dietary PAM (C3 plant origin) and dietary sugars (C4 plant origin) to trace the source of PAM in the brain. This method avoids the use of radioactive tracers and allows for the investigation of PAM origin in a more physiological setting. The study also used RNA sequencing to identify pathways involved in maintaining brain PAM levels.

Existing research on the origin of brain palmitic acid (PAM) during development is limited and shows inconsistencies with findings in adults. Studies using labeled PAM in artificially reared rat pups indicated that the brain synthesizes PAM entirely endogenously. In contrast, adult rat and mouse studies suggested that dietary PAM can enter brain tissue lipids. This discrepancy highlights the need for further investigation into the developmental trajectory of brain PAM origin. Previous work from the authors' group utilized compound-specific isotope analysis (CSIA) to study brain PAM origin in adult mice, revealing that a significant portion of the brain PAM pool is maintained by DNL from dietary sugars. This study aimed to extend this work to the developmental period, when PAM accretion is rapid.

The study used 32 four-week-old BALB/c dams, randomly assigned to isocaloric diets varying in palmitic acid (PAM) content: low (<2%), medium (~47%), and high (>95%). The diets were isocaloric, differing only in fatty acid composition. The dams were bred, and their offspring were maintained on the same diet until weaning at postnatal day (P) 21, then until P35. Male pups (and female pups when available) were euthanized at P0, P10, P21, and P35, and brain and liver tissue samples were collected. Total PAM levels and δ¹³C-PAM values were measured using gas chromatography-flame ionization detection (GC-FID) and gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS), respectively. RNA sequencing (RNA-Seq) was conducted on liver and brain tissue at P35 to identify differentially expressed genes and pathways. Maternal behavior (nest building) and pup sensorimotor development (geotaxis and righting reflex tests) were also assessed. Statistical analyses included two-way ANOVA, one-way ANOVA, and Kruskal-Wallis tests, depending on data distribution. Principal component analysis (PCA), differential gene expression analysis, gene set enrichment analysis (GSEA), and weighted gene co-expression network analysis (WGCNA) were used for RNA-Seq data analysis.

Despite significant dietary PAM differences, total brain PAM levels in male pups remained consistent across all time points. In contrast, liver PAM levels reflected dietary intake. Analysis of δ¹³C-PAM revealed that brain PAM was significantly enriched in ¹³C, closer to the isotopic signature of dietary sugars than dietary PAM, suggesting that DNL from dietary sugars is the primary source of brain PAM. This effect was more pronounced in mice fed the low-PAM diet. Liver δ¹³C-PAM also showed enrichment, indicating that DNL from dietary sugars maintains the liver PAM pool as well. RNA sequencing at P35 revealed that hepatic, but not brain, gene expression was significantly altered by diet. Specifically, genes involved in hepatic DNL were upregulated in mice fed the low-PAM diet compared to the high-PAM diet. Pathway analysis identified multiple pathways related to lipid metabolism that were differentially expressed between the high- and low-PAM groups. Network analysis confirmed the involvement of genes central to de novo lipogenesis in the liver. No significant effects of diet were observed on maternal behavior or pup sensorimotor development.

The findings demonstrate that hepatic DNL compensates for low dietary PAM intake, ensuring sufficient brain PAM supply during development. The enrichment of ¹³C in both brain and liver PAM strongly supports the role of DNL from dietary sugars. The lack of significant changes in brain gene expression suggests that the brain relies on a basal level of DNL, which is not significantly modulated by dietary PAM. The liver's metabolic flexibility in responding to dietary changes highlights its crucial role in maintaining brain fatty acid homeostasis. The results are consistent with previous studies showing that the liver, but not the brain, upregulates fatty acid synthesis in response to dietary changes. These findings have significant implications for understanding brain development and the interplay between diet and brain lipid metabolism.

This study demonstrates that hepatic de novo lipogenesis from dietary sugars plays a major role in supplying brain palmitic acid during development, particularly when dietary intake is low. The liver's metabolic flexibility in response to dietary PAM highlights the importance of hepatic lipid metabolism in maintaining brain fatty acid homeostasis. Future research should investigate the regulatory mechanisms controlling hepatic DNL during development and explore the contribution of other substrates to brain acetyl-CoA pools.

The study focused primarily on male pups, limiting the generalizability of findings to female pups. The RNA-Seq data were collected only at P35, preventing a complete understanding of the developmental trajectory of gene expression changes. While the study controlled for several factors, the inability to blind the investigator to diet assignment could introduce a potential bias. Finally, the study did not directly quantify the contribution of other molecules to brain acetyl-CoA pools in addition to dietary sugars, which may have influenced the observed δ¹³C values.