Figure 9. Maternal high-fat diet (HFD) feeding impairs neural development in SIRT1 deficient embryos.

(A) Maternal HFD feeding reduces body weight of embryos. Maternal HFD feeding was performed 4-8 weeks before pregnancy (pre-feeding) as described in Materials and methods. Body weight of E14.5 and E18.5 embryos were measured (*p<0.05, **p<0.01, ***p<0.001). (B) SIRT1 KO embryos have reduced expression of Smpdl3b in brains. The mRNA levels of Smpdl3b in brain of E18.5 embryos from chow fed dams or HFD fed dams were analyzed by qPCR (n = 6 embryos, *p<0.05, **p<0.01). (C) Maternal HFD feeding induces sphingomyelin accumulation in brains of SIRT1 KO embryos. Maternal HFD feeding was performed 4-8 weeks before pregnancy (pre-feeding) as described in Materials and methods. Brains from E18.5 embryos were dissected into three parts and the endogenous sphingomyelins were extracted and measured (n = 6 embryos, *p<0.05). (D) Maternal HFD feeding induces defective expression of neural markers in brains of SIRT1 KO embryos. The mRNA levels of indicated neural markers in brain of E18.5 embryos from chow fed dams or HFD fed dams were analyzed by qPCR (n = 6 embryos, *p<0.05, **p<0.01, ***p<0.001). (E) SIRT1 regulates sphingomyelin degradation and neural differentiation of mESCs through c-Myc and EZH2. SIRT1 is highly expressed in mESCs cells, where it functions to promote association of c-Myc and recruitment of Pol II to activate transcription of Smpdl3b gene and subsequent sphingomyelin degradation. This action of SIRT1 is important for maintenance of a proper membrane fluidity for normal neural differentiation in response to nutritional/developmental cues. Deletion of SIRT1 causes hyperacetylation and instability of c-Myc, leading to Pol II depletion and transcriptional repression of Smpdl3b. SIRT1 deficiency-induced hyperacetylation and stabilization of EZH2 likely enforce this transcriptional suppression by adding H3K27me3 mark. This transcriptional repression of Smpdl3b is associated with accumulation of sphingomyelin, which increases membrane fluidity and impairs neural differentiation. Light blue squares: H3K4me3; Light green circles: H3K27me3; Ac: acetylation.