Vitamin D Receptor Activation in Liver Macrophages Ameliorates Hepatic Inflammation, Steatosis, and Insulin Resistance in Mice.

BACKGROUND AND AIMS: Obesity-induced chronic inflammation is a key component in the pathogenesis of nonalcoholic fatty liver disease (NAFLD) and insulin resistance. Increased secretion of proinflammatory cytokines by macrophages in metabolic tissues promotes disease progression. In the diet-induced obesity (DIO) mouse model, activation of liver resident macrophages, or Kupffer cells (KCs), drives inflammatory responses, which recruits circulating macrophages and promotes fatty liver development, and ultimately contributes to impaired hepatic insulin sensitivity. Hepatic macrophages express the highest level of vitamin D receptors (VDRs) among nonparenchymal cells, whereas VDR expression is very low in hepatocytes. VDR activation exerts anti-inflammatory effects in immune cells. APPROACH AND RESULTS: Here we found that VDR activation exhibits strong anti-inflammatory effects in mouse hepatic macrophages, including those isolated from DIO livers, and mice with genetic loss of Vdr developed spontaneous hepatic inflammation at 6 months of age. Under the chronic inflammation conditions of the DIO model, VDR activation by the vitamin D analog calcipotriol reduced liver inflammation and hepatic steatosis, significantly improving insulin sensitivity. The hyperinsulinemic euglycemic clamp revealed that VDR activation greatly increased the glucose infusion rate, while hepatic glucose production was remarkably decreased. Glucose uptake in muscle and adipose did not show similar effects, suggesting that improved hepatic insulin sensitivity is the primary contributor to the beneficial effects of VDR activation. Finally, specifically ablating liver macrophages by treatment with clodronate liposomes largely abolished the beneficial metabolic effects of calcipotriol, confirming that VDR activation in liver macrophages is required for the antidiabetic effect. CONCLUSIONS: Activation of liver macrophage VDRs by vitamin D ligands ameliorates liver inflammation, steatosis and insulin resistance. Our results suggest therapeutic paradigms for treatment of NAFLD and type 2 diabetes mellitus.

A group 3 medulloblastoma stem cell program is maintained by OTX2-mediated alternative splicing.

OTX2 is a transcription factor and known driver in medulloblastoma (MB), where it is amplified in a subset of tumours and overexpressed in most cases of group 3 and group 4 MB. Here we demonstrate a noncanonical role for OTX2 in group 3 MB alternative splicing. OTX2 associates with the large assembly of splicing regulators complex through protein-protein interactions and regulates a stem cell splicing program. OTX2 can directly or indirectly bind RNA and this may be partially independent of its DNA regulatory functions. OTX2 controls a pro-tumorigenic splicing program that is mirrored in human cerebellar rhombic lip origins. Among the OTX2-regulated differentially spliced genes, PPHLN1 is expressed in the most primitive rhombic lip stem cells, and targeting PPHLN1 splicing reduces tumour growth and enhances survival in vivo. These findings identify OTX2-mediated alternative splicing as a major determinant of cell fate decisions that drive group 3 MB progression.

Defining the mammalian coactivation of hepatic 12-h clock and lipid metabolism.

The 12-h clock coordinates lipid homeostasis, energy metabolism, and stress rhythms via the transcriptional regulator XBP1. However, the biochemical and physiological bases for integrated control of the 12-h clock and diverse metabolic pathways remain unclear. Here, we show that steroid receptor coactivator SRC-3 coactivates XBP1 transcription and regulates hepatic 12-h cistrome and gene rhythmicity. Mice lacking SRC-3 show abnormal 12-h rhythms in hepatic transcription, metabolic functions, systemic energetics, and rate-limiting lipid metabolic processes, including triglyceride, phospholipid, and cardiolipin pathways. Notably, 12-h clock coactivation is not only preserved, with its cistromic activation priming ahead of the zeitgeber cue of light, but concomitant with rhythmic remodeling in the absence of food. These findings reveal that SRC-3 integrates the mammalian 12-h clock, energy metabolism, and membrane and lipid homeostasis and demonstrates a role for the 12-h clock machinery as an active transcriptional mechanism in anticipating physiological and metabolic energy needs and stresses.

XBP1 links the 12-hour clock to NAFLD and regulation of membrane fluidity and lipid homeostasis.

A distinct 12-hour clock exists in addition to the 24-hour circadian clock to coordinate metabolic and stress rhythms. Here, we show that liver-specific ablation of X-box binding protein 1 (XBP1) disrupts the hepatic 12-hour clock and promotes spontaneous non-alcoholic fatty liver disease (NAFLD). We show that hepatic XBP1 predominantly regulates the 12-hour rhythmicity of gene transcription in the mouse liver and demonstrate that perturbation of the 12-hour clock, but not the core circadian clock, is associated with the onset and progression of this NAFLD phenotype. Mechanistically, we provide evidence that the spliced form of XBP1 (XBP1s) binds to the hepatic 12-hour cistrome to directly regulate the 12-hour clock, with a periodicity paralleling the harmonic activation of the 12-hour oscillatory transcription of many rate-limiting metabolic genes known to have perturbations in human metabolic disease. Functionally, we show that Xbp1 ablation significantly reduces cellular membrane fluidity and impairs lipid homeostasis via rate-limiting metabolic processes in fatty acid monounsaturated and phospholipid remodeling pathways. These findings reveal that genetic disruption of the hepatic 12-hour clock links to the onset and progression of NAFLD development via transcriptional regulator XBP1, and demonstrate a role for XBP1 and the 12-hour clock in the modulation of phospholipid composition and the maintenance of lipid homeostasis.

Murine neonatal intravascular injections: modeling newborn disease.

The ability to perform murine neonatal intravascular injections likely will prove useful in studying many newborn-specific disease states that are modeled in mice. Unfortunately, effective intravascular injection in the neonatal mouse has been limited by developmental immaturity and small size. To establish a mouse model of neonatal intravascular injection, C57Bl/6 pups between birth and 6 d of age were injected with a buffered solution containing cells or vehicle alone. For both external jugular and superficial temporal vein injections, a 2-member team was used to position the pup, insert the needle, and perfuse the injectate. For superficial temporal vein injections, the vascular anatomy was visualized by using transillumination. After injection into the jugular or superficial temporal vein, the survival rate to adulthood was 100% (n = 30 pups per group), with no long-term complications. Occasional extravasation of injectate was well tolerated, allowing for serial injections (n = 40 pups). Intravascular access was confirmed by using fluorescent dye perfusion studies and cellular engraftment analysis. The 2 techniques are safe and reproducible methods of obtaining intravascular access via the external jugular and superficial temporal veins in newborn mice. These methods provide a mechanism for delivering a wide variety of substances, ranging from aqueous solutions to suspensions.