AIMTo investigate the roles of liver X receptors (LXR) in the lipid composition and gene expression regulation in the murine caput epididymidis. LXR are nuclear receptors for oxysterols, molecules derived from cholesterol metabolism that are present in mammals as two isoforms: LXRalpha, which is more specifically expressed in lipid-metabolising tissues, such as liver, adipose and steroidogenic tissues, and macrophages, whereas LXRbeta is ubiquitous. Their importance in reproductive physiology has been sustained by the fact that male mice in which the function of both LXR has been disrupted have fertility disturbances starting at the age of 5 months, leading to complete sterility by the age of 9 months. These defects are associated with epididymal epithelial degeneration in caput segments one and two, and with a sperm midpiece fragility, leading to the presence of isolated sperm heads and flagella when luminal contents are recovered from the cauda epididymidis.

METHODSThe lipid composition of the caput epididymidis of wild-type and LXR-deficient mice was assessed using oil red O staining on tissue cryosections and lipid extraction followed by high performance liquid chromatography or gas chromatography. Gene expression was checked by quantitative real time polymerase chain reaction.

RESULTSUsing LXR-deficient mice, we showed an alteration of the lipid composition of the caput epididymidis as well as a significantly decreased expression of the genes encoding SREBP1c, SCD1 and SCD2, involved in fatty acid metabolism.

CONCLUSIONAltogether, these results show that LXR are important regulators of epididymal function, and play a critical role in the lipid maturation processes occurring during sperm epididymal maturation.

Animals ; DNA Primers ; DNA-Binding Proteins ; deficiency ; genetics ; physiology ; Epididymis ; cytology ; physiology ; Epithelial Cells ; physiology ; Fatty Acids ; metabolism ; Homeostasis ; Lipids ; physiology ; Liver X Receptors ; Male ; Mice ; Mice, Knockout ; Orphan Nuclear Receptors ; Polymerase Chain Reaction ; Receptors, Cytoplasmic and Nuclear ; deficiency ; genetics ; physiology

Considerable controversy exists in determining the role of peroxisome proliferator-activated receptor-alpha(PPARalpha) on obesity. Previous reports demonstrated that PPARalpha is a critical modulator of lipid homeostasis, but the overt, obese phenotypic characterization in the strain of PPAR deficient (PPARalpha-/-) mice is influenced by other factors, including diet and genetics. Therefore, it is necessary to establish the phenotypic characterization of PPARalpha-/- mice prior to the obesity-related study. In this study, we observed phenotype of PPARalpha-/- mice on mixed genetic background (C57BL/6Nx129/Sv) fed a high fat diet for 16 weeks. PPARalpha-/- mice, regardless of sex, raised body growth rate significantly comparing with wild type and showed male-specific fatty change in the liver. They were shown to lack hepatic induction of PPARalpha target genes encoding enzymes for fatty acid beta-oxidation.
Adipose Tissue/metabolism ; Animals ; Body Weight ; Cholesterol/blood ; Crosses, Genetic ; Dietary Fats/*administration & dosage ; Female ; Histocytochemistry ; Liver/enzymology/metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; Obesity/genetics/*metabolism ; Phenotype ; RNA/chemistry/genetics ; Receptors, Cytoplasmic and Nuclear/*deficiency/genetics/metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Specific Pathogen-Free Organisms ; Transcription Factors/*deficiency/genetics/metabolism ; Triglycerides/blood

Use of adenoviruses as vehicle for gene therapy requires that target cells express appropriate receptors such as coxsakievirus and adenovirus receptor (CAR). We show here that CAR-deficiency in cancer cells, that limits adenoviral gene delivery, can be overcome by using adenovirus complexed with the liposome, Ad-PEGPE [1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(poly-ethylene glycol)-2000]. We first confirmed that CT-26 mouse colon cancer cells are deficient in CAR by RT-PCR, and then showed that CT-26 cells infected with Ad-GFP/PEGPE exhibited highly enhanced expression of green fluorescent protein (GFP), compared with those infected with Ad-GFP. GFP expression depends on the dose of liposome and adenovirus. Luciferase expression in livers treated with Ad-luc/PEGPE was about 1,000-fold less than those infected with Ad-luc. In a liver metastasis mouse tumor model developed by intrasplenic injection of CT-26 cells, luciferase expression following i.v. injection of Ad-luc/PEGPE was significantly higher in tumors than in adjacent non-neoplastic liver. Following systemic administration of Ad-GFP/PEGPE, GFP expression increased in tumors more than in adjacent liver while the reverse was true following administration of Ad-GFP. In the latter case, GFP expression was higher in liver than in tumors. This study demonstrates that systemic delivery of PEGPE-adenovirus complex is an effective tool of adenoviral delivery as it overcomes limitation due to CAR deficiency of target cells while reducing hepatic uptake and enhancing adenoviral gene expression in tumors.
*Adenoviridae/genetics/metabolism ; Animals ; Colonic Neoplasms/*genetics/metabolism/pathology/*therapy ; Dose-Response Relationship, Drug ; Gene Therapy ; *Gene Transfer Techniques ; Genetic Vectors ; Green Fluorescent Proteins/genetics ; Liposomes/administration & dosage/chemistry/pharmacokinetics/*therapeutic use ; Liver/drug effects/metabolism ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Transgenic ; NIH 3T3 Cells ; Phosphatidylethanolamines/administration & dosage ; Polyethylene Glycols/administration & dosage ; Receptors, Cytoplasmic and Nuclear/deficiency/genetics ; Receptors, Virus/deficiency/*genetics ; Transcription Factors/deficiency/genetics ; Tumor Cells, Cultured