Obesity is an independent risk factor for cardiac failure. Obesity promotes excessive deposition of fat in adipose and non adipose tissues. Intramyocardial lipid overload is a relatively common finding in non-ischemic heart failure, especially in obese and diabetic patients, and promotes lipoapoptosis that contributes to the alteration of cardiac function. Lipoprotein production has been proposed as a heart-protective mechanism through the unloading of surplus cellular lipids. We previously analyzed the heart transcriptome in a dog nutritional model of obesity and we identified a new apolipoprotein, regulated by obesity in heart, which is the subject of this study. We detected this new protein in lipoproteins HDL, LDL and VLDL. We designated it Apolipoprotein O. Apolipoprotein O is a 198 amino acids protein that contains a 23 amino acids long signal peptide. The apolipoprotein O gene is expressed in a set of human tissues. Confocal immunofluorescence microscopy colocalized apolipoprotein O and perilipins, a cellular marker of the lipid droplet. Chondroitinase ABC deglycosylation analysis or cell incubation with p-nitrophenyl-B-D-xyloside indicated that apolipoprotein O belongs to the proteoglycan family. Naringenin or CP-346086 treatments indicated that apolipoprotein O secretion requires microsomal triglyceride transfer protein activity. Apolipoprotein O gene expression is up-regulated in the human diabetic heart. Apolipoprotein O promoted cholesterol efflux from macrophage cells. To our knowledge, apolipoprotein O is the first chondroitine sulfate chain containing apolipoprotein. Apolipoprotein O may be involved in myocardium protective mechanisms against lipid accumulation or it may have specific properties mediated by its unique glycosylation pattern.