ORIGINAL ARTICLE Leptin-deficient (ob/ob) mouse ovaries show fatty degeneration, enhanced apoptosis and decreased expression of steroidogenic acute regulatory enzyme H Serke 1,3 , M Nowicki 1,3 , J Kosacka 2 , T Schro ¨ der 1 , N Klo ¨ ting 2 , M Blu ¨ her 2 , S Kallendrusch 1 and K Spanel-Borowski 1 OBJECTIVE: Leptin-deficient (ob/ob) mice are obese and infertile. Dysfunctions of the ovaries are preferentially related to leptin-deficiency. DESIGN: Morphological and molecular biological obesity-dependent changes in ob/ob ovaries. SUBJECTS: Ovaries were obtained from three-month-old mice either homozygote (ob/ob) and heterozygote (ob/ þ ) or wild-type (C57BL6, WT) for the investigation by light and electron microscopy, as well as for western blot analysis of lectin-like oxidised low density lipoprotein receptor (LOX-1), Toll-like receptor 4 (TLR4), CD36, cleaved caspase-3, microtubule-associated protein light chain 3 (LC3), and the steroidogenic acute regulatory protein (StAR). RESULTS: Compared with control ovaries with corpora lutea, ob/ob ovaries lacked corpora lutea, follicular atresia was at a higher rate; lipid droplets accumulated in follicle cells and in the oocyte with damaged mitochondria; the basement membrane of follicles was thickened. LOX-1 and CD36 expressions were comparable for all three groups. Ob/ob ovaries showed significantly higher levels of TLR4 and cleaved caspase-3 than the ones from the control groups. The high LC3-II/I ratio in the WT and ob/ þ ovaries was related to the presence of corpora lutea. The StAR protein was lower in the ob/ob ovaries signifying reduced steroidogenesis. CONCLUSIONS: Excessive lipid storage causes disorders of ovarian function in ob/ob mice. The local lipid overload leads to advanced follicular atresia with apoptosis and defect steroidogenesis. We suggest that the changes in lipid metabolism lead to increased oxidative stress and thereby, they are an important reason of anovulation and infertility. International Journal of Obesity advance online publication, 15 November 2011; doi:10.1038/ijo.2011.220 Keywords: ob/ob; obese mice; oxidative stress; LOX-1; Toll-like receptor 4; infertility INTRODUCTION Mice with deficiency of the leptin gene Lep ob /Lep ob (ob/ob) are obese and infertile. 1,2 They are generally accepted animal model for obesity and type 2 diabetes. Deficiency in leptin signalling in the hypothalamus leads to increased adiposity, severe insulin resistance and diminished fertility. Interestingly, fertility and insulin sensitivity were restored with leptin application, but not by calorie restriction or weight loss. 3 Leptin mediates appetite and food intake as well as changes in gonadotropin releasing hormone secretion. Ob/ob mice of young reproductive age suffer from lack in gonadotropins associated with diminishing fertility. Leptin appears to affect the ovary directly by modulating preovulatory events like steroidogenesis, meiotic maturation and follicle rupture. 4-6 Leptin could act in an auto/paracrine manner, because the adipokine is also produced in follicle cells and cells of the corpus luteum, which express the long and short form of the leptin receptor. 7 Absence of leptin should be accompanied by low levels of steroidogenic enzymes in the ob/ob ovary, as deduced from decreased sex steroid serum levels. 8 The shortage in intraovarian estrogens might lead to increased follicular atresia through activation of the Fas/Fas ligand system apparently active in ob/ob mouse follicles. 9 In ob/ob mice, obesity depends on hyperphagy, decreased metabolism and energy expenditure. Highly augmented serum levels of triglycerides, free fatty acids, oxidised lipoproteins, insulin and glucose are the pathophysiological consequences in obese persons. 10 In ob/ob mice, increase of these serum factors is associated with mild diabetes in the ob/ob mouse 11,12 and with excessively accumulated lipid droplets in cells of follicles and of the ovarian cortex. 8,13 The striking intraovarian lipogenesis is probably responsible for mitochondrial dysfunction with the augmented release of reactive oxygen species (ROS) leading to oxidative stress and cellular damage. The ROS production could be involved in the increased rate of follicular atresia and recognised by indirect parameters responding to oxidative stress. Scavenger receptors such as lectin-like oxidised low density lipoprotein receptor (LOX-1), the Toll-like receptor 4 (TLR4) and CD36 are specific for oxidised lipoproteins. 14 - 17 They are expressed on endothelial cells, leukocyte subtypes and on granulosa cells from human preovulatory follicles. 15,18 The oxidised lipoproteins- dependent activation of these receptors either triggers apoptosis or survival autophagy in endothelial or granulosa cells. 14,18,19 Ob/ob ovaries could respond to lipid droplet accumulation by upregulation of LOX-1 and/or TLR4 resulting in the cell damage. If so, follicular atresia is not because of leptin-deficiency only. 9,20 The present objectives were to study whether follicular atresia is associated with changes in LOX-1, TLR4 and CD36 expression in ovaries of ob/ob mice compared with wild-type (WT) or heterozygote (ob/ þ ) mice. The expression of cleaved caspase-3, Received 25 March 2011; revised 5 October 2011; accepted 10 October 2011 1 Institute of Anatomy, University of Leipzig, Leipzig, Germany and 2 Department of Internal Medicine III, University of Leipzig, Leipzig, Germany. Correspondence: Dr rer. nat. H Serke, Institute of Anatomy, University of Leipzig, Liebigstrasse 13, Leipzig 04103, Germany. E-mail: heike.serke@medizin.uni-leipzig.de 3 These authors contributed equally to this work. International Journal of Obesity (2011), 1 - 7 & 2011 Macmillan Publishers Limited All rights reserved 0307-0565/11 www.nature.com/ijo