Cathepsin K Null Mice Show Reduced Adiposity during the Rapid Accumulation of Fat Stores Marcella Funicello 1 , Michela Novelli 2 , Maurizio Ragni 1 , Teresa Vottari 1 , Cesare Cocuzza 1 , Joaquin Soriano-Lopez 1 , Chiara Chiellini 1 , Federico Boschi 3 , Pasquina Marzola 3 , Pellegrino Masiello 2 , Paul Saftig 4 , Ferruccio Santini 5 , Rene St-Jacques 6 , Sylvie Desmarais 7 , Nicolas Morin 7 , Joseph Mancini 7 , M. David Percival 7 , Aldo Pinchera 5 , Margherita Maffei 1 * 1 Dulbecco Telethon Institute at Department of Endocrinology and Metabolism, University Hospital of Pisa, Pisa, Italy, 2 Department of Experimental Pathology, Medical Biotechnologies, Infectivology and Epidemiology, University of Pisa, Pisa, Italy, 3 Department of Morphological-Biomedical Sciences, Human Anatomy and Histology Section, Medical Faculty, University of Verona, Verona, Italy, 4 Biochemical Institute, Christian-Albrechts University, Kiel, Germany, 5 Department of Endocrinology and Metabolism, University Hospital of Pisa, Pisa, Italy, 6 Department of Pharmacology, Merck Frosst Centre for Therapeutic Research, Kirkland, Quebec, Canada, 7 Department of Biochemistry and Molecular Biology, Merck Frosst Centre for Therapeutic Research, Kirkland, Quebec, Canada Growing evidences indicate that proteases are implicated in adipogenesis and in the onset of obesity. We previously reported that the cysteine protease cathepsin K (ctsk) is overexpressed in the white adipose tissue (WAT) of obese individuals. We herein characterized the WAT and the metabolic phenotype of ctsk deficient animals (ctsk2/2). When the growth rate of ctsk2/2 was compared to that of the wild type animals (WT), we could establish a time window (5–8 weeks of age) within which ctsk2/ 2display significantly lower body weight and WAT size as compared to WT. Such a difference was not observable in older mice. Upon treatment with high fat diet (HFD) for 12 weeks ctsk2/2 gained significantly less weight than WT and showed reduced brown adipose tissue, liver mass and a lower percentage of body fat. Plasma triglycerides, cholesterol and leptin were significantly lower in HFD-fed-ctsk2/2 as compared to HFD-fed WT animals. Adipocyte lipolysis rates were increased in both young and HFD-fed-ctsk2/2, as compared to WT. Carnitine palmitoyl transferase-1 activity, was higher in mitochondria isolated from the WAT of HFD treated ctsk2/2 as compared to WT. Together, these data indicate that ctsk ablation in mice results in reduced body fat content under conditions requiring a rapid accumulation of fat stores. This observation could be partly explained by an increased release and/or utilization of FFA and by an augmented ratio of lipolysis/lipogenesis. These results also demonstrate that under a HFD, ctsk deficiency confers a partial resistance to the development of dyslipidemia. Citation: Funicello M, Novelli M, Ragni M, Vottari T, Cocuzza C, et al (2007) Cathepsin K Null Mice Show Reduced Adiposity during the Rapid Accumulation of Fat Stores. PLoS ONE 2(8): e683. doi:10.1371/journal.pone.0000683 INTRODUCTION Cathepsin K (ctsk) is a member of the papain-like cysteine protease family. Members of this family are generally lysosomal enzymes although some, including ctsk are also found secreted from the cell. Ctsk has potent proteolytic activities against several extracellular matrix components such as collagen I and II, elastase, osteonectin and osteopontin [1–3]. Ctsk is most highly expressed in osteoclasts and its major recognized function is in the process of bone remodelling. However, broader potential roles for ctsk have been revealed by the identification of ctsk in a number of other cell and tissue types, including activated macrophages[4], thyroid [5], lung[6], atheroma [7,8], and skin [9]. Ctsk mRNA and protein were localized in samples of primary breast carcinoma [10] and prostate cancer [11]and it was suggested that this protease, like metalloproteases, may contribute to the invasive potential of certain cancer cells through the digestion of the extracellular matrix. Several recent reports link cysteine cathepsins with metabolic function. Taleb et al. [12] found that cathepsin S (ctss) gene expression is augmented in adipose tissue of obese humans and defined it as a novel marker of adiposity. Given the involvement of this enzyme in the development of atherosclerotic lesions Rodgers et al. [13], propose that ctss is a molecular link between obesity and atherosclerosis. Inactivation of cathepsin B (ctsb) protects against diet-induced fatty liver disease, as ctsb2/2 mice placed on a high carbohydrate diet developed marked obesity, but not the severe hepatomegaly, steatosis and associated dysmetabolic syndrome observed in the wild type (WT) controls placed on the same diet [14]. An even larger body of results indicate that metalloproteases (MMP) also play an important role in obesity and that their activities are important for the early events leading to adipogenesis. Chavey et al. [15] found that MMP-2, MMP-3, MMP-12, MMP-14, MMP-19, and TIMP-1, the endogenous MMP inhibitor, are strongly induced in obese adipose tissues compared with lean tissues. In addition, they and others found that pharmacological inhibition of MMP-2 and MMP-9 decreased adipose conversion in vivo [16] and in vitro [17,18]. We have recently reported that ctsk is relatively highly expressed in the white adipose tissue (WAT) and that it is a marker of adiposity as its transcript is induced in murine and human obesity [19]. In addition, we found that ctsk is a novel marker of adipogenesis, since Academic Editor: Alessandro Bartolomucci, University of Parma, Italy Received April 3, 2007; Accepted June 25, 2007; Published August 1, 2007 Copyright: ß 2007 Funicello et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Margherita Maffei is an Assistant Telethon Scientist. MF, MR, TV, CC, JSL were supported by a Telethon fellowship. This work was supported by Telethon Foundation (grant TCP99016) and Compagnia di San Paolo. Competing Interests: Rene St-Jacques, Sylvie Desmarais, Nicolas Morin, Joseph Mancini and M. David Percival are employees of Merck and Co., Inc. which is investigating Cathepsin K inhibitors as potential therapeutics. All the other authors do not disclose any competing interest. * To whom correspondence should be addressed. E-mail: mmaffei@dti.telethon. it PLoS ONE | www.plosone.org 1 August 2007 | Issue 8 | e683