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FJ
EXPRESS SUMMARY ARTICLE The Full-length version of this article is also available, published online June 28, 2005 as doi:10.1096/fj.05-3673fje. |
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* INSERM, "Avenir" team, Paris; University Paris 6, Paris, France; CHRU Pitié Salpétrière, Hôtel-Dieu Nutrition Department, Centre de Recherche en Nutrition Humaine, Paris, France.
INSERM U680, Paris, France; CHRU Tenon hospital, Biochemistry Department, Paris, France.
INSERM U586, Louis Bugnard Institute; University Paul Sabatier, Toulouse France; CHRU Toulouse France, Toulouse, France.
¥ Paris-Nord University, LIM & BIO, Bobigny, France;
¥¥ Hôtel-Dieu Surgery Department, Paris, France; and
# CHRU Pitié Salpétrière, Hôtel-Dieu hospital, Hormonology Department, Paris, France
1Correspondence: INSERM Avenir, Nutrition Department, Hôtel-Dieu, Place du parvis Notre-Dame, 75004 Paris. E-mail: karine.clement{at}htd.ap-hop-paris.fr
SPECIFIC AIMS
It is now well established that adipose tissue produces and secretes a variety of bioactive molecules into the bloodstream. In obesity, the circulating concentrations of many of these molecules is altered and some contribute to the development of complications associated with obesity such as diabetes and cardiovascular diseases. However, knowledge regarding the contribution of molecules originated from adipose tissue that could target vascular endothelium is scarce. This question has recently attracted considerable interest particularly by the identification of atherogenic or anti-atherogenic functions of the known adipose tissue biomarkers such as leptin, adiponectin, or PAI-1.
The purpose of our study was to identify candidate biomarkers overexpressed in adipose tissue of obese subjects that could link enlarged adipose mass to atherosclerosis. We used gene expression profiling at large scale on subcutaneous adipose tissue biopsies obtained from lean and obese individuals. This strategy led to the discovery of cathepsin S as a biomolecule expressed in human adipose tissue and whose gene expression was strongly correlated with body mass index (BMI). Cathepsin S is an elastolytic cysteine protease that has been implicated in the development of atherosclerosis in animal models and humans. These observations prompted us to follow up with characterization of cathepsin S expression and regulation in human adipose tissue. We performed a series of experiments to determine whether the cathepsin S protein was secreted by adipose tissue and from which cellular fraction (i.e., adipocytes or cells of stroma vascular fraction). Next we determined whether cathepsin S adipose tissue production was subject to regulation by proinflammatory factors as described in cells that express cathepsin S such as macrophage and smooth muscle cells. Finally, to investigate the physiological relevance of increased adipose tissue cathepsin S expression in obesity, we measured the circulating levels of cathepsin S and its endogenous inhibitor cystatin C in obese and nonobese subjects.
PRINCIPAL FINDINGS
1. Comparison of gene profiling in scWAT of 28 obese vs. 11 lean subjects
Using cDNA microarrays with accurate method of statistical analysis, we isolated 25 genes whose expression was significantly overexpressed in scWAT of obese vs. nonobese subjects and strongly correlated with BMI. Among these genes, cathepsin S was unique in its biochemical and enzymatic properties and in its potential adverse influence on vascular function.
2. Cathepsin S is overexpressed in adipose tissue of obese subjects
We confirmed by real time RT-PCR the differential expression of scWAT cathepsin S mRNA in an independent set of 7 lean and 19 obese women. The level of mRNA cathepsin S was 4.6-fold higher in scWAT of obese vs. lean subjects (Fig. 1
A). In this series of subjects, we checked the level of cathepsin K expression, which was also increased in scWAT of obese, but to a much extent than cathepsin S. The gene expression level of cathepsin S in scWAT positively correlated with body weight, BMI, total fat mass, and plasma triglyceride levels. The cathepsin S protein was also increased in scWAT of obese than nonobese subjects as assessed by Western blot and immunohistochemistry (Fig. 1B
).
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3. Cathepsin S secretion is increased by proinflammatory factors in adipose tissue
Cathepsin S was secreted at a rate of 14 ng. mL–1/g tissue 24 h–1 by adipose explants from 5 obese individuals. The rate of leptin secretion was in the same range (6.5 ng. mL–1/g tissue 24 h–1). The cathepsin S secretion is increased by proinflammatory factors such as lipopolysaccharide (LPS), IL-1ß, and TNF-
.
4. Cathepsin S is mainly secreted by adipocytes
To determine the cellular source of cathepsin S, adipocytes and the stroma vascular fraction (SVF) were prepared from adipose tissue. Cathepsin S secretion was determined in each fraction and normalized to the weight of adipose biopsy. The rate of cathepsin S secretion was 2-fold higher in adipocytes than in SVF cells. Moreover, the cathepsin S mRNA was increased in adipocyte cells of obese individuals.
5. Cathepsin S circulating levels are increased in obese subjects
We examined circulating levels of cathepsin S with ELISA test in 25 obese and 10 nonobese subjects. Serum cathepsin S levels were significantly 3-fold higher in obese than in nonobese subjects (Fig. 2
A). There was a significant correlation between circulating cathepsin S and body weight, BMI, fat mass, and plasma triglycerides. Cystatin C is an endogenous cathepsin S inhibitor that has been suggested to play role in protection against cardiovascular diseases. The ratio cathepsin S/cystatin C may be a determinant of cathepsin S biological effect. The ratio cathepsin S/cystatin C was significantly higher in obese than in nonobese subjects (Fig. 2B
).
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CONCLUSIONS AND SIGNIFICANCE
Because of the recognition that obesity is a major health threat in Western countries, the effort to understand the pathophysiology of expanded adipose tissue particularly its impact on development of cardiovascular diseases, has become crucial. This study identified a potential candidate biomolecule capable of linking obesity to atherogenesis starting from gene expression profiling comparison. We compared the adipose tissue expression profiling of 28 obese and 11 nonobese subjects, providing an extensive view for identifying new biomarkers. This strategy led to the discovery of cathepsin S as a biomolecule expressed by human adipose tissue in proportion with BMI. To our knowledge, this is the first report of cathepsin S expression in this tissue. We showed that cathepsin S is secreted by adipose tissue and increased in the circulation of obese subjects. The increased adipose production of cathepsin S likely contributed to the increase in circulating cathepsin S levels in obese individuals.
Modulations in extracellular matrix (ECM) components of adipose tissue are thought to be associated with fat mass expansion. Since cathepsin S is capable of degrading ECM components of adipose tissue such as laminin, collagens and chondroitin sulfate proteoglycan, we propose that cathepsin S could have local effects on remodeling of adipose tissue ECM.
In conclusion, we identified a novel marker of adiposity named cathepsin S. Based on our observations in obese patients and given the potential deleterious effect of cathepsin S excess on the arterial wall, we propose that cathepsin S represents a plausible molecular link between enlarged fat mass and atherosclerosis (Fig. 3
). This opens new avenues for examination of this protein in well-defined clinical populations to evaluate its predictive value of vascular diseases in obesity.
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FOOTNOTES
To read the full text of this article, go to http://www.fasebj.org/cgi/doi/10.1096/fj.05-3675fje;
This article has been cited by other articles:
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  J. Y. Kim, Y. Wu, and C. M. Smas Characterization of ScAP-23, a new cell line from murine subcutaneous adipose tissue, identifies genes for the molecular definition of preadipocytes Physiol Genomics, October 19, 2007; 31(2): 328 - 342. [Abstract] [Full Text] [PDF]
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 S. P. M. Lutgens, K. B. J. M. Cleutjens, M. J. A. P. Daemen, and S. Heeneman Cathepsin cysteine proteases in cardiovascular disease FASEB J, October 1, 2007; 21(12): 3029 - 3041. [Abstract] [Full Text] [PDF]
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 S. Taleb, R. Cancello, C. Poitou, C. Rouault, P. Sellam, P. Levy, J.-L. Bouillot, C. Coussieu, A. Basdevant, M. Guerre-Millo, et al. Weight Loss Reduces Adipose Tissue Cathepsin S and Its Circulating Levels in Morbidly Obese Women J. Clin. Endocrinol. Metab., March 1, 2006; 91(3): 1042 - 1047. [Abstract] [Full Text] [PDF]
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