Karin Müller-Decker

Cyclooxygenase-2 Controls Energy Homeostasis in Mice by de Novo Recruitment of Brown Adipocytes

Fat-Burning Fat In mammals, fat exists in two forms—the well-known white adipose tissue (WAT), which stores energy and is associated with obesity, and the lesser-known brown adipose tissue (BAT), which burns energy to generate heat. BATs role in human physiology was once thought to be restricted to newborns, but the recent discovery that adults also harbor functional BAT has re-ignited interest in the factors regulating BAT development and their potential as targets for anti-obesity therapies. Vegiopoulos et al. (p. 1158, published online 6 May; see the Perspective Ishibashi and Seale) now show that cyclooxygenase-2 (COX-2), an enzyme critical to prostaglandin synthesis, triggers fat progenitor cells in mice to differentiate into BAT rather than WAT. Mice overexpressing COX-2 displayed increased energy expenditure and were protected from diet-induced obesity. In mice, the development of energy-burning brown fat is regulated by an enzyme that is critical for prostaglandin synthesis. Obesity results from chronic energy surplus and excess lipid storage in white adipose tissue (WAT). In contrast, brown adipose tissue (BAT) efficiently burns lipids through adaptive thermogenesis. Studying mouse models, we show that cyclooxygenase (COX)–2, a rate-limiting enzyme in prostaglandin (PG) synthesis, is a downstream effector of β-adrenergic signaling in WAT and is required for the induction of BAT in WAT depots. PG shifted the differentiation of defined mesenchymal progenitors toward a brown adipocyte phenotype. Overexpression of COX-2 in WAT induced de novo BAT recruitment in WAT, increased systemic energy expenditure, and protected mice against high-fat diet–induced obesity. Thus, COX-2 appears integral to de novo BAT recruitment, which suggests that the PG pathway regulates systemic energy homeostasis.

RAGE signaling sustains inflammation and promotes tumor development

A broad range of experimental and clinical evidence has highlighted the central role of chronic inflammation in promoting tumor development. However, the molecular mechanisms converting a transient inflammatory tissue reaction into a tumor-promoting microenvironment remain largely elusive. We show that mice deficient for the receptor for advanced glycation end-products (RAGE) are resistant to DMBA/TPA-induced skin carcinogenesis and exhibit a severe defect in sustaining inflammation during the promotion phase. Accordingly, RAGE is required for TPA-induced up-regulation of proinflammatory mediators, maintenance of immune cell infiltration, and epidermal hyperplasia. RAGE-dependent up-regulation of its potential ligands S100a8 and S100a9 supports the existence of an S100/RAGE-driven feed-forward loop in chronic inflammation and tumor promotion. Finally, bone marrow chimera experiments revealed that RAGE expression on immune cells, but not keratinocytes or endothelial cells, is essential for TPA-induced dermal infiltration and epidermal hyperplasia. We show that RAGE signaling drives the strength and maintenance of an inflammatory reaction during tumor promotion and provide direct genetic evidence for a novel role for RAGE in linking chronic inflammation and cancer.

Transgenic cyclooxygenase-2 overexpression sensitizes mouse skin for carcinogenesis

Genetic and pharmacological evidence suggests that overexpression of cyclooxygenase-2 (COX-2) is critical for epithelial carcinogenesis and provides a major target for cancer chemoprevention by nonsteroidal antiinflammatory drugs. Transgenic mouse lines with keratin 5 promoter-driven COX-2 overexpression in basal epidermal cells exhibit a preneoplastic skin phenotype. As shown here, this phenotype depends on the level of COX-2 expression and COX-2-mediated prostaglandin accumulation. The transgenics did not develop skin tumors spontaneously but did so after a single application of an initiating dose of the carcinogen 7,12-dimethylbenz[a]anthracene (DMBA). Long-term treatment with the tumor promoter phorbol 12-myristate 13-acetate, as required for tumorigenesis in wild-type mice, was not necessary for transgenics. The ratios of squamous cell carcinomas to papillomas and of sebaceous gland adenomas to papillomas plus squamous cell carcinomas were increased markedly in transgenic mice treated with DMBA alone compared with DMBA/phorbol 12-myristate 13-acetate-treated transgenic and wild-type mice. Thus, COX-2 overexpression, which leads to high levels of epidermal prostaglandin E2, prostaglandin F2α, and 15-deoxyΔ12,14-PGJ2, is insufficient for tumor induction but transforms epidermis into an “autopromoted” state, i.e., dramatically sensitizes the tissue for genotoxic carcinogens.

Abnormal differentiation of epidermis in transgenic mice constitutively expressing cyclooxygenase-2 in skin

In prostanoid biosynthesis, the first two steps are catalyzed by cyclooxygenases (COX). In mice and humans, deregulated expression of COX-2, but not of COX-1, is characteristic of epithelial tumors, including squamous cell carcinomas of skin. To explore the function of COX-2 in epidermis, a keratin 5 promoter was used to direct COX-2 expression to the basal cells of interfollicular epidermis and the pilosebaceous appendage of transgenic mouse skin. COX-2 overexpression in the expected locations, resulting in increased prostaglandin levels in epidermis and plasma, correlated with a pronounced skin phenotype. Heterozygous transgenic mice exhibited a reduced hair follicle density. Moreover, postnatally hair follicle morphogenesis and thinning of interfollicular dorsal epidermis were delayed. Adult transgenics showed a body-site-dependent sparse coat of greasy hair, the latter caused by sebaceous gland hyperplasia and increased epicutaneous sebum levels. In tail skin, hyperplasia of scale epidermis reflecting an increased number of viable and cornified cell layers was observed. Hyperplasia was a result of a disturbed program of epidermal differentiation rather than an increased proliferation rate, as reflected by the strong suppression of keratin 10, involucrin, and loricrin expression in suprabasal cells. Further pathological signs were loss of cell polarity, mainly of basal keratinocytes, epidermal invaginations into the dermis, and formation of horn perls. Invaginating hyperplastic lobes were surrounded by CD31-positive vessels. These results demonstrate a causal relationship between transgenic COX-2 expression in basal keratinocytes and epidermal hyperplasia as well as dysplastic features at discrete body sites.

Enzymes involved in the biosynthesis of leukotriene B4 and prostaglandin E2 are active in sebaceous glands.

The expression of enzymes involved in leukotriene and prostaglandin signalling pathways, of interleukins 6 and 8 and of peroxisome proliferator-activated receptors in sebaceous glands of acne-involved facial skin was compared with those of non-involved skin of acne patients and of healthy individuals. Moreover, 5-lipoxygenase and leukotriene A4 hydrolase were expressed at mRNA and protein levels in vivo and in SZ95 sebocytes in vitro (leukotriene A4 hydrolase > 5-lipoxygenase), while 15-lipoxygenase-1 was only detected in cultured sebocytes. Cyclooxygenase-1 and cyclooxygenase-2 were also present. Peroxisome proliferator-activated receptors were constitutively expressed. Enhanced 5-lipoxygenase, cyclooxygenase 2 and interleukin 6 expression was detected in acne-involved facial skin. Arachidonic acid stimulated leukotriene B4 and interleukin 6 release as well as prostaglandin E2 biosynthesis in SZ95 sebocytes, induced abundant increase in neutral lipids and down-regulated peroxisome proliferator-activated receptor-α, but not receptor-γ1 mRNA levels, which were the predominant peroxisome proliferator-activated receptor isotypes in SZ95 sebocytes. In conclusion, human sebocytes possess the enzyme machinery for functional leukotriene and prostaglandin pathways. A comprehensive link between inflammation and sebaceous lipid synthesis is provided.

Prostaglandin-H-synthase isozyme expression in normal and neoplastic human skin

Expression of prostaglandin‐H‐synthase (PGHS) isozymes was analyzed in 50 biopsies of normal human skin and of pre‐malignant and malignant skin lesions, by means of quantitative RT‐PCR, immunoprecipitation and Western blotting, as well as immunohistochemistry. Normal skin constitutively expressed PGHS‐1 in all cell layers of the epidermis, in endothelial cells of small blood vessels and in sweat‐gland epithelium. PGHS‐2 expression was very low and restricted to a few keratinocytes of the interfollicular and follicular epidermis. Steady‐state concentrations of PGHS‐1 and PGHS‐2 mRNA were similar in normal skin and in basal‐cell carcinomas, but PGHS‐1 mRNA was reduced and PGHS‐2 mRNA was elevated in actinic keratoses, squamous‐cell carcinomas and keratoacanthomas. PGHS‐1 protein was detected in all tumor biopsies, being occasionally increased in basal‐cell carcinomas. High amounts of PGHS‐2 protein were found in actinic keratoses, squamous‐cell carcinomas and keratoacanthomas, but not in basal‐cell carcinomas. Four malignant melanomas included in this study contained PGHS‐1 but no PGHS‐2 protein. Immunohistochemical analysis of the biopsies identified keratinocytes, in addition to cells of inflammatory infiltrates and of dendritic morphology, as the major PGHS‐expressing cell types. PGHS‐2‐specific signals were spread throughout the epidermal part of actinic keratoses and squamous‐cell carcinomas. These data suggest that constitutive up‐regulation of PGHS‐2 expression is a consistent pre‐malignant event in squamous‐cell cancer development in man, as it is in animal models of skin carcinogenesis. Thus, pre‐cancerous lesions such as actinic keratoses present a likely target for chemoprevention of skin cancer by selective PGHS‐2 inhibitors. Int. J. Cancer 82:648–656, 1999.

Localization of prostaglandin H synthase isoenzymes in murine epidermal tumors: Suppression of skin tumor promotion by inhibition of prostaglandin H synthase-2

The growth factor– and phorbol ester–inducible prostaglandin H synthase (PGHS)‐2 has been found to be constitutively overexpressed in epidermal tumors generated by the initiation‐promotion protocol in murine skin, whereas the expression of PGHS‐1 does not change under these conditions. In this paper we report the intra‐tumor distribution of the aberrantly expressed PGHS‐2 and the cancer chemopreventive activity of a specific PGHS‐2 inhibitor. By immunohistochemical methods using isoenzyme‐specific antibodies, we found that the PGHS‐1 protein was expressed in keratinocytes and Langerhans cells dispersed throughout the epithelial part of papillomas and squamous cell carcinomas and in inflammatory infiltrates occasionally seen in these tumors. A uniform pattern of PGHS‐2 expression was observed in the basal keratinocytes of papillomas and in the follicular keratinocytes of carcinomas. In addition, Langerhans cells as well as tumor‐associated inflammatory infiltrates exhibited PGHS‐2–specific immunoreactivity. PGHS‐2–catalyzed prostaglandin synthesis stimulated by the phorbol ester 12‐O‐tetradecanoylphorbol‐13 acetate (TPA) in mouse epidermis in vivo was dose‐dependently suppressed by topical administration of SC‐58125, a specific PGHS‐2 inhibitor. TPA‐induced edema formation, epidermal DNA synthesis, and mitotic activity were not impaired by SC‐58125 applied at a dose that inhibited TPA‐induced prostaglandin E2 synthesis. However, the repetitive epicutaneous administration of SC‐58125 substantially and significantly suppressed papilloma development. Malignant progression of papillomas was slightly retarded by the drug. These results indicate that aberrant expression of PGHS‐2 in epidermal tumors may be a relevant target for prevention of epidermal cancer development in experimental animals and that the PGHS‐2–specific inhibitor SC‐58125, which is a potent inhibitor of tumor promotion in mouse skin, may be important for cancer chemoprevention in humans as well. Mol. Carcinog. 23:36–44, 1998.

The cyclooxygenase-2-mediated prostaglandin signaling is causally related to epithelial carcinogenesis.

Epidemiologic, pharmacologic, clinical, and experimental studies document the importance of prostaglandin (PG) signaling in cancer development, including non‐melanoma skin cancer lesions in humans and mice. First of all, enzymes involved in PG biosynthesis, such as cyclooxygenase (COX)‐2 and/or membrane prostaglandin E synthase (mPGES)‐1, were found to be overexpressed in a wide range of premalignant and malignant epithelial tumors, including those of the skin, breast, esophagus, stomach, colorectum, pancreas, and bladder. On the other hand, 15‐hydroxy‐prostaglandin dehydrogenase (15‐PGDH), which is involved in the degradation pathway of PG including PGE2, thus counteracting the activities of COX‐2 and PGES, was found to be downregulated in human epithelial tumors, indicating a tumor suppressor activity of this enzyme. Most remarkably, genetic studies showed that mice, which are deficient in COX‐2 and/or PGES are resistant to the development of cancer of skin, colon, and stomach. In contrast, the forced overexpression of COX‐2 in proliferative compartments of simple or stratified epithelia such as skin epidermis, urinary bladder, mammary gland, and pancreas results in spontaneous hyperplasia and dysplasia in transgenic mice. In skin, the pathological changes are found to be due to an abnormal process of terminal differentiation, while in other tissues, hyperproliferation seems to be the main contributer to the pre‐invasive neoplasms. Moreover, the COX‐2 transgenic mouse lines are sensitized for cancer development.

Production of sophorolipids from whey

Abstract Sophorolipids, obtained by a two-stage process starting from deproteinized whey concentrate using Cryptococcus curvatus ATCC 20509 and Candida bombicola ATCC 22214, were compared to products from one-stage processes, using different lipidic compounds as substrates. Results showed that above all carbon source and not cultivation conditions had a distinct influence on the composition of the crude product mixture and therefore on the physicochemical and biological properties of the sophorolipids, such as, for example, surface activity, cytotoxicity and stability against hydrolases. The results were completed by corresponding data for purified mono- and diacetylated (17-hydroxyoctadecenoic)-1′,4′′-lactonized sophorolipids. Crude sophorolipid mixtures showed moderate to good surface active properties (SFTmin 39 mN m−1, CMC 130 mg l−1), water solubilities (2–3 g l−1) and low cytotoxicities (LC50 300–700 mg l−1). In contrast, purified sophorolipids were more surface active (SFTmin 36 mN m−1, CMC 10 mg l−1), less water soluble (max. 70 mg l−1) and showed stronger cytotoxic effects (LC50 15 mg l−1). Incubation of crude sophorolipid mixtures with different hydrolases demonstrated that treatment with commercially available lipases such as from Candida rugosa and Mucor miehei distinctly reduced the surface active properties of the sophorolipids, while treatment with porcine liver esterase and glycosidases had no effect.

cAMP-dependent Signaling Regulates the Adipogenic Effect of n-6 Polyunsaturated Fatty Acids

The effect of n-6 polyunsaturated fatty acids (n-6 PUFAs) on adipogenesis and obesity is controversial. Using in vitro cell culture models, we show that n-6 PUFAs was pro-adipogenic under conditions with base-line levels of cAMP, but anti-adipogenic when the levels of cAMP were elevated. The anti-adipogenic action of n-6 PUFAs was dependent on a cAMP-dependent protein kinase-mediated induction of cyclooxygenase expression and activity. We show that n-6 PUFAs were pro-adipogenic when combined with a high carbohydrate diet, but non-adipogenic when combined with a high protein diet in mice. The high protein diet increased the glucagon/insulin ratio, leading to elevated cAMP-dependent signaling and induction of cyclooxygenase-mediated prostaglandin synthesis. Mice fed the high protein diet had a markedly lower feed efficiency than mice fed the high carbohydrate diet. Yet, oxygen consumption and apparent heat production were similar. Mice on a high protein diet had increased hepatic expression of PGC-1α (peroxisome proliferator-activated receptor γ coactivator 1α) and genes involved in energy-demanding processes like urea synthesis and gluconeogenesis. We conclude that cAMP signaling is pivotal in regulating the adipogenic effect of n-6 PUFAs and that diet-induced differences in cAMP levels may explain the ability of n-6 PUFAs to either enhance or counteract adipogenesis and obesity.